Baxter, J.S.
Brough, R.
Krastev, D.B.
Song, F.
Sridhar, S.
Gulati, A.
Alexander, J.
Roumeliotis, T.I.
Kozik, Z.
Choudhary, J.S.
Haider, S.
Pettitt, S.J.
Tutt, A.N.
Lord, C.J.
(2024). Cancer-associated FBXW7 loss is synthetic lethal with pharmacological targeting of CDC7. Mol oncol,
Vol.18
(2),
pp. 369-385.
show abstract
full text
The F-box and WD repeat domain containing 7 (FBXW7) tumour suppressor gene encodes a substrate-recognition subunit of Skp, cullin, F-box (SCF)-containing complexes. The tumour-suppressive role of FBXW7 is ascribed to its ability to drive ubiquitination and degradation of oncoproteins. Despite this molecular understanding, therapeutic approaches that target defective FBXW7 have not been identified. Using genome-wide clustered regularly interspaced short palindromic repeats (CRISPR)-Cas9 screens, focussed RNA-interference screens and whole and phospho-proteome mass spectrometry profiling in multiple FBXW7 wild-type and defective isogenic cell lines, we identified a number of FBXW7 synthetic lethal targets, including proteins involved in the response to replication fork stress and proteins involved in replication origin firing, such as cell division cycle 7-related protein kinase (CDC7) and its substrate, DNA replication complex GINS protein SLD5 (GINS4). The CDC7 synthetic lethal effect was confirmed using small-molecule inhibitors. Mechanistically, FBXW7/CDC7 synthetic lethality is dependent upon the replication factor telomere-associated protein RIF1 (RIF1), with RIF1 silencing reversing the FBXW7-selective effects of CDC7 inhibition. The delineation of FBXW7 synthetic lethal effects we describe here could serve as the starting point for subsequent drug discovery and/or development in this area..
Harvey-Jones, E.
Raghunandan, M.
Robbez-Masson, L.
Magraner-Pardo, L.
Alaguthurai, T.
Yablonovitch, A.
Yen, J.
Xiao, H.
Brough, R.
Frankum, J.
Song, F.
Yeung, J.
Savy, T.
Gulati, A.
Alexander, J.
Kemp, H.
Starling, C.
Konde, A.
Marlow, R.
Cheang, M.
Proszek, P.
Hubank, M.
Cai, M.
Trendell, J.
Lu, R.
Liccardo, R.
Ravindran, N.
Llop-Guevara, A.
Rodriguez, O.
Balmana, J.
Lukashchuk, N.
Dorschner, M.
Drusbosky, L.
Roxanis, I.
Serra, V.
Haider, S.
Pettitt, S.J.
Lord, C.J.
Tutt, A.N.
(2024). Longitudinal profiling identifies co-occurring BRCA1/2 reversions, TP53BP1, RIF1 and PAXIP1 mutations in PARP inhibitor-resistant advanced breast cancer. Ann oncol,
Vol.35
(4),
pp. 364-380.
show abstract
full text
BACKGROUND: Resistance to therapies that target homologous recombination deficiency (HRD) in breast cancer limits their overall effectiveness. Multiple, preclinically validated, mechanisms of resistance have been proposed, but their existence and relative frequency in clinical disease are unclear, as is how to target resistance. PATIENTS AND METHODS: Longitudinal mutation and methylation profiling of circulating tumour (ct)DNA was carried out in 47 patients with metastatic BRCA1-, BRCA2- or PALB2-mutant breast cancer treated with HRD-targeted therapy who developed progressive disease-18 patients had primary resistance and 29 exhibited response followed by resistance. ctDNA isolated at multiple time points in the patient treatment course (before, on-treatment and at progression) was sequenced using a novel >750-gene intron/exon targeted sequencing panel. Where available, matched tumour biopsies were whole exome and RNA sequenced and also used to assess nuclear RAD51. RESULTS: BRCA1/2 reversion mutations were present in 60% of patients and were the most prevalent form of resistance. In 10 cases, reversions were detected in ctDNA before clinical progression. Two new reversion-based mechanisms were identified: (i) intragenic BRCA1/2 deletions with intronic breakpoints; and (ii) intragenic BRCA1/2 secondary mutations that formed novel splice acceptor sites, the latter being confirmed by in vitro minigene reporter assays. When seen before commencing subsequent treatment, reversions were associated with significantly shorter time to progression. Tumours with reversions retained HRD mutational signatures but had functional homologous recombination based on RAD51 status. Although less frequent than reversions, nonreversion mechanisms [loss-of-function (LoF) mutations in TP53BP1, RIF1 or PAXIP1] were evident in patients with acquired resistance and occasionally coexisted with reversions, challenging the notion that singular resistance mechanisms emerge in each patient. CONCLUSIONS: These observations map the prevalence of candidate drivers of resistance across time in a clinical setting, information with implications for clinical management and trial design in HRD breast cancers..
Alexander, J.
Schipper, K.
Nash, S.
Brough, R.
Kemp, H.
Iacovacci, J.
Isacke, C.
Natrajan, R.
Sawyer, E.
Lord, C.J.
Haider, S.
(2024). Pathway-based signatures predict patient outcome, chemotherapy benefit and synthetic lethal dependencies in invasive lobular breast cancer. Br j cancer,
.
show abstract
BACKGROUND: Invasive Lobular Carcinoma (ILC) is a morphologically distinct breast cancer subtype that represents up to 15% of all breast cancers. Compared to Invasive Breast Carcinoma of No Special Type (IBC-NST), ILCs exhibit poorer long-term outcome and a unique pattern of metastasis. Despite these differences, the systematic discovery of robust prognostic biomarkers and therapeutically actionable molecular pathways in ILC remains limited. METHODS: Pathway-centric multivariable models using statistical machine learning were developed and tested in seven retrospective clinico-genomic cohorts (n = 996). Further external validation was performed using a new RNA-Seq clinical cohort of aggressive ILCs (n = 48). RESULTS AND CONCLUSIONS: mRNA dysregulation scores of 25 pathways were strongly prognostic in ILC (FDR-adjusted P < 0.05). Of these, three pathways including Cell-cell communication, Innate immune system and Smooth muscle contraction were also independent predictors of chemotherapy response. To aggregate these findings, a multivariable machine learning predictor called PSILC was developed and successfully validated for predicting overall and metastasis-free survival in ILC. Integration of PSILC with CRISPR-Cas9 screening data from breast cancer cell lines revealed 16 candidate therapeutic targets that were synthetic lethal with high-risk ILCs. This study provides interpretable prognostic and predictive biomarkers of ILC which could serve as the starting points for targeted drug discovery for this disease..
Tien, J.C.
Chang, Y.
Zhang, Y.
Chou, J.
Cheng, Y.
Wang, X.
Yang, J.
Mannan, R.
Shah, P.
Wang, X.-.
Todd, A.J.
Eyunni, S.
Cheng, C.
Rebernick, R.J.
Xiao, L.
Bao, Y.
Neiswender, J.
Brough, R.
Pettitt, S.J.
Cao, X.
Miner, S.J.
Zhou, L.
Wu, Y.-.
Labanca, E.
Wang, Y.
Parolia, A.
Cieslik, M.
Robinson, D.R.
Wang, Z.
Feng, F.Y.
Lord, C.J.
Ding, K.
Chinnaiyan, A.M.
(2024). CDK12 Loss Promotes Prostate Cancer Development While Exposing Vulnerabilities to Paralog-Based Synthetic Lethality. Biorxiv,
.
show abstract
Biallelic loss of cyclin-dependent kinase 12 (CDK12) defines a unique molecular subtype of metastatic castration-resistant prostate cancer (mCRPC). It remains unclear, however, whether CDK12 loss per se is sufficient to drive prostate cancer development-either alone, or in the context of other genetic alterations-and whether CDK12-mutant tumors exhibit sensitivity to specific pharmacotherapies. Here, we demonstrate that tissue-specific Cdk12 ablation is sufficient to induce preneoplastic lesions and robust T cell infiltration in the mouse prostate. Allograft-based CRISPR screening demonstrated that Cdk12 loss is positively associated with Trp53 inactivation but negatively associated with Pten inactivation-akin to what is observed in human mCRPC. Consistent with this, ablation of Cdk12 in prostate organoids with concurrent Trp53 loss promotes their proliferation and ability to form tumors in mice, while Cdk12 knockout in the Pten-null prostate cancer mouse model abrogates tumor growth. Bigenic Cdk12 and Trp53 loss allografts represent a new syngeneic model for the study of androgen receptor (AR)-positive, luminal prostate cancer. Notably, Cdk12/Trp53 loss prostate tumors are sensitive to immune checkpoint blockade. Cdk12-null organoids (either with or without Trp53 co-ablation) and patient-derived xenografts from tumors with CDK12 inactivation are highly sensitive to inhibition or degradation of its paralog kinase, CDK13. Together, these data identify CDK12 as a bona fide tumor suppressor gene with impact on tumor progression and lends support to paralog-based synthetic lethality as a promising strategy for treating CDK12-mutant mCRPC..
Harvey-Jones, E.J.
Lord, C.J.
Tutt, A.N.
(2023). Systemic Therapy for Hereditary Breast Cancers. Hematol oncol clin north am,
Vol.37
(1),
pp. 203-224.
show abstract
full text
Approximately 5% to 10% of all breast cancers are hereditary; many of which are caused by pathogenic variants in genes required for homologous recombination, including BRCA1 and BRCA2. Here we discuss systemic treatment for such breast cancers, including approved chemotherapeutic approaches and also targeted treatment approaches using poly-(ADP ribose) polymerase inhibitors. We also discuss experimental approaches to treating hereditary breast cancer, including new small molecule DNA repair inhibitors and also immunomodulatory agents. Finally, we discuss how drug resistance emerges in patients with hereditary breast cancer, how this might be delayed or prevented, and how biomarker-adapted treatment is molding the future management of hereditary breast cancer..
Bland, P.
Saville, H.
Wai, P.T.
Curnow, L.
Muirhead, G.
Nieminuszczy, J.
Ravindran, N.
John, M.B.
Hedayat, S.
Barker, H.E.
Wright, J.
Yu, L.
Mavrommati, I.
Read, A.
Peck, B.
Allen, M.
Gazinska, P.
Pemberton, H.N.
Gulati, A.
Nash, S.
Noor, F.
Guppy, N.
Roxanis, I.
Pratt, G.
Oldreive, C.
Stankovic, T.
Barlow, S.
Kalirai, H.
Coupland, S.E.
Broderick, R.
Alsafadi, S.
Houy, A.
Stern, M.-.
Pettit, S.
Choudhary, J.S.
Haider, S.
Niedzwiedz, W.
Lord, C.J.
Natrajan, R.
(2023). SF3B1 hotspot mutations confer sensitivity to PARP inhibition by eliciting a defective replication stress response. Nat genet,
Vol.55
(8),
pp. 1311-1323.
show abstract
full text
SF3B1 hotspot mutations are associated with a poor prognosis in several tumor types and lead to global disruption of canonical splicing. Through synthetic lethal drug screens, we identify that SF3B1 mutant (SF3B1MUT) cells are selectively sensitive to poly (ADP-ribose) polymerase inhibitors (PARPi), independent of hotspot mutation and tumor site. SF3B1MUT cells display a defective response to PARPi-induced replication stress that occurs via downregulation of the cyclin-dependent kinase 2 interacting protein (CINP), leading to increased replication fork origin firing and loss of phosphorylated CHK1 (pCHK1; S317) induction. This results in subsequent failure to resolve DNA replication intermediates and G2/M cell cycle arrest. These defects are rescued through CINP overexpression, or further targeted by a combination of ataxia-telangiectasia mutated and PARP inhibition. In vivo, PARPi produce profound antitumor effects in multiple SF3B1MUT cancer models and eliminate distant metastases. These data provide the rationale for testing the clinical efficacy of PARPi in a biomarker-driven, homologous recombination proficient, patient population..
Pettitt, S.J.
Shao, N.
Zatreanu, D.
Frankum, J.
Bajrami, I.
Brough, R.
Krastev, D.B.
Roumeliotis, T.I.
Choudhary, J.S.
Lorenz, S.
Rust, A.
de Bono, J.S.
Yap, T.A.
Tutt, A.N.
Lord, C.J.
(2023). A HUWE1 defect causes PARP inhibitor resistance by modulating the BRCA1-∆11q splice variant. Oncogene,
Vol.42
(36),
pp. 2701-2709.
show abstract
full text
Although PARP inhibitors (PARPi) now form part of the standard-of-care for the treatment of homologous recombination defective cancers, de novo and acquired resistance limits their overall effectiveness. Previously, overexpression of the BRCA1-∆11q splice variant has been shown to cause PARPi resistance. How cancer cells achieve increased BRCA1-∆11q expression has remained unclear. Using isogenic cells with different BRCA1 mutations, we show that reduction in HUWE1 leads to increased levels of BRCA1-∆11q and PARPi resistance. This effect is specific to cells able to express BRCA1-∆11q (e.g. BRCA1 exon 11 mutant cells) and is not seen in BRCA1 mutants that cannot express BRCA1-∆11q, nor in BRCA2 mutant cells. As well as increasing levels of BRCA1-∆11q protein in exon 11 mutant cells, HUWE1 silencing also restores RAD51 nuclear foci and platinum salt resistance. HUWE1 catalytic domain mutations were also seen in a case of PARPi resistant, BRCA1 exon 11 mutant, high grade serous ovarian cancer. These results suggest how elevated levels of BRCA1-∆11q and PARPi resistance can be achieved, identify HUWE1 as a candidate biomarker of PARPi resistance for assessment in future clinical trials and illustrate how some PARPi resistance mechanisms may only operate in patients with particular BRCA1 mutations..
Ryan, C.J.
Devakumar, L.P.
Pettitt, S.J.
Lord, C.J.
(2023). Complex synthetic lethality in cancer. Nat genet,
Vol.55
(12),
pp. 2039-2048.
show abstract
The concept of synthetic lethality has been widely applied to identify therapeutic targets in cancer, with varying degrees of success. The standard approach normally involves identifying genetic interactions between two genes, a driver and a target. In reality, however, most cancer synthetic lethal effects are likely complex and also polygenic, being influenced by the environment in addition to involving contributions from multiple genes. By acknowledging and delineating this complexity, we describe in this article how the success rate in cancer drug discovery and development could be improved..
Zelceski, A.
Francica, P.
Lingg, L.
Mutlu, M.
Stok, C.
Liptay, M.
Alexander, J.
Baxter, J.S.
Brough, R.
Gulati, A.
Haider, S.
Raghunandan, M.
Song, F.
Sridhar, S.
Forment, J.V.
O'Connor, M.J.
Davies, B.R.
van Vugt, M.A.
Krastev, D.B.
Pettitt, S.J.
Tutt, A.N.
Rottenberg, S.
Lord, C.J.
(2023). MND1 and PSMC3IP control PARP inhibitor sensitivity in mitotic cells. Cell rep,
Vol.42
(5),
p. 112484.
show abstract
full text
The PSMC3IP-MND1 heterodimer promotes meiotic D loop formation before DNA strand exchange. In genome-scale CRISPR-Cas9 mutagenesis and interference screens in mitotic cells, depletion of PSMC3IP or MND1 causes sensitivity to poly (ADP-Ribose) polymerase inhibitors (PARPi) used in cancer treatment. PSMC3IP or MND1 depletion also causes ionizing radiation sensitivity. These effects are independent of PSMC3IP/MND1's role in mitotic alternative lengthening of telomeres. PSMC3IP- or MND1-depleted cells accumulate toxic RAD51 foci in response to DNA damage, show impaired homology-directed DNA repair, and become PARPi sensitive, even in cells lacking both BRCA1 and TP53BP1. Epistasis between PSMC3IP-MND1 and BRCA1/BRCA2 defects suggest that abrogated D loop formation is the cause of PARPi sensitivity. Wild-type PSMC3IP reverses PARPi sensitivity, whereas a PSMC3IP p.Glu201del mutant associated with D loop defects and ovarian dysgenesis does not. These observations suggest that meiotic proteins such as MND1 and PSMC3IP have a greater role in mitotic DNA repair..
Pettitt, S.J.
Ryan, C.J.
Lord, C.J.
(2023). Exploiting Cancer Synthetic Lethality in Cancer-Lessons Learnt from PARP Inhibitors. Cancer treat res,
Vol.186,
pp. 13-23.
show abstract
PARP inhibitors now have proven utility in the treatment of homologous recombination (HR) defective cancers. These drugs, and the synthetic lethality effect they exploit, have not only taught us how to approach the treatment of HR defective cancers but have also illuminated how resistance to a synthetic lethal approach can occur, how cancer-associated synthetic lethal effects are perhaps more complex than we imagine, how the better use of biomarkers could improve the success of treatment and even how drug resistance might be targeted. Here, we discuss some of the lessons learnt from the study of PARP inhibitor synthetic lethality and how these lessons might have wider application. Specifically, we discuss the concept of synthetic lethal penetrance, phenocopy effects in cancer such as BRCAness, synthetic lethal resistance, the polygenic and complex nature of synthetic lethal interactions, how evolutionary double binds could be exploited in treatment as well as future horizons for the field..
Krastev, D.B.
Li, S.
Sun, Y.
Wicks, A.J.
Hoslett, G.
Weekes, D.
Badder, L.M.
Knight, E.G.
Marlow, R.
Pardo, M.C.
Yu, L.
Talele, T.T.
Bartek, J.
Choudhary, J.S.
Pommier, Y.
Pettitt, S.J.
Tutt, A.N.
Ramadan, K.
Lord, C.J.
(2022). The ubiquitin-dependent ATPase p97 removes cytotoxic trapped PARP1 from chromatin. Nat cell biol,
Vol.24
(1),
pp. 62-73.
show abstract
full text
Poly (ADP-ribose) polymerase (PARP) inhibitors elicit antitumour activity in homologous recombination-defective cancers by trapping PARP1 in a chromatin-bound state. How cells process trapped PARP1 remains unclear. Using wild-type and a trapping-deficient PARP1 mutant combined with rapid immunoprecipitation mass spectrometry of endogenous proteins and Apex2 proximity labelling, we delineated mass spectrometry-based interactomes of trapped and non-trapped PARP1. These analyses identified an interaction between trapped PARP1 and the ubiquitin-regulated p97 ATPase/segregase. We found that following trapping, PARP1 is SUMOylated by PIAS4 and subsequently ubiquitylated by the SUMO-targeted E3 ubiquitin ligase RNF4, events that promote recruitment of p97 and removal of trapped PARP1 from chromatin. Small-molecule p97-complex inhibitors, including a metabolite of the clinically used drug disulfiram (CuET), prolonged PARP1 trapping and enhanced PARP inhibitor-induced cytotoxicity in homologous recombination-defective tumour cells and patient-derived tumour organoids. Together, these results suggest that p97 ATPase plays a key role in the processing of trapped PARP1 and the response of tumour cells to PARP inhibitors..
Llorca-Cardenosa, M.J.
Aronson, L.I.
Krastev, D.B.
Nieminuszczy, J.
Alexander, J.
Song, F.
Dylewska, M.
Broderick, R.
Brough, R.
Zimmermann, A.
Zenke, F.T.
Gurel, B.
Riisnaes, R.
Ferreira, A.
Roumeliotis, T.
Choudhary, J.
Pettitt, S.J.
de Bono, J.
Cervantes, A.
Haider, S.
Niedzwiedz, W.
Lord, C.J.
Chong, I.Y.
(2022). SMG8/SMG9 Heterodimer Loss Modulates SMG1 Kinase to Drive ATR Inhibitor Resistance. Cancer res,
Vol.82
(21),
pp. 3962-3973.
show abstract
full text
UNLABELLED: Gastric cancer represents the third leading cause of global cancer mortality and an area of unmet clinical need. Drugs that target the DNA damage response, including ATR inhibitors (ATRi), have been proposed as novel targeted agents in gastric cancer. Here, we sought to evaluate the efficacy of ATRi in preclinical models of gastric cancer and to understand how ATRi resistance might emerge as a means to identify predictors of ATRi response. A positive selection genome-wide CRISPR-Cas9 screen identified candidate regulators of ATRi resistance in gastric cancer. Loss-of-function mutations in either SMG8 or SMG9 caused ATRi resistance by an SMG1-mediated mechanism. Although ATRi still impaired ATR/CHK1 signaling in SMG8/9-defective cells, other characteristic responses to ATRi exposure were not seen, such as changes in ATM/CHK2, γH2AX, phospho-RPA, or 53BP1 status or changes in the proportions of cells in S- or G2-M-phases of the cell cycle. Transcription/replication conflicts (TRC) elicited by ATRi exposure are a likely cause of ATRi sensitivity, and SMG8/9-defective cells exhibited a reduced level of ATRi-induced TRCs, which could contribute to ATRi resistance. These observations suggest ATRi elicits antitumor efficacy in gastric cancer but that drug resistance could emerge via alterations in the SMG8/9/1 pathway. SIGNIFICANCE: These findings reveal how cancer cells acquire resistance to ATRi and identify pathways that could be targeted to enhance the overall effectiveness of these inhibitors..
Vergote, I.
González-Martín, A.
Ray-Coquard, I.
Harter, P.
Colombo, N.
Pujol, P.
Lorusso, D.
Mirza, M.R.
Brasiuniene, B.
Madry, R.
Brenton, J.D.
Ausems, M.G.
Büttner, R.
Lambrechts, D.
European experts’ consensus group,
(2022). European experts consensus: BRCA/homologous recombination deficiency testing in first-line ovarian cancer. Ann oncol,
Vol.33
(3),
pp. 276-287.
show abstract
full text
BACKGROUND: Homologous recombination repair (HRR) enables fault-free repair of double-stranded DNA breaks. HRR deficiency is predicted to occur in around half of high-grade serous ovarian carcinomas. Ovarian cancers harbouring HRR deficiency typically exhibit sensitivity to poly-ADP ribose polymerase inhibitors (PARPi). Current guidelines recommend a range of approaches for genetic testing to identify predictors of sensitivity to PARPi in ovarian cancer and to identify genetic predisposition. DESIGN: To establish a European-wide consensus for genetic testing (including the genetic care pathway), decision making and clinical management of patients with recently diagnosed advanced ovarian cancer, and the validity of biomarkers to predict the effectiveness of PARPi in the first-line setting. The collaborative European experts' consensus group consisted of a steering committee (n = 14) and contributors (n = 84). A (modified) Delphi process was used to establish consensus statements based on a systematic literature search, conducted according to the Preferred Reporting Items for Systematic Reviews and Meta-analyses guidelines. RESULTS: A consensus was reached on 34 statements amongst 98 caregivers (including oncologists, pathologists, clinical geneticists, genetic researchers, and patient advocates). The statements concentrated on (i) the value of testing for BRCA1/2 mutations and HRR deficiency testing, including when and whom to test; (ii) the importance of developing new and better HRR deficiency tests; (iii) the importance of germline non-BRCA HRR and mismatch repair gene mutations for predicting familial risk, but not for predicting sensitivity to PARPi, in the first-line setting; (iv) who should be able to inform patients about genetic testing, and what training and education should these caregivers receive. CONCLUSION: These consensus recommendations, from a multidisciplinary panel of experts from across Europe, provide clear guidance on the use of BRCA and HRR deficiency testing for recently diagnosed patients with advanced ovarian cancer..
Tarantino, D.
Walker, C.
Weekes, D.
Pemberton, H.
Davidson, K.
Torga, G.
Frankum, J.
Mendes-Pereira, A.M.
Prince, C.
Ferro, R.
Brough, R.
Pettitt, S.J.
Lord, C.J.
Grigoriadis, A.
Nj Tutt, A.
(2022). Functional screening reveals HORMAD1-driven gene dependencies associated with translesion synthesis and replication stress tolerance. Oncogene,
Vol.41
(32),
pp. 3969-3977.
show abstract
full text
HORMAD1 expression is usually restricted to germline cells, but it becomes mis-expressed in epithelial cells in ~60% of triple-negative breast cancers (TNBCs), where it is associated with elevated genomic instability (1). HORMAD1 expression in TNBC is bimodal with HORMAD1-positive TNBC representing a biologically distinct disease group. Identification of HORMAD1-driven genetic dependencies may uncover novel therapies for this disease group. To study HORMAD1-driven genetic dependencies, we generated a SUM159 cell line model with doxycycline-inducible HORMAD1 that replicated genomic instability phenotypes seen in HORMAD1-positive TNBC (1). Using small interfering RNA screens, we identified candidate genes whose depletion selectively inhibited the cellular growth of HORMAD1-expressing cells. We validated five genes (ATR, BRIP1, POLH, TDP1 and XRCC1), depletion of which led to reduced cellular growth or clonogenic survival in cells expressing HORMAD1. In addition to the translesion synthesis (TLS) polymerase POLH, we identified a HORMAD1-driven dependency upon additional TLS polymerases, namely POLK, REV1, REV3L and REV7. Our data confirms that out-of-context somatic expression of HORMAD1 can lead to genomic instability and reveals that HORMAD1 expression induces dependencies upon replication stress tolerance pathways, such as translesion synthesis. Our data also suggest that HORMAD1 expression could be a patient selection biomarker for agents targeting replication stress..
Van Baelen, K.
Geukens, T.
Maetens, M.
Tjan-Heijnen, V.
Lord, C.J.
Linn, S.
Bidard, F.-.
Richard, F.
Yang, W.W.
Steele, R.E.
Pettitt, S.J.
Van Ongeval, C.
De Schepper, M.
Isnaldi, E.
Nevelsteen, I.
Smeets, A.
Punie, K.
Voorwerk, L.
Wildiers, H.
Floris, G.
Vincent-Salomon, A.
Derksen, P.W.
Neven, P.
Senkus, E.
Sawyer, E.
Kok, M.
Desmedt, C.
(2022). Current and future diagnostic and treatment strategies for patients with invasive lobular breast cancer. Ann oncol,
Vol.33
(8),
pp. 769-785.
show abstract
full text
BACKGROUND: Invasive lobular breast cancer (ILC) is the second most common type of breast cancer after invasive breast cancer of no special type (NST), representing up to 15% of all breast cancers. DESIGN: Latest data on ILC are presented, focusing on diagnosis, molecular make-up according to the European Society for Medical Oncology Scale for Clinical Actionability of molecular Targets (ESCAT) guidelines, treatment in the early and metastatic setting and ILC-focused clinical trials. RESULTS: At the imaging level, magnetic resonance imaging-based and novel positron emission tomography/computed tomography-based techniques can overcome the limitations of currently used imaging techniques for diagnosing ILC. At the pathology level, E-cadherin immunohistochemistry could help improving inter-pathologist agreement. The majority of patients with ILC do not seem to benefit as much from (neo-)adjuvant chemotherapy as patients with NST, although chemotherapy might be required in a subset of high-risk patients. No differences in treatment efficacy are seen for anti-human epidermal growth factor receptor 2 (HER2) therapies in the adjuvant setting and cyclin-dependent kinases 4 and 6 inhibitors in the metastatic setting. The clinical utility of the commercially available prognostic gene expression-based tests is unclear for patients with ILC. Several ESCAT alterations differ in frequency between ILC and NST. Germline BRCA1 and PALB2 alterations are less frequent in patients with ILC, while germline CDH1 (gene coding for E-cadherin) alterations are more frequent in patients with ILC. Somatic HER2 mutations are more frequent in ILC, especially in metastases (15% ILC versus 5% NST). A high tumour mutational burden, relevant for immune checkpoint inhibition, is more frequent in ILC metastases (16%) than in NST metastases (5%). Tumours with somatic inactivating CDH1 mutations may be vulnerable for treatment with ROS1 inhibitors, a concept currently investigated in early and metastatic ILC. CONCLUSION: ILC is a unique malignancy based on its pathological and biological features leading to differences in diagnosis as well as in treatment response, resistance and targets as compared to NST..
Baxter, J.S.
Zatreanu, D.
Pettitt, S.J.
Lord, C.J.
(2022). Resistance to DNA repair inhibitors in cancer. Mol oncol,
Vol.16
(21),
pp. 3811-3827.
show abstract
full text
The DNA damage response (DDR) represents a complex network of proteins which detect and repair DNA damage, thereby maintaining the integrity of the genome and preventing the transmission of mutations and rearranged chromosomes to daughter cells. Faults in the DDR are a known driver and hallmark of cancer. Furthermore, inhibition of DDR enzymes can be used to treat the disease. This is exemplified by PARP inhibitors (PARPi) used to treat cancers with defects in the homologous recombination DDR pathway. A series of novel DDR targets are now also under pre-clinical or clinical investigation, including inhibitors of ATR kinase, WRN helicase or the DNA polymerase/helicase Polθ (Pol-Theta). Drug resistance is a common phenomenon that impairs the overall effectiveness of cancer treatments and there is already some understanding of how resistance to PARPi occurs. Here, we discuss how an understanding of PARPi resistance could inform how resistance to new drugs targeting the DDR emerges. We also discuss potential strategies that could limit the impact of these therapy resistance mechanisms in cancer..
Lüscher, B.
Ahel, I.
Altmeyer, M.
Ashworth, A.
Bai, P.
Chang, P.
Cohen, M.
Corda, D.
Dantzer, F.
Daugherty, M.D.
Dawson, T.M.
Dawson, V.L.
Deindl, S.
Fehr, A.R.
Feijs, K.L.
Filippov, D.V.
Gagné, J.-.
Grimaldi, G.
Guettler, S.
Hoch, N.C.
Hottiger, M.O.
Korn, P.
Kraus, W.L.
Ladurner, A.
Lehtiö, L.
Leung, A.K.
Lord, C.J.
Mangerich, A.
Matic, I.
Matthews, J.
Moldovan, G.-.
Moss, J.
Natoli, G.
Nielsen, M.L.
Niepel, M.
Nolte, F.
Pascal, J.
Paschal, B.M.
Pawłowski, K.
Poirier, G.G.
Smith, S.
Timinszky, G.
Wang, Z.-.
Yélamos, J.
Yu, X.
Zaja, R.
Ziegler, M.
(2022). ADP-ribosyltransferases, an update on function and nomenclature. Febs j,
Vol.289
(23),
pp. 7399-7410.
show abstract
full text
ADP-ribosylation, a modification of proteins, nucleic acids, and metabolites, confers broad functions, including roles in stress responses elicited, for example, by DNA damage and viral infection and is involved in intra- and extracellular signaling, chromatin and transcriptional regulation, protein biosynthesis, and cell death. ADP-ribosylation is catalyzed by ADP-ribosyltransferases (ARTs), which transfer ADP-ribose from NAD+ onto substrates. The modification, which occurs as mono- or poly-ADP-ribosylation, is reversible due to the action of different ADP-ribosylhydrolases. Importantly, inhibitors of ARTs are approved or are being developed for clinical use. Moreover, ADP-ribosylhydrolases are being assessed as therapeutic targets, foremost as antiviral drugs and for oncological indications. Due to the development of novel reagents and major technological advances that allow the study of ADP-ribosylation in unprecedented detail, an increasing number of cellular processes and pathways are being identified that are regulated by ADP-ribosylation. In addition, characterization of biochemical and structural aspects of the ARTs and their catalytic activities have expanded our understanding of this protein family. This increased knowledge requires that a common nomenclature be used to describe the relevant enzymes. Therefore, in this viewpoint, we propose an updated and broadly supported nomenclature for mammalian ARTs that will facilitate future discussions when addressing the biochemistry and biology of ADP-ribosylation. This is combined with a brief description of the main functions of mammalian ARTs to illustrate the increasing diversity of mono- and poly-ADP-ribose mediated cellular processes..
Serra, V.
Wang, A.T.
Castroviejo-Bermejo, M.
Polanska, U.M.
Palafox, M.
Herencia-Ropero, A.
Jones, G.N.
Lai, Z.
Armenia, J.
Michopoulos, F.
Llop-Guevara, A.
Brough, R.
Gulati, A.
Pettitt, S.J.
Bulusu, K.C.
Nikkilä, J.
Wilson, Z.
Hughes, A.
Wijnhoven, P.W.
Ahmed, A.
Bruna, A.
Gris-Oliver, A.
Guzman, M.
Rodríguez, O.
Grueso, J.
Arribas, J.
Cortés, J.
Saura, C.
Lau, A.
Critchlow, S.
Dougherty, B.
Caldas, C.
Mills, G.B.
Barrett, J.C.
Forment, J.V.
Cadogan, E.
Lord, C.J.
Cruz, C.
Balmaña, J.
O'Connor, M.J.
(2022). Identification of a Molecularly-Defined Subset of Breast and Ovarian Cancer Models that Respond to WEE1 or ATR Inhibition, Overcoming PARP Inhibitor Resistance. Clin cancer res,
Vol.28
(20),
pp. 4536-4550.
show abstract
full text
PURPOSE: PARP inhibitors (PARPi) induce synthetic lethality in homologous recombination repair (HRR)-deficient tumors and are used to treat breast, ovarian, pancreatic, and prostate cancers. Multiple PARPi resistance mechanisms exist, most resulting in restoration of HRR and protection of stalled replication forks. ATR inhibition was highlighted as a unique approach to reverse both aspects of resistance. Recently, however, a PARPi/WEE1 inhibitor (WEE1i) combination demonstrated enhanced antitumor activity associated with the induction of replication stress, suggesting another approach to tackling PARPi resistance. EXPERIMENTAL DESIGN: We analyzed breast and ovarian patient-derived xenoimplant models resistant to PARPi to quantify WEE1i and ATR inhibitor (ATRi) responses as single agents and in combination with PARPi. Biomarker analysis was conducted at the genetic and protein level. Metabolite analysis by mass spectrometry and nucleoside rescue experiments ex vivo were also conducted in patient-derived models. RESULTS: Although WEE1i response was linked to markers of replication stress, including STK11/RB1 and phospho-RPA, ATRi response associated with ATM mutation. When combined with olaparib, WEE1i could be differentiated from the ATRi/olaparib combination, providing distinct therapeutic strategies to overcome PARPi resistance by targeting the replication stress response. Mechanistically, WEE1i sensitivity was associated with shortage of the dNTP pool and a concomitant increase in replication stress. CONCLUSIONS: Targeting the replication stress response is a valid therapeutic option to overcome PARPi resistance including tumors without an underlying HRR deficiency. These preclinical insights are now being tested in several clinical trials where the PARPi is administered with either the WEE1i or the ATRi..
Rouleau-Turcotte, É.
Krastev, D.B.
Pettitt, S.J.
Lord, C.J.
Pascal, J.M.
(2022). Captured snapshots of PARP1 in the active state reveal the mechanics of PARP1 allostery. Mol cell,
Vol.82
(16),
pp. 2939-2951.e5.
show abstract
full text
PARP1 rapidly detects DNA strand break damage and allosterically signals break detection to the PARP1 catalytic domain to activate poly(ADP-ribose) production from NAD+. PARP1 activation is characterized by dynamic changes in the structure of a regulatory helical domain (HD); yet, there are limited insights into the specific contributions that the HD makes to PARP1 allostery. Here, we have determined crystal structures of PARP1 in isolated active states that display specific HD conformations. These captured snapshots and biochemical analysis illustrate HD contributions to PARP1 multi-domain and high-affinity interaction with DNA damage, provide novel insights into the mechanics of PARP1 allostery, and indicate how HD active conformations correspond to alterations in the catalytic region that reveal the active site to NAD+. Our work deepens the understanding of PARP1 catalytic activation, the dynamics of the binding site of PARP inhibitor compounds, and the mechanisms regulating PARP1 retention on DNA damage..
Chabanon, R.M.
Morel, D.
Eychenne, T.
Colmet-Daage, L.
Bajrami, I.
Dorvault, N.
Garrido, M.
Meisenberg, C.
Lamb, A.
Ngo, C.
Hopkins, S.R.
Roumeliotis, T.I.
Jouny, S.
Hénon, C.
Kawai-Kawachi, A.
Astier, C.
Konde, A.
Del Nery, E.
Massard, C.
Pettitt, S.J.
Margueron, R.
Choudhary, J.S.
Almouzni, G.
Soria, J.-.
Deutsch, E.
Downs, J.A.
Lord, C.J.
Postel-Vinay, S.
(2021). PBRM1 Deficiency Confers Synthetic Lethality to DNA Repair Inhibitors in Cancer. Cancer res,
Vol.81
(11),
pp. 2888-2902.
show abstract
Inactivation of Polybromo 1 (PBRM1), a specific subunit of the PBAF chromatin remodeling complex, occurs frequently in cancer, including 40% of clear cell renal cell carcinomas (ccRCC). To identify novel therapeutic approaches to targeting PBRM1-defective cancers, we used a series of orthogonal functional genomic screens that identified PARP and ATR inhibitors as being synthetic lethal with PBRM1 deficiency. The PBRM1/PARP inhibitor synthetic lethality was recapitulated using several clinical PARP inhibitors in a series of in vitro model systems and in vivo in a xenograft model of ccRCC. In the absence of exogenous DNA damage, PBRM1-defective cells exhibited elevated levels of replication stress, micronuclei, and R-loops. PARP inhibitor exposure exacerbated these phenotypes. Quantitative mass spectrometry revealed that multiple R-loop processing factors were downregulated in PBRM1-defective tumor cells. Exogenous expression of the R-loop resolution enzyme RNase H1 reversed the sensitivity of PBRM1-deficient cells to PARP inhibitors, suggesting that excessive levels of R-loops could be a cause of this synthetic lethality. PARP and ATR inhibitors also induced cyclic GMP-AMP synthase/stimulator of interferon genes (cGAS/STING) innate immune signaling in PBRM1-defective tumor cells. Overall, these findings provide the preclinical basis for using PARP inhibitors in PBRM1-defective cancers. SIGNIFICANCE: This study demonstrates that PARP and ATR inhibitors are synthetic lethal with the loss of PBRM1, a PBAF-specific subunit, thus providing the rationale for assessing these inhibitors in patients with PBRM1-defective cancer. GRAPHICAL ABSTRACT: http://cancerres.aacrjournals.org/content/canres/81/11/2888/F1.large.jpg..
Baxter, J.S.
Johnson, N.
Tomczyk, K.
Gillespie, A.
Maguire, S.
Brough, R.
Fachal, L.
Michailidou, K.
Bolla, M.K.
Wang, Q.
Dennis, J.
Ahearn, T.U.
Andrulis, I.L.
Anton-Culver, H.
Antonenkova, N.N.
Arndt, V.
Aronson, K.J.
Augustinsson, A.
Becher, H.
Beckmann, M.W.
Behrens, S.
Benitez, J.
Bermisheva, M.
Bogdanova, N.V.
Bojesen, S.E.
Brenner, H.
Brucker, S.Y.
Cai, Q.
Campa, D.
Canzian, F.
Castelao, J.E.
Chan, T.L.
Chang-Claude, J.
Chanock, S.J.
Chenevix-Trench, G.
Choi, J.-.
Clarke, C.L.
NBCS Collaborators,
Colonna, S.
Conroy, D.M.
Couch, F.J.
Cox, A.
Cross, S.S.
Czene, K.
Daly, M.B.
Devilee, P.
Dörk, T.
Dossus, L.
Dwek, M.
Eccles, D.M.
Ekici, A.B.
Eliassen, A.H.
Engel, C.
Fasching, P.A.
Figueroa, J.
Flyger, H.
Gago-Dominguez, M.
Gao, C.
García-Closas, M.
García-Sáenz, J.A.
Ghoussaini, M.
Giles, G.G.
Goldberg, M.S.
González-Neira, A.
Guénel, P.
Gündert, M.
Haeberle, L.
Hahnen, E.
Haiman, C.A.
Hall, P.
Hamann, U.
Hartman, M.
Hatse, S.
Hauke, J.
Hollestelle, A.
Hoppe, R.
Hopper, J.L.
Hou, M.-.
kConFab Investigators,
ABCTB Investigators,
Ito, H.
Iwasaki, M.
Jager, A.
Jakubowska, A.
Janni, W.
John, E.M.
Joseph, V.
Jung, A.
Kaaks, R.
Kang, D.
Keeman, R.
Khusnutdinova, E.
Kim, S.-.
Kosma, V.-.
Kraft, P.
Kristensen, V.N.
Kubelka-Sabit, K.
Kurian, A.W.
Kwong, A.
Lacey, J.V.
Lambrechts, D.
Larson, N.L.
Larsson, S.C.
Le Marchand, L.
Lejbkowicz, F.
Li, J.
Long, J.
Lophatananon, A.
Lubiński, J.
Mannermaa, A.
Manoochehri, M.
Manoukian, S.
Margolin, S.
Matsuo, K.
Mavroudis, D.
Mayes, R.
Menon, U.
Milne, R.L.
Mohd Taib, N.A.
Muir, K.
Muranen, T.A.
Murphy, R.A.
Nevanlinna, H.
O'Brien, K.M.
Offit, K.
Olson, J.E.
Olsson, H.
Park, S.K.
Park-Simon, T.-.
Patel, A.V.
Peterlongo, P.
Peto, J.
Plaseska-Karanfilska, D.
Presneau, N.
Pylkäs, K.
Rack, B.
Rennert, G.
Romero, A.
Ruebner, M.
Rüdiger, T.
Saloustros, E.
Sandler, D.P.
Sawyer, E.J.
Schmidt, M.K.
Schmutzler, R.K.
Schneeweiss, A.
Schoemaker, M.J.
Shah, M.
Shen, C.-.
Shu, X.-.
Simard, J.
Southey, M.C.
Stone, J.
Surowy, H.
Swerdlow, A.J.
Tamimi, R.M.
Tapper, W.J.
Taylor, J.A.
Teo, S.H.
Teras, L.R.
Terry, M.B.
Toland, A.E.
Tomlinson, I.
Truong, T.
Tseng, C.-.
Untch, M.
Vachon, C.M.
van den Ouweland, A.M.
Wang, S.S.
Weinberg, C.R.
Wendt, C.
Winham, S.J.
Winqvist, R.
Wolk, A.
Wu, A.H.
Yamaji, T.
Zheng, W.
Ziogas, A.
Pharoah, P.D.
Dunning, A.M.
Easton, D.F.
Pettitt, S.J.
Lord, C.J.
Haider, S.
Orr, N.
Fletcher, O.
(2021). Functional annotation of the 2q35 breast cancer risk locus implicates a structural variant in influencing activity of a long-range enhancer element. Am j hum genet,
Vol.108
(7),
pp. 1190-1203.
show abstract
full text
A combination of genetic and functional approaches has identified three independent breast cancer risk loci at 2q35. A recent fine-scale mapping analysis to refine these associations resulted in 1 (signal 1), 5 (signal 2), and 42 (signal 3) credible causal variants at these loci. We used publicly available in silico DNase I and ChIP-seq data with in vitro reporter gene and CRISPR assays to annotate signals 2 and 3. We identified putative regulatory elements that enhanced cell-type-specific transcription from the IGFBP5 promoter at both signals (30- to 40-fold increased expression by the putative regulatory element at signal 2, 2- to 3-fold by the putative regulatory element at signal 3). We further identified one of the five credible causal variants at signal 2, a 1.4 kb deletion (esv3594306), as the likely causal variant; the deletion allele of this variant was associated with an average additional increase in IGFBP5 expression of 1.3-fold (MCF-7) and 2.2-fold (T-47D). We propose a model in which the deletion allele of esv3594306 juxtaposes two transcription factor binding regions (annotated by estrogen receptor alpha ChIP-seq peaks) to generate a single extended regulatory element. This regulatory element increases cell-type-specific expression of the tumor suppressor gene IGFBP5 and, thereby, reduces risk of estrogen receptor-positive breast cancer (odds ratio = 0.77, 95% CI 0.74-0.81, p = 3.1 × 10-31)..
Dreyer, S.B.
Upstill-Goddard, R.
Paulus-Hock, V.
Paris, C.
Lampraki, E.-.
Dray, E.
Serrels, B.
Caligiuri, G.
Rebus, S.
Plenker, D.
Galluzzo, Z.
Brunton, H.
Cunningham, R.
Tesson, M.
Nourse, C.
Bailey, U.-.
Jones, M.
Moran-Jones, K.
Wright, D.W.
Duthie, F.
Oien, K.
Evers, L.
McKay, C.J.
McGregor, G.A.
Gulati, A.
Brough, R.
Bajrami, I.
Pettitt, S.
Dziubinski, M.L.
Candido, J.
Balkwill, F.
Barry, S.T.
Grützmann, R.
Rahib, L.
Johns, A.
Pajic, M.
Froeling, F.E.
Beer, P.
Musgrove, E.A.
Petersen, G.M.
Ashworth, A.
Frame, M.C.
Crawford, H.C.
Simeone, D.M.
Lord, C.
Mukhopadhyay, D.
Pilarsky, C.
Tuveson, D.A.
Cooke, S.L.
Jamieson, N.B.
Morton, J.P.
Sansom, O.J.
Bailey, P.J.
Biankin, A.V.
Chang, D.K.
Allison, S.
Bailey, P.J.
Bailey, U.-.
Biankin, A.V.
Beraldi, D.
Brunton, H.
Caligiuri, G.
Cameron, E.
Chang, D.K.
Cooke, S.L.
Cunningham, R.
Dreyer, S.
Grimwood, P.
Kelly, S.
Lampraki, E.-.
Marshall, J.
Martin, S.
McDade, B.
McElroy, D.
Musgrove, E.A.
Nourse, C.
Paulus-Hock, V.
Ramsay, D.
Upstill-Goddard, R.
Wright, D.
Jones, M.D.
Evers, L.
Rebus, S.
Rahib, L.
Serrels, B.
Hair, J.
Jamieson, N.B.
McKay, C.J.
Westwood, P.
Williams, N.
Duthie, F.
Biankin, A.V.
Johns, A.L.
Mawson, A.
Chang, D.K.
Scarlett, C.J.
Brancato, M.-.
Rowe, S.J.
Simpson, S.H.
Martyn-Smith, M.
Thomas, M.T.
Chantrill, L.A.
Chin, V.T.
Chou, A.
Cowley, M.J.
Humphris, J.L.
Jones, M.D.
Mead, R.S.
Nagrial, A.M.
Pajic, M.
Pettit, J.
Pinese, M.
Rooman, I.
Wu, J.
Tao, J.
DiPietro, R.
Watson, C.
Steinmann, A.
Lee, H.C.
Wong, R.
Pinho, A.V.
Giry-Laterriere, M.
Daly, R.J.
Musgrove, E.A.
Sutherland, R.L.
Grimmond, S.M.
Waddell, N.
Kassahn, K.S.
Miller, D.K.
Wilson, P.J.
Patch, A.-.
Song, S.
Harliwong, I.
Idrisoglu, S.
Nourse, C.
Nourbakhsh, E.
Manning, S.
Wani, S.
Gongora, M.
Anderson, M.
Holmes, O.
Leonard, C.
Taylor, D.
Wood, S.
Xu, C.
Nones, K.
Fink, J.L.
Christ, A.
Bruxner, T.
Cloonan, N.
Newell, F.
Pearson, J.V.
Bailey, P.
Quinn, M.
Nagaraj, S.
Kazakoff, S.
Waddell, N.
Krisnan, K.
Quek, K.
Wood, D.
Samra, J.S.
Gill, A.J.
Pavlakis, N.
Guminski, A.
Toon, C.
Asghari, R.
Merrett, N.D.
Pavey, D.
Das, A.
Cosman, P.H.
Ismail, K.
O’Connnor, C.
Lam, V.W.
McLeod, D.
Pleass, H.C.
Richardson, A.
James, V.
Kench, J.G.
Cooper, C.L.
Joseph, D.
Sandroussi, C.
Crawford, M.
Gallagher, J.
Texler, M.
Forest, C.
Laycock, A.
Epari, K.P.
Ballal, M.
Fletcher, D.R.
Mukhedkar, S.
Spry, N.A.
DeBoer, B.
Chai, M.
Zeps, N.
Beilin, M.
Feeney, K.
Nguyen, N.Q.
Ruszkiewicz, A.R.
Worthley, C.
Tan, C.P.
Debrencini, T.
Chen, J.
Brooke-Smith, M.E.
Papangelis, V.
Tang, H.
Barbour, A.P.
Clouston, A.D.
Martin, P.
O’Rourke, T.J.
Chiang, A.
Fawcett, J.W.
Slater, K.
Yeung, S.
Hatzifotis, M.
Hodgkinson, P.
Christophi, C.
Nikfarjam, M.
Mountain, A.
Biobank, V.C.
Eshleman, J.R.
Hruban, R.H.
Maitra, A.
Iacobuzio-Donahue, C.A.
Schulick, R.D.
Wolfgang, C.L.
Morgan, R.A.
Hodgin, M.
Scarpa, A.
Lawlor, R.T.
Beghelli, S.
Corbo, V.
Scardoni, M.
Bassi, C.
Tempero, M.A.
Biankin, A.V.
Grimmond, S.M.
Chang, D.K.
Musgrove, E.A.
Jones, M.D.
Nourse, C.
Jamieson, N.B.
Graham, J.S.
Biankin, A.V.
Chang, D.K.
Jamieson, N.B.
Graham, J.S.
(2021). Targeting DNA Damage Response and Replication Stress in Pancreatic Cancer. Gastroenterology,
Vol.160
(1),
pp. 362-377.e13.
Parkhitko, A.A.
Singh, A.
Hsieh, S.
Hu, Y.
Binari, R.
Lord, C.J.
Hannenhalli, S.
Ryan, C.J.
Perrimon, N.
(2021). Cross-species identification of PIP5K1-, splicing- and ubiquitin-related pathways as potential targets for RB1-deficient cells. Plos genet,
Vol.17
(2),
p. e1009354.
show abstract
full text
The RB1 tumor suppressor is recurrently mutated in a variety of cancers including retinoblastomas, small cell lung cancers, triple-negative breast cancers, prostate cancers, and osteosarcomas. Finding new synthetic lethal (SL) interactions with RB1 could lead to new approaches to treating cancers with inactivated RB1. We identified 95 SL partners of RB1 based on a Drosophila screen for genetic modifiers of the eye phenotype caused by defects in the RB1 ortholog, Rbf1. We validated 38 mammalian orthologs of Rbf1 modifiers as RB1 SL partners in human cancer cell lines with defective RB1 alleles. We further show that for many of the RB1 SL genes validated in human cancer cell lines, low activity of the SL gene in human tumors, when concurrent with low levels of RB1 was associated with improved patient survival. We investigated higher order combinatorial gene interactions by creating a novel Drosophila cancer model with co-occurring Rbf1, Pten and Ras mutations, and found that targeting RB1 SL genes in this background suppressed the dramatic tumor growth and rescued fly survival whilst having minimal effects on wild-type cells. Finally, we found that drugs targeting the identified RB1 interacting genes/pathways, such as UNC3230, PYR-41, TAK-243, isoginkgetin, madrasin, and celastrol also elicit SL in human cancer cell lines. In summary, we identified several high confidence, evolutionarily conserved, novel targets for RB1-deficient cells that may be further adapted for the treatment of human cancer..
Neeb, A.
Herranz, N.
Arce-Gallego, S.
Miranda, S.
Buroni, L.
Yuan, W.
Athie, A.
Casals, T.
Carmichael, J.
Rodrigues, D.N.
Gurel, B.
Rescigno, P.
Rekowski, J.
Welti, J.
Riisnaes, R.
Gil, V.
Ning, J.
Wagner, V.
Casanova-Salas, I.
Cordoba, S.
Castro, N.
Fenor de la Maza, M.D.
Seed, G.
Chandran, K.
Ferreira, A.
Figueiredo, I.
Bertan, C.
Bianchini, D.
Aversa, C.
Paschalis, A.
Gonzalez, M.
Morales-Barrera, R.
Suarez, C.
Carles, J.
Swain, A.
Sharp, A.
Gil, J.
Serra, V.
Lord, C.
Carreira, S.
Mateo, J.
de Bono, J.S.
(2021). Advanced Prostate Cancer with ATM Loss: PARP and ATR Inhibitors. Eur urol,
Vol.79
(2),
pp. 200-211.
show abstract
full text
BACKGROUND: Deleterious ATM alterations are found in metastatic prostate cancer (PC); PARP inhibition has antitumour activity against this subset, but only some ATM loss PCs respond. OBJECTIVE: To characterise ATM-deficient lethal PC and to study synthetic lethal therapeutic strategies for this subset. DESIGN, SETTING, AND PARTICIPANTS: We studied advanced PC biopsies using validated immunohistochemical (IHC) and next-generation sequencing (NGS) assays. In vitro cell line models modified using CRISPR-Cas9 to impair ATM function were generated and used in drug-sensitivity and functional assays, with validation in a patient-derived model. OUTCOME MEASUREMENTS AND STATISTICAL ANALYSIS: ATM expression by IHC was correlated with clinical outcome using Kaplan-Meier curves and log-rank test; sensitivity to different drug combinations was assessed in the preclinical models. RESULTS AND LIMITATIONS: Overall, we detected ATM IHC loss in 68/631 (11%) PC patients in at least one biopsy, with synchronous and metachronous intrapatient heterogeneity; 46/71 (65%) biopsies with ATM loss had ATM mutations or deletions by NGS. ATM IHC loss was not associated with worse outcome from advanced disease, but ATM loss was associated with increased genomic instability (NtAI:number of subchromosomal regions with allelic imbalance extending to the telomere, p = 0.005; large-scale transitions, p = 0.05). In vitro, ATM loss PC models were sensitive to ATR inhibition, but had variable sensitivity to PARP inhibition; superior antitumour activity was seen with combined PARP and ATR inhibition in these models. CONCLUSIONS: ATM loss in PC is not always detected by targeted NGS, associates with genomic instability, and is most sensitive to combined ATR and PARP inhibition. PATIENT SUMMARY: Of aggressive prostate cancers, 10% lose the DNA repair gene ATM; this loss may identify a distinct prostate cancer subtype that is most sensitive to the combination of oral drugs targeting PARP and ATR..
Bajrami, I.
Walker, C.
Krastev, D.B.
Weekes, D.
Song, F.
Wicks, A.J.
Alexander, J.
Haider, S.
Brough, R.
Pettitt, S.J.
Tutt, A.N.
Lord, C.J.
(2021). Sirtuin inhibition is synthetic lethal with BRCA1 or BRCA2 deficiency. Commun biol,
Vol.4
(1),
p. 1270.
show abstract
full text
PARP enzymes utilise NAD+ as a co-substrate for their enzymatic activity. Inhibition of PARP1 is synthetic lethal with defects in either BRCA1 or BRCA2. In order to assess whether other genes implicated in NAD+ metabolism were synthetic lethal with BRCA1 or BRCA2 gene defects, we carried out a genetic screen, which identified a synthetic lethality between BRCA1 and genetic inhibition of either of two sirtuin (SIRT) enzymes, SIRT1 or SIRT6. This synthetic lethal interaction was replicated using small-molecule SIRT inhibitors and was associated with replication stress and increased cellular PARylation, in contrast to the decreased PARylation associated with BRCA-gene/PARP inhibitor synthetic lethality. SIRT/BRCA1 synthetic lethality was reversed by genetic ablation of either PARP1 or the histone PARylation factor-coding gene HPF1, implicating PARP1/HPF1-mediated serine ADP-ribosylation as part of the mechanistic basis of this synthetic lethal effect. These observations suggest that PARP1/HPF1-mediated serine ADP-ribosylation, when driven by SIRT inhibition, can inadvertently inhibit the growth of BRCA-gene mutant cells..
Sumanasuriya, S.
Seed, G.
Parr, H.
Christova, R.
Pope, L.
Bertan, C.
Bianchini, D.
Rescigno, P.
Figueiredo, I.
Goodall, J.
Fowler, G.
Flohr, P.
Mehra, N.
Neeb, A.
Rekowski, J.
Eisenberger, M.
Sartor, O.
Oudard, S.
Geffriaud-Ricouard, C.
Ozatilgan, A.
Chadjaa, M.
Macé, S.
Lord, C.
Baxter, J.
Pettitt, S.
Lambros, M.
Sharp, A.
Mateo, J.
Carreira, S.
Yuan, W.
de Bono, J.S.
(2021). Elucidating Prostate Cancer Behaviour During Treatment via Low-pass Whole-genome Sequencing of Circulating Tumour DNA. Eur urol,
Vol.80
(2),
pp. 243-253.
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BACKGROUND: Better blood tests to elucidate the behaviour of metastatic castration-resistant prostate cancer (mCRPC) are urgently needed to drive therapeutic decisions. Plasma cell-free DNA (cfDNA) comprises normal and circulating tumour DNA (ctDNA). Low-pass whole-genome sequencing (lpWGS) of ctDNA can provide information on mCRPC behaviour. OBJECTIVE: To validate and clinically qualify plasma lpWGS for mCRPC. DESIGN, SETTING, AND PARTICIPANTS: Plasma lpWGS data were obtained for mCRPC patients consenting to optional substudies of two prospective phase 3 trials (FIRSTANA and PROSELICA). In FIRSTANA, chemotherapy-naïve patients were randomised to treatment with docetaxel (75 mg/m2) or cabazitaxel (20 or 25 mg/m2). In PROSELICA, patients previously treated with docetaxel were randomised to 20 or 25 mg/m2 cabazitaxel. lpWGS data were generated from 540 samples from 188 mCRPC patients acquired at four different time points (screening, cycle 1, cycle 4, and end of study). OUTCOME MEASUREMENTS AND STATISTICAL ANALYSIS: lpWGS data for ctDNA were evaluated for prognostic, response, and tumour genomic measures. Associations with response and survival data were determined for tumour fraction. Genomic biomarkers including large-scale transition (LST) scores were explored in the context of prior treatments. RESULTS AND LIMITATIONS: Plasma tumour fraction was prognostic for overall survival in univariable and stratified multivariable analyses (hazard ratio 1.75, 95% confidence interval 1.08-2.85; p = 0.024) and offered added value compared to existing biomarkers (C index 0.722 vs 0.709; p = 0.021). Longitudinal changes were associated with drug response. PROSELICA samples were enriched for LSTs (p = 0.029) indicating genomic instability, and this enrichment was associated with prior abiraterone and enzalutamide treatment but not taxane or radiation therapy. Higher LSTs were correlated with losses of RB1/RNASEH2B, independent of BRCA2 loss. CONCLUSIONS: Plasma lpWGS of ctDNA describes CRPC behaviour, providing prognostic and response data of clinical relevance. The added prognostic value of the ctDNA fraction over established biomarkers should be studied further. PATIENT SUMMARY: We studied tumour DNA in blood samples from patients with prostate cancer. We found that levels of tumour DNA in blood were indicative of disease prognosis, and that changes after treatment could be detected. We also observed a "genetic scar" in the results that was associated with certain previous treatments. This test allows an assessment of tumour activity that can complement existing tests, offer insights into drug response, and detect clinically relevant genetic changes..
Chong, I.Y.
Starling, N.
Rust, A.
Alexander, J.
Aronson, L.
Llorca-Cardenosa, M.
Chauhan, R.
Chaudry, A.
Kumar, S.
Fenwick, K.
Assiotis, I.
Matthews, N.
Begum, R.
Wotherspoon, A.
Terlizzo, M.
Watkins, D.
Chau, I.
Lord, C.J.
Haider, S.
Rao, S.
Cunningham, D.
(2021). The Mutational Concordance of Fixed Formalin Paraffin Embedded and Fresh Frozen Gastro-Oesophageal Tumours Using Whole Exome Sequencing. J clin med,
Vol.10
(2).
show abstract
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UNLABELLED: 1. BACKGROUND: The application of massively parallel sequencing has led to the identification of aberrant druggable pathways and somatic mutations within therapeutically relevant genes in gastro-oesophageal cancer. Given the widespread use of formalin-fixed paraffin-embedded (FFPE) samples in the study of this disease, it would be beneficial, especially for the purposes of biomarker evaluation, to assess the concordance between comprehensive exome-wide sequencing data from archival FFPE samples originating from a prospective clinical study and those derived from fresh-frozen material. 2. METHODS: We analysed whole-exome sequencing data to define the mutational concordance of 16 matched fresh-frozen and FFPE gastro-oesophageal tumours (N = 32) from a prospective clinical study. We assessed DNA integrity prior to sequencing and then identified coding mutations in genes that have previously been implicated in other cancers. In addition, we calculated the mutant-allele heterogeneity (MATH) for these samples. 3. RESULTS: Although there was increased degradation of DNA in FFPE samples compared with frozen samples, sequencing data from only two FFPE samples failed to reach an adequate mapping quality threshold. Using a filtering threshold of mutant read counts of at least ten and a minimum of 5% variant allele frequency (VAF) we found that there was a high median mutational concordance of 97% (range 80.1-98.68%) between fresh-frozen and FFPE gastro-oesophageal tumour-derived exomes. However, the majority of FFPE tumours had higher mutant-allele heterogeneity (MATH) scores when compared with corresponding frozen tumours (p < 0.001), suggesting that FFPE-based exome sequencing is likely to over-represent tumour heterogeneity in FFPE samples compared to fresh-frozen samples. Furthermore, we identified coding mutations in 120 cancer-related genes, including those associated with chromatin remodelling and Wnt/β-catenin and Receptor Tyrosine Kinase signalling. 4. CONCLUSIONS: These data suggest that comprehensive genomic data can be generated from exome sequencing of selected DNA samples extracted from archival FFPE gastro-oesophageal tumour tissues within the context of prospective clinical trials..
Carreira, S.
Porta, N.
Arce-Gallego, S.
Seed, G.
Llop-Guevara, A.
Bianchini, D.
Rescigno, P.
Paschalis, A.
Bertan, C.
Baker, C.
Goodall, J.
Miranda, S.
Riisnaes, R.
Figueiredo, I.
Ferreira, A.
Pereira, R.
Crespo, M.
Gurel, B.
Nava Rodrigues, D.
Pettitt, S.J.
Yuan, W.
Serra, V.
Rekowski, J.
Lord, C.J.
Hall, E.
Mateo, J.
de Bono, J.S.
(2021). Biomarkers Associating with PARP Inhibitor Benefit in Prostate Cancer in the TOPARP-B Trial. Cancer discov,
Vol.11
(11),
pp. 2812-2827.
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PARP inhibitors are approved for treating advanced prostate cancers (APC) with various defective DNA repair genes; however, further studies to clinically qualify predictive biomarkers are warranted. Herein we analyzed TOPARP-B phase II clinical trial samples, evaluating whole-exome and low-pass whole-genome sequencing and IHC and IF assays evaluating ATM and RAD51 foci (testing homologous recombination repair function). BRCA1/2 germline and somatic pathogenic mutations associated with similar benefit from olaparib; greater benefit was observed with homozygous BRCA2 deletion. Biallelic, but not monoallelic, PALB2 deleterious alterations were associated with clinical benefit. In the ATM cohort, loss of ATM protein by IHC was associated with a better outcome. RAD51 foci loss identified tumors with biallelic BRCA and PALB2 alterations while most ATM- and CDK12-altered APCs had higher RAD51 foci levels. Overall, APCs with homozygous BRCA2 deletion are exceptional responders; PALB2 biallelic loss and loss of ATM IHC expression associated with clinical benefit. SIGNIFICANCE: Not all APCs with DNA repair defects derive similar benefit from PARP inhibition. Most benefit was seen among patients with BRCA2 homozygous deletions, biallelic loss of PALB2, and loss of ATM protein. Loss of RAD51 foci, evaluating homologous recombination repair function, was found primarily in tumors with biallelic BRCA1/2 and PALB2 alterations.This article is highlighted in the In This Issue feature, p. 2659..
Chabanon, R.M.
Rouanne, M.
Lord, C.J.
Soria, J.-.
Pasero, P.
Postel-Vinay, S.
(2021). Targeting the DNA damage response in immuno-oncology: developments and opportunities. Nat rev cancer,
Vol.21
(11),
pp. 701-717.
show abstract
Immunotherapy has revolutionized cancer treatment and substantially improved patient outcome with regard to multiple tumour types. However, most patients still do not benefit from such therapies, notably because of the absence of pre-existing T cell infiltration. DNA damage response (DDR) deficiency has recently emerged as an important determinant of tumour immunogenicity. A growing body of evidence now supports the concept that DDR-targeted therapies can increase the antitumour immune response by (1) promoting antigenicity through increased mutability and genomic instability, (2) enhancing adjuvanticity through the activation of cytosolic immunity and immunogenic cell death and (3) favouring reactogenicity through the modulation of factors that control the tumour-immune cell synapse. In this Review, we discuss the interplay between the DDR and anticancer immunity and highlight how this dynamic interaction contributes to shaping tumour immunogenicity. We also review the most innovative preclinical approaches that could be used to investigate such effects, including recently developed ex vivo systems. Finally, we highlight the therapeutic opportunities presented by the exploitation of the DDR-anticancer immunity interplay, with a focus on those in early-phase clinical development..
Banerjee, S.
Stewart, J.
Porta, N.
Toms, C.
Leary, A.
Lheureux, S.
Khalique, S.
Tai, J.
Attygalle, A.
Vroobel, K.
Lord, C.J.
Natrajan, R.
Bliss, J.
(2021). ATARI trial: ATR inhibitor in combination with olaparib in gynecological cancers with ARID1A loss or no loss (ENGOT/GYN1/NCRI). Int j gynecol cancer,
Vol.31
(11),
pp. 1471-1475.
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BACKGROUND: ARID1A (AT-rich interactive domain containing protein 1A) loss-of-function mutations have been reported in gynecological cancers, including rarer subtypes such as clear cell carcinoma. Preclinical studies indicate that ARID1A mutant cancers display sensitivity to ATR inhibition while tumors without ARID1A mutations may be sensitive to Ataxia telangiectasia and Rad3 related (ATR) inhibitors in combination with poly-ADP ribose polymerase (PARP) inhibitors. PRIMARY OBJECTIVE: To determine whether the ATR inhibitor, ceralasertib, has clinical activity as a single agent and in combination with the PARP inhibitor, olaparib, in patients with ARID1A 'loss' and 'no loss' clear cell carcinomas and other relapsed gynecological cancers. STUDY HYPOTHESIS: ARID1A deficient clear cell carcinoma of the ovary or endometrium is sensitive to ATR inhibition, while the combination of ATR and PARP inhibition has activity in other gynecological tumors, irrespective of ARID1A status. TRIAL DESIGN: ATARI (ENGOT/GYN1/NCRI) is a multicenter, international, proof-of-concept, phase II, parallel cohort trial assessing ceralasertib activity as a single agent and in combination with olaparib in ARID1A stratified gynecological cancers. Patients with relapsed ovarian/endometrial clear cell carcinoma with ARID1A loss will receive ceralasertib monotherapy (cohort 1A). Relapsed ovarian/endometrial clear cell carcinoma patients with no ARID1A loss (cohort 2) or patients with other histological subtypes (endometrioid, carcinosarcoma, cervical) (cohort 3) will receive combination therapy (olaparib/ceralasertib). Treatment will continue until disease progression. MAJOR INCLUSION/EXCLUSION CRITERIA: Patients with histologically confirmed recurrent clear cell (ovarian, endometrial, or endometriosis related), endometrioid (ovarian, endometrial, or endometriosis related), cervical (adenocarcinomas or squamous), or carcinosarcomas (ovarian or endometrial) are eligible. Patients progressing after ≥1 prior platinum with evidence of measurable (RECIST v1.1) radiological disease progression since last systemic anticancer therapy and prior to trial entry are eligible. Previous ATR or PARP inhibitor treatment is not permissible. PRIMARY ENDPOINT: Best overall objective response rate (RECIST v1.1). SAMPLE SIZE: A minimum of 40 and a maximum of 116. ESTIMATED DATES FOR COMPLETING ACCRUAL AND PRESENTING RESULTS: Accrual is anticipated to be complete by the second quarter of 2022, with reporting of results by the fourth quarter of 2022. Overall accrual targets and reporting timelines are dependent on individual cohort progression to stage 2. TRIAL REGISTRATION NUMBER: NCT0405269..
Zatreanu, D.
Robinson, H.M.
Alkhatib, O.
Boursier, M.
Finch, H.
Geo, L.
Grande, D.
Grinkevich, V.
Heald, R.A.
Langdon, S.
Majithiya, J.
McWhirter, C.
Martin, N.M.
Moore, S.
Neves, J.
Rajendra, E.
Ranzani, M.
Schaedler, T.
Stockley, M.
Wiggins, K.
Brough, R.
Sridhar, S.
Gulati, A.
Shao, N.
Badder, L.M.
Novo, D.
Knight, E.G.
Marlow, R.
Haider, S.
Callen, E.
Hewitt, G.
Schimmel, J.
Prevo, R.
Alli, C.
Ferdinand, A.
Bell, C.
Blencowe, P.
Bot, C.
Calder, M.
Charles, M.
Curry, J.
Ekwuru, T.
Ewings, K.
Krajewski, W.
MacDonald, E.
McCarron, H.
Pang, L.
Pedder, C.
Rigoreau, L.
Swarbrick, M.
Wheatley, E.
Willis, S.
Wong, A.C.
Nussenzweig, A.
Tijsterman, M.
Tutt, A.
Boulton, S.J.
Higgins, G.S.
Pettitt, S.J.
Smith, G.C.
Lord, C.J.
(2021). Polθ inhibitors elicit BRCA-gene synthetic lethality and target PARP inhibitor resistance. Nat commun,
Vol.12
(1),
p. 3636.
show abstract
full text
To identify approaches to target DNA repair vulnerabilities in cancer, we discovered nanomolar potent, selective, low molecular weight (MW), allosteric inhibitors of the polymerase function of DNA polymerase Polθ, including ART558. ART558 inhibits the major Polθ-mediated DNA repair process, Theta-Mediated End Joining, without targeting Non-Homologous End Joining. In addition, ART558 elicits DNA damage and synthetic lethality in BRCA1- or BRCA2-mutant tumour cells and enhances the effects of a PARP inhibitor. Genetic perturbation screening revealed that defects in the 53BP1/Shieldin complex, which cause PARP inhibitor resistance, result in in vitro and in vivo sensitivity to small molecule Polθ polymerase inhibitors. Mechanistically, ART558 increases biomarkers of single-stranded DNA and synthetic lethality in 53BP1-defective cells whilst the inhibition of DNA nucleases that promote end-resection reversed these effects, implicating these in the synthetic lethal mechanism-of-action. Taken together, these observations describe a drug class that elicits BRCA-gene synthetic lethality and PARP inhibitor synergy, as well as targeting a biomarker-defined mechanism of PARPi-resistance..
Hewitt, G.
Borel, V.
Segura-Bayona, S.
Takaki, T.
Ruis, P.
Bellelli, R.
Lehmann, L.C.
Sommerova, L.
Vancevska, A.
Tomas-Loba, A.
Zhu, K.
Cooper, C.
Fugger, K.
Patel, H.
Goldstone, R.
Schneider-Luftman, D.
Herbert, E.
Stamp, G.
Brough, R.
Pettitt, S.
Lord, C.J.
West, S.C.
Ahel, I.
Ahel, D.
Chapman, J.R.
Deindl, S.
Boulton, S.J.
(2021). Defective ALC1 nucleosome remodeling confers PARPi sensitization and synthetic lethality with HRD. Mol cell,
Vol.81
(4),
pp. 767-783.e11.
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Chromatin is a barrier to efficient DNA repair, as it hinders access and processing of certain DNA lesions. ALC1/CHD1L is a nucleosome-remodeling enzyme that responds to DNA damage, but its precise function in DNA repair remains unknown. Here we report that loss of ALC1 confers sensitivity to PARP inhibitors, methyl-methanesulfonate, and uracil misincorporation, which reflects the need to remodel nucleosomes following base excision by DNA glycosylases but prior to handover to APEX1. Using CRISPR screens, we establish that ALC1 loss is synthetic lethal with homologous recombination deficiency (HRD), which we attribute to chromosome instability caused by unrepaired DNA gaps at replication forks. In the absence of ALC1 or APEX1, incomplete processing of BER intermediates results in post-replicative DNA gaps and a critical dependence on HR for repair. Hence, targeting ALC1 alone or as a PARP inhibitor sensitizer could be employed to augment existing therapeutic strategies for HRD cancers..
Tutt, A.N.
Garber, J.E.
Kaufman, B.
Viale, G.
Fumagalli, D.
Rastogi, P.
Gelber, R.D.
de Azambuja, E.
Fielding, A.
Balmaña, J.
Domchek, S.M.
Gelmon, K.A.
Hollingsworth, S.J.
Korde, L.A.
Linderholm, B.
Bandos, H.
Senkus, E.
Suga, J.M.
Shao, Z.
Pippas, A.W.
Nowecki, Z.
Huzarski, T.
Ganz, P.A.
Lucas, P.C.
Baker, N.
Loibl, S.
McConnell, R.
Piccart, M.
Schmutzler, R.
Steger, G.G.
Costantino, J.P.
Arahmani, A.
Wolmark, N.
McFadden, E.
Karantza, V.
Lakhani, S.R.
Yothers, G.
Campbell, C.
Geyer, C.E.
OlympiA Clinical Trial Steering Committee and Investigators,
(2021). Adjuvant Olaparib for Patients with BRCA1- or BRCA2-Mutated Breast Cancer. N engl j med,
Vol.384
(25),
pp. 2394-2405.
show abstract
full text
BACKGROUND: Poly(adenosine diphosphate-ribose) polymerase inhibitors target cancers with defects in homologous recombination repair by synthetic lethality. New therapies are needed to reduce recurrence in patients with BRCA1 or BRCA2 germline mutation-associated early breast cancer. METHODS: We conducted a phase 3, double-blind, randomized trial involving patients with human epidermal growth factor receptor 2 (HER2)-negative early breast cancer with BRCA1 or BRCA2 germline pathogenic or likely pathogenic variants and high-risk clinicopathological factors who had received local treatment and neoadjuvant or adjuvant chemotherapy. Patients were randomly assigned (in a 1:1 ratio) to 1 year of oral olaparib or placebo. The primary end point was invasive disease-free survival. RESULTS: A total of 1836 patients underwent randomization. At a prespecified event-driven interim analysis with a median follow-up of 2.5 years, the 3-year invasive disease-free survival was 85.9% in the olaparib group and 77.1% in the placebo group (difference, 8.8 percentage points; 95% confidence interval [CI], 4.5 to 13.0; hazard ratio for invasive disease or death, 0.58; 99.5% CI, 0.41 to 0.82; P<0.001). The 3-year distant disease-free survival was 87.5% in the olaparib group and 80.4% in the placebo group (difference, 7.1 percentage points; 95% CI, 3.0 to 11.1; hazard ratio for distant disease or death, 0.57; 99.5% CI, 0.39 to 0.83; P<0.001). Olaparib was associated with fewer deaths than placebo (59 and 86, respectively) (hazard ratio, 0.68; 99% CI, 0.44 to 1.05; P = 0.02); however, the between-group difference was not significant at an interim-analysis boundary of a P value of less than 0.01. Safety data were consistent with known side effects of olaparib, with no excess serious adverse events or adverse events of special interest. CONCLUSIONS: Among patients with high-risk, HER2-negative early breast cancer and germline BRCA1 or BRCA2 pathogenic or likely pathogenic variants, adjuvant olaparib after completion of local treatment and neoadjuvant or adjuvant chemotherapy was associated with significantly longer survival free of invasive or distant disease than was placebo. Olaparib had limited effects on global patient-reported quality of life. (Funded by the National Cancer Institute and AstraZeneca; OlympiA ClinicalTrials.gov number, NCT02032823.)..
Khalique, S.
Nash, S.
Mansfield, D.
Wampfler, J.
Attygale, A.
Vroobel, K.
Kemp, H.
Buus, R.
Cottom, H.
Roxanis, I.
Jones, T.
von Loga, K.
Begum, D.
Guppy, N.
Ramagiri, P.
Fenwick, K.
Matthews, N.
Hubank, M.J.
Lord, C.J.
Haider, S.
Melcher, A.
Banerjee, S.
Natrajan, R.
(2021). Quantitative Assessment and Prognostic Associations of the Immune Landscape in Ovarian Clear Cell Carcinoma. Cancers (basel),
Vol.13
(15).
show abstract
full text
Ovarian clear cell carcinoma (OCCC) is a rare subtype of epithelial ovarian cancer characterised by a high frequency of loss-of-function ARID1A mutations and a poor response to chemotherapy. Despite their generally low mutational burden, an intratumoural T cell response has been reported in a subset of OCCC, with ARID1A purported to be a biomarker for the response to the immune checkpoint blockade independent of micro-satellite instability (MSI). However, assessment of the different immune cell types and spatial distribution specifically within OCCC patients has not been described to date. Here, we characterised the immune landscape of OCCC by profiling a cohort of 33 microsatellite stable OCCCs at the genomic, gene expression and histological level using targeted sequencing, gene expression profiling using the NanoString targeted immune panel, and multiplex immunofluorescence to assess the spatial distribution and abundance of immune cell populations at the protein level. Analysis of these tumours and subsequent independent validation identified an immune-related gene expression signature associated with risk of recurrence of OCCC. Whilst histological quantification of tumour-infiltrating lymphocytes (TIL, Salgado scoring) showed no association with the risk of recurrence or ARID1A mutational status, the characterisation of TILs via multiplexed immunofluorescence identified spatial differences in immunosuppressive cell populations in OCCC. Tumour-associated macrophages (TAM) and regulatory T cells were excluded from the vicinity of tumour cells in low-risk patients, suggesting that high-risk patients have a more immunosuppressive microenvironment. We also found that TAMs and cytotoxic T cells were also excluded from the vicinity of tumour cells in ARID1A-mutated OCCCs compared to ARID1A wild-type tumours, suggesting that the exclusion of these immune effectors could determine the host response of ARID1A-mutant OCCCs to therapy. Overall, our study has provided new insights into the immune landscape and prognostic associations in OCCC and suggest that tailored immunotherapeutic approaches may be warranted for different subgroups of OCCC patients..
Khalique, S.
Pettitt, S.J.
Kelly, G.
Tunariu, N.
Natrajan, R.
Banerjee, S.
Lord, C.J.
(2020). Longitudinal analysis of a secondary BRCA2 mutation using digital droplet PCR. J pathol clin res,
Vol.6
(1),
pp. 3-11.
show abstract
full text
Development of resistance to platinum and poly(ADP-ribose) polymerase inhibitors via secondary BRCA gene mutations that restore functional homologous recombination has been observed in a number of cancer types. Here we report a case of somatic BRCA2 mutation in a patient with high grade serous ovarian carcinoma. A secondary mutation predicted to restore the BRCA2 open reading frame was detected at low frequency (2.3%) in whole exome sequencing of a peritoneal biopsy at disease progression after treatment that included carboplatin and olaparib. We used digital droplet PCR (ddPCR) to verify the presence and frequency of this mutation in the biopsy sample at progression and also used this approach to assess the presence of the secondary mutation in preceding biopsies at diagnosis and first relapse. We found no evidence for the secondary mutation being present prior to the final progression biopsy, suggesting that this mutation was acquired late in the course of treatment. ddPCR provides a sensitive and specific technique to investigate the presence of low frequency mutations in a time series of biopsies..
Zandarashvili, L.
Langelier, M.-.
Velagapudi, U.K.
Hancock, M.A.
Steffen, J.D.
Billur, R.
Hannan, Z.M.
Wicks, A.J.
Krastev, D.B.
Pettitt, S.J.
Lord, C.J.
Talele, T.T.
Pascal, J.M.
Black, B.E.
(2020). Structural basis for allosteric PARP-1 retention on DNA breaks. Science,
Vol.368
(6486).
show abstract
The success of poly(ADP-ribose) polymerase-1 (PARP-1) inhibitors (PARPi) to treat cancer relates to their ability to trap PARP-1 at the site of a DNA break. Although different forms of PARPi all target the catalytic center of the enzyme, they have variable abilities to trap PARP-1. We found that several structurally distinct PARPi drive PARP-1 allostery to promote release from a DNA break. Other inhibitors drive allostery to retain PARP-1 on a DNA break. Further, we generated a new PARPi compound, converting an allosteric pro-release compound to a pro-retention compound and increasing its ability to kill cancer cells. These developments are pertinent to clinical applications where PARP-1 trapping is either desirable or undesirable..
Khalique, S.
Lord, C.J.
Banerjee, S.
Natrajan, R.
(2020). Translational genomics of ovarian clear cell carcinoma. Semin cancer biol,
Vol.61,
pp. 121-131.
show abstract
full text
Ovarian clear cell carcinomas (OCCC) are rare aggressive, chemo-resistant tumours comprising approximately 13% of all epithelial ovarian cancers, which have distinct clinical and molecular features, when compared to other gynaecological malignancies. At present, there are no specific licensed targeted therapies for OCCC, although a number of candidate targets have been identified. This review focuses on recent knowledge underpinning our understanding of the pathogenesis of OCCC including direct and synthetic-lethal therapeutic strategies in particular focussing on ARID1A deficiency. We also discuss current targeted clinical trials and immunotherapeutic approaches..
King, D.
Li, X.D.
Almeida, G.S.
Kwok, C.
Gravells, P.
Harrison, D.
Burke, S.
Hallsworth, A.
Jamin, Y.
George, S.
Robinson, S.P.
Lord, C.J.
Poon, E.
Yeomanson, D.
Chesler, L.
Bryant, H.E.
(2020). MYCN expression induces replication stress and sensitivity to PARP inhibition in neuroblastoma. Oncotarget,
Vol.11
(23),
pp. 2141-2159.
show abstract
full text
This study investigates the influence expression of the MYCN oncogene has on the DNA damage response, replication fork progression and sensitivity to PARP inhibition in neuroblastoma. In a panel of neuroblastoma cell lines, MYCN amplification or MYCN expression resulted in increased cell death in response to a range of PARP inhibitors (niraparib, veliparib, talazoparib and olaparib) compared to the response seen in non-expressing/amplified cells. MYCN expression slowed replication fork speed and increased replication fork stalling, an effect that was amplified by PARP inhibition or PARP1 depletion. Increased DNA damage seen was specifically induced in S-phase cells. Importantly, PARP inhibition caused a significant increase in the survival of mice bearing MYCN expressing tumours in a transgenic murine model of MYCN expressing neuroblastoma. Olaparib also sensitized MYCN expressing cells to camptothecin- and temozolomide-induced cell death to a greater degree than non-expressing cells. In summary, MYCN expression leads to increased replication stress in neuroblastoma cells. This effect is exaggerated by inhibition of PARP, resulting in S-phase specific DNA damage and ultimately increased tumour cell death. PARP inhibition alone or in combination with classical chemotherapeutics is therefore a potential therapeutic strategy for neuroblastoma and may be more effective in MYCN expressing tumours..
George, S.L.
Lorenzi, F.
King, D.
Hartlieb, S.
Campbell, J.
Pemberton, H.
Toprak, U.H.
Barker, K.
Tall, J.
da Costa, B.M.
van den Boogaard, M.L.
Dolman, M.E.
Molenaar, J.J.
Bryant, H.E.
Westermann, F.
Lord, C.J.
Chesler, L.
(2020). Therapeutic vulnerabilities in the DNA damage response for the treatment of ATRX mutant neuroblastoma. Ebiomedicine,
Vol.59,
p. 102971.
show abstract
full text
BACKGROUND: In neuroblastoma, genetic alterations in ATRX, define a distinct poor outcome patient subgroup. Despite the need for new therapies, there is a lack of available models and a dearth of pre-clinical research. METHODS: To evaluate the impact of ATRX loss of function (LoF) in neuroblastoma, we utilized CRISPR-Cas9 gene editing to generate neuroblastoma cell lines isogenic for ATRX. We used these and other models to identify therapeutically exploitable synthetic lethal vulnerabilities associated with ATRX LoF. FINDINGS: In isogenic cell lines, we found that ATRX inactivation results in increased DNA damage, homologous recombination repair (HRR) defects and impaired replication fork processivity. In keeping with this, high-throughput compound screening showed selective sensitivity in ATRX mutant cells to multiple PARP inhibitors and the ATM inhibitor KU60019. ATRX mutant cells also showed selective sensitivity to the DNA damaging agents, sapacitabine and irinotecan. HRR deficiency was also seen in the ATRX deleted CHLA-90 cell line, and significant sensitivity demonstrated to olaparib/irinotecan combination therapy in all ATRX LoF models. In-vivo sensitivity to olaparib/irinotecan was seen in ATRX mutant but not wild-type xenografts. Finally, sustained responses to olaparib/irinotecan therapy were seen in an ATRX deleted neuroblastoma patient derived xenograft. INTERPRETATION: ATRX LoF results in specific DNA damage repair defects that can be therapeutically exploited. In ATRX LoF models, preclinical sensitivity is demonstrated to olaparib and irinotecan, a combination that can be rapidly translated into the clinic. FUNDING: This work was supported by Christopher's Smile, Neuroblastoma UK, Cancer Research UK, and the Royal Marsden Hospital NIHR BRC..
Yeow, Z.Y.
Lambrus, B.G.
Marlow, R.
Zhan, K.H.
Durin, M.-.
Evans, L.T.
Scott, P.M.
Phan, T.
Park, E.
Ruiz, L.A.
Moralli, D.
Knight, E.G.
Badder, L.M.
Novo, D.
Haider, S.
Green, C.M.
Tutt, A.N.
Lord, C.J.
Chapman, J.R.
Holland, A.J.
(2020). Targeting TRIM37-driven centrosome dysfunction in 17q23-amplified breast cancer. Nature,
Vol.585
(7825),
pp. 447-452.
show abstract
full text
Genomic instability is a hallmark of cancer, and has a central role in the initiation and development of breast cancer1,2. The success of poly-ADP ribose polymerase inhibitors in the treatment of breast cancers that are deficient in homologous recombination exemplifies the utility of synthetically lethal genetic interactions in the treatment of breast cancers that are driven by genomic instability3. Given that defects in homologous recombination are present in only a subset of breast cancers, there is a need to identify additional driver mechanisms for genomic instability and targeted strategies to exploit these defects in the treatment of cancer. Here we show that centrosome depletion induces synthetic lethality in cancer cells that contain the 17q23 amplicon, a recurrent copy number aberration that defines about 9% of all primary breast cancer tumours and is associated with high levels of genomic instability4-6. Specifically, inhibition of polo-like kinase 4 (PLK4) using small molecules leads to centrosome depletion, which triggers mitotic catastrophe in cells that exhibit amplicon-directed overexpression of TRIM37. To explain this effect, we identify TRIM37 as a negative regulator of centrosomal pericentriolar material. In 17q23-amplified cells that lack centrosomes, increased levels of TRIM37 block the formation of foci that comprise pericentriolar material-these foci are structures with a microtubule-nucleating capacity that are required for successful cell division in the absence of centrosomes. Finally, we find that the overexpression of TRIM37 causes genomic instability by delaying centrosome maturation and separation at mitotic entry, and thereby increases the frequency of mitotic errors. Collectively, these findings highlight TRIM37-dependent genomic instability as a putative driver event in 17q23-amplified breast cancer and provide a rationale for the use of centrosome-targeting therapeutic agents in treating these cancers..
Yap, T.A.
Kristeleit, R.
Michalarea, V.
Pettitt, S.J.
Lim, J.S.
Carreira, S.
Roda, D.
Miller, R.
Riisnaes, R.
Miranda, S.
Figueiredo, I.
Rodrigues, D.N.
Ward, S.
Matthews, R.
Parmar, M.
Turner, A.
Tunariu, N.
Chopra, N.
Gevensleben, H.
Turner, N.C.
Ruddle, R.
Raynaud, F.I.
Decordova, S.
Swales, K.E.
Finneran, L.
Hall, E.
Rugman, P.
Lindemann, J.P.
Foxley, A.
Lord, C.J.
Banerji, U.
Plummer, R.
Basu, B.
Lopez, J.S.
Drew, Y.
de Bono, J.S.
(2020). Phase I Trial of the PARP Inhibitor Olaparib and AKT Inhibitor Capivasertib in Patients with BRCA1/2- and Non-BRCA1/2-Mutant Cancers. Cancer discov,
Vol.10
(10),
pp. 1528-1543.
show abstract
full text
Preclinical studies have demonstrated synergy between PARP and PI3K/AKT pathway inhibitors in BRCA1 and BRCA2 (BRCA1/2)-deficient and BRCA1/2-proficient tumors. We conducted an investigator-initiated phase I trial utilizing a prospective intrapatient dose- escalation design to assess two schedules of capivasertib (AKT inhibitor) with olaparib (PARP inhibitor) in 64 patients with advanced solid tumors. Dose expansions enrolled germline BRCA1/2-mutant tumors, or BRCA1/2 wild-type cancers harboring somatic DNA damage response (DDR) or PI3K-AKT pathway alterations. The combination was well tolerated. Recommended phase II doses for the two schedules were: olaparib 300 mg twice a day with either capivasertib 400 mg twice a day 4 days on, 3 days off, or capivasertib 640 mg twice a day 2 days on, 5 days off. Pharmacokinetics were dose proportional. Pharmacodynamic studies confirmed phosphorylated (p) GSK3β suppression, increased pERK, and decreased BRCA1 expression. Twenty-five (44.6%) of 56 evaluable patients achieved clinical benefit (RECIST complete response/partial response or stable disease ≥ 4 months), including patients with tumors harboring germline BRCA1/2 mutations and BRCA1/2 wild-type cancers with or without DDR and PI3K-AKT pathway alterations. SIGNIFICANCE: In the first trial to combine PARP and AKT inhibitors, a prospective intrapatient dose- escalation design demonstrated safety, tolerability, and pharmacokinetic-pharmacodynamic activity and assessed predictive biomarkers of response/resistance. Antitumor activity was observed in patients harboring tumors with germline BRCA1/2 mutations and BRCA1/2 wild-type cancers with or without somatic DDR and/or PI3K-AKT pathway alterations.This article is highlighted in the In This Issue feature, p. 1426..
Pettitt, S.J.
Frankum, J.R.
Punta, M.
Lise, S.
Alexander, J.
Chen, Y.
Yap, T.A.
Haider, S.
Tutt, A.N.
Lord, C.J.
(2020). Clinical BRCA1/2 Reversion Analysis Identifies Hotspot Mutations and Predicted Neoantigens Associated with Therapy Resistance. Cancer discov,
Vol.10
(10),
pp. 1475-1488.
show abstract
full text
Reversion mutations in BRCA1 or BRCA2 are associated with resistance to PARP inhibitors and platinum. To better understand the nature of these mutations, we collated, codified, and analyzed more than 300 reversions. This identified reversion "hotspots" and "deserts" in regions encoding the N and C terminus, respectively, of BRCA2, suggesting that pathogenic mutations in these regions may be at higher or lower risk of reversion. Missense and splice-site pathogenic mutations in BRCA1/2 also appeared less likely to revert than truncating mutations. Most reversions were <100 bp deletions. Although many deletions exhibited microhomology, this was not universal, suggesting that multiple DNA-repair processes cause reversion. Finally, we found that many reversions were predicted to encode immunogenic neopeptides, suggesting a route to the treatment of reverted disease. As well as providing a freely available database for the collation of future reversion cases, these observations have implications for how drug resistance might be managed in BRCA-mutant cancers. SIGNIFICANCE: Reversion mutations in BRCA genes are a major cause of clinical platinum and PARP inhibitor resistance. This analysis of all reported clinical reversions suggests that the position of BRCA2 mutations affects the risk of reversion. Many reversions are also predicted to encode tumor neoantigens, providing a potential route to targeting resistance.This article is highlighted in the In This Issue feature, p. 1426..
Brunton, H.
Caligiuri, G.
Cunningham, R.
Upstill-Goddard, R.
Bailey, U.-.
Garner, I.M.
Nourse, C.
Dreyer, S.
Jones, M.
Moran-Jones, K.
Wright, D.W.
Paulus-Hock, V.
Nixon, C.
Thomson, G.
Jamieson, N.B.
McGregor, G.A.
Evers, L.
McKay, C.J.
Gulati, A.
Brough, R.
Bajrami, I.
Pettitt, S.J.
Dziubinski, M.L.
Barry, S.T.
Grützmann, R.
Brown, R.
Curry, E.
Glasgow Precision Oncology Laboratory,
Australian Pancreatic Cancer Genome Initiative,
Pajic, M.
Musgrove, E.A.
Petersen, G.M.
Shanks, E.
Ashworth, A.
Crawford, H.C.
Simeone, D.M.
Froeling, F.E.
Lord, C.J.
Mukhopadhyay, D.
Pilarsky, C.
Grimmond, S.E.
Morton, J.P.
Sansom, O.J.
Chang, D.K.
Bailey, P.J.
Biankin, A.V.
(2020). HNF4A and GATA6 Loss Reveals Therapeutically Actionable Subtypes in Pancreatic Cancer. Cell rep,
Vol.31
(6),
p. 107625.
show abstract
full text
Pancreatic ductal adenocarcinoma (PDAC) can be divided into transcriptomic subtypes with two broad lineages referred to as classical (pancreatic) and squamous. We find that these two subtypes are driven by distinct metabolic phenotypes. Loss of genes that drive endodermal lineage specification, HNF4A and GATA6, switch metabolic profiles from classical (pancreatic) to predominantly squamous, with glycogen synthase kinase 3 beta (GSK3β) a key regulator of glycolysis. Pharmacological inhibition of GSK3β results in selective sensitivity in the squamous subtype; however, a subset of these squamous patient-derived cell lines (PDCLs) acquires rapid drug tolerance. Using chromatin accessibility maps, we demonstrate that the squamous subtype can be further classified using chromatin accessibility to predict responsiveness and tolerance to GSK3β inhibitors. Our findings demonstrate that distinct patterns of chromatin accessibility can be used to identify patient subgroups that are indistinguishable by gene expression profiles, highlighting the utility of chromatin-based biomarkers for patient selection in the treatment of PDAC..
Miller, R.E.
Leary, A.
Scott, C.L.
Serra, V.
Lord, C.J.
Bowtell, D.
Chang, D.K.
Garsed, D.W.
Jonkers, J.
Ledermann, J.A.
Nik-Zainal, S.
Ray-Coquard, I.
Shah, S.P.
Matias-Guiu, X.
Swisher, E.M.
Yates, L.R.
(2020). ESMO recommendations on predictive biomarker testing for homologous recombination deficiency and PARP inhibitor benefit in ovarian cancer. Ann oncol,
Vol.31
(12),
pp. 1606-1622.
show abstract
BACKGROUND: Homologous recombination repair deficiency (HRD) is a frequent feature of high-grade serous ovarian, fallopian tube and peritoneal carcinoma (HGSC) and is associated with sensitivity to PARP inhibitor (PARPi) therapy. HRD testing provides an opportunity to optimise PARPi use in HGSC but methodologies are diverse and clinical application remains controversial. MATERIALS AND METHODS: To define best practice for HRD testing in HGSC the ESMO Translational Research and Precision Medicine Working Group launched a collaborative project that incorporated a systematic review approach. The main aims were to (i) define the term 'HRD test'; (ii) provide an overview of the biological rationale and the level of evidence supporting currently available HRD tests; (iii) provide recommendations on the clinical utility of HRD tests in clinical management of HGSC. RESULTS: A broad range of repair genes, genomic scars, mutational signatures and functional assays are associated with a history of HRD. Currently, the clinical validity of HRD tests in ovarian cancer is best assessed, not in terms of biological HRD status per se, but in terms of PARPi benefit. Clinical trials evidence supports the use of BRCA mutation testing and two commercially available assays that also incorporate genomic instability for identifying subgroups of HGSCs that derive different magnitudes of benefit from PARPi therapy, albeit with some variation by clinical scenario. These tests can be used to inform treatment selection and scheduling but their use is limited by a failure to consistently identify a subgroup of patients who derive no benefit from PARPis in most studies. Existing tests lack negative predictive value and inadequately address the complex and dynamic nature of the HRD phenotype. CONCLUSIONS: Currently available HRD tests are useful for predicting likely magnitude of benefit from PARPis but better biomarkers are urgently needed to better identify current homologous recombination proficiency status and stratify HGSC management..
Yap, T.A.
O'Carrigan, B.
Penney, M.S.
Lim, J.S.
Brown, J.S.
de Miguel Luken, M.J.
Tunariu, N.
Perez-Lopez, R.
Rodrigues, D.N.
Riisnaes, R.
Figueiredo, I.
Carreira, S.
Hare, B.
McDermott, K.
Khalique, S.
Williamson, C.T.
Natrajan, R.
Pettitt, S.J.
Lord, C.J.
Banerji, U.
Pollard, J.
Lopez, J.
de Bono, J.S.
(2020). Phase I Trial of First-in-Class ATR Inhibitor M6620 (VX-970) as Monotherapy or in Combination With Carboplatin in Patients With Advanced Solid Tumors. J clin oncol,
Vol.38
(27),
pp. 3195-3204.
show abstract
full text
PURPOSE: Preclinical studies demonstrated that ATR inhibition can exploit synthetic lethality (eg, in cancer cells with impaired compensatory DNA damage responses through ATM loss) as monotherapy and combined with DNA-damaging drugs such as carboplatin. PATIENTS AND METHODS: This phase I trial assessed the ATR inhibitor M6620 (VX-970) as monotherapy (once or twice weekly) and combined with carboplatin (carboplatin on day 1 and M6620 on days 2 and 9 in 21-day cycles). Primary objectives were safety, tolerability, and maximum tolerated dose; secondary objectives included pharmacokinetics and antitumor activity; exploratory objectives included pharmacodynamics in timed paired tumor biopsies. RESULTS: Forty patients were enrolled; 17 received M6620 monotherapy, which was safe and well tolerated. The recommended phase II dose (RP2D) for once- or twice-weekly administration was 240 mg/m2. A patient with metastatic colorectal cancer harboring molecular aberrations, including ATM loss and an ARID1A mutation, achieved RECISTv1.1 complete response and maintained this response, with a progression-free survival of 29 months at last assessment. Twenty-three patients received M6620 with carboplatin, with mechanism-based hematologic toxicities at higher doses, requiring dose delays and reductions. The RP2D for combination therapy was M6620 90 mg/m2 with carboplatin AUC5. A patient with advanced germline BRCA1 ovarian cancer achieved RECISTv1.1 partial response and Gynecologic Cancer Intergroup CA125 response despite being platinum refractory and PARP inhibitor resistant. An additional 15 patients had RECISTv1.1 stable disease as best response. Pharmacokinetics were dose proportional and exceeded preclinical efficacious levels. Pharmacodynamic studies demonstrated substantial inhibition of phosphorylation of CHK1, the downstream ATR substrate. CONCLUSION: To our knowledge, this report is the first of an ATR inhibitor as monotherapy and combined with carboplatin. M6620 was well tolerated, with target engagement and preliminary antitumor responses observed..
Lord, C.J.
Quinn, N.
Ryan, C.J.
(2020). Integrative analysis of large-scale loss-of-function screens identifies robust cancer-associated genetic interactions. Elife,
Vol.9.
show abstract
full text
Genetic interactions, including synthetic lethal effects, can now be systematically identified in cancer cell lines using high-throughput genetic perturbation screens. Despite this advance, few genetic interactions have been reproduced across multiple studies and many appear highly context-specific. Here, by developing a new computational approach, we identified 220 robust driver-gene associated genetic interactions that can be reproduced across independent experiments and across non-overlapping cell line panels. Analysis of these interactions demonstrated that: (i) oncogene addiction effects are more robust than oncogene-related synthetic lethal effects; and (ii) robust genetic interactions are enriched among gene pairs whose protein products physically interact. Exploiting the latter observation, we used a protein-protein interaction network to identify robust synthetic lethal effects associated with passenger gene alterations and validated two new synthetic lethal effects. Our results suggest that protein-protein interaction networks can be used to prioritise therapeutic targets that will be more robust to tumour heterogeneity..
Nyamundanda, G.
Eason, K.
Guinney, J.
Lord, C.J.
Sadanandam, A.
(2020). A Machine-Learning Tool Concurrently Models Single Omics and Phenome Data for Functional Subtyping and Personalized Cancer Medicine. Cancers (basel),
Vol.12
(10).
show abstract
full text
One of the major challenges in defining clinically-relevant and less heterogeneous tumor subtypes is assigning biological and/or clinical interpretations to etiological (intrinsic) subtypes. Conventional clustering/subtyping approaches often fail to define such subtypes, as they involve several discrete steps. Here we demonstrate a unique machine-learning method, phenotype mapping (PhenMap), which jointly integrates single omics data with phenotypic information using three published breast cancer datasets (n = 2045). The PhenMap framework uses a modified factor analysis method that is governed by a key assumption that, features from different omics data types are correlated due to specific "hidden/mapping" variables (context-specific mapping variables (CMV)). These variables can be simultaneously modeled with phenotypic data as covariates to yield functional subtypes and their associated features (e.g., genes) and phenotypes. In one example, we demonstrate the identification and validation of six novel "functional" (discrete) subtypes with differential responses to a cyclin-dependent kinase (CDK)4/6 inhibitor and etoposide by jointly integrating transcriptome profiles with four different drug response data from 37 breast cancer cell lines. These robust subtypes are also present in patient breast tumors with different prognosis. In another example, we modeled patient gene expression profiles and clinical covariates together to identify continuous subtypes with clinical/biological implications. Overall, this genome-phenome machine-learning integration tool, PhenMap identifies functional and phenotype-integrated discrete or continuous subtypes with clinical translational potential..
Quist, J.
Mirza, H.
Cheang, M.C.
Telli, M.L.
O'Shaughnessy, J.A.
Lord, C.J.
Tutt, A.N.
Grigoriadis, A.
(2019). A Four-gene Decision Tree Signature Classification of Triple-negative Breast Cancer: Implications for Targeted Therapeutics. Mol cancer ther,
Vol.18
(1),
pp. 204-212.
show abstract
full text
The molecular complexity of triple-negative breast cancers (TNBCs) provides a challenge for patient management. We set out to characterize this heterogeneous disease by combining transcriptomics and genomics data, with the aim of revealing convergent pathway dependencies with the potential for treatment intervention. A Bayesian algorithm was used to integrate molecular profiles in two TNBC cohorts, followed by validation using five independent cohorts (n = 1,168), including three clinical trials. A four-gene decision tree signature was identified, which robustly classified TNBCs into six subtypes. All four genes in the signature (EXO1, TP53BP2, FOXM1, and RSU1) are associated with either genomic instability, malignant growth, or treatment response. One of the six subtypes, MC6, encompassed the largest proportion of tumors (∼50%) in early diagnosed TNBCs. In TNBC patients with metastatic disease, the MC6 proportion was reduced to 25%, and was independently associated with a higher response rate to platinum-based chemotherapy. In TNBC cell line data, platinum sensitivity was recapitulated, and a sensitivity to the inhibition of the phosphatase PPM1D was revealed. Molecularly, MC6-TNBCs displayed high levels of telomeric allelic imbalances, enrichment of CD4+ and CD8+ immune signatures, and reduced expression of genes negatively regulating the MAPK signaling pathway. These observations suggest that our integrative classification approach may identify TNBC patients with discernible and theoretically pharmacologically tractable features that merit further studies in prospective trials..
Chabanon, R.M.
Muirhead, G.
Krastev, D.B.
Adam, J.
Morel, D.
Garrido, M.
Lamb, A.
Hénon, C.
Dorvault, N.
Rouanne, M.
Marlow, R.
Bajrami, I.
Cardeñosa, M.L.
Konde, A.
Besse, B.
Ashworth, A.
Pettitt, S.J.
Haider, S.
Marabelle, A.
Tutt, A.N.
Soria, J.-.
Lord, C.J.
Postel-Vinay, S.
(2019). PARP inhibition enhances tumor cell-intrinsic immunity in ERCC1-deficient non-small cell lung cancer. J clin invest,
Vol.129
(3),
pp. 1211-1228.
show abstract
full text
The cyclic GMP-AMP synthase/stimulator of IFN genes (cGAS/STING) pathway detects cytosolic DNA to activate innate immune responses. Poly(ADP-ribose) polymerase inhibitors (PARPi) selectively target cancer cells with DNA repair deficiencies such as those caused by BRCA1 mutations or ERCC1 defects. Using isogenic cell lines and patient-derived samples, we showed that ERCC1-defective non-small cell lung cancer (NSCLC) cells exhibit an enhanced type I IFN transcriptomic signature and that low ERCC1 expression correlates with increased lymphocytic infiltration. We demonstrated that clinical PARPi, including olaparib and rucaparib, have cell-autonomous immunomodulatory properties in ERCC1-defective NSCLC and BRCA1-defective triple-negative breast cancer (TNBC) cells. Mechanistically, PARPi generated cytoplasmic chromatin fragments with characteristics of micronuclei; these were found to activate cGAS/STING, downstream type I IFN signaling, and CCL5 secretion. Importantly, these effects were suppressed in PARP1-null TNBC cells, suggesting that this phenotype resulted from an on-target effect of PARPi on PARP1. PARPi also potentiated IFN-γ-induced PD-L1 expression in NSCLC cell lines and in fresh patient tumor cells; this effect was enhanced in ERCC1-deficient contexts. Our data provide a preclinical rationale for using PARPi as immunomodulatory agents in appropriately molecularly selected populations..
Oser, M.G.
Fonseca, R.
Chakraborty, A.A.
Brough, R.
Spektor, A.
Jennings, R.B.
Flaifel, A.
Novak, J.S.
Gulati, A.
Buss, E.
Younger, S.T.
McBrayer, S.K.
Cowley, G.S.
Bonal, D.M.
Nguyen, Q.-.
Brulle-Soumare, L.
Taylor, P.
Cairo, S.
Ryan, C.J.
Pease, E.J.
Maratea, K.
Travers, J.
Root, D.E.
Signoretti, S.
Pellman, D.
Ashton, S.
Lord, C.J.
Barry, S.T.
Kaelin, W.G.
(2019). Cells Lacking the RB1 Tumor Suppressor Gene Are Hyperdependent on Aurora B Kinase for Survival. Cancer discov,
Vol.9
(2),
pp. 230-247.
show abstract
full text
Small cell lung cancer (SCLC) accounts for 15% of lung cancers and is almost always linked to inactivating RB1 and TP53 mutations. SCLC frequently responds, albeit briefly, to chemotherapy. The canonical function of the RB1 gene product RB1 is to repress the E2F transcription factor family. RB1 also plays both E2F-dependent and E2F-independent mitotic roles. We performed a synthetic lethal CRISPR/Cas9 screen in an RB1 -/- SCLC cell line that conditionally expresses RB1 to identify dependencies that are caused by RB1 loss and discovered that RB1 -/- SCLC cell lines are hyperdependent on multiple proteins linked to chromosomal segregation, including Aurora B kinase. Moreover, we show that an Aurora B kinase inhibitor is efficacious in multiple preclinical SCLC models at concentrations that are well tolerated in mice. These results suggest that RB1 loss is a predictive biomarker for sensitivity to Aurora B kinase inhibitors in SCLC and perhaps other RB1 -/- cancers. SIGNIFICANCE: SCLC is rarely associated with actionable protooncogene mutations. We did a CRISPR/Cas9-based screen that showed that RB1 -/- SCLC are hyperdependent on AURKB, likely because both genes control mitotic fidelity, and confirmed that Aurora B kinase inhibitors are efficacious against RB1 -/- SCLC tumors in mice at nontoxic doses.See related commentary by Dick and Li, p. 169.This article is highlighted in the In This Issue feature, p. 151..
Pettitt, S.J.
Lord, C.J.
(2019). Dissecting PARP inhibitor resistance with functional genomics. Curr opin genet dev,
Vol.54,
pp. 55-63.
show abstract
full text
The poly-(ADP-ribose) polymerase (PARP) inhibitor (PARPi) olaparib was the first licenced cancer drug that targeted an inherited form of cancer, namely ovarian cancers caused by germline BRCA1 or BRCA2 gene mutations. Multiple different PARPi have now been approved for use in a wider group of gynaecological cancers as well as for the treatment of BRCA-gene mutant breast cancer. Despite these advances, resistance to PARPi is a common clinical phenotype. Understanding, at the molecular level, how tumour cells respond to PARPi has the potential to inform how these drugs should be used clinically and since the discovery of this drug class, multiple different functional genomic strategies have been employed to dissect PARPi sensitivity and resistance. These have included genetic perturbation via classical gene targeting, gene silencing by siRNA or shRNA or transposon mutagenesis techniques. Recently, CRISPR-Cas9-based mutagenesis has greatly expanded the available range of relevant preclinical models and the precision of mutagenesis. Here, we review how these approaches have been used either in low-throughput, hypothesis-testing experiments or in the setting of large, hypothesis-generating, genetic screens aimed at understanding the molecular basis of PARPi sensitivity and resistance..
Tonnessen-Murray, C.A.
Frey, W.D.
Rao, S.G.
Shahbandi, A.
Ungerleider, N.A.
Olayiwola, J.O.
Murray, L.B.
Vinson, B.T.
Chrisey, D.B.
Lord, C.J.
Jackson, J.G.
(2019). Chemotherapy-induced senescent cancer cells engulf other cells to enhance their survival. J cell biol,
Vol.218
(11),
pp. 3827-3844.
show abstract
full text
In chemotherapy-treated breast cancer, wild-type p53 preferentially induces senescence over apoptosis, resulting in a persisting cell population constituting residual disease that drives relapse and poor patient survival via the senescence-associated secretory phenotype. Understanding the properties of tumor cells that allow survival after chemotherapy treatment is paramount. Using time-lapse and confocal microscopy to observe interactions of cells in treated tumors, we show here that chemotherapy-induced senescent cells frequently engulf both neighboring senescent or nonsenescent tumor cells at a remarkable frequency. Engulfed cells are processed through the lysosome and broken down, and cells that have engulfed others obtain a survival advantage. Gene expression analysis showed a marked up-regulation of conserved macrophage-like program of engulfment in chemotherapy-induced senescent cell lines and tumors. Our data suggest compelling explanations for how senescent cells persist in dormancy, how they manage the metabolically expensive process of cytokine production that drives relapse in those tumors that respond the worst, and a function for their expanded lysosomal compartment..
Mateo, J.
Lord, C.J.
Serra, V.
Tutt, A.
Balmaña, J.
Castroviejo-Bermejo, M.
Cruz, C.
Oaknin, A.
Kaye, S.B.
de Bono, J.S.
(2019). A decade of clinical development of PARP inhibitors in perspective. Ann oncol,
Vol.30
(9),
pp. 1437-1447.
show abstract
full text
Genomic instability is a hallmark of cancer, and often is the result of altered DNA repair capacities in tumour cells. DNA damage repair defects are common in different cancer types; these alterations can also induce tumour-specific vulnerabilities that can be exploited therapeutically. In 2009, a first-in-man clinical trial of the poly(ADP-ribose) polymerase (PARP) inhibitor olaparib clinically validated the synthetic lethal interaction between inhibition of PARP1, a key sensor of DNA damage, and BRCA1/BRCA2 deficiency. In this review, we summarize a decade of PARP inhibitor clinical development, a work that has resulted in the registration of several PARP inhibitors in breast (olaparib and talazoparib) and ovarian cancer (olaparib, niraparib and rucaparib, either alone or following platinum chemotherapy as maintenance therapy). Over the past 10 years, our knowledge on the mechanism of action of PARP inhibitor as well as how tumours become resistant has been extended, and we summarise this work here. We also discuss opportunities for expanding the precision medicine approach with PARP inhibitors, identifying a wider population who could benefit from this drug class. This includes developing and validating better predictive biomarkers for patient stratification, mainly based on homologous recombination defects beyond BRCA1/BRCA2 mutations, identifying DNA repair deficient tumours in other cancer types such as prostate or pancreatic cancer, or by designing combination therapies with PARP inhibitors..
Davidson, M.
Aronson, L.I.
Howard-Reeves, J.
Bryant, H.
Cutts, R.J.
Hulkki-Wilson, S.
Kouvelakis, K.
Kalaitzaki, E.
Watkins, D.
Starling, N.
Rao, S.
Cardenosa, M.L.
Begum, R.
Rana, I.
Lazaro-Alcausi, R.
Terlizzo, M.
Wotherspoon, A.
Brown, G.
Swansbury, J.
Lord, C.J.
Cunningham, D.
Chau, I.
Chong, I.Y.
(2019). Clonal diversity of MYC amplification evaluated by fluorescent in situ hybridisation and digital droplet polymerase chain reaction in oesophagogastric cancer: Results from a prospective clinical trial screening programme. Eur j cancer,
Vol.122,
pp. 12-21.
show abstract
INTRODUCTION: The MYC proto-oncogene is among the most commonly dysregulated genes in human cancers. We report screening data from the iMYC trial, an ongoing phase II study assessing ibrutinib monotherapy in advanced pretreated MYC- and/or HER2-amplified oesophagogastric cancer, representing the first attempt to prospectively identify MYC amplifications in this tumour type for the purposes of therapeutic targeting. METHODS: Screening utilising a fluorescent in situ hybridisation (FISH) assay for assessment of tumour MYC amplification has been instituted. An experimental digital droplet polymerase chain reaction (ddPCR) assay to assess MYC amplification in both tumour and circulating-tumour (ct)DNA has been developed and investigated. RESULTS: One hundred thirty-five archival tumour specimens have undergone successful FISH analysis with 23% displaying evidence of MYC amplification. Intertumour heterogeneity was observed, with the percentage of cancer cells harbouring MYC amplification ranging widely between samples (median 51%, range 11-94%). Intratumoural clonal diversity of MYC amplification was also observed, with a significant degree of variance in amplification ratios (Bartlett's test for equal variance p < 0.001), and an association between greater variance in MYC amplification and improved outcome with prior first-line chemotherapy. ddPCR was most accurate in quantifying MYC amplification in tumour-derived DNA from cases with a high proportion (>70%) of amplified cells within the tumour specimen but was not reliable in samples containing a low proportion of amplified cells or in ctDNA. CONCLUSIONS: Our results illustrate the utility of FISH to assess MYC amplification prospectively for a biomarker-selected trial by providing reliable and reproducible results in real time, with a high degree of heterogeneity of MYC amplification observed. We show that ddPCR can potentially detect high-level MYC amplifications in tumour tissue..
Menon, M.
Elliott, R.
Bowers, L.
Balan, N.
Rafiq, R.
Costa-Cabral, S.
Munkonge, F.
Trinidade, I.
Porter, R.
Campbell, A.D.
Johnson, E.R.
Esdar, C.
Buchstaller, H.-.
Leuthner, B.
Rohdich, F.
Schneider, R.
Sansom, O.
Wienke, D.
Ashworth, A.
Lord, C.J.
(2019). A novel tankyrase inhibitor, MSC2504877, enhances the effects of clinical CDK4/6 inhibitors. Sci rep,
Vol.9
(1),
p. 201.
show abstract
full text
Inhibition of the PARP superfamily tankyrase enzymes suppresses Wnt/β-catenin signalling in tumour cells. Here, we describe here a novel, drug-like small molecule inhibitor of tankyrase MSC2504877 that inhibits the growth of APC mutant colorectal tumour cells. Parallel siRNA and drug sensitivity screens showed that the clinical CDK4/6 inhibitor palbociclib, causes enhanced sensitivity to MSC2504877. This tankyrase inhibitor-CDK4/6 inhibitor combinatorial effect is not limited to palbociclib and MSC2504877 and is elicited with other CDK4/6 inhibitors and toolbox tankyrase inhibitors. The addition of MSC2504877 to palbociclib enhances G1 cell cycle arrest and cellular senescence in tumour cells. MSC2504877 exposure suppresses the upregulation of Cyclin D2 and Cyclin E2 caused by palbociclib and enhances the suppression of phospho-Rb, providing a mechanistic explanation for these effects. The combination of MSC2504877 and palbociclib was also effective in suppressing the cellular hyperproliferative phenotype seen in Apc defective intestinal stem cells in vivo. However, the presence of an oncogenic Kras p.G12D mutation in mice reversed the effects of the MSC2504877/palbociclib combination, suggesting one molecular route that could lead to drug resistance..
Chabanon, R.M.
Soria, J.-.
Lord, C.J.
Postel-Vinay, S.
(2019). Beyond DNA repair: the novel immunological potential of PARP inhibitors. Mol cell oncol,
Vol.6
(2),
p. 1585170.
show abstract
full text
Loss of excision repair cross-complementation group 1 (ERCC1), frequently found in lung cancer, and mutations in breast cancer type 1/2 susceptibility genes (BRCA1/2), often found in ovarian, breast and prostate cancers, confer sensitivity to poly-(ADP-ribose) polymerase inhibitors (PARPi). Our work, and that of others, shows that PARPi selectively trigger tumor cell-autonomous immune phenotypes in ERCC1- or BRCA-defective contexts. This suggests that PARPi, used in appropriately selected populations, might mediate their therapeutic effects by potentiating anti-tumor immunity..
Naidoo, K.
Wai, P.T.
Maguire, S.L.
Daley, F.
Haider, S.
Kriplani, D.
Campbell, J.
Mirza, H.
Grigoriadis, A.
Tutt, A.
Moseley, P.M.
Abdel-Fatah, T.M.
Chan, S.Y.
Madhusudan, S.
Rhaka, E.A.
Ellis, I.O.
Lord, C.J.
Yuan, Y.
Green, A.R.
Natrajan, R.
(2018). Evaluation of CDK12 Protein Expression as a Potential Novel Biomarker for DNA Damage Response-Targeted Therapies in Breast Cancer. Mol cancer ther,
Vol.17
(1),
pp. 306-315.
show abstract
full text
Disruption of Cyclin-Dependent Kinase 12 (CDK12) is known to lead to defects in DNA repair and sensitivity to platinum salts and PARP1/2 inhibitors. However, CDK12 has also been proposed as an oncogene in breast cancer. We therefore aimed to assess the frequency and distribution of CDK12 protein expression by IHC in independent cohorts of breast cancer and correlate this with outcome and genomic status. We found that 21% of primary unselected breast cancers were CDK12 high, and 10.5% were absent, by IHC. CDK12 positivity correlated with HER2 positivity but was not an independent predictor of breast cancer-specific survival taking HER2 status into account; however, absent CDK12 protein expression significantly correlated with a triple-negative phenotype. Interestingly, CDK12 protein absence was associated with reduced expression of a number of DDR proteins including ATR, Ku70/Ku80, PARP1, DNA-PK, and γH2AX, suggesting a novel mechanism of CDK12-associated DDR dysregulation in breast cancer. Our data suggest that diagnostic IHC quantification of CDK12 in breast cancer is feasible, with CDK12 absence possibly signifying defective DDR function. This may have important therapeutic implications, particularly for triple-negative breast cancers. Mol Cancer Ther; 17(1); 306-15. ©2017 AACR..
Nava Rodrigues, D.
Rescigno, P.
Liu, D.
Yuan, W.
Carreira, S.
Lambros, M.B.
Seed, G.
Mateo, J.
Riisnaes, R.
Mullane, S.
Margolis, C.
Miao, D.
Miranda, S.
Dolling, D.
Clarke, M.
Bertan, C.
Crespo, M.
Boysen, G.
Ferreira, A.
Sharp, A.
Figueiredo, I.
Keliher, D.
Aldubayan, S.
Burke, K.P.
Sumanasuriya, S.
Fontes, M.S.
Bianchini, D.
Zafeiriou, Z.
Teixeira Mendes, L.S.
Mouw, K.
Schweizer, M.T.
Pritchard, C.C.
Salipante, S.
Taplin, M.-.
Beltran, H.
Rubin, M.A.
Cieslik, M.
Robinson, D.
Heath, E.
Schultz, N.
Armenia, J.
Abida, W.
Scher, H.
Lord, C.
D'Andrea, A.
Sawyers, C.L.
Chinnaiyan, A.M.
Alimonti, A.
Nelson, P.S.
Drake, C.G.
Van Allen, E.M.
de Bono, J.S.
(2018). Immunogenomic analyses associate immunological alterations with mismatch repair defects in prostate cancer. J clin invest,
Vol.128
(10),
pp. 4441-4453.
show abstract
full text
BACKGROUND: Understanding the integrated immunogenomic landscape of advanced prostate cancer (APC) could impact stratified treatment selection. METHODS: Defective mismatch repair (dMMR) status was determined by either loss of mismatch repair protein expression on IHC or microsatellite instability (MSI) by PCR in 127 APC biopsies from 124 patients (Royal Marsden [RMH] cohort); MSI by targeted panel next-generation sequencing (MSINGS) was then evaluated in the same cohort and in 254 APC samples from the Stand Up To Cancer/Prostate Cancer Foundation (SU2C/PCF). Whole exome sequencing (WES) data from this latter cohort were analyzed for pathogenic MMR gene variants, mutational load, and mutational signatures. Transcriptomic data, available for 168 samples, was also performed. RESULTS: Overall, 8.1% of patients in the RMH cohort had some evidence of dMMR, which associated with decreased overall survival. Higher MSINGS scores associated with dMMR, and these APCs were enriched for higher T cell infiltration and PD-L1 protein expression. Exome MSINGS scores strongly correlated with targeted panel MSINGS scores (r = 0.73, P < 0.0001), and higher MSINGS scores associated with dMMR mutational signatures in APC exomes. dMMR mutational signatures also associated with MMR gene mutations and increased immune cell, immune checkpoint, and T cell-associated transcripts. APC with dMMR mutational signatures overexpressed a variety of immune transcripts, including CD200R1, BTLA, PD-L1, PD-L2, ADORA2A, PIK3CG, and TIGIT. CONCLUSION: These data could impact immune target selection, combination therapeutic strategy selection, and selection of predictive biomarkers for immunotherapy in APC. FUNDING: We acknowledge funding support from Movember, Prostate Cancer UK, The Prostate Cancer Foundation, SU2C, and Cancer Research UK..
Touat, M.
Sourisseau, T.
Dorvault, N.
Chabanon, R.M.
Garrido, M.
Morel, D.
Krastev, D.B.
Bigot, L.
Adam, J.
Frankum, J.R.
Durand, S.
Pontoizeau, C.
Souquère, S.
Kuo, M.-.
Sauvaigo, S.
Mardakheh, F.
Sarasin, A.
Olaussen, K.A.
Friboulet, L.
Bouillaud, F.
Pierron, G.
Ashworth, A.
Lombès, A.
Lord, C.J.
Soria, J.-.
Postel-Vinay, S.
(2018). DNA repair deficiency sensitizes lung cancer cells to NAD+ biosynthesis blockade. J clin invest,
Vol.128
(4),
pp. 1671-1687.
show abstract
full text
Synthetic lethality is an efficient mechanism-based approach to selectively target DNA repair defects. Excision repair cross-complementation group 1 (ERCC1) deficiency is frequently found in non-small-cell lung cancer (NSCLC), making this DNA repair protein an attractive target for exploiting synthetic lethal approaches in the disease. Using unbiased proteomic and metabolic high-throughput profiling on a unique in-house-generated isogenic model of ERCC1 deficiency, we found marked metabolic rewiring of ERCC1-deficient populations, including decreased levels of the metabolite NAD+ and reduced expression of the rate-limiting NAD+ biosynthetic enzyme nicotinamide phosphoribosyltransferase (NAMPT). We also found reduced NAMPT expression in NSCLC samples with low levels of ERCC1. These metabolic alterations were a primary effect of ERCC1 deficiency, and caused selective exquisite sensitivity to small-molecule NAMPT inhibitors, both in vitro - ERCC1-deficient cells being approximately 1,000 times more sensitive than ERCC1-WT cells - and in vivo. Using transmission electronic microscopy and functional metabolic studies, we found that ERCC1-deficient cells harbor mitochondrial defects. We propose a model where NAD+ acts as a regulator of ERCC1-deficient NSCLC cell fitness. These findings open therapeutic opportunities that exploit a yet-undescribed nuclear-mitochondrial synthetic lethal relationship in NSCLC models, and highlight the potential for targeting DNA repair/metabolic crosstalks for cancer therapy..
Bajrami, I.
Marlow, R.
van de Ven, M.
Brough, R.
Pemberton, H.N.
Frankum, J.
Song, F.
Rafiq, R.
Konde, A.
Krastev, D.B.
Menon, M.
Campbell, J.
Gulati, A.
Kumar, R.
Pettitt, S.J.
Gurden, M.D.
Cardenosa, M.L.
Chong, I.
Gazinska, P.
Wallberg, F.
Sawyer, E.J.
Martin, L.-.
Dowsett, M.
Linardopoulos, S.
Natrajan, R.
Ryan, C.J.
Derksen, P.W.
Jonkers, J.
Tutt, A.N.
Ashworth, A.
Lord, C.J.
(2018). E-Cadherin/ROS1 Inhibitor Synthetic Lethality in Breast Cancer. Cancer discov,
Vol.8
(4),
pp. 498-515.
show abstract
full text
The cell adhesion glycoprotein E-cadherin (CDH1) is commonly inactivated in breast tumors. Precision medicine approaches that exploit this characteristic are not available. Using perturbation screens in breast tumor cells with CRISPR/Cas9-engineered CDH1 mutations, we identified synthetic lethality between E-cadherin deficiency and inhibition of the tyrosine kinase ROS1. Data from large-scale genetic screens in molecularly diverse breast tumor cell lines established that the E-cadherin/ROS1 synthetic lethality was not only robust in the face of considerable molecular heterogeneity but was also elicited with clinical ROS1 inhibitors, including foretinib and crizotinib. ROS1 inhibitors induced mitotic abnormalities and multinucleation in E-cadherin-defective cells, phenotypes associated with a defect in cytokinesis and aberrant p120 catenin phosphorylation and localization. In vivo, ROS1 inhibitors produced profound antitumor effects in multiple models of E-cadherin-defective breast cancer. These data therefore provide the preclinical rationale for assessing ROS1 inhibitors, such as the licensed drug crizotinib, in appropriately stratified patients.Significance: E-cadherin defects are common in breast cancer but are currently not targeted with a precision medicine approach. Our preclinical data indicate that licensed ROS1 inhibitors, including crizotinib, should be repurposed to target E-cadherin-defective breast cancers, thus providing the rationale for the assessment of these agents in molecularly stratified phase II clinical trials. Cancer Discov; 8(4); 498-515. ©2018 AACR.This article is highlighted in the In This Issue feature, p. 371..
Khalique, S.
Naidoo, K.
Attygalle, A.D.
Kriplani, D.
Daley, F.
Lowe, A.
Campbell, J.
Jones, T.
Hubank, M.
Fenwick, K.
Matthews, N.
Rust, A.G.
Lord, C.J.
Banerjee, S.
Natrajan, R.
(2018). Optimised ARID1A immunohistochemistry is an accurate predictor of ARID1A mutational status in gynaecological cancers. J pathol clin res,
Vol.4
(3),
pp. 154-166.
show abstract
full text
ARID1A is a tumour suppressor gene that is frequently mutated in clear cell and endometrioid carcinomas of the ovary and endometrium and is an important clinical biomarker for novel treatment approaches for patients with ARID1A defects. However, the accuracy of ARID1A immunohistochemistry (IHC) as a surrogate for mutation status has not fully been established for patient stratification in clinical trials. Here we tested whether ARID1A IHC could reliably predict ARID1A mutations identified by next-generation sequencing. Three commercially available antibodies - EPR13501 (Abcam), D2A8U (Cell Signaling), and HPA005456 (Sigma) - were optimised for IHC using cell line models and human tissue, and screened across a cohort of 45 gynaecological tumours. IHC was scored independently by three pathologists using an immunoreactive score. ARID1A mutation status was assessed using two independent sequencing platforms and the concordance between ARID1A mutation and protein expression was evaluated using Receiver Operating Characteristic statistics. Overall, 21 ARID1A mutations were identified in 14/43 assessable tumours (33%), the majority of which were predicted to be deleterious. Mutations were identified in 6/17 (35%) ovarian clear cell carcinomas, 5/8 (63%) ovarian endometrioid carcinomas, 2/5 (40%) endometrial carcinomas, and 1/7 (14%) carcinosarcomas. ROC analysis identified greater than 95% concordance between mutation status and IHC using a modified immunoreactive score for all three antibodies allowing a definitive cut-point for ARID1A mutant status to be calculated. Comprehensive assessment of concordance of ARID1A IHC and mutation status identified EPR13501 as an optimal antibody, with 100% concordance between ARID1A mutation status and protein expression, across different gynaecological histological subtypes. It delivered the best inter-rater agreement between all pathologists, as well as a clear cost-benefit advantage. This could allow patients to be accurately stratified based on their ARID1A IHC status into early phase clinical trials..
Noordermeer, S.M.
Adam, S.
Setiaputra, D.
Barazas, M.
Pettitt, S.J.
Ling, A.K.
Olivieri, M.
Álvarez-Quilón, A.
Moatti, N.
Zimmermann, M.
Annunziato, S.
Krastev, D.B.
Song, F.
Brandsma, I.
Frankum, J.
Brough, R.
Sherker, A.
Landry, S.
Szilard, R.K.
Munro, M.M.
McEwan, A.
Goullet de Rugy, T.
Lin, Z.-.
Hart, T.
Moffat, J.
Gingras, A.-.
Martin, A.
van Attikum, H.
Jonkers, J.
Lord, C.J.
Rottenberg, S.
Durocher, D.
(2018). The shieldin complex mediates 53BP1-dependent DNA repair. Nature,
Vol.560
(7716),
pp. 117-121.
show abstract
full text
53BP1 is a chromatin-binding protein that regulates the repair of DNA double-strand breaks by suppressing the nucleolytic resection of DNA termini1,2. This function of 53BP1 requires interactions with PTIP3 and RIF14-9, the latter of which recruits REV7 (also known as MAD2L2) to break sites10,11. How 53BP1-pathway proteins shield DNA ends is currently unknown, but there are two models that provide the best potential explanation of their action. In one model the 53BP1 complex strengthens the nucleosomal barrier to end-resection nucleases12,13, and in the other 53BP1 recruits effector proteins with end-protection activity. Here we identify a 53BP1 effector complex, shieldin, that includes C20orf196 (also known as SHLD1), FAM35A (SHLD2), CTC-534A2.2 (SHLD3) and REV7. Shieldin localizes to double-strand-break sites in a 53BP1- and RIF1-dependent manner, and its SHLD2 subunit binds to single-stranded DNA via OB-fold domains that are analogous to those of RPA1 and POT1. Loss of shieldin impairs non-homologous end-joining, leads to defective immunoglobulin class switching and causes hyper-resection. Mutations in genes that encode shieldin subunits also cause resistance to poly(ADP-ribose) polymerase inhibition in BRCA1-deficient cells and tumours, owing to restoration of homologous recombination. Finally, we show that binding of single-stranded DNA by SHLD2 is critical for shieldin function, consistent with a model in which shieldin protects DNA ends to mediate 53BP1-dependent DNA repair..
Chong, I.Y.
Aronson, L.
Bryant, H.
Gulati, A.
Campbell, J.
Elliott, R.
Pettitt, S.
Wilkerson, P.
Lambros, M.B.
Reis-Filho, J.S.
Ramessur, A.
Davidson, M.
Chau, I.
Cunningham, D.
Ashworth, A.
Lord, C.J.
(2018). Mapping genetic vulnerabilities reveals BTK as a novel therapeutic target in oesophageal cancer. Gut,
Vol.67
(10),
pp. 1780-1792.
show abstract
full text
OBJECTIVE: Oesophageal cancer is the seventh most common cause of cancer-related death worldwide. Disease relapse is frequent and treatment options are limited. DESIGN: To identify new biomarker-defined therapeutic approaches for patients with oesophageal cancer, we integrated the genomic profiles of 17 oesophageal tumour-derived cell lines with drug sensitivity data from small molecule inhibitor profiling, identifying drug sensitivity effects associated with cancer driver gene alterations. We also interrogated recently described RNA interference screen data for these tumour cell lines to identify candidate genetic dependencies or vulnerabilities that could be exploited as therapeutic targets. RESULTS: By integrating the genomic features of oesophageal tumour cell lines with siRNA and drug screening data, we identified a series of candidate targets in oesophageal cancer, including a sensitivity to inhibition of the kinase BTK in MYC amplified oesophageal tumour cell lines. We found that this genetic dependency could be elicited with the clinical BTK/ERBB2 kinase inhibitor, ibrutinib. In both MYC and ERBB2 amplified tumour cells, ibrutinib downregulated ERK-mediated signal transduction, cMYC Ser-62 phosphorylation and levels of MYC protein, and elicited G1 cell cycle arrest and apoptosis, suggesting that this drug could be used to treat biomarker-selected groups of patients with oesophageal cancer. CONCLUSIONS: BTK represents a novel candidate therapeutic target in oesophageal cancer that can be targeted with ibrutinib. On the basis of this work, a proof-of-concept phase II clinical trial evaluating the efficacy of ibrutinib in patients with MYC and/or ERBB2 amplified advanced oesophageal cancer is currently underway (NCT02884453). TRIAL REGISTRATION NUMBER: NCT02884453; Pre-results..
Brough, R.
Gulati, A.
Haider, S.
Kumar, R.
Campbell, J.
Knudsen, E.
Pettitt, S.J.
Ryan, C.J.
Lord, C.J.
(2018). Identification of highly penetrant Rb-related synthetic lethal interactions in triple negative breast cancer. Oncogene,
Vol.37
(43),
pp. 5701-5718.
show abstract
full text
Although defects in the RB1 tumour suppressor are one of the more common driver alterations found in triple-negative breast cancer (TNBC), therapeutic approaches that exploit this have not been identified. By integrating molecular profiling data with data from multiple genetic perturbation screens, we identified candidate synthetic lethal (SL) interactions associated with RB1 defects in TNBC. We refined this analysis by identifying the highly penetrant effects, reasoning that these would be more robust in the face of molecular heterogeneity and would represent more promising therapeutic targets. A significant proportion of the highly penetrant RB1 SL effects involved proteins closely associated with RB1 function, suggesting that this might be a defining characteristic. These included nuclear pore complex components associated with the MAD2 spindle checkpoint protein, the kinase and bromodomain containing transcription factor TAF1, and multiple components of the SCFSKP Cullin F box containing complex. Small-molecule inhibition of SCFSKP elicited an increase in p27Kip levels, providing a mechanistic rationale for RB1 SL. Transcript expression of SKP2, a SCFSKP component, was elevated in RB1-defective TNBCs, suggesting that in these tumours, SKP2 activity might buffer the effects of RB1 dysfunction..
Pettitt, S.J.
Krastev, D.B.
Brandsma, I.
Dréan, A.
Song, F.
Aleksandrov, R.
Harrell, M.I.
Menon, M.
Brough, R.
Campbell, J.
Frankum, J.
Ranes, M.
Pemberton, H.N.
Rafiq, R.
Fenwick, K.
Swain, A.
Guettler, S.
Lee, J.-.
Swisher, E.M.
Stoynov, S.
Yusa, K.
Ashworth, A.
Lord, C.J.
(2018). Genome-wide and high-density CRISPR-Cas9 screens identify point mutations in PARP1 causing PARP inhibitor resistance. Nat commun,
Vol.9
(1),
p. 1849.
show abstract
full text
Although PARP inhibitors (PARPi) target homologous recombination defective tumours, drug resistance frequently emerges, often via poorly understood mechanisms. Here, using genome-wide and high-density CRISPR-Cas9 "tag-mutate-enrich" mutagenesis screens, we identify close to full-length mutant forms of PARP1 that cause in vitro and in vivo PARPi resistance. Mutations both within and outside of the PARP1 DNA-binding zinc-finger domains cause PARPi resistance and alter PARP1 trapping, as does a PARP1 mutation found in a clinical case of PARPi resistance. This reinforces the importance of trapped PARP1 as a cytotoxic DNA lesion and suggests that PARP1 intramolecular interactions might influence PARPi-mediated cytotoxicity. PARP1 mutations are also tolerated in cells with a pathogenic BRCA1 mutation where they result in distinct sensitivities to chemotherapeutic drugs compared to other mechanisms of PARPi resistance (BRCA1 reversion, 53BP1, REV7 (MAD2L2) mutation), suggesting that the underlying mechanism of PARPi resistance that emerges could influence the success of subsequent therapies..
Ryan, C.J.
Bajrami, I.
Lord, C.J.
(2018). Synthetic Lethality and Cancer - Penetrance as the Major Barrier. Trends cancer,
Vol.4
(10),
pp. 671-683.
show abstract
full text
Synthetic lethality has long been proposed as an approach for targeting genetic defects in tumours. Despite a decade of screening efforts, relatively few robust synthetic lethal targets have been identified. Improved genetic perturbation techniques, including CRISPR/Cas9 gene editing, have resulted in renewed enthusiasm for searching for synthetic lethal effects in cancer. An implicit assumption behind this enthusiasm is that the lack of reproducibly identified targets can be attributed to limitations of RNAi technologies. We argue here that a bigger hurdle is that most synthetic lethal interactions (SLIs) are not highly penetrant, in other words they are not robust to the extensive molecular heterogeneity seen in tumours. We outline strategies for identifying and prioritising SLIs that are most likely to be highly penetrant..
Holme, H.
Gulati, A.
Brough, R.
Fleuren, E.D.
Bajrami, I.
Campbell, J.
Chong, I.Y.
Costa-Cabral, S.
Elliott, R.
Fenton, T.
Frankum, J.
Jones, S.E.
Menon, M.
Miller, R.
Pemberton, H.N.
Postel-Vinay, S.
Rafiq, R.
Selfe, J.L.
von Kriegsheim, A.
Munoz, A.G.
Rodriguez, J.
Shipley, J.
van der Graaf, W.T.
Williamson, C.T.
Ryan, C.J.
Pettitt, S.
Ashworth, A.
Strauss, S.J.
Lord, C.J.
(2018). Chemosensitivity profiling of osteosarcoma tumour cell lines identifies a model of BRCAness. Sci rep,
Vol.8
(1),
p. 10614.
show abstract
full text
Osteosarcoma (OS) is an aggressive sarcoma, where novel treatment approaches are required. Genomic studies suggest that a subset of OS, including OS tumour cell lines (TCLs), exhibit genomic loss of heterozygosity (LOH) patterns reminiscent of BRCA1 or BRCA2 mutant tumours. This raises the possibility that PARP inhibitors (PARPi), used to treat BRCA1/2 mutant cancers, could be used to target OS. Using high-throughput drug sensitivity screening we generated chemosensitivity profiles for 79 small molecule inhibitors, including three clinical PARPi. Drug screening was performed in 88 tumour cell lines, including 18 OS TCLs. This identified known sensitivity effects in OS TCLs, such as sensitivity to FGFR inhibitors. When compared to BRCA1/2 mutant TCLs, OS TCLs, with the exception of LM7, were PARPi resistant, including those with previously determined BRCAness LoH profiles. Post-screen validation experiments confirmed PARPi sensitivity in LM7 cells as well as a defect in the ability to form nuclear RAD51 foci in response to DNA damage. LM7 provides one OS model for the study of PARPi sensitivity through a potential defect in RAD51-mediated DNA repair. The drug sensitivity dataset we generated in 88 TCLs could also serve as a resource for the study of drug sensitivity effects in OS..
Barazas, M.
Annunziato, S.
Pettitt, S.J.
de Krijger, I.
Ghezraoui, H.
Roobol, S.J.
Lutz, C.
Frankum, J.
Song, F.F.
Brough, R.
Evers, B.
Gogola, E.
Bhin, J.
van de Ven, M.
van Gent, D.C.
Jacobs, J.J.
Chapman, R.
Lord, C.J.
Jonkers, J.
Rottenberg, S.
(2018). The CST Complex Mediates End Protection at Double-Strand Breaks and Promotes PARP Inhibitor Sensitivity in BRCA1-Deficient Cells. Cell rep,
Vol.23
(7),
pp. 2107-2118.
show abstract
full text
Selective elimination of BRCA1-deficient cells by inhibitors of poly(ADP-ribose) polymerase (PARP) is a prime example of the concept of synthetic lethality in cancer therapy. This interaction is counteracted by the restoration of BRCA1-independent homologous recombination through loss of factors such as 53BP1, RIF1, and REV7/MAD2L2, which inhibit end resection of DNA double-strand breaks (DSBs). To identify additional factors involved in this process, we performed CRISPR/SpCas9-based loss-of-function screens and selected for factors that confer PARP inhibitor (PARPi) resistance in BRCA1-deficient cells. Loss of members of the CTC1-STN1-TEN1 (CST) complex were found to cause PARPi resistance in BRCA1-deficient cells in vitro and in vivo. We show that CTC1 depletion results in the restoration of end resection and that the CST complex may act downstream of 53BP1/RIF1. These data suggest that, in addition to its role in protecting telomeres, the CST complex also contributes to protecting DSBs from end resection..
Krastev, D.B.
Pettitt, S.J.
Campbell, J.
Song, F.
Tanos, B.E.
Stoynov, S.S.
Ashworth, A.
Lord, C.J.
(2018). Coupling bimolecular PARylation biosensors with genetic screens to identify PARylation targets. Nat commun,
Vol.9
(1),
p. 2016.
show abstract
full text
Poly (ADP-ribose)ylation is a dynamic protein modification that regulates multiple cellular processes. Here, we describe a system for identifying and characterizing PARylation events that exploits the ability of a PBZ (PAR-binding zinc finger) protein domain to bind PAR with high-affinity. By linking PBZ domains to bimolecular fluorescent complementation biosensors, we developed fluorescent PAR biosensors that allow the detection of temporal and spatial PARylation events in live cells. Exploiting transposon-mediated recombination, we integrate the PAR biosensor en masse into thousands of protein coding genes in living cells. Using these PAR-biosensor "tagged" cells in a genetic screen we carry out a large-scale identification of PARylation targets. This identifies CTIF (CBP80/CBP20-dependent translation initiation factor) as a novel PARylation target of the tankyrase enzymes in the centrosomal region of cells, which plays a role in the distribution of the centrosomal satellites..
Ashworth, A.
Lord, C.J.
(2018). Synthetic lethal therapies for cancer: what's next after PARP inhibitors?. Nat rev clin oncol,
.
show abstract
full text
The genetic concept of synthetic lethality has now been validated clinically through the demonstrated efficacy of poly(ADP-ribose) polymerase (PARP) inhibitors for the treatment of cancers in individuals with germline loss-of-function mutations in either BRCA1 or BRCA2. Three different PARP inhibitors have now been approved for the treatment of patients with BRCA-mutant ovarian cancer and one for those with BRCA-mutant breast cancer; these agents have also shown promising results in patients with BRCA-mutant prostate cancer. Here, we describe a number of other synthetic lethal interactions that have been discovered in cancer. We discuss some of the underlying principles that might increase the likelihood of clinical efficacy and how new computational and experimental approaches are now facilitating the discovery and validation of synthetic lethal interactions. Finally, we make suggestions on possible future directions and challenges facing researchers in this field..
Witkiewicz, A.K.
Chung, S.
Brough, R.
Vail, P.
Franco, J.
Lord, C.J.
Knudsen, E.S.
(2018). Targeting the Vulnerability of RB Tumor Suppressor Loss in Triple-Negative Breast Cancer. Cell rep,
Vol.22
(5),
pp. 1185-1199.
show abstract
full text
Approximately 30% of triple-negative breast cancers (TNBCs) exhibit functional loss of the RB tumor suppressor, suggesting a target for precision intervention. Here, we use drug screens to identify agents specifically antagonized by the retinoblastoma tumor suppressor (RB) using CDK4/6 inhibitors. A number of candidate RB-synthetic lethal small molecules were identified, including anti-helmenthics, chemotherapeutic agents, and small-molecule inhibitors targeting DNA-damage checkpoints (e.g., CHK) and chromosome segregation (e.g., PLK1). Counter-screens using isogenic TNBC tumor cell lines and cell panels with varying endogenous RB statuses confirmed that therapeutic effects were robust and selective for RB loss of function. By analyzing TNBC clinical specimens, RB-deficient tumors were found to express high levels of CHK1 and PLK1. Loss of RB specifically resulted in loss of checkpoint functions governing DNA replication, yielding increased drug sensitivity. Xenograft models demonstrated RB-selective efficacy of CHK inhibitors. This study supports the possibility of selectively targeting RB loss in the treatment of TNBC..
Pettitt, S.J.
Lord, C.J.
(2018). PARP inhibitors and breast cancer: highlights and hang-ups. Expert review of precision medicine and drug development,
Vol.3
(2),
pp. 83-94.
full text
Campbell, J.
Ryan, C.J.
Lord, C.J.
(2018). Identifying Genetic Dependencies in Cancer by Analyzing siRNA Screens in Tumor Cell Line Panels. Methods mol biol,
Vol.1711,
pp. 83-99.
show abstract
full text
Loss-of-function screening using RNA interference or CRISPR approaches can be used to identify genes that specific tumor cell lines depend upon for survival. By integrating the results from screens in multiple cell lines with molecular profiling data, it is possible to associate the dependence upon specific genes with particular molecular features (e.g., the mutation of a cancer driver gene, or transcriptional or proteomic signature). Here, using a panel of kinome-wide siRNA screens in osteosarcoma cell lines as an example, we describe a computational protocol for analyzing loss-of-function screens to identify genetic dependencies associated with particular molecular features. We describe the steps required to process the siRNA screen data, integrate the results with genotypic information to identify genetic dependencies, and finally the integration of protein-protein interaction data to interpret these dependencies..
Pettitt, S.J.
Krastev, D.B.
Pemberton, H.N.
Fontebasso, Y.
Frankum, J.
Rehman, F.L.
Brough, R.
Song, F.
Bajrami, I.
Rafiq, R.
Wallberg, F.
Kozarewa, I.
Fenwick, K.
Armisen-Garrido, J.
Swain, A.
Gulati, A.
Campbell, J.
Ashworth, A.
Lord, C.J.
(2017). Genome-wide barcoded transposon screen for cancer drug sensitivity in haploid mouse embryonic stem cells. Sci data,
Vol.4,
p. 170020.
show abstract
full text
We describe a screen for cellular response to drugs that makes use of haploid embryonic stem cells. We generated ten libraries of mutants with piggyBac gene trap transposon integrations, totalling approximately 100,000 mutant clones. Random barcode sequences were inserted into the transposon vector to allow the number of cells bearing each insertion to be measured by amplifying and sequencing the barcodes. These barcodes were associated with their integration sites by inverse PCR. We exposed these libraries to commonly used cancer drugs and profiled changes in barcode abundance by Ion Torrent sequencing in order to identify mutations that conferred sensitivity. Drugs tested included conventional chemotherapeutics as well as targeted inhibitors of topoisomerases, poly(ADP-ribose) polymerase (PARP), Hsp90 and WEE1..
Jansen, S.
Geuer, S.
Pfundt, R.
Brough, R.
Ghongane, P.
Herkert, J.C.
Marco, E.J.
Willemsen, M.H.
Kleefstra, T.
Hannibal, M.
Shieh, J.T.
Lynch, S.A.
Flinter, F.
FitzPatrick, D.R.
Gardham, A.
Bernhard, B.
Ragge, N.
Newbury-Ecob, R.
Bernier, R.
Kvarnung, M.
Magnusson, E.A.
Wessels, M.W.
van Slegtenhorst, M.A.
Monaghan, K.G.
de Vries, P.
Veltman, J.A.
Deciphering Developmental Disorders Study,
Lord, C.J.
Vissers, L.E.
de Vries, B.B.
(2017). De Novo Truncating Mutations in the Last and Penultimate Exons of PPM1D Cause an Intellectual Disability Syndrome. Am j hum genet,
Vol.100
(4),
pp. 650-658.
show abstract
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Intellectual disability (ID) is a highly heterogeneous disorder involving at least 600 genes, yet a genetic diagnosis remains elusive in ∼35%-40% of individuals with moderate to severe ID. Recent meta-analyses statistically analyzing de novo mutations in >7,000 individuals with neurodevelopmental disorders highlighted mutations in PPM1D as a possible cause of ID. PPM1D is a type 2C phosphatase that functions as a negative regulator of cellular stress-response pathways by mediating a feedback loop of p38-p53 signaling, thereby contributing to growth inhibition and suppression of stress-induced apoptosis. We identified 14 individuals with mild to severe ID and/or developmental delay and de novo truncating PPM1D mutations. Additionally, deep phenotyping revealed overlapping behavioral problems (ASD, ADHD, and anxiety disorders), hypotonia, broad-based gait, facial dysmorphisms, and periods of fever and vomiting. PPM1D is expressed during fetal brain development and in the adult brain. All mutations were located in the last or penultimate exon, suggesting escape from nonsense-mediated mRNA decay. Both PPM1D expression analysis and cDNA sequencing in EBV LCLs of individuals support the presence of a stable truncated transcript, consistent with this hypothesis. Exposure of cells derived from individuals with PPM1D truncating mutations to ionizing radiation resulted in normal p53 activation, suggesting that p53 signaling is unaffected. However, a cell-growth disadvantage was observed, suggesting a possible effect on the stress-response pathway. Thus, we show that de novo truncating PPM1D mutations in the last and penultimate exons cause syndromic ID, which provides additional insight into the role of cell-cycle checkpoint genes in neurodevelopmental disorders..
Goodall, J.
Mateo, J.
Yuan, W.
Mossop, H.
Porta, N.
Miranda, S.
Perez-Lopez, R.
Dolling, D.
Robinson, D.R.
Sandhu, S.
Fowler, G.
Ebbs, B.
Flohr, P.
Seed, G.
Rodrigues, D.N.
Boysen, G.
Bertan, C.
Atkin, M.
Clarke, M.
Crespo, M.
Figueiredo, I.
Riisnaes, R.
Sumanasuriya, S.
Rescigno, P.
Zafeiriou, Z.
Sharp, A.
Tunariu, N.
Bianchini, D.
Gillman, A.
Lord, C.J.
Hall, E.
Chinnaiyan, A.M.
Carreira, S.
de Bono, J.S.
TOPARP-A investigators,
(2017). Circulating Cell-Free DNA to Guide Prostate Cancer Treatment with PARP Inhibition. Cancer discov,
Vol.7
(9),
pp. 1006-1017.
show abstract
full text
Biomarkers for more precise patient care are needed in metastatic prostate cancer. We have reported a phase II trial (TOPARP-A) of the PARP inhibitor olaparib in metastatic prostate cancer, demonstrating antitumor activity associating with homologous recombination DNA repair defects. We now report targeted and whole-exome sequencing of serial circulating cell-free DNA (cfDNA) samples collected during this trial. Decreases in cfDNA concentration independently associated with outcome in multivariable analyses (HR for overall survival at week 8: 0.19; 95% CI, 0.06-0.56; P = 0.003). All tumor tissue somatic DNA repair mutations were detectable in cfDNA; allele frequency of somatic mutations decreased selectively in responding patients (χ2P < 0.001). At disease progression, following response to olaparib, multiple subclonal aberrations reverting germline and somatic DNA repair mutations (BRCA2, PALB2) back in frame emerged as mechanisms of resistance. These data support the role of liquid biopsies as a predictive, prognostic, response, and resistance biomarker in metastatic prostate cancer.Significance: We report prospectively planned, serial, cfDNA analyses from patients with metastatic prostate cancer treated on an investigator-initiated phase II trial of olaparib. These analyses provide predictive, prognostic, response, and resistance data with "second hit" mutations first detectable at disease progression, suggesting clonal evolution from treatment-selective pressure and platinum resistance. Cancer Discov; 7(9); 1006-17. ©2017 AACR.See related commentary by Domchek, p. 937See related article by Kondrashova et al., p. 984See related article by Quigley et al., p. 999This article is highlighted in the In This Issue feature, p. 920..
Dréan, A.
Williamson, C.T.
Brough, R.
Brandsma, I.
Menon, M.
Konde, A.
Garcia-Murillas, I.
Pemberton, H.N.
Frankum, J.
Rafiq, R.
Badham, N.
Campbell, J.
Gulati, A.
Turner, N.C.
Pettitt, S.J.
Ashworth, A.
Lord, C.J.
(2017). Modeling Therapy Resistance in BRCA1/2-Mutant Cancers. Mol cancer ther,
Vol.16
(9),
pp. 2022-2034.
show abstract
full text
Although PARP inhibitors target BRCA1- or BRCA2-mutant tumor cells, drug resistance is a problem. PARP inhibitor resistance is sometimes associated with the presence of secondary or "revertant" mutations in BRCA1 or BRCA2 Whether secondary mutant tumor cells are selected for in a Darwinian fashion by treatment is unclear. Furthermore, how PARP inhibitor resistance might be therapeutically targeted is also poorly understood. Using CRISPR mutagenesis, we generated isogenic tumor cell models with secondary BRCA1 or BRCA2 mutations. Using these in heterogeneous in vitro culture or in vivo xenograft experiments in which the clonal composition of tumor cell populations in response to therapy was monitored, we established that PARP inhibitor or platinum salt exposure selects for secondary mutant clones in a Darwinian fashion, with the periodicity of PARP inhibitor administration and the pretreatment frequency of secondary mutant tumor cells influencing the eventual clonal composition of the tumor cell population. In xenograft studies, the presence of secondary mutant cells in tumors impaired the therapeutic effect of a clinical PARP inhibitor. However, we found that both PARP inhibitor-sensitive and PARP inhibitor-resistant BRCA2 mutant tumor cells were sensitive to AZD-1775, a WEE1 kinase inhibitor. In mice carrying heterogeneous tumors, AZD-1775 delivered a greater therapeutic benefit than olaparib treatment. This suggests that despite the restoration of some BRCA1 or BRCA2 gene function in "revertant" tumor cells, vulnerabilities still exist that could be therapeutically exploited. Mol Cancer Ther; 16(9); 2022-34. ©2017 AACR..
Quigley, D.
Alumkal, J.J.
Wyatt, A.W.
Kothari, V.
Foye, A.
Lloyd, P.
Aggarwal, R.
Kim, W.
Lu, E.
Schwartzman, J.
Beja, K.
Annala, M.
Das, R.
Diolaiti, M.
Pritchard, C.
Thomas, G.
Tomlins, S.
Knudsen, K.
Lord, C.J.
Ryan, C.
Youngren, J.
Beer, T.M.
Ashworth, A.
Small, E.J.
Feng, F.Y.
(2017). Analysis of Circulating Cell-Free DNA Identifies Multiclonal Heterogeneity of BRCA2 Reversion Mutations Associated with Resistance to PARP Inhibitors. Cancer discov,
Vol.7
(9),
pp. 999-1005.
show abstract
full text
Approximately 20% of metastatic prostate cancers harbor mutations in genes required for DNA repair by homologous recombination repair (HRR) such as BRCA2 HRR defects confer synthetic lethality to PARP inhibitors (PARPi) such as olaparib and talazoparib. In ovarian or breast cancers, olaparib resistance has been associated with HRR restoration, including by BRCA2 mutation reversion. Whether similar mechanisms operate in prostate cancer, and could be detected in liquid biopsies, is unclear. Here, we identify BRCA2 reversion mutations associated with olaparib and talazoparib resistance in patients with prostate cancer. Analysis of circulating cell-free DNA (cfDNA) reveals reversion mutation heterogeneity not discernable from a single solid-tumor biopsy and potentially allows monitoring for the emergence of PARPi resistance.Significance: The mechanisms of clinical resistance to PARPi in DNA repair-deficient prostate cancer have not been described. Here, we show BRCA2 reversion mutations in patients with prostate cancer with metastatic disease who developed resistance to talazoparib and olaparib. Furthermore, we show that PARPi resistance is highly multiclonal and that cfDNA allows monitoring for PARPi resistance. Cancer Discov; 7(9); 999-1005. ©2017 AACR.See related commentary by Domchek, p. 937See related article by Kondrashova et al., p. 984See related article by Goodall et al., p. 1006This article is highlighted in the In This Issue feature, p. 920..
Brandsma, I.
Fleuren, E.D.
Williamson, C.T.
Lord, C.J.
(2017). Directing the use of DDR kinase inhibitors in cancer treatment. Expert opin investig drugs,
Vol.26
(12),
pp. 1341-1355.
show abstract
full text
INTRODUCTION: Defects in the DNA damage response (DDR) drive the development of cancer by fostering DNA mutation but also provide cancer-specific vulnerabilities that can be exploited therapeutically. The recent approval of three different PARP inhibitors for the treatment of ovarian cancer provides the impetus for further developing targeted inhibitors of many of the kinases involved in the DDR, including inhibitors of ATR, ATM, CHEK1, CHEK2, DNAPK and WEE1. Areas covered: We summarise the current stage of development of these novel DDR kinase inhibitors, and describe which predictive biomarkers might be exploited to direct their clinical use. Expert opinion: Novel DDR inhibitors present promising candidates in cancer treatment and have the potential to elicit synthetic lethal effects. In order to fully exploit their potential and maximize their utility, identifying highly penetrant predictive biomarkers of single agent and combinatorial DDR inhibitor sensitivity are critical. Identifying the optimal drug combination regimens that could used with DDR inhibitors is also a key objective..
Jones, S.E.
Fleuren, E.D.
Frankum, J.
Konde, A.
Williamson, C.T.
Krastev, D.B.
Pemberton, H.N.
Campbell, J.
Gulati, A.
Elliott, R.
Menon, M.
Selfe, J.L.
Brough, R.
Pettitt, S.J.
Niedzwiedz, W.
van der Graaf, W.T.
Shipley, J.
Ashworth, A.
Lord, C.J.
(2017). ATR Is a Therapeutic Target in Synovial Sarcoma. Cancer res,
Vol.77
(24),
pp. 7014-7026.
show abstract
full text
Synovial sarcoma (SS) is an aggressive soft-tissue malignancy characterized by expression of SS18-SSX fusions, where treatment options are limited. To identify therapeutically actionable genetic dependencies in SS, we performed a series of parallel, high-throughput small interfering RNA (siRNA) screens and compared genetic dependencies in SS tumor cells with those in >130 non-SS tumor cell lines. This approach revealed a reliance of SS tumor cells upon the DNA damage response serine/threonine protein kinase ATR. Clinical ATR inhibitors (ATRi) elicited a synthetic lethal effect in SS tumor cells and impaired growth of SS patient-derived xenografts. Oncogenic SS18-SSX family fusion genes are known to alter the composition of the BAF chromatin-remodeling complex, causing ejection and degradation of wild-type SS18 and the tumor suppressor SMARCB1. Expression of oncogenic SS18-SSX fusion proteins caused profound ATRi sensitivity and a reduction in SS18 and SMARCB1 protein levels, but an SSX18-SSX1 Δ71-78 fusion containing a C-terminal deletion did not. ATRi sensitivity in SS was characterized by an increase in biomarkers of replication fork stress (increased γH2AX, decreased replication fork speed, and increased R-loops), an apoptotic response, and a dependence upon cyclin E expression. Combinations of cisplatin or PARP inhibitors enhanced the antitumor cell effect of ATRi, suggesting that either single-agent ATRi or combination therapy involving ATRi might be further assessed as candidate approaches for SS treatment. Cancer Res; 77(24); 7014-26. ©2017 AACR..
Rivera, B.
Di Iorio, M.
Frankum, J.
Nadaf, J.
Fahiminiya, S.
Arcand, S.L.
Burk, D.L.
Grapton, D.
Tomiak, E.
Hastings, V.
Hamel, N.
Wagener, R.
Aleynikova, O.
Giroux, S.
Hamdan, F.F.
Dionne-Laporte, A.
Zogopoulos, G.
Rousseau, F.
Berghuis, A.M.
Provencher, D.
Rouleau, G.A.
Michaud, J.L.
Mes-Masson, A.-.
Majewski, J.
Bens, S.
Siebert, R.
Narod, S.A.
Akbari, M.R.
Lord, C.J.
Tonin, P.N.
Orthwein, A.
Foulkes, W.D.
(2017). Functionally Null RAD51D Missense Mutation Associates Strongly with Ovarian Carcinoma. Cancer res,
Vol.77
(16),
pp. 4517-4529.
show abstract
full text
RAD51D is a key player in DNA repair by homologous recombination (HR), and RAD51D truncating variant carriers have an increased risk for ovarian cancer. However, the contribution of nontruncating RAD51D variants to cancer predisposition remains uncertain. Using deep sequencing and case-control genotyping studies, we show that in French Canadians, the missense RAD51D variant c.620C>T;p.S207L is highly prevalent and is associated with a significantly increased risk for ovarian high-grade serous carcinoma (HGSC; 3.8% cases vs. 0.2% controls). The frequency of the p.S207L variant did not significantly differ from that of controls in breast, endometrial, pancreas, or colorectal adenocarcinomas. Functionally, we show that this mutation impairs HR by disrupting the RAD51D-XRCC2 interaction and confers PARP inhibitor sensitivity. These results highlight the importance of a functional RAD51D-XRCC2 interaction to promote HR and prevent the development of HGSC. This study identifies c.620C>T;p.S207L as the first bona fide pathogenic RAD51D missense cancer susceptibility allele and supports the use of targeted PARP-inhibitor therapies in ovarian cancer patients carrying deleterious missense RAD51D variants. Cancer Res; 77(16); 4517-29. ©2017 AACR..
Fleuren, E.D.
Vlenterie, M.
van der Graaf, W.T.
Hillebrandt-Roeffen, M.H.
Blackburn, J.
Ma, X.
Chan, H.
Magias, M.C.
van Erp, A.
van Houdt, L.
Cebeci, S.A.
van de Ven, A.
Flucke, U.E.
Heyer, E.E.
Thomas, D.M.
Lord, C.J.
Marini, K.D.
Vaghjiani, V.
Mercer, T.R.
Cain, J.E.
Wu, J.
Versleijen-Jonkers, Y.M.
Daly, R.J.
(2017). Phosphoproteomic Profiling Reveals ALK and MET as Novel Actionable Targets across Synovial Sarcoma Subtypes. Cancer res,
Vol.77
(16),
pp. 4279-4292.
show abstract
full text
Despite intensive multimodal treatment of sarcomas, a heterogeneous group of malignant tumors arising from connective tissue, survival remains poor. Candidate-based targeted treatments have demonstrated limited clinical success, urging an unbiased and comprehensive analysis of oncogenic signaling networks to reveal therapeutic targets and personalized treatment strategies. Here we applied mass spectrometry-based phosphoproteomic profiling to the largest and most heterogeneous set of sarcoma cell lines characterized to date and identified novel tyrosine phosphorylation patterns, enhanced tyrosine kinases in specific subtypes, and potential driver kinases. ALK was identified as a novel driver in the Aska-SS synovial sarcoma (SS) cell line via expression of an ALK variant with a large extracellular domain deletion (ALKΔ2-17). Functional ALK dependency was confirmed in vitro and in vivo with selective inhibitors. Importantly, ALK immunopositivity was detected in 6 of 43 (14%) of SS patient specimens, one of which exhibited an ALK rearrangement. High PDGFRα phosphorylation also characterized SS cell lines, which was accompanied by enhanced MET activation in Yamato-SS cells. Although Yamato-SS cells were sensitive to crizotinib (ALK/MET-inhibitor) but not pazopanib (VEGFR/PDGFR-inhibitor) monotherapy in vitro, synergistic effects were observed upon drug combination. In vivo, both drugs were individually effective, with pazopanib efficacy likely attributable to reduced angiogenesis. MET or PDGFRα expression was detected in 58% and 84% of SS patients, respectively, with coexpression in 56%. Consequently, our integrated approach has led to the identification of ALK and MET as promising therapeutic targets in SS. Cancer Res; 77(16); 4279-92. ©2017 AACR..
Ryan, C.J.
Kennedy, S.
Bajrami, I.
Matallanas, D.
Lord, C.J.
(2017). A Compendium of Co-regulated Protein Complexes in Breast Cancer Reveals Collateral Loss Events. Cell syst,
Vol.5
(4),
pp. 399-409.e5.
show abstract
full text
Protein complexes are responsible for the bulk of activities within the cell, but how their behavior and abundance varies across tumors remains poorly understood. By combining proteomic profiles of breast tumors with a large-scale protein-protein interaction network, we have identified a set of 285 high-confidence protein complexes whose subunits have highly correlated protein abundance across tumor samples. We used this set to identify complexes that are reproducibly under- or overexpressed in specific breast cancer subtypes. We found that mutation or deletion of one subunit of a co-regulated complex was often associated with a collateral reduction in protein expression of additional complex members. This collateral loss phenomenon was typically evident from proteomic, but not transcriptomic, profiles, suggesting post-transcriptional control. Mutation of the tumor suppressor E-cadherin (CDH1) was associated with a collateral loss of members of the adherens junction complex, an effect we validated using an engineered model of E-cadherin loss..
Bridgett, S.
Campbell, J.
Lord, C.J.
Ryan, C.J.
(2017). CancerGD: A Resource for Identifying and Interpreting Genetic Dependencies in Cancer. Cell syst,
Vol.5
(1),
pp. 82-86.e3.
show abstract
full text
Genes whose function is selectively essential in the presence of cancer-associated genetic aberrations represent promising targets for the development of precision therapeutics. Here, we present CancerGD, a resource that integrates genotypic profiling with large-scale loss-of-function genetic screens in tumor cell lines to identify such genetic dependencies. CancerGD provides tools for searching, visualizing, and interpreting these genetic dependencies through the integration of functional interaction networks. CancerGD includes different screen types (siRNA, shRNA, CRISPR), and we describe a simple format for submitting new datasets..
Nikkilä, J.
Kumar, R.
Campbell, J.
Brandsma, I.
Pemberton, H.N.
Wallberg, F.
Nagy, K.
Scheer, I.
Vertessy, B.G.
Serebrenik, A.A.
Monni, V.
Harris, R.S.
Pettitt, S.J.
Ashworth, A.
Lord, C.J.
(2017). Elevated APOBEC3B expression drives a kataegic-like mutation signature and replication stress-related therapeutic vulnerabilities in p53-defective cells. Br j cancer,
Vol.117
(1),
pp. 113-123.
show abstract
full text
BACKGROUND: Elevated APOBEC3B expression in tumours correlates with a kataegic pattern of localised hypermutation. We assessed the cellular phenotypes associated with high-level APOBEC3B expression and the influence of p53 status on these phenotypes using an isogenic system. METHODS: We used RNA interference of p53 in cells with inducible APOBEC3B and assessed DNA damage response (DDR) biomarkers. The mutational effects of APOBEC3B were assessed using whole-genome sequencing. In vitro small-molecule inhibitor sensitivity profiling was used to identify candidate therapeutic vulnerabilities. RESULTS: Although APOBEC3B expression increased the incorporation of genomic uracil, invoked DDR biomarkers and caused cell cycle arrest, inactivation of p53 circumvented APOBEC3B-induced cell cycle arrest without reversing the increase in genomic uracil or DDR biomarkers. The continued expression of APOBEC3B in p53-defective cells not only caused a kataegic mutational signature but also caused hypersensitivity to small-molecule DDR inhibitors (ATR, CHEK1, CHEK2, PARP, WEE1 inhibitors) as well as cisplatin/ATR inhibitor and ATR/PARP inhibitor combinations. CONCLUSIONS: Although loss of p53 might allow tumour cells to tolerate elevated APOBEC3B expression, continued expression of this enzyme might impart a number of therapeutic vulnerabilities upon tumour cells..
Lord, C.J.
Ashworth, A.
(2016). BRCAness revisited. Nat rev cancer,
Vol.16
(2),
pp. 110-120.
show abstract
Over the past 20 years, there has been considerable progress in our understanding of the biological functions of the BRCA1 and BRCA2 cancer susceptibility genes. This has led to the development of new therapeutic approaches that target tumours with loss-of-function mutations in either BRCA1 or BRCA2. Tumours that share molecular features of BRCA-mutant tumours - that is, those with 'BRCAness' - may also respond to similar therapeutic approaches. Several paradigm shifts require a reassessment of the concept of BRCAness, how this property is assayed and its relevance to our understanding of tumour biology and the treatment of cancer..
Beck, D.
Zobel, J.
Barber, R.
Evans, S.
Lezina, L.
Allchin, R.L.
Blades, M.
Elliott, R.
Lord, C.J.
Ashworth, A.
Porter, A.C.
Wagner, S.D.
(2016). Synthetic Lethal Screen Demonstrates That a JAK2 Inhibitor Suppresses a BCL6-dependent IL10RA/JAK2/STAT3 Pathway in High Grade B-cell Lymphoma. J biol chem,
Vol.291
(32),
pp. 16686-16698.
show abstract
full text
We demonstrate the usefulness of synthetic lethal screening of a conditionally BCL6-deficient Burkitt lymphoma cell line, DG75-AB7, with a library of small molecules to determine survival pathways suppressed by BCL6 and suggest mechanism-based treatments for lymphoma. Lestaurtinib, a JAK2 inhibitor and one of the hits from the screen, repressed survival of BCL6-deficient cells in vitro and reduced growth and proliferation of xenografts in vivo BCL6 deficiency in DG75-AB7 induced JAK2 mRNA and protein expression and STAT3 phosphorylation. Surface IL10RA was elevated by BCL6 deficiency, and blockade of IL10RA repressed STAT3 phosphorylation. Therefore, we define an IL10RA/JAK2/STAT3 pathway each component of which is repressed by BCL6. We also show for the first time that JAK2 is a direct BCL6 target gene; BCL6 bound to the JAK2 promoter in vitro and was enriched by ChIP-seq. The place of JAK2 inhibitors in the treatment of diffuse large B-cell lymphoma has not been defined; we suggest that JAK2 inhibitors might be most effective in poor prognosis ABC-DLBCL, which shows higher levels of IL10RA, JAK2, and STAT3 but lower levels of BCL6 than GC-DLBCL and might be usefully combined with novel approaches such as inhibition of IL10RA..
Rajan, N.
Andersson, M.K.
Sinclair, N.
Fehr, A.
Hodgson, K.
Lord, C.J.
Kazakov, D.V.
Vanecek, T.
Ashworth, A.
Stenman, G.
(2016). Overexpression of MYB drives proliferation of CYLD-defective cylindroma cells. J pathol,
Vol.239
(2),
pp. 197-205.
show abstract
full text
Cutaneous cylindroma is an adnexal tumour with apocrine differentiation. A predisposition to multiple cylindromas is seen in patients with Brooke-Spiegler syndrome, who carry germline mutations in the tumour suppressor gene CYLD. Previous studies of inherited cylindromas have highlighted the frequent presence of bi-allelic truncating CYLD mutations as a recurrent driver mutation. We have previously shown that sporadic cylindromas express either MYB-NFIB fusion transcripts or show evidence of MYB activation in the absence of such fusions. Here, we investigated inherited cylindromas from several families with germline CYLD mutations for the presence of MYB activation. Strikingly, none of the inherited CYLD-defective (n = 23) tumours expressed MYB-NFIB fusion transcripts. However, MYB expression was increased in the majority of tumours (69%) and global gene expression analysis revealed that well-established MYB target genes were up-regulated in CYLD-defective tumours. Moreover, knock-down of MYB expression caused a significant reduction in cylindroma cell proliferation, suggesting that MYB is also a key player and oncogenic driver in inherited cylindromas. Taken together, our findings suggest molecular heterogeneity in the pathogenesis of sporadic and inherited cutaneous cylindromas, with convergence on MYB activation. © 2016 The Authors. The Journal of Pathology published by John Wiley & Sons Ltd on behalf of Pathological Society of Great Britain and Ireland..
Miller, R.E.
Brough, R.
Bajrami, I.
Williamson, C.T.
McDade, S.
Campbell, J.
Kigozi, A.
Rafiq, R.
Pemberton, H.
Natrajan, R.
Joel, J.
Astley, H.
Mahoney, C.
Moore, J.D.
Torrance, C.
Gordan, J.D.
Webber, J.T.
Levin, R.S.
Shokat, K.M.
Bandyopadhyay, S.
Lord, C.J.
Ashworth, A.
(2016). Synthetic Lethal Targeting of ARID1A-Mutant Ovarian Clear Cell Tumors with Dasatinib. Mol cancer ther,
Vol.15
(7),
pp. 1472-1484.
show abstract
full text
New targeted approaches to ovarian clear cell carcinomas (OCCC) are needed, given the limited treatment options in this disease and the poor response to standard chemotherapy. Using a series of high-throughput cell-based drug screens in OCCC tumor cell models, we have identified a synthetic lethal (SL) interaction between the kinase inhibitor dasatinib and a key driver in OCCC, ARID1A mutation. Imposing ARID1A deficiency upon a variety of human or mouse cells induced dasatinib sensitivity, both in vitro and in vivo, suggesting that this is a robust synthetic lethal interaction. The sensitivity of ARID1A-deficient cells to dasatinib was associated with G1-S cell-cycle arrest and was dependent upon both p21 and Rb. Using focused siRNA screens and kinase profiling, we showed that ARID1A-mutant OCCC tumor cells are addicted to the dasatinib target YES1. This suggests that dasatinib merits investigation for the treatment of patients with ARID1A-mutant OCCC. Mol Cancer Ther; 15(7); 1472-84. ©2016 AACR..
Dhillon, K.K.
Bajrami, I.
Taniguchi, T.
Lord, C.J.
(2016). Synthetic lethality: the road to novel therapies for breast cancer. Endocr relat cancer,
Vol.23
(10),
pp. T39-T55.
show abstract
full text
When the BRCA1 and BRCA2 tumour suppressor genes were identified in the early 1990s, the immediate implications of mapping, cloning and delineating the sequence of these genes were that individuals in families with a BRCA gene mutation could be tested for the presence of a mutation and their risk of developing cancer could be predicted. Over time though, the discovery of BRCA1 and BRCA2 has had a much greater influence than many might have imagined. In this review, we discuss how the discovery of BRCA1 and BRCA2 has not only provided an understanding of the molecular processes that drive tumourigenesis but also reignited an interest in therapeutically exploiting loss-of-function alterations in tumour suppressor genes..
Maguire, S.L.
Peck, B.
Wai, P.T.
Campbell, J.
Barker, H.
Gulati, A.
Daley, F.
Vyse, S.
Huang, P.
Lord, C.J.
Farnie, G.
Brennan, K.
Natrajan, R.
(2016). Three-dimensional modelling identifies novel genetic dependencies associated with breast cancer progression in the isogenic MCF10 model. J pathol,
Vol.240
(3),
pp. 315-328.
show abstract
full text
The initiation and progression of breast cancer from the transformation of the normal epithelium to ductal carcinoma in situ (DCIS) and invasive disease is a complex process involving the acquisition of genetic alterations and changes in gene expression, alongside microenvironmental and recognized histological alterations. Here, we sought to comprehensively characterise the genomic and transcriptomic features of the MCF10 isogenic model of breast cancer progression, and to functionally validate potential driver alterations in three-dimensional (3D) spheroids that may provide insights into breast cancer progression, and identify targetable alterations in conditions more similar to those encountered in vivo. We performed whole genome, exome and RNA sequencing of the MCF10 progression series to catalogue the copy number and mutational and transcriptomic landscapes associated with progression. We identified a number of predicted driver mutations (including PIK3CA and TP53) that were acquired during transformation of non-malignant MCF10A cells to their malignant counterparts that are also present in analysed primary breast cancers from The Cancer Genome Atlas (TCGA). Acquisition of genomic alterations identified MYC amplification and previously undescribed RAB3GAP1-HRAS and UBA2-PDCD2L expressed in-frame fusion genes in malignant cells. Comparison of pathway aberrations associated with progression showed that, when cells are grown as 3D spheroids, they show perturbations of cancer-relevant pathways. Functional interrogation of the dependency on predicted driver events identified alterations in HRAS, PIK3CA and TP53 that selectively decreased cell growth and were associated with progression from preinvasive to invasive disease only when cells were grown as spheroids. Our results have identified changes in the genomic repertoire in cell lines representative of the stages of breast cancer progression, and demonstrate that genetic dependencies can be uncovered when cells are grown in conditions more like those in vivo. The MCF10 progression series therefore represents a good model with which to dissect potential biomarkers and to evaluate therapeutic targets involved in the progression of breast cancer. © 2016 The Authors. The Journal of Pathology published by John Wiley & Sons Ltd on behalf of Pathological Society of Great Britain and Ireland..
Dréan, A.
Lord, C.J.
Ashworth, A.
(2016). PARP inhibitor combination therapy. Crit rev oncol hematol,
Vol.108,
pp. 73-85.
show abstract
full text
In 2014, olaparib (Lynparza) became the first PARP (Poly(ADP-ribose) polymerase) inhibitor to be approved for the treatment of cancer. When used as single agents, PARP inhibitors can selectively target tumour cells with BRCA1 or BRCA2 tumour suppressor gene mutations through synthetic lethality. However, PARP inhibition also shows considerable promise when used together with other therapeutic agents. Here, we summarise both the pre-clinical and clinical evidence for the utility of such combinations and discuss the future prospects and challenges for PARP inhibitor combinatorial therapies..
Williamson, C.T.
Miller, R.
Pemberton, H.N.
Jones, S.E.
Campbell, J.
Konde, A.
Badham, N.
Rafiq, R.
Brough, R.
Gulati, A.
Ryan, C.J.
Francis, J.
Vermulen, P.B.
Reynolds, A.R.
Reaper, P.M.
Pollard, J.R.
Ashworth, A.
Lord, C.J.
(2016). ATR inhibitors as a synthetic lethal therapy for tumours deficient in ARID1A. Nat commun,
Vol.7,
p. 13837.
show abstract
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Identifying genetic biomarkers of synthetic lethal drug sensitivity effects provides one approach to the development of targeted cancer therapies. Mutations in ARID1A represent one of the most common molecular alterations in human cancer, but therapeutic approaches that target these defects are not yet clinically available. We demonstrate that defects in ARID1A sensitize tumour cells to clinical inhibitors of the DNA damage checkpoint kinase, ATR, both in vitro and in vivo. Mechanistically, ARID1A deficiency results in topoisomerase 2A and cell cycle defects, which cause an increased reliance on ATR checkpoint activity. In ARID1A mutant tumour cells, inhibition of ATR triggers premature mitotic entry, genomic instability and apoptosis. The data presented here provide the pre-clinical and mechanistic rationale for assessing ARID1A defects as a biomarker of single-agent ATR inhibitor response and represents a novel synthetic lethal approach to targeting tumour cells..
Campbell, J.
Ryan, C.J.
Brough, R.
Bajrami, I.
Pemberton, H.N.
Chong, I.Y.
Costa-Cabral, S.
Frankum, J.
Gulati, A.
Holme, H.
Miller, R.
Postel-Vinay, S.
Rafiq, R.
Wei, W.
Williamson, C.T.
Quigley, D.A.
Tym, J.
Al-Lazikani, B.
Fenton, T.
Natrajan, R.
Strauss, S.J.
Ashworth, A.
Lord, C.J.
(2016). Large-Scale Profiling of Kinase Dependencies in Cancer Cell Lines. Cell rep,
Vol.14
(10),
pp. 2490-2501.
show abstract
full text
One approach to identifying cancer-specific vulnerabilities and therapeutic targets is to profile genetic dependencies in cancer cell lines. Here, we describe data from a series of siRNA screens that identify the kinase genetic dependencies in 117 cancer cell lines from ten cancer types. By integrating the siRNA screen data with molecular profiling data, including exome sequencing data, we show how vulnerabilities/genetic dependencies that are associated with mutations in specific cancer driver genes can be identified. By integrating additional data sets into this analysis, including protein-protein interaction data, we also demonstrate that the genetic dependencies associated with many cancer driver genes form dense connections on functional interaction networks. We demonstrate the utility of this resource by using it to predict the drug sensitivity of genetically or histologically defined subsets of tumor cell lines, including an increased sensitivity of osteosarcoma cell lines to FGFR inhibitors and SMAD4 mutant tumor cells to mitotic inhibitors..
Herrera-Abreu, M.T.
Palafox, M.
Asghar, U.
Rivas, M.A.
Cutts, R.J.
Garcia-Murillas, I.
Pearson, A.
Guzman, M.
Rodriguez, O.
Grueso, J.
Bellet, M.
Cortés, J.
Elliott, R.
Pancholi, S.
Baselga, J.
Dowsett, M.
Martin, L.-.
Turner, N.C.
Serra, V.
(2016). Early Adaptation and Acquired Resistance to CDK4/6 Inhibition in Estrogen Receptor-Positive Breast Cancer. Cancer res,
Vol.76
(8),
pp. 2301-2313.
show abstract
Small-molecule inhibitors of the CDK4/6 cell-cycle kinases have shown clinical efficacy in estrogen receptor (ER)-positive metastatic breast cancer, although their cytostatic effects are limited by primary and acquired resistance. Here we report that ER-positive breast cancer cells can adapt quickly to CDK4/6 inhibition and evade cytostasis, in part, via noncanonical cyclin D1-CDK2-mediated S-phase entry. This adaptation was prevented by cotreatment with hormone therapies or PI3K inhibitors, which reduced the levels of cyclin D1 (CCND1) and other G1-S cyclins, abolished pRb phosphorylation, and inhibited activation of S-phase transcriptional programs. Combined targeting of both CDK4/6 and PI3K triggered cancer cell apoptosis in vitro and in patient-derived tumor xenograft (PDX) models, resulting in tumor regression and improved disease control. Furthermore, a triple combination of endocrine therapy, CDK4/6, and PI3K inhibition was more effective than paired combinations, provoking rapid tumor regressions in a PDX model. Mechanistic investigations showed that acquired resistance to CDK4/6 inhibition resulted from bypass of cyclin D1-CDK4/6 dependency through selection of CCNE1 amplification or RB1 loss. Notably, although PI3K inhibitors could prevent resistance to CDK4/6 inhibitors, they failed to resensitize cells once resistance had been acquired. However, we found that cells acquiring resistance to CDK4/6 inhibitors due to CCNE1 amplification could be resensitized by targeting CDK2. Overall, our results illustrate convergent mechanisms of early adaptation and acquired resistance to CDK4/6 inhibitors that enable alternate means of S-phase entry, highlighting strategies to prevent the acquisition of therapeutic resistance to these agents. Cancer Res; 76(8); 2301-13. ©2016 AACR..
Costa-Cabral, S.
Brough, R.
Konde, A.
Aarts, M.
Campbell, J.
Marinari, E.
Riffell, J.
Bardelli, A.
Torrance, C.
Lord, C.J.
Ashworth, A.
(2016). CDK1 Is a Synthetic Lethal Target for KRAS Mutant Tumours. Plos one,
Vol.11
(2),
p. e0149099.
show abstract
full text
Activating KRAS mutations are found in approximately 20% of human cancers but no RAS-directed therapies are currently available. Here we describe a novel, robust, KRAS synthetic lethal interaction with the cyclin dependent kinase, CDK1. This was discovered using parallel siRNA screens in KRAS mutant and wild type colorectal isogenic tumour cells and subsequently validated in a genetically diverse panel of 26 colorectal and pancreatic tumour cell models. This established that the KRAS/CDK1 synthetic lethality applies in tumour cells with either amino acid position 12 (p.G12V, pG12D, p.G12S) or amino acid position 13 (p.G13D) KRAS mutations and can also be replicated in vivo in a xenograft model using a small molecule CDK1 inhibitor. Mechanistically, CDK1 inhibition caused a reduction in the S-phase fraction of KRAS mutant cells, an effect also characterised by modulation of Rb, a master control of the G1/S checkpoint. Taken together, these observations suggest that the KRAS/CDK1 interaction is a robust synthetic lethal effect worthy of further investigation..
Vollan, H.K.
Rueda, O.M.
Chin, S.-.
Curtis, C.
Turashvili, G.
Shah, S.
Lingjærde, O.C.
Yuan, Y.
Ng, C.K.
Dunning, M.J.
Dicks, E.
Provenzano, E.
Sammut, S.
McKinney, S.
Ellis, I.O.
Pinder, S.
Purushotham, A.
Murphy, L.C.
Kristensen, V.N.
METABRIC Group,
Brenton, J.D.
Pharoah, P.D.
Børresen-Dale, A.-.
Aparicio, S.
Caldas, C.
(2015). A tumor DNA complex aberration index is an independent predictor of survival in breast and ovarian cancer. Mol oncol,
Vol.9
(1),
pp. 115-127.
show abstract
full text
Complex focal chromosomal rearrangements in cancer genomes, also called "firestorms", can be scored from DNA copy number data. The complex arm-wise aberration index (CAAI) is a score that captures DNA copy number alterations that appear as focal complex events in tumors, and has potential prognostic value in breast cancer. This study aimed to validate this DNA-based prognostic index in breast cancer and test for the first time its potential prognostic value in ovarian cancer. Copy number alteration (CNA) data from 1950 breast carcinomas (METABRIC cohort) and 508 high-grade serous ovarian carcinomas (TCGA dataset) were analyzed. Cases were classified as CAAI positive if at least one complex focal event was scored. Complex alterations were frequently localized on chromosome 8p (n = 159), 17q (n = 176) and 11q (n = 251). CAAI events on 11q were most frequent in estrogen receptor positive (ER+) cases and on 17q in estrogen receptor negative (ER-) cases. We found only a modest correlation between CAAI and the overall rate of genomic instability (GII) and number of breakpoints (r = 0.27 and r = 0.42, p < 0.001). Breast cancer specific survival (BCSS), overall survival (OS) and ovarian cancer progression free survival (PFS) were used as clinical end points in Cox proportional hazard model survival analyses. CAAI positive breast cancers (43%) had higher mortality: hazard ratio (HR) of 1.94 (95%CI, 1.62-2.32) for BCSS, and of 1.49 (95%CI, 1.30-1.71) for OS. Representations of the 70-gene and the 21-gene predictors were compared with CAAI in multivariable models and CAAI was independently significant with a Cox adjusted HR of 1.56 (95%CI, 1.23-1.99) for ER+ and 1.55 (95%CI, 1.11-2.18) for ER- disease. None of the expression-based predictors were prognostic in the ER- subset. We found that a model including CAAI and the two expression-based prognostic signatures outperformed a model including the 21-gene and 70-gene signatures but excluding CAAI. Inclusion of CAAI in the clinical prognostication tool PREDICT significantly improved its performance. CAAI positive ovarian cancers (52%) also had worse prognosis: HRs of 1.3 (95%CI, 1.1-1.7) for PFS and 1.3 (95%CI, 1.1-1.6) for OS. This study validates CAAI as an independent predictor of survival in both ER+ and ER- breast cancer and reveals a significant prognostic value for CAAI in high-grade serous ovarian cancer..
Aarts, M.
Bajrami, I.
Herrera-Abreu, M.T.
Elliott, R.
Brough, R.
Ashworth, A.
Lord, C.J.
Turner, N.C.
(2015). Functional Genetic Screen Identifies Increased Sensitivity to WEE1 Inhibition in Cells with Defects in Fanconi Anemia and HR Pathways. Mol cancer ther,
Vol.14
(4),
pp. 865-876.
show abstract
full text
WEE1 kinase regulates CDK1 and CDK2 activity to facilitate DNA replication during S-phase and to prevent unscheduled entry into mitosis. WEE1 inhibitors synergize with DNA-damaging agents that arrest cells in S-phase by triggering direct mitotic entry without completing DNA synthesis, resulting in catastrophic chromosome fragmentation and apoptosis. Here, we investigated how WEE1 inhibition could be best exploited for cancer therapy by performing a functional genetic screen to identify novel determinants of sensitivity to WEE1 inhibition. Inhibition of kinases that regulate CDK activity, CHK1 and MYT1, synergized with WEE1 inhibition through both increased replication stress and forced mitotic entry of S-phase cells. Loss of multiple components of the Fanconi anemia (FA) and homologous recombination (HR) pathways, in particular DNA helicases, sensitized to WEE1 inhibition. Silencing of FA/HR genes resulted in excessive replication stress and nucleotide depletion following WEE1 inhibition, which ultimately led to increased unscheduled mitotic entry. Our results suggest that cancers with defects in FA and HR pathways may be targeted by WEE1 inhibition, providing a basis for a novel synthetic lethal strategy for cancers harboring FA/HR defects..
Watkins, J.
Weekes, D.
Shah, V.
Gazinska, P.
Joshi, S.
Sidhu, B.
Gillett, C.
Pinder, S.
Vanoli, F.
Jasin, M.
Mayrhofer, M.
Isaksson, A.
Cheang, M.C.
Mirza, H.
Frankum, J.
Lord, C.J.
Ashworth, A.
Vinayak, S.
Ford, J.M.
Telli, M.L.
Grigoriadis, A.
Tutt, A.N.
(2015). Genomic Complexity Profiling Reveals That HORMAD1 Overexpression Contributes to Homologous Recombination Deficiency in Triple-Negative Breast Cancers. Cancer discov,
Vol.5
(5),
pp. 488-505.
show abstract
full text
UNLABELLED: Triple-negative breast cancers (TNBC) are characterized by a wide spectrum of genomic alterations, some of which might be caused by defects in DNA repair processes such as homologous recombination (HR). Despite this understanding, associating particular patterns of genomic instability with response to therapy has been challenging. Here, we show that allelic-imbalanced copy-number aberrations (AiCNA) are more prevalent in TNBCs that respond to platinum-based chemotherapy, thus providing a candidate predictive biomarker for this disease. Furthermore, we show that a high level of AiCNA is linked with elevated expression of a meiosis-associated gene, HORMAD1. Elevated HORMAD1 expression suppresses RAD51-dependent HR and drives the use of alternative forms of DNA repair, the generation of AiCNAs, as well as sensitizing cancer cells to HR-targeting therapies. Our data therefore provide a mechanistic association between HORMAD1 expression, a specific pattern of genomic instability, and an association with response to platinum-based chemotherapy in TNBC. SIGNIFICANCE: Previous studies have shown correlation between mutational "scars" and sensitivity to platinums extending beyond associations with BRCA1/2 mutation, but do not elucidate the mechanism. Here, a novel allele-specific copy-number characterization of genome instability identifies and functionally validates the inappropriate expression of the meiotic gene HORMAD1 as a driver of HR deficiency in TNBC, acting to induce allelic imbalance and moderate platinum and PARP inhibitor sensitivity with implications for the use of such "scars" and expression of meiotic genes as predictive biomarkers..
Francis, J.C.
Melchor, L.
Campbell, J.
Kendrick, H.
Wei, W.
Armisen-Garrido, J.
Assiotis, I.
Chen, L.
Kozarewa, I.
Fenwick, K.
Swain, A.
Smalley, M.J.
Lord, C.J.
Ashworth, A.
(2015). Whole-exome DNA sequence analysis of Brca2- and Trp53-deficient mouse mammary gland tumours. J pathol,
Vol.236
(2),
pp. 186-200.
show abstract
Germline mutations in the tumour suppressor BRCA2 predispose to breast, ovarian and a number of other human cancers. Brca2-deficient mouse models are used for preclinical studies but the pattern of genomic alterations in these tumours has not yet been described in detail. We have performed whole-exome DNA sequencing analysis of mouse mammary tumours from Blg-Cre Brca2(f/f) Trp53(f/f) animals, a model of BRCA2-deficient human cancer. We also used the sequencing data to estimate DNA copy number alterations in these tumours and identified a recurrent copy number gain in Met, which has been found amplified in other mouse mammary cancer models. Through a comparative genomic analysis, we identified several mouse Blg-Cre Brca2(f/f) Trp53(f/f) mammary tumour somatic mutations in genes that are also mutated in human cancer, but few of these genes have been found frequently mutated in human breast cancer. A more detailed analysis of these somatic mutations revealed a set of genes that are mutated in human BRCA2 mutant breast and ovarian tumours and that are also mutated in mouse Brca2-null, Trp53-null mammary tumours. Finally, a DNA deletion surrounded by microhomology signature found in human BRCA1/2-deficient cancers was not common in the genome of these mouse tumours. Although a useful model, there are some differences in the genomic landscape of tumours arising in Blg-Cre Brca2(f/f) Trp53(f/f) mice compared to human BRCA-mutated breast cancers. Therefore, this needs to be taken into account in the use of this model..
Kwon, Y.-.
Petrie, K.
Leibovitch, B.A.
Zeng, L.
Mezei, M.
Howell, L.
Gil, V.
Christova, R.
Bansal, N.
Yang, S.
Sharma, R.
Ariztia, E.V.
Frankum, J.
Brough, R.
Sbirkov, Y.
Ashworth, A.
Lord, C.J.
Zelent, A.
Farias, E.
Zhou, M.-.
Waxman, S.
(2015). Selective Inhibition of SIN3 Corepressor with Avermectins as a Novel Therapeutic Strategy in Triple-Negative Breast Cancer. Mol cancer ther,
Vol.14
(8),
pp. 1824-1836.
show abstract
full text
Triple-negative breast cancers (TNBC) lacking estrogen, progesterone, and HER2 receptors account for 10% to 20% of breast cancer and are indicative of poor prognosis. The development of effective treatment strategies therefore represents a pressing unmet clinical need. We previously identified a molecularly targeted approach to target aberrant epigenetics of TNBC using a peptide corresponding to the SIN3 interaction domain (SID) of MAD. SID peptide selectively blocked binding of SID-containing proteins to the paired α-helix (PAH2) domain of SIN3, resulting in epigenetic and transcriptional modulation of genes associated with epithelial-mesenchymal transition (EMT). To find small molecule inhibitor (SMI) mimetics of SID peptide, we performed an in silico screen for PAH2 domain-binding compounds. This led to the identification of the avermectin macrocyclic lactone derivatives selamectin and ivermectin (Mectizan) as candidate compounds. Both selamectin and ivermectin phenocopied the effects of SID peptide to block SIN3-PAH2 interaction with MAD, induce expression of CDH1 and ESR1, and restore tamoxifen sensitivity in MDA-MB-231 human and MMTV-Myc mouse TNBC cells in vitro. Treatment with selamectin or ivermectin led to transcriptional modulation of genes associated with EMT and maintenance of a cancer stem cell phenotype in TNBC cells. This resulted in impairment of clonogenic self-renewal in vitro and inhibition of tumor growth and metastasis in vivo. Underlining the potential of avermectins in TNBC, pathway analysis revealed that selamectin also modulated the expression of therapeutically targetable genes. Consistent with this, an unbiased drug screen in TNBC cells identified selamectin-induced sensitization to a number of drugs, including those targeting modulated genes..
Frankum, J.
Moudry, P.
Brough, R.
Hodny, Z.
Ashworth, A.
Bartek, J.
Lord, C.J.
(2015). Complementary genetic screens identify the E3 ubiquitin ligase CBLC, as a modifier of PARP inhibitor sensitivity. Oncotarget,
Vol.6
(13),
pp. 10746-10758.
show abstract
full text
Based on a series of basic, preclinical and clinical studies, the Poly (ADP-ribose) Polymerase 1 (PARP1) inhibitor, olaparib, has recently been approved for use in ovarian cancer patients with BRCA1 or BRCA2 mutations. By identifying novel predictive biomarkers of tumour cell sensitivity to olaparib, it is possible that the utility of PARP inhibitors could be extended beyond this patient subgroup. Many of the known genetic determinants of PARP inhibitor response have key roles in DNA damage response (DDR) pathways. Although protein ubiquitylation is known to play an important role in regulating the DDR, the exact mechanisms by which this occurs are not fully understood. Using two parallel RNA interference-based screening approaches, we identified the E3 ubiquitin ligase, CBLC, as a candidate biomarker of response to olaparib. We validated this observation by demonstrating that silencing of CBLC causes increased sensitivity to olaparib in breast cancer cell line models and that defective homologous recombination (HR) DNA repair is the likely cause. This data provides an example of how defects in the ubiquitin machinery have the potential to influence the response of tumour cells to PARP inhibitors..
Conti, A.
Majorini, M.T.
Elliott, R.
Ashworth, A.
Lord, C.J.
Cancelliere, C.
Bardelli, A.
Seneci, P.
Walczak, H.
Delia, D.
Lecis, D.
(2015). Oncogenic KRAS sensitizes premalignant, but not malignant cells, to Noxa-dependent apoptosis through the activation of the MEK/ERK pathway. Oncotarget,
Vol.6
(13),
pp. 10994-11008.
show abstract
full text
KRAS is mutated in about 20-25% of all human cancers and especially in pancreatic, lung and colorectal tumors. Oncogenic KRAS stimulates several pro-survival pathways, but it also triggers the trans-activation of pro-apoptotic genes. In our work, we show that G13D mutations of KRAS activate the MAPK pathway, and ERK2, but not ERK1, up-regulates Noxa basal levels. Accordingly, premalignant epithelial cells are sensitized to various cytotoxic compounds in a Noxa-dependent manner. In contrast to these findings, colorectal cancer cell sensitivity to treatment is independent of KRAS status and Noxa levels are not up-regulated in the presence of mutated KRAS despite the fact that ERK2 still promotes Noxa expression. We therefore speculated that other survival pathways are counteracting the pro-apoptotic effect of mutated KRAS and found that the inhibition of AKT restores sensitivity to treatment, especially in presence of oncogenic KRAS. In conclusion, our work suggests that the pharmacological inhibition of the pathways triggered by mutated KRAS could also switch off its oncogene-activated pro-apoptotic stimulation. On the contrary, the combination of chemotherapy to inhibitors of specific pro-survival pathways, such as the one controlled by AKT, could enhance treatment efficacy by exploiting the pro-death stimulation derived by oncogene activation..
Mateo, J.
Carreira, S.
Sandhu, S.
Miranda, S.
Mossop, H.
Perez-Lopez, R.
Nava Rodrigues, D.
Robinson, D.
Omlin, A.
Tunariu, N.
Boysen, G.
Porta, N.
Flohr, P.
Gillman, A.
Figueiredo, I.
Paulding, C.
Seed, G.
Jain, S.
Ralph, C.
Protheroe, A.
Hussain, S.
Jones, R.
Elliott, T.
McGovern, U.
Bianchini, D.
Goodall, J.
Zafeiriou, Z.
Williamson, C.T.
Ferraldeschi, R.
Riisnaes, R.
Ebbs, B.
Fowler, G.
Roda, D.
Yuan, W.
Wu, Y.-.
Cao, X.
Brough, R.
Pemberton, H.
A'Hern, R.
Swain, A.
Kunju, L.P.
Eeles, R.
Attard, G.
Lord, C.J.
Ashworth, A.
Rubin, M.A.
Knudsen, K.E.
Feng, F.Y.
Chinnaiyan, A.M.
Hall, E.
de Bono, J.S.
(2015). DNA-Repair Defects and Olaparib in Metastatic Prostate Cancer. N engl j med,
Vol.373
(18),
pp. 1697-1708.
show abstract
full text
BACKGROUND: Prostate cancer is a heterogeneous disease, but current treatments are not based on molecular stratification. We hypothesized that metastatic, castration-resistant prostate cancers with DNA-repair defects would respond to poly(adenosine diphosphate [ADP]-ribose) polymerase (PARP) inhibition with olaparib. METHODS: We conducted a phase 2 trial in which patients with metastatic, castration-resistant prostate cancer were treated with olaparib tablets at a dose of 400 mg twice a day. The primary end point was the response rate, defined either as an objective response according to Response Evaluation Criteria in Solid Tumors, version 1.1, or as a reduction of at least 50% in the prostate-specific antigen level or a confirmed reduction in the circulating tumor-cell count from 5 or more cells per 7.5 ml of blood to less than 5 cells per 7.5 ml. Targeted next-generation sequencing, exome and transcriptome analysis, and digital polymerase-chain-reaction testing were performed on samples from mandated tumor biopsies. RESULTS: Overall, 50 patients were enrolled; all had received prior treatment with docetaxel, 49 (98%) had received abiraterone or enzalutamide, and 29 (58%) had received cabazitaxel. Sixteen of 49 patients who could be evaluated had a response (33%; 95% confidence interval, 20 to 48), with 12 patients receiving the study treatment for more than 6 months. Next-generation sequencing identified homozygous deletions, deleterious mutations, or both in DNA-repair genes--including BRCA1/2, ATM, Fanconi's anemia genes, and CHEK2--in 16 of 49 patients who could be evaluated (33%). Of these 16 patients, 14 (88%) had a response to olaparib, including all 7 patients with BRCA2 loss (4 with biallelic somatic loss, and 3 with germline mutations) and 4 of 5 with ATM aberrations. The specificity of the biomarker suite was 94%. Anemia (in 10 of the 50 patients [20%]) and fatigue (in 6 [12%]) were the most common grade 3 or 4 adverse events, findings that are consistent with previous studies of olaparib. CONCLUSIONS: Treatment with the PARP inhibitor olaparib in patients whose prostate cancers were no longer responding to standard treatments and who had defects in DNA-repair genes led to a high response rate. (Funded by Cancer Research UK and others; ClinicalTrials.gov number, NCT01682772; Cancer Research UK number, CRUK/11/029.)..
Lord, C.J.
Tutt, A.N.
Ashworth, A.
(2015). Synthetic lethality and cancer therapy: lessons learned from the development of PARP inhibitors. Annu rev med,
Vol.66,
pp. 455-470.
show abstract
The genetic concept of synthetic lethality, in which the combination or synthesis of mutations in multiple genes results in cell death, provides a framework to design novel therapeutic approaches to cancer. Already there are promising indications, from clinical trials exploiting this concept by using poly(ADP-ribose) polymerase (PARP) inhibitors in patients with germline BRCA1 or BRCA2 gene mutations, that this approach could be beneficial. We discuss the biological rationale for BRCA-PARP synthetic lethality, how the synthetic lethal approach is being assessed in the clinic, and how mechanisms of resistance are starting to be dissected. Applying the synthetic lethal concept to target non-BRCA-mutant cancers also has clear potential, and we discuss how some of the principles learned in developing PARP inhibitors might also drive the development of additional genetic approaches..
Elliott, R.J.
Jarvis, A.
Rajasekaran, M.B.
Menon, M.
Bowers, L.
Boffey, R.
Bayford, M.
Firth-Clark, S.
Key, R.
Aqil, R.
Kirton, S.B.
Niculescu-Duvaz, D.
Fish, L.
Lopes, F.
McLeary, R.
Trindade, I.
Vendrell, E.
Munkonge, F.
Porter, R.
Perrior, T.
Springer, C.
Oliver, A.W.
Pearl, L.H.
Ashworth, A.
Lord, C.J.
(2015). Design and discovery of 3-aryl-5-substituted-isoquinolin-1-ones as potent tankyrase inhibitors. Medchemcomm,
Vol.6
(9),
pp. 1687-1692.
Bajrami, I.
Frankum, J.R.
Konde, A.
Miller, R.E.
Rehman, F.L.
Brough, R.
Campbell, J.
Sims, D.
Rafiq, R.
Hooper, S.
Chen, L.
Kozarewa, I.
Assiotis, I.
Fenwick, K.
Natrajan, R.
Lord, C.J.
Ashworth, A.
(2014). Genome-wide profiling of genetic synthetic lethality identifies CDK12 as a novel determinant of PARP1/2 inhibitor sensitivity. Cancer res,
Vol.74
(1),
pp. 287-297.
show abstract
Small-molecule inhibitors of PARP1/2, such as olaparib, have been proposed to serve as a synthetic lethal therapy for cancers that harbor BRCA1 or BRCA2 mutations. Indeed, in clinical trials, PARP1/2 inhibitors elicit sustained antitumor responses in patients with germline BRCA gene mutations. In hypothesizing that additional genetic determinants might direct use of these drugs, we conducted a genome-wide synthetic lethal screen for candidate olaparib sensitivity genes. In support of this hypothesis, the set of identified genes included known determinants of olaparib sensitivity, such as BRCA1, RAD51, and Fanconi's anemia susceptibility genes. In addition, the set included genes implicated in established networks of DNA repair, DNA cohesion, and chromatin remodeling, none of which were known previously to confer sensitivity to PARP1/2 inhibition. Notably, integration of the list of candidate sensitivity genes with data from tumor DNA sequencing studies identified CDK12 deficiency as a clinically relevant biomarker of PARP1/2 inhibitor sensitivity. In models of high-grade serous ovarian cancer (HGS-OVCa), CDK12 attenuation was sufficient to confer sensitivity to PARP1/2 inhibition, suppression of DNA repair via homologous recombination, and reduced expression of BRCA1. As one of only nine genes known to be significantly mutated in HGS-OVCa, CDK12 has properties that should confirm interest in its use as a biomarker, particularly in ongoing clinical trials of PARP1/2 inhibitors and other agents that trigger replication fork arrest..
Murugaesu, N.
Iravani, M.
van Weverwijk, A.
Ivetic, A.
Johnson, D.A.
Antonopoulos, A.
Fearns, A.
Jamal-Hanjani, M.
Sims, D.
Fenwick, K.
Mitsopoulos, C.
Gao, Q.
Orr, N.
Zvelebil, M.
Haslam, S.M.
Dell, A.
Yarwood, H.
Lord, C.J.
Ashworth, A.
Isacke, C.M.
(2014). An in vivo functional screen identifies ST6GalNAc2 sialyltransferase as a breast cancer metastasis suppressor. Cancer discov,
Vol.4
(3),
pp. 304-317.
show abstract
To interrogate the complex mechanisms involved in the later stages of cancer metastasis, we designed a functional in vivo RNA interference (RNAi) screen combined with next-generation sequencing. Using this approach, we identified the sialyltransferase ST6GalNAc2 as a novel breast cancer metastasis suppressor. Mechanistically, ST6GalNAc2 silencing alters the profile of O-glycans on the tumor cell surface, facilitating binding of the soluble lectin galectin-3. This then enhances tumor cell retention and emboli formation at metastatic sites leading to increased metastatic burden, events that can be completely blocked by galectin-3 inhibition. Critically, elevated ST6GALNAC2, but not galectin-3, expression in estrogen receptor-negative breast cancers significantly correlates with reduced frequency of metastatic events and improved survival. These data demonstrate that the prometastatic role of galectin-3 is regulated by its ability to bind to the tumor cell surface and highlight the potential of monitoring ST6GalNAc2 expression to stratify patients with breast cancer for treatment with galectin-3 inhibitors..
Natrajan, R.
Wilkerson, P.M.
Marchiò, C.
Piscuoglio, S.
Ng, C.K.
Wai, P.
Lambros, M.B.
Samartzis, E.P.
Dedes, K.J.
Frankum, J.
Bajrami, I.
Kopec, A.
Mackay, A.
A'hern, R.
Fenwick, K.
Kozarewa, I.
Hakas, J.
Mitsopoulos, C.
Hardisson, D.
Lord, C.J.
Kumar-Sinha, C.
Ashworth, A.
Weigelt, B.
Sapino, A.
Chinnaiyan, A.M.
Maher, C.A.
Reis-Filho, J.S.
(2014). Characterization of the genomic features and expressed fusion genes in micropapillary carcinomas of the breast. J pathol,
Vol.232
(5),
pp. 553-565.
show abstract
full text
Micropapillary carcinoma (MPC) is a rare histological special type of breast cancer, characterized by an aggressive clinical behaviour and a pattern of copy number aberrations (CNAs) distinct from that of grade- and oestrogen receptor (ER)-matched invasive carcinomas of no special type (IC-NSTs). The aims of this study were to determine whether MPCs are underpinned by a recurrent fusion gene(s) or mutations in 273 genes recurrently mutated in breast cancer. Sixteen MPCs were subjected to microarray-based comparative genomic hybridization (aCGH) analysis and Sequenom OncoCarta mutation analysis. Eight and five MPCs were subjected to targeted capture and RNA sequencing, respectively. aCGH analysis confirmed our previous observations about the repertoire of CNAs of MPCs. Sequencing analysis revealed a spectrum of mutations similar to those of luminal B IC-NSTs, and recurrent mutations affecting mitogen-activated protein kinase family genes and NBPF10. RNA-sequencing analysis identified 17 high-confidence fusion genes, eight of which were validated and two of which were in-frame. No recurrent fusions were identified in an independent series of MPCs and IC-NSTs. Forced expression of in-frame fusion genes (SLC2A1-FAF1 and BCAS4-AURKA) resulted in increased viability of breast cancer cells. In addition, genomic disruption of CDK12 caused by out-of-frame rearrangements was found in one MPC and in 13% of HER2-positive breast cancers, identified through a re-analysis of publicly available massively parallel sequencing data. In vitro analyses revealed that CDK12 gene disruption results in sensitivity to PARP inhibition, and forced expression of wild-type CDK12 in a CDK12-null cell line model resulted in relative resistance to PARP inhibition. Our findings demonstrate that MPCs are neither defined by highly recurrent mutations in the 273 genes tested, nor underpinned by a recurrent fusion gene. Although seemingly private genetic events, some of the fusion transcripts found in MPCs may play a role in maintenance of a malignant phenotype and potentially offer therapeutic opportunities..
McDade, S.S.
Patel, D.
Moran, M.
Campbell, J.
Fenwick, K.
Kozarewa, I.
Orr, N.J.
Lord, C.J.
Ashworth, A.A.
McCance, D.J.
(2014). Genome-wide characterization reveals complex interplay between TP53 and TP63 in response to genotoxic stress. Nucleic acids res,
Vol.42
(10),
pp. 6270-6285.
show abstract
full text
In response to genotoxic stress the TP53 tumour suppressor activates target gene expression to induce cell cycle arrest or apoptosis depending on the extent of DNA damage. These canonical activities can be repressed by TP63 in normal stratifying epithelia to maintain proliferative capacity or drive proliferation of squamous cell carcinomas, where TP63 is frequently overexpressed/amplified. Here we use ChIP-sequencing, integrated with microarray analysis, to define the genome-wide interplay between TP53 and TP63 in response to genotoxic stress in normal cells. We reveal that TP53 and TP63 bind to overlapping, but distinct cistromes of sites through utilization of distinctive consensus motifs and that TP53 is constitutively bound to a number of sites. We demonstrate that cisplatin and adriamycin elicit distinct effects on TP53 and TP63 binding events, through which TP53 can induce or repress transcription of an extensive network of genes by direct binding and/or modulation of TP63 activity. Collectively, this results in a global TP53-dependent repression of cell cycle progression, mitosis and DNA damage repair concomitant with activation of anti-proliferative and pro-apoptotic canonical target genes. Further analyses reveal that in the absence of genotoxic stress TP63 plays an important role in maintaining expression of DNA repair genes, loss of which results in defective repair..
Ryan, C.J.
Lord, C.J.
Ashworth, A.
(2014). DAISY: picking synthetic lethals from cancer genomes. Cancer cell,
Vol.26
(3),
pp. 306-308.
show abstract
full text
A better understanding of genetic interactions in cancer might help identify new therapeutic approaches that exploit the concept of synthetic lethality. Ruppin and colleagues have developed a new computational method, DAISY, that predicts such interactions and potentially facilitates the delineation and validation of comprehensive genetic interaction networks..
Rajan, N.
Elliott, R.J.
Smith, A.
Sinclair, N.
Swift, S.
Lord, C.J.
Ashworth, A.
(2014). The cylindromatosis gene product, CYLD, interacts with MIB2 to regulate Notch signalling. Oncotarget,
Vol.5
(23),
pp. 12126-12140.
full text
Gao, S.
Bajrami, I.
Verrill, C.
Kigozi, A.
Ouaret, D.
Aleksic, T.
Asher, R.
Han, C.
Allen, P.
Bailey, D.
Feller, S.
Kashima, T.
Athanasou, N.
Blay, J.-.
Schmitz, S.
Machiels, J.-.
Upile, N.
Jones, T.M.
Thalmann, G.
Ashraf, S.Q.
Wilding, J.L.
Bodmer, W.F.
Middleton, M.R.
Ashworth, A.
Lord, C.J.
Macaulay, V.M.
(2014). Dsh homolog DVL3 mediates resistance to IGFIR inhibition by regulating IGF-RAS signaling. Cancer res,
Vol.74
(20),
pp. 5866-5877.
show abstract
Drugs that inhibit insulin-like growth factor 1 (IGFI) receptor IGFIR were encouraging in early trials, but predictive biomarkers were lacking and the drugs provided insufficient benefit in unselected patients. In this study, we used genetic screening and downstream validation to identify the WNT pathway element DVL3 as a mediator of resistance to IGFIR inhibition. Sensitivity to IGFIR inhibition was enhanced specifically in vitro and in vivo by genetic or pharmacologic blockade of DVL3. In breast and prostate cancer cells, sensitization tracked with enhanced MEK-ERK activation and relied upon MEK activity and DVL3 expression. Mechanistic investigations showed that DVL3 is present in an adaptor complex that links IGFIR to RAS, which includes Shc, growth factor receptor-bound-2 (Grb2), son-of-sevenless (SOS), and the tumor suppressor DAB2. Dual DVL and DAB2 blockade synergized in activating ERKs and sensitizing cells to IGFIR inhibition, suggesting a nonredundant role for DVL3 in the Shc-Grb2-SOS complex. Clinically, tumors that responded to IGFIR inhibition contained relatively lower levels of DVL3 protein than resistant tumors, and DVL3 levels in tumors correlated inversely with progression-free survival in patients treated with IGFIR antibodies. Because IGFIR does not contain activating mutations analogous to EGFR variants associated with response to EGFR inhibitors, we suggest that IGF signaling achieves an equivalent integration at the postreceptor level through adaptor protein complexes, influencing cellular dependence on the IGF axis and identifying a patient population with potential to benefit from IGFIR inhibition..
Wetterskog, D.
Shiu, K.-.
Chong, I.
Meijer, T.
Mackay, A.
Lambros, M.
Cunningham, D.
Reis-Filho, J.S.
Lord, C.J.
Ashworth, A.
(2014). Identification of novel determinants of resistance to lapatinib in ERBB2-amplified cancers. Oncogene,
Vol.33
(8),
pp. 966-976.
show abstract
The gene encoding the receptor tyrosine kinase ERBB2, also known as HER2, is amplified and/or overexpressed in up to 15% of breast cancers. These tumours are characterised by an aggressive phenotype and poor clinical outcome. Although therapies targeted at ERBB2 have proven effective, many patients fail to respond to treatment or become resistant and the reasons for this are still largely unknown. Using a high-throughput functional screen we assessed whether genes found to be recurrently amplified and overexpressed in ERBB2+ve breast cancers mediate resistance to the ERBB2-targeted agent lapatinib. Lapatinib-resistant ERBB2-amplified breast cancer cell lines were screened, in the presence or absence of lapatinib, with an RNA interference library targeting 369 genes recurrently amplified and overexpressed in both ERBB2-amplified breast cancer tumours and cell lines. Small interfering RNAs targeting a number of genes caused sensitivity to lapatinib in this context. The mechanisms of resistance conferred by the identified genes were further investigated and in the case of NIBP (TRAPPC9), lapatinib resistance was found to be mediated through NF-κB signalling. Our results indicate that specific amplified and/ or overexpressed genes found in ERBB2-amplified breast cancer may mediate response to ERBB2-targeting agents..
Shiu, K.-.
Wetterskog, D.
Mackay, A.
Natrajan, R.
Lambros, M.
Sims, D.
Bajrami, I.
Brough, R.
Frankum, J.
Sharpe, R.
Marchio, C.
Horlings, H.
Reyal, F.
van der Vijver, M.
Turner, N.
Reis-Filho, J.S.
Lord, C.J.
Ashworth, A.
(2014). Integrative molecular and functional profiling of ERBB2-amplified breast cancers identifies new genetic dependencies. Oncogene,
Vol.33
(5),
pp. 619-631.
show abstract
Overexpression of the receptor tyrosine kinase ERBB2 (also known as HER2) occurs in around 15% of breast cancers and is driven by amplification of the ERBB2 gene. ERBB2 amplification is a marker of poor prognosis, and although anti-ERBB2-targeted therapies have shown significant clinical benefit, de novo and acquired resistance remains an important problem. Genomic profiling has demonstrated that ERBB2+ve breast cancers are distinguished from ER+ve and 'triple-negative' breast cancers by harbouring not only the ERBB2 amplification on 17q12, but also a number of co-amplified genes on 17q12 and amplification events on other chromosomes. Some of these genes may have important roles in influencing clinical outcome, and could represent genetic dependencies in ERBB2+ve cancers and therefore potential therapeutic targets. Here, we describe an integrated genomic, gene expression and functional analysis to determine whether the genes present within amplicons are critical for the survival of ERBB2+ve breast tumour cells. We show that only a fraction of the ERBB2-amplified breast tumour lines are truly addicted to the ERBB2 oncogene at the mRNA level and display a heterogeneous set of additional genetic dependencies. These include an addiction to the transcription factor gene TFAP2C when it is amplified and overexpressed, suggesting that TFAP2C represents a genetic dependency in some ERBB2+ve breast cancer cells..
Ang, J.E.
Gourley, C.
Powell, C.B.
High, H.
Shapira-Frommer, R.
Castonguay, V.
De Greve, J.
Atkinson, T.
Yap, T.A.
Sandhu, S.
Banerjee, S.
Chen, L.-.
Friedlander, M.L.
Kaufman, B.
Oza, A.M.
Matulonis, U.
Barber, L.J.
Kozarewa, I.
Fenwick, K.
Assiotis, I.
Campbell, J.
Chen, L.
de Bono, J.S.
Gore, M.E.
Lord, C.J.
Ashworth, A.
Kaye, S.B.
(2013). Efficacy of chemotherapy in BRCA1/2 mutation carrier ovarian cancer in the setting of PARP inhibitor resistance: a multi-institutional study. Clin cancer res,
Vol.19
(19),
pp. 5485-5493.
show abstract
PURPOSE: Preclinical data suggest that exposure to PARP inhibitors (PARPi) may compromise benefit to subsequent chemotherapy, particularly platinum-based regimens, in patients with BRCA1/2 mutation carrier ovarian cancer (PBMCOC), possibly through the acquisition of secondary BRCA1/2 mutations. The efficacy of chemotherapy in the PARPi-resistant setting was therefore investigated. EXPERIMENTAL DESIGN: We conducted a retrospective review of PBMCOC who received chemotherapy following disease progression on olaparib, administered at ≥200 mg twice daily for one month or more. Tumor samples were obtained in the post-olaparib setting where feasible and analyzed by massively parallel sequencing. RESULTS: Data were collected from 89 patients who received a median of 3 (range 1-11) lines of pre-olaparib chemotherapy. The overall objective response rate (ORR) to post-olaparib chemotherapy was 36% (24 of 67 patients) by Response Evaluation Criteria in Solid Tumors (RECIST) and 45% (35 of 78) by RECIST and/or Gynecologic Cancer InterGroup (GCIG) CA125 criteria with median progression-free survival (PFS) and overall survival (OS) of 17 weeks [95% confidence interval (CI), 13-21] and 34 weeks (95% CI, 26-42), respectively. For patients receiving platinum-based chemotherapy, ORRs were 40% (19 of 48) and 49% (26/53), respectively, with a median PFS of 22 weeks (95% CI, 15-29) and OS of 45 weeks (95% CI, 15-75). An increased platinum-to-platinum interval was associated with an increased OS and likelihood of response following post-olaparib platinum. No evidence of secondary BRCA1/2 mutation was detected in tumor samples of six PARPi-resistant patients [estimated frequency of such mutations adjusted for sample size: 0.125 (95%-CI: 0-0.375)]. CONCLUSIONS: Heavily pretreated PBMCOC who are PARPi-resistant retain the potential to respond to subsequent chemotherapy, including platinum-based agents. These data support the further development of PARPi in PBMCOC..
Neijenhuis, S.
Bajrami, I.
Miller, R.
Lord, C.J.
Ashworth, A.
(2013). Identification of miRNA modulators to PARP inhibitor response. Dna repair (amst),
Vol.12
(6),
pp. 394-402.
show abstract
Based on the principle of synthetic lethality, PARP inhibitors have been shown to be very effective in killing cells deficient in homologous recombination (HR), such as those bearing mutations in BRCA1/2. However, questions regarding their wider use persist and other determinants of responsiveness to PARP inhibitor remain to be fully explored. MicroRNAs (miRNAs) are small non-coding RNAs, which serve as post-transcriptional regulators of gene expression and are involved in a wide variety of cellular processes, including the DNA damage response (DDR). However, little is known about whether miRNAs might influence sensitivity to PARP inhibitors. To investigate this, we performed a high throughput miRNA mimetic screen, which identified several miRNAs whose over-expression results in sensitization to the clinical PARP inhibitor olaparib. In particular, our findings indicate that hsa-miR-107 and hsa-miR-222 regulate the DDR and sensitise tumour cells to olaparib by repressing expression of RAD51, thus impairing DSB repair by HR. Moreover, elevated expression of hsa-miR-107 has been observed in a subset of ovarian clear cell carcinomas, which correlates with PARP inhibitor sensitivity and reduced RAD51 expression. Taken together, these observations raise the possibility that these miRNAs could be used as biomarkers to identify patients that may benefit from treatment with PARP inhibitors..
Barber, L.J.
Sandhu, S.
Chen, L.
Campbell, J.
Kozarewa, I.
Fenwick, K.
Assiotis, I.
Rodrigues, D.N.
Reis Filho, J.S.
Moreno, V.
Mateo, J.
Molife, L.R.
De Bono, J.
Kaye, S.
Lord, C.J.
Ashworth, A.
(2013). Secondary mutations in BRCA2 associated with clinical resistance to a PARP inhibitor. J pathol,
Vol.229
(3),
pp. 422-429.
show abstract
PARP inhibitors (PARPi) for the treatment of BRCA1 or BRCA2 deficient tumours are currently the focus of seminal clinical trials exploiting the concept of synthetic lethality. Although clinical resistance to PARPi has been described, the mechanism underlying this has not been elucidated. Here, we investigate tumour material from patients who had developed resistance to the PARPi olaparib, subsequent to showing an initial clinical response. Massively parallel DNA sequencing of treatment-naive and post-olaparib treatment biopsies identified tumour-specific BRCA2 secondary mutations in olaparib-resistant metastases. These secondary mutations restored full-length BRCA2 protein, and most likely cause olaparib resistance by re-establishing BRCA2 function in the tumour cells..
Sandhu, S.K.
Omlin, A.
Hylands, L.
Miranda, S.
Barber, L.J.
Riisnaes, R.
Reid, A.H.
Attard, G.
Chen, L.
Kozarewa, I.
Gevensleben, H.
Campbell, J.
Fenwick, K.
Assiotis, I.
Olmos, D.
Yap, T.A.
Fong, P.
Tunariu, N.
Koh, D.
Molife, L.R.
Kaye, S.
Lord, C.J.
Ashworth, A.
de Bono, J.
(2013). Poly (ADP-ribose) polymerase (PARP) inhibitors for the treatment of advanced germline BRCA2 mutant prostate cancer. Ann oncol,
Vol.24
(5),
pp. 1416-1418.
Bjerke, L.
Mackay, A.
Nandhabalan, M.
Burford, A.
Jury, A.
Popov, S.
Bax, D.A.
Carvalho, D.
Taylor, K.R.
Vinci, M.
Bajrami, I.
McGonnell, I.M.
Lord, C.J.
Reis, R.M.
Hargrave, D.
Ashworth, A.
Workman, P.
Jones, C.
(2013). Histone H3 3 Mutations Drive Pediatric Glioblastoma through Upregulation of MYCN. Cancer discovery,
Vol.3
(5),
pp. 512-519.
Pettitt, S.J.
Rehman, F.L.
Bajrami, I.
Pemberton, H.
Brough, R.
Kozarewa, I.
Lord, C.J.
Ashworth, A.
(2013). A transposon-based genetic screen in haploid mouse embryonic stem cells identifies Parp1 as a major mediator of olaparib toxicity. Molecular cancer therapeutics,
Vol.12
(5).
Fontebasso, Y.
Lord, C.J.
Ashworth, A.
(2013). Induced aggravation of oxidative DNA damage as a targeted synthetic lethal approach in mismatch repair-deficient tumors. Molecular cancer therapeutics,
Vol.12
(5).
Bjerke, L.
Mackay, A.
Nandhabalan, M.
Burford, A.
Jury, A.
Popov, S.
Bax, D.A.
Carvalho, D.
Taylor, K.R.
Vinci, M.
Bajrami, I.
McGonnell, I.M.
Lord, C.J.
Reis, R.M.
Hargrave, D.
Ashworth, A.
Workman, P.
Jones, C.
(2013). Histone H3 3 mutations drive pediatric glioblastoma through upregulation of MYCN. Cancer discov,
Vol.3
(5),
pp. 512-519.
show abstract
UNLABELLED: Children and young adults with glioblastoma (GBM) have a median survival rate of only 12 to 15 months, and these GBMs are clinically and biologically distinct from histologically similar cancers in older adults. They are defined by highly specific mutations in the gene encoding the histone H3.3 variant H3F3A , occurring either at or close to key residues marked by methylation for regulation of transcription—K27 and G34. Here, we show that the cerebral hemisphere-specific G34 mutation drives a distinct expression signature through differential genomic binding of the K36 trimethylation mark (H3K36me3). The transcriptional program induced recapitulates that of the developing forebrain, and involves numerous markers of stem-cell maintenance, cell-fate decisions, and self-renewal.Critically, H3F3A G34 mutations cause profound upregulation of MYCN , a potent oncogene that is causative of GBMs when expressed in the correct developmental context. This driving aberration is selectively targetable in this patient population through inhibiting kinases responsible for stabilization of the protein. SIGNIFICANCE: We provide the mechanistic explanation for how the fi rst histone gene mutation inhuman disease biology acts to deliver MYCN, a potent tumorigenic initiator, into a stem-cell compartment of the developing forebrain, selectively giving rise to incurable cerebral hemispheric GBM. Using synthetic lethal approaches to these mutant tumor cells provides a rational way to develop novel and highly selective treatment strategies.
Hewish, M.
Martin, S.A.
Elliott, R.
Cunningham, D.
Lord, C.J.
Ashworth, A.
(2013). Cytosine-based nucleoside analogs are selectively lethal to DNA mismatch repair-deficient tumour cells by enhancing levels of intracellular oxidative stress. Br j cancer,
Vol.108
(4),
pp. 983-992.
show abstract
BACKGROUND: DNA mismatch repair deficiency is present in a significant proportion of a number of solid tumours and is associated with distinct clinical behaviour. METHODS: To identify the therapeutic agents that might show selectivity for mismatch repair-deficient tumour cells, we screened a pair of isogenic MLH1-deficient and MLH1-proficient tumour cell lines with a library of clinically used drugs. To test the generality of hits in the screen, selective agents were retested in cells deficient in the MSH2 mismatch repair gene. RESULTS: We identified cytarabine and other related cytosine-based nucleoside analogues as being selectively toxic to MLH1 and MSH2-deficient tumour cells. The selective cytotoxicity we observed was likely caused by increased levels of cellular oxidative stress, as it could be abrogated by antioxidants. CONCLUSION: We propose that cytarabine-based chemotherapy regimens may represent a tumour-selective treatment strategy for mismatch repair-deficient cancers..
Lord, C.
Pollak, J.
(2013). The Pitfalls of Representation as Claims-Making in the European Union1. Journal of european integration,
Vol.35
(5),
pp. 517-530.
Mereniuk, T.R.
El Gendy, M.A.
Mendes-Pereira, A.M.
Lord, C.J.
Ghosh, S.
Foley, E.
Ashworth, A.
Weinfeld, M.
(2013). Synthetic lethal targeting of PTEN-deficient cancer cells using selective disruption of polynucleotide kinase/phosphatase. Mol cancer ther,
Vol.12
(10),
pp. 2135-2144.
show abstract
A recent screen of 6,961 siRNAs to discover possible synthetic lethal partners of the DNA repair protein polynucleotide kinase/phosphatase (PNKP) led to the identification of the potent tumor suppressor phosphatase and tensin homolog deleted on chromosome 10 (PTEN). Here, we have confirmed the PNKP/PTEN synthetic lethal partnership in a variety of different cell lines including the PC3 prostate cancer cell line, which is naturally deficient in PTEN. We provide evidence that codepletion of PTEN and PNKP induces apoptosis. In HCT116 colon cancer cells, the loss of PTEN is accompanied by an increased background level of DNA double-strand breaks, which accumulate in the presence of an inhibitor of PNKP DNA 3'-phosphatase activity. Complementation of PC3 cells with several well-characterized mutated PTEN cDNAs indicated that the critical function of PTEN required to prevent toxicity induced by an inhibitor of PNKP is most likely associated with its cytoplasmic lipid phosphatase activity. Finally, we show that modest inhibition of PNKP in a PTEN knockout background enhances cellular radiosensitivity, suggesting that such a "synthetic sickness" approach involving the combination of PNKP inhibition with radiotherapy may be applicable to PTEN-deficient tumors..
Chong, I.Y.
Cunningham, D.
Barber, L.J.
Campbell, J.
Chen, L.
Kozarewa, I.
Fenwick, K.
Assiotis, I.
Guettler, S.
Garcia-Murillas, I.
Awan, S.
Lambros, M.
Starling, N.
Wotherspoon, A.
Stamp, G.
Gonzalez-de-Castro, D.
Benson, M.
Chau, I.
Hulkki, S.
Nohadani, M.
Eltahir, Z.
Lemnrau, A.
Orr, N.
Rao, S.
Lord, C.J.
Ashworth, A.
(2013). The genomic landscape of oesophagogastric junctional adenocarcinoma. Journal of pathology,
Vol.231
(3),
pp. 301-310.
Lord, C.J.
Ashworth, A.
(2013). Mechanisms of resistance to therapies targeting BRCA-mutant cancers. Nat med,
Vol.19
(11),
pp. 1381-1388.
show abstract
Synthetic lethality provides a potential mechanistic framework for the therapeutic targeting of genetic and functional deficiencies in cancers and is now being explored widely. The first clinical exemplification of synthetic lethality in cancer has been the exploitation of inhibitors of poly-(ADP-ribose) polymerase (PARP) for the treatment of cancers with defects in the BRCA1 or BRCA2 tumor suppressor proteins, which are involved in the repair of DNA damage. Although this approach has shown promise, multiple potential resistance mechanisms have been identified. In this Perspective, we discuss these mechanisms and their relevance to the development of selective therapies for BRCA-deficient cancers..
Natrajan, R.C.
Leonidou, A.
Brough, R.
Frankum, J.
Wai, P.T.
Ng, C.K.
Reis-Filho, J.S.
Lord, C.J.
Ashworth, A.
(2013). Integrated genomic analyses of members of protein kinase C family identifies subtype specific alterations as novel therapeutic targets. Cancer research,
Vol.73.
Shen, Y.
Rehman, F.L.
Feng, Y.
Boshuizen, J.
Bajrami, I.
Elliott, R.
Wang, B.
Lord, C.J.
Post, L.E.
Ashworth, A.
(2013). BMN 673, a novel and highly potent PARP1/2 inhibitor for the treatment of human cancers with DNA repair deficiency. Clin cancer res,
Vol.19
(18),
pp. 5003-5015.
show abstract
PURPOSE: PARP1/2 inhibitors are a class of anticancer agents that target tumor-specific defects in DNA repair. Here, we describe BMN 673, a novel, highly potent PARP1/2 inhibitor with favorable metabolic stability, oral bioavailability, and pharmacokinetic properties. EXPERIMENTAL DESIGN: Potency and selectivity of BMN 673 was determined by biochemical assays. Anticancer activity either as a single-agent or in combination with other antitumor agents was evaluated both in vitro and in xenograft cancer models. RESULTS: BMN 673 is a potent PARP1/2 inhibitor (PARP1 IC50 = 0.57 nmol/L), but it does not inhibit other enzymes that we have tested. BMN 673 exhibits selective antitumor cytotoxicity and elicits DNA repair biomarkers at much lower concentrations than earlier generation PARP1/2 inhibitors (such as olaparib, rucaparib, and veliparib). In vitro, BMN 673 selectively targeted tumor cells with BRCA1, BRCA2, or PTEN gene defects with 20- to more than 200-fold greater potency than existing PARP1/2 inhibitors. BMN 673 is readily orally bioavailable, with more than 40% absolute oral bioavailability in rats when dosed in carboxylmethyl cellulose. Oral administration of BMN 673 elicited remarkable antitumor activity in vivo; xenografted tumors that carry defects in DNA repair due to BRCA mutations or PTEN deficiency were profoundly sensitive to oral BMN 673 treatment at well-tolerated doses in mice. Synergistic or additive antitumor effects were also found when BMN 673 was combined with temozolomide, SN38, or platinum drugs. CONCLUSION: BMN 673 is currently in early-phase clinical development and represents a promising PARP1/2 inhibitor with potentially advantageous features in its drug class..
Ruark, E.
Snape, K.
Humburg, P.
Loveday, C.
Bajrami, I.
Brough, R.
Rodrigues, D.N.
Renwick, A.
Seal, S.
Ramsay, E.
Duarte, S.D.
Rivas, M.A.
Warren-Perry, M.
Zachariou, A.
Campion-Flora, A.
Hanks, S.
Murray, A.
Ansari Pour, N.
Douglas, J.
Gregory, L.
Rimmer, A.
Walker, N.M.
Yang, T.-.
Adlard, J.W.
Barwell, J.
Berg, J.
Brady, A.F.
Brewer, C.
Brice, G.
Chapman, C.
Cook, J.
Davidson, R.
Donaldson, A.
Douglas, F.
Eccles, D.
Evans, D.G.
Greenhalgh, L.
Henderson, A.
Izatt, L.
Kumar, A.
Lalloo, F.
Miedzybrodzka, Z.
Morrison, P.J.
Paterson, J.
Porteous, M.
Rogers, M.T.
Shanley, S.
Walker, L.
Gore, M.
Houlston, R.
Brown, M.A.
Caufield, M.J.
Deloukas, P.
McCarthy, M.I.
Todd, J.A.
Breast and Ovarian Cancer Susceptibility Collaboration,
Wellcome Trust Case Control Consortium,
Turnbull, C.
Reis-Filho, J.S.
Ashworth, A.
Antoniou, A.C.
Lord, C.J.
Donnelly, P.
Rahman, N.
(2013). Mosaic PPM1D mutations are associated with predisposition to breast and ovarian cancer. Nature,
Vol.493
(7432),
pp. 406-410.
show abstract
Improved sequencing technologies offer unprecedented opportunities for investigating the role of rare genetic variation in common disease. However, there are considerable challenges with respect to study design, data analysis and replication. Using pooled next-generation sequencing of 507 genes implicated in the repair of DNA in 1,150 samples, an analytical strategy focused on protein-truncating variants (PTVs) and a large-scale sequencing case-control replication experiment in 13,642 individuals, here we show that rare PTVs in the p53-inducible protein phosphatase PPM1D are associated with predisposition to breast cancer and ovarian cancer. PPM1D PTV mutations were present in 25 out of 7,781 cases versus 1 out of 5,861 controls (P = 1.12 × 10(-5)), including 18 mutations in 6,912 individuals with breast cancer (P = 2.42 × 10(-4)) and 12 mutations in 1,121 individuals with ovarian cancer (P = 3.10 × 10(-9)). Notably, all of the identified PPM1D PTVs were mosaic in lymphocyte DNA and clustered within a 370-base-pair region in the final exon of the gene, carboxy-terminal to the phosphatase catalytic domain. Functional studies demonstrate that the mutations result in enhanced suppression of p53 in response to ionizing radiation exposure, suggesting that the mutant alleles encode hyperactive PPM1D isoforms. Thus, although the mutations cause premature protein truncation, they do not result in the simple loss-of-function effect typically associated with this class of variant, but instead probably have a gain-of-function effect. Our results have implications for the detection and management of breast and ovarian cancer risk. More generally, these data provide new insights into the role of rare and of mosaic genetic variants in common conditions, and the use of sequencing in their identification..
Postel-Vinay, S.
Bajrami, I.
Friboulet, L.
Elliott, R.
Fontebasso, Y.
Dorvault, N.
Olaussen, K.A.
André, F.
Soria, J.-.
Lord, C.J.
Ashworth, A.
(2013). A high-throughput screen identifies PARP1/2 inhibitors as a potential therapy for ERCC1-deficient non-small cell lung cancer. Oncogene,
Vol.32
(47),
pp. 5377-5387.
show abstract
Excision repair cross-complementation group 1 (ERCC1) is a DNA repair enzyme that is frequently defective in non-small cell lung cancer (NSCLC). Although low ERCC1 expression correlates with platinum sensitivity, the clinical effectiveness of platinum therapy is limited, highlighting the need for alternative treatment strategies. To discover new mechanism-based therapeutic strategies for ERCC1-defective tumours, we performed high-throughput drug screens in an isogenic NSCLC model of ERCC1 deficiency and dissected the mechanism underlying ERCC1-selective effects by studying molecular biomarkers of tumour cell response. The high-throughput screens identified multiple clinical poly (ADP-ribose) polymerase 1 and 2 (PARP1/2) inhibitors, such as olaparib (AZD-2281), niraparib (MK-4827) and BMN 673, as being selective for ERCC1 deficiency. We observed that ERCC1-deficient cells displayed a significant delay in double-strand break repair associated with a profound and prolonged G₂/M arrest following PARP1/2 inhibitor treatment. Importantly, we found that ERCC1 isoform 202, which has recently been shown to mediate platinum sensitivity, also modulated PARP1/2 sensitivity. A PARP1/2 inhibitor-synthetic lethal siRNA screen revealed that ERCC1 deficiency was epistatic with homologous recombination deficiency. However, ERCC1-deficient cells did not display a defect in RAD51 foci formation, suggesting that ERCC1 might be required to process PARP1/2 inhibitor-induced DNA lesions before DNA strand invasion. PARP1 silencing restored PARP1/2 inhibitor resistance in ERCC1-deficient cells but had no effect in ERCC1-proficient cells, supporting the hypothesis that PARP1 might be required for the ERCC1 selectivity of PARP1/2 inhibitors. This study suggests that PARP1/2 inhibitors as a monotherapy could represent a novel therapeutic strategy for NSCLC patients with ERCC1-deficient tumours..
Pettitt, S.J.
Rehman, F.L.
Bajrami, I.
Brough, R.
Wallberg, F.
Kozarewa, I.
Fenwick, K.
Assiotis, I.
Chen, L.
Campbell, J.
Lord, C.J.
Ashworth, A.
(2013). A genetic screen using the PiggyBac transposon in haploid cells identifies Parp1 as a mediator of olaparib toxicity. Plos one,
Vol.8
(4),
p. e61520.
show abstract
Genetic perturbation screens have the potential to dissect a wide range of cellular phenotypes. Such screens have historically been difficult in diploid mammalian cells. The recent derivation of haploid embryonic stem cells provides an opportunity to cause loss of function mutants with a random mutagen in a mammalian cell with a normal genetic background. We describe an approach to genetic screens that exploits the highly active piggyBac transposon in haploid mammalian cells. As an example of haploid transposon (HTP) screening, we apply this approach to identifying determinants of cancer drug toxicity and resistance. In a screen for 6-thioguanine resistance we recovered components of the DNA mismatch repair pathway, a known requirement for toxicity. In a further screen for resistance to the clinical poly(ADP-ribose) polymerase (PARP) inhibitor olaparib we recovered multiple Parp1 mutants. Our results show that olaparib toxicity to normal cells is mediated predominantly via Parp1, and suggest that the clinical side effects of olaparib may be on target. The transposon mutant libraries are stable and can be readily reused to screen other drugs. The screening protocol described has several advantages over other methods such as RNA interference: it is rapid and low cost, and mutations can be easily reverted to establish causality..
Turajlic, S.
Furney, S.J.
Lambros, M.B.
Mitsopoulos, C.
Kozarewa, I.
Geyer, F.C.
Mackay, A.
Hakas, J.
Zvelebil, M.
Lord, C.J.
Ashworth, A.
Thomas, M.
Stamp, G.
Larkin, J.
Reis-Filho, J.S.
Marais, R.
(2012). Whole genome sequencing of matched primary and metastatic acral melanomas. Genome res,
Vol.22
(2),
pp. 196-207.
show abstract
Next generation sequencing has enabled systematic discovery of mutational spectra in cancer samples. Here, we used whole genome sequencing to characterize somatic mutations and structural variation in a primary acral melanoma and its lymph node metastasis. Our data show that the somatic mutational rates in this acral melanoma sample pair were more comparable to the rates reported in cancer genomes not associated with mutagenic exposure than in the genome of a melanoma cell line or the transcriptome of melanoma short-term cultures. Despite the perception that acral skin is sun-protected, the dominant mutational signature in these samples is compatible with damage due to ultraviolet light exposure. A nonsense mutation in ERCC5 discovered in both the primary and metastatic tumors could also have contributed to the mutational signature through accumulation of unrepaired dipyrimidine lesions. However, evidence of transcription-coupled repair was suggested by the lower mutational rate in the transcribed regions and expressed genes. The primary and the metastasis are highly similar at the level of global gene copy number alterations, loss of heterozygosity and single nucleotide variation (SNV). Furthermore, the majority of the SNVs in the primary tumor were propagated in the metastasis and one nonsynonymous coding SNV and one splice site mutation appeared to arise de novo in the metastatic lesion..
Walker, B.A.
Wardell, C.P.
Melchor, L.
Hulkki, S.
Potter, N.E.
Johnson, D.C.
Fenwick, K.
Kozarewa, I.
Gonzalez, D.
Lord, C.J.
Ashworth, A.
Davies, F.E.
Morgan, G.J.
(2012). Intraclonal heterogeneity and distinct molecular mechanisms characterize the development of t(4;14) and t(11;14) myeloma. Blood,
Vol.120
(5),
pp. 1077-1086.
show abstract
We have used whole exome sequencing to compare a group of presentation t(4;14) with t(11;14) cases of myeloma to define the mutational landscape. Each case was characterized by a median of 24.5 exonic nonsynonymous single-nucleotide variations, and there was a consistently higher number of mutations in the t(4;14) group, but this number did not reach statistical significance. We show that the transition and transversion rates in the 2 subgroups are similar, suggesting that there was no specific mechanism leading to mutation differentiating the 2 groups. Only 3% of mutations were seen in both groups, and recurrently mutated genes include NRAS, KRAS, BRAF, and DIS3 as well as DNAH5, a member of the axonemal dynein family. The pattern of mutation in each group was distinct, with the t(4;14) group being characterized by deregulation of chromatin organization, actin filament, and microfilament movement. Recurrent RAS pathway mutations identified subclonal heterogeneity at a mutational level in both groups, with mutations being present as either dominant or minor subclones. The presence of subclonal diversity was confirmed at a single-cell level using other tumor-acquired mutations. These results are consistent with a distinct molecular pathogenesis underlying each subgroup and have important impacts on targeted treatment strategies. The Medical Research Council Myeloma IX trial is registered under ISRCTN68454111..
Hewish, M.
Stamp, G.
Puckey, L.
Shanley, S.
Costello, C.
Webb, J.
Hulkki Wilson, S.
Gonzalez de Castro, D.
Barbachano, Y.
Saffery, C.
Swanton, C.
Lord, C.J.
Reis-Filho, J.S.
Ashworth, A.
Chau, I.
Cunningham, D.
(2012). Impact of mismatch repair (MMR) testing on colorectal cancer (CRC) oncology clinical practice. J clin oncol,
Vol.30
(4_suppl),
p. 603.
show abstract
603 Background: MMR deficiency (dMMR) has been reported in 15% of CRC, but with a lower frequency in advanced disease. Most cases are due to sporadic MLH1 promoter hypermethylation (often with BRAF mutations), with a minority reflecting germline mutations in MLH1, MSH2, PMS2, or MSH6 (Lynch Syndrome [LS]). The Revised Bethesda Guidelines (RBG) are one means of selecting individuals at risk of LS for further assessment, but will miss a proportion of cases. METHODS: We screened all consenting patients for eligibility for CRC trials recruiting specific genetic aberrations, which included MMR assessment. RESULTS: Of 314 patients, immunohistochemistry (IHC) for MMR protein expression is complete on 171. Staining was reduced/absent in 19.3% of tests, and heterogeneous in 12.1%. The dMMR rate was 6.4%. 2 dMMR patients* were identified as at risk of LS, and referred to genetics by their treating clinician before IHC results were known. However 4 other cases† were not referred, and an underlying predisposition would have been missed without this unbiased approach. 4 patients developed metastatic disease, with none experiencing a partial response to chemotherapy thus far. (Table.) Conclusions: This data is representative of a practice with a high proportion of metastatic disease. It suggests that within oncology, an unbiased screening approach for LS is preferable. Whilst the RBG detect the majority of cases, they may be underutilised as other management issues take precedence in oncology clinics. A cost-effective alternative may be the introduction of a nurse-led programme to identify cases at risk, as is being introduced at our centre. A spectrum of clinical behavior exists amongst metastatic dMMR CRC, and larger numbers will reveal if this affects therapeutic response. [Table: see text]..
Postel-Vinay, S.
Vanhecke, E.
Olaussen, K.A.
Lord, C.J.
Ashworth, A.
Soria, J.-.
(2012). The potential of exploiting DNA-repair defects for optimizing lung cancer treatment. Nature reviews clinical oncology,
Vol.9
(3),
pp. 144-155.
Natrajan, R.
Mackay, A.
Lambros, M.B.
Weigelt, B.
Wilkerson, P.M.
Manie, E.
Grigoriadis, A.
A'Hern, R.
van der Groep, P.
Kozarewa, I.
Popova, T.
Mariani, O.
Turaljic, S.
Furney, S.J.
Marais, R.
Rodruigues, D.-.
Flora, A.C.
Wai, P.
Pawar, V.
McDade, S.
Carroll, J.
Stoppa-Lyonnet, D.
Green, A.R.
Ellis, I.O.
Swanton, C.
van Diest, P.
Delattre, O.
Lord, C.J.
Foulkes, W.D.
Vincent-Salomon, A.
Ashworth, A.
Stern, M.H.
Reis-Filho, J.S.
(2012). A whole-genome massively parallel sequencing analysis of BRCA1 mutant oestrogen receptor-negative and -positive breast cancers. J pathol,
Vol.227
(1),
pp. 29-41.
show abstract
full text
BRCA1 encodes a tumour suppressor protein that plays pivotal roles in homologous recombination (HR) DNA repair, cell-cycle checkpoints, and transcriptional regulation. BRCA1 germline mutations confer a high risk of early-onset breast and ovarian cancer. In more than 80% of cases, tumours arising in BRCA1 germline mutation carriers are oestrogen receptor (ER)-negative; however, up to 15% are ER-positive. It has been suggested that BRCA1 ER-positive breast cancers constitute sporadic cancers arising in the context of a BRCA1 germline mutation rather than being causally related to BRCA1 loss-of-function. Whole-genome massively parallel sequencing of ER-positive and ER-negative BRCA1 breast cancers, and their respective germline DNAs, was used to characterize the genetic landscape of BRCA1 cancers at base-pair resolution. Only BRCA1 germline mutations, somatic loss of the wild-type allele, and TP53 somatic mutations were recurrently found in the index cases. BRCA1 breast cancers displayed a mutational signature consistent with that caused by lack of HR DNA repair in both ER-positive and ER-negative cases. Sequencing analysis of independent cohorts of hereditary BRCA1 and sporadic non-BRCA1 breast cancers for the presence of recurrent pathogenic mutations and/or homozygous deletions found in the index cases revealed that DAPK3, TMEM135, KIAA1797, PDE4D, and GATA4 are potential additional drivers of breast cancers. This study demonstrates that BRCA1 pathogenic germline mutations coupled with somatic loss of the wild-type allele are not sufficient for hereditary breast cancers to display an ER-negative phenotype, and has led to the identification of three potential novel breast cancer genes (ie DAPK3, TMEM135, and GATA4)..
Turnbull, C.
Loveday, C.
Ramsay, E.
Hughes, D.
Ruark, E.
Frankum, J.R.
Warren-Perry, M.
Snape, K.
Eccles, D.
Evans, D.G.
Renwick, A.
Seal, S.
Lord, C.J.
Ashworth, A.
Reis-Filho, J.S.
Antoniou, A.C.
Rahman, N.
(2012). Newly discovered high-intermediate penetrance ovarian cancer susceptibility genes: clinical testing of RAD51D and RAD51C may have significant utility. International journal of gynecological cancer,
Vol.22,
pp. S42-S42.
Furney, S.J.
Turajlic, S.
Fenwick, K.
Lambros, M.B.
MacKay, A.
Ricken, G.
Mitsopoulos, C.
Kozarewa, I.
Hakas, J.
Zvelebil, M.
Lord, C.J.
Ashworth, A.
Reis-Filho, J.S.
Herlyn, M.
Murata, H.
Marais, R.
(2012). Genomic characterisation of acral melanoma cell lines. Pigment cell melanoma res,
Vol.25
(4),
pp. 488-492.
show abstract
Acral melanoma is a rare melanoma subtype with distinct epidemiological, clinical and genetic features. To determine if acral melanoma cell lines are representative of this melanoma subtype, six lines were analysed by whole-exome sequencing and array comparative genomic hybridisation. We demonstrate that the cell lines display a mutation rate that is comparable to that of published primary and metastatic acral melanomas and observe a mutational signature suggestive of UV-induced mutagenesis in two of the cell lines. Mutations were identified in oncogenes and tumour suppressors previously linked to melanoma including BRAF, NRAS, KIT, PTEN and TP53, in cancer genes not previously linked to melanoma and in genes linked to DNA repair such as BRCA1 and BRCA2. Our findings provide strong circumstantial evidence to suggest that acral melanoma cell lines and acral tumours share genetic features in common and that these cells are therefore valuable tools to investigate the biology of this aggressive melanoma subtype. Data are available at: http://rock.icr.ac.uk/collaborations/Furney_et_al_2012/..
Stratford, A.L.
Reipas, K.
Hu, K.
Fotovati, A.
Brough, R.
Frankum, J.
Takhar, M.
Watson, P.
Ashworth, A.
Lord, C.J.
Lasham, A.
Print, C.G.
Dunn, S.E.
(2012). Targeting p90 Ribosomal S6 Kinase Eliminates Tumor-Initiating Cells by Inactivating Y-Box Binding Protein-1 in Triple-Negative Breast Cancers. Stem cells,
Vol.30
(7),
pp. 1338-1348.
Brough, R.
Bajrami, I.
Vatcheva, R.
Natrajan, R.
Reis-Filho, J.S.
Lord, C.J.
Ashworth, A.
(2012). APRIN is a cell cycle specific BRCA2-interacting protein required for genome integrity and a predictor of outcome after chemotherapy in breast cancer. Embo journal,
Vol.31
(5),
pp. 1160-1176.
McDade, S.S.
Henry, A.E.
Pivato, G.P.
Kozarewa, I.
Mitsopoulos, C.
Fenwick, K.
Assiotis, I.
Hakas, J.
Zvelebil, M.
Orr, N.
Lord, C.J.
Patel, D.
Ashworth, A.
McCance, D.J.
(2012). Genome-wide analysis of p63 binding sites identifies AP-2 factors as co-regulators of epidermal differentiation. Nucleic acids res,
Vol.40
(15),
pp. 7190-7206.
show abstract
The p63 transcription factor (TP63) is critical in development, growth and differentiation of stratifying epithelia. This is highlighted by the severity of congenital abnormalities caused by TP63 mutations in humans, the dramatic phenotypes in knockout mice and de-regulation of TP63 expression in neoplasia altering the tumour suppressive roles of the TP53 family. In order to define the normal role played by TP63 and provide the basis for better understanding how this network is perturbed in disease, we used chromatin immunoprecipitation combined with massively parallel sequencing (ChIP-seq) to identify >7500 high-confidence TP63-binding regions across the entire genome, in primary human neonatal foreskin keratinocytes (HFKs). Using integrative strategies, we demonstrate that only a subset of these sites are bound by TP53 in response to DNA damage. We identify a role for TP63 in transcriptional regulation of multiple genes genetically linked to cleft palate and identify AP-2alpha (TFAP2A) as a co-regulator of a subset of these genes. We further demonstrate that AP-2gamma (TFAP2C) can bind a subset of these regions and that acute depletion of either TFAP2A or TFAP2C alone is sufficient to reduce terminal differentiation of organotypic epidermal skin equivalents, indicating overlapping physiological functions with TP63..
Iorns, E.
Ward, T.M.
Dean, S.
Jegg, A.
Thomas, D.
Murugaesu, N.
Sims, D.
Mitsopoulos, C.
Fenwick, K.
Kozarewa, I.
Naceur-Lombarelli, C.
Zvelebil, M.
Isacke, C.M.
Lord, C.J.
Ashworth, A.
Hnatyszyn, H.J.
Pegram, M.
Lippman, M.
(2012). Whole genome in vivo RNAi screening identifies the leukemia inhibitory factor receptor as a novel breast tumor suppressor. Breast cancer res treat,
Vol.135
(1),
pp. 79-91.
show abstract
Cancer is caused by mutations in oncogenes and tumor suppressor genes, resulting in the deregulation of processes fundamental to the normal behavior of cells. The identification and characterization of oncogenes and tumor suppressors has led to new treatment strategies that have significantly improved cancer outcome. The advent of next generation sequencing has allowed the elucidation of the fine structure of cancer genomes, however, the identification of pathogenic changes is complicated by the inherent genomic instability of cancer cells. Therefore, functional approaches for the identification of novel genes involved in the initiation and development of tumors are critical. Here we report the first whole human genome in vivo RNA interference screen to identify functionally important tumor suppressor genes. Using our novel approach, we identify previously validated tumor suppressor genes including TP53 and MNT, as well as several novel candidate tumor suppressor genes including leukemia inhibitory factor receptor (LIFR). We show that LIFR is a key novel tumor suppressor, whose deregulation may drive the transformation of a significant proportion of human breast cancers. These results demonstrate the power of genome wide in vivo RNAi screens as a method for identifying novel genes regulating tumorigenesis..
Daemen, A.
Wolf, D.M.
Korkola, J.E.
Griffith, O.L.
Frankum, J.R.
Brough, R.
Jakkula, L.R.
Wang, N.J.
Natrajan, R.
Reis-Filho, J.S.
Lord, C.J.
Ashworth, A.
Spellman, P.T.
Gray, J.W.
van't Veer, L.J.
(2012). Cross-platform pathway-based analysis identifies markers of response to the PARP inhibitor olaparib. Breast cancer research and treatment,
Vol.135
(2),
pp. 505-517.
Bajrami, I.
Kigozi, A.
Van Weverwijk, A.
Brough, R.
Frankum, J.
Lord, C.J.
Ashworth, A.
(2012). Synthetic lethality of PARP and NAMPT inhibition in triple-negative breast cancer cells. Embo mol med,
Vol.4
(10),
pp. 1087-1096.
show abstract
PARP inhibitors have been proposed as a potential targeted therapy for patients with triple-negative (ER-, PR-, HER2-negative) breast cancers. However, it is as yet unclear as to whether single agent or combination therapy using PARP inhibitors would be most beneficial. To better understand the mechanisms that determine the response to PARP inhibitors, we investigated whether enzymes involved in metabolism of the PARP substrate, β-NAD(+) , might alter the response to a clinical PARP inhibitor. Using an olaparib sensitization screen in a triple-negative (TN) breast cancer model, we identified nicotinamide phosphoribosyltransferase (NAMPT) as a non-redundant modifier of olaparib response. NAMPT is a rate-limiting enzyme involved in the generation of the PARP substrate β-NAD(+) and the suppression of β-NAD(+) levels by NAMPT inhibition most likely explains these observations. Importantly, the combination of a NAMPT small molecule inhibitor, FK866, with olaparib inhibited TN breast tumour growth in vivo to a greater extent than either single agent alone suggesting that assessing NAMPT/PARP inhibitor combinations for the treatment of TN breast cancer may be warranted..
Rehman, F.L.
Lord, C.J.
Ashworth, A.
(2012). The Promise of Combining Inhibition of PI3K and PARP as Cancer Therapy. Cancer discovery,
Vol.2
(11),
pp. 982-984.
Riffell, J.L.
Lord, C.J.
Ashworth, A.
(2012). Tankyrase-targeted therapeutics: expanding opportunities in the PARP family. Nat rev drug discov,
Vol.11
(12),
pp. 923-936.
show abstract
The poly(ADP-ribose) polymerase (PARP) protein superfamily has wide-ranging roles in cellular processes such as DNA repair and WNT signalling. Efforts to pharmacologically target PARP enzymes have largely focused on PARP1 and the closely related PARP2, but recent work highlighting the role of another family member, tankyrase 1 (TANK1; also known as PARP5A and ARTD5), in the control of WNT signalling has fuelled interest in the development of additional inhibitors to target this enzyme class. Tankyrase function is also implicated in other processes such as the regulation of telomere length, lung fibrogenesis and myelination, suggesting that tankyrase inhibitors could have broad clinical utility. Here, we discuss the biology of tankyrases and the discovery of tankyrase-specific inhibitors. We also consider the challenges that lie ahead for the clinical development of PARP family inhibitors in general..
Mackay, A.G.
Natrajan, R.
Maher, C.
Lambros, M.
Weigelt, B.
Nava-Rodrigues, D.
Wilkerson, P.
CampionFlora, A.
Kozarewa, I.
Fenwick, K.
Campbell, J.
Chen, L.
Lord, C.J.
Gauthier, A.
Pawar, V.
Mariani, O.
Vincent-Salomon, A.
Reis-Filho, J.
(2012). Integrative genomic and transcriptomic analysis of metaplastic carcinomas of the breast. Cancer research,
Vol.72.
Ng, C.K.
Gauthier, A.
Mackay, A.
Lambros, M.B.
Rodrigues, D.N.
Arnoud, L.
Lacroix-Triki, M.
Penault-Llorca, F.
Baranzelli, M.C.
Sastre-Garau, X.
Lord, C.J.
Zvelebil, M.
Mitsopoulos, C.
Ashworth, A.
Natrajan, R.
Weigelt, B.
Delattre, O.
Cottu, P.
Reis-Filho, J.S.
Vincent-Salomon, A.
(2012). Genomic characterisation of invasive breast cancers with heterogeneous HER2 gene amplification. Cancer research,
Vol.72.
Murugaesu, N.
Iravani, M.
Johnson, D.
Antonopoulos, A.
Sims, D.
Fenwick, K.
Mitsopoulos, C.
Gao, Q.
Orr, N.
Zvelebil, M.
Haslam, S.
Dell, A.
Lord, C.
Ashworth, A.
Isacke, C.
(2012). An in vivo functional screen to identify metastasis suppressor genes. Cancer research,
Vol.72.
Lord, C.J.
Ashworth, A.
(2012). The DNA damage response and cancer therapy. Nature,
Vol.481
(7381),
pp. 287-294.
show abstract
Genomic instability is one of the most pervasive characteristics of tumour cells and is probably the combined effect of DNA damage, tumour-specific DNA repair defects, and a failure to stop or stall the cell cycle before the damaged DNA is passed on to daughter cells. Although these processes drive genomic instability and ultimately the disease process, they also provide therapeutic opportunities. A better understanding of the cellular response to DNA damage will not only inform our knowledge of cancer development but also help to refine the classification as well as the treatment of the disease..
Mendes-Pereira, A.M.
Sims, D.
Dexter, T.
Fenwick, K.
Assiotis, I.
Kozarewa, I.
Mitsopoulos, C.
Hakas, J.
Zvelebil, M.
Lord, C.J.
Ashworth, A.
(2012). Genome-wide functional screen identifies a compendium of genes affecting sensitivity to tamoxifen. Proc natl acad sci u s a,
Vol.109
(8),
pp. 2730-2735.
show abstract
Therapies that target estrogen signaling have made a very considerable contribution to reducing mortality from breast cancer. However, resistance to tamoxifen remains a major clinical problem. Here we have used a genome-wide functional profiling approach to identify multiple genes that confer resistance or sensitivity to tamoxifen. Combining whole-genome shRNA screening with massively parallel sequencing, we have profiled the impact of more than 56,670 RNA interference reagents targeting 16,487 genes on the cellular response to tamoxifen. This screen, along with subsequent validation experiments, identifies a compendium of genes whose silencing causes tamoxifen resistance (including BAP1, CLPP, GPRC5D, NAE1, NF1, NIPBL, NSD1, RAD21, RARG, SMC3, and UBA3) and also a set of genes whose silencing causes sensitivity to this endocrine agent (C10orf72, C15orf55/NUT, EDF1, ING5, KRAS, NOC3L, PPP1R15B, RRAS2, TMPRSS2, and TPM4). Multiple individual genes, including NF1, a regulator of RAS signaling, also correlate with clinical outcome after tamoxifen treatment..
Mendes-Pereira, A.M.
Lord, C.J.
Ashworth, A.
(2012). NLK Is a Novel Therapeutic Target for PTEN Deficient Tumour Cells. Plos one,
Vol.7
(10).
Orr, N.
Lemnrau, A.
Cooke, R.
Fletcher, O.
Tomczyk, K.
Jones, M.
Johnson, N.
Lord, C.J.
Mitsopoulos, C.
Zvelebil, M.
McDade, S.S.
Buck, G.
Blancher, C.
Consortium, K.C.
Trainer, A.H.
James, P.A.
Bojesen, S.E.
Bokmand, S.
Nevanlinna, H.
Mattson, J.
Friedman, E.
Laitman, Y.
Palli, D.
Masala, G.
Zanna, I.
Ottini, L.
Giannini, G.
Hollestelle, A.
Ouweland, A.M.
Novakovic, S.
Krajc, M.
Gago-Dominguez, M.
Castelao, J.E.
Olsson, H.
Hedenfalk, I.
Easton, D.F.
Pharoah, P.D.
Dunning, A.M.
Bishop, D.T.
Neuhausen, S.L.
Steele, L.
Houlston, R.S.
Garcia-Closas, M.
Ashworth, A.
Swerdlow, A.J.
(2012). Genome-wide association study identifies a common variant in RAD51B associated with male breast cancer risk. Nat genet,
Vol.44,
pp. 1182-1184.
show abstract
We conducted a genome-wide association study of male breast cancer comprising 823 cases and 2,795 controls of European ancestry, with validation in independent sample sets totaling 438 cases and 474 controls. A SNP in RAD51B at 14q24.1 was significantly associated with male breast cancer risk (P = 3.02 x 10(-13); odds ratio (OR) = 1.57). We also refine association at 16q12.1 to a SNP within TOX3 (P = 3.87 x 10(-15); OR = 1.50)..
Kozarewa, I.
Rosa-Rosa, J.M.
Wardell, C.P.
Walker, B.A.
Fenwick, K.
Assiotis, I.
Mitsopoulos, C.
Zvelebil, M.
Morgan, G.J.
Ashworth, A.
Lord, C.J.
(2012). A modified method for whole exome resequencing from minimal amounts of starting DNA. Plos one,
Vol.7
(3),
p. e32617.
show abstract
Next generation DNA sequencing (NGS) technologies have revolutionized the pace at which whole genome and exome sequences can be generated. However, despite these advances, many of the methods for targeted resequencing, such as the generation of high-depth exome sequences, are somewhat limited by the relatively large amounts of starting DNA that are normally required. In the case of tumour analysis this is particularly pertinent as many tumour biopsies often return submicrogram quantities of DNA, especially when tumours are microdissected prior to analysis. Here, we present a method for exome capture and resequencing using as little as 50 ng of starting DNA. The sequencing libraries generated by this minimal starting amount (MSA-Cap) method generate datasets that are comparable to standard amount (SA) whole exome libraries that use three micrograms of starting DNA. This method, which can be performed in most laboratories using commonly available reagents, has the potential to enhance large scale profiling efforts such as the resequencing of tumour exomes..
Martin, S.A.
Hewish, M.
Sims, D.
Lord, C.J.
Ashworth, A.
(2011). Parallel high-throughput RNA interference screens identify PINK1 as a potential therapeutic target for the treatment of DNA mismatch repair-deficient cancers. Cancer res,
Vol.71
(5),
pp. 1836-1848.
show abstract
Synthetic lethal approaches to cancer treatment have the potential to deliver relatively large therapeutic windows and therefore significant patient benefit. To identify potential therapeutic approaches for cancers deficient in DNA mismatch repair (MMR), we have carried out parallel high-throughput RNA interference screens using tumor cell models of MSH2- and MLH1-related MMR deficiency. We show that silencing of the PTEN-induced putative kinase 1 (PINK1), is synthetically lethal with MMR deficiency in cells with MSH2, MLH1, or MSH6 dysfunction. Inhibition of PINK1 in an MMR-deficient background results in an elevation of reactive oxygen species and the accumulation of both nuclear and mitochondrial oxidative DNA lesions, which likely limit cell viability. Therefore, PINK1 represents a potential therapeutic target for the treatment of cancers characterized by MMR deficiency caused by a range of different gene deficiencies..
Ashworth, A.
Lord, C.J.
Reis-Filho, J.S.
(2011). Genetic interactions in cancer progression and treatment. Cell,
Vol.145
(1),
pp. 30-38.
show abstract
As cancer cell genomes are unveiled at a breathtaking pace, the genetic principles at play in cancer are emerging in all their complexity, prompting the assessment of classical genetic interaction models. Here, we discuss the implications of these findings for cancer progression and heterogeneity and for the development of new therapeutic approaches..
Cerone, M.A.
Burgess, D.J.
Naceur-Lombardelli, C.
Lord, C.J.
Ashworth, A.
(2011). High-throughput RNAi screening reveals novel regulators of telomerase. Cancer res,
Vol.71
(9),
pp. 3328-3340.
show abstract
Telomerase is considered an attractive anticancer target on the basis of its common and specific activation in most human cancers. While direct telomerase inhibition is being explored as a therapeutic strategy, alternative strategies to target regulators of telomerase that could disrupt telomere maintenance and cancer cell proliferation are not yet available. Here, we report the findings of a high-throughput functional RNA interference screen to globally profile the contribution of kinases to telomerase activity (TA). This analysis identified a number of novel telomerase modulators, including ERK8 kinase, whose inhibition reduces TA and elicited characteristics of telomere dysfunction. Given that kinases represent attractive drug targets, we addressed the therapeutic implications of our findings, such as demonstrating how limiting TA via kinase blockade could sensitize cells to inhibition of the telomere-associated protein tankyrase. Taken together, our findings suggest novel combinatorial approaches to targeting telomere maintenance as a strategy for cancer therapy..
Brough, R.
Frankum, J.R.
Costa-Cabral, S.
Lord, C.J.
Ashworth, A.
(2011). Searching for synthetic lethality in cancer. Curr opin genet dev,
Vol.21
(1),
pp. 34-41.
show abstract
The incentive to develop personalised therapy for cancer treatment is driven by the premise that it will increase therapeutic efficacy and reduce toxicity. Understanding the underlying cellular and molecular basis of the disease has been extremely important in the design of these novel therapies; however, identifying new drug targets for personalised therapies remains problematic. This review describes how the biological concept of synthetic lethality has been successfully implemented to identify new therapeutic approaches and targets in models from yeast through to human cells. We also discuss how recent technical advances combined with an increased understanding of the complexity of cellular networks may facilitate therapeutic advances in the future..
Rajan, N.
Burn, J.
Langtry, J.
Sieber-Blum, M.
Lord, C.J.
Ashworth, A.
(2011). Transition from cylindroma to spiradenoma in CYLD-defective tumours is associated with reduced DKK2 expression. J pathol,
Vol.224
(3),
pp. 309-321.
show abstract
Patients carrying heterozygous germline truncating mutations in the CYLD gene develop multiple primary hair follicle-related tumours. A highly patterned tumour, termed cylindroma, and a highly disorganized tumour, termed spiradenoma, may both develop in the same patient. Furthermore, histological features of both tumour types have been described within the same tumour specimen. We used three-dimensional computer-aided reconstruction of these tumours to demonstrate contiguous growth of cylindromas into spiradenomas, thus suggesting a transition between the two tumour types. To explore factors that may influence cutaneous tumour patterning, genome-wide transcriptomic analysis of 32 CYLD-defective tumours was performed. Overexpression of the Wnt/β-catenin signalling pathway was observed relative to normal perilesional tissue. Morphometric analysis was used to investigate the relationship between Wnt pathway-related gene expression and tumour organization. This revealed an association between reduced Dickkopf 2 (DKK2-a negative regulator of the Wnt/β-catenin signalling pathway) expression and loss of tumour patterning. Reduced DKK2 expression was associated with methylation of the DKK2 gene promoter in the majority of tumour samples assayed. RNA interference-mediated silencing of DKK2 expression in cylindroma primary cell cultures caused an increase in colony formation, cell viability, and anchorage-independent growth. Using these data, we propose a model where epigenetic programming may influence tumour patterning in patients with CYLD mutations..
Loveday, C.
Turnbull, C.
Ramsay, E.
Hughes, D.
Ruark, E.
Frankum, J.R.
Bowden, G.
Kalmyrzaev, B.
Warren-Perry, M.
Snape, K.
Adlard, J.W.
Barwell, J.
Berg, J.
Brady, A.F.
Brewer, C.
Brice, G.
Chapman, C.
Cook, J.
Davidson, R.
Donaldson, A.
Douglas, F.
Greenhalgh, L.
Henderson, A.
Izatt, L.
Kumar, A.
Lalloo, F.
Miedzybrodzka, Z.
Morrison, P.J.
Paterson, J.
Porteous, M.
Rogers, M.T.
Shanley, S.
Walker, L.
Breast Cancer Susceptibility Collaboration (UK),
Eccles, D.
Evans, D.G.
Renwick, A.
Seal, S.
Lord, C.J.
Ashworth, A.
Reis-Filho, J.S.
Antoniou, A.C.
Rahman, N.
(2011). Germline mutations in RAD51D confer susceptibility to ovarian cancer. Nat genet,
Vol.43
(9),
pp. 879-882.
show abstract
full text
Recently, RAD51C mutations were identified in families with breast and ovarian cancer. This observation prompted us to investigate the role of RAD51D in cancer susceptibility. We identified eight inactivating RAD51D mutations in unrelated individuals from 911 breast-ovarian cancer families compared with one inactivating mutation identified in 1,060 controls (P = 0.01). The association found here was principally with ovarian cancer, with three mutations identified in the 59 pedigrees with three or more individuals with ovarian cancer (P = 0.0005). The relative risk of ovarian cancer for RAD51D mutation carriers was estimated to be 6.30 (95% CI 2.86-13.85, P = 4.8 × 10(-6)). By contrast, we estimated the relative risk of breast cancer to be 1.32 (95% CI 0.59-2.96, P = 0.50). These data indicate that RAD51D mutation testing may have clinical utility in individuals with ovarian cancer and their families. Moreover, we show that cells deficient in RAD51D are sensitive to treatment with a PARP inhibitor, suggesting a possible therapeutic approach for cancers arising in RAD51D mutation carriers..
Rovillain, E.
Mansfield, L.
Lord, C.J.
Ashworth, A.
Jat, P.S.
(2011). An RNA interference screen for identifying downstream effectors of the p53 and pRB tumour suppressor pathways involved in senescence. Bmc genomics,
Vol.12,
p. 355.
show abstract
BACKGROUND: Cellular senescence is an irreversible cell cycle arrest that normal cells undergo in response to progressive shortening of telomeres, changes in telomeric structure, oncogene activation or oxidative stress and acts as an important tumour suppressor mechanism. RESULTS: To identify the downstream effectors of the p53-p21 and p16-pRB tumour suppressor pathways crucial for mediating entry into senescence, we have carried out a loss-of-function RNA interference screen in conditionally immortalised human fibroblasts that can be induced to rapidly undergo senescence, whereas in primary cultures senescence is stochastic and occurs asynchronously. These cells are immortal but undergo a rapid irreversible arrest upon activation of the p53-p21 and p16-pRB pathways that can be readily bypassed upon their inactivation. The primary screen identified 112 known genes including p53 and another 29 shRNAmirs targetting as yet unidentified loci. Comparison of these known targets with genes known to be up-regulated upon senescence in these cells, by micro-array expression profiling, identified 4 common genes TMEM9B, ATXN10, LAYN and LTBP2/3. Direct silencing of these common genes, using lentiviral shRNAmirs, bypassed senescence in the conditionally immortalised cells. CONCLUSION: The senescence bypass screen identified TMEM9B, ATXN10, LAYN and LTBP2/3 as novel downstream effectors of the p53-p21 and p16-pRB tumour suppressor pathways. Although none of them has previously been linked to cellular senescence, TMEM9B has been suggested to be an upstream activator of NF-κB signalling which has been found to have a causal role in promoting senescence. Future studies will focus on determining on how many of the other primary hits also have a casual role in senescence and what is the mechanism of action..
Brough, R.
Frankum, J.R.
Sims, D.
Mackay, A.
Mendes-Pereira, A.M.
Bajrami, I.
Costa-Cabral, S.
Rafiq, R.
Ahmad, A.S.
Cerone, M.A.
Natrajan, R.
Sharpe, R.
Shiu, K.-.
Wetterskog, D.
Dedes, K.J.
Lambros, M.B.
Rawjee, T.
Linardopoulos, S.
Reis-Filho, J.S.
Turner, N.C.
Lord, C.J.
Ashworth, A.
(2011). Functional viability profiles of breast cancer. Cancer discov,
Vol.1
(3),
pp. 260-273.
show abstract
UNLABELLED: The design of targeted therapeutic strategies for cancer has largely been driven by the identification of tumor-specific genetic changes. However, the large number of genetic alterations present in tumor cells means that it is difficult to discriminate between genes that are critical for maintaining the disease state and those that are merely coincidental. Even when critical genes can be identified, directly targeting these is often challenging, meaning that alternative strategies such as exploiting synthetic lethality may be beneficial. To address these issues, we have carried out a functional genetic screen in >30 commonly used models of breast cancer to identify genes critical to the growth of specific breast cancer subtypes. In particular, we describe potential new therapeutic targets for PTEN-mutated cancers and for estrogen receptor-positive breast cancers. We also show that large-scale functional profiling allows the classification of breast cancers into subgroups distinct from established subtypes. SIGNIFICANCE: Despite the wealth of molecular profiling data that describe breast tumors and breast tumor cell models, our understanding of the fundamental genetic dependencies in this disease is relatively poor. Using high-throughput RNA interference screening of a series of pharmacologically tractable genes, we have generated comprehensive functional viability profiles for a wide panel of commonly used breast tumor cell models. Analysis of these profiles identifies a series of novel genetic dependencies, including that of PTEN-null breast tumor cells upon mitotic checkpoint kinases, and provides a framework upon which additional dependencies and candidate therapeutic targets may be identified..
Wilkerson, P.M.
Dedes, K.J.
Wetterskog, D.
Mackay, A.
Lambros, M.B.
Mansour, M.
Frankum, J.
Lord, C.J.
Natrajan, R.
Ashworth, A.
Reis-Filho, J.S.
(2011). Functional characterization of EMSY gene amplification in human cancers. J pathol,
Vol.225
(1),
pp. 29-42.
show abstract
The 11q13-q14 locus is frequently amplified in human cancers, with a complex structure harbouring multiple potential oncogenic drivers. The EMSY gene has been proposed as a driver of the third core of the 11q13-q14 amplicon. This gene encodes a protein reported to be a BRCA2-binding partner, which when over-expressed would lead to impairment of BRCA2 functions and could constitute a mechanism for BRCA2 inactivation in non-hereditary breast and ovarian cancers. We hypothesized that if EMSY amplification abrogates BRCA2 functions, cells harbouring this aberration would be unable to elicit competent homologous recombination DNA repair and, therefore, may have increased sensitivity to genotoxic therapies and potent PARP inhibitors. Microarray-based comparative genomic hybridization of cell lines from distinct tumour sites, including breast, ovary, pancreas, oesophagus, lung and the oral cavity, led to the identification of 10 cell lines with EMSY amplification and 18 without. EMSY amplification was shown to correlate with EMSY mRNA levels, although not all cell lines harbouring EMSY amplification displayed EMSY mRNA or protein over-expression. RNA interference-mediated silencing of EMSY did not lead to a reduction in cell viability in tumour models harbouring EMSY amplification. Cell lines with and without EMSY amplification displayed a similar ability to elicit RAD51 foci in response to DNA damaging agents, and similar sensitivity to cisplatin and olaparib. Taken together, this suggests that EMSY is unlikely to be a driver of the 11q13-q14 amplicon and does not have a dominant role in modulating the response to agents targeting cells with defective homologous recombination..
Vaughan, S.
Coward, J.I.
Bast, R.C.
Berchuck, A.
Berek, J.S.
Brenton, J.D.
Coukos, G.
Crum, C.C.
Drapkin, R.
Etemadmoghadam, D.
Friedlander, M.
Gabra, H.
Kaye, S.B.
Lord, C.J.
Lengyel, E.
Levine, D.A.
McNeish, I.A.
Menon, U.
Mills, G.B.
Nephew, K.P.
Oza, A.M.
Sood, A.K.
Stronach, E.A.
Walczak, H.
Bowtell, D.D.
Balkwill, F.R.
(2011). Rethinking ovarian cancer: recommendations for improving outcomes. Nature reviews cancer,
Vol.11
(10),
pp. 719-7.
Rajan, N.
Elliott, R.
Clewes, O.
Mackay, A.
Reis-Filho, J.S.
Burn, J.
Langtry, J.
Sieber-Blum, M.
Lord, C.J.
Ashworth, A.
(2011). Dysregulated TRK signalling is a therapeutic target in CYLD defective tumours. Oncogene,
Vol.30
(41),
pp. 4243-4260.
show abstract
Individuals with germline mutations in the tumour-suppressor gene CYLD are at high risk of developing disfiguring cutaneous appendageal tumours, the defining tumour being the highly organised cylindroma. Here, we analysed CYLD mutant tumour genomes by array comparative genomic hybridisation and gene expression microarray analysis. CYLD mutant tumours were characterised by an absence of copy-number aberrations apart from LOH chromosome 16q, the genomic location of the CYLD gene. Gene expression profiling of CYLD mutant tumours showed dysregulated tropomyosin kinase (TRK) signalling, with overexpression of TRKB and TRKC in tumours when compared with perilesional skin. Immunohistochemical analysis of a tumour microarray showed strong membranous TRKB and TRKC staining in cylindromas, as well as elevated levels of ERK phosphorylation and BCL2 expression. Membranous TRKC overexpression was also observed in 70% of sporadic BCCs. RNA interference-mediated silencing of TRKB and TRKC, as well as treatment with the small-molecule TRK inhibitor lestaurtinib, reduced colony formation and proliferation in 3D primary cell cultures established from CYLD mutant tumours. These results suggest that TRK inhibition could be used as a strategy to treat tumours with loss of functional CYLD..
Hewish, M.
Lord, C.J.
Martin, S.A.
Cunningham, D.
Ashworth, A.
(2011). High-throughput drug screens identify novel synthetic lethal interactions with MLH1-deficient cancers. Cancer research,
Vol.71.
Wetterskog, D.
Shiu, K.-.
Chong, I.
Meijer, T.
Natrajan, R.
Lord, C.J.
Ashworth, A.
Reis-Filho, J.S.
(2011). Identification of novel genes and pathways involved in resistance to HER2-targeting agents in breast cancer. Cancer research,
Vol.71.
Natrajan, R.
Maher, C.A.
Lambros, M.B.
Pawar, V.
Wetterskog, D.
Marchio, C.
Barbashina, V.
Geyer, F.C.
Fenwick, K.
Kozarewa, I.
Mackay, A.
Hakas, J.
Mitsopoulos, K.
Hardisson, D.
Sapino, A.
Vincent-Salomon, A.
Lord, C.J.
Chinniayan, A.
Ashworth, A.
Reis-Filho, J.S.
(2011). Massively parallel RNA sequencing analysis of micropapillary carcinomas of the breast. Cancer research,
Vol.71.
Daemen, A.
Wolf, D.M.
Korkola, J.E.
Griffith, O.L.
Frankum, J.R.
Jakkula, L.R.
Wang, N.J.
Natrajan, R.
Reis-Filho, J.S.
Lord, C.J.
Ashworth, A.
Gray, J.W.
Spellman, P.T.
van't Veer, L.
(2011). Identification of Biomarkers in Breast Cancer for Prediction of Response to PARP Inhibitor Olaparib. Cancer research,
Vol.71.
Wilkerson, P.
Dedes, K.J.
Wetterskog, D.
Natrajan, R.
Lambros, M.B.
Mackay, A.
Lord, C.J.
Ashworth, A.
Reis-Filho, J.S.
(2011). EMSY amplification and overexpression is not associated with defective homologous recombination and does not predict sensitivity to cisplatin or PARP inhibitors. Cancer research,
Vol.71.
Ha, K.C.
Lalonde, E.
Li, L.
Cavallone, L.
Natrajan, R.
Lambros, M.B.
Mitsopoulos, C.
Hakas, J.
Kozarewa, I.
Fenwick, K.
Lord, C.J.
Ashworth, A.
Vincent-Salomon, A.
Basik, M.
Reis-Filho, J.S.
Majewski, J.
Foulkes, W.D.
(2011). Identification of gene fusion transcripts by transcriptome sequencing in BRCA1-mutated breast cancers and cell lines. Bmc med genomics,
Vol.4,
p. 75.
show abstract
BACKGROUND: Gene fusions arising from chromosomal translocations have been implicated in cancer. However, the role of gene fusions in BRCA1-related breast cancers is not well understood. Mutations in BRCA1 are associated with an increased risk for breast cancer (up to 80% lifetime risk) and ovarian cancer (up to 50%). We sought to identify putative gene fusions in the transcriptomes of these cancers using high-throughput RNA sequencing (RNA-Seq). METHODS: We used Illumina sequencing technology to sequence the transcriptomes of five BRCA1-mutated breast cancer cell lines, three BRCA1-mutated primary tumors, two secretory breast cancer primary tumors and one non-tumorigenic breast epithelial cell line. Using a bioinformatics approach, our initial attempt at discovering putative gene fusions relied on analyzing single-end reads and identifying reads that aligned across exons of two different genes. Subsequently, latter samples were sequenced with paired-end reads and at longer cycles (producing longer reads). We then refined our approach by identifying misaligned paired reads, which may flank a putative gene fusion junction. RESULTS: As a proof of concept, we were able to identify two previously characterized gene fusions in our samples using both single-end and paired-end approaches. In addition, we identified three novel in-frame fusions, but none were recurrent. Two of the candidates, WWC1-ADRBK2 in HCC3153 cell line and ADNP-C20orf132 in a primary tumor, were confirmed by Sanger sequencing and RT-PCR. RNA-Seq expression profiling of these two fusions showed a distinct overexpression of the 3' partner genes, suggesting that its expression may be under the control of the 5' partner gene's regulatory elements. CONCLUSIONS: In this study, we used both single-end and paired-end sequencing strategies to discover gene fusions in breast cancer transcriptomes with BRCA1 mutations. We found that the use of paired-end reads is an effective tool for transcriptome profiling of gene fusions. Our findings suggest that while gene fusions are present in some BRCA1-mutated breast cancers, they are infrequent and not recurrent. However, private fusions may still be valuable as potential patient-specific biomarkers for diagnosis and treatment..
Tanas, M.R.
Sboner, A.
Oliveira, A.M.
Erickson-Johnson, M.R.
Hespelt, J.
Hanwright, P.J.
Flanagan, J.
Luo, Y.
Fenwick, K.
Natrajan, R.
Mitsopoulos, C.
Zvelebil, M.
Hoch, B.L.
Weiss, S.W.
Debiec-Rychter, M.
Sciot, R.
West, R.B.
Lazar, A.J.
Ashworth, A.
Reis-Filho, J.S.
Lord, C.J.
Gerstein, M.B.
Rubin, M.A.
Rubin, B.P.
(2011). Identification of a disease-defining gene fusion in epithelioid hemangioendothelioma. Sci transl med,
Vol.3
(98),
p. 98ra82.
show abstract
Integrating transcriptomic sequencing with conventional cytogenetics, we identified WWTR1 (WW domain-containing transcription regulator 1) (3q25) and CAMTA1 (calmodulin-binding transcription activator 1) (1p36) as the two genes involved in the t(1;3)(p36;q25) chromosomal translocation that is characteristic of epithelioid hemangioendothelioma (EHE), a vascular sarcoma. This WWTR1/CAMTA1 gene fusion is under the transcriptional control of the WWTR1 promoter and encodes a putative chimeric transcription factor that joins the amino terminus of WWTR1, a protein that is highly expressed in endothelial cells, in-frame to the carboxyl terminus of CAMTA1, a protein that is normally expressed only in brain. Thus, CAMTA1 expression is activated inappropriately through a promoter-switch mechanism. The gene fusion is present in virtually all EHEs tested but is absent from all other vascular neoplasms, demonstrating it to be a disease-defining genetic alteration. A sensitive and specific break-apart fluorescence in situ hybridization assay was also developed to detect the translocation and will assist in the evaluation of this diagnostically challenging neoplasm. The chimeric WWTR1/CAMTA1 transcription factor may represent a therapeutic target for EHE and offers the opportunity to shed light on the functions of two poorly characterized proteins..
Barber, L.J.
Rosa Rosa, J.M.
Kozarewa, I.
Fenwick, K.
Assiotis, I.
Mitsopoulos, C.
Sims, D.
Hakas, J.
Zvelebil, M.
Lord, C.J.
Ashworth, A.
(2011). Comprehensive genomic analysis of a BRCA2 deficient human pancreatic cancer. Plos one,
Vol.6
(7),
p. e21639.
show abstract
Capan-1 is a well-characterised BRCA2-deficient human cell line isolated from a liver metastasis of a pancreatic adenocarcinoma. Here we report a genome-wide assessment of structural variations and high-depth exome characterization of single nucleotide variants and small insertion/deletions in Capan-1. To identify potential somatic and tumour-associated variations in the absence of a matched-normal cell line, we devised a novel method based on the analysis of HapMap samples. We demonstrate that Capan-1 has one of the most rearranged genomes sequenced to date. Furthermore, small insertions and deletions are detected more frequently in the context of short sequence repeats than in other genomes. We also identify a number of novel mutations that may represent genetic changes that have contributed to tumour progression. These data provide insight into the genomic effects of loss of BRCA2 function..
Sims, D.
Mendes-Pereira, A.M.
Frankum, J.
Burgess, D.
Cerone, M.-.
Lombardelli, C.
Mitsopoulos, C.
Hakas, J.
Murugaesu, N.
Isacke, C.M.
Fenwick, K.
Assiotis, I.
Kozarewa, I.
Zvelebil, M.
Ashworth, A.
Lord, C.J.
(2011). High-throughput RNA interference screening using pooled shRNA libraries and next generation sequencing. Genome biology,
Vol.12
(10).
Martin, S.A.
Hewish, M.
Lord, C.J.
Ashworth, A.
(2010). Genomic instability and the selection of treatments for cancer. J pathol,
Vol.220
(2),
pp. 281-289.
show abstract
A critical link exists between DNA mutation and chromosomal rearrangements (genomic instability) and cancer development. This genomic instability can manifest itself as small changes at the nucleotide level or as gross chromosomal alterations. Mutations in the genes that encode DNA damage response proteins are responsible for a variety of genomic instability syndromes including hereditary non-polyposis colorectal carcinoma, Bloom's syndrome, ataxia-telangiectasia, BRCA-associated breast and ovarian cancers and Fanconi anaemia. Similarly, epigenetic silencing of genes associated with the maintenance of genomic stability have also been implicated in the pathogenesis of cancer. Here, we discuss how different tumours may be classified not only by tumour site but also by the type of underlying genetic instability. This type of classification may assist in the optimization of existing treatment regimens as well as informing the development of new therapeutic approaches..
Martin, S.A.
Lord, C.J.
Ashworth, A.
(2010). Therapeutic Targeting of the DNA Mismatch Repair Pathway. Clinical cancer research,
Vol.16
(21),
pp. 5107-7.
Hewish, M.
Lord, C.J.
Martin, S.A.
Cunningham, D.
Ashworth, A.
(2010). Mismatch repair deficient colorectal cancer in the era of personalized treatment. Nat rev clin oncol,
Vol.7
(4),
pp. 197-208.
show abstract
The molecular and genetic subtyping of cancer has allowed the emergence of individualized therapies. This approach could potentially deliver treatments that have both increased efficacy as well as reduced toxicity. A well-defined subtype of colorectal cancer (CRC) is characterized by a deficiency in the mismatch repair (MMR) pathway. MMR deficiency not only contributes to the pathogenesis of a large proportion of CRC, but also determines the response to many of the drugs that are frequently used to treat this disease. In this Review we describe the MMR deficient phenotype and discuss how a deficiency in this DNA repair process may impact on the management of CRC, including surgery, adjuvant chemotherapy and the choice of systemic agents for the palliation of advanced disease. We also discuss how the DNA repair defect in MMR deficient CRC could be exploited in the development of novel therapeutic strategies..
Sourisseau, T.
Maniotis, D.
McCarthy, A.
Tang, C.
Lord, C.J.
Ashworth, A.
Linardopoulos, S.
(2010). Aurora-A expressing tumour cells are deficient for homology-directed DNA double strand-break repair and sensitive to PARP inhibition. Embo mol med,
Vol.2
(4),
pp. 130-142.
show abstract
The protein kinase Aurora-A is a major regulator of the cell cycle that orchestrates mitotic entry and is required for the assembly of a functional mitotic spindle. Overexpression of Aurora-A has been strongly linked with oncogenesis and this has led to considerable efforts at therapeutic targeting of the kinase activity of this protein. However, the exact mechanism by which Aurora-A promotes oncogenesis remains unclear. Here, we show that Aurora-A modulates the repair of DNA double-strand breaks (DSBs). Aurora-A expression inhibits RAD51 recruitment to DNA DSBs, decreases DSB repair by homologous recombination and sensitizes cancer cells to PARP inhibition. This impairment of RAD51 function requires inhibition of CHK1 by Polo-like kinase 1 (PLK1). These results identify a novel function of Aurora-A in modulating the response to DNA DSB that likely contributes to carcinogenesis and suggest a novel therapeutic approach to the treatment of cancers overexpressing this protein..
Natrajan, R.
Weigelt, B.
Mackay, A.
Geyer, F.C.
Grigoriadis, A.
Tan, D.S.
Jones, C.
Lord, C.J.
Vatcheva, R.
Rodriguez-Pinilla, S.M.
Palacios, J.
Ashworth, A.
Reis-Filho, J.S.
(2010). An integrative genomic and transcriptomic analysis reveals molecular pathways and networks regulated by copy number aberrations in basal-like, HER2 and luminal cancers. Breast cancer res treat,
Vol.121
(3),
pp. 575-589.
show abstract
Breast cancer is a heterogeneous disease caused by the accumulation of genetic changes in neoplastic cells. We hypothesised that molecular subtypes of breast cancer may be driven by specific constellations of genes whose expression is regulated by gene copy number aberrations. To address this question, we analysed a series of 48 microdissected grade III ductal carcinomas using high resolution microarray comparative genomic hybridisation and mRNA expression arrays. There were 5,931 genes whose expression significantly correlates with copy number identified; out of these, 1,897 genes were significantly differentially expressed between basal-like, HER2 and luminal tumours. Ingenuity Pathway Analysis (IPA) revealed that 'G1/S cell cycle regulation' and 'BRCA1 in DNA damage control' pathways were significantly enriched for genes whose expression correlates with copy number and are differentially expressed between the molecular subtypes of breast cancer. IPA of genes whose expression significantly correlates with copy number in each molecular subtype individually revealed that canonical pathways involved in oestrogen receptor (ER) signalling and DNA repair are enriched for these genes. We also identified 32, 157 and 265 genes significantly overexpressed when amplified in basal-like, HER2 and luminal cancers, respectively. These lists include known and novel potential therapeutic targets (e.g. HER2 and PPM1D in HER2 cancers). Our results provide strong circumstantial evidence that different patterns of genetic aberrations in distinct molecular subtypes of breast cancer contribute to their specific transcriptomic profiles and that biological phenomena characteristic of each subtype (e.g. proliferation, HER2 and ER signalling) may be driven by specific patterns of copy number aberrations..
Lord, C.
Pollak, J.
(2010). Representation and Accountability: Communicating Tubes?. West european politics,
Vol.33
(5),
pp. 968-988.
Lambros, M.B.
Natrajan, R.
Geyer, F.C.
Lopez-Garcia, M.A.
Dedes, K.J.
Savage, K.
Lacroix-Triki, M.
Jones, R.L.
Lord, C.J.
Linardopoulos, S.
Ashworth, A.
Reis-Filho, J.S.
(2010). PPM1D gene amplification and overexpression in breast cancer: a qRT-PCR and chromogenic in situ hybridization study. Mod pathol,
Vol.23
(10),
pp. 1334-1345.
show abstract
PPM1D (protein phosphatase magnesium-dependent 1δ) maps to the 17q23.2 amplicon and is amplified in ∼8% of breast cancers. The PPM1D gene encodes a serine threonine phosphatase, which is involved in the regulation of several tumour suppressor pathways, including the p53 pathway. Along with others, we have recently shown that PPM1D is one of the drivers of the 17q23.2 amplicon and a promising therapeutic target. Here we investigate whether PPM1D is overexpressed when amplified in breast cancers and the correlations between PPM1D overexpression and amplification with clinicopathological features and survival of breast cancer patients from a cohort of 245 patients with invasive breast cancer treated with therapeutic surgery followed by adjuvant anthracycline-based chemotherapy. mRNA was extracted from representative sections of tumours containing >50% of tumour cells and subjected to TaqMan quantitative real-time PCR using primers for PPM1D and for two housekeeping genes. PPM1D overexpression was defined as the top quartile of expression levels. Chromogenic in situ hybridization with in-house-generated probes for PPM1D was performed. Amplification was defined as >50% of cancer cells with >5 signals per nucleus/large gene clusters. PPM1D overexpression and amplification were found in 25 and 6% of breast cancers, respectively. All cases harbouring PPM1D amplification displayed PPM1D overexpression. PPM1D overexpression was inversely correlated with expression of TOP2A, EGFR and cytokeratins 5/6 and 17. PPM1D amplification was significantly associated with HER2 overexpression, and HER2, TOP2A and CCND1 amplification. No association between PPM1D gene amplification and PPM1D mRNA overexpression with survival was observed. In conclusion, PPM1D is consistently overexpressed when amplified; however, PPM1D overexpression is more pervasive than gene amplification. PPM1D overexpression and amplification are associated with tumours displaying luminal or HER2 phenotypes. Co-amplification of PPM1D and HER2/TOP2A and CCND1 are not random events and may suggest the presence of a 'firestorm' genetic profile..
Lord, C.J.
Ashworth, A.
(2010). Biology-driven cancer drug development: back to the future. Bmc biology,
Vol.8,
p. 12.
Rehman, F.L.
Lord, C.J.
Ashworth, A.
(2010). Synthetic lethal approaches to breast cancer therapy. Nat rev clin oncol,
Vol.7
(12),
pp. 718-724.
show abstract
The promise of personalized therapy for breast cancer is that therapeutic efficacy will be increased while toxic effects are reduced to a minimum. To achieve this goal, there is now an emphasis on the design of therapies that are based not only on the clinical manifestations of the disease, but also on the underlying molecular and cellular biology of cancer. However, identifying targets for personalized therapies in breast cancer is challenging. Here, we describe how biological concepts such as synthetic lethality and oncogene addiction can be used to identify new therapeutic targets and approaches. We discuss the current clinical developments in implementing synthetic lethality therapies, and highlight new ways in which this approach could be used to target specific subsets of breast cancer..
de Plater, L.
Lauge, A.
Guyader, C.
Poupon, M.-.
Assayag, F.
de Cremoux, P.
Vincent-Salomon, A.
Stoppa-Lyonnet, D.
Sigal-Zafrani, B.
Fontaine, J.-.
Brough, R.
Lord, C.J.
Ashworth, A.
Cottu, P.
Decaudin, D.
Marangoni, E.
(2010). Establishment and characterisation of a new breast cancer xenograft obtained from a woman carrying a germline BRCA2 mutation. British journal of cancer,
Vol.103
(8),
pp. 1192-9.
Dedes, K.J.
Wetterskog, D.
Mendes-Pereira, A.M.
Natrajan, R.
Lambros, M.B.
Geyer, F.C.
Vatcheva, R.
Savage, K.
Mackay, A.
Lord, C.J.
Ashworth, A.
Reis-Filho, J.S.
(2010). PTEN deficiency in endometrioid endometrial adenocarcinomas predicts sensitivity to PARP inhibitors. Sci transl med,
Vol.2
(53),
p. 53ra75.
show abstract
PTEN (phosphatase and tensin homolog) loss of function is the most common genetic aberration in endometrioid endometrial carcinomas. In addition to its well-described role in cell signaling, PTEN is involved in the maintenance of genomic stability. Loss of PTEN function causes defects in repair of DNA double-strand breaks by homologous recombination and, therefore, sensitizes cells to inhibition of the poly(adenosine diphosphate ribose) polymerase (PARP). Here, we determined the PTEN status of eight endometrioid endometrial carcinoma cell lines and correlated it with in vitro sensitivity to the PARP inhibitor KU0058948. PTEN-deficient cells showed a significantly greater sensitivity to KU0058948 than the two endometrioid endometrial carcinoma cell lines with wild-type PTEN. The cell lines lacking PTEN expression were unable to elicit a homologous recombination damage response as assayed by RAD51 focus function (a marker of competent homologous recombination DNA repair) upon irradiation and treatment with PARP inhibitors. PTEN silencing in PTEN wild-type Hec-1b cells resulted in reduced RAD51 foci formation after DNA damage and increased sensitivity to PARP inhibition. PTEN reexpression in PTEN-null cell lines resulted in enhanced RAD51 foci formation and in relative resistance to KU0058948. Given that up to 80% of endometrioid endometrial cancers lack PTEN expression, our results suggest that PARP inhibitors may be therapeutically useful for a subset of endometrioid endometrial cancers..
Graeser, M.
McCarthy, A.
Lord, C.J.
Savage, K.
Hills, M.
Salter, J.
Orr, N.
Parton, M.
Smith, I.E.
Reis-Filho, J.S.
Dowsett, M.
Ashworth, A.
Turner, N.C.
(2010). A marker of homologous recombination predicts pathologic complete response to neoadjuvant chemotherapy in primary breast cancer. Clin cancer res,
Vol.16
(24),
pp. 6159-6168.
show abstract
PURPOSE: To assess the prevalence of defective homologous recombination (HR)-based DNA repair in sporadic primary breast cancers, examine the clincopathologic features that correlate with defective HR and the relationship with neoadjuvant chemotherapy response. EXPERIMENTAL DESIGN: We examined a cohort of 68 patients with sporadic primary breast cancer who received neoadjuvant anthracylcine-based chemotherapy, with core biopsies taken 24 hours after the first cycle of chemotherapy. We assessed RAD51 focus formation, a marker of HR competence, by immunofluorescence in postchemotherapy biopsies along with geminin as a marker of proliferative cells. We assessed the RAD51 score as the proportion of proliferative cells with RAD51 foci. RESULTS: A low RAD51 score was present in 26% of cases (15/57, 95% CI: 15%-40%). Low RAD51 score correlated with high histologic grade (P = 0.031) and high baseline Ki67 (P = 0.005). Low RAD51 score was more frequent in triple-negative breast cancers than in ER- and/or HER2-positive breast cancer (67% vs. 19% respectively; P = 0.0036). Low RAD51 score was strongly predictive of pathologic complete response (pathCR) to chemotherapy, with 33% low RAD51 score cancers achieving pathCR compared with 3% of other cancers (P = 0.011). CONCLUSIONS: Our results suggest that defective HR, as indicated by low RAD51 score, may be one of the factors that underlie sensitivity to anthracycline-based chemotherapy. Defective HR is frequent in triple-negative breast cancer, but it is also present in a subset of other subtypes, identifying breast cancers that may benefit from therapies that target defective HR such as PARP inhibitors..
Martin, S.A.
McCabe, N.
Mullarkey, M.
Cummins, R.
Burgess, D.J.
Nakabeppu, Y.
Oka, S.
Kay, E.
Lord, C.J.
Ashworth, A.
(2010). DNA polymerases as potential therapeutic targets for cancers deficient in the DNA mismatch repair proteins MSH2 or MLH1. Cancer cell,
Vol.17
(3),
pp. 235-248.
show abstract
Synthetic sickness/lethality (SSL) can be exploited to develop therapeutic strategies for cancer. Deficiencies in the tumor suppressor proteins MLH1 and MSH2 have been implicated in cancer. Here we demonstrate that deficiency in MSH2 is SSL with inhibition of the DNA polymerase POLB, whereas deficiency in MLH1 is SSL with DNA polymerase POLG inhibition. Both SSLs led to the accumulation of 8-oxoG oxidative DNA lesions. MSH2/POLB SSL caused nuclear 8-oxoG accumulation, whereas MLH1/POLG SSL led to a rise in mitochondrial 8-oxoG levels. Both SSLs were rescued by silencing the adenine glycosylase MUTYH, suggesting that lethality could be caused by the formation of lethal DNA breaks upon 8-oxoG accumulation. These data suggest targeted, mechanism-based therapeutic approaches..
Dedes, K.J.
Wetterskog, D.
Mendes-Pereira, A.M.
Vatcheva, R.
Natrajan, R.
Lambros, M.B.
Lord, C.J.
Ashworth, A.
Reis-Filho, J.S.
(2010). Preclinical evaluation of PARP inhibition as a treatment for endometrioid endometrial carcinomas. Journal of clinical oncology,
Vol.28
(15).
Hewish, M.
Saffery, C.
Barbachano, Y.
Wotherspoon, A.
Brown, G.
Martin, S.A.
Lord, C.J.
Chau, I.
Ashworth, A.
Cunningham, D.
(2010). MESH: Phase II trial of methotrexate as a synthetic lethal therapy for metastatic MSH2-deficient colorectal and other tumors. Journal of clinical oncology,
Vol.28
(15).
Iorns, E.
Lord, C.J.
Ashworth, A.
(2009). Parallel RNAi and compound screens identify the PDK1 pathway as a target for tamoxifen sensitization. Biochemical journal,
Vol.417,
pp. 361-10.
McCabe, N.
Cerone, M.A.
Ohishi, T.
Seimiya, H.
Lord, C.J.
Ashworth, A.
(2009). Targeting Tankyrase 1 as a therapeutic strategy for BRCA-associated cancer. Oncogene,
Vol.28
(11),
pp. 1465-6.
Lord, C.J.
Martin, S.A.
Ashworth, A.
(2009). RNA interference screening demystified. Journal of clinical pathology,
Vol.62
(3),
pp. 195-6.
Tan, D.S.
Lambros, M.B.
Rayter, S.
Natrajan, R.
Vatcheva, R.
Gao, Q.
Marchio, C.
Geyer, F.C.
Savage, K.
Parry, S.
Fenwick, K.
Tamber, N.
Mackay, A.
Dexter, T.
Jameson, C.
McCluggage, W.G.
Williams, A.
Graham, A.
Faratian, D.
El-Bahrawy, M.
Paige, A.J.
Gabra, H.
Gore, M.E.
Zvelebil, M.
Lord, C.J.
Kaye, S.B.
Ashworth, A.
Reis-Filho, J.S.
(2009). PPM1D Is a Potential Therapeutic Target in Ovarian Clear Cell Carcinomas. Clinical cancer research,
Vol.15
(7),
pp. 2269-12.
Houlihan, S.J.
Simpson, S.H.
Cave, A.J.
Flook, N.W.
Hurlburt, M.E.
Lord, C.J.
Smith, L.L.
Sternberg, H.H.
(2009). Hypertension treatment and control rates: chart review in an academic family medicine clinic. Can fam physician,
Vol.55
(7),
pp. 735-741.
show abstract
OBJECTIVE: To characterize hypertension management in an academic family medicine clinic. DESIGN: Cross-sectional chart review. SETTING: Academic family medicine clinic in Edmonton, Alta. PARTICIPANTS: A total of 210 patients with 1 or more visits for hypertension during the previous 3 years. MAIN OUTCOME MEASURES: Patient characteristics, current antihypertensive therapies, most recent blood pressure measurements, and compelling indications according to the 2006 Canadian Hypertension Education Program recommendations. RESULTS: A total of 185 subjects (88%) were prescribed antihypertensive medications, and 89 (42%) had controlled hypertension. Younger subjects, people with diabetes, and people not receiving antihypertensive medication therapy appeared less likely to have controlled hypertension. There were 76 subjects (36%) prescribed 1 antihypertensive medication, 65 subjects (31%) prescribed 2 antihypertensive medications, and 44 (21%) prescribed 3 or more antihypertensive medications. Angiotensin-converting enzyme inhibitors were prescribed for 51% of the subjects, diuretics for 47%, beta-blockers for 27%, calcium channel blockers for 23%, angiotensin receptor blockers for 20%, and alpha-blockers for 1%. CONCLUSION: Hypertension treatment and control rates in this academic family medicine clinic appear to be better than those in the general population. Following the principles of a continuous quality improvement process, this information will serve as an important foundation for identifying areas to improve hypertension management in the clinic..
Iorns, E.
Lord, C.J.
Grigoriadis, A.
McDonald, S.
Fenwick, K.
MacKay, A.
Mein, C.A.
Natrajan, R.
Savage, K.
Tamber, N.
Reis-Filho, J.S.
Turner, N.C.
Ashworth, A.
(2009). Integrated Functional, Gene Expression and Genomic Analysis for the Identification of Cancer Targets. Plos one,
Vol.4
(4),
p. 11.
Oliver, A.W.
Swift, S.
Lord, C.J.
Ashworth, A.
Pearl, L.H.
(2009). Structural basis for recruitment of BRCA2 by PALB2. Embo reports,
Vol.10
(9),
pp. 990-7.
Iorns, E.
Martens-de Kemp, S.R.
Lord, C.J.
Ashworth, A.
(2009). CRK7 modifies the MAPK pathway and influences the response to endocrine therapy. Carcinogenesis,
Vol.30
(10),
pp. 1696-6.
McCarthy, A.
Lord, C.J.
Savage, K.
Grigoriadis, A.
Smith, D.P.
Weigelt, B.
Reis-Filho, J.S.
Ashworth, A.
(2009). Conditional deletion of the Lkb1 gene in the mouse mammary gland induces tumour formation. J pathol,
Vol.219
(3),
pp. 306-316.
show abstract
Heterozygous germline mutations in the LKB1 (STK11) gene cause Peutz-Jeghers syndrome (PJS), an autosomal dominant disorder characterized by hamartomatous polyposis of the gastrointestinal tract and an increased risk of colorectal, breast, and other cancers. To model the role of LKB1 mutation in mammary tumourigenesis, we have used a conditional gene targeting strategy to generate a mouse in which exons encoding the kinase domain of Lkb1 were deleted specifically in the mammary gland. Mammary gland tumours developed in these mice with a latency of 46-85 weeks and occurred in the thoracic or inguinal glands. These tumours were grade 2 invasive ductal carcinomas or solid papillary carcinomas with histological features similar to those described in breast cancers arising in patients with PJS. This mouse model of Lkb1 deficiency provides a potentially useful tool to investigate the role of Lkb1 in tumourigenesis and to guide the development of therapeutic approaches..
Mackay, A.
Tamber, N.
Fenwick, K.
Iravani, M.
Grigoriadis, A.
Dexter, T.
Lord, C.J.
Reis-Filho, J.S.
Ashworth, A.
(2009). A high-resolution integrated analysis of genetic and expression profiles of breast cancer cell lines. Breast cancer res treat,
Vol.118
(3),
pp. 481-498.
show abstract
Tumour cell lines derived from breast cancer patients constitute one of the cornerstones of breast cancer research. To characterise breast cancer cell lines at the genetic level, we have developed a full tiling path bacterial artificial chromosome (BAC) array collection for comparative genomic hybridisation (aCGH). This aCGH BAC collection covers 98% of the entire human genome at a resolution of 40-60 kbp. We have used this platform alongside an in-house produced 17 K cDNA microarray set to characterise the genetic and transcriptomic profiles of 24 breast cancer cell lines, as well as cell types derived from non-diseased breast. We demonstrate that breast cancer cell lines have genomic and transcriptomic features that recapitulate those of primary breast cancers and can be reliably subclassified into basal-like and luminal subgroups. By overlaying aCGH and transcriptomic data, we have identified 753 genes whose expression correlate with copy number; this list comprised numerous oncogenes recurrently amplified and overexpressed in breast cancer (e.g., HER2, MYC, CCND1 and AURKA). Finally, we demonstrate that although breast cancer cell lines have genomic features usually found in grade III breast cancers (i.e., gains of 1q, 8q and 20q), basal-like and luminal cell lines are characterised by distinct genomic aberrations..
Lord, C.J.
Ashworth, A.
(2009). Bringing DNA Repair in Tumors into Focus. Clinical cancer research,
Vol.15
(10),
pp. 3241-3.
Natrajan, R.
Lambros, M.B.
Rodríguez-Pinilla, S.M.
Moreno-Bueno, G.
Tan, D.S.
Marchió, C.
Vatcheva, R.
Rayter, S.
Mahler-Araujo, B.
Fulford, L.G.
Hungermann, D.
Mackay, A.
Grigoriadis, A.
Fenwick, K.
Tamber, N.
Hardisson, D.
Tutt, A.
Palacios, J.
Lord, C.J.
Buerger, H.
Ashworth, A.
Reis-Filho, J.S.
(2009). Tiling path genomic profiling of grade 3 invasive ductal breast cancers. Clin cancer res,
Vol.15
(8),
pp. 2711-2722.
show abstract
PURPOSE: To characterize the molecular genetic profiles of grade 3 invasive ductal carcinomas of no special type using high-resolution microarray-based comparative genomic hybridization (aCGH) and to identify recurrent amplicons harboring putative therapeutic targets associated with luminal, HER-2, and basal-like tumor phenotypes. EXPERIMENTAL DESIGN: Ninety-five grade 3 invasive ductal carcinomas of no special type were classified into luminal, HER-2, and basal-like subgroups using a previously validated immunohistochemical panel. Tumor samples were microdissected and subjected to aCGH using a tiling path 32K BAC array platform. Selected regions of recurrent amplification were validated by means of in situ hybridization. Expression of genes pertaining to selected amplicons was investigated using quantitative real-time PCR and gene silencing was done using previously validated short hairpin RNA constructs. RESULTS: We show that basal-like and HER-2 tumors are characterized by "sawtooth" and "firestorm" genetic patterns, respectively, whereas luminal cancers were more heterogeneous. Apart from confirming known amplifications associated with basal-like (1q21, 10p, and 12p), luminal (8p12, 11q13, and 11q14), and HER-2 (17q12) cancers, we identified previously unreported recurrent amplifications associated with each molecular subgroup: 19q12 in basal-like, 1q32.1 in luminal, and 14q12 in HER-2 cancers. PPM1D gene amplification (17q23.2) was found in 20% and 8% of HER-2 and luminal cancers, respectively. Silencing of PPM1D by short hairpin RNA resulted in selective loss of viability in tumor cell lines harboring the 17q23.2 amplification. CONCLUSIONS: Our results show the power of aCGH analysis in unraveling the genetic profiles of specific subgroups of cancer and for the identification of novel therapeutic targets..
Brough, R.
Wei, D.
Leulier, S.
Lord, C.J.
Rong, Y.S.
Ashworth, A.
(2008). Functional analysis of Drosophila melanogaster BRCA2 in DNA repair. Dna repair (amst),
Vol.7
(1),
pp. 10-19.
show abstract
The human BRCA2 cancer susceptibility protein functions in double-strand DNA break repair by homologous recombination and this pathway is conserved in the fly Drosophila melanogaster. Although a potential Drosophila melanogaster BRCA2 orthologue (dmbrca2; CG30169) has been identified by sequence similarity, no functional data addressing the role of this protein in DNA repair is available. Here, we demonstrate that depletion of dmbrca2 from Drosophila cells induces sensitivity to DNA damage induced by irradiation or treatment with hydroxyurea. Dmbrca2 physically interacts with dmrad51 (spnA) and the two proteins become recruited to nuclear foci after DNA damage. A functional assay for DNA repair demonstrated that in flies dmbrca2 plays a role in double-strand break repair by gene conversion. Finally, we show that depletion of dmbrca2 in cells is synthetically lethal with deficiency in other DNA repair proteins including dmparp. The conservation of the function of BRCA2 in Drosophila will allow the analysis of this key DNA repair protein in a genetically tractable organism potentially illuminating mechanisms of carcinogenesis and aiding the development of therapeutic agents..
Lord, C.J.
McDonald, S.
Swift, S.
Turner, N.C.
Ashworth, A.
(2008). A high-throughput RNA interference screen for DNA repair determinants of PARP inhibitor sensitivity. Dna repair (amst),
Vol.7
(12),
pp. 2010-2019.
show abstract
Synthetic lethality is an attractive strategy for the design of novel therapies for cancer. Using this approach we have previously demonstrated that inhibition of the DNA repair protein, PARP1, is synthetically lethal with deficiency of either of the breast cancer susceptibility proteins, BRCA1 and BRCA2. This observation is most likely explained by the inability of BRCA deficient cells to repair DNA damage by homologous recombination (HR) and has led to the clinical trials of potent PARP inhibitors for the treatment of BRCA mutation-associated cancer. To identify further determinants of PARP inhibitor response, we took a high-throughput genetic approach. We tested each of the genes recognised as having a role in DNA repair using short-interfering RNA (siRNA) and assessed the sensitivity of siRNA transfected cells to a potent PARP inhibitor, KU0058948. The validity of this approach was confirmed by the identification of known genetic determinants of PARP inhibitor sensitivity, including genes involved in HR. Novel determinants of PARP inhibitor response were also identified, including the transcription coupled DNA repair (TCR) proteins DDB1 and XAB2. These results suggest that DNA repair pathways other than HR may determine sensitivity to PARP inhibitors and highlight the likelihood that ostensibly distinct DNA repair pathways cooperate to maintain genomic stability and cellular viability. Furthermore, the identification of these novel determinants may eventually guide the optimal use of PARP inhibitors in the clinic..
Lord, C.J.
Iorns, E.
Ashworth, A.
(2008). Dissecting resistance to endocrine therapy in breast cancer. Cell cycle,
Vol.7
(13),
pp. 1895-4.
Iorns, E.
Turner, N.C.
Elliott, R.
Syed, N.
Garrone, O.
Gasco, M.
Tutt, A.N.
Crook, T.
Lord, C.J.
Ashworth, A.
(2008). Identification of CDK10 as an important determinant of resistance to endocrine therapy for breast cancer. Cancer cell,
Vol.13
(2),
pp. 91-104.
show abstract
Therapies that target estrogen signaling have transformed the treatment of breast cancer. However, the effectiveness of these agents is limited by the development of resistance. Here, an RNAi screen was used to identify modifiers of tamoxifen sensitivity. We demonstrate that CDK10 is an important determinant of resistance to endocrine therapies and show that CDK10 silencing increases ETS2-driven transcription of c-RAF, resulting in MAPK pathway activation and loss of tumor cell reliance upon estrogen signaling. Patients with ER alpha-positive tumors that express low levels of CDK10 relapse early on tamoxifen, demonstrating the clinical significance of these observations. The association of low levels of CDK10 with methylation of the CDK10 promoter suggests a mechanism by which CDK10 expression is reduced in tumors..
Martin, S.A.
Lord, C.J.
Ashworth, A.
(2008). DNA repair deficiency as a therapeutic target in cancer. Curr opin genet dev,
Vol.18
(1),
pp. 80-86.
show abstract
Inhibitors of DNA repair proteins have been used in cancer therapy, mostly to potentiate the effects of cytotoxic agents. However, tumor cells frequently exhibit deficiencies in the signalling or repair of DNA damage. These deficiencies probably contribute to pathogenesis of the disease, but they also present an opportunity to target the tumor. Recently, inhibitors of poly(ADP-ribose) polymerase (PARP) have been shown to be highly selective for tumor cells with defects in the repair of double-strand DNA breaks (DSBs) by homologous recombination, particularly in the context of BRCA1 or BRCA2 mutation. It seems likely that other DNA repair processes can be targeted in a similar manner. These synthetic lethal approaches highlight how an understanding of DNA repair processes can be used in the development of novel cancer treatments..
Turner, N.C.
Lord, C.J.
Iorns, E.
Brough, R.
Swift, S.
Elliott, R.
Rayter, S.
Tutt, A.N.
Ashworth, A.
(2008). A synthetic lethal siRNA screen identifying genes mediating sensitivity to a PARP inhibitor. Embo journal,
Vol.27
(9),
pp. 1368-10.
Lord, C.J.
Ashworth, A.
(2008). Targeted therapy for cancer using PARP inhibitors. Current opinion in pharmacology,
Vol.8
(4),
pp. 363-7.
Marchiò, C.
Natrajan, R.
Shiu, K.K.
Lambros, M.B.
Rodriguez-Pinilla, S.M.
Tan, D.S.
Lord, C.J.
Hungermann, D.
Fenwick, K.
Tamber, N.
Mackay, A.
Palacios, J.
Sapino, A.
Buerger, H.
Ashworth, A.
Reis-Filho, J.S.
(2008). The genomic profile of HER2-amplified breast cancers: the influence of ER status. J pathol,
Vol.216
(4),
pp. 399-407.
show abstract
Expression profiling studies have suggested that HER2-amplified breast cancers constitute a heterogeneous group that may be subdivided according to their ER status: HER2-amplified ER-positive breast carcinomas that fall into the luminal B cluster; and HER2-amplified ER-negative cancers which form a distinct molecular subgroup, known as the erbB2 or HER2 subgroup. ER-negative breast cancer differs significantly from ER-positive disease in the pattern, type, and complexity of genetic aberrations. Here we have compared the genomic profiles of ER-positive and ER-negative HER2-amplified cancers using tiling path microarray-based comparative genomic hybridization (aCGH). Validation of the differentially amplified regions was performed in an independent series of 70 HER2-amplified breast cancers. Although HER2-amplified cancers had remarkably complex patterns of molecular genetic aberrations, ER-positive and ER-negative HER2-amplified breast carcinomas shared most molecular genetic features as defined by aCGH. Genome-wide Fisher's exact test analysis revealed that less than 1.5% of the genome was significantly differentially gained or lost in ER-positive versus ER-negative HER2-amplified cancers. However, two regions of amplification were significantly associated with ER-positive carcinomas, one of which mapped to 17q21.2 and encompassed GJC1, IGFBP4, TNS4, and TOP2A. Chromogenic in situ hybridization analysis of an independent validation series confirmed the association between ER status and TOP2A amplification. In conclusion, although hormone receptor status does not determine the overall genetic profile of HER2-amplified breast cancers, specific genetic aberrations may be characteristic of subgroups of HER2 breast cancers..
Rayter, S.
Elliott, R.
Travers, J.
Rowlands, M.G.
Richardson, T.B.
Boxall, K.
Jones, K.
Linardopoulos, S.
Workman, P.
Aherne, W.
Lord, C.J.
Ashworth, A.
(2008). A chemical inhibitor of PPM1D that selectively kills cells overexpressing PPM1D. Oncogene,
Vol.27
(8),
pp. 1036-1044.
show abstract
The PPM1D gene is aberrantly amplified in a range of common cancers and encodes a protein phosphatase that is a potential therapeutic target. However, the issue of whether inhibition of PPM1D in human tumour cells that overexpress this protein compromises their viability has not yet been fully addressed. We show here, using an RNA interference (RNAi) approach, that inhibition of PPM1D can indeed reduce the viability of human tumour cells and that this effect is selective; tumour cell lines that overexpress PPM1D are sensitive to PPM1D inhibition whereas cell lines with normal levels are not. Loss of viability associated with PPM1D RNAi in human tumour cells occurs via the activation of the kinase P38. To identify chemical inhibitors of PPM1D, a high-throughput screening of a library of small molecules was performed. This strategy successfully identified a compound that selectively reduces viability of human tumour cell lines that overexpress PPM1D. As expected of a specific inhibitor, the toxicity to PPM1D overexpressing cell lines after inhibitor treatment is P38 dependent. These results further validate PPM1D as a therapeutic target and identify a proof-of-principle small molecule inhibitor..
Edwards, S.L.
Brough, R.
Lord, C.J.
Natrajan, R.
Vatcheva, R.
Levine, D.A.
Boyd, J.
Reis-Filho, J.S.
Ashworth, A.
(2008). Resistance to therapy caused by intragenic deletion in BRCA2. Nature,
Vol.451
(7182),
pp. 1111-1115.
show abstract
Cells with loss of BRCA2 function are defective in homologous recombination (HR) and are highly sensitive to inhibitors of poly(ADP-ribose) polymerase (PARP), which provides the basis for a new therapeutic approach. Here we show that resistance to PARP inhibition can be acquired by deletion of a mutation in BRCA2. We derived PARP-inhibitor-resistant (PIR) clones from the human CAPAN1 pancreatic cancer cell line, which carries the protein-truncating c.6174delT frameshift mutation. PIR clones could form DNA-damage-induced RAD51 nuclear foci and were able to limit genotoxin-induced genomic instability, both hallmarks of a competent HR pathway. New BRCA2 isoforms were expressed in the resistant lines as a result of intragenic deletion of the c.6174delT mutation and restoration of the open reading frame (ORF). Reconstitution of BRCA2-deficient cells with these revertant BRCA2 alleles rescued PARP inhibitor sensitivity and HR deficiency. Most of the deletions in BRCA2 were associated with small tracts of homology, and possibly arose from error-prone repair caused by BRCA2 deficiency. Similar ORF-restoring mutations were present in carboplatin-resistant ovarian tumours from c.6174delT mutation carriers. These observations have implications for understanding drug resistance in BRCA mutation carriers as well as in defining functionally important domains within BRCA2..
Komander, D.
Lord, C.J.
Scheel, H.
Swift, S.
Hofmann, K.
Ashworth, A.
Barford, D.
(2008). The structure of the CYLD USP domain explains its specificity for Lys63-linked polyubiquitin and reveals a B box module. Mol cell,
Vol.29
(4),
pp. 451-464.
show abstract
The tumor suppressor CYLD antagonizes NF-kappaB and JNK signaling by disassembly of Lys63-linked ubiquitin chains synthesized in response to cytokine stimulation. Here we describe the crystal structure of the CYLD USP domain, revealing a distinctive architecture that provides molecular insights into its specificity toward Lys63-linked polyubiquitin. We identify regions of the USP domain responsible for this specificity and demonstrate endodeubiquitinase activity toward such chains. Pathogenic truncations of the CYLD C terminus, associated with the hypertrophic skin tumor cylindromatosis, disrupt the USP domain, accounting for loss of CYLD catalytic activity. A small zinc-binding B box domain, similar in structure to other crossbrace Zn-binding folds--including the RING domain found in E3 ubiquitin ligases--is inserted within the globular core of the USP domain. Biochemical and functional characterization of the B box suggests a role as a protein-interaction module that contributes to determining the subcellular localization of CYLD..
Lord, C.J.
Ashworth, A.
(2007). RAD51, BRCA2 and DNA repair: a partial resolution. Nature structural & molecular biology,
Vol.14
(6),
pp. 461-2.
Iorns, E.
Lord, C.J.
Turner, N.
Ashworth, A.
(2007). Utilizing RNA interference to enhance cancer drug discovery. Nat rev drug discov,
Vol.6
(7),
pp. 556-568.
show abstract
With the development of RNA interference (RNAi) libraries, systematic and cost-effective genome-wide loss-of-function screens can now be carried out with the aim of assessing the role of specific genes in neoplastic phenotypes, and the rapid identification of novel drug targets. Here, we discuss the existing applications of RNAi in cancer drug discovery and highlight areas in this process that may benefit from this technology in the future..
Gudmundsdottir, K.
Lord, C.J.
Ashworth, A.
(2007). The proteasome is involved in determining differential utilization of double-strand break repair pathways. Oncogene,
Vol.26
(54),
pp. 7601-6.
Smalley, M.J.
Iravani, M.
Leao, M.
Grigoriadis, A.
Kendrick, H.
Dexter, T.
Fenwick, K.
Regan, J.L.
Britt, K.
McDonald, S.
Lord, C.J.
Mackay, A.
Ashworth, A.
(2007). Regulator of G-protein signalling 2 mRNA is differentially expressed in mammary epithelial subpopulations and over-expressed in the majority of breast cancers. Breast cancer res,
Vol.9
(6),
p. R85.
show abstract
INTRODUCTION: To understand which signalling pathways become deregulated in breast cancer, it is necessary to identify functionally significant gene expression patterns in the stem, progenitor, transit amplifying and differentiated cells of the mammary epithelium. We have previously used the markers 33A10, CD24 and Sca-1 to identify mouse mammary epithelial cell subpopulations. We now investigate the relationship between cells expressing these markers and use gene expression microarray analysis to identify genes differentially expressed in the cell populations. METHODS: Freshly isolated primary mouse mammary epithelial cells were separated on the basis of staining with the 33A10 antibody and an alpha-Sca-1 antibody. The populations identified were profiled using gene expression microarray analysis. Gene expression patterns were confirmed on normal mouse and human mammary epithelial subpopulations and were examined in a panel of breast cancer samples and cell lines. RESULTS: Analysis of the separated populations demonstrated that Sca-1- 33A10High stained cells were estrogen receptor alpha (Esr1)- luminal epithelial cells, whereas Sca-1+ 33A10Low/- stained cells were a mix of nonepithelial cells and Esr1+ epithelial cells. Analysis of the gene expression data identified the gene Rgs2 (regulator of G-protein signalling 2) as being highly expressed in the Sca-1- 33A10Low/- population, which included myoepithelial/basal cells. RGS2 has previously been described as a regulator of angiotensin II receptor signalling. Gene expression analysis by quantitative real-time RT-PCR of cells separated on the basis of CD24 and Sca-1 expression confirmed that Rgs2 was more highly expressed in mouse myoepithelial/basal mammary cells than luminal cells. This expression pattern was conserved in normal human breast cells. Functional analysis demonstrated RGS2 to be a modulator of oxytocin receptor signalling. The potential significance of RGS2 expression in breast cancer was demonstrated by semi-quantitative RT-PCR analysis, data mining and quantitative real-time RT-PCR approaches, which showed that RGS2 was expressed in the majority of solid breast cancers at much higher levels than in normal human mammary cells. CONCLUSION: Molecular analysis of prospectively isolated mammary epithelial cells identified RGS2 as a modulator of oxytocin receptor signalling, which is highly expressed in the myoepithelial cells. The RGS2 gene, but not the oxytocin receptor, was also shown to be over-expressed in the majority of breast cancers, identifying the product of this gene, or the pathway(s) it regulates, as potentially significant therapeutic targets..
Friedmann, B.J.
Caplin, M.
Savic, B.
Shah, T.
Lord, C.J.
Ashworth, A.
Hartley, J.A.
Hochhauser, D.
(2006). Interaction of the epidermal growth factor receptor and the DNA-dependent protein kinase pathway following gefitinib treatment. Molecular cancer therapeutics,
Vol.5
(2),
pp. 209-10.
Lord, C.J.
Garrett, M.D.
Ashworth, A.
(2006). Targeting the double-strand DNA break repair pathway as a therapeutic strategy. Clinical cancer research,
Vol.12
(15),
pp. 4463-6.
Arias-Lopez, C.
Lazaro-Trueba, I.
Kerr, P.
Lord, C.J.
Dexter, T.
Iravani, M.
Ashworth, A.
Silva, A.
(2006). P53 modulates homologous recombination by transcriptional regulation of the RAD51 gene. Embo reports,
Vol.7
(2),
pp. 219-6.
full text
Clarke, J.C.
Honey, E.M.
Bekker, E.
Snyman, L.C.
Raymond, R.M.
Lord, C.
Brophy, P.D.
(2006). A novel nonsense mutation in the EYA1 gene associated with branchio-oto-renal/branchiootic syndrome in an Afrikaner kindred. Clin genet,
Vol.70
(1),
pp. 63-67.
show abstract
Branchio-oto-renal (BOR) syndrome is an autosomal dominant disorder characterized by the associations of hearing loss, branchial arch defects and renal anomalies. Branchiootic (BO) syndrome is a related disorder that presents without the highly variable characteristic renal anomalies of BOR syndrome. Dominant mutations in the human homologue of the Drosophila eyes absent gene (EYA1) are frequently the cause of both BOR and BO syndromes. We report a South African family of Afrikaner descent with affected individuals presenting with pre-auricular abnormalities and either hearing loss or bilateral absence of the kidneys. Genetic analysis of the pedigree detected a novel EYA1 heterozygous nonsense mutation in affected family members but not in unaffected family members or a random DNA panel. Through mutational analysis, we conclude that this particular mutation is the cause of BOR/BO syndrome in this family as a result of a truncation of the EYA1 protein that ablates the critical EYA homologous region. To the best of our knowledge, this is the first case of BOR/BO syndrome reported in Africa or in those of the Afrikaner descent..
McCabe, N.
Turner, N.C.
Lord, C.J.
Kluzek, K.
Bialkowska, A.
Swift, S.
Giavara, S.
O'Connor, M.J.
Tutt, A.N.
Zdzienicka, M.Z.
Smith, G.C.
Ashworth, A.
(2006). Deficiency in the repair of DNA damage by homologous recombination and sensitivity to poly(ADP-ribose) polymerase inhibition. Cancer research,
Vol.66
(16),
pp. 8109-7.
McCabe, N.
Lord, C.J.
Tutt, A.N.
Martin, N.M.
Smith, G.C.
Ashworth, A.
(2005). BRCA2-deficient CAPAN-1 cells are extremely sensitive to the inhibition of poly (ADP-Ribose) polymerase. Cancer biology & therapy,
Vol.4
(9),
pp. 934-3.
Farmer, H.
McCabe, N.
Lord, C.J.
Tutt, A.N.
Johnson, D.A.
Richardson, T.B.
Santarosa, M.
Dillon, K.J.
Hickson, I.
Knights, C.
Martin, N.M.
Jackson, S.P.
Smith, G.C.
Ashworth, A.
(2005). Targeting the DNA repair defect in BRCA mutant cells as a therapeutic strategy. Nature,
Vol.434
(7035),
pp. 917-5.
Gudmundsdottir, K.
Lord, C.J.
Witt, E.
Tutt, A.N.
Ashworth, A.
(2004). DSS1 is required for RAD51 focus formation and genomic stability in mammalian cells. Embo rep,
Vol.5
(10),
pp. 989-993.
show abstract
full text
BRCA2 is a breast cancer susceptibility gene implicated in the repair of double-strand breaks by homologous recombination with RAD51. BRCA2 associates with a 70-amino-acid protein, DSS1, but the functional significance of this interaction has remained unclear. Recently, deficiency of a DSS1 orthologue in the fungus Ustilago maydis has been shown to cause a defect in recombinational DNA repair. Here we have investigated the consequences of DSS1 depletion in mammalian cells. We show that like BRCA2, DSS1 is required for DNA damage-induced RAD51 focus formation and for the maintenance of genomic stability, indicating a function conserved from lower eukaryotes to humans. However, DSS1 seems to be not required for BRCA2 or RAD51 stability or for BRCA2 and RAD51 to interact, raising the possibility that DSS1 may be required for the BRCA2-RAD51 complex to become associated with sites of DNA damage..
Brook, F.A.
Evans, E.P.
Lord, C.J.
Lyons, P.A.
Rainbow, D.B.
Howlett, S.K.
Wicker, L.S.
Todd, J.A.
Gardner, R.L.
(2003). The derivation of highly germline-competent embryonic stem cells containing NOD-derived genome. Diabetes,
Vol.52
(1),
pp. 205-208.
show abstract
It would be extremely advantageous to the analysis of disease mechanisms in the spontaneous mouse model of type 1 diabetes, the nonobese diabetic (NOD) strain, if genes in this strain could be modified in vivo using embryonic stem (ES) cells and homologous recombination. However, a NOD ES cell line with adequate germline transmission has not yet been reported. We report the development of highly germline-competent ES cell lines from the F1 hybrid of NOD and 129 for use in NOD gene targeting. Consequently, we developed ES cell lines derived from (NOD x 129)F1 x 129 backcross 1 mice, which were intercrossed to select for homozygosity of particular regions of NOD genome known to contain disease loci..
Penha-Gonçalves, C.
Moule, C.
Smink, L.J.
Howson, J.
Gregory, S.
Rogers, J.
Lyons, P.A.
Suttie, J.J.
Lord, C.J.
Peterson, L.B.
Todd, J.A.
Wicker, L.S.
(2003). Identification of a structurally distinct CD101 molecule encoded in the 950-kb Idd10 region of NOD mice. Diabetes,
Vol.52
(6),
pp. 1551-1556.
show abstract
Genes affecting autoimmune type 1 diabetes susceptibility in the nonobese diabetic (NOD) mouse (Idd loci) have been mapped using a congenic strain breeding strategy. In the present study, we used a combination of BAC clone contig construction, polymorphism analysis of DNA from congenic strains, and sequence mining of the human orthologous region to generate an integrated map of the Idd10 region on mouse chromosome 3. We found seven genes and one pseudogene in the 950-kb Idd10 region. Although all seven genes in the interval are Idd10 candidates, we suggest the gene encoding the EWI immunoglobulin subfamily member EWI-101 (Cd101) as the most likely Idd10 candidate because of the previously reported immune-associated properties of the human CD101 molecule. Additional support for the candidacy of Cd101 is the presence of 17 exonic single-nucleotide polymorphisms that differ between the NOD and B6 sequences, 10 causing amino acid substitutions in the predicted CD101 protein. Four of these 10 substitutions are nonconservative, 2 of which could potentially alter N-linked glycosylation. Considering our results together with those previous reports that antibodies recognizing human CD101 modulate human T-cell and dendritic cell function, there is now justification to test whether the alteration of CD101 function affects autoimmune islet destruction..
Warren, M.
Lord, C.J.
Masabanda, J.
Griffin, D.
Ashworth, A.
(2003). Phenotypic effects of heterozygosity for a BRCA2 mutation. Hum mol genet,
Vol.12
(20),
pp. 2645-2656.
show abstract
Heterozygous carriers of mutations in the BRCA2 gene have a high risk of developing breast and other cancers. In these individuals, BRCA2 appears to act as a tumour suppressor gene, in that loss of the wild type allele is frequently observed within tumours, leading to loss of BRCA2 function. Because BRCA2 functions in DNA repair via homologous recombination, this leads to genomic instability. However, it is unclear whether loss of the wild type allele is stochastic or if heterozygosity for BRCA2 mutation carries a phenotype that contributes to tumorigenic progression. Here we demonstrate that, in a specific vertebrate cell type, the chicken B cell line DT40, heterozygosity for a BRCA2 mutation has a distinct phenotype. This is characterized by a reduced growth rate, increased cell death, heightened sensitivity to specific DNA damaging agents and reduced RAD51 focus formation after irradiation. Thus in certain cell types, genome instability might be driven directly by heterozygosity for BRCA2 mutation..
Lyons, P.A.
Armitage, N.
Lord, C.J.
Phillips, M.S.
Todd, J.A.
Peterson, L.B.
Wicker, L.S.
(2001). Mapping by genetic interaction - High-resolution congenic mapping of the type 1 diabetes loci Idd10 and Idd18 in the NOD mouse. Diabetes,
Vol.50
(11),
pp. 2633-5.
Lord, C.J.
Howlett, S.
Lyons, P.A.
Peterson, L.B.
Wicker, L.S.
Todd, J.A.
(2001). The murine type 1 diabetes loci, Idd1, Idd3, Idd5, Idd9, and Idd17/10/18, do not control thymic CD4-CD8-/TCRalphabeta+ deficiency in the nonobese diabetic mouse. Mamm genome,
Vol.12
(2),
pp. 175-176.
Cordell, H.J.
Todd, J.A.
Hill, N.J.
Lord, C.J.
Lyons, P.A.
Peterson, L.B.
Wicker, L.S.
Clayton, D.G.
(2001). Statistical modeling of interlocus interactions in a complex disease: rejection of the multiplicative model of epistasis in type 1 diabetes. Genetics,
Vol.158
(1),
pp. 357-367.
show abstract
In general, common diseases do not follow a Mendelian inheritance pattern. To identify disease mechanisms and etiology, their genetic dissection may be assisted by evaluation of linkage in mouse models of human disease. Statistical modeling of multiple-locus linkage data from the nonobese diabetic (NOD) mouse model of type 1 diabetes has previously provided evidence for epistasis between alleles of several Idd (insulin-dependent diabetes) loci. The construction of NOD congenic strains containing selected segments of the diabetes-resistant strain genome allows analysis of the joint effects of alleles of different loci in isolation, without the complication of other segregating Idd loci. In this article, we analyze data from congenic strains carrying two chromosome intervals (a double congenic strain) for two pairs of loci: Idd3 and Idd10 and Idd3 and Idd5. The joint action of both pairs is consistent with models of additivity on either the log odds of the penetrance, or the liability scale, rather than with the previously proposed multiplicative model of epistasis. For Idd3 and Idd5 we would also not reject a model of additivity on the penetrance scale, which might indicate a disease model mediated by more than one pathway leading to beta-cell destruction and development of diabetes. However, there has been confusion between different definitions of interaction or epistasis as used in the biological, statistical, epidemiological, and quantitative and human genetics fields. The degree to which statistical analyses can elucidate underlying biologic mechanisms may be limited and may require prior knowledge of the underlying etiology..
Lyons, P.A.
Armitage, N.
Argentina, F.
Denny, P.
Hill, N.J.
Lord, C.J.
Wilusz, M.B.
Peterson, L.B.
Wicker, L.S.
Todd, J.A.
(2000). Congenic mapping of the type 1 diabetes locus, Idd3, to a 780-kb region of mouse chromosome 3: identification of a candidate segment of ancestral DNA by haplotype mapping. Genome res,
Vol.10
(4),
pp. 446-453.
show abstract
Type 1 diabetes in the nonobese diabetic (NOD) mouse arises as a consequence of T cell-mediated destruction of the insulin-producing beta cells of the pancreas. Although little is known of the events that initiate and subsequently drive beta-cell destruction it is clear that the entire process is under complex genetic control. At present 19 loci have been mapped that influence the development of diabetes either at the level of initiation of insulitis or at the level of progression from insulitis to overt diabetes, or both. Previously, we have mapped one of these loci, Idd3, to a 0.35-cM interval on proximal mouse chromosome 3. In the present study we have narrowed the map position of this locus to an interval of 0.15 cM by a combination of novel congenic strains and an ancestral haplotype analysis approach. We have constructed a physical contig in bacterial artificial chromosome (BAC) clones across the minimal interval. Restriction mapping of the BAC contig placed the maximum size of the Idd3 interval at 780 kb between the markers D3Nds36 and D3Nds76. To refine further the Idd3 interval we developed a series of novel single nucleotide polymorphisms (SNPs) and carried out haplotype analysis on DNA from mouse strains known to carry either Idd3 susceptibility or protective alleles. This haplotype analysis identified a 145-kb segment of ancestral DNA between the microsatellite marker D3Nds6 and the SNP 81.3. One haplotype of this ancestral segment of DNA is found in mouse strains carrying an Idd3 susceptibility allele and another is found in mouse strains carrying an Idd3 protective allelle. Within the 780-kb congenically defined interval this 145-kb segment represents the most likely location for Idd3. The Il2 gene, which encodes the cytokine interleukin 2 (IL2), maps to this interval and is a strong candidate for Idd3. To investigate whether sequence variation exists in the promoter region of the Il2 gene, which might alter its expression, we sequenced the promoter region of the Il2 gene from mouse strains carrying either an Idd3 susceptibility or resistance allele. Two sequence variants were identified, neither of which fell in known regulatory elements within the Il2 promoter. In agreement with this observation steady-state Il2 mRNA levels showed no variation between susceptible and resistant mouse strains. These data suggest that the profound protection from diabetes seen in congenic mice carrying an Idd3 protective allele is unlikely to be due to differences in the level of expression of the Il2 gene. Instead, all of the current data support our hypothesis that Idd3 corresponds to amino acid variation at the amino terminus of Il2..
Podolin, P.L.
Wilusz, M.B.
Cubbon, R.M.
Pajvani, U.
Lord, C.J.
Todd, J.A.
Peterson, L.B.
Wicker, L.S.
Lyons, P.A.
(2000). Differential glycosylation of interleukin 2, the molecular basis for the NOD Idd3 type 1 diabetes gene?. Cytokine,
Vol.12
(5),
pp. 477-482.
show abstract
The insulin-dependent diabetes (Idd) gene, Idd3, has been localised to a 0.35 cM region of chromosome 3 containing the structural gene for the cytokine interleukin 2 (IL-2). While variation of the N-terminal amino acid sequence of IL-2 has been shown to correlate with Idd3 allelic variation, differences in induction of proliferation by IL-2 allotypes have not been detected. In the current study, we examined the electrophoretic migration of IL-2 allotypes and have found two distinct patterns, consistent with differences in glycosylation, that correlate with diabetes-resistance and susceptibility. These findings strongly suggest that IL-2 variants may be functionally distinct..
Lyons, P.A.
Hancock, W.W.
Denny, P.
Lord, C.J.
Hill, N.J.
Armitage, N.
Siegmund, T.
Todd, J.A.
Phillips, M.S.
Hess, J.F.
Chen, S.L.
Fischer, P.A.
Peterson, L.B.
Wicker, L.S.
(2000). The NOD Idd9 genetic interval influences the pathogenicity of insulitis and contains molecular variants of Cd30, Tnfr2, and Cd137. Immunity,
Vol.13
(1),
pp. 107-115.
show abstract
Previous analyses of NOD mice have shown that some genes control the development of both insulitis and diabetes, while other loci influence diabetes without reducing insulitis. Evidence for the existence of a gene only influencing diabetes, Idd9 on mouse chromosome 4, is provided here by the development of a novel congenic mouse strain, NOD.B10 Idd9. NOD.B10 Idd9 mice display profound resistance to diabetes even though nearly all develop insulitis. Subcongenic analysis has demonstrated that alleles of at least three B10 genes, Idd9.1, Idd9.2, and Idd9.3 are required to produce Idd9-mediated diabetes resistance. Candidate genes with amino acid differences between the NOD and B10 strains have been localized to the 5.6 cM Idd9.2 interval (Tnfr2, Cd30) and to the 2.0 cM Idd9.3 interval (Cd137)..
Podolin, P.L.
Denny, P.
Armitage, N.
Lord, C.J.
Hill, N.J.
Levy, E.R.
Peterson, L.B.
Todd, J.A.
Wicker, L.S.
Lyons, P.A.
(1998). Localization of two insulin-dependent diabetes (Idd) genes to the Idd10 region on mouse chromosome 3. Mamm genome,
Vol.9
(4),
pp. 283-286.
show abstract
Multiple genes control the development of autoimmune diabetes both in humans and in the nonobese diabetic (NOD) strain of mouse. Previously, three insulin-dependent diabetes (Idd) genes, Idd3, Idd10, and Idd17, were localized to mouse Chromosome (Chr) 3. The B10- or B6-derived resistance alleles at Idd10 and Idd3 together provide the NOD mouse with nearly complete protection from diabetes. In the present study, the 10.2-cM region encoding Idd10 was defined further with newly developed congenic strains. A locus, located in the centromeric 2.1 cM of the 10.2 cM region, contributed to the Idd10 trait. However, this locus did not account for the full effect of Idd10, suggesting the presence of a second gene in the distal portion of the 10.2-cM region. This second gene is designated as Idd18 and is localized to a 5.1-cM region. The resolution of the originally defined Idd3 locus into at least four separate loci, Idd3, Idd10, Idd17, and Idd18, illustrates the complex polygenic nature of diabetes..
Denny, P.
Lord, C.J.
Hill, N.J.
Goy, J.V.
Levy, E.R.
Podolin, P.L.
Peterson, L.B.
Wicker, L.S.
Todd, J.A.
Lyons, P.A.
(1997). Mapping of the IDDM locus Idd3 to a 0 35-cM interval containing the interleukin-2 gene. Diabetes,
Vol.46
(4),
pp. 695-700.
show abstract
Currently, 16 loci that contribute to the development of IDDM in the NOD mouse have been mapped by linkage analysis. To fine map these loci, we used congenic mapping. Using this approach, we localized the Idd3 locus to a 0.35-cM interval on chromosome 3 containing the Il2 gene. Segregation analysis of the known variations within this interval indicated that only one variant, a serine-to-proline substitution at position 6 of the mature interleukin-2 (IL-2) protein, consistently segregates with IDDM in crosses between NOD and a series of nondiabetic mouse strains. These data, taken together with the immunomodulatory role of IL-2, provide circumstantial evidence in support of the hypothesis that Idd3 is an allelic variation of the Il2 gene, or a variant in strong linkage disequilibrium..
Barnardo, M.C.
Bunce, M.
Lord, C.J.
Welsh, K.I.
(1997). HLA-B*5603: sequence of a novel hybrid allele comprising B*56 and B*4601 segments. Tissue antigens,
Vol.49
(5),
pp. 496-498.
Podolin, P.L.
Denny, P.
Lord, C.J.
Hill, N.J.
Todd, J.A.
Peterson, L.B.
Wicker, L.S.
Lyons, P.A.
(1997). Congenic mapping of the insulin-dependent diabetes (Idd) gene, Idd10, localizes two genes mediating the Idd10 effect and eliminates the candidate Fcgr1. J immunol,
Vol.159
(4),
pp. 1835-1843.
show abstract
The development of autoimmune diabetes in the nonobese diabetic (NOD) mouse is under the control of multiple insulin-dependent diabetes (Idd) genes. The Idd3 gene, originally defined as a broad peak of linkage on mouse chromosome 3, was subsequently identified as two genes, Idd3 and Idd10, separated by at least 20 cM. The resistance alleles of Idd3 and Idd10 individually confer only partial protection from diabetes but, in combination, result in profound resistance to disease due to an epistatic genetic interaction. In this study, we used newly developed congenic strains to further localize Idd10. Surprisingly, we found that Idd10 itself comprises at least two linked loci: Idd10 and the newly designated Idd17. Idd17 was localized to a 1.1-cM region between D3Mit26 and D3Mit40, proximal to Fcgr1, a candidate gene encoding the high affinity Fc receptor for IgG. Idd10 was localized to a 10-cM region between D3Mit213 and D3Mit106, distal to Fcgr1. Thus, Fcgr1 was excluded as a candidate for either Idd10 or Idd17, despite the fact that the NOD strain expresses a mutant form of the receptor. Interestingly, although Idd10 and Idd17 participate in a genetic interaction with each other, Idd10 but not Idd17 participates in the genetic interaction with Idd3. Our study on chromosome 3 begins to reveal the extent of the polygenic nature of autoimmune diabetes, and demonstrates that the use of congenic strains is an effective mapping strategy, even in the dissection of multiple, linked genes with subtle effects..
Lord, C.J.
Bohlander, S.K.
Hopes, E.A.
Montague, C.T.
Hill, N.J.
Prins, J.B.
Renjilian, R.J.
Peterson, L.B.
Wicker, L.S.
Todd, J.A.
(1995). Mapping the diabetes polygene Idd3 on mouse chromosome 3 by use of novel congenic strains. Mamm genome,
Vol.6
(9),
pp. 563-570.
show abstract
Development of novel congenic mouse strains has allowed us to better define the location of the diabetogenic locus, Idd3, on Chromosome (Chr) 3. Congenic strains were identified by use of published and newly developed microsatellite markers, their genomes fingerprinted by a rapid, fluorescence-based approach, and their susceptibility to type 1 diabetes evaluated. The maximum interval containing Idd3 is now approximately 4 cM..
Mendes-Pereira, A.M.
Martin, S.A.
Brough, R.
McCarthy, A.
Taylor, J.R.
Kim, J.-.
Waldman, T.
Lord, C.J.
Ashworth, A.
Synthetic lethal targeting of PTEN mutant cells with PARP inhibitors. Embo molecular medicine,
Vol.1
(6-7),
pp. 315-8.
Martin, S.A.
McCarthy, A.
Barber, L.J.
Burgess, D.J.
Parry, S.
Lord, C.J.
Ashworth, A.
Methotrexate induces oxidative DNA damage and is selectively lethal to tumour cells with defects in the DNA mismatch repair gene MSH2. Embo molecular medicine,
Vol.1
(6-7),
pp. 323-15.
Lord, C.J.
Ashworth, A.
PARP inhibitors: Synthetic lethality in the clinic. Science,
Vol.355
(6330),
pp. 1152-1158.
show abstract
full text
PARP inhibitors (PARPi), a cancer therapy targeting poly(ADP-ribose) polymerase, are the first clinically approved drugs designed to exploit synthetic lethality, a genetic concept proposed nearly a century ago. Tumors arising in patients who carry germline mutations in either BRCA1 or BRCA2 are sensitive to PARPi because they have a specific type of DNA repair defect. PARPi also show promising activity in more common cancers that share this repair defect. However, as with other targeted therapies, resistance to PARPi arises in advanced disease. In addition, determining the optimal use of PARPi within drug combination approaches has been challenging. Nevertheless, the preclinical discovery of PARPi synthetic lethality and the route to clinical approval provide interesting lessons for the development of other therapies. Here, we discuss current knowledge of PARP inhibitors and potential ways to maximize their clinical effectiveness..