GISTIC2.0 facilitates sensitive and confident localization of the targets of focal somatic copy-number alteration in human cancers

Prostate cancer is the most common type of cancer in men in the UK and USA. Cancers develop when cells in the body acquire genetic mutations that allow the cells to grow rapidly and form a mass known as a tumor. Prostate cancer cells from different individuals can carry different genetic mutations, which affects whether the disease progresses and how the tumors respond to medical treatments. This genetic variety arises in cancer cells partly from a phenomenon known as genomic instability, in which DNA mutations accumulate due to defects in DNA repair. Genetic studies of biopsies taken from human prostate cancers have shown that genomic instability causes chromosomes—the structures in which the cell's DNA is organized—to break and then be stuck back together haphazardly. As a result, fragments of chromosomes can end up in the wrong position, be duplicated, or be lost altogether. All of these mutations could spur on the growth of the tumor. However, it is currently not clear why some prostate cancers are more genomically unstable than others, or what exactly causes this instability. Boysen, Barbieri et al. studied prostate cancer cells taken from patients before they started medical treatment. The experiments show that the cancer cells with high levels of genomic instability also often had mutations in a gene that encodes a protein called SPOP. These mutations occur in about 10 percent of men with prostate cancer and appear early in the development of the tumors. Next, they studied the SPOP protein in zebrafish (which is nearly identical to human SPOP), as well as in mouse and human cells. The experiments show that SPOP normally helps the cell to accurately repair DNA that has been damaged. Mutations in SPOP change the DNA repair process, which lead to genomic instability by increasing the likelihood that broken chromosomes will be stuck back together incorrectly. Further experiments tested drugs known as PARP inhibitors on mouse and human prostate cancer cells. The drugs, which have been recently tested successfully in patients with prostate cancer, block a different method of DNA repair that operates separately to the one that involves SPOP. When both of these pathways were inactivated—one by the SPOP mutation, the other by the drug—the cancer cells died more quickly. Therefore, men that are diagnosed with types of prostate cancer in which the gene that encodes SPOP is mutated might benefit from treatment with PARP inhibitors or other therapies that affect DNA repair.

Somatic copy number alterations (SCNAs) are a pervasive trait of human cancers that contributes to tumorigenesis by affecting the dosage of multiple genes at the same time. In the past decade, The Cancer Genome Atlas (TCGA) and the International Cancer Genome Consortium (ICGC) initiatives have generated and made publicly available SCNA genomic profiles from thousands of tumor samples across multiple cancer types. Here, we present a comprehensive analysis of 853,218 SCNAs across 10,729 tumor samples belonging to 32 cancer types using TCGA data. We then discuss current models for how SCNAs likely arise during carcinogenesis and how genomic SCNA profiles can inform clinical practice. Lastly, we highlight open questions in the field of cancer-associated SCNAs.

Introduction: Aneuploidy is a cancer hallmark that includes broad somatic copy-number alterations (SCNAs), being whole chromosome- or arm-level events, or smaller focal SCNAs. Pan-cancer studies suggest that tumor broad and focal SCNAs are linked to distinctive molecular/clinical traits, and broad SCNAs may potentially interfere with tumor immune infiltrates. However, the impact of SCNA genomic loads in hepatocellular carcinoma (HCC) is still unresolved. Here we have assessed broad and focal SCNA burdens in HCC to unveil associations with clinic-molecular characteristics and immune cell profiles. Method: The study includes 520 paired tumor/adjacent surgically resected HCC samples: 150 of a training cohort (HEPTROMIC) and 370 of a validation cohort (TCGA). Tumor ploidy and SCNA segments were extracted from SNP array data using ASCAT and SAASCNV. We applied the CNApp tool (bioinfo.ciberehd.org/CNApp) to extract a Broad SCNA Score (BCS) and Focal SCNA Score (FCS) to assess broad and focal SCNA loads of each sample. Broad and focal alterations were defined as those spanning ≥50% and <50% of a chromosome arm, respectively. Subsequently, the scores were integrated with a) gene expression data, b) clinic-pathological data, and c) the composition of the tumor immune infiltrate, determined using the Immunophenoscore. Results: HCC tumors characterized by a low BCS (25% of Heptromic, 15% of TCGA) were associated with the HCC Immune class and up-regulation of genes related to inflammation, active infiltrate signaling, antigen presentation and cytolytic activity (FDR<0.1, p<0.05). Conversely, tumors with high BCS (25% in Heptromic, 45% in TCGA) were linked to polyploidy and TP53 loss of function, were enriched in proliferation and DNA repair gene signatures and presented up-regulation of genes from immune suppressor cells and cytokines. On the other hand, while tumors with high FCS (25% in Heptromic, 49% in TCGA) were associated with TP53 loss of function, up-regulation of genes related to proliferation and progenitor cells and gene expression signatures suggesting increased tumor aggressiveness. Conversely, low-intermediate FCS tumors displayed higher β-catenin pathway activity, with enrichment in CTNNB1 mutations. FSS was not associated with immunity. Conclusions: Broad are more informative than focal SCNA burdens in terms of molecular features and immune status of HCC tumors. Those tumors characterized by chromosomal stability (low broad SCNAs loads) are enriched in antitumor immune response and antigenicity traits and therefore might correspond to those tumors responding to checkpoint inhibitors. Citation Format: Roger Esteban-Fabró, Laia Bassaganyas, Sara Torrecilla, Agrin Moeini, Sebastià Franch-Expósito, Maria Vila-Casadesús, Ferran Nadeu, Daniela Sia, Itziar Salaverria, Laia Cabellos, Roser Pinyol, Jordi Camps, Vicenzo Mazzaferro, Vicenzo Mazzaferro, Josep M Llovet. Aneuploidy profiles in hepatocellular carcinoma and their impact on tumor progression and immune features [abstract]. In: Proceedings of the American Association for Cancer Research Annual Meeting 2019; 2019 Mar 29-Apr 3; Atlanta, GA. Philadelphia (PA): AACR; Cancer Res 2019;79(13 Suppl):Abstract nr 3095.

BackgroundWhile RB1 loss initiates retinoblastoma development, additional somatic copy number alterations (SCNAs) can drive tumor progression. Although SCNAs have been identified with good concordance between studies at a cytoband resolution, accurate identification of single genes for all recurrent SCNAs is still challenging. This study presents a comprehensive meta-analysis of genome-wide SCNAs integrated with gene expression profiling data, narrowing down the list of plausible retinoblastoma driver genes.MethodsWe performed SCNA profiling of 45 primary retinoblastoma samples and eight retinoblastoma cell lines by high-resolution microarrays. We combined our data with genomic, clinical and histopathological data of ten published genome-wide SCNA studies, which strongly enhanced the power of our analyses (N = 310).ResultsComprehensive recurrence analysis of SCNAs in all studies integrated with gene expression data allowed us to reduce candidate gene lists for 1q, 2p, 6p, 7q and 13q to a limited gene set. Besides the well-established driver genes RB1 (13q-loss) and MYCN (2p-gain) we identified CRB1 and NEK7 (1q-gain), SOX4 (6p-gain) and NUP205 (7q-gain) as novel retinoblastoma driver candidates. Depending on the sample subset and algorithms used, alternative candidates were identified including MIR181 (1q-gain) and DEK (6p gain). Remarkably, our study showed that copy number gains rarely exceeded change of one copy, even in pure tumor samples with 100% homozygosity at the RB1 locus (N = 34), which is indicative for intra-tumor heterogeneity. In addition, profound between-tumor variability was observed that was associated with age at diagnosis and differentiation grades.InterpretationSince focal alterations at commonly altered chromosome regions were rare except for 2p24.3 (MYCN), further functional validation of the oncogenic potential of the described candidate genes is now required. For further investigations, our study provides a refined and revised set of candidate retinoblastoma driver genes.

Thanks to a never-before detailed view of the human genome, the last decade has brought to light the notion of DNA copy number variation (CNV) as the pivotal force contributing to population genomic diversity and evolution. It is as well clear now that cancer typically results in loosened control over genomic integrity and that the acquisition of somatic copy number alterations (SCNAs), whether confined to specific genes or affecting entire chromosome arms, is likely to be a fundamental prerequisite to the adaptive pressure that drives oncogenesis. This review gives a brief overview of key developments in genome-wide SCNA profiling, with specific emphasis on array-based techniques and deep-sequencing, which indeed enabled us to identify the large majority of genomic regions undergoing frequent alteration in human cancers and defining recognizable clinical phenotype. Alongside with the prospective to take advantage for future personalized precision medicine, high-throughput SCNA analysis have already proven diagnostic and prognostic potential, particularly for those clinically unpredictable and therapy-refractory tumors, such us cutaneous melanoma. Biomedical Reviews 2013; 24: 11-24.

Background: Characterization of focal somatic copy number alterations (SCNAs) has led to the identification of many cancer genes, yet similar investigations of arm-level SCNAs remain challenging. The identity of driver genes within these broad SCNAs remains a critical unanswered question in cancer genetics. One of the most frequent arm-level SCNAs is 9p loss, which contains the tumor suppressor gene (TSG) CDKN2A. It is also believed that other TSGs exist on 9p, though their identity has yet to be revealed. Methods: We analyzed every arm-level SCNA in 28 cancer types from The Cancer Genome Atlas (TCGA) to clarify the relative impact of 9p loss across cancer. We also performed a multi-tiered genomic dissection of chromosome 9p in 540 patients from 3 independent lower grade glioma (LGG) datasets (TCGA, REMBRANDT, MSKCC) to pinpoint genetic loci that are tied to tumor aggressiveness and poor survival. Focal and arm-level SCNAs were determined by GISTIC2.0. As per recently proposed criteria, 3 LGG subtypes were clustered by IDH mutation and 1p/19q deletion status. Survival analyses were performed using log-rank tests or Cox regression. Results: We found that chromosome 9p loss is one of the most frequent and prognostic arm-level events across 28 TCGA cancer types. Of these, the strongest 9p survival association was found in LGG. We performed a large-scale associations test of 376 important clinical and molecular variables and revealed that 9p loss was only associated with 6q or 9q loss, and 9p loss alone was sufficient to predict poor prognosis. On subtype-specific analysis, 9p loss predicted worse overall survival (OS) in both IDH mutant LGG subtypes but not IDH wild-type (IDHwt). To identify underlying drivers on 9p, we identified 87 genes most frequently targeted by 9p loss. Additional genetic events were rare except for homozygous deletion (HD) at 9p21.3, which contains CDKN2A. In contrast, mRNA/miRNA expression at all gene loci was much more variable and poorly correlated to copy number status. Therefore, pan-LGG and subtype-specific gene expression analyses were used to identify several drivers of tumor aggressiveness, notably KLHL9, MTAP, PLAA, and PTPRD. Although CDKN2A HD was linked to worse OS in a pan-LGG analysis, this event significantly co-occurred with the LGG IDHwt subtype, a group with GBM-like survival outcomes. Further, CDKN2A copy number status and expression did not predict worse OS in any subtype. Discussion: We characterize the nature of 9p loss in LGG, pinpoint genes involved in worse survival, and redefine the role of the tumor suppressor CDKN2A. These results provide new critical insight into long-standing questions about the nature of chromosome 9p loss and establish a framework for examining other arm-level SCNAs in cancer. Citation Format: David M. Roy, Logan A. Walsh, Alexis Desrichard, Jianjiong Gao, Promita Bose, Jason T. Huse, William Lee, Timothy A. Chan. Multidimensional genomic dissection of chromosome 9p in glioma. [abstract]. In: Proceedings of the 106th Annual Meeting of the American Association for Cancer Research; 2015 Apr 18-22; Philadelphia, PA. Philadelphia (PA): AACR; Cancer Res 2015;75(15 Suppl):Abstract nr LB-169. doi:10.1158/1538-7445.AM2015-LB-169

The ubiquitin and ubiquitin-like protein modifications contribute to functional changes of substrate proteins and thus regulate a range of biological processes including cancer.1 Ubiquitin-fold modifier 1 (UFM1) is a newly identified ubiquitin-like modification with essential biological functions.2, 3 Like ubiquitination, UFM1 modification (UFMylation) is catalyzed by the dynamic E1–E2–E3 enzymatic reaction. Unlike ubiquitination which encompasses several E1s and different E2s (>50) and E3s (>600), only one of each enzyme, UBA5 (E1-like), UFC1 (E2-like), and UFL1 (E3-like), has been identified for the UFMylation reaction.2, 3 The proteases UFM1-specific cysteine proteases 1 (UFSP1) and 2 (UFSP2) execute the maturation of the UFM1 precursor and de-UFMylation process.2, 3 In addition, the satellite components DDRGK1 and CDK5RAP3 are considered as key regulators of the UFMylation system.3-5 Recent studies suggest that UFMylation plays critical roles in diverse cellular processes.3 However, the roles of UFMylation in tumorigenesis have not been systematically explored. In this study, we comprehensively analysed the genomic alterations of these eight UFMylation family genes across the Cancer Genome Atlas (TCGA) data cohort. (Data S1). We performed GISTIC2.0 analysis and identified a total of 55 recurrent and focal somatic copy number alterations (SCNA) events in UFMylation family genes across the whole TCGA cohort with 33 cancer types (Figure 1A). Among the UFMylation genes, UFSP2 was frequently deleted in 14 cancer types. We calculated the frequencies of copy number gain or loss of UFMylation genes in each cancer type (Figure 1B). We found UFSP2 (31%), UFM1 (31%) and UFL1 (28%) showed the highest average frequency for copy number loss, whereas UFC1 (34%), UFSP1 (34%) and DDRGK1 (30%) had the highest average alteration frequency for copy number gain (Figure 1B). Similar SCNAs patterns of the UFMylation genes were observed by using the ICGC Data Portal (Figure S1 and Data S2). Interestingly, we found that the UFSP2 copy number was mainly heterozygous loss in tumours and cancer cell lines (Figure S2), and homozygous deletions were rarely detected, indicating a critical biological function and possible haploinsufficiency of the UFSP2 gene (Figure 1A and Data S3). To further explore the SCNA patterns of UFMylation genes, we calculated the high-level alteration of SCNAs and found that 11.08% of TCGA samples have high-level copy number alterations in at least one of the eight genes. Noteworthy, the high-level amplifications and deletions of UFMylation genes appeared to be occurred in a pattern of mutually exclusive (Figure 1C), suggesting UFMylation genes share common biological functions. However, we noticed that most of UFMylation genes exhibit low frequencies of somatic mutations (<5%) and transcript fusions (<.1%) (Figure S3 and Data S4–S7). Gene expression profiles analysed based on the RNA sequencing data from TCGA database (Data S8 and S9) indicate that UFMylation genes were ubiquitously expressed (Figure S4). Although most UFMylation genes generally have low mutation frequencies in cancers, we found that UFSP2 is frequently mutated in colon adenocarcinoma and uterine corpus endometrial carcinoma. The high recurrent copy number loss and frequent somatic mutations of UFSP2 suggest that UFSP2 may function as a tumour suppressor. It has been previously demonstrated that knockdown UFSP2 promoted breast cancer cell growth and tumour formation, suggesting that UFSP2 is a tumour suppressor in breast cancer.6 In addition, we found that the levels of UFSP2 mRNA were significantly lower in 11 cancer types (Figure S5A). Using human tissue microarrays, we confirmed that the expression of UFSP2 was significantly reduced in cancer tissues (Figures 2A and S6). To further validate the potential tumour suppressor function of UFSP2 in colon cancer, we examined the effects of UFSP2 knockdown on cell growth of colon cancer cells HT29 and HCT116, and observed that knockdown UFSP2 expression significantly promoted growth rates of colon cancer cells and its anchorage-independent cell growth. (Figure 2B–G). Significantly, knockdown UFSP2 expression significantly promoted the growth of xenograft tumours from UFSP2 depleted HT29 cells (Figure 2H–J). In addition, we observed that total UFMyaltion levels were increased in both UFSP2 depleted cells and xenograft tumours (Figure S7). These findings suggested that genomic alterations of UFSP2 were associated with the functional involvement of UFMylation in human colon cancer. Furthermore, our GSEA analysis indicates that loss of UFSP2 is mainly associated with the pathways in DNA replication, cell cycle, spliceosome, ribosome and mismatch repair (Figure S5C–D and Data S10). This is in line with previous reports that the UFMylation is critically involved in DNA damage, cell cycle, and ribosome protein modification.3, 4, 7, 8 We confirmed that knockdown UFSP2 increased the expression of some marker genes in DNA replication (PCNA and MCM2), cell cycle (CDK4 and CCND1), and ribosome protein (RPL26) (Figure S5E). It has been reported that MCM2, CDK4, and PCNA are interacted with UFM1 and are potential targets of the UFMylation.7 Thus, we speculate that UFSP2 may contribute to the tumorigenesis by modulating key regulators of cell cycle, DNA replication or protein biogenesis through Ufmylation modification. Together, our experimental findings revealed that knockdown UFSP2 expression significantly promotes the growth of colon cancer cells both in vitro and in vivo, suggesting that UFSP2 is a potential tumour suppressor in colon cancers. Given that UFMylation genes exhibit a high frequency of SCNA with a mutually exclusive pattern in common adult cancers, this suggests that involvement of UFMylation in cancer may be cell type or tissue specific. In line with this, it has been previously reported that CDK5RAP3 may function as a tumour suppressor9 in HNSCC and as an oncogene in liver cancer.10 In summary, we found that UFMylation family genes have a high frequency of SCNAs. Especially, UFSP2 is recurrently and focally deletion in a total of 14 human common cancers with the highest alteration score. We further demonstrated that depleted UFSP2 expression significantly promotes the growth of tumour cells both in vitro and in vivo, suggesting that UFSP2 may function as a tumour suppressor in colon cancers. Our integrated genomic analysis and functional studies provide insights in understanding this new post-translational modifier in cancer and potential therapeutic opportunities. We are grateful to Dr. Shuo Mou for bioinformatics analysis assistance, and Dr. Yongkang Zou for helpful discussions. This work was supported by grants from the National Natural Science Foundation of China (31730020), and the Hangzhou Science and Technology Bureau (20182014B01). Please note: The publisher is not responsible for the content or functionality of any supporting information supplied by the authors. Any queries (other than missing content) should be directed to the corresponding author for the article.

Endometrial serous carcinoma (ESC) represents the most aggressive subtype of endometrial carcinoma (EC). According to The Cancer Genome Atlas (TCGA), ESC exhibits a genomic profile characterised by frequent TP53 mutations and somatic copy-number alterations (SCNA). Several studies have suggested the role of intratumour heterogeneity (ITH) in tumour progression and therapy resistance, highlighting ITH as a challenge for personalised medicine. ITH is described as the co-existence of clonal and subclonal cellular populations within a single tumour. To date, the extent and prevalence of ITH in ESC have not been fully evaluated. The aim of this study was to address ITH analysis in ESC. We performed a descriptive integrated molecular approach using targeted sequencing and multiplex ligation-dependent probe amplification (MLPA) to identify mutations and SCNA patterns, respectively. Eight ESC were examined, selecting three tumour regions per case and their corresponding normal tissue. For targeted sequencing a gene panel of 40 genes based on TCGA and other survey data was performed. For MLPA different probe mixes were used to detect SCNA in 106 genes. Analysis of mutations and SCNA were performed in each sample and comparative analysis of the three tumour regions was also conducted. Targeted sequencing showed that mutations in TP53, PIK3CA and PPP2R1A were ubiquitous in all tumour regions. Moreover, MLPA results demonstrated a high frequency of SCNA, according to the already known presence of genomic instability in ESC. Unlike the homogeneous distribution of somatic mutations, SCNA exhibited ITH affecting targetable genes such as ERBB2. Our study suggests that somatic gene copy-number alterations are the main source of ITH in ESC.

Pan-cancer studies of somatic copy number alterations (SCNAs) have demonstrated shared SCNAs across cancer types, but whether these shared SCNAs vary by race-ethnicity has not been explored. Utilizing data from The Cancer Genome Atlas (TCGA), we identified SCNAs in breast and prostate tumors, two cancers with racially-disparate outcomes, and then tested for differences in SCNA magnitude by self-reported African American and European American race-ethnicity, as well as by regional chromosomal African ancestry within African Americans. GISTIC2 was applied to high density SNP array data to map SCNA regions in 712 European and 174 African American female breast tumors and 267 European and 42 African American prostate tumors derived from the TCGA dataset. For each tumor, SCNA magnitude was quantified by the area under the logarithm-base 2 copy number curve, and the germline ancestral origin of SCNAs was inferred using RFMix. A linear model was used to assess the association between SCNA magnitude and race-ethnicity (or regional African ancestry) while adjusting age-at-diagnosis and tumor severity. Race-differentiated SCNAs common to breast and prostate were found at chromosomes 5q11-21, 6q12-14, 6q16-22, 8q21-24, 11q22, 13q12-21, and 16q21-24, with 8q21-24 being the only amplification. African American breast and prostate tumors had higher magnitude alterations in the regions on 5q11-21, 8q21-24, 11q22, and 13q12-21, and among African Americans, this higher magnitude at 8q21-24 and 13q12-21 was consistent with increasing regional African ancestry. Within these regions with higher magnitude SCNAs in African Americans, expression analysis revealed 18 cancer genes, including RB1 and PVT1, differentially expressed by race-ethnicity in both tumors types that were consistent with the observed SCNA differences. While differences in SCNAs by race-ethnicity have been studied in single cancers, this is the first study to identify race-differentiated SCNAs shared by two hormonally-driven cancers and to explore the potential of germline genetic ancestry as a mechanism leading to this differentiation. The differentially expressed genes within SCNAs common to both tumor types could provide further insight into the racially disparate outcomes in breast and prostate cancers. Citation Format: Yalei Chen, Jia Li, Sudha Sadasivan, Ruicong She, Indrani Datta, Dhananjay Chitale, Nilesh Gupta, Melissa B. Davis, Craig G. Rogers, Lisa A. Newman, Pamela L. Paris, Benjamin A. Rybicki, Albert M. Levin. Breast and prostate cancers harbor common somatic copy number alterations that consistently differ by race-ethnicity [abstract]. In: Proceedings of the American Association for Cancer Research Annual Meeting 2018; 2018 Apr 14-18; Chicago, IL. Philadelphia (PA): AACR; Cancer Res 2018;78(13 Suppl):Abstract nr 4231.

BackgroundSomatic copy number alterations (SCNAs) are an important class of genomic alteration in cancer. They are frequently observed in cancer samples, with studies showing that, on average, SCNAs affect 34% of a cancer cell’s genome. Furthermore, SCNAs have been shown to be major drivers of tumour development and have been associated with response to therapy and prognosis. Large-scale cancer genome studies suggest that tumours are driven by somatic copy number alterations (SCNAs) or single-nucleotide variants (SNVs). Despite the frequency of SCNAs and their clinical relevance, the use of genomics assays in the clinic is biased towards targeted gene panels, which identify SNVs but provide limited scope to detect SCNAs throughout the genome. There is a need for a comparably low-cost and simple method for high-resolution SCNA profiling.ResultsWe present conliga, a fully probabilistic method that infers SCNA profiles from a low-cost, simple, and clinically-relevant assay (FAST-SeqS). When applied to 11 high-purity oesophageal adenocarcinoma samples, we obtain good agreement (Spearman’s rank correlation coefficient, rs=0.94) between conliga’s inferred SCNA profiles using FAST-SeqS data (approximately £14 per sample) and those inferred by ASCAT using high-coverage WGS (gold-standard). We find that conliga outperforms CNVkit (rs=0.89), also applied to FAST-SeqS data, and is comparable to QDNAseq (rs=0.96) applied to low-coverage WGS, which is approximately four-fold more expensive, more laborious and less clinically-relevant. By performing an in silico dilution series experiment, we find that conliga is particularly suited to detecting SCNAs in low tumour purity samples. At two million reads per sample, conliga is able to detect SCNAs in all nine samples at 3% tumour purity and as low as 0.5% purity in one sample. Crucially, we show that conliga’s hidden state information can be used to decide when a sample is abnormal or normal, whereas CNVkit and QDNAseq cannot provide this critical information.ConclusionsWe show that conliga provides high-resolution SCNA profiles using a convenient, low-cost assay. We believe conliga makes FAST-SeqS a more clinically valuable assay as well as a useful research tool, enabling inexpensive and fast copy number profiling of pre-malignant and cancer samples.

Clinical Impact of Somatic Copy Number Alterations in Circulating Tumor DNA from Diverse Lymphoma Subtypes

ABSTRACTCancer stem cells (CSCs) are considered to be responsible for tumorigenesis and cancer relapse. EpCAMhighCD44+ tumor cells are putative colorectal CSCs that express high levels of stem cell genes, while the EpCAMhighCD44− population mostly contains differentiated tumor cells (DTCs). This study aims to determine whether single CSC (EpCAMhighCD44+) and DTC (EpCAMhighCD44−) can be distinguished in terms of somatic copy number alterations (SCNAs). We applied fluorescence-activated cell sorting to isolate the CD45−EpCAMhighCD44+ and CD45−EpCAMhighCD44− populations from two primary colon tumors, on which low-coverage single-cell whole-genome sequencing (WGS) was then performed ∼0.1x depth. We compared the SCNAs of the CSCs and DTCs at single-cell resolution. In total, 47 qualified single cells of the two populations underwent WGS. The single-cell SCNA profiles showed that there were obvious SCNAs in both the CSCs and DTCs of each patient, and each patient had a specific copy number alteration pattern. Hierarchical clustering and correlation analysis both showed that the SCNA profiles of CSCs and DTCs from the same patient had similar SCNA pattern, while there were regional differences in the CSCs and DTCs in certain patient. SCNAs of CSCs in the same patient were highly reproducible. Our data suggest that major SCNAs occurred at an early stage and were inherited steadily. The similarity of ubiquitous SCNAs between the CSCs and DTCs might have arisen from lineage differentiation. CSCs from the same patient had reproducible SCNA profiles, indicating that gain or loss in certain chromosome is required for colon cancer development.

African Americans (AAs) tend to have more aggressive diseases and worse outcomes for many cancer types compared to European Americans (EAs). Large structural changes on chromosomes, such as somatic copy number alterations (SCNAs), are commonly present in many cancers and associated with clinical outcomes. While studies focusing on single tumor types have identified racial differences in SCNA frequencies and their possible association with cancer outcomes, pan-cancer race-differentiated SCNAs and their clinical implications have not been fully explored. Focusing on two of the most common hormonally driven cancers, breast cancer in women and prostate cancer in men, and using data from The Cancer Genome Atlas (TCGA), we first identified recurrent SCNAs using GISTIC2 in AAs and EAs separately within each cancer. For each GISTIC2 identified SCNA region, we calculated the magnitude deviation from the expected null value (i.e. copy number (CN) = 2) using the area under the log2(CN) (cnAUC) for each tumor. Modeling cnAUC as the outcome variable, we identified SCNAs that differed between AAs and EAs in breast cancers (n=58 SCNAs; permutation p&amp;lt;10-4) and prostate cancers (n=78 SCNAs; permutation p=0.006). Six of nine race-differentiated SCNAs common to breast and prostate cancers had consistent magnitude differences by race across both tumor types, and all six were of higher magnitude in AAs. These six race-differentiated SCNAs were found at chromosomes 5q11.2-q14.1, 5q15-q21.1, 8q21.11-q21.13, 8q21.3-q24.3, 11q22.3, and 13q12.3-q21.3 with the chromosome 8q regions being the only amplifications. Unsupervised hierarchical clustering of the SCNAs in these six race-differentiated regions identified three patient groups in each tumor type with significant survival differences. Of particular importance, within triple negative breast cancers, these SCNA patient groups showed the most significant difference in progression-free survival (p=0.019). While there was a higher frequency of AAs among the SCNA-defined patient groups with worse survival, no significant racial difference in survival was observed within each SCNA-defined patient group. This observation supports the idea that racial disparities in breast and prostate cancer outcomes might be in part due to racial differences in tumor biology that drive structural chromosomal changes shared across these common tumor types. Our ability to derive SCNA defined patient groups associated with survival based on race-differentiated genomic aberrations demonstrates both their clinical relevance and the need to understand the biologic mechanisms that give rise to them. Citation Format: Yalei Chen, Sudha M Sadasivan, Ruicong She, Indrani Datta, Kanika Taneja, Chitale Dhananjay, Nilesh Gupta, Melissa B Davis, Lisa A Newman, Craig G Rogers, Pamela L Paris, Jia Li, Benjamin A Rybicki, Albert M Levin. Breast and prostate cancers harbor common somatic copy number alterations that consistently differ by race and are associated with survival [abstract]. In: Proceedings of the Twelfth AACR Conference on the Science of Cancer Health Disparities in Racial/Ethnic Minorities and the Medically Underserved; 2019 Sep 20-23; San Francisco, CA. Philadelphia (PA): AACR; Cancer Epidemiol Biomarkers Prev 2020;29(6 Suppl_2):Abstract nr B066.

A powerful way to discover key genes playing causal roles in oncogenesis is to identify genomic regions that undergo frequent alteration in human cancers. Here, we report high-resolution analyses of somatic copy-number alterations (SCNAs) from 3131 cancer specimens, belonging largely to 26 histological types. We identify 158 regions of focal SCNA that are altered at significant frequency across multiple cancer types, of which 122 cannot be explained by the presence of a known cancer target gene located within these regions. Several gene families are enriched among these regions of focal SCNA, including the BCL2 family of apoptosis regulators and the NF-κB pathway. We show that cancer cells harboring amplifications surrounding the MCL1 and BCL2L1 anti-apoptotic genes depend upon expression of these genes for survival. Finally, we demonstrate that a large majority of SCNAs identified in individual cancer types are present in multiple cancer types.

5511 Background: Endometrial cancer is classified by tumor stage, histologic subtype and grade. However, a substantial proportion of presumed non-high risk cases recur, supporting the need for improved tools of prognostication. Methods: Using clinical and Affymetrix SNP 6.0 data from The Cancer Genome Atlas (TCGA) endometrial carcinoma project, we identified 4 somatic copy number alteration (SCNA) subtypes, established their prognostic value and validated them in an independent, population-based cohort from Norway. Patients had endometrioid, uterine papillary serous carcinoma (UPSC) or mixed histology tumors. Progression-free survival (PFS) was defined as time from diagnosis to recurrence or progression, and estimated by the Kaplan-Meier method. Results: Four groups of SCNA patterns were identified using hierarchical clustering: low SCNA, moderate SCNA, SCNA dominated by 1q amplification (1q amplified) and high SCNA level (serous-like). Their prognostic value was assessed in all TCGA patients (N = 292) and in a low risk subset with endometrioid histology, stage 1 disease (N = 210). In the full TCGA cohort, patients with low SCNA (reference group) had excellent 2-year PFS of 94%, while for moderate SCNA it was 84% (hazard ratio[HR] 2.7, p = .08). The 1q amplified and serous-like groups had significantly worse outcomes with 2-year PFS of 74% (HR 5.9, p= .002) and 74% (HR 6.0, p &lt;.001), respectively. On multivariable analysis, adjusting for variables including stage and grade, 1q amplified and serous-like SCNA patterns remained independently prognostic (respectively, adjusted HR 6.2, p = .002 and 4.7, p = .02). Similar results were found in the low risk subset. The prognostic value of the SCNA patterns was validated in an independent group of patients with low risk disease (N = 57). 5-year PFS was 91% for low SCNA, 83% for moderate SCNA (HR 2.0, p = .58), 72% for 1q amplified (HR 3.7, p = .11) and 50% for the serous-like SCNA group (HR 6.7, p = .04). Conclusions: Four subtypes of DNA SCNA patterns in endometrial cancer were identified and validated to be prognostic of outcome. These novel biomarkers may be useful in guiding therapeutic decisions, and shed insight on the biology of more, or less, aggressive endometrial cancer.