Bubendorf, Lukas

Intratumoral heterogeneity and clonal evolution are pivotal in the progression and metastasis of melanoma. However, when combined with variable tumor cellularity, intratumoral heterogeneity limits the sensitivity and accuracy of uncovering a cancer’s clonal evolution. In this study, we combined fluorescence-activated cell sorting (FACS) with whole-exome sequencing (WES) to investigate the clonal composition and evolutionary patterns in seven melanoma biopsies obtained from three patients, each having both primary site and metastatic samples. We employed a multiparameter ploidy sorting approach to isolate tumor populations based on DNA ploidy and melanoma biomarkers (SOX10 or S100), enabling us to investigate clonal evolution with high resolution. Our approach increased the mean tumor purity from 70% (range 19–88%) in unsorted material to 91% (range 87–96%) post-sorting. Our findings revealed significant inter- and intratumor heterogeneity, with one patient exhibiting two genomically distinct clonal tumor populations within a single primary site biopsy, each giving rise to different metastases. Our findings highlight the critical role of intratumoral heterogeneity and clonal evolution in melanoma, especially when analyzing tumor trajectories. The unique combination of multiparameter FACS and WES provides a powerful method for identifying clonal populations and reconstructing clonal evolution. This study provides valuable insights into the clonal architecture of melanoma and lays the groundwork for future research with larger patient groups.

AbstractVariable tumor cellularity can limit sensitivity and precision in comparative genomics because differences in tumor content can result in misclassifying truncal mutations as region‐specific private mutations in stroma‐rich regions, especially when studying tissue specimens of mediocre tumor cellularity such as lung adenocarcinomas (LUADs). To address this issue, we refined a nuclei flow‐sorting approach by sorting nuclei based on ploidy and the LUAD lineage marker thyroid transcription factor 1 and applied this method to investigate genome‐wide somatic copy number aberrations (SCNAs) and mutations of 409 cancer genes in 39 tumor populations obtained from 16 primary tumors and 21 matched metastases. This approach increased the mean tumor purity from 54% (range 7–89%) of unsorted material to 92% (range 79–99%) after sorting. Despite this rise in tumor purity, we detected limited genetic heterogeneity between primary tumors and their metastases. In fact, 88% of SCNAs and 80% of mutations were propagated from primary tumors to metastases and low allele frequency mutations accounted for much of the mutational heterogeneity. Even though the presence of SCNAs indicated a history of chromosomal instability (CIN) in all tumors, metastases did not have more SCNAs than primary tumors. Moreover, tumors with biallelic TP53 or ATM mutations had high numbers of SCNAs, yet they were associated with a low interlesional genetic heterogeneity. The results of our study thus provide evidence that most macroevolutionary events occur in primary tumors before metastatic dissemination and advocate for a limited degree of CIN over time and space in this cohort of LUADs. Sampling of primary tumors thus may suffice to detect most mutations and SCNAs. In addition, metastases but not primary tumors had seeded additional metastases in three of four patients; this provides a genomic rational for surgical treatment of such oligometastatic LUADs. Copyright © 2018 Pathological Society of Great Britain and Ireland. Published by John Wiley & Sons, Ltd.

AbstractUnderstanding the evolutionary mechanisms and genomic events leading to castration‐resistant (CR) prostate cancer (PC) is key to improve the outcome of this otherwise deadly disease. Here, we delineated the tumour history of seven patients progressing to castration resistance by analysing matched prostate cancer tissues before and after castration. We performed genomic profiling of DNA content‐based flow‐sorted populations in order to define the different evolutionary patterns. In one patient, we discovered that a catastrophic genomic event, known as chromothripsis, resulted in multiple CRPC tumour populations with distinct, potentially advantageous copy number aberrations, including an amplification of FK506 binding protein 4 (FKBP4, also known as FKBP52), a protein enhancing the transcriptional activity of androgen receptor signalling. Analysis of FKBP4 protein expression in more than 500 prostate cancer samples revealed increased expression in CRPC in comparison to hormone‐naïve (HN) PC. Moreover, elevated FKBP4 expression was associated with poor survival of patients with HNPC. We propose FKBP4 amplification and overexpression as a selective advantage in the process of tumour evolution and as a potential mechanism associated with the development of CRPC. Furthermore, FKBP4 interaction with androgen receptor may provide a potential therapeutic target in PC. Copyright © 2018 Pathological Society of Great Britain and Ireland. Published by John Wiley & Sons, Ltd.