Landscape Of Genomic Alterations Research Articles

Clinically advanced renal cell carcinoma (RCC) and renal sarcoma (RSC) in young patients: A comprehensive genomic profiling (CGP) study.

5066 Background: We queried whether the landscape of genomic alterations (GA) would differ in patients with metastatic RCC under 40 years of age (under40) and patients 40 years of age or older (over40). Methods: FFPE tissues from 2,128 clinically advanced RCC and 25 RSC underwent hybrid-capture based CGP to evaluate all classes of GA. Samples were classified at time of sequencing as the following RCC subtypes: clear cell (ccRCC), papillary (papRCC), chromophobe (chrRCC), medullary (medRCC), collecting duct (cdRC), sarcomatoid (sarcRCC) and NOS (nosRCC). Tumor mutational burden (TMB) was determined on up to 1.1 Mbp of sequenced DNA. Tumor cell PD-L1 expression was determined by IHC (Dako 22C3). Results: The male preponderance increased in the over40 patients. The GA/tumor increased in the over40 cohorts except for medRCC. Similarly, TMB was consistently higher in over40 groups. MSI high status was virtually absent. PD-L1 expression, available only in small subsets, was generally absent although 44% high positive staining in sarcRCC was noteworthy. Differences in GA in under40 vs over40 RCC were seen and included increased PBRM1 and SETD2 GA in over40 vs under40 ccRCC; increased C DKN2A/B and TERT and decreased FH GA in over40 vs under40 papRCC; increased TP53 and decreased VHL, BAP1, SETD2 and PTEN in over40 vs under40 chrRCC; increased TP53, PTEN and TERT GA with decreased NF2 GA in over40 vs under 40 sarcRCC; and increased TP53, VHL and TERT in over40 vs under40 nosRCC. Changes in GA in under40 vs over40 medRCC, cdRCC and RSC were noted but insufficient cases prevented further evaluation. Conclusions: When separately evaluated by under/over 40 years of age, CGP of clinically advanced RCC demonstrates differences in genomic landscapes with over40 cases featuring increasing male preponderance, higher GA/tumor, higher TMB and increases in a variety of GA. These findings may play important roles in the planning of future clinical trials designed to personalize the treatment of metastatic RCC. [Table: see text]

Comprehensive profiling of genomic in 80 Chinese patients with bladder cancer.

e17024 Background: Bladder cancer (BLCA) is the ninth most common malignant disease and the thirteenth most common cause of cancer death worldwide, To understand the genomic landscape of Chinese BLCA patients, 80 cases were analyzed by comprehensive genomic profiling (CGP) for genomic alterations and genome wide signatures. Methods: Formalin Fixed Paraffin Embedded (FFPE) tumor and matched blood samples of 80 Chinese BLCA patients were obtained for next-generation sequencing (NGS)-based 642 cancer genes panel assay and bladder cancer genomic profiling was evaluated. Results: Collectively, in our Chinese BLCA cohort, multiple histological subtypes, including Non-muscle invasive bladder cancer (NMIBC) (20/80, 25%), muscle invasive bladder cancer (MIBC) (49/80, 61.25%), and other subtypes (11/80, 13.75%). The top ranked somatic gene alterations in Chinese BLCA patients were TP53 (49%), TERT (34%), KMT2D (33%), KMD6A (25%), CDKN2A (23%), FGFR3 (23%), FLG (19%), PIK3CA (19%), CCDN1 (18%), RB1 (18%). Some FGFR2/3 gene fusions were detected such as FGFR2-WDR11 and FGFR3-TACC3. Interestingly, some co-mutations were identified in our cohort, such as ERBB3 was co-mutated with MUC17, FAT1 was co-mutated with RB1 etc. (p < 0.01). Out of 15 most common canonical pathways, potentially targetable genomic alterations were mainly identified in RTK-RAS (74.68%), DNA damage response (53.16%), TP53 (53.16%),chromatin remodeling (51.90%), PI3K (44.30%), NOTCH (41.77%), Cell Cycle(32.91%), Hippo (29.11%) and WNT (29.1%) pathway. Additionally, chromatin remodeling and NOTCH pathways were commonly co-altered with MYC pathway, TP53、Hippo and NOTCH pathways were significantly co-altered with DNA damage response pathway. Across samples, 33 Chinese BLCA patients had high tumor mutational burden (TMB-H) ( > 10 mutations/Mb). Only 2 Chinese patients had high microsatellite instability (MSI-H). Conclusions: Our results displayed the landscape of genomic alterations in 80 Chinese BLCA patients. The genomic alterations identified in our study may provide an opportunity to discover potential strategies for targeted and immunotherapy in BLCA. Future studies should account for these genomic alterations.

Comprehensive genomic profiling of 216 Chinese patients with renal cell carcinoma.

3532 Background: Renal cell carcinoma (RCC), a global public health problem, has exhibited a gradual rise in incidence. Unfortunately, the scarcity of effective biomarkers in the clinical became a major limitation of the progress of biological therapies. Therefore, it is imperative to accurately comprehend RCC genomic profiling for exploring its clinical treatment strategies. Methods: Formalin Fixed Paraffin Embedded (FFPE) tumor and matched blood samples of 216 Chinese RCC patients were obtained for next-generation sequencing (NGS)-based 620 cancer genes panel assay and RCC genomic profiling was evaluated. Results: In our Chinese RCC cohort, multiple histological subtypes, encompassing clear cell (96/216, 44.44%), papillary (14/216, 6.48%), chromophobe (2/216, 0.93%), and undefined subtypes (104/216, 48.15%) were included. The top ranked genomic alterations in Chinese RCC patients were VHL (45.83%), PBRM1 (17.1%), BAP1(13.89%), TP53 (10.65%), SETD2 (9.29%), MTOR (8.67%), ARID1A (5.6%), PTEN (5.09%). Interestingly, BAP1, PBRM1 were co-mutated with VHL, and MET, NF2 were mutually exclusive with VHL (all p < 0.05). Of these patients, 87.9% (190/216) of RCC patients had at least one genomic alteration, indicating the potential clinical benefits of targeted therapies. Out of 15 most common canonical pathways, potentially targetable genomic alterations were mainly identified in HIF (45.83%), chromatin remodeling (42.59%), PI3K/AKT (24.07%), DNA damage response (19.44%), RTK-RAS (18.06%), TP53 (15.74%), NOTCH (9.72%), Hippo (9.72%), and WNT (5.56%) pathway. Additionally, HIF pathway was commonly co-altered with chromatin remodeling pathway, and WNT and NOTCH pathways were significantly co-altered with TP53 pathway. Rare mutation types such as CDKN2A-DMRTA1 and HOOK1-ALK were also detected. Across samples, the median TMB was 2.2 (0-18.6) mutations/Mb. Only 2 Chinese RCC patients had TMB-high (> 10 mutations/Mb). Conclusions: Our results displayed the landscape of genomic alterations in 216 Chinese RCC patients. The genomic alterations identified in our study may provide an opportunity to discover potential strategies for targeted and immunotherapy in RCC. Future studies should account for these genomic alterations.

Genomic profiling of breast cancer brain metastases reveals targetable alterations.

2525 Background: Genomic characterization of breast cancer brain metastases (BCBMs) has thus far been limited. The objective of this study was to describe the landscape of genomic alterations in patients (pts) with BCBMs. Methods: Targeted next-generation DNA sequencing of > 300 cancer-related genes (OncoPanel) was prospectively performed on primary and metastatic (met) tumors in 321 pts with a diagnosis of BCBM between August 2016 and April 2019 at Dana-Farber Cancer Institute (table). Enrichment analysis of genomic alterations was performed using a two-sided Fisher exact test and differences in tumor mutation burden (TMB) between groups were assessed using two-sided Mann-Whitney U test. Multiple comparison correction was performed using the Benjamini-Hochberg procedure. Results: All subtypes were represented in BCBM (25 HR+/HER2-; 24 HR+/HER2+; 27 HR-/HER2+; 18 TNBC; 5 unknown; n = 99) and extracranial (EC) samples: (96 HR+/HER2-; 32 HR+/HER2+; 22 HR-/HER2+; 41 TNBC; 31 unknown; n = 222). BCBMs were found most commonly to have mutations or copy number alterations in TP53, ERBB2, PIK3CA, GATA3, PTEN, ESR1, CDH1, BRCA2, ARID1A, BRCA1 (>5% frequency, table). Two pts acquired ERBB2 amplification (amp) between the matched primary breast sample and brain met. In pair-wise comparisons of BCBMs to unmatched primaries or EC mets, only ERBB2 amp was significantly enriched (table, † = adjusted p < 0.05). There was no significant difference in TMB between BCBM and EC mets (median 9.12 vs 7.26, p = 0.15). In contrast, TMB was significantly higher in BCBMs compared to unmatched primaries (median 9.12 vs 7.26, p=0.005). Conclusions: BCBMs display similar mutations and copy number alterations compared to primary tumors and EC mets in pts with BCBM. These data suggest that BCBMs contain actionable genomic alterations that are most often also reflected in EC disease. Alterations in ERBB2, PIK3CA/PTEN, and BRCA1/2 represent potentially targetable alterations in pts with BCBM. [Table: see text]

The landscape of genomic alterations detected in serial circulating tumor DNA (ctDNA) in clinical progressive metastatic breast cancer.

1084 Background: Metastatic breast cancer (MBC) is associated with genomic evolution, representing a challenge at clinical progression. While tissue and blood next-generation sequencing (NGS) allows for the baseline detection of alterations, non-invasive longitudinal assessment of ctDNA can provide a tool for monitoring tumor evolution. We characterized genomic changes using serial ctDNA testing in patients with clinical progression. Methods: Patient data was obtained under an IRB-approved protocol and ctDNA was collected at Northwestern University between 2015-2019. All ctDNA samples were analyzed using the Guardant360 NGS assay. Of 255 patients with MBC, 86 had at least two serial ctDNA collections with the second collection drawn at first progression (P1) by imaging and clinical assessment. Participants were followed until second clinical progression (P2). We analyzed type of alterations, mutant allele frequency (MAF), number of alterations (NOA), and sites of disease on imaging in close proximity to ctDNA evaluation. Matched pairs variations in MAF and NOA at P1 and P2 were tested through Wilcoxon test. Results: We identified 44 HR+, 20 HER2+ and 22 TNBC cases. Median lines of therapy were 3 (interquartile range (IQR): 1-6) for HR+, 3 (IQR: 1-5) for HER2+, and 2 (IQR: 1-4) for TNBC. The most likely alterations between baseline to P1 were TP53 (p < 0.0075), PIK3CA (p < 0.0126), AR (p < 0.0126), FGFR1 (p < 0.0455) and ESR1 (p < 0.0143). In the HR+ subset , ESR1 was statistically more likely at P1. ESR1 at P1 was also associated with development of new liver lesions (p < 0.0320). ERBB2 mutation at P1 was associated with new lung (p < 0.0050) or bone lesions (p < 0.0030). Increase in NOA was observed between baseline and P1 (p < 0.0001), P1 and P2 (p < 0.0001), and baseline to P2 (p < 0.0004). MAF was increased between baseline and P2 (p < 0.0480). Conclusions: Serial ctDNA testing identified resistance alterations ( TP53, PIK3CA, AR, ESR1, FGFR1), with some mutations indicating new sites of disease ( ESR1, ERBB2). Heterogeneity of ctDNA was significantly associated with progressive disease. Prospective evaluation of the impact of serial ctDNA testing on treatment decisions is needed to expand the role of precision medicine in MBC. [Table: see text]

PD47-01 GENOMIC DETERMINANTS OF NESTED VARIANT UROTHELIAL CARCINOMA

You have accessJournal of UrologyBladder Cancer: Basic Research & Pathophysiology IV (PD47)1 Apr 2020PD47-01 GENOMIC DETERMINANTS OF NESTED VARIANT UROTHELIAL CARCINOMA Timothy Clinton*, Karissa Whiting, Vitor Silva, Qiang Li, Nima Almassi, Aditya Bagrodia, Eugene Pietzak, Guido Dalbagni, Gopa Iyer, Bernard Bochner, Irina Ostrovnaya, Anuradha Gopalan, Yingbei Chen, Samson Fine, Satish Tickoo, Maria Arcila, Victor Reuter, David Solit, and Hikmat Al-Ahmadie Timothy Clinton*Timothy Clinton* More articles by this author , Karissa WhitingKarissa Whiting More articles by this author , Vitor SilvaVitor Silva More articles by this author , Qiang LiQiang Li More articles by this author , Nima AlmassiNima Almassi More articles by this author , Aditya BagrodiaAditya Bagrodia More articles by this author , Eugene PietzakEugene Pietzak More articles by this author , Guido DalbagniGuido Dalbagni More articles by this author , Gopa IyerGopa Iyer More articles by this author , Bernard BochnerBernard Bochner More articles by this author , Irina OstrovnayaIrina Ostrovnaya More articles by this author , Anuradha GopalanAnuradha Gopalan More articles by this author , Yingbei ChenYingbei Chen More articles by this author , Samson FineSamson Fine More articles by this author , Satish TickooSatish Tickoo More articles by this author , Maria ArcilaMaria Arcila More articles by this author , Victor ReuterVictor Reuter More articles by this author , David SolitDavid Solit More articles by this author , and Hikmat Al-AhmadieHikmat Al-Ahmadie More articles by this author View All Author Informationhttps://doi.org/10.1097/JU.0000000000000934.01AboutPDF ToolsAdd to favoritesDownload CitationsTrack CitationsPermissionsReprints ShareFacebookLinked InTwitterEmail Abstract INTRODUCTION AND OBJECTIVE: Nested variant urothelial carcinoma (NVUC) is a historically aggressive variant of bladder cancer with management reliant on radical cystectomy given unclear effectiveness of chemotherapy. Given the aggressive behavior of NVUC, the identification of pathogenic and potentially targetable genomic alterations could suggest novel treatment paradigms for this rare histologic variant. METHODS: Query of our institutional database identified 81 cases of clinically localized NVUC. Each case was re-reviewed by GU pathologists to confirm the histology. Targeted capture sequencing was performed to define the spectrum of genomic alterations in known and putative cancer genes. Clinical and sequencing data was then compared between the nested variant cases and a prospective cohort of clinically localized high-grade urothelial carcinomas, not otherwise specified (UC NOS). RESULTS: On central re-review, tumors were classified as either classic NVUC (N=53) or high-grade urothelial carcinoma with nested features (N=28). A cohort of UC NOS patients (N=600) was used as a comparator. Median follow-up of the overall cohort was 4.3 years. Classic NVUC was associated with higher risk of cancer specific mortality compared to UC NOS on multivariate analysis (HR 1.76, p=0.04). Tumor mutational burden was lower in classic NVUC tumors versus UC NOS. Evaluation of 136 cancer-associated genes identified differences in mutational frequencies across the 3 cohorts (Table 1) with RhoA and FOXA1 alterations enriched in NVUC. CONCLUSIONS: Patients with NVUC have significantly higher risk of cancer specific mortality than patients with UC NOS. While there was significant overlap between the landscape of genomic alterations in NVUC and UC NOS tumors, NVUC tumors were enriched for RhoA and FOXA1 alterations among others. Source of Funding: This work was supported by the Sidney Kimmel Center for Prostate and Urologic Cancers, the Michael and Zena Wiener for Therapeutics Program in Bladder Cancer, Pin Down Bladder Cancer, Cycle for Survival, the Marie-Josee and Henry R. Kravis Center for Molecular Oncology, NIH/NCATS Grant Number UL1-TR002384, the National Cancer Institute Cancer Center Core Grant Number P30-CA008748 and by SPORE in Bladder Cancer P50-CA221745. © 2020 by American Urological Association Education and Research, Inc.FiguresReferencesRelatedDetails Volume 203Issue Supplement 4April 2020Page: e925-e925 Advertisement Copyright & Permissions© 2020 by American Urological Association Education and Research, Inc.MetricsAuthor Information Timothy Clinton* More articles by this author Karissa Whiting More articles by this author Vitor Silva More articles by this author Qiang Li More articles by this author Nima Almassi More articles by this author Aditya Bagrodia More articles by this author Eugene Pietzak More articles by this author Guido Dalbagni More articles by this author Gopa Iyer More articles by this author Bernard Bochner More articles by this author Irina Ostrovnaya More articles by this author Anuradha Gopalan More articles by this author Yingbei Chen More articles by this author Samson Fine More articles by this author Satish Tickoo More articles by this author Maria Arcila More articles by this author Victor Reuter More articles by this author David Solit More articles by this author Hikmat Al-Ahmadie More articles by this author Expand All Advertisement PDF downloadLoading ...

A landscape of genomic alterations at the root of a near-untreatable tuberculosis epidemic

BackgroundAtypical Beijing genotype Mycobacterium tuberculosis strains are widespread in South Africa and have acquired resistance to up to 13 drugs on multiple occasions. It is puzzling that these strains have retained fitness and transmissibility despite the potential fitness cost associated with drug resistance mutations.MethodsWe conducted Illumina sequencing of 211 Beijing genotype M. tuberculosis isolates to facilitate the detection of genomic features that may promote acquisition of drug resistance and restore fitness in highly resistant atypical Beijing forms. Phylogenetic and comparative genomic analysis was done to determine changes that are unique to the resistant strains that also transmit well. Minimum inhibitory concentration (MIC) determination for streptomycin and bedaquiline was done for a limited number of isolates to demonstrate a difference in MIC between isolates with and without certain variants.ResultsPhylogenetic analysis confirmed that two clades of atypical Beijing strains have independently developed resistance to virtually all the potent drugs included in standard (pre-bedaquiline) drug-resistant TB treatment regimens. We show that undetected drug resistance in a progenitor strain was likely instrumental in this resistance acquisition. In this cohort, ethionamide (ethA A381P) resistance would be missed in first-line drug-susceptible isolates, and streptomycin (gidB L79S) resistance may be missed due to an MIC close to the critical concentration. Subsequent inadequate treatment historically led to amplification of resistance and facilitated spread of the strains. Bedaquiline resistance was found in a small number of isolates, despite lack of exposure to the drug. The highly resistant clades also carry inhA promoter mutations, which arose after ethA and katG mutations. In these isolates, inhA promoter mutations do not alter drug resistance, suggesting a possible alternative role.ConclusionThe presence of the ethA mutation in otherwise susceptible isolates from ethionamide-naïve patients demonstrates that known exposure is not an adequate indicator of drug susceptibility. Similarly, it is demonstrated that bedaquiline resistance can occur without exposure to the drug. Inappropriate treatment regimens, due to missed resistance, leads to amplification of resistance, and transmission. We put these results into the context of current WHO treatment regimens, underscoring the risks of treatment without knowledge of the full drug resistance profile.

Genomic determinants of nested variant urothelial carcinoma.

538 Background: Nested variant urothelial carcinoma (NVUC) is a historically aggressive variant of bladder cancer with management reliant on radical cystectomy given unclear effectiveness of chemotherapy. Given the aggressive behavior of NVUC, the identification of pathogenic and potentially targetable genomic alterations could suggest novel treatment paradigms for this rare histologic variant. Methods: Query of our institutional database identified 81 cases of clinically localized NVUC. Each case was re-reviewed by GU pathologists to confirm the histology. Targeted capture sequencing was performed to define the spectrum of genomic alterations in known and putative cancer genes. Clinical and sequencing data was then compared between the nested variant cases and a prospective cohort of clinically localized high-grade urothelial carcinomas, not otherwise specified (UC NOS). Results: On central re-review, tumors were classified as either classic NVUC (N=53) or high-grade urothelial carcinoma with nested features (N=28). A cohort of UC NOS patients (N=600) was used as a comparator. Median follow-up of the overall cohort was 4.3 years. Classic NVUC was associated with higher risk of cancer specific mortality compared to UC NOS on multivariate analysis (HR 1.76, p=0.04). Tumor mutational burden was lower in classic NVUC tumors versus UC NOS. Evaluation of 136 cancer-associated genes identified differences in mutational frequencies across the 3 cohorts (Table) with RhoA and FOXA1 alterations enriched in NVUC. Conclusions: Patients with NVUC have significantly higher risk of cancer specific mortality than patients with UC NOS. While there was significant overlap between the landscape of genomic alterations in NVUC and UC NOS tumors, NVUC tumors were enriched for RhoA and FOXA1 alterations among others.[Table: see text]

Integrated characterisation of cancer genes identifies key molecular biomarkers in stomach adenocarcinoma

AimsGastric cancer is one of the leading causes for cancer mortality. Recent studies have defined the landscape of genomic alterations of gastric cancer and their association with clinical outcomes. However,...

Association of a novel circulating tumor DNA next-generating sequencing platform with circulating tumor cells (CTCs) and CTC clusters in metastatic breast cancer

PurposeLiquid biopsies, including circulating tumor DNA (ctDNA) and circulating tumor cells (CTCs), can be used to understand disease prognosis, tumor heterogeneity, and dynamic response to treatment in metastatic breast cancer (MBC). We explored a novel, 180-gene ctDNA panel and the association of this platform with CTCs and CTC clusters.MethodsA total of 40 samples from 22 patients with MBC were included in the study. For the primary analysis, all patients had ctDNA sequencing using the PredicinePLUS™ platform. CTCs and CTC clusters were examined using the CellSearch™ System. Clinical and pathological variables were reported using descriptive analyses. Associations between CTC count and specific genomic alterations were tested using the Mann-Whitney U test.ResultsOf 43 sequenced patients, 40 (93%) had at least one detectable genomic alteration with a median of 6 (range 1–22). Fifty-seven different genes were altered, and the landscape of genomic alterations was representative of MBC, including the commonly encountered alterations TP53, PTEN, PIK3CA, ATM, BRCA1, CCND1, ESR1, and MYC. In patients with predominantly hormone-receptor-positive MBC, the number of CTCs was significantly associated with alterations in ESR1 (P < 0.005), GATA3 (P < 0.05), CDH1 (P < 0.0005), and CCND1 (P < 0.05) (Mann-Whitney U test). Thirty-six percent of patients had CTC clusters, which were associated with alterations in CDH1, CCND1, and BRCA1 (all P < 0.05, Mann-Whitney U test). In an independent validation cohort, CTC enumeration confirmed significant associations with ESR1 and GATA3, while CTC clusters were significantly associated with CDH1.ConclusionsWe report on a novel ctDNA platform that detected genomic alterations in the vast majority of tested patients, further indicating potential clinical utility for capturing disease heterogeneity and for disease monitoring. Detection of CTCs and CTC clusters was associated with particular genomic profiles.

Circulating Tumor DNA Profiling of Advanced Biliary Tract Cancers.

Recent advances in molecular diagnostic technologies have allowed for the evaluation of solid tumor malignancies via noninvasive blood sampling, including circulating tumor DNA (ctDNA) profiling. We sought to characterize the ctDNA genomic alteration landscape in patients with biliary tract cancers (BTCs). From January 2015 to February 2018, 124 patients with BTC at the Mayo Clinic Comprehensive Cancer Center underwent ctDNA testing using a clinically available assay. The majority of samples (n = 122) were tested using the 73-gene panel that includes somatic genomic targets, including complete or critical exon coverage in 30 and 40 genes, respectively, and in some, amplifications, fusions, and indels. A total of 138 samples were included, with approximately 70% of patients having intrahepatic BTC. All patients had locally advanced or metastatic BTC. Samples with one or more alterations, when variants of unknown significance were excluded, numbered 105 (76%). Each sample contained, on average, three alterations with a median allelic fraction of 0.52%. The overall landscape of alterations is summarized in Figures 1 and 2. After excluding variants of unknown significance, therapeutically relevant alterations were observed in 76 patients (55%), including BRAF mutations, ERBB2 amplifications, FGFR2 fusions, FGFR2 mutations, and IDH1 mutations seen in 21% of patients. A different spectrum of alterations was observed in patients with early-onset BTC (younger than age 50 years) compared with older patients (older than age 50 years). Data on ctDNA in BTC is currently limited. Our study, the largest cohort reported to date to our knowledge, demonstrates the feasibility of ctDNA testing in this disease. We provide a foundation upon which the field can continue to grow.

The Genomic Landscape of Childhood Acute Lymphoblastic Leukemia

Introduction: Although recent studies have refined the classification of B-progenitor and T-lineage acute lymphoblastic leukemia into gene-expression based subgroups, a comprehensive integration of significantly mutated genes and pathways for each subgroup is needed to understand disease etiology. Methods: We studied 2789 children, adolescents and young adults (AYA) with newly diagnosed B-ALL (n=2,322 cases) or T-ALL (n=467) treated on Children's Oncology Group (n=1,872) and St. Jude Children's Research Hospital trials (n=917). The cohort comprised childhood NCI standard-risk (41.8%; age range 1-9.99 yrs, WBC ≤ 50,000/ml), childhood NCI high-risk (44.5%; age range ≥10 to 15.99 yrs) and AYA (9.9%; age range 16-30.7 yrs). Genomic analysis was performed on tumor and matched-remission samples using whole transcriptome sequencing (RNA-seq; tumor only; n=1,922), whole exome sequencing (n=1,659), whole genome sequencing (n=757), and single nucleotide polymorphism array (n=1,909). Results: For B-ALL, 2104 cases (90.6%) were classified into 26 subgroups based on RNA-seq gene expression data and aneuploidy or other gross chromosomal abnormalities (iAMP21, Down syndrome, dicentric), deregulation of known transcription factors by rearrangement or mutation (PAX5 P80R, IKZF1 N159Y), or activation of kinase alterations (Ph+, Ph-like). For T-ALL, cases were classified into 9 previously described subtypes based on dysregulation of transcription factor genes and gene expression. In 1,659 cases subject to exome sequencing (1259 B-ALL, 405 T-ALL) we identified 18,954 nonsynonymous single nucleotide variants (SNV) and 2,329 insertion-deletion mutations (indels) in 8,985 genes. Overall, 161 potential driver genes were identified by the mutation-significance detection tool MutSigCV or by presence of pathogenic variants in known cancer genes. Integration of sequence mutations and DNA copy number alteration data in B-ALL identified 7 recurrently mutated pathways: transcriptional regulation (40.6%), cell cycle and tumor suppression (38.0%), B-cell development (34.5%), epigenetic regulation (24.7%), Ras signaling (33.0%), JAK-STAT signaling (12.0%) and protein modification (ubiquitination or SUMOylation, 5.0%). The top 10 genes altered by deletion or mutation in B-ALL were CDKN2A/B (30.1%), ETV6 (27.0%), PAX5 (24.6%), CDKN1B (20.3%), IKZF1 (17.6%), KRAS (16.5%), NRAS (14.6%), BTG1 (7.5%) histone genes on chromosome 6 (6.9%) and FLT3 (6.1%), and for T-ALL, CDKN2A/B (74.7%), NOTCH1 (68.2%), FBXW7 (21.3%), PTEN (20.5%) and PHF6 (18.2%) (Figure 1A). We identified 17 putative novel driver genes involved in ubiquitination (UBE2D3, UBE2A, UHRF1, and USP1), SUMOylation (SAE1, UBE2I), transcriptional regulation (ZMYM2, HMGB1), immune function (B2M), migration (CXCR4), epigenetic regulation (DOT1L) and mitochondrial function (LETM1). We also observed variation in the frequency of genes and pathways altered across B-ALL subtypes (Figure 1B). Interestingly, alteration of SAE1 and UBA2, novel genes that form a heterodimeric complex important for SUMOylation, and UHRF1 were enriched in ETV6-RUNX1 cases. Deletions of LETM1, ZMYM2 and CHD4 were associated with near haploid and low hypodiploid cases. Deletion of histone genes on chromosome 6 and alterations of HDAC7 were enriched in Ph+ and Ph-like ALL. Mutations in the RNA-binding protein ZFP36L2 were observed in PAX5alt, DUX4 and MEF2D subgroups. Genomic subtypes were prognostic. ETV6-RUNX1, hyperdiploid, DUX4 and ZNF384 ALL were associated with good outcome (5-yr EFS 91.1%, 87.2%, 91.9% and 85.7%, respectively), ETV6-RUNX1-like, iAMP21, low hyperdiploid, PAX5 P80R and PAX5alt were associated with intermediate outcome (5-yr EFS 68.6%, 72.2%, 70.8%, 77.0% and 70.9%, respectively), whilst KMT2A, MEF2D, Ph-like CRLF2 and Ph-like other conferred a poor prognosis (55.5%, 67.1%, 51.5% and 62.1%, respectively). TCF3-HLF and near haploid had the worst outcome with 5-yr EFS rates of 27.3% and 47.2%, respectively. Conclusions: These findings provide a comprehensive landscape of genomic alterations in childhood ALL. The associations of mutations with ALL subtypes highlights the need for specific patterns of cooperating mutations in the development of leukemia, which may help identify vulnerabilities for therapy intervention. Disclosures Gastier-Foster: Bristol Myers Squibb (BMS): Other: Commercial Research; Incyte Corporation: Other: Commercial Research. Willman:to come: Patents &amp; Royalties; to come: Membership on an entity's Board of Directors or advisory committees; to come: Research Funding. Raetz:Pfizer: Research Funding. Borowitz:Beckman Coulter: Honoraria. Zweidler-McKay:ImmunoGen: Employment. Angiolillo:Servier Pharmaceuticals: Consultancy. Relling:Servier Pharmaceuticals: Research Funding. Hunger:Jazz: Honoraria; Amgen: Consultancy, Equity Ownership; Bristol Myers Squibb: Consultancy; Novartis: Consultancy. Loh:Medisix Therapeutics, Inc.: Membership on an entity's Board of Directors or advisory committees. Mullighan:Amgen: Honoraria, Other: speaker, sponsored travel; Loxo Oncology: Research Funding; AbbVie: Research Funding; Pfizer: Honoraria, Other: speaker, sponsored travel, Research Funding; Illumina: Honoraria, Membership on an entity's Board of Directors or advisory committees, Other: sponsored travel.

Identification of driver genes and somatic mutations in cell-free DNA of patients with pulmonary lymphangioleiomyomatosis.

Next-generation sequencing of cell-free circulating DNA (cfDNA) has emerged as promising technique for identifying minimally invasive genomic profiling of tumor cells recently. However, it remains relatively unknown in LAM disease. In our study, paired cfDNA and genomic DNA (gDNA) in blood samples were obtained from 23 LAM patients and seven healthy controls to explore mutations profiles of targeted 70 cancer-related genes. As results, log2-based allele frequencies of mutations in cfDNA were significantly different from those of gDNA. By comparing the mutual mutations identified both in cfDNA and gDNA, a significant correlation was also observed. After removing mutations in gDNA, distinct somatic mutation profiles of cfDNA were observed in LAM patients. Forty of 70 targeted genes had recurrent mutations, of which ATM, BRCA2 and APC showed the highest frequency. Based on the mutation, correlation network constructed of 40 mutated genes, 11 hub genes bearing intensive interactions were highlighted, including BRCA1, BRCA2, RAD50, RB1, NF1, APC, MLH3, ATM, PDGFRA, PALB2 and BLM. Expression of the hub genes showed significant clusters between LAM patients and controls and that RAD50 and BRCA2 had the strongest associations with subject phenotypes. Myogenesis and estrogen response were confirmed to be positively regulated in LAM patients. Collectively, our study provided a landscape of genomic alterations in LAM and discovered several potential driver genes, that is, BRCA2 and RAD50, which shed a substantial light on the clinical application of key molecular markers and potential therapy targets for precision diagnosis and treatment in the future.

Abstract 5134: Landscape of genomic alterations across solid tumors based on a comprehensive clinical sequencing analysis of 5355 Chinese cancer patients

Abstract Background: Precision oncology highlights the individualization of diagnosis, prognosis and treatment based on molecular stratification of cancer patients (pts). Genetic patterns in many types of cancer vary among geographical regions, leading to variation in evidence-based recommendations for clinical management of pts. Genomic alterations (GAs) of solid tumors from Chinese pts have not been fully studied across all cancer types. Methods: Tumor tissues and matched blood from 5355 Chinese pts were collected and sequenced by a next generation sequencing based 450-gene panel assay. About 60% of the samples were FFPE tissues. In our study, 59% were males and 41% were females with a median age of 58 years old (range 1-92). The major cancer types were NSCLC (36%), CRC (8%), HCC (7%), gastric cancer (5%), pancreatic cancer (5%), and also cholangiocarcinoma, esophageal carcinoma, SCLC, gallbladder carcinoma, ovarian cancer, breast cancer, head and neck cancer, sarcoma, renal carcinoma, uterine neoplasms and others. 1% were pediatric patients . Single base substitutions, short and long insertions/deletions (indels), copy number alterations (CNA), fusions/rearrangements, TMB and MSI status were assessed by NGS. Results: On average, our study identified 7 mutations per patient. Gene rearrangements and long indels (50bp - 2k) were detected in 15% and 5% of the cancers. In total, 58% of pts harbored GAs in druggable genes defined as targets of FDA approved drugs. Highest mutation frequencies were found in TP53 (60%), EGFR (23%), KRAS (18%), CDKN2A (13%), TERT (12%), PIK3CA (11%), APC (11%), LRP1B (10%), ARID1A (9%), and RB1 (8%) across all tumor types. The frequency of other druggable genes included BRCA1/2 (5%), BRAF (3%), HER2 amplifications (3%), MET amplifications (2%), ALK fusions (2%), RET fusions (0.7%), ROS1 fusions (0.7%), FGFR fusions (0.7%), NTRK fusion (0.6%). Frequency of GAs in druggable genes varied among different tumor types. For instance, EGFR mutations occurred in 49% of NSCLC, 7% of SCLC, 2% of gastric cancer, and 2% of CRC. NTRK fusions enriched in sarcoma (4%), gastric cancer (2%) and CRC (1%). The median TMB of the whole cohort was 4.6 muts/Mb and 6% had TMB higher than 20 muts/Mb. Top 3 tumors with highest median TMB were SCLC, CRC and gastric cancer, while pancreatic cancer and sarcoma had low TMB. As a pan-cancer biomarker for immunotherapy, pts with MSI-H accounted for 1.3% of solid tumors, including 9% of uterine neoplasms, 8% of CRC, 6% of gastric cancer and 2% of ICC. Conclusions: In summary, we have constructed a large-scale data set including somatic mutations, CNAs, fusions/rearrangements, TMB and MSI status from 5355 Chinese pts with more than 19 tumor types, which provided an extensive understanding of disease-specific indicators, outcomes and therapeutic strategies for both targeted and immune therapies in Chinese pts. Citation Format: Ming Yao, Minghui Wang, Mingwu Chen, Tao Shou, Jingyu Cao, Hui Chen, Aodi Wang, Lijuan Chen, Jinwei Hu, Shuirong Zhang, Kai Wang. Landscape of genomic alterations across solid tumors based on a comprehensive clinical sequencing analysis of 5355 Chinese cancer patients [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 5134.

Abstract 2722: The genomic landscape of high-grade serous ovarian cancer in long-term survivors

Abstract Purpose: The majority of patients with high-grade serous ovarian cancer (HGSC) develop progressive disease following primary treatment, with a five-year survival rate of ~30%. However, a subset of patients have an extraordinary response to treatment and ~15% survive more than ten years (long-term survivors). The Multidisciplinary Ovarian Cancer Outcomes Group (MOCOG) aims to uncover factors that influence long-term survival of HGSC patients. Here, we investigated the genomic and immunologic determinants of exceptional survival of this deadly disease. Experimental Design: Patient characteristics and clinical histories were evaluated to identify patients diagnosed with advanced stage (Stage IIIC/IV) and histopathologically confirmed HGSC with greater than 10-year overall survival. Whole-genome sequencing (WGS) was performed on primary tumors (median 78x coverage) and germline samples (median 39x coverage) of 55 long-term survivors. Primary tumor samples were also characterised by RNA sequencing, DNA methylation profiling and immunohistochemistry. Results: A total 38 (69%) of long-term surviving patients had residual disease following surgery, suggestive of highly chemo-sensitive disease. Most patients (41, 75%) were alive at last follow-up and 26 (47%) were progression-free. Somatic mutation burden was higher in primary tumors of long-term survivors relative to controls (316 unselected HGSC patients in The Cancer Genome Atlas). Genome-wide mutational signatures were predominantly Signature 3 (associated with homologous recombination deficiency), Signature 1 (age related) and Signatures 5, 8 and 16 (unknown etiology). Inactivation of the tumor suppressor RB1 by structural rearrangements or homozygous deletion was frequent in long-term survivors, with 33% of tumors showing associated loss of RB1 protein expression by immunohistochemistry compared to 13% of unselected HGSC controls (n=207; P = 0.001). Staining of adjacent tumor tissue revealed that RB1 loss was associated with increased numbers of PD-1+ tumor-infiltrating lymphocytes (P = 0.015) and MHC class I on tumor cells (P = 0.002). In an independent HGSC cohort (n=847), RB1 protein loss was associated with prolonged survival (HR: 0.75, P &amp;lt; 0.001) compared to patients with RB1 positive tumors. Furthermore, co-occurrence of germline mutations in BRCA1 or BRCA2 and RB1 loss was associated with a significantly longer overall survival compared to patients with retained RB1 protein expression and no germline BRCA mutation (HR: 0.44, P &amp;lt; 0.001). Conclusions: This study delineates the full landscape of genomic alterations in HGSC of long-term survivors. Our findings indicate that specific mutations might be associated with enhanced host immune responses and long-term survival. Citation Format: Dale W. Garsed, Ahwan Pandey, Sian Fereday, Kathryn Alsop, Maartje C. Wouters, Flurina Saner, Jessica A. Beach, Katy Milne, Catherine J. Kennedy, Joy Hendley, Nadia Traficante, Celeste L. Pearce, Malcolm C. Pike, Multidisciplinary Ovarian Cancer Outcomes Group, Susan J. Ramus, Martin Köbel, Brad H. Nelson, Ellen L. Goode, Anna deFazio, David D. Bowtell. The genomic landscape of high-grade serous ovarian cancer in long-term survivors [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 2722.

Abstract 3912: The PI3K/mTOR dual inhibitor PF-04691502 shows antitumor activity in Sezary cells and in a xenograft mouse model

Abstract Introduction: Cutaneous T-cell Lymphomas (CTCLs) are extranodal non-Hodgkin’s lymphomas deriving from neoplastic transformation of T lymphocytes that home to the skin. Mycosis fungoides (MF) is the most diffuse form, usually having an indolent course but presenting in some cases as an aggressive disease with a poor prognosis. Sézary syndrome (SS) is a rare and aggressive variant, with erythroderma, leukemia, and lymph node involvement. Despite recent advances, CTCLs are still incurable cancers and the clinical symptoms are highly disabling. Among the landscape of genomic alterations identified in CTCL, we and others have described alterations of the PI3K pathway members (loss of PTEN, LKB1 and PDCD4, and gain of p70S6K). As a consequence, several SS patients show hyper-activation of PI3K/AKT/mTOR pathway. Hence, inhibition of this pathway represents a potential therapeutic choice against CTCL. Objective: In this work, we investigated the therapeutic potential of the dual-PI3K/mTOR inhibitor PF-04691502 (hereafter PF-502) against CTCL. Materials and Methods: PF-502 therapeutic efficacy was evaluated in four CTCL cell lines and in patient-derived SS cells, for its ability to inhibit cell growth, to induce apoptosis and to inhibit chemotaxis toward the chemokine SDF-1. Moreover, to evaluate PF-502 therapeutic effects in vivo we used a xenograft mouse model, consisting of nude mice transplanted with the CTCL-derived HH cell line. Results: PF-502 inhibited the cell growth of all the CTCL cell lines (0.07-0.32 µM, IC50 range), and of tumor cells derived from six SS patients (2.3 µM, mean IC50). The mechanism of action of PF-502 in CTCL cell lines was a combination of apoptosis induction and G1 block, while in SS patients it was mostly based on a strong apoptosis induction. Of note, PF-502 showed an only modest effect in T-lymphocytes from 3 healthy donors. PF-502 inhibited also cell migration toward the chemokine SDF-1, in both the systems. This result might have an important clinical implication because we have recently demonstrated that malignant lymphocytes are activated by the skin microenvironment. Thus, blocking the homing to the skin could affect the disease course. Finally, PF-502 has a robust antitumor activity in the xenograft mouse model, by reducing tumor volume (400 mm3 in treated mice vs 936 mm3 in controls) as well as tumor weight (0.2 g in treated mice vs 0.56 g in controls). Moreover, the Kaplan-Meier curve, in these mice, revealed that PF-502 treatment prolongs survival (P&amp;lt;0.005). Conclusions: Our results strongly support the therapeutic potential of targeting PI3K/AKT/mTOR pathway in CTCL. Citation Format: Antonella Bresin, Cristina Cristofoletti, Francesca Monzo, Elisabetta Caprini, Mauro Helmer Citterich, Alessandra Frezzolini, Alessandro Monopoli, Roberto Benucci, Maria Cantonetti, Enrico Scala, Giandomenico Russo, Maria Grazia Narducci. The PI3K/mTOR dual inhibitor PF-04691502 shows antitumor activity in Sezary cells and in a xenograft mouse model [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 3912.

Genomic somatic alterations of human epidermal growth factor-2 (HER2) gene: A pan-cancer analysis.

3112 Background: Human epidermal growth receptor 2 (HER2) is a well-known oncogenic drive gene with multiple targeted therapeutic options. In this study, we aim to assess the landscape of HER2 alterations in solid tumors and evaluate the feasibility of circulating tumor DNA (ctDNA) tested by next-generation sequence (NGS) as a tool to detect HER2 alterations. Methods: Alterations of HER2 by NGS (Illumina NextSeq 500) were queried in 3D Medicines database. The mean depth of tissue and circulating tumor DNA (ctDNA) test was 500X and 5000X, respectively. 11,013 patients tested using tumor tissue and 6,970 patients tested using ctDNA were included in this analysis. Results: Of 11,013 patients tested using tumor tissue, any HER2 and known or likely deleterious HER2 mutations were identified in 739 (6.7%) and 531 (4.8%) patients, respectively. Of 531 patients who carried known or likely deleterious HER2 mutations, 263 (49.5%) had HER2 amplification and 259 (48.8%) had single nucleotide variations (SNVs). Across all tumor types, breast cancer was found to have the highest frequency of HER2 amplification (14.9%, 48/323), followed by gastric cancer (6.6%, 31/470) and biliary tract cancer (5.8%, 33/571). Moreover, 11% (8/73) of duodenal cancer, 4.5% (7/154) of urothelial cancer, 3.8% (18/470) of gastric cancer, 3.1% (142/4555) of lung cancer, 2.9% (17/571) of biliary tract cancer, 2.8% (44/1562) of colorectal cancer and 2.7% (9/323) of breast cancer carried known or likely deleterious HER2 SNVs. Of 6970 patients tested using ctDNA, any HER2 and known or likely deleterious HER2 mutations were identified in 592 (8.5%) and 277 (4.0%) patients, respectively. In the ctDNA cohort, 15.7% (36/230) of breast cancer and 3.1% (5/161) of biliary tract cancer carried HER2 amplification. However, 11.6% (20/173) of gastric cancer had HER2 amplification tested by ctDNA which was higher than that tested using tissue. Furthermore, 5.6% (13/230) of breast cancer, 4.5% (2/44) of urothelial cancer, 3.4% (6/173), 2.5% of biliary tract cancer and 2.0% (94/4586) lung cancer harbored known or likely deleterious HER2 SNVs in ctDNA cohort. Conclusions: HER2 alterations existed across tumor types and the landscape of genomic alterations in HER2 gene varied according to different type of tumor. In addition, ctDNA can be used as a potential tool to detect HER2 alterations.

A landscape of circulating tumor DNA in esophageal adenocarcinoma and squamous cell carcinoma.

4070 Background: Esophageal cancer (EC) is a lethal malignancy with limited treatment options. Genomic analyses have led to the elucidation of numerous dysregulated genes in esophageal adenocarcinoma (AC) and squamous cell carcinoma (SCC), and the potential for advancement of targeted therapies in this disease. Data regarding circulating tumor DNA (ctDNA) plasma analysis in EC in real-world clinical practice is limited. Methods: We performed ctDNA next-generation sequencing (NGS) analysis in patients (pts) with EC (January 2015- February 2018). ctDNA analysis was performed using Guardant 360 (Guardant Health, CA) which detects single nucleotide variants and insertion/deletion mutations, and specific amplifications and fusions, in up to 73 different genes. The mutant allele fraction (MAF) for detected alterations was calculated relative to wild type in ctDNA. Therapeutic relevance was defined as alterations within OncoKB levels 1-3B and R1. Results: Among 450 pts, 487 total samples were analyzed (77% AC, 31% SCC). ctDNA NGS revealed at least one genomic alteration (excluding variants of uncertain significance and synonymous mutations) in 81% of pts (90% AC, 88% SCC). Median number of alterations per AC patient was 4 [range, 1-59] and a median MAF of 0.84% (range, 0.02% - 83.7%); SCC was 5 [range, 1-26], with a median MAF of 0.99% (range, 0.01% - 85.2%). The total number of unique alterations was 1,162. The most commonly altered genes in AC: TP53 (70%), KRAS (20%), ERBB2 (18%), EGFR (16%), PIK3CA (16%); in SCC: TP53 (88%), PIK3CA (24%), CCND1 (23%), KRAS (21%), EGFR 15%). Therapeutically relevant alterations will be described. Conclusions: ctDNA plasma profiling of pts with EC is a feasible alternative and non-invasive method to gather comprehensive genomic data. Further large comparison studies to assess landscape of genomic alterations observed through ctDNA versus tissue-based assays, in addition to studies of targeted therapy outcomes based on ctDNA-detected alterations, are needed.

Mutational landscape of T-cell lymphoma in mice lacking the DNA mismatch repair gene Mlh1: no synergism with ionizing radiation.

Biallelic germline mutations in the DNA mismatch repair gene MLH1 lead to constitutional mismatch repair-deficiency syndrome and an increased risk for childhood hematopoietic malignancies, including lymphoma and leukemia. To examine how Mlh1 dysfunction promotes lymphoma as well as the influence of ionizing radiation (IR) exposure, we used an Mlh1-/- mouse model and whole-exome sequencing to assess genomic alterations in 23 T-cell lymphomas, including 8 spontaneous and 15 IR-associated lymphomas. Exposure to IR accelerated T-cell lymphoma induction in the Mlh1-/- mice, and whole-exome sequencing revealed that IR exposure neither increased the number of mutations nor altered the mutation spectrum of the lymphomas. Frequent mutations were evident in genes encoding transcription factors (e.g. Ikzf1, Trp53, Bcl11b), epigenetic regulators (e.g. Suv420h1, Ep300, Kmt2d), transporters (e.g. Rangap1, Kcnj16), extracellular matrix (e.g. Megf6, Lrig1), cell motility (e.g. Argef19, Dnah17), protein kinase cascade (e.g. Ptpro, Marcks) and in genes involved in NOTCH (e.g. Notch1), and PI3K/AKT (e.g. Pten, Akt2) signaling pathways in both spontaneous and IR-associated lymphomas. Frameshift mutations in mononucleotide repeat sequences within the genes Trp53, Ep300, Kmt2d, Notch1, Pten and Marcks were newly identified in the lymphomas. The lymphomas also exhibited a few chromosomal abnormalities. The results establish a landscape of genomic alterations in spontaneous and IR-associated lymphomas that occur in the context of mismatch repair dysfunction and suggest potential targets for cancer treatment.

Cerebrospinal fluid cell-free tumour DNA as a liquid biopsy for primary brain tumours and central nervous system metastases.

Challenges in obtaining tissue specimens from patients with brain tumours limit the diagnosis and molecular characterisation and impair the development of better therapeutic approaches. The analysis of cell-free tumour DNA in plasma (considered a liquid biopsy) has facilitated the characterisation of extra-cranial tumours. However, cell-free tumour DNA in plasma is limited in quantity and may not reliably capture the landscape of genomic alterations of brain tumours. Here, we review recent work assessing the relevance of cell-free tumour DNA from cerebrospinal fluid in the characterisation of brain cancer. We focus on the advances in the use of the cerebrospinal fluid as a source of cell-free tumour DNA to facilitate diagnosis, reveal actionable genomic alterations, monitor responses to therapy, and capture tumour heterogeneity in patients with primary brain tumours and brain and leptomeningeal metastases. Profiling cerebrospinal fluid cell-free tumour DNA provides the opportunity to precisely acquire and monitor genomic information in real time and guide precision therapies.