Fu, X.
Zhao, Y.
Lopez, J.I.
Rowan, A.
Au, L.
Fendler, A.
Hazell, S.
Xu, H.
Horswell, S.
Shepherd, S.T.
Spencer, C.E.
Spain, L.
Byrne, F.
Stamp, G.
O'Brien, T.
Nicol, D.
Augustine, M.
Chandra, A.
Rudman, S.
Toncheva, A.
Furness, A.J.
Pickering, L.
Kumar, S.
Koh, D.-.
Messiou, C.
Dafydd, D.A.
Orton, M.R.
Doran, S.J.
Larkin, J.
Swanton, C.
Sahai, E.
Litchfield, K.
Turajlic, S.
TRACERx Renal Consortium,
Bates, P.A.
(2022). Spatial patterns of tumour growth impact clonal diversification in a computational model and the TRACERx Renal study. Nat ecol evol,
Vol.6
(1),
pp. 88-102.
show abstract
full text
Genetic intra-tumour heterogeneity fuels clonal evolution, but our understanding of clinically relevant clonal dynamics remain limited. We investigated spatial and temporal features of clonal diversification in clear cell renal cell carcinoma through a combination of modelling and real tumour analysis. We observe that the mode of tumour growth, surface or volume, impacts the extent of subclonal diversification, enabling interpretation of clonal diversity in patient tumours. Specific patterns of proliferation and necrosis explain clonal expansion and emergence of parallel evolution and microdiversity in tumours. In silico time-course studies reveal the appearance of budding structures before detectable subclonal diversification. Intriguingly, we observe radiological evidence of budding structures in early-stage clear cell renal cell carcinoma, indicating that future clonal evolution may be predictable from imaging. Our findings offer a window into the temporal and spatial features of clinically relevant clonal evolution..
Shum, B.
Larkin, J.
Turajlic, S.
(2022). Predictive biomarkers for response to immune checkpoint inhibition. Semin cancer biol,
Vol.79,
pp. 4-17.
show abstract
Immune checkpoint inhibitors have transformed the prognosis and treatment paradigm of many cancer types, through the potential for durable responses. However, the majority of patients still do not benefit. Response to checkpoint inhibition is determined by dynamic host, tumour and tumour microenvironment factors that display spatial and temporal variability, but our understanding of these interactions is incomplete. Through investigating biomarkers of resistance and response, opportunities arise to discover new therapeutic targets and shape personalised treatment strategies. Here we review approved and emerging biomarkers of response to immune checkpoint inhibitors, in particular the recent rapid progress in host and tumour genomics. It is unlikely that a single biomarker will precisely predict response, but multivariate multiomic markers may provide a balanced assessment of these factors and more accurately identify patients who will benefit. Further efforts are required to translate these groundbreaking discoveries into novel therapeutics and biomarker driven clinical trials, to provide durable treatment response to a greater population of patients..
Siddiqui, M.S.
Lai, Z.M.
Spain, L.
Greener, V.
Turajlic, S.
Larkin, J.
Morganstein, D.L.
(2021). Predicting development of ipilimumab-induced hypophysitis: utility of T4 and TSH index but not TSH. J endocrinol invest,
Vol.44
(1),
pp. 195-203.
show abstract
full text
PURPOSE: Ipilimumab, a monoclonal antibody inhibiting CLTA-4, is an established treatment in metastatic melanoma, either alone or in combination with nivolumab, and results in immune mediated adverse events, including endocrinopathy. Hypophysitis is one of the most common endocrine abnormalities. An early recognition of hypophysitis may prevent life threatening consequences of hypopituitarism; therefore, biomarkers to predict which patients will develop hypophysitis would have clinical utility. Recent studies suggested that a decline in TSH may serve as an early marker of IH. This study was aimed at assessing the utility of thyroid function tests in predicting development of hypophysitis. METHODS: A retrospective cohort study was performed for all patients (n = 308) treated with ipilimumab either as a monotherapy or in combination with nivolumab for advanced melanoma at the Royal Marsden Hospital from 2010 to 2016. Thyroid function tests, other pituitary function tests and Pituitary MRIs were used to identify those with hypophysitis. RESULTS AND CONCLUSIONS: Ipilimumab-induced hypophysitis (IH) was diagnosed in 25 patients (8.15%). A decline in TSH was observed in hypophysitis cohort during the first three cycles but it did not reach statistical significance (P = 0.053). A significant fall in FT4 (P < 0.001), TSH index (P < 0.001) and standardised TSH index (P < 0.001) prior to cycles 3 and 4 in hypophysitis cohort was observed. TSH is not useful in predicting development of IH. FT4, TSH index and standardised TSH index may be valuable but a high index of clinical suspicion remains paramount in early detection of hypophysitis..
Tippu, Z.
Au, L.
Turajlic, S.
(2021). Evolution of Renal Cell Carcinoma. Eur urol focus,
Vol.7
(1),
pp. 148-151.
show abstract
A diverse range of clinical phenotypes are observed in clear-cell renal cell carcinoma (ccRCC) with resultant therapeutic implications. Molecular characterisation underpinning the diversity observed has been qualified, where the near ubiquitous loss of the short arm of chromosome 3 precedes distinct evolutionary trajectories involving different sequences and combinations of genetic events. Primary tumours are characterised by varying degrees of intratumoural heterogeneity and chromosomal complexity associated with distinct metastatic phenotypes. An evolutionary understanding reconciles the diverse clinical phenotypes observed in metastatic ccRCC and has the potential to impact patient management. PATIENT SUMMARY: Re-tracing the evolutionary paths taken by clear-cell kidney cancer through analyses at the gene level gives an insight into genetic changes that are correlated to metastatic potential. This may provide a framework to guide therapeutic interventions, especially in identifying candidates for surgical intervention..
Edwards, C.L.
Comito, F.
Agraso Busto, S.
Harland, C.
Turajlic, S.
Larkin, J.
Heelan, K.
Fearfield, L.
(2021). Cutaneous toxicities in patients with melanoma receiving checkpoint inhibitor therapy: a retrospective review The experience of a single large specialist institution. Clin exp dermatol,
Vol.46
(2),
pp. 338-341.
show abstract
Checkpoint inhibitor (CPI) therapy has significantly improved overall survival for metastatic melanoma, and is now approved for use in the adjuvant setting. Modulating the immune system is recognized to cause cutaneous immune-related adverse events (irAEs). We conducted a retrospective observational cohort study of adult patients with melanoma at our tertiary referral centre, who received CPI therapy from 2006 to March 2018. This is the single largest study of cutaneous irAEs occurring on CPI therapy in patients with melanoma to date and encompasses 12 years. The results showed that cutaneous toxicity occurs in 24% of patients but is generally manageable, with < 5% patients discontinuing treatment..
Litchfield, K.
Reading, J.L.
Puttick, C.
Thakkar, K.
Abbosh, C.
Bentham, R.
Watkins, T.B.
Rosenthal, R.
Biswas, D.
Rowan, A.
Lim, E.
Al Bakir, M.
Turati, V.
Guerra-Assunção, J.A.
Conde, L.
Furness, A.J.
Saini, S.K.
Hadrup, S.R.
Herrero, J.
Lee, S.-.
Van Loo, P.
Enver, T.
Larkin, J.
Hellmann, M.D.
Turajlic, S.
Quezada, S.A.
McGranahan, N.
Swanton, C.
(2021). Meta-analysis of tumor- and T cell-intrinsic mechanisms of sensitization to checkpoint inhibition. Cell,
Vol.184
(3),
pp. 596-614.e14.
show abstract
full text
Checkpoint inhibitors (CPIs) augment adaptive immunity. Systematic pan-tumor analyses may reveal the relative importance of tumor-cell-intrinsic and microenvironmental features underpinning CPI sensitization. Here, we collated whole-exome and transcriptomic data for >1,000 CPI-treated patients across seven tumor types, utilizing standardized bioinformatics workflows and clinical outcome criteria to validate multivariable predictors of CPI sensitization. Clonal tumor mutation burden (TMB) was the strongest predictor of CPI response, followed by total TMB and CXCL9 expression. Subclonal TMB, somatic copy alteration burden, and histocompatibility leukocyte antigen (HLA) evolutionary divergence failed to attain pan-cancer significance. Dinucleotide variants were identified as a source of immunogenic epitopes associated with radical amino acid substitutions and enhanced peptide hydrophobicity/immunogenicity. Copy-number analysis revealed two additional determinants of CPI outcome supported by prior functional evidence: 9q34 (TRAF2) loss associated with response and CCND1 amplification associated with resistance. Finally, single-cell RNA sequencing (RNA-seq) of clonal neoantigen-reactive CD8 tumor-infiltrating lymphocytes (TILs), combined with bulk RNA-seq analysis of CPI-responding tumors, identified CCR5 and CXCL13 as T-cell-intrinsic markers of CPI sensitivity..
Rini, B.
Abel, E.J.
Albiges, L.
Bex, A.
Brugarolas, J.
Bukowski, R.M.
Coleman, J.A.
Drake, C.G.
Figlin, R.A.
Futreal, A.
Hammers, H.
Powles, T.
Rathmell, W.K.
Ricketts, C.J.
Turajlic, S.
Wood, C.G.
Leibovich, B.C.
(2021). Summary from the Kidney Cancer Association's Inaugural Think Thank: Coalition for a Cure. Clin genitourin cancer,
Vol.19
(2),
pp. 167-175.
show abstract
Close to 74,000 cases of renal cell carcinoma (RCC) are diagnosed each year in the United States. The past 2 decades have shown great developments in surgical techniques, targeted therapy and immunotherapy agents, and longer complete response rates. However, without a global cure, there is still room for further advancement in improving patient care in this space. To address some of the gaps restricting this progress, the Kidney Cancer Association brought together a group of 27 specialists across the areas of clinical care, research, industry, and advocacy at the inaugural "Think Tank: Coalition for a Cure" session. Topics addressed included screening, imaging, rarer RCC subtypes, combination drug therapy options, and patient response. This commentary summarizes the discussion of these topics and their respective clinical challenges, along with a proposal of projects for collaboration in overcoming those needs and making a greater impact on care for patients with RCC moving forward..
Bailey, C.
Black, J.R.
Reading, J.L.
Litchfield, K.
Turajlic, S.
McGranahan, N.
Jamal-Hanjani, M.
Swanton, C.
(2021). Tracking Cancer Evolution through the Disease Course. Cancer discov,
Vol.11
(4),
pp. 916-932.
show abstract
During cancer evolution, constituent tumor cells compete under dynamic selection pressures. Phenotypic variation can be observed as intratumor heterogeneity, which is propagated by genome instability leading to mutations, somatic copy-number alterations, and epigenomic changes. TRACERx was set up in 2014 to observe the relationship between intratumor heterogeneity and patient outcome. By integrating multiregion sequencing of primary tumors with longitudinal sampling of a prospectively recruited patient cohort, cancer evolution can be tracked from early- to late-stage disease and through therapy. Here we review some of the key features of the studies and look to the future of the field. SIGNIFICANCE: Cancers evolve and adapt to environmental challenges such as immune surveillance and treatment pressures. The TRACERx studies track cancer evolution in a clinical setting, through primary disease to recurrence. Through multiregion and longitudinal sampling, evolutionary processes have been detailed in the tumor and the immune microenvironment in non-small cell lung cancer and clear-cell renal cell carcinoma. TRACERx has revealed the potential therapeutic utility of targeting clonal neoantigens and ctDNA detection in the adjuvant setting as a minimal residual disease detection tool primed for translation into clinical trials..
Gudd, C.L.
Au, L.
Triantafyllou, E.
Shum, B.
Liu, T.
Nathwani, R.
Kumar, N.
Mukherjee, S.
Dhar, A.
Woollard, K.J.
Yone, Y.
Pinato, D.J.
Thursz, M.R.
Goldin, R.D.
Gore, M.E.
Larkin, J.
Khamri, W.
Antoniades, C.G.
Turajlic, S.
Possamai, L.A.
(2021). Activation and transcriptional profile of monocytes and CD8+ T cells are altered in checkpoint inhibitor-related hepatitis. J hepatol,
Vol.75
(1),
pp. 177-189.
show abstract
full text
BACKGROUND & AIMS: Checkpoint inhibitor-related hepatitis (CPI-Hep) is an emerging clinical challenge. We aimed to gain insights into the immunopathology of CPI-Hep by comprehensively characterising myeloid and lymphoid subsets. METHODS: CPI-treated patients with or without related hepatitis (CPI-Hep; n = 22 and CPI-noHep; n = 7) were recruited. Phenotypic and transcriptional profiling of peripheral immune subsets was performed and compared with 19 healthy controls (HCs). In vitro monocyte-derived macrophages (MoMFs) were assessed for activation and cytokine production. CD163, CCR2, CD68, CD3, CD8 and granzyme B expression was assessed using immunohistochemistry/immunofluorescence (n = 4). RESULTS: A significant total monocyte depletion was observed in CPI-Hep compared with HCs (p = 0.04), along with a proportionate increase in the classical monocyte population (p = 0.0002) and significant upregulation of CCR2, CD163 and downregulation of CCR7. Soluble CD163 levels were significantly elevated in CPI-Hep compared with HCs (p <0.0001). In vitro MoMFs from CPI-Hep showed enhanced production of pro-inflammatory cytokines. CD8+ T cells demonstrated increased perforin, granzyme B, ICOS and HLA-DR expression in CPI-Hep. Transcriptional profiling indicated the presence of activated monocyte and enhanced effector CD8+ T cell populations in CPI-Hep. Immunohistochemistry demonstrated co-localisation of CD8+/granzyme B+ T cells with CD68+CCR2+/CD68+CD163+ macrophages in CPI-Hep liver tissue. CONCLUSIONS: CPI-Hep is associated with activation of peripheral monocytes and an enhanced cytotoxic, effector CD8+ T cell phenotype. These changes were reflected by liver inflammation composed of CD163+/CCR2+ macrophages and CD8+ T cells. LAY SUMMARY: Some patients who receive immunotherapy for cancer develop liver inflammation, which requires cessation of cancer treatment. Herein, we describe ways in which the white blood cells of patients who develop liver inflammation differ from those of patients who receive the same immunotherapy but do not experience liver-related side effects. Targeting some of the pathways we identify may help to prevent or manage this side effect and facilitate cancer treatment..
Rzeniewicz, K.
Larkin, J.
Menzies, A.M.
Turajlic, S.
(2021). Immunotherapy use outside clinical trial populations: never say never?. Ann oncol,
Vol.32
(7),
pp. 866-880.
show abstract
BACKGROUND: Based on favourable outcomes in clinical trials, immune checkpoint inhibitors (ICIs), most notably programmed cell death protein 1 (PD-1) and cytotoxic T-lymphocyte antigen-4 (CTLA-4) inhibitors, are now widely used across multiple cancer types. However, due to their strict inclusion and exclusion criteria, clinical studies often do not address challenges presented by non-trial populations. DESIGN: This review summarises available data on the efficacy and safety of ICIs in trial-ineligible patients, including those with autoimmune disease, chronic viral infections, organ transplants, organ dysfunction, poor performance status, and brain metastases, as well as the elderly, children, and those who are pregnant. In addition, we review data concerning other real-world challenges with ICIs, including timing of therapy switch, relationships to radiotherapy or surgery, re-treatment after an immune-related toxicity, vaccinations in patients on ICIs, and current experience around ICI and coronavirus disease-19. Where possible, we provide recommendations to aid the often-difficult decision-making process in those settings. CONCLUSIONS: Data suggest that ICIs are often active and have an acceptable safety profile in the populations described above, with the exception of PD-1 inhibitors in solid organ transplant recipients. Decisions about whether to treat with ICIs should be personalised and require multidisciplinary input and careful counselling of patients with respect to potential risks and benefits. Clinical judgements need to be carefully weighed, considering factors such as underlying cancer type, feasibility of alternative treatment options, or activity in trial-eligible patients..
Zhao, Y.
Fu, X.
Lopez, J.I.
Rowan, A.
Au, L.
Fendler, A.
Hazell, S.
Xu, H.
Horswell, S.
Shepherd, S.T.
Spain, L.
Byrne, F.
Stamp, G.
O'Brien, T.
Nicol, D.
Augustine, M.
Chandra, A.
Rudman, S.
Toncheva, A.
Pickering, L.
Sahai, E.
Larkin, J.
Bates, P.A.
Swanton, C.
Turajlic, S.
TRACERx Renal Consortium,
Litchfield, K.
(2021). Selection of metastasis competent subclones in the tumour interior. Nat ecol evol,
Vol.5
(7),
pp. 1033-1045.
show abstract
full text
The genetic evolutionary features of solid tumour growth are becoming increasingly well described, but the spatial and physical nature of subclonal growth remains unclear. Here, we utilize 102 macroscopic whole-tumour images from clear cell renal cell carcinoma patients, with matched genetic and phenotypic data from 756 biopsies. Utilizing a digital image processing pipeline, a renal pathologist marked the boundaries between tumour and normal tissue and extracted positions of boundary line and biopsy regions to X and Y coordinates. We then integrated coordinates with genomic data to map exact spatial subclone locations, revealing how genetically distinct subclones grow and evolve spatially. We observed a phenotype of advanced and more aggressive subclonal growth in the tumour centre, characterized by an elevated burden of somatic copy number alterations and higher necrosis, proliferation rate and Fuhrman grade. Moreover, we found that metastasizing subclones preferentially originate from the tumour centre. Collectively, these observations suggest a model of accelerated evolution in the tumour interior, with harsh hypoxic environmental conditions leading to a greater opportunity for driver somatic copy number alterations to arise and expand due to selective advantage. Tumour subclone growth is predominantly spatially contiguous in nature. We found only two cases of subclone dispersal, one of which was associated with metastasis. The largest subclones spatially were dominated by driver somatic copy number alterations, suggesting that a large selective advantage can be conferred to subclones upon acquisition of these alterations. In conclusion, spatial dynamics is strongly associated with genomic alterations and plays an important role in tumour evolution..
Switzer, B.
Haanen, J.
Lorigan, P.C.
Puzanov, I.
Turajlic, S.
(2021). Clinical and immunologic implications of COVID-19 in patients with melanoma and renal cell carcinoma receiving immune checkpoint inhibitors. J immunother cancer,
Vol.9
(7).
show abstract
full text
The clinical and immunologic implications of the SARS-CoV-2 pandemic for patients with cancer receiving systemic anticancer therapy have introduced a multitude of clinical challenges and academic controversies. This review summarizes the current evidence, discussion points, and recommendations regarding the use of immune checkpoint inhibitors (ICIs) in patients with cancer during the SARS-CoV-2 pandemic, with a focus on patients with melanoma and renal cell carcinoma (RCC). More specifically, we summarize the theoretical concepts and available objective data regarding the relationships between ICIs and the antiviral immune response, along with recommended clinical approaches to the management of melanoma and RCC patient cohorts receiving ICIs throughout the course of the COVID-19 pandemic. Additional insights regarding the use of ICIs in the setting of current and upcoming COVID-19 vaccines and broader implications toward future pandemics are also discussed..
Alexander, J.L.
Ibraheim, H.
Sheth, B.
Little, J.
Khan, M.S.
Richards, C.
Hunter, N.
Chauhan, D.
Ratnakumaran, R.
McHugh, K.
Pinato, D.J.
Nathan, P.
Choy, J.
Crusz, S.M.
Furness, A.
Turajlic, S.
Pickering, L.
Larkin, J.
Teare, J.P.
Papa, S.
Speight, A.
Sharma, A.
Powell, N.
(2021). Clinical outcomes of patients with corticosteroid refractory immune checkpoint inhibitor-induced enterocolitis treated with infliximab. J immunother cancer,
Vol.9
(7).
show abstract
full text
INTRODUCTION: Immune checkpoint inhibitors (CPIs) have changed the treatment landscape for many cancers, but also cause severe inflammatory side effects including enterocolitis. CPI-induced enterocolitis is treated empirically with corticosteroids, and infliximab (IFX) is used in corticosteroid-refractory cases. However, robust outcome data for these patients are scarce. METHODS: We conducted a multicenter (six cancer centers), cohort study of outcomes in patients treated with IFX for corticosteroid-refractory CPI-induced enterocolitis between 2007 and 2020. The primary outcome was corticosteroid-free clinical remission (CFCR) with Common Terminology Criteria for Adverse Events (CTCAE) grade 0 for diarrhea at 12 weeks after IFX initiation. We also assessed cancer outcomes at 1 year using RECIST V1.1 criteria. RESULTS: 127 patients (73 male; median age 59 years) were treated with IFX for corticosteroid-refractory CPI-induced enterocolitis. Ninety-six (75.6%) patients had diarrhea CTCAE grade >2 and 115 (90.6%) required hospitalization for colitis. CFCR was 41.2% at 12 weeks and 50.9% at 26 weeks. In multivariable logistic regression, IFX-resistant enterocolitis was associated with rectal bleeding (OR 0.19; 95% CI 0.04 to 0.80; p=0.03) and absence of colonic crypt abscesses (OR 2.16; 95% CI 1.13 to 8.05; p=0.03). Cancer non-progression was significantly more common in patients with IFX-resistant enterocolitis (64.4%) as compared with patients with IFX-responsive enterocolitis (37.5%; p=0.013). CONCLUSION: This is the largest study to date reporting outcomes of IFX therapy in patients with corticosteroid-refractory CPI-induced enterocolitis. Using predefined robust endpoints, we have demonstrated that fewer than half of patients achieved CFCR. Our data also indicate that cancer outcomes may be better in patients developing prolonged and severe inflammatory side effects of CPI therapy..
Au, L.
Fendler, A.
Shepherd, S.T.
Rzeniewicz, K.
Cerrone, M.
Byrne, F.
Carlyle, E.
Edmonds, K.
Del Rosario, L.
Shon, J.
Haynes, W.A.
Ward, B.
Shum, B.
Gordon, W.
Gerard, C.L.
Xie, W.
Joharatnam-Hogan, N.
Young, K.
Pickering, L.
Furness, A.J.
Larkin, J.
Harvey, R.
Kassiotis, G.
Gandhi, S.
Crick COVID-19 Consortium,
Swanton, C.
Fribbens, C.
Wilkinson, K.A.
Wilkinson, R.J.
Lau, D.K.
Banerjee, S.
Starling, N.
Chau, I.
CAPTURE Consortium,
Turajlic, S.
(2021). Cytokine release syndrome in a patient with colorectal cancer after vaccination with BNT162b2. Nat med,
Vol.27
(8),
pp. 1362-1366.
show abstract
full text
Patients with cancer are currently prioritized in coronavirus disease 2019 (COVID-19) vaccination programs globally, which includes administration of mRNA vaccines. Cytokine release syndrome (CRS) has not been reported with mRNA vaccines and is an extremely rare immune-related adverse event of immune checkpoint inhibitors. We present a case of CRS that occurred 5 d after vaccination with BTN162b2 (tozinameran)-the Pfizer-BioNTech mRNA COVID-19 vaccine-in a patient with colorectal cancer on long-standing anti-PD-1 monotherapy. The CRS was evidenced by raised inflammatory markers, thrombocytopenia, elevated cytokine levels (IFN-γ/IL-2R/IL-18/IL-16/IL-10) and steroid responsiveness. The close temporal association of vaccination and diagnosis of CRS in this case suggests that CRS was a vaccine-related adverse event; with anti-PD1 blockade as a potential contributor. Overall, further prospective pharmacovigillence data are needed in patients with cancer, but the benefit-risk profile remains strongly in favor of COVID-19 vaccination in this population..
Au, L.
Hatipoglu, E.
Robert de Massy, M.
Litchfield, K.
Beattie, G.
Rowan, A.
Schnidrig, D.
Thompson, R.
Byrne, F.
Horswell, S.
Fotiadis, N.
Hazell, S.
Nicol, D.
Shepherd, S.T.
Fendler, A.
Mason, R.
Del Rosario, L.
Edmonds, K.
Lingard, K.
Sarker, S.
Mangwende, M.
Carlyle, E.
Attig, J.
Joshi, K.
Uddin, I.
Becker, P.D.
Sunderland, M.W.
Akarca, A.
Puccio, I.
Yang, W.W.
Lund, T.
Dhillon, K.
Vasquez, M.D.
Ghorani, E.
Xu, H.
Spencer, C.
López, J.I.
Green, A.
Mahadeva, U.
Borg, E.
Mitchison, M.
Moore, D.A.
Proctor, I.
Falzon, M.
Pickering, L.
Furness, A.J.
Reading, J.L.
Salgado, R.
Marafioti, T.
Jamal-Hanjani, M.
PEACE Consortium,
Kassiotis, G.
Chain, B.
Larkin, J.
Swanton, C.
Quezada, S.A.
Turajlic, S.
TRACERx Renal Consortium,
(2021). Determinants of anti-PD-1 response and resistance in clear cell renal cell carcinoma. Cancer cell,
Vol.39
(11),
pp. 1497-1518.e11.
show abstract
full text
ADAPTeR is a prospective, phase II study of nivolumab (anti-PD-1) in 15 treatment-naive patients (115 multiregion tumor samples) with metastatic clear cell renal cell carcinoma (ccRCC) aiming to understand the mechanism underpinning therapeutic response. Genomic analyses show no correlation between tumor molecular features and response, whereas ccRCC-specific human endogenous retrovirus expression indirectly correlates with clinical response. T cell receptor (TCR) analysis reveals a significantly higher number of expanded TCR clones pre-treatment in responders suggesting pre-existing immunity. Maintenance of highly similar clusters of TCRs post-treatment predict response, suggesting ongoing antigen engagement and survival of families of T cells likely recognizing the same antigens. In responders, nivolumab-bound CD8+ T cells are expanded and express GZMK/B. Our data suggest nivolumab drives both maintenance and replacement of previously expanded T cell clones, but only maintenance correlates with response. We hypothesize that maintenance and boosting of a pre-existing response is a key element of anti-PD-1 mode of action..
Mendis, S.
Ealing, J.
Larkin, J.
Turajlic, S.
Carr, A.
Bronstein, A.
Kaski, D.
(2021). Isolated imbalance due to bilateral vestibular failure following immune checkpoint inhibitor administration: two cases. Eur j cancer,
Vol.156,
pp. 187-189.
Fendler, A.
Shepherd, S.T.
Au, L.
Wilkinson, K.A.
Wu, M.
Byrne, F.
Cerrone, M.
Schmitt, A.M.
Joharatnam-Hogan, N.
Shum, B.
Tippu, Z.
Rzeniewicz, K.
Boos, L.A.
Harvey, R.
Carlyle, E.
Edmonds, K.
Del Rosario, L.
Sarker, S.
Lingard, K.
Mangwende, M.
Holt, L.
Ahmod, H.
Korteweg, J.
Foley, T.
Bazin, J.
Gordon, W.
Barber, T.
Emslie-Henry, A.
Xie, W.
Gerard, C.L.
Deng, D.
Wall, E.C.
Agua-Doce, A.
Namjou, S.
Caidan, S.
Gavrielides, M.
MacRae, J.I.
Kelly, G.
Peat, K.
Kelly, D.
Murra, A.
Kelly, K.
O'Flaherty, M.
Dowdie, L.
Ash, N.
Gronthoud, F.
Shea, R.L.
Gardner, G.
Murray, D.
Kinnaird, F.
Cui, W.
Pascual, J.
Rodney, S.
Mencel, J.
Curtis, O.
Stephenson, C.
Robinson, A.
Oza, B.
Farag, S.
Leslie, I.
Rogiers, A.
Iyengar, S.
Ethell, M.
Messiou, C.
Cunningham, D.
Chau, I.
Starling, N.
Turner, N.
Welsh, L.
van As, N.
Jones, R.L.
Droney, J.
Banerjee, S.
Tatham, K.C.
O'Brien, M.
Harrington, K.
Bhide, S.
Okines, A.
Reid, A.
Young, K.
Furness, A.J.
Pickering, L.
Swanton, C.
Crick COVID-19 Consortium,
Gandhi, S.
Gamblin, S.
Bauer, D.L.
Kassiotis, G.
Kumar, S.
Yousaf, N.
Jhanji, S.
Nicholson, E.
Howell, M.
Walker, S.
Wilkinson, R.J.
Larkin, J.
Turajlic, S.
CAPTURE Consortium,
(2021). Adaptive immunity and neutralizing antibodies against SARS-CoV-2 variants of concern following vaccination in patients with cancer: the CAPTURE study. Nat cancer,
Vol.2
(12),
pp. 1305-1320.
show abstract
full text
Coronavirus disease 2019 (COVID-19) antiviral response in a pan-tumor immune monitoring (CAPTURE) ( NCT03226886 ) is a prospective cohort study of COVID-19 immunity in patients with cancer. Here we evaluated 585 patients following administration of two doses of BNT162b2 or AZD1222 vaccines, administered 12 weeks apart. Seroconversion rates after two doses were 85% and 59% in patients with solid and hematological malignancies, respectively. A lower proportion of patients had detectable titers of neutralizing antibodies (NAbT) against severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) variants of concern (VOC) versus wild-type (WT) SARS-CoV-2. Patients with hematological malignancies were more likely to have undetectable NAbT and had lower median NAbT than those with solid cancers against both SARS-CoV-2 WT and VOC. By comparison with individuals without cancer, patients with hematological, but not solid, malignancies had reduced neutralizing antibody (NAb) responses. Seroconversion showed poor concordance with NAbT against VOC. Previous SARS-CoV-2 infection boosted the NAb response including against VOC, and anti-CD20 treatment was associated with undetectable NAbT. Vaccine-induced T cell responses were detected in 80% of patients and were comparable between vaccines or cancer types. Our results have implications for the management of patients with cancer during the ongoing COVID-19 pandemic..
Fendler, A.
Au, L.
Shepherd, S.T.
Byrne, F.
Cerrone, M.
Boos, L.A.
Rzeniewicz, K.
Gordon, W.
Shum, B.
Gerard, C.L.
Ward, B.
Xie, W.
Schmitt, A.M.
Joharatnam-Hogan, N.
Cornish, G.H.
Pule, M.
Mekkaoui, L.
Ng, K.W.
Carlyle, E.
Edmonds, K.
Rosario, L.D.
Sarker, S.
Lingard, K.
Mangwende, M.
Holt, L.
Ahmod, H.
Stone, R.
Gomes, C.
Flynn, H.R.
Agua-Doce, A.
Hobson, P.
Caidan, S.
Howell, M.
Wu, M.
Goldstone, R.
Crawford, M.
Cubitt, L.
Patel, H.
Gavrielides, M.
Nye, E.
Snijders, A.P.
MacRae, J.I.
Nicod, J.
Gronthoud, F.
Shea, R.L.
Messiou, C.
Cunningham, D.
Chau, I.
Starling, N.
Turner, N.
Welsh, L.
van As, N.
Jones, R.L.
Droney, J.
Banerjee, S.
Tatham, K.C.
Jhanji, S.
O'Brien, M.
Curtis, O.
Harrington, K.
Bhide, S.
Bazin, J.
Robinson, A.
Stephenson, C.
Slattery, T.
Khan, Y.
Tippu, Z.
Leslie, I.
Gennatas, S.
Okines, A.
Reid, A.
Young, K.
Furness, A.J.
Pickering, L.
Gandhi, S.
Gamblin, S.
Swanton, C.
Crick COVID-19 Consortium,
Nicholson, E.
Kumar, S.
Yousaf, N.
Wilkinson, K.A.
Swerdlow, A.
Harvey, R.
Kassiotis, G.
Larkin, J.
Wilkinson, R.J.
Turajlic, S.
CAPTURE consortium,
(2021). Functional antibody and T cell immunity following SARS-CoV-2 infection, including by variants of concern, in patients with cancer: the CAPTURE study. Nat cancer,
Vol.2
(12),
pp. 1321-1337.
show abstract
full text
Patients with cancer have higher COVID-19 morbidity and mortality. Here we present the prospective CAPTURE study, integrating longitudinal immune profiling with clinical annotation. Of 357 patients with cancer, 118 were SARS-CoV-2 positive, 94 were symptomatic and 2 died of COVID-19. In this cohort, 83% patients had S1-reactive antibodies and 82% had neutralizing antibodies against wild type SARS-CoV-2, whereas neutralizing antibody titers against the Alpha, Beta and Delta variants were substantially reduced. S1-reactive antibody levels decreased in 13% of patients, whereas neutralizing antibody titers remained stable for up to 329 days. Patients also had detectable SARS-CoV-2-specific T cells and CD4+ responses correlating with S1-reactive antibody levels, although patients with hematological malignancies had impaired immune responses that were disease and treatment specific, but presented compensatory cellular responses, further supported by clinical recovery in all but one patient. Overall, these findings advance the understanding of the nature and duration of the immune response to SARS-CoV-2 in patients with cancer..
Gallegos, L.L.
Gilchrist, A.
Spain, L.
Stanislaw, S.
Hill, S.M.
Primus, V.
Jones, C.
Agrawal, S.
Tippu, Z.
Barhoumi, A.
Noel-Storr, G.
Alexander, N.R.
Turajlic, S.
(2021). A protocol for representative sampling of solid tumors to improve the accuracy of sequencing results. Star protoc,
Vol.2
(3),
p. 100624.
show abstract
full text
Owing to spatial segregation of tumor subclones, solid tumor sampling using formalin-fixed, paraffin-embedded blocks is often inadequate to represent the genomic heterogeneity of solid tumors. We present an approach, representative sampling, to dissect and homogenize leftover residual surgical tissue prior to sequencing. We also detail optional tumor cell enrichment and DNA preparation. This method, applicable only to surgically removed tumors with leftover tissue, facilitates robust sampling to avoid missing or over-representing actionable variants. For complete details on the use and execution of this protocol, please refer to Litchfield et al. (2020)..
Fu, X.
Zhao, Y.
Lopez, J.
Rowan, A.
Au, L.
Fendler, A.
Hazell, S.
Xu, H.
Horswell, S.
Shepherd, S.
Spain, L.
Byrne, F.
Stamp, G.
O'Brien, T.
Nicol, D.
Augustine, M.
Chandra, A.
Rudman, S.
Toncheva, A.
Furness, A.
Pickering, L.
Orton, M.
Doran, S.
Koh, D.-.
Messiou, C.
Dafydd, D.A.
Kumar, S.
Larkin, J.
Swanton, C.
Sahai, E.
Litchfield, K.
Turajlic, S.
Bates, P.
(2021). Spatial patterns of tumour growth impact clonal diversification: computational modelling and evidence in the TRACERx Renal study. ,
.
show abstract
Abstract
Intra-tumour genetic heterogeneity (ITH) fuels cancer evolution. The role of clonal diversity and genetic complexity in the progression of clear-cell renal cell carcinomas (ccRCCs) has been characterised, but the ability to predict clinically relevant evolutionary trajectories remains limited. Here, towards enhancing this ability, we investigated spatial features of clonal diversification through a combined computational modelling and experimental analysis in the TRACERx Renal study. We observe through modelling that spatial patterns of tumour growth impact the extent and trajectory of subclonal diversification. Moreover, subpopulations with high clonal diversity, and parallel evolution events, are frequently observed near the tumour margin. In-silico time-course studies further showed that budding structures on the tumour surface could indicate future steps of subclonal evolution. Such structures were evident radiologically in 15 early-stage ccRCCs, raising the possibility that spatially resolved sampling of these regions, when combined with sequencing, may enable identification of evolutionary potential in early-stage tumours..
Buck, M.D.
Poirier, E.Z.
Cardoso, A.
Frederico, B.
Canton, J.
Barrell, S.
Beale, R.
Byrne, R.
Caidan, S.
Crawford, M.
Cubitt, L.
Gandhi, S.
Goldstone, R.
Grant, P.R.
Gulati, K.
Hindmarsh, S.
Howell, M.
Hubank, M.
Instrell, R.
Jiang, M.
Kassiotis, G.
Lu, W.-.
MacRae, J.I.
Martini, I.
Miller, D.
Moore, D.
Nastouli, E.
Nicod, J.
Nightingale, L.
Olsen, J.
Oomatia, A.
O'Reilly, N.
Rideg, A.
Song, O.-.
Strange, A.
Swanton, C.
Turajlic, S.
Wu, M.
Reis e Sousa, C.
Crick COVID-19 Consortium,
(2021). SARS-CoV-2 detection by a clinical diagnostic RT-LAMP assay. Wellcome open res,
Vol.6,
p. 9.
show abstract
full text
The ongoing pandemic of SARS-CoV-2 calls for rapid and cost-effective methods to accurately identify infected individuals. The vast majority of patient samples is assessed for viral RNA presence by RT-qPCR. Our biomedical research institute, in collaboration between partner hospitals and an accredited clinical diagnostic laboratory, established a diagnostic testing pipeline that has reported on more than 252,000 RT-qPCR results since its commencement at the beginning of April 2020. However, due to ongoing demand and competition for critical resources, alternative testing strategies were sought. In this work, we present a clinically-validated procedure for high-throughput SARS-CoV-2 detection by RT-LAMP that is robust, reliable, repeatable, specific, and inexpensive..
Hodzic, E.
Shrestha, R.
Malikic, S.
Collins, C.C.
Litchfield, K.
Turajlic, S.
Sahinalp, S.C.
(2020). Identification of conserved evolutionary trajectories in tumors. Bioinformatics,
Vol.36
(Suppl_1),
pp. i427-i435.
show abstract
MOTIVATION: As multi-region, time-series and single-cell sequencing data become more widely available; it is becoming clear that certain tumors share evolutionary characteristics with others. In the last few years, several computational methods have been developed with the goal of inferring the subclonal composition and evolutionary history of tumors from tumor biopsy sequencing data. However, the phylogenetic trees that they report differ significantly between tumors (even those with similar characteristics). RESULTS: In this article, we present a novel combinatorial optimization method, CONETT, for detection of recurrent tumor evolution trajectories. Our method constructs a consensus tree of conserved evolutionary trajectories based on the information about temporal order of alteration events in a set of tumors. We apply our method to previously published datasets of 100 clear-cell renal cell carcinoma and 99 non-small-cell lung cancer patients and identify both conserved trajectories that were reported in the original studies, as well as new trajectories. AVAILABILITY AND IMPLEMENTATION: CONETT is implemented in C++ and available at https://github.com/ehodzic/CONETT. SUPPLEMENTARY INFORMATION: Supplementary data are available at Bioinformatics online..
O'Reilly, A.
Hughes, P.
Mann, J.
Lai, Z.
Teh, J.J.
Mclean, E.
Edmonds, K.
Lingard, K.
Chauhan, D.
Lynch, J.
Au, L.
Ludlow, A.
Pattison, N.
Wiseman, T.
Turajlic, S.
Gore, M.
Larkin, J.
Husson, O.
(2020). An immunotherapy survivor population: health-related quality of life and toxicity in patients with metastatic melanoma treated with immune checkpoint inhibitors. Support care cancer,
Vol.28
(2),
pp. 561-570.
show abstract
full text
PURPOSE: The immune checkpoint inhibitors (ICIs) have resulted in subgroups of patients with metastatic melanoma achieving high-quality durable responses. Metastatic melanoma survivors are a new population in the era of cancer survivorship. The aim of this study was to evaluate metastatic melanoma survivors in terms of health-related quality of life (HRQoL), immune-related adverse events (irAEs) and exposure to immunosuppressive agents in a large single centre in the UK. METHODS: We defined the survivor population as patients with a diagnosis of metastatic melanoma who achieved a durable response to an ICI and had been followed-up for a minimum of 12 months from initiation of ICI without disease progression. HRQoL was assessed using SF-36. Electronic health records were accessed to collect data on demographics, treatments, irAEs and survival. HRQoL data was compared with two norm-based datasets. RESULTS: Eighty-four metastatic melanoma survivors were eligible and 87% (N = 73) completed the SF-36. ICI-related toxicity of any grade occurred in 92% of patients and 43% had experienced a grade 3 or 4 toxicity. Almost half (49%) of the patients required steroids for the treatment of ICI-related toxicity, whilst 14% required treatment with an immunosuppressive agent beyond steroids. Melanoma survivors had statistically significant lower HRQoL scores with regard to physical, social and physical role functioning and general health compared with the normative population. There was a trend towards inferior scores in patients with previous exposure to ipilimumab compared with those never exposed to ipilimumab. CONCLUSIONS: Our results show that metastatic melanoma survivors have potentially experienced significant ICI-related toxicity and experience significant impairments in specific HRQoL domains. Future service planning is required to meet this population's unique survivorship needs..
Carbone, F.
Huang, J.K.
Perelli, L.
Poggetto, E.D.
Gutschner, T.
Tomihara, H.
Soeung, M.
Lam, T.N.
Xia, R.
Zhu, C.
Song, X.
Zhang, J.
Sircar, K.
Malouf, G.G.
Sgambato, A.
Karam, J.A.
Gao, J.
Jonasch, E.
Viale, A.
Draetta, G.F.
Futreal, A.
Bakouny, Z.
Van Allen, E.M.
Choueiri, T.
Msaouel, P.
Litchfield, K.
Turajlic, S.
Heffernan, T.P.
Chen, Y.B.
DiNatale, R.G.
Hakimi, A.A.
Bristow, C.A.
Tannir, N.M.
Carugo, A.
Genovese, G.
(2020). 9p21 Loss Defines the Evolutionary Patterns of Aggressive Renal Cell Carcinomas. ,
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show abstract
AbstractDedifferentiation and acquisition of chromosomal instability in renal cell carcinoma portends dismal prognosis and aggressive clinical behavior. However, the absence of reliable experimental models dramatically impacts the understanding of mechanisms underlying malignant progression. Here we established an in vivo genetic platform to rapidly generate somatic mosaic genetically engineerd immune-competent mouse models of renal tumors, recapitulating the genomic and phenotypic features of these malignancies. Leveraging somatic chromosomal engineering, we demonstrated that ablation of the murine locus syntenic to human 9p21 drives the rapid expansion of aggressive mesenchymal clones with prominent metastatic behavior, characterized by early emergence of chromosomal instability, whole-genome duplication, and conserved patterns of aneuploidy. This model of punctuated equilibrium provides a remarkable example of cross-species convergent evolution.SignificanceTo better understand the role of 9p21 in malignant progression, we generated a somatic mosaic GEMM of renal cancer, capturing the histological, genomic and evolutionary features of human disease. With this technology we demonstrated a critica role of 9p21 loss in metastatic evolution of RCC and provide a unique tool for testing new therapeutic treatments..
Litchfield, K.
Stanislaw, S.
Spain, L.
Gallegos, L.L.
Rowan, A.
Schnidrig, D.
Rosenbaum, H.
Harle, A.
Au, L.
Hill, S.M.
Tippu, Z.
Thomas, J.
Thompson, L.
Xu, H.
Horswell, S.
Barhoumi, A.
Jones, C.
Leith, K.F.
Burgess, D.L.
Watkins, T.B.
Lim, E.
Birkbak, N.J.
Lamy, P.
Nordentoft, I.
Dyrskjøt, L.
Pickering, L.
Hazell, S.
Jamal-Hanjani, M.
PEACE Consortium,
Larkin, J.
Swanton, C.
Alexander, N.R.
Turajlic, S.
(2020). Representative Sequencing: Unbiased Sampling of Solid Tumor Tissue. Cell rep,
Vol.31
(5),
p. 107550.
show abstract
full text
Although thousands of solid tumors have been sequenced to date, a fundamental under-sampling bias is inherent in current methodologies. This is caused by a tissue sample input of fixed dimensions (e.g., 6 mm biopsy), which becomes grossly under-powered as tumor volume scales. Here, we demonstrate representative sequencing (Rep-Seq) as a new method to achieve unbiased tumor tissue sampling. Rep-Seq uses fixed residual tumor material, which is homogenized and subjected to next-generation sequencing. Analysis of intratumor tumor mutation burden (TMB) variability shows a high level of misclassification using current single-biopsy methods, with 20% of lung and 52% of bladder tumors having at least one biopsy with high TMB but low clonal TMB overall. Misclassification rates by contrast are reduced to 2% (lung) and 4% (bladder) when a more representative sampling methodology is used. Rep-Seq offers an improved sampling protocol for tumor profiling, with significant potential for improved clinical utility and more accurate deconvolution of clonal structure..
Powell, N.
Ibraheim, H.
Raine, T.
Speight, R.A.
Papa, S.
Brain, O.
Green, M.
Samaan, M.A.
Spain, L.
Yousaf, N.
Hunter, N.
Eldridge, L.
Pavlidis, P.
Irving, P.
Hayee, B.
Turajlic, S.
Larkin, J.
Lindsay, J.O.
Gore, M.
(2020). British Society of Gastroenterology endorsed guidance for the management of immune checkpoint inhibitor-induced enterocolitis. Lancet gastroenterol hepatol,
Vol.5
(7),
pp. 679-697.
show abstract
full text
Immune checkpoint inhibitors are a novel class of cancer treatment that have improved outcomes for a subset of cancer patients. They work by antagonising inhibitory immune pathways, thereby augmenting immune-mediated antitumour responses. However, immune activation is not cancer-specific and often results in the activation of immune cells in non-cancer tissues, resulting in off-target immune-mediated injury and organ dysfunction. Diarrhoea and gastrointestinal tract inflammation are common and sometimes serious side-effects of this type of therapy. Prompt recognition of gastrointestinal toxicity and, in many cases, rapid institution of anti-inflammatory or biologic therapy (or both) is required to reverse these complications. Management of organ-specific complications benefits from multidisciplinary input, including engagement with gastroenterologists for optimal management of immune checkpoint inhibitor-induced enterocolitis. In this British Society of Gastroenterology endorsed guidance document, we have developed a consensus framework for the investigation and management of immune checkpoint inhibitor-induced enterocolitis..
Favara, D.M.
Spain, L.
Au, L.
Clark, J.
Daniels, E.
Diem, S.
Chauhan, D.
Turajlic, S.
Powell, N.
Larkin, J.M.
Yousaf, N.
(2020). Five-year review of corticosteroid duration and complications in the management of immune checkpoint inhibitor-related diarrhoea and colitis in advanced melanoma. Esmo open,
Vol.5
(4).
show abstract
full text
BACKGROUND: Immune-related diarrhoea/colitis (ir-D/C) is a common adverse event of immune checkpoint inhibitor (ICI) therapy. Guidelines recommend corticosteroid (CS) treatment; however, the average treatment duration for ir-D/C remains poorly defined. METHODS: All advanced melanoma patients treated with ICI therapy at the Royal Marsden Hospital between 2011 and 2016 were reviewed to identify ir-D/C cases alongside clinical variables. RESULTS: 117 any-grade ir-D/C episodes occurred in 109 (21%) patients out of a total of 519 patients treated (ipilimumab=77 episodes, anti-PD1=17 (nivolumab or pembrolizumab), ipilimumab and nivolumab=23 (ipi+nivo)) (seven patients had ir-D/C more than once on different lines of treatment) and >/=grade 3 ir-D/C occurred most frequently (63/519 patients (12%) vs 29/519 (5%) grade 1, and 25/519 (5%) grade 2). Median onset (days) of all-grade ir-D/C after starting ICI therapy was 41 for ipilimumab (IQR 24 to 59, n=77), 91 for anti-PD1 (IQR 46 to 355, n=17) and 45 for ipi+nivo (IQR 24 to 67, n=23). In 71/117 (61%) patients, ir-D/C episodes were treated with CS (17% grade 2; 79% grade 3/4): 54 being steroid-responsive; 17 being steroid-refractory and received additional anti-tumor necrosis factor (TNF) treatment. Median grade 3 ir-D/C CS duration was similar across treatments, averaging 58 days. Median overall CS duration (days) was longer in the grade 3/4 D/C steroid-refractory group (94 vs 45 days). Infection developed in 11/71 (15%) CS recipients and in 6/17 (35%) anti-TNF recipients. In 65/117 (55%) patients, ir-D/C episodes were investigated with flexible sigmoidoscopy. Of these patients, 38/65 (58%) had macroscopic colitis and 12/65 (18%) had microscopic colitis. The steroid-refractory group had more macroscopic changes, 13/17 (76%), than the steroid-responsive group, 22/41 (54%). CONCLUSION: Rates of grade 3 ir-D/C were higher than reported in clinical trials. The 58-day median duration of CS therapy for grade 3 ir-D/C places a significant number of patients at risk of complications. We demonstrate that microscopic colitis is an important subgroup, advocating biopsies in ir-D/C even with macroscopically normal bowel..
Hodzic, E.
Shrestha, R.
Malikic, S.
Collins, C.C.
Litchfield, K.
Turajlic, S.
Sahinalp, S.C.
(2020). Identification of Conserved Evolutionary Trajectories in Tumors. ,
.
show abstract
AbstractMotivationAs multi-region, time-series, and single cell sequencing data become more widely available, it is becoming clear that certain tumors share evolutionary characteristics with others. In the last few years, several computational methods have been developed with the goal of inferring the subclonal composition and evolutionary history of tumors from tumor biopsy sequencing data. However, the phylogenetic trees that they report differ significantly between tumors (even those with similar characteristics).ResultsIn this paper, we present a novel combinatorial optimization method, CONETT, for detection of recurrent tumor evolution trajectories. Our method constructs a consensus tree of conserved evolutionary trajectories based on the information about temporal order of alteration events in a set of tumors. We apply our method to previously published datasets of 100 clear-cell renal cell carcinoma and 99 non-small-cell lung cancer patients and identify both conserved trajectories that were reported in the original studies, as well as new trajectories.AvailabilityCONETT is implemented in C++ and available at https://github.com/ehodzic/CONETT..
Comito, F.
Leslie, I.
Boos, L.
Furness, A.
Pickering, L.
Turajlic, S.
Larkin, J.
(2020). Oligoprogression After Checkpoint Inhibition in Metastatic Melanoma Treated With Locoregional Therapy: A Single-center Retrospective Analysis. J immunother,
Vol.43
(8),
pp. 250-255.
show abstract
Checkpoint inhibitors (CPIs) have demonstrated a heterogenous spectrum of response and disease progression that may not be fully captured by conventional response criteria, such as a limited degree of progression, known as oligoprogression, which could benefit from local treatment. We retrospectively analyzed data from all patients diagnosed with metastatic melanoma, who received CPI between January 2006 and March 2018 at Royal Marsden. We enrolled 36 patients who experienced progression in a maximum of 3 metastatic sites, after achieving disease control from therapy with CPI, and were radically treated with the locoregional approach. We carried out Kaplan-Meier analysis to obtain progression free-survival post-first oligoprogression (PFS-PO1), overall survival (OS) post-first oligoprogression, and OS estimates. The median time to oligoprogression from the start of CPI was 12 months. At a median follow-up of 34 months, the median PFS-PO1 was 32 months, with 50% of patients not progressed at the time of the data cutoff. The median OS-post-first oligoprogression was not reached. At a median follow-up of 52 months (from the first cycle of CPI), the median OS was not reached, with 75% of patients alive at the time of analysis. Univariate and multivariate analyses demonstrated that baseline American Joint Committee on Cancer stage IV M1a or M1b is associated with a longer PFS-PO1 compared with stage M1c or M1d. We observed that local therapy for oligoprogression after CPI can result in durable disease control, suggesting that locoregional treatment should be considered in patients being treated with immunotherapy. However, prospective evaluation, perhaps in randomized trials, is needed..
Watkins, T.B.
Lim, E.L.
Petkovic, M.
Elizalde, S.
Birkbak, N.J.
Wilson, G.A.
Moore, D.A.
Grönroos, E.
Rowan, A.
Dewhurst, S.M.
Demeulemeester, J.
Dentro, S.C.
Horswell, S.
Au, L.
Haase, K.
Escudero, M.
Rosenthal, R.
Bakir, M.A.
Xu, H.
Litchfield, K.
Lu, W.T.
Mourikis, T.P.
Dietzen, M.
Spain, L.
Cresswell, G.D.
Biswas, D.
Lamy, P.
Nordentoft, I.
Harbst, K.
Castro-Giner, F.
Yates, L.R.
Caramia, F.
Jaulin, F.
Vicier, C.
Tomlinson, I.P.
Brastianos, P.K.
Cho, R.J.
Bastian, B.C.
Dyrskjøt, L.
Jönsson, G.B.
Savas, P.
Loi, S.
Campbell, P.J.
Andre, F.
Luscombe, N.M.
Steeghs, N.
Tjan-Heijnen, V.C.
Szallasi, Z.
Turajlic, S.
Jamal-Hanjani, M.
Van Loo, P.
Bakhoum, S.F.
Schwarz, R.F.
McGranahan, N.
Swanton, C.
(2020). Pervasive chromosomal instability and karyotype order in tumour evolution. Nature,
Vol.587
(7832),
pp. 126-132.
show abstract
full text
Chromosomal instability in cancer consists of dynamic changes to the number and structure of chromosomes1,2. The resulting diversity in somatic copy number alterations (SCNAs) may provide the variation necessary for tumour evolution1,3,4. Here we use multi-sample phasing and SCNA analysis of 1,421 samples from 394 tumours across 22 tumour types to show that continuous chromosomal instability results in pervasive SCNA heterogeneity. Parallel evolutionary events, which cause disruption in the same genes (such as BCL9, MCL1, ARNT (also known as HIF1B), TERT and MYC) within separate subclones, were present in 37% of tumours. Most recurrent losses probably occurred before whole-genome doubling, that was found as a clonal event in 49% of tumours. However, loss of heterozygosity at the human leukocyte antigen (HLA) locus and loss of chromosome 8p to a single haploid copy recurred at substantial subclonal frequencies, even in tumours with whole-genome doubling, indicating ongoing karyotype remodelling. Focal amplifications that affected chromosomes 1q21 (which encompasses BCL9, MCL1 and ARNT), 5p15.33 (TERT), 11q13.3 (CCND1), 19q12 (CCNE1) and 8q24.1 (MYC) were frequently subclonal yet appeared to be clonal within single samples. Analysis of an independent series of 1,024 metastatic samples revealed that 13 focal SCNAs were enriched in metastatic samples, including gains in chromosome 8q24.1 (encompassing MYC) in clear cell renal cell carcinoma and chromosome 11q13.3 (encompassing CCND1) in HER2+ breast cancer. Chromosomal instability may enable the continuous selection of SCNAs, which are established as ordered events that often occur in parallel, throughout tumour evolution..
Litchfield, K.
Reading, J.L.
Lim, E.L.
Xu, H.
Liu, P.
Al-Bakir, M.
Wong, Y.N.
Rowan, A.
Funt, S.A.
Merghoub, T.
Perkins, D.
Lauss, M.
Svane, I.M.
Jönsson, G.
Herrero, J.
Larkin, J.
Quezada, S.A.
Hellmann, M.D.
Turajlic, S.
Swanton, C.
(2020). Escape from nonsense-mediated decay associates with anti-tumor immunogenicity. Nat commun,
Vol.11
(1),
p. 3800.
show abstract
full text
Frameshift insertion/deletions (fs-indels) are an infrequent but highly immunogenic mutation subtype. Although fs-indels are degraded through the nonsense-mediated decay (NMD) pathway, we hypothesise that some fs-indels escape degradation and elicit anti-tumor immune responses. Using allele-specific expression analysis, expressed fs-indels are enriched in genomic positions predicted to escape NMD, and associated with higher protein expression, consistent with degradation escape (NMD-escape). Across four independent melanoma cohorts, NMD-escape mutations are significantly associated with clinical-benefit to checkpoint inhibitor (CPI) therapy (Pmeta = 0.0039). NMD-escape mutations are additionally found to associate with clinical-benefit in the low-TMB setting. Furthermore, in an adoptive cell therapy treated melanoma cohort, NMD-escape mutation count is the most significant biomarker associated with clinical-benefit. Analysis of functional T cell reactivity screens from personalized vaccine studies shows direct evidence of fs-indel derived neoantigens eliciting immune response, particularly those with highly elongated neo open reading frames. NMD-escape fs-indels represent an attractive target for biomarker optimisation and immunotherapy design..
Zhao, Y.
Fu, X.
Lopez, J.
Rowan, A.
Au, L.
Fendler, A.
Hazell, S.
Xu, H.
Horswell, S.
Shephard, S.
Spain, L.
Byrne, F.
Stamp, G.
O'Brien, T.
Nicol, D.
Augustine, M.
Chandra, A.
Rudman, S.
Toncheva, A.
Pickering, L.
Larkin, J.
Sahai, E.
Bates, P.
Swanton, C.
Turajlic, S.
Litchfield, K.
(2020). Selection of metastasis competent subclones in the tumour interior: TRACERx renal. ,
.
show abstract
Abstract
While the genetic evolutionary features of solid tumour growth are becoming increasingly described, the spatial and physical nature of subclonal growth remains unclear. Here we utilise 102 macroscopic whole tumour images from clear cell renal cell carcinoma (ccRCC) patients, with matched genetic and phenotypic data from 756 biopsies. Utilising a digital image processing pipeline the boundaries between tumour and normal tissue were marked by a renal pathologist, and positions of boundary line and biopsy regions were extracted to X- and Y-coordinates. The coordinates were then integrated with genomic data to map exact spatial subclone locations, revealing how genetically distinct subclones grow and evolve spatially. A phenotype of advanced and more aggressive subclonal growth was present in the tumour centre, characterised by an elevated burden of somatic copy number alterations, higher necrosis, proliferation rate and Fuhrman grade. Moreover, metastasising subclones were found to preferentially originate from the tumour centre. Collectively these observations suggest a model of accelerated evolution in the tumour interior, with harsh hypoxic environmental conditions leading to heightened cellular turnover and greater opportunity for driver SCNAs to arise and expand due to selective advantage. Tumour subclone growth was found to be predominantly spatially contiguous in nature, with subclone dispersal a rare event found in two cases, which notably was associated with metastasis. In terms of genetic events, the largest subclones spatially were dominated by driver somatic copy number alterations, suggesting a large selective advantage can be conferred to subclones upon acquisition of these alterations. In conclusion, spatial dynamics is strongly associated with genomic alterations and plays an important role in tumour evolution..
Shepherd, S.T.
Litchfield, K.
Turajlic, S.
(2019). Searching for the needle in the haystack: deconvoluting the evolutionary dynamics of residual disease in human glioblastoma. Ann oncol,
Vol.30
(3),
pp. 355-357.
Turajlic, S.
(2019). Standing on the shoulders of giants. Nat med,
Vol.25
(3),
p. 357.
Turajlic, S.
Sottoriva, A.
Graham, T.
Swanton, C.
(2019). Resolving genetic heterogeneity in cancer. Nat rev genet,
Vol.20
(7),
pp. 404-416.
show abstract
full text
To a large extent, cancer conforms to evolutionary rules defined by the rates at which clones mutate, adapt and grow. Next-generation sequencing has provided a snapshot of the genetic landscape of most cancer types, and cancer genomics approaches are driving new insights into cancer evolutionary patterns in time and space. In contrast to species evolution, cancer is a particular case owing to the vast size of tumour cell populations, chromosomal instability and its potential for phenotypic plasticity. Nevertheless, an evolutionary framework is a powerful aid to understand cancer progression and therapy failure. Indeed, such a framework could be applied to predict individual tumour behaviour and support treatment strategies..
Amaria, R.N.
Menzies, A.M.
Burton, E.M.
Scolyer, R.A.
Tetzlaff, M.T.
Antdbacka, R.
Ariyan, C.
Bassett, R.
Carter, B.
Daud, A.
Faries, M.
Fecher, L.A.
Flaherty, K.T.
Gershenwald, J.E.
Hamid, O.
Hong, A.
Kirkwood, J.M.
Lo, S.
Margolin, K.
Messina, J.
Postow, M.A.
Rizos, H.
Ross, M.I.
Rozeman, E.A.
Saw, R.P.
Sondak, V.
Sullivan, R.J.
Taube, J.M.
Thompson, J.F.
van de Wiel, B.A.
Eggermont, A.M.
Davies, M.A.
International Neoadjuvant Melanoma Consortium members,
Ascierto, P.A.
Spillane, A.J.
van Akkooi, A.C.
Wargo, J.A.
Blank, C.U.
Tawbi, H.A.
Long, G.V.
(2019). Neoadjuvant systemic therapy in melanoma: recommendations of the International Neoadjuvant Melanoma Consortium. Lancet oncol,
Vol.20
(7),
pp. e378-e389.
show abstract
Advances in the treatment of metastatic melanoma have improved responses and survival. However, many patients continue to experience resistance or toxicity to treatment, highlighting a crucial need to identify biomarkers and understand mechanisms of response and toxicity. Neoadjuvant therapy for regional metastases might improve operability and clinical outcomes over upfront surgery and adjuvant therapy, and has become an established role for drug development and biomarker discovery in other cancers (including locally advanced breast cancer, head and neck squamous cell carcinomas, gastroesophageal cancer, and anal cancer). Patients with clinically detectable stage III melanoma are ideal candidates for neoadjuvant therapy, because they represent a high-risk patient population with poor outcomes when treated with upfront surgery alone. Neoadjuvant therapy is now an active area of research for melanoma with numerous completed and ongoing trials (since 2014) with disparate designs, endpoints, and analyses under investigation. We have, therefore, established the International Neoadjuvant Melanoma Consortium with experts in medical oncology, surgical oncology, pathology, radiation oncology, radiology, and translational research to develop recommendations for investigating neoadjuvant therapy in melanoma to align future trial designs and correlative analyses. Alignment and consistency of neoadjuvant trials will facilitate optimal data organisation for future regulatory review and strengthen translational research across the melanoma disease continuum..
Ferronika, P.
Hof, J.
Kats-Ugurlu, G.
Sijmons, R.H.
Terpstra, M.M.
de Lange, K.
Leliveld-Kors, A.
Westers, H.
Kok, K.
(2019). Comprehensive Profiling of Primary and Metastatic ccRCC Reveals a High Homology of the Metastases to a Subregion of the Primary Tumour. Cancers (basel),
Vol.11
(6).
show abstract
While intratumour genetic heterogeneity of primary clear cell renal cell carcinoma (ccRCC) is well characterized, the genomic profiles of metastatic ccRCCs are seldom studied. We profiled the genomes and transcriptomes of a primary tumour and matched metastases to better understand the evolutionary processes that lead to metastasis. In one ccRCC patient, four regions of the primary tumour, one region of the thrombus in the inferior vena cava, and four lung metastases (including one taken after pegylated (PEG)-interferon therapy) were analysed separately. Each sample was analysed for copy number alterations and somatic mutations by whole exome sequencing. We also evaluated gene expression profiles for this patient and 15 primary tumour and 15 metastasis samples from four additional patients. Copy number profiles of the index patient showed two distinct subgroups: one consisted of three primary tumours with relatively minor copy number changes, the other of a primary tumour, the thrombus, and the lung metastases, all with a similar copy number pattern and tetraploid-like characteristics. Somatic mutation profiles indicated parallel clonal evolution with similar numbers of private mutations in each primary tumour and metastatic sample. Expression profiling of the five patients revealed significantly changed expression levels of 57 genes between primary tumours and metastases, with enrichment in the extracellular matrix cluster. The copy number profiles suggest a punctuated evolution from a subregion of the primary tumour. This process, which differentiated the metastases from the primary tumours, most likely occurred rapidly, possibly even before metastasis formation. The evolutionary patterns we deduced from the genomic alterations were also reflected in the gene expression profiles..
Schnidrig, D.
Turajlic, S.
Litchfield, K.
(2019). Tumour mutational burden: primary versus metastatic tissue creates systematic bias. Immuno-oncology technology,
Vol.4,
pp. 8-14.
Spain, L.
Tippu, Z.
Larkin, J.M.
Carr, A.
Turajlic, S.
(2019). How we treat neurological toxicity from immune checkpoint inhibitors. Esmo open,
Vol.4
(Suppl 4),
p. e000540.
show abstract
full text
Neurological adverse events from immune checkpoint inhibition are increasingly recognised, especially with combination anti-cytotoxic T-lymphocyte antigen 4 (CTLA4) and anti-programmed death receptor 1 (anti-PD-1) therapies. Their presenting symptoms and signs are often subacute and highly variable, reflecting the numerous components of the nervous system. Given the risk of substantial morbidity and mortality, it is important to inform patients of symptoms that may be of concern, and to assess any suspected toxicity promptly. As with other immune-related adverse events, the cornerstone of management is administration of corticosteroids. Specialist neurology input is vital in this group of patients to guide appropriate investigations and tailor treatment strategies..
Turajlic, S.
Swanton, C.
Boshoff, C.
(2018). Kidney cancer: The next decade. J exp med,
Vol.215
(10),
pp. 2477-2479.
show abstract
Chris Boshoff, Senior Vice President of Immuno-Oncology, Translational and Early Development at Pfizer, and colleagues Samra Turajlic and Charles Swanton from the Francis Crick Institute and University College London give us their personal point of view on new insights and future therapeutic approaches for renal cancer..
Arce Vargas, F.
Furness, A.J.
Litchfield, K.
Joshi, K.
Rosenthal, R.
Ghorani, E.
Solomon, I.
Lesko, M.H.
Ruef, N.
Roddie, C.
Henry, J.Y.
Spain, L.
Ben Aissa, A.
Georgiou, A.
Wong, Y.N.
Smith, M.
Strauss, D.
Hayes, A.
Nicol, D.
O'Brien, T.
Mårtensson, L.
Ljungars, A.
Teige, I.
Frendéus, B.
TRACERx Melanoma,
TRACERx Renal,
TRACERx Lung consortia,
Pule, M.
Marafioti, T.
Gore, M.
Larkin, J.
Turajlic, S.
Swanton, C.
Peggs, K.S.
Quezada, S.A.
(2018). Fc Effector Function Contributes to the Activity of Human Anti-CTLA-4 Antibodies. Cancer cell,
Vol.33
(4),
pp. 649-663.e4.
show abstract
full text
With the use of a mouse model expressing human Fc-gamma receptors (FcγRs), we demonstrated that antibodies with isotypes equivalent to ipilimumab and tremelimumab mediate intra-tumoral regulatory T (Treg) cell depletion in vivo, increasing the CD8+ to Treg cell ratio and promoting tumor rejection. Antibodies with improved FcγR binding profiles drove superior anti-tumor responses and survival. In patients with advanced melanoma, response to ipilimumab was associated with the CD16a-V158F high affinity polymorphism. Such activity only appeared relevant in the context of inflamed tumors, explaining the modest response rates observed in the clinical setting. Our data suggest that the activity of anti-CTLA-4 in inflamed tumors may be improved through enhancement of FcγR binding, whereas poorly infiltrated tumors will likely require combination approaches..
Tio, M.
Rai, R.
Ezeoke, O.M.
McQuade, J.L.
Zimmer, L.
Khoo, C.
Park, J.J.
Spain, L.
Turajlic, S.
Ardolino, L.
Yip, D.
Goldinger, S.M.
Cohen, J.V.
Millward, M.
Atkinson, V.
Kane, A.Y.
Ascierto, P.A.
Garbe, C.
Gutzmer, R.
Johnson, D.B.
Rizvi, H.A.
Joshua, A.M.
Hellmann, M.D.
Long, G.V.
Menzies, A.M.
(2018). Anti-PD-1/PD-L1 immunotherapy in patients with solid organ transplant, HIV or hepatitis B/C infection. Eur j cancer,
Vol.104,
pp. 137-144.
show abstract
full text
BACKGROUND: Anti-programmed cell death protein 1/programmed death ligand 1 (PD-1/PD-L1) immunotherapy is now routinely used to treat several cancers. Clinical trials have excluded several populations, including patients with solid organ transplant, HIV infection and hepatitis B/C infection. We examined the safety outcomes of these populations treated with anti-PD-1/PD-L1 treatment in a multicentre retrospective study. METHODS: Patients from 16 centres with advanced cancer and solid organ transplant, HIV infection or hepatitis B/C infection were included. Demographic, tumour, treatment, toxicity and outcome data were recorded. RESULTS: Forty-six patients were included for analysis, with a median age of 60 years, and the majority of patients diagnosed with melanoma (72%). Among six patients with solid organ transplants, two graft rejections occurred, with one resulting in death, whereas two patients achieved partial responses. There were four responses in 12 patients with HIV infection. In 14 patients with hepatitis B, there were three responses, and similarly, there were three responses in 14 patients with hepatitis C. There was no unexpected toxicity in any viral infection group or an increase in viral load. CONCLUSION: Patients with HIV or hepatitis B/C infections treated with anti-PD-1/PD-L1 immunotherapy may respond to treatment without increased toxicity. Given the risk of graft rejection in solid organ transplant patients and also the potential for response, the role of anti-PD-1/PD-L1 immunotherapy needs to be carefully considered..
Turajlic, S.
Xu, H.
Litchfield, K.
Rowan, A.
Horswell, S.
Chambers, T.
O'Brien, T.
Lopez, J.I.
Watkins, T.B.
Nicol, D.
Stares, M.
Challacombe, B.
Hazell, S.
Chandra, A.
Mitchell, T.J.
Au, L.
Eichler-Jonsson, C.
Jabbar, F.
Soultati, A.
Chowdhury, S.
Rudman, S.
Lynch, J.
Fernando, A.
Stamp, G.
Nye, E.
Stewart, A.
Xing, W.
Smith, J.C.
Escudero, M.
Huffman, A.
Matthews, N.
Elgar, G.
Phillimore, B.
Costa, M.
Begum, S.
Ward, S.
Salm, M.
Boeing, S.
Fisher, R.
Spain, L.
Navas, C.
Grönroos, E.
Hobor, S.
Sharma, S.
Aurangzeb, I.
Lall, S.
Polson, A.
Varia, M.
Horsfield, C.
Fotiadis, N.
Pickering, L.
Schwarz, R.F.
Silva, B.
Herrero, J.
Luscombe, N.M.
Jamal-Hanjani, M.
Rosenthal, R.
Birkbak, N.J.
Wilson, G.A.
Pipek, O.
Ribli, D.
Krzystanek, M.
Csabai, I.
Szallasi, Z.
Gore, M.
McGranahan, N.
Van Loo, P.
Campbell, P.
Larkin, J.
Swanton, C.
TRACERx Renal Consortium,
(2018). Deterministic Evolutionary Trajectories Influence Primary Tumor Growth: TRACERx Renal. Cell,
Vol.173
(3),
pp. 595-610.e11.
show abstract
full text
The evolutionary features of clear-cell renal cell carcinoma (ccRCC) have not been systematically studied to date. We analyzed 1,206 primary tumor regions from 101 patients recruited into the multi-center prospective study, TRACERx Renal. We observe up to 30 driver events per tumor and show that subclonal diversification is associated with known prognostic parameters. By resolving the patterns of driver event ordering, co-occurrence, and mutual exclusivity at clone level, we show the deterministic nature of clonal evolution. ccRCC can be grouped into seven evolutionary subtypes, ranging from tumors characterized by early fixation of multiple mutational and copy number drivers and rapid metastases to highly branched tumors with >10 subclonal drivers and extensive parallel evolution associated with attenuated progression. We identify genetic diversity and chromosomal complexity as determinants of patient outcome. Our insights reconcile the variable clinical behavior of ccRCC and suggest evolutionary potential as a biomarker for both intervention and surveillance..
Turajlic, S.
Xu, H.
Litchfield, K.
Rowan, A.
Chambers, T.
Lopez, J.I.
Nicol, D.
O'Brien, T.
Larkin, J.
Horswell, S.
Stares, M.
Au, L.
Jamal-Hanjani, M.
Challacombe, B.
Chandra, A.
Hazell, S.
Eichler-Jonsson, C.
Soultati, A.
Chowdhury, S.
Rudman, S.
Lynch, J.
Fernando, A.
Stamp, G.
Nye, E.
Jabbar, F.
Spain, L.
Lall, S.
Guarch, R.
Falzon, M.
Proctor, I.
Pickering, L.
Gore, M.
Watkins, T.B.
Ward, S.
Stewart, A.
DiNatale, R.
Becerra, M.F.
Reznik, E.
Hsieh, J.J.
Richmond, T.A.
Mayhew, G.F.
Hill, S.M.
McNally, C.D.
Jones, C.
Rosenbaum, H.
Stanislaw, S.
Burgess, D.L.
Alexander, N.R.
Swanton, C.
PEACE,
TRACERx Renal Consortium,
(2018). Tracking Cancer Evolution Reveals Constrained Routes to Metastases: TRACERx Renal. Cell,
Vol.173
(3),
pp. 581-594.e12.
show abstract
full text
Clear-cell renal cell carcinoma (ccRCC) exhibits a broad range of metastatic phenotypes that have not been systematically studied to date. Here, we analyzed 575 primary and 335 metastatic biopsies across 100 patients with metastatic ccRCC, including two cases sampledat post-mortem. Metastatic competence was afforded by chromosome complexity, and we identify 9p loss as a highly selected event driving metastasis and ccRCC-related mortality (p = 0.0014). Distinct patterns of metastatic dissemination were observed, including rapid progression to multiple tissue sites seeded by primary tumors of monoclonal structure. By contrast, we observed attenuated progression in cases characterized by high primary tumor heterogeneity, with metastatic competence acquired gradually and initial progression to solitary metastasis. Finally, we observed early divergence of primitive ancestral clones and protracted latency of up to two decades as a feature of pancreatic metastases..
Mitchell, T.J.
Turajlic, S.
Rowan, A.
Nicol, D.
Farmery, J.H.
O'Brien, T.
Martincorena, I.
Tarpey, P.
Angelopoulos, N.
Yates, L.R.
Butler, A.P.
Raine, K.
Stewart, G.D.
Challacombe, B.
Fernando, A.
Lopez, J.I.
Hazell, S.
Chandra, A.
Chowdhury, S.
Rudman, S.
Soultati, A.
Stamp, G.
Fotiadis, N.
Pickering, L.
Au, L.
Spain, L.
Lynch, J.
Stares, M.
Teague, J.
Maura, F.
Wedge, D.C.
Horswell, S.
Chambers, T.
Litchfield, K.
Xu, H.
Stewart, A.
Elaidi, R.
Oudard, S.
McGranahan, N.
Csabai, I.
Gore, M.
Futreal, P.A.
Larkin, J.
Lynch, A.G.
Szallasi, Z.
Swanton, C.
Campbell, P.J.
TRACERx Renal Consortium,
(2018). Timing the Landmark Events in the Evolution of Clear Cell Renal Cell Cancer: TRACERx Renal. Cell,
Vol.173
(3),
pp. 611-623.e17.
show abstract
full text
Clear cell renal cell carcinoma (ccRCC) is characterized by near-universal loss of the short arm of chromosome 3, deleting several tumor suppressor genes. We analyzed whole genomes from 95 biopsies across 33 patients with clear cell renal cell carcinoma. We find hotspots of point mutations in the 5' UTR of TERT, targeting a MYC-MAX-MAD1 repressor associated with telomere lengthening. The most common structural abnormality generates simultaneous 3p loss and 5q gain (36% patients), typically through chromothripsis. This event occurs in childhood or adolescence, generally as the initiating event that precedes emergence of the tumor's most recent common ancestor by years to decades. Similar genomic changes drive inherited ccRCC. Modeling differences in age incidence between inherited and sporadic cancers suggests that the number of cells with 3p loss capable of initiating sporadic tumors is no more than a few hundred. Early development of ccRCC follows well-defined evolutionary trajectories, offering opportunity for early intervention..
Lim, K.H.
Spain, L.
Barker, C.
Georgiou, A.
Walls, G.
Gore, M.
Turajlic, S.
Board, R.
Larkin, J.M.
Lorigan, P.
(2018). Contemporary outcomes from the use of regular imaging to detect relapse in high-risk cutaneous melanoma. Esmo open,
Vol.3
(2),
p. e000317.
show abstract
full text
BACKGROUND: Agreement on the utility of imaging follow-up in patients with high-risk melanoma is lacking. A UK consensus statement recommends a surveillance schedule of CT or positron-emission tomography-CT and MRI brain (every 6 months for 3 years, then annually in years 4 and 5) as well as clinical examination for high-risk resected Stages II and III cutaneous melanoma. Our aim was to assess patterns of relapse and whether imaging surveillance could be of clinical benefit. PATIENTS AND METHODS: A retrospective study of patients enrolled between July 2013 and June 2015 from three UK tertiary cancer centres followed-up according to this protocol was undertaken. We evaluated time-to-recurrence (TTR), recurrence-free survival (RFS), method of detection and characteristics of recurrence, treatment received and overall survival (OS). RESULTS: A total of 173 patients were included. Most (79%) had treated Stages IIIB and IIIC disease. With a median follow-up of 23.3 months, 82 patients (47%) had relapsed. Median TTR was 10.1 months and median RFS was 21.2 months. The majority of recurrences (66%) were asymptomatic and detected by scheduled surveillance scan. Fifty-six (68%) patients recurred with Stage IV disease, with a median OS of 25.3 months; 26 (31.7%) patients had a locoregional recurrence, median OS not reached (P=0.016). Patients who underwent surgery at recurrence for either Stage III (27%) or IV (18%) disease did not reach their median OS. The median OS for the 33 patients (40%) who received systemic therapy was 12.9 months. CONCLUSION: Imaging appears to reliably detect subclinical disease and identify patients suitable for surgery, conferring favourable outcomes. The short median TTR provides rationale to intensify imaging schedule in the first year of surveillance. The poor OS of patients treated with systemic therapy probably reflects the relatively inferior treatment options during this time and requires further evaluation in the current era..
Flynn, M.
Pickering, L.
Larkin, J.
Turajlic, S.
(2018). Immune-checkpoint inhibitors in melanoma and kidney cancer: from sequencing to rational selection. Ther adv med oncol,
Vol.10,
p. 1758835918777427.
show abstract
full text
Immune-checkpoint inhibitors (ICPIs), including antibodies against cytotoxic T-lymphocyte associated antigen 4 and programmed cell death protein 1, have been shown to induce durable complete responses in a proportion of patients in the first-line and refractory setting in advanced melanoma and renal cell carcinoma. In fact, there are several lines of both targeted agents and ICPI that are now feasible treatment options. However, survival in the metastatic setting continues to be poor and there remains a need for improved therapeutic approaches. In order to enhance patient selection for the most appropriate next line of therapy, better predictive biomarkers of responsiveness will need to be developed in tandem with technologies to identify mechanisms of ICPI resistance. Adaptive, biomarker-driven trials will drive this evolution. The combination of ICPI with specific chemotherapies, targeted therapies and other immuno-oncology (IO) drugs in order to circumvent ICPI resistance and enhance efficacy is discussed. Recent data support the role for both targeted therapies and ICPI in the adjuvant setting of melanoma and targeted therapies in the adjuvant setting for renal cell carcinoma, which may influence the consideration of treatment on subsequent relapse. Approaches to select the optimal treatment sequences for these patients will need to be refined..
Spain, L.
Walls, G.
Messiou, C.
Turajlic, S.
Gore, M.
Larkin, J.
(2017). Efficacy and toxicity of rechallenge with combination immune checkpoint blockade in metastatic melanoma: a case series. Cancer immunol immunother,
Vol.66
(1),
pp. 113-117.
show abstract
full text
BACKGROUND: The efficacy and potential toxicity of rechallenge with combination ipilimumab and nivolumab has not been described. Retreatment of patients with immune checkpoint inhibitors in the setting of prior significant toxicity lacks evidence-based guidance. METHODS: We present the first three, consecutive patients who received re-treatment with combination ipilimumab and nivolumab for metastatic melanoma managed at our institution. RESULTS: Rechallenge with combination ipilimumab and nivolumab in the setting of prior grade 3 toxicity with initial combination therapy is feasible, and responses are seen. We highlight the fact that grade 3 toxicity is likely to recur, but if so, can be manageable. CONCLUSIONS: Retreatment with ipi + nivo may be considered an option in carefully selected, well-informed patients. More research is required to delineate the benefits and risks with this approach..
Jamal-Hanjani, M.
Wilson, G.A.
McGranahan, N.
Birkbak, N.J.
Watkins, T.B.
Veeriah, S.
Shafi, S.
Johnson, D.H.
Mitter, R.
Rosenthal, R.
Salm, M.
Horswell, S.
Escudero, M.
Matthews, N.
Rowan, A.
Chambers, T.
Moore, D.A.
Turajlic, S.
Xu, H.
Lee, S.-.
Forster, M.D.
Ahmad, T.
Hiley, C.T.
Abbosh, C.
Falzon, M.
Borg, E.
Marafioti, T.
Lawrence, D.
Hayward, M.
Kolvekar, S.
Panagiotopoulos, N.
Janes, S.M.
Thakrar, R.
Ahmed, A.
Blackhall, F.
Summers, Y.
Shah, R.
Joseph, L.
Quinn, A.M.
Crosbie, P.A.
Naidu, B.
Middleton, G.
Langman, G.
Trotter, S.
Nicolson, M.
Remmen, H.
Kerr, K.
Chetty, M.
Gomersall, L.
Fennell, D.A.
Nakas, A.
Rathinam, S.
Anand, G.
Khan, S.
Russell, P.
Ezhil, V.
Ismail, B.
Irvin-Sellers, M.
Prakash, V.
Lester, J.F.
Kornaszewska, M.
Attanoos, R.
Adams, H.
Davies, H.
Dentro, S.
Taniere, P.
O'Sullivan, B.
Lowe, H.L.
Hartley, J.A.
Iles, N.
Bell, H.
Ngai, Y.
Shaw, J.A.
Herrero, J.
Szallasi, Z.
Schwarz, R.F.
Stewart, A.
Quezada, S.A.
Le Quesne, J.
Van Loo, P.
Dive, C.
Hackshaw, A.
Swanton, C.
TRACERx Consortium,
(2017). Tracking the Evolution of Non-Small-Cell Lung Cancer. N engl j med,
Vol.376
(22),
pp. 2109-2121.
show abstract
full text
BACKGROUND: Among patients with non-small-cell lung cancer (NSCLC), data on intratumor heterogeneity and cancer genome evolution have been limited to small retrospective cohorts. We wanted to prospectively investigate intratumor heterogeneity in relation to clinical outcome and to determine the clonal nature of driver events and evolutionary processes in early-stage NSCLC. METHODS: In this prospective cohort study, we performed multiregion whole-exome sequencing on 100 early-stage NSCLC tumors that had been resected before systemic therapy. We sequenced and analyzed 327 tumor regions to define evolutionary histories, obtain a census of clonal and subclonal events, and assess the relationship between intratumor heterogeneity and recurrence-free survival. RESULTS: We observed widespread intratumor heterogeneity for both somatic copy-number alterations and mutations. Driver mutations in EGFR, MET, BRAF, and TP53 were almost always clonal. However, heterogeneous driver alterations that occurred later in evolution were found in more than 75% of the tumors and were common in PIK3CA and NF1 and in genes that are involved in chromatin modification and DNA damage response and repair. Genome doubling and ongoing dynamic chromosomal instability were associated with intratumor heterogeneity and resulted in parallel evolution of driver somatic copy-number alterations, including amplifications in CDK4, FOXA1, and BCL11A. Elevated copy-number heterogeneity was associated with an increased risk of recurrence or death (hazard ratio, 4.9; P=4.4×10-4), which remained significant in multivariate analysis. CONCLUSIONS: Intratumor heterogeneity mediated through chromosome instability was associated with an increased risk of recurrence or death, a finding that supports the potential value of chromosome instability as a prognostic predictor. (Funded by Cancer Research UK and others; TRACERx ClinicalTrials.gov number, NCT01888601 .)..
Spain, L.
Walls, G.
Julve, M.
O'Meara, K.
Schmid, T.
Kalaitzaki, E.
Turajlic, S.
Gore, M.
Rees, J.
Larkin, J.
(2017). Neurotoxicity from immune-checkpoint inhibition in the treatment of melanoma: a single centre experience and review of the literature. Ann oncol,
Vol.28
(2),
pp. 377-385.
show abstract
full text
BACKGROUND: Treatment with immune checkpoint inhibitors (ICPi) has greatly improved survival for patients with advanced melanoma in recent years. Anti-CTLA-4 and anti-PD1 antibodies have been approved following large Phase III trials. Immune-related neurological toxicity of varying severity has been reported in the literature. The cumulative incidence of neurotoxicity among ipilimumab, nivolumab and pembrolizumab is reported as <1% in published clinical trials. We aimed to identify the incidence of neurotoxicity in our institution across anti-CTLA4 and anti-PD-1 antibodies, including the combination of ipilimumab with nivolumab. We also review the existing literature and propose an investigation and management algorithm. METHODS: All patients with advanced melanoma treated with ipilimumab, nivolumab, pembrolizumab or the combination of ipilimumab and nivolumab (ipi + nivo), managed at the Royal Marsden Hospital between September 2010 and December 2015, including patients on (published) clinical trials were included. Medical records for each patient were reviewed and information on neurotoxicity recorded. A systematic search strategy was performed to collate existing reports of neurological toxicity. RESULTS: In total, 413 immunotherapy treatment episodes in 352 patients were included, with median follow-up of 26.7 months. Ten cases of neurotoxicity were recorded, affecting 2.8% of patients overall, ranging from grade 1 to 4, affecting both central and peripheral nervous systems. A rate of 14% was noted with ipi + nivo. Three of five patients commenced on corticosteroids responded to these. Six patients had made a full recovery at the time of reporting. A favorable radiological response was found in 7 of the 10 cases. Unusual presentations are described in detail. CONCLUSIONS: Neurological toxicity is not uncommon, and may be more frequent in patients treated with combination ipi + nivo. Patterns of presentation and response to treatment are varied. A prompt and considered approach is required to optimize outcomes in this group of patients..
Turajlic, S.
Swanton, C.
(2017). Implications of cancer evolution for drug development. Nat rev drug discov,
Vol.16
(7),
pp. 441-442.
full text
Turajlic, S.
Litchfield, K.
Xu, H.
Rosenthal, R.
McGranahan, N.
Reading, J.L.
Wong, Y.N.
Rowan, A.
Kanu, N.
Al Bakir, M.
Chambers, T.
Salgado, R.
Savas, P.
Loi, S.
Birkbak, N.J.
Sansregret, L.
Gore, M.
Larkin, J.
Quezada, S.A.
Swanton, C.
(2017). Insertion-and-deletion-derived tumour-specific neoantigens and the immunogenic phenotype: a pan-cancer analysis. Lancet oncol,
Vol.18
(8),
pp. 1009-1021.
show abstract
full text
BACKGROUND: The focus of tumour-specific antigen analyses has been on single nucleotide variants (SNVs), with the contribution of small insertions and deletions (indels) less well characterised. We investigated whether the frameshift nature of indel mutations, which create novel open reading frames and a large quantity of mutagenic peptides highly distinct from self, might contribute to the immunogenic phenotype. METHODS: We analysed whole-exome sequencing data from 5777 solid tumours, spanning 19 cancer types from The Cancer Genome Atlas. We compared the proportion and number of indels across the cohort, with a subset of results replicated in two independent datasets. We assessed in-silico tumour-specific neoantigen predictions by mutation type with pan-cancer analysis, together with RNAseq profiling in renal clear cell carcinoma cases (n=392), to compare immune gene expression across patient subgroups. Associations between indel burden and treatment response were assessed across four checkpoint inhibitor datasets. FINDINGS: We observed renal cell carcinomas to have the highest proportion (0·12) and number of indel mutations across the pan-cancer cohort (p<2·2 × 10-16), more than double the median proportion of indel mutations in all other cancer types examined. Analysis of tumour-specific neoantigens showed that enrichment of indel mutations for high-affinity binders was three times that of non-synonymous SNV mutations. Furthermore, neoantigens derived from indel mutations were nine times enriched for mutant specific binding, as compared with non-synonymous SNV derived neoantigens. Immune gene expression analysis in the renal clear cell carcinoma cohort showed that the presence of mutant-specific neoantigens was associated with upregulation of antigen presentation genes, which correlated (r=0·78) with T-cell activation as measured by CD8-positive expression. Finally, analysis of checkpoint inhibitor response data revealed frameshift indel count to be significantly associated with checkpoint inhibitor response across three separate melanoma cohorts (p=4·7 × 10-4). INTERPRETATION: Renal cell carcinomas have the highest pan-cancer proportion and number of indel mutations. Evidence suggests indels are a highly immunogenic mutational class, which can trigger an increased abundance of neoantigens and greater mutant-binding specificity. FUNDING: Cancer Research UK, UK National Institute for Health Research (NIHR) at the Royal Marsden Hospital National Health Service Foundation Trust, Institute of Cancer Research and University College London Hospitals Biomedical Research Centres, the UK Medical Research Council, the Rosetrees Trust, Novo Nordisk Foundation, the Prostate Cancer Foundation, the Breast Cancer Research Foundation, the European Research Council..
Litchfield, K.
Turajlic, S.
Swanton, C.
(2017). The GENIE Is Out of the Bottle: Landmark Cancer Genomics Dataset Released. Cancer discov,
Vol.7
(8),
pp. 796-798.
show abstract
full text
In this issue of Cancer Discovery, an overview of the AACR Project GENIE, a landmark study in cancer genomics, is presented by The AACR Project GENIE Consortium. A summary of the goals and objectives of this ambitious program is provided, together with an analysis of the phase I cohort of 19,000 samples. Cancer Discov; 7(8); 796-8. ©2017 AACR.See related article by The AACR Project GENIE Consortium, p. 818..
TRACERx Renal consortium,
(2017). TRACERx Renal: tracking renal cancer evolution through therapy. Nat rev urol,
Vol.14
(10),
pp. 575-576.
Arce Vargas, F.
Furness, A.J.
Solomon, I.
Joshi, K.
Mekkaoui, L.
Lesko, M.H.
Miranda Rota, E.
Dahan, R.
Georgiou, A.
Sledzinska, A.
Ben Aissa, A.
Franz, D.
Werner Sunderland, M.
Wong, Y.N.
Henry, J.Y.
O'Brien, T.
Nicol, D.
Challacombe, B.
Beers, S.A.
Melanoma TRACERx Consortium,
Renal TRACERx Consortium,
Lung TRACERx Consortium,
Turajlic, S.
Gore, M.
Larkin, J.
Swanton, C.
Chester, K.A.
Pule, M.
Ravetch, J.V.
Marafioti, T.
Peggs, K.S.
Quezada, S.A.
(2017). Fc-Optimized Anti-CD25 Depletes Tumor-Infiltrating Regulatory T Cells and Synergizes with PD-1 Blockade to Eradicate Established Tumors. Immunity,
Vol.46
(4),
pp. 577-586.
show abstract
full text
CD25 is expressed at high levels on regulatory T (Treg) cells and was initially proposed as a target for cancer immunotherapy. However, anti-CD25 antibodies have displayed limited activity against established tumors. We demonstrated that CD25 expression is largely restricted to tumor-infiltrating Treg cells in mice and humans. While existing anti-CD25 antibodies were observed to deplete Treg cells in the periphery, upregulation of the inhibitory Fc gamma receptor (FcγR) IIb at the tumor site prevented intra-tumoral Treg cell depletion, which may underlie the lack of anti-tumor activity previously observed in pre-clinical models. Use of an anti-CD25 antibody with enhanced binding to activating FcγRs led to effective depletion of tumor-infiltrating Treg cells, increased effector to Treg cell ratios, and improved control of established tumors. Combination with anti-programmed cell death protein-1 antibodies promoted complete tumor rejection, demonstrating the relevance of CD25 as a therapeutic target and promising substrate for future combination approaches in immune-oncology..
Abbosh, C.
Birkbak, N.J.
Wilson, G.A.
Jamal-Hanjani, M.
Constantin, T.
Salari, R.
Le Quesne, J.
Moore, D.A.
Veeriah, S.
Rosenthal, R.
Marafioti, T.
Kirkizlar, E.
Watkins, T.B.
McGranahan, N.
Ward, S.
Martinson, L.
Riley, J.
Fraioli, F.
Al Bakir, M.
Grönroos, E.
Zambrana, F.
Endozo, R.
Bi, W.L.
Fennessy, F.M.
Sponer, N.
Johnson, D.
Laycock, J.
Shafi, S.
Czyzewska-Khan, J.
Rowan, A.
Chambers, T.
Matthews, N.
Turajlic, S.
Hiley, C.
Lee, S.M.
Forster, M.D.
Ahmad, T.
Falzon, M.
Borg, E.
Lawrence, D.
Hayward, M.
Kolvekar, S.
Panagiotopoulos, N.
Janes, S.M.
Thakrar, R.
Ahmed, A.
Blackhall, F.
Summers, Y.
Hafez, D.
Naik, A.
Ganguly, A.
Kareht, S.
Shah, R.
Joseph, L.
Marie Quinn, A.
Crosbie, P.A.
Naidu, B.
Middleton, G.
Langman, G.
Trotter, S.
Nicolson, M.
Remmen, H.
Kerr, K.
Chetty, M.
Gomersall, L.
Fennell, D.A.
Nakas, A.
Rathinam, S.
Anand, G.
Khan, S.
Russell, P.
Ezhil, V.
Ismail, B.
Irvin-Sellers, M.
Prakash, V.
Lester, J.F.
Kornaszewska, M.
Attanoos, R.
Adams, H.
Davies, H.
Oukrif, D.
Akarca, A.U.
Hartley, J.A.
Lowe, H.L.
Lock, S.
Iles, N.
Bell, H.
Ngai, Y.
Elgar, G.
Szallasi, Z.
Schwarz, R.F.
Herrero, J.
Stewart, A.
Quezada, S.A.
Peggs, K.S.
Van Loo, P.
Dive, C.
Lin, C.J.
Rabinowitz, M.
Aerts, H.J.
Hackshaw, A.
Shaw, J.A.
Zimmermann, B.G.
TRACERx consortium,
PEACE consortium,
Swanton, C.
(2017). Phylogenetic ctDNA analysis depicts early-stage lung cancer evolution. Nature,
Vol.545
(7655),
pp. 446-451.
show abstract
The early detection of relapse following primary surgery for non-small-cell lung cancer and the characterization of emerging subclones, which seed metastatic sites, might offer new therapeutic approaches for limiting tumour recurrence. The ability to track the evolutionary dynamics of early-stage lung cancer non-invasively in circulating tumour DNA (ctDNA) has not yet been demonstrated. Here we use a tumour-specific phylogenetic approach to profile the ctDNA of the first 100 TRACERx (Tracking Non-Small-Cell Lung Cancer Evolution Through Therapy (Rx)) study participants, including one patient who was also recruited to the PEACE (Posthumous Evaluation of Advanced Cancer Environment) post-mortem study. We identify independent predictors of ctDNA release and analyse the tumour-volume detection limit. Through blinded profiling of postoperative plasma, we observe evidence of adjuvant chemotherapy resistance and identify patients who are very likely to experience recurrence of their lung cancer. Finally, we show that phylogenetic ctDNA profiling tracks the subclonal nature of lung cancer relapse and metastasis, providing a new approach for ctDNA-driven therapeutic studies..
McGranahan, N.
Rosenthal, R.
Hiley, C.T.
Rowan, A.J.
Watkins, T.B.
Wilson, G.A.
Birkbak, N.J.
Veeriah, S.
Van Loo, P.
Herrero, J.
Swanton, C.
TRACERx Consortium,
(2017). Allele-Specific HLA Loss and Immune Escape in Lung Cancer Evolution. Cell,
Vol.171
(6),
pp. 1259-1271.e11.
show abstract
Immune evasion is a hallmark of cancer. Losing the ability to present neoantigens through human leukocyte antigen (HLA) loss may facilitate immune evasion. However, the polymorphic nature of the locus has precluded accurate HLA copy-number analysis. Here, we present loss of heterozygosity in human leukocyte antigen (LOHHLA), a computational tool to determine HLA allele-specific copy number from sequencing data. Using LOHHLA, we find that HLA LOH occurs in 40% of non-small-cell lung cancers (NSCLCs) and is associated with a high subclonal neoantigen burden, APOBEC-mediated mutagenesis, upregulation of cytolytic activity, and PD-L1 positivity. The focal nature of HLA LOH alterations, their subclonal frequencies, enrichment in metastatic sites, and occurrence as parallel events suggests that HLA LOH is an immune escape mechanism that is subject to strong microenvironmental selection pressures later in tumor evolution. Characterizing HLA LOH with LOHHLA refines neoantigen prediction and may have implications for our understanding of resistance mechanisms and immunotherapeutic approaches targeting neoantigens. VIDEO ABSTRACT..
Spain, L.
Higgins, R.
Gopalakrishnan, K.
Turajlic, S.
Gore, M.
Larkin, J.
(2016). Acute renal allograft rejection after immune checkpoint inhibitor therapy for metastatic melanoma. Ann oncol,
Vol.27
(6),
pp. 1135-1137.
show abstract
Immune checkpoint inhibitors such as ipilimumab and nivolumab improve survival in patients with advanced melanoma and are increasingly available to clinicians for use in the clinic. Their safety in organ transplant recipients is not well defined but published case reports describing treatment with ipilimumab have not been complicated by graft rejection. No cases of anti-programmed cell death protein 1 administration are reported in this group. We describe a case of acute graft rejection in a kidney transplant recipient after treatment with nivolumab, after progression on ipilimumab. Potential factors increasing the risk of graft rejection in this case are discussed, in particular the contribution of nivolumab..
Turajlic, S.
Swanton, C.
(2016). Metastasis as an evolutionary process. Science,
Vol.352
(6282),
pp. 169-175.
show abstract
Therapeutic advances in oncology have not fully translated to the treatment of metastatic disease, which remains largely incurable. Metastatic subclones can emerge both early and late in the life of the primary tumor. A better understanding of the genetic evolution of metastatic disease has the potential to reveal differences in the therapeutic vulnerabilities of primary and metastatic tumors, shed light on the temporal patterns of and routes to metastatic colonization, and provide insight into the biology of the metastatic process. Here we review recent comparative studies of primary and metastatic tumors, including data suggesting that macroevolutionary shifts (the onset of chromosomal instability) contribute to the evolution of metastatic disease. We also discuss the practical challenges associated with these studies and how they might be overcome..
Turajlic, S.
Larkin, J.
(2016). Systemic treatment of advanced papillary renal cell carcinoma: Where next?. Eur j cancer,
Vol.69,
pp. 223-225.
full text
Turajlic, S.
Larkin, J.
Swanton, C.
(2015). SnapShot: Renal Cell Carcinoma. Cell,
Vol.163
(6),
pp. 1556-1556.e1.
show abstract
This SnapShot summarizes current knowledge about the key features in mutational landscape, major pathways, and tumor evolution and heterogeneity in renal cell carcinoma, as well as the most recent advances in therapeutic development. To view this SnapShot, open or download the PDF..
Turajlic, S.
McGranahan, N.
Swanton, C.
(2015). Inferring mutational timing and reconstructing tumour evolutionary histories. Biochim biophys acta,
Vol.1855
(2),
pp. 264-275.
show abstract
Cancer evolution can be considered within a Darwinian framework. Both micro and macro-evolutionary theories can be applied to understand tumour progression and treatment failure. Owing to cancers' complexity and heterogeneity the rules of tumour evolution, such as the role of selection, remain incompletely understood. The timing of mutational events during tumour evolution presents diagnostic, prognostic and therapeutic opportunities. Here we review the current sampling and computational approaches for inferring mutational timing and the evidence from next generation sequencing-informed data on mutational timing across all tumour types. We discuss how this knowledge can be used to illuminate the genes and pathways that drive cancer initiation and relapse; and to support drug development and clinical trial design..
Gulati, S.
Turajlic, S.
Larkin, J.
Bates, P.A.
Swanton, C.
(2015). Relapse models for clear cell renal carcinoma. Lancet oncol,
Vol.16
(8),
pp. e376-e378.
Soultati, A.
Stares, M.
Swanton, C.
Larkin, J.
Turajlic, S.
(2015). How should clinicians address intratumour heterogeneity in clear cell renal cell carcinoma?. Curr opin urol,
Vol.25
(5),
pp. 358-366.
show abstract
PURPOSE OF REVIEW: Despite the availability of multiple targeted therapies, the 5-year survival rate of patients with metastatic clear cell renal cell carcinoma (ccRCC) rarely exceeds 10%. Recent insights into the mutational landscape and evolutionary dynamics of ccRCC have offered up a plausible explanation for these outcomes. The purpose of this review is to link the research findings to potential changes in clinical practice. RECENT FINDINGS: Intratumour heterogeneity (ITH) dominates the evolutionary landscape in ccRCC at the genetic, transcriptomic and proteomic level. Spatial and temporal separation of tumour subclones within the primary tumour as well as between primary and metastatic sites has been demonstrated at single nucleotide resolution. In the cases analysed to date, approximately two-thirds of somatic mutations are not shared between multiple biopsies from the same primary tumour. Very few of the key disease-driving events are shared across all primary tumour regions (with the exception of VHL and loss of chromosome 3p), whereas the majority are restricted to one or more tumour regions (TP53, SETD2, BAP1, PTEN, mTOR, PIK3CA and KDM5C). SUMMARY: ITH must be considered in the management of ccRCC with respect to diagnostic procedures, prognostic and predictive biomarkers and drug development..
Turajlic, S.
Larkin, J.
Swanton, C.
(2015). Academically led clinical trials: challenges and opportunities. Ann oncol,
Vol.26
(10),
pp. 2010-2011.
Turajlic, S.
Swanton, C.
(2015). Gastrointestinal cancer: Tracking tumour evolution through liquid biopsy. Nat rev clin oncol,
Vol.12
(10),
pp. 565-566.
Turajlic, S.
Furney, S.J.
Stamp, G.
Rana, S.
Ricken, G.
Oduko, Y.
Saturno, G.
Springer, C.
Hayes, A.
Gore, M.
Larkin, J.
Marais, R.
(2014). Whole-genome sequencing reveals complex mechanisms of intrinsic resistance to BRAF inhibition. Ann oncol,
Vol.25
(5),
pp. 959-967.
show abstract
BACKGROUND: BRAF is mutated in ∼42% of human melanomas (COSMIC. http://www.sanger.ac.uk/genetics/CGP/cosmic/) and pharmacological BRAF inhibitors such as vemurafenib and dabrafenib achieve dramatic responses in patients whose tumours harbour BRAF(V600) mutations. Objective responses occur in ∼50% of patients and disease stabilisation in a further ∼30%, but ∼20% of patients present primary or innate resistance and do not respond. Here, we investigated the underlying cause of treatment failure in a patient with BRAF mutant melanoma who presented primary resistance. METHODS: We carried out whole-genome sequencing and single nucleotide polymorphism (SNP) array analysis of five metastatic tumours from the patient. We validated mechanisms of resistance in a cell line derived from the patient's tumour. RESULTS: We observed that the majority of the single-nucleotide variants identified were shared across all tumour sites, but also saw site-specific copy-number alterations in discrete cell populations at different sites. We found that two ubiquitous mutations mediated resistance to BRAF inhibition in these tumours. A mutation in GNAQ sustained mitogen-activated protein kinase (MAPK) signalling, whereas a mutation in PTEN activated the PI3 K/AKT pathway. Inhibition of both pathways synergised to block the growth of the cells. CONCLUSIONS: Our analyses show that the five metastases arose from a common progenitor and acquired additional alterations after disease dissemination. We demonstrate that a distinct combination of mutations mediated primary resistance to BRAF inhibition in this patient. These mutations were present in all five tumours and in a tumour sample taken before BRAF inhibitor treatment was administered. Inhibition of both pathways was required to block tumour cell growth, suggesting that combined targeting of these pathways could have been a valid therapeutic approach for this patient..
Furney, S.J.
Turajlic, S.
Stamp, G.
Thomas, J.M.
Hayes, A.
Strauss, D.
Gavrielides, M.
Xing, W.
Gore, M.
Larkin, J.
Marais, R.
(2014). The mutational burden of acral melanoma revealed by whole-genome sequencing and comparative analysis. Pigment cell melanoma res,
Vol.27
(5),
pp. 835-838.
show abstract
Acral melanoma is a subtype of melanoma with distinct epidemiological, clinical and mutational profiles. To define the genomic alterations in acral melanoma, we conducted whole-genome sequencing and SNP array analysis of five metastatic tumours and their matched normal genomes. We identified the somatic mutations, copy number alterations and structural variants in these tumours and combined our data with published studies to identify recurrently mutated genes likely to be the drivers of acral melanomagenesis. We compared and contrasted the genomic landscapes of acral, mucosal, uveal and common cutaneous melanoma to reveal the distinctive mutational characteristics of each subtype..
Sanchez-Laorden, B.
Viros, A.
Girotti, M.R.
Pedersen, M.
Saturno, G.
Zambon, A.
Niculescu-Duvaz, D.
Turajlic, S.
Hayes, A.
Gore, M.
Larkin, J.
Lorigan, P.
Cook, M.
Springer, C.
Marais, R.
(2014). BRAF inhibitors induce metastasis in RAS mutant or inhibitor-resistant melanoma cells by reactivating MEK and ERK signaling. Sci signal,
Vol.7
(318),
p. ra30.
show abstract
Melanoma is a highly metastatic and lethal form of skin cancer. The protein kinase BRAF is mutated in about 40% of melanomas, and BRAF inhibitors improve progression-free and overall survival in these patients. However, after a relatively short period of disease control, most patients develop resistance because of reactivation of the RAF-ERK (extracellular signal-regulated kinase) pathway, mediated in many cases by mutations in RAS. We found that BRAF inhibition induces invasion and metastasis in RAS mutant melanoma cells through a mechanism mediated by the reactivation of the MEK (mitogen-activated protein kinase kinase)-ERK pathway, increased expression and secretion of interleukin 8, and induction of protease-dependent invasion. These events were accompanied by a cell morphology switch from predominantly rounded to predominantly elongated cells. We also observed similar responses in BRAF inhibitor-resistant melanoma cells. These data show that BRAF inhibitors can induce melanoma cell invasion and metastasis in tumors that develop resistance to these drugs..
Girotti, M.R.
Pedersen, M.
Sanchez-Laorden, B.
Viros, A.
Turajlic, S.
Niculescu-Duvaz, D.
Zambon, A.
Sinclair, J.
Hayes, A.
Gore, M.
Lorigan, P.
Springer, C.
Larkin, J.
Jorgensen, C.
Marais, R.
(2013). Inhibiting EGF receptor or SRC family kinase signaling overcomes BRAF inhibitor resistance in melanoma. Cancer discov,
Vol.3
(2),
pp. 158-167.
show abstract
UNLABELLED: We generated cell lines resistant to BRAF inhibitors and show that the EGF receptor (EGFR)-SRC family kinase (SFK)-STAT3 signaling pathway was upregulated in these cells. In addition to driving proliferation of resistant cells, this pathway also stimulated invasion and metastasis. EGFR inhibitors cooperated with BRAF inhibitors to block the growth of the resistant cells in vitro and in vivo, and monotherapy with the broad specificity tyrosine kinase inhibitor dasatinib blocked growth and metastasis in vivo. We analyzed tumors from patients with intrinsic or acquired resistance to vemurafenib and observed increased EGFR and SFK activity. Furthermore, dasatinib blocked the growth and metastasis of one of the resistant tumors in immunocompromised mice. Our data show that BRAF inhibitor-mediated activation of EGFR-SFK-STAT3 signaling can mediate resistance in patients with BRAF-mutant melanoma. We describe 2 treatments that seem to overcome this resistance and could deliver therapeutic efficacy in patients with drug-resistant BRAF-mutant melanoma. SIGNIFICANCE: Therapies that target the driver oncogenes in cancer can achieve remarkable responses if patients are stratified for treatment. However, as with conventional therapies, patients often develop acquired resistance to targeted therapies, and a proportion of patients are intrinsically resistant and fail to respond despite the presence of an appropriate driver oncogene mutation. We found that the EGFR/SFK pathway mediated resistance to vemurafenib in BRAF -mutant melanoma and that BRAF and EGFR or SFK inhibition blocked proliferation and invasion of these resistant tumors, providing potentially effective therapeutic options for these patients..
Khattak, M.
Fisher, R.
Turajlic, S.
Larkin, J.
(2013). Targeted therapy and immunotherapy in advanced melanoma: an evolving paradigm. Ther adv med oncol,
Vol.5
(2),
pp. 105-118.
show abstract
Metastatic melanoma is one of the most challenging malignancies to treat and often has a poor outcome. Until recently, systemic treatment options were limited, with poor response rates and no survival advantage. However, the treatment of metastatic melanoma has been revolutionized by developments in targeted therapy and immunotherapy; the BRAF inhibitor, vemurafenib, and anticytotoxic T-lymphocyte antigen 4 antibody, ipilimumab, are the first agents to demonstrate a survival benefit. Despite the success of these treatments, most patients eventually progress, and research into response and resistance mechanisms, rationally designed combination therapies and evaluation of the role of these agents in the adjuvant setting is critically important..
Turajlic, S.
Ali, Z.
Yousaf, N.
Larkin, J.
(2013). Phase I/II RAF kinase inhibitors in cancer therapy. Expert opin investig drugs,
Vol.22
(6),
pp. 739-749.
show abstract
INTRODUCTION: Aberrant activation of RAF signalling is a frequent finding in human cancers. BRAF is the only RAF family member that is commonly mutated, whilst CRAF and ARAF play important roles in the signal transduction from mutant RAS. BRAF-specific inhibitors have been more effective in the treatment of BRAF-mutant melanoma than BRAF-mutant thyroid and colorectal cancers. AREAS COVERED: The review summarises the experience with RAF kinase inhibitors, including efficacy, modes of acquired resistance, and the mechanism behind the progression of pre-malignant RAS-mutant lesions observed with RAF kinase inhibitors. The authors review all the completed and ongoing Phase I or II clinical trials of RAF kinase inhibitors and discuss in detail the rationale behind the combinatorial approaches. EXPERT OPINION: The success of RAF kinase inhibitors has demonstrated the necessity of genotype-driven treatment selection for cancer patients. The spectrum of responses in different tumour types is explained by feedback events that are determined by cell lineage. Dissection of these events and the mechanisms of acquired resistance will determine the appropriate combination therapies. Ongoing characterisation of RAS-MAPK regulation in malignant cells may aid the development of novel agents that have greater potency for the inhibition of activated RAF kinase, and lesser propensity for promotion of RAS-mutant tumours..
Furney, S.J.
Turajlic, S.
Stamp, G.
Nohadani, M.
Carlisle, A.
Thomas, J.M.
Hayes, A.
Strauss, D.
Gore, M.
van den Oord, J.
Larkin, J.
Marais, R.
(2013). Genome sequencing of mucosal melanomas reveals that they are driven by distinct mechanisms from cutaneous melanoma. J pathol,
Vol.230
(3),
pp. 261-269.
show abstract
Mucosal melanoma displays distinct clinical and epidemiological features compared to cutaneous melanoma. Here we used whole genome and whole exome sequencing to characterize the somatic alterations and mutation spectra in the genomes of ten mucosal melanomas. We observed somatic mutation rates that are considerably lower than occur in sun-exposed cutaneous melanoma, but comparable to the rates seen in cancers not associated with exposure to known mutagens. In particular, the mutation signatures are not indicative of ultraviolet light- or tobacco smoke-induced DNA damage. Genes previously reported as mutated in other cancers were also mutated in mucosal melanoma. Notably, there were substantially more copy number and structural variations in mucosal melanoma than have been reported in cutaneous melanoma. Thus, mucosal and cutaneous melanomas are distinct diseases with discrete genetic features. Our data suggest that different mechanisms underlie the genesis of these diseases and that structural variations play a more important role in mucosal than in cutaneous melanomagenesis..
Furney, S.J.
Pedersen, M.
Gentien, D.
Dumont, A.G.
Rapinat, A.
Desjardins, L.
Turajlic, S.
Piperno-Neumann, S.
de la Grange, P.
Roman-Roman, S.
Stern, M.-.
Marais, R.
(2013). SF3B1 mutations are associated with alternative splicing in uveal melanoma. Cancer discov,
Vol.3
(10),
pp. 1122-1129.
show abstract
full text
UNLABELLED: Uveal melanoma, the most common eye malignancy, causes severe visual morbidity and is fatal in approximately 50% of patients. Primary uveal melanoma can be cured by surgery or radiotherapy, but the metastatic disease is treatment refractory. To understand comprehensively uveal melanoma genetics, we conducted single-nucleotide polymorphism arrays and whole-genome sequencing on 12 primary uveal melanomas. We observed only approximately 2,000 predicted somatic single-nucleotide variants per tumor and low levels of aneuploidy. We did not observe an ultraviolet radiation DNA damage signature, but identified SF3B1 mutations in three samples and a further 15 mutations in an extension cohort of 105 samples. SF3B1 mutations were associated with good prognosis and were rarely coincident with BAP1 mutations. SF3B1 encodes a component of the spliceosome, and RNA sequencing revealed that SF3B1 mutations were associated with differential alternative splicing of protein coding genes, including ABCC5 and UQCC, and of the long noncoding RNA CRNDE. SIGNIFICANCE: Our data show that despite its dismal prognosis, uveal melanoma is a relatively simple genetic disease characterized by recurrent chromosomal losses and gains and a low mutational burden. We show that SF3B1 is recurrently mutated in uveal melanoma, and the mutations are associated with aberrant alternative splicing..
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..
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)..
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/..
Larkin, J.M.
Turajlic, S.
Nathan, P.D.
Lorigan, P.
Stamp, G.
Gonzalez de Castro, D.
Martin, N.
Griffiths, J.
Edmonds, K.
Sarker, S.
James, M.G.
A'Hern, R.
Coombes, G.
Snowdon, C.
Bliss, J.M.
Gore, M.E.
Marais, R.
(2011). A phase II trial of nilotinib in the treatment of patients with KIT mutated advanced acral and mucosal melanoma (NICAM). J clin oncol,
Vol.29
(15_suppl),
p. TPS229.
show abstract
TPS229 Background: Recent studies have indicated that KIT mutations occur in approximately 20% of acral and mucosal melanomas (Curtin, al. 2006; Beadling, et al. 2008). Several case reports have shown that KIT mutated melanomas can respond to treatment with KIT inhibitors (Hodi et al. 2008; Lutzky et al. 2008; Quintas-Cardama et al. 2008; Handolias et al. 2010; Itoh, et al. 2010; Satzger et al. 2010; Terheyden et al. 2010; Woodman and Davies 2010) suggesting that such tumours critically depend on upregulated KIT signalling. Nilotinib, which potently inhibits the tyrosine kinase activity of BCR-ABL and KIT, has been evaluated in patients with advanced KIT mutated GIST and has shown clinical efficacy at well tolerated doses (Demetri et al. 2009; Montemurro et al. 2009; Schlemmer et al. 2010). METHODS: NICAM is a single arm 2-stage open label phase II study of nilotinib in KIT mutated advanced mucosal or acral melanoma. Patients with KIT mutations known to confer resistance to nilotinib are excluded. The primary objective in this feasibility study is to evaluate the efficacy of nilotinib in KIT mutated advanced mucosal and acral melanoma with a primary endpoint of progression free survival at 6 months. Secondary endpoints include response rate at 12 weeks; overall survival; toxicity of treatment; correlation between the response to nilotinib and KIT genotype, KIT gene copy number and KIT expression. In addition, the following will be explored: 1. Suppression of phosphorylation of KIT and downstream pathways on day 15 tumour biopsies; 2. Possible mechanisms of resistance by analysis of biopsies obtained at disease progression; 3. The use of circulating tumour DNA for non-invasive KIT mutational testing; 4. Genetic lesions that potentially cooperate with oncogenic KIT in melanoma (using Next Generation Sequencing). All patients are treated until disease progression or unacceptable toxicity. If 2 or more of the first 9 patients are progression free at 6 months, patient entry will continue into the second stage until a total of 24 patients have been recruited. To date, 45 patients have been registered and screened for KIT mutation and 5 patients have entered the study..
Perez, A.
Turajlic, S.
Szyszko, T.
O'Doherty, M.
Calonje, E.
Harries, M.
Acland, K.
(2010). Generalized melanosis and melanuria in a patient with metastatic melanoma. Clin exp dermatol,
Vol.35
(3),
pp. e37-e39.
Buck, M.D.
Poirier, E.Z.
Cardoso, A.
Frederico, B.
Canton, J.
Barrell, S.
Beale, R.
Byrne, R.
Caidan, S.
Crawford, M.
Cubitt, L.
Gandhi, S.
Goldstone, R.
Grant, P.R.
Gulati, K.
Hindmarsh, S.
Howell, M.
Hubank, M.
Instrell, R.
Jiang, M.
Kassiotis, G.
Lu, W.-.
MacRae, J.I.
Martini, I.
Miller, D.
Moore, D.
Nastouli, E.
Nicod, J.
Nightingale, L.
Olsen, J.
Oomatia, A.
O'Reilly, N.
Rideg, A.
Song, O.-.
Strange, A.
Swanton, C.
Turajlic, S.
Wu, M.
Reis e Sousa, C.
SARS-CoV-2 detection by a clinical diagnostic RT-LAMP assay. Wellcome open research,
Vol.6,
pp. 9-9.
show abstract
full text
The ongoing pandemic of SARS-CoV-2 calls for rapid and cost-effective methods to accurately identify infected individuals. The vast majority of patient samples is assessed for viral RNA presence by RT-qPCR. Our biomedical research institute, in collaboration between partner hospitals and an accredited clinical diagnostic laboratory, established a diagnostic testing pipeline that has reported on more than 252,000 RT-qPCR results since its commencement at the beginning of April 2020. However, due to ongoing demand and competition for critical resources, alternative testing strategies were sought. In this work, we present a clinically-validated procedure for high-throughput SARS-CoV-2 detection by RT-LAMP in 25 minutes that is robust, reliable, repeatable, sensitive, specific, and inexpensive..
Litchfield, D.K.
Stanislaw, S.
Spain, L.
Gallegos, L.
Rowan, A.
Schnidrig, D.
Rosenbaum, H.
Harle, A.
Au, L.
Hill, S.
Tippu, Z.
Thomas, J.
Xu, H.
Horswell, S.
Barhoumi, A.
Jones, C.
Leith, K.F.
Burgess, D.L.
Watkins, T.B.
Lim, E.
Birkbak, N.J.
Lamy, P.
Nordentoft, I.
Dyrskjøt, L.
Hazell, S.
Jamal-Hanjani, M.
Larkin, J.
Swanton, C.
Alexander, N.R.
Turajlic, S.
Representative Sequencing: Unbiased Sampling of Solid Tumor Tissue. Ssrn electronic journal,
.