Clonal Heterogeneity and Tumor Evolution: Past, Present, and the Future

Intratumor heterogeneity, which fosters tumor evolution, is a key challenge in cancer medicine. Here, we review data and technologies that have revealed intra-tumor heterogeneity across cancer types and the dynamics, constraints, and contingencies inherent to tumor evolution. We emphasize the importance of macro-evolutionary leaps, often involving large-scale chromosomal alterations, in driving tumor evolution and metastasis and consider the role of the tumor microenvironment in engendering heterogeneity and drug resistance. We suggest that bold approaches to drug development, harnessing the adaptive properties of the immune-microenvironment while limiting those of the tumor, combined with advances in clinical trial-design, will improve patient outcome

Circulating tumor DNA analyses reveal novel resistance mechanisms to CDK inhibition in metastatic breast cancer

Cyclin-dependent kinase (CDK) 4/6 inhibition has been demonstrated to improve progression-free survival (PFS) in patients with human epidermal growth factor receptor 2 (HER2−), hormone receptor positive (HR+) in advanced breast cancer [1–3]. Palbociclib, ribociclib and abemaciclib are orally bioavailable selective CDK 4/6 inhibitors. These small molecules likely bind the ATP-binding pocket within the CDK4/6 protein kinases thereby inhibiting phosphorylation of retinoblastoma tumour suppressor protein (Rb). In its hypophosphorylated state Rb remains bound to E2F thereby preventing progression through the G1-S-cell cycle checkpoint [4]. The mechanism behind the observed efficacy of CDK inhibition in metastatic breast cancer may relate to a dependence of HR+ breast cancer on CDK4/6 activity to override Rb mediated repression of cell cycle progression (Figure 1) [5]

Intra-tumor heterogeneity: lessons from microbial evolution and clinical implications

Multiple subclonal populations of tumor cells can coexist within the same tumor. This intra-tumor heterogeneity will have clinical implications and it is therefore important to identify factors that drive or suppress such heterogeneous tumor progression. Evolutionary biology can provide important insights into this process. In particular, experimental evolution studies of microbial populations, which exist as clonal populations that can diversify into multiple subclones, have revealed important evolutionary processes driving heterogeneity within a population. There are transferrable lessons that can be learnt from these studies that will help us to understand the process of intra-tumor heterogeneity in the clinical setting. In this review, we summarize drivers of microbial diversity that have been identified, such as mutation rate and environmental influences, and discuss how knowledge gained from microbial experimental evolution studies may guide us to identify and understand important selective factors that promote intra-tumor heterogeneity. Furthermore, we discuss how these factors could be used to direct and optimize research efforts to improve patient care, focusing on therapeutic resistance. Finally, we emphasize the need for longitudinal studies to address the impact of these potential tumor heterogeneity-promoting factors on drug resistance, metastatic potential and clinical outcome

Spatial and temporal diversity in genomic instability processes defines lung cancer evolution

PMCID: PMC4636050.-- et al.Spatial and temporal dissection of the genomic changes occurring during the evolution of human non-small cell lung cancer (NSCLC) may help elucidate the basis for its dismal prognosis.We sequenced 25 spatially distinct regions from seven operable NSCLCs and found evidence of branched evolution, with driver mutations arising before and after subclonal diversification. There was pronounced intratumor heterogeneity in copy number alterations, translocations, and mutations associated with APOBEC cytidine deaminase activity. Despite maintained carcinogen exposure, tumors from smokers showed a relative decrease in smoking-related mutations over time, accompanied by an increase in APOBEC-associated mutations. In tumors from former smokers, genome-doubling occurred within a smoking-signature context before subclonal diversification, which suggested that a long period of tumor latency had preceded clinical detection. The regionally separated driver mutations, coupled with the relentless and heterogeneous nature of the genome instability processes, are likely to confound treatment success in NSCLC.E.B. is a Rosetrees Trust fellow; M.J.H. has a Cancer Research UK fellowship; N.Mu. received funding from the Rosetrees Trust; M.G. is funded by the UK Medical Research Council; I.V. is funded by Spanish Ministerio de Economía y Competitividad subprograma Ramón y Cajal; R.C.R. and D.M.R. are partly funded by the Cambridge Biomedical Research Centre and Cancer Research UK Cancer Centre; P.V.L. is a postdoctoral researcher of the Research Foundation—Flanders (FWO); S.M.J. is a Wellcome Senior Fellow in Clinical Science; and C.S. is a senior Cancer Research UK clinical research fellow and is funded by Cancer Research UK, the Rosetrees Trust, European Union Framework Programme 7 (projects PREDICT and RESPONSIFY, ID:259303), the Prostate Cancer Foundation, the European Research Council and the Breast Cancer Research Foundation. This research is supported by the National Institute for Health Research University College London Hospitals Biomedical Research Centre.Peer Reviewe

deconstructSigs: delineating mutational processes in single tumors distinguishes DNA repair deficiencies and patterns of carcinoma evolution

BACKGROUND: Analysis of somatic mutations provides insight into the mutational processes that have shaped the cancer genome, but such analysis currently requires large cohorts. We develop deconstructSigs, which allows the identification of mutational signatures within a single tumor sample. RESULTS: Application of deconstructSigs identifies samples with DNA repair deficiencies and reveals distinct and dynamic mutational processes molding the cancer genome in esophageal adenocarcinoma compared to squamous cell carcinomas. CONCLUSIONS: deconstructSigs confers the ability to define mutational processes driven by environmental exposures, DNA repair abnormalities, and mutagenic processes in individual tumors with implications for precision cancer medicine

APOBEC mutagenesis in drug resistance and immune escape in HIV and cancer evolution

The APOBEC mutational signature has only recently been detected in a multitude of cancers through next-generation sequencing. In contrast, APOBEC has been a focus of virology research for over a decade. Many lessons learnt regarding APOBEC within virology are likely to be applicable to cancer. In this review, we explore the parallels between the role of APOBEC enzymes in HIV and cancer evolution. We discuss data supporting the role of APOBEC mutagenesis in creating HIV genome heterogeneity, drug resistance, and immune escape variants. We hypothesize similar functions of APOBEC will also hold true in cancer

Differential binding affinity of mutated peptides for MHC class I is a predictor of survival in advanced lung cancer and melanoma

Background: Cancer mutations generate novel (neo-)peptides recognised by T cells, but the determinants of recognition are not well characterised. The difference in predicted class I major histocompatibility complex (MHC-I) binding affinity between wild-type and corresponding mutant peptides (differential agretopicity index; DAI) may reflect clinically relevant cancer peptide immunogenicity. Our aim was to explore the relationship between DAI, measures of immune infiltration and patient outcomes in advanced cancer. Patients and methods: Cohorts of patients with advanced non-small-cell lung cancer (NSCLC; LUAD, n = 66) and melanoma (SKCM, n = 72) were obtained from The Cancer Genome Atlas. Three additional cohorts of immunotherapy treated patients with advanced melanoma (total n = 131) and NSCLC (n = 31) were analysed. Neopeptides and their clonal status were defined using genomic data. MHC-I binding affinity was predicted for each neopeptide and DAI values summarised as the sample mean DAI. Correlations between mean DAI and markers of immune activity were evaluated using measures of lymphocyte infiltration and immune gene expression. Results: In univariate and multivariate analyses, mean DAI significantly correlated with overall survival in 3/5 cohorts, with evidence of superiority over nonsynonymous mutational and neoantigen burden. In these cohorts, the effect was seen for mean DAI of clonal but not subclonal peptides. In SKCM, the association between mean DAI and survival bordered significance (P = 0.068), reaching significance in an immunotherapy-treated melanoma cohort (P = 0.003). Mean DAI but not mutational nor neoantigen burden was positively correlated with independently derived markers of immune infiltration in both SKCM (P = 0.027) and LUAD (P = 0.024). Conclusions: The association between mean DAI, survival and measures of immune activity support the hypothesis that DAI is a determinant of cancer peptide immunogenicity. Investigation of DAI as a marker of immunologically relevant peptides in further datasets and future clinical studies of neoantigen based immunotherapies is warranted

Expansion of airway basal epithelial cells from primary human non-small cell lung cancer tumors

Pre-clinical non-small cell lung cancer (NSCLC) models are poorly representative of the considerable inter- and intra-tumor heterogeneity of the disease in patients. Primary cell-based in vitro models of NSCLC are therefore desirable for novel therapy development and personalized cancer medicine. Methods have been described to generate rapidly proliferating epithelial cell cultures from multiple human epithelia using 3T3-J2 feeder cell culture in the presence of Y-27632, a RHO-associated protein kinase (ROCK) inhibitor, in what are known as "conditional reprograming conditions" (CRC) or 3T3+Y. In some cancer studies, variations of this methodology have allowed primary tumor cell expansion across a number of cancer types but other studies have demonstrated the preferential expansion of normal epithelial cells from tumors in such conditions. Here, we report our experience regarding the derivation of primary NSCLC cell cultures from 12 lung adenocarcinoma patients enrolled in the Tracking Cancer Evolution through Therapy (TRACERx) clinical study and discuss these in the context of improving the success rate for in vitro cultivation of cells from NSCLC tumors. This article is protected by copyright. All rights reserved

Cyclin D mediates tolerance of genome-doubling in cancers with functional p53

BACKGROUND: Aneuploidy and chromosomal instability (CIN) are common features of human malignancy that fuel genetic heterogeneity. Although tolerance to tetraploidization, an intermediate state that further exacerbates CIN, is frequently mediated by TP53 dysfunction, we find that some genome-doubled tumours retain wild-type TP53. We sought to understand how tetraploid cells with a functional p53/p21-axis tolerate genome-doubling events. METHOD: We performed quantitative proteomics in a diploid/tetraploid pair within a system of multiple independently derived TP53 wild-type tetraploid clones arising spontaneously from a diploid progenitor. We characterized adapted and acute tetraploidization in a variety of flow cytometry and biochemical assays and tested our findings against human tumours through bioinformatics analysis of the TCGA dataset. RESULTS: Cyclin D1 was found to be specifically overexpressed in early but not late passage tetraploid clones, and this overexpression was sufficient to promote tolerance to spontaneous and pharmacologically induced tetraploidy. We provide evidence that this role extends to D-type cyclins and their overexpression confers specific proliferative advantage to tetraploid cells. We demonstrate that tetraploid clones exhibit elevated levels of functional p53 and p21 but override the p53/p21 checkpoint by elevated expression of cyclin D1, via a stoichiometry-dependent and CDK activity-independent mechanism. Tetraploid cells do not exhibit increased sensitivity to abemaciclib, suggesting that cyclin D-overexpressing tumours might not be specifically amenable to treatment with CDK4/6 inhibitors. CONCLUSION: Our study suggests that D-type cyclin overexpression is an acute event, permissive for rapid adaptation to a genome-doubled state in TP53 wild-type tumours and that its overexpression is dispensable in later stages of tumour progression