Abstract 3522: Population estimates of prostate cancer risk and prognosis for carriers of germline pathogenic variants in disease implicated genes, using 200,000 UK Biobank whole exomes

67 Background: Germline mutations in DNA Damage Response (DDR) genes such as BRCA2 and ATM have been associated with prostate cancer risk and aggressiveness. These associations are largely based on studies that ascertain for cancer diagnosis and family history and provide risk estimates with limited population-level accuracy. Here we evaluate the clinical significance of germline pathogenic variants in 20 DDR genes and a high-risk susceptibility coding variant in HOXB13 using whole exome sequences from (1) the UK Biobank (UKBB), a population-based cohort (300,000 participants) and (2) patients recruited in AZ clinical trials. Methods: Whole exomes from 6,987 prostate cancer patients (5,921 UKBB and 1,066 AZ) and 88,499 cancer-free males were analysed. Known and novel pathogenic variants were identified and associations with disease risk, age of onset, family history, response to hormonal therapy and overall survival were estimated (multiplicity corrected P value < 0.005). Results: HOXB13 G48E (1.36%), ATM (1.03%) and BRCA2 (0.99%) were the largest contributors to prostate cancer risk, each conferring an increase of ~4-fold, followed by CHEK2 with a moderate contribution (0.46%). No significant contributions to prostate cancer risk were observed for any of the other genes analysed. Family history of prostate cancer was not significantly enriched in any of the gene subpopulations of prostate cancer carriers and compared with non-carriers, there was no significant difference in the median age of disease onset. Analysis of clinical outcomes showed that BRCA2 carriers had a 4-fold increased risk of death (Cox PH HR p = 4.11E-12) and poorer overall survival (Log-Rank p = 4.60E-14), with 88% dying from prostate cancer compared to 49% of non- BRCA2 carriers (UKBB analysis). BRCA2 pathogenic mutations were also associated with early failure to hormonal therapy (Cox PH HR 2.38; p = 9.48E-04; AZ cohort analysis). HOXB13, ATM and CHEK2 mutations were not significantly associated with clinical outcomes. Conclusions: This is the largest study to date providing population-based estimates of prostate cancer risk and prognosis, highlighting BRCA2 carriers as a population in clinical need of early identification and targeted intervention. Updated analyses with data from the full 450,000 UKBB participants (9,000 prostate cancers) will be presented.

Decision letter: Pan-cancer association of DNA repair deficiencies with whole-genome mutational patterns

Editor's evaluation: Pan-cancer association of DNA repair deficiencies with whole-genome mutational patterns

11567 Background: Deleterious mutations in DDR genes are frequently associated with response to poly(ADP-ribose) polymerase (PARP) inhibitors and platinum chemotherapy. However, much remains unknown about their association with specific molecular signaling pathways. We report the prevalence of pathogenic variants in DDR genes and their co-alteration with other somatic variants. Methods: Targeted exome sequencing of 201 genes was performed in 1,189 patients (pts) with advanced solid tumors enrolled in a molecular testing protocol (NCT01772771), using matched normal and tumor DNA. We assessed germline and somatic alterations in 15 cancer-related DDR genes, their co-occuring genomic alterations and the tumor mutation burden (TMB), defined as number of somatic non-synonymous mutations. Results: A total of 124 pathogenic or likely pathogenic variants in DDR genes were identified in 111/1189 (9%) pts with 57% of these alterations being somatic. These variants were found in the following genes: ATM 17 pts (1.4%); BAP1 5 pts (0.4%); BRCA1 18 pts (1.5%); BRCA2 17pts (1.4%); CHEK1 8 pts (0.7%); CHEK2 16 pts (1.3%); ERCC3 4 pts (0.3%); ERCC4 2 pts (0.2%); ERCC5 3 pts (0.2%); MLH1 4pts (0.3%); MSH2 8 pts (0.7%); MSH6 6 pts (0.5%); PALB2 3 pts (0.2%) and RAD51 1pt (0.1%). DDR alterations were found more frequently in the following tumor types tested: breast 14%, colorectal 12%, melanoma 8%, glioblastoma 6% and ovarian 6%. The most relevant somatic co-alterations with DDR mutations were activation of the PI3K/AKT/mTOR pathway through mutations or copy-number variations in AKT1, MTOR, NF1, PIK3CA, PIK3R1, PTEN, TSC1 and TSC2 (p = 0.008). Patients with deleterious variants in mismatch excision repair genes (MLH1, MSH2 or MSH6) had a significantly higher TMB than all other patients enrolled (median TMB = 62 vs 5, p = 0.002). Patients with somatic pathogenic DDR variants had a significantly higher TMB (median = 13) compared to patients with germline DDR variants (median = 5) (p = 0.004). Conclusions: The association of DNA repair mutations with alterations in signaling pathways provide rationale for novel therapeutic combinations. Variations in TMB based on distinct types of DDR gene alterations may have implications for immunotherapy.

Background: Detecting pathogenic intronic variants resulting in aberrant splicing remains a challenge in routine genetic testing. We describe germline whole-exome sequencing (WES) analyses and apply in silico predictive tools of familial ovarian cancer (OC) cases reported clinically negative for pathogenic BRCA1 and BRCA2 variants. Methods: WES data from 27 familial OC cases reported clinically negative for pathogenic BRCA1 and BRCA2 variants and 53 sporadic early-onset OC cases were analyzed for pathogenic variants in BRCA1 or BRCA2. WES data from carriers of pathogenic BRCA1 or BRCA2 variants were analyzed for pathogenic variants in 10 other OC predisposing genes. Loss of heterozygosity analysis was performed on tumor DNA from variant carriers. Results:BRCA1 c.5407-25T>A intronic variant, identified in two affected sisters and one sporadic OC case, is predicted to create a new splice effecting transcription of BRCA1. WES data from BRCA1 c.5407-25T>A carriers showed no evidence of pathogenic variants in other OC predisposing genes. Sequencing the tumor DNA from the variant carrier showed complete loss of the wild-type allele. Conclusions: The findings support BRCA1 c.5407-25T>A as a likely pathogenic variant and highlight the importance of investigating intronic sequences as causal variants in OC families where the involvement of BRCA1 is highly suggestive.

Rare Hypomorphic Sucrase Isomaltase Variants in Relation to Irritable Bowel Syndrome Risk in UK Biobank

Oral cavity squamous cell carcinoma (OSCC) is one of the most common cancers in Taiwan and worldwide. Except cigarette smoking and alcohol drinking, areca quid (AQ) chewing is also the major risk factor of OSCC in Southeast Asia including Taiwan. Despite better methods of diagnosis and new treatment procedures, there has not been a significant improvement in the five year survival rate for OSCC in the past 30 years. To explore some clues for clinical management of OSCC, we carried out whole genome loss of heterozygosity (LOH) and copy number alterations (CNAs) analysis and found high frequency of minimal deleted region (MDR) in11q13.4-q25 and significant CNA region in 11q22.3-q24.3. In this region, several DNA damage response (DDR) genes are located including MRE11A, ATM, H2AFX and CHEK1. The DDR pathway represents a complex network of multiple signaling pathways involving cell cycle checkpoints, DNA repairs, transcriptional programs, and apoptosis, through which cells maintain genomic integrity following various endogenous or environmental stress. To investigate the clinical significance of DDR pathway in OSCC, CNAs and LOH of ATM, MRE11A, and CHEK1 gene were analyzed in the present study. A total number of 324 OSCCs were first examined for CNAs using TaqMan CNV analysis and the copy number neutral cases were then examined for LOH using TaqMan SNP analysis and denaturing high performance liquid chromatography (DHPLC). Preliminary results indicated that only 8% (26/324) of OSCCs did not have any alterations in these 3 genes. The frequency of loss of MRE11A, ATM and CHEK1 gene was 7.10% (23/324), 15.74% (51/324) and 13.89% (45/324), respectively. Loss of MRE11A, ATM, and CHEK1 was associated with tumor differentiation and lymph node metastasis. The frequency of LOH in MRE11A, ATM, and CHEK1 gene was 37.84% (42/111), 50% (61/122) and 58.28% (95/163), respectively and LOH of MRE11A was associated with tumor differentiation. Combined the loss and LOH as the alteration event, we found that alterations of MRE11A, ATM, and CHEK1 gene were associated with tumor differentiation. Alterations of MRE11A and ATM were associated with early lymph node metastasis, while alterations of CHEK1 were associated with late lymph node metastasis. Alterations of MRE11A, ATM, and CHEK1 gene were associated with poor disease-free survival (DFS) and overall survival (OS) and gene loss had a stronger effect than LOH. Furthermore, alterations of MRE11A/ATM/CHEK1 as a whole were strongly associated with poor DFS and OS. These results confirmed our previous MDR findings and indicated that MRE11A/ATM/CHEK1 pathway might play a crucial role in the development and progression of OSCCs. Citation Format: Huei-Tzu Chien, Shiang-Fu Huang, I-How Chen, Chun-Ta Liao, Hung-Ming Wang, Ling-Ling Hsieh. Clinical significance of MRE11A/ATM/CHEK1 DNA damage response genes in oral cavity squamous cell carcinomas. [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 3868. doi:10.1158/1538-7445.AM2015-3868

1075 Background: In the phase 3 TROPiCS-02 study, SG, a Trop-2–directed antibody-drug conjugate coupled to SN-38 as the payload, demonstrated clinically meaningful improvement in survival outcomes over chemotherapy in patients (pts) with pretreated HR+/HER2- mBC. SN-38 leads to double-stranded DNA damage, therefore we hypothesized that in tumors with defective DDR machinery, SG may provoke synthetic lethality. We report on comprehensive genomic analysis of DDR gene variants and the impact on SG clinical efficacy in this patient population. Methods: Pts with HR+/HER2– mBC who received prior taxane, endocrine therapy (ET), CDK4/6 inhibitor, and 2-4 prior lines of chemotherapy were randomized to receive SG (10 mg/kg IV days 1 and 8, every 21 days) or treatment of physician’s choice (TPC) until disease progression or unacceptable toxicity. DDR gene deleterious variants were identified using whole exome sequencing (WES) on archival or screening tumor tissues based on 142 DDR pathway genes annotated in the Kyoto Encyclopedia Genes and Genomes (KEGG) database and Human DNA Repair Gene database; association between these variants and clinical outcomes was evaluated using Cox regression model as Hazard Ratio (HR) with 95% Confidence Intervals (CI). Results: Of 543 patients included in the intent-to-treat (ITT) population, WES data was available for 195 (36%) – clinicopathological features were similar between ITT and WES dataset (median age = 57 vs 58 for SG and 55 vs 55 for TPC; prior lines of chemotherapy >2 lines = 59% vs 57% for SG and 58% vs 55% for TPC). Estrogen Receptor (ER)>10% and prior lines of chemotherapy were comparable between WT and MUT in SG arm and in TPC arm. Overall, 114 (58%) of tumor samples had ≥1 deleterious alteration with BRCA2, PRKDC, ATM being the most common alterations. The primary results are included (Table). Although all patients demonstrated improved efficacy with SG vs TPC, patients with DDR mutant MBC had numerically greater benefit in PFS vs WT (MUT HR= 0.61; WT HR= 0.76) and OS (MUT HR = 0.68; WT HR= 0.82). Conclusions: While SG benefit over TPC was observed in both DDR WT and DDR MUT HR+/HER2– mBC, numerically greater benefit was observed for patients with DDR deficient tumors, suggesting possible synergy between the DDR pathway and SG’s anti-tumor effect. Further study of the synergistic effects of SG in combination with agents targeting DDR pathway are warranted. Clinical trial information: NCT03901339 . [Table: see text]

e15608 Background: DNA damage response (DDR) genes are essential to maintain genomic integrity. Increasing evidence showed that mutations in DDR genes were related to tumor mutation load (TMB), microsatellite instability (MSI) and clinical benefit from immunotherapy. TMB and MSI are predictive biomarkers for immunotherapy. However, the mutation spectrum of genes involved in the DDR pathways and its association with TMB and MSI in Chinese colorectal cancer patients are still ambiguous. Methods: The genomic information of 728 Chinese colorectal cancer patients without germline mutation of Lynch syndrome related genes was obtained from HapLab database. HaploX WESPlus gene panel (an upgraded version of the standard WES) was performed to analyze the genomic data of patients. Results: 630 out of 728 patients (86.54%) had at least one mutation of the DDR genes. The frequently mutated DDR genes included TP53 (54.95%), BRCA2 (18.27%), PARP4 (13.46%), MSH2 (9.62%), XPC (7.55%), RAD54L2 (7.28%), ATM (6.73%), HMGB1 (5.77%), REV3L (5.49%) and POLD3 (5.36%). The TMB in patients with one, two, three and more than three mutations of DDR genes increased sequentially. The proportion of MSI-H in patients with more than three mutations of DDR genes (20.9%) was significantly higher than that in patients with less than 3 mutations of DDR genes (0.38%). Among the frequently mutated DDR genes, mutations of BRCA2, PARP4, MSH2, XPC, RAD54L2, ATM, REV3L or POLD3 were associated with higher TMB, mutations of MSH2, RAD54L2, ATM, REV3L or POLD3 were associated with higher proportion of MSI-H. The patients with mutation in each of eight DDR pathways showed significantly higher TMB and proportion of MSI-H than who without mutation of DDR genes. Conclusions: This study revealed the mutation profile of DDR genes in Chinese colorectal cancer patients. The mutation of DDR genes was closely related to TMB and MSI status. This result indicated the mutation of DDR genes may be a promising biomarker for immunotherapy in Chinese colorectal cancer patients.

4522 Background: Neoadjuvant cisplatin-based chemo followed by radical cystectomy (RC) is a standard of care treatment for pts with MIBC. DDR gene mts, including within ERCC2, a DNA helicase implicated in cisplatin sensitivity in MIBC, have been associated with higher pathologic (path) downstaging ( < pT2) and complete response (pT0) at RC and improved overall survival (OS) in retrospective series. S1314 randomized pts to one of 2 chemo regimens (dose dense MVAC or Gem/Cis) followed by RC. We sought to correlate ERCC2 and other DDR gene mts with response and survival in MIBC pts enrolled onto this prospective trial. Methods: Tumor and matched germline DNA from evaluable pts enrolled onto S1314 underwent exon capture sequencing of 505 cancer-associated genes (MSK-IMPACT). Both deleterious (del) mts and any mts in 9 DDR genes (ERCC2, ERCC5, BRCA1, BRCA2, RECQL4, ATM, ATR, RAD51C, FANCC) were correlated with clinical outcomes. The prespecified analyses included the association of mts with < pT2 and pT0 by logistic regression analysis and with progression-free survival (PFS) and OS by Cox proportional hazards regression. Results: 179 patients (median 61 years, 85% male, 87% white, and 87% clinical stage T2) who received >2 cycles of chemo and were evaluable for path response were included in the analysis. The pT0 rate was 28% and < pT2 was 41%. Del mts in ERCC2 were detected in 26 (14%) pts followed by ATM (n = 12, 7%), ATR (n = 3) and BRCA2 (n = 2). ERCC2 mts were associated with statistically significantly higher path responses with a 54% pT0 rate and 62% downstaging rate. Patients with any del mts had higher path response rates (51% pT0, 56% < pT2) and better PFS (Table) with a median follow-up of 53 months. There was a non-significant trend towards improved OS. Conclusions: In pts managed with neoadjuvant chemo and RC on S1314, both ERCC2 mts and del DDR gene mts correlated with pathologic response. Any del DDR gene mt was associated with improved PFS. These results are in line with retrospective analyses displaying a correlation between DDR gene mts and neoadjuvant chemosensitivity in MIBC and support ongoing genomically-informed organ sparing trials.[Table: see text]

Background: Minimal data exists for the utilization of the Oncotype Dx® assay specifically in breast cancers associated with BRCA1/2 pathogenic variants (PVs). It is unknown whether estrogen receptor positive (ER+) breast cancer associated with an inherited BRCA1 or BRCA2 (BRCA1/2) PV is more aggressive than disease seen in patients who do not carry an inherited PV, and whether there are differences between BRCA1 and BRCA2. In prostate cancer patients with inherited cancer predisposition due to a BRCA2 PV, more aggressive cancers are observed, which influences first-line treatment. Limited data exists for the optimal management of early stage ER+ breast cancer in BRCA1/2 PV carriers. Comparing Recurrence Score® (RS) results in ER+ breast cancer patients with an inherited BRCA1/2 PV (cases) versus matched patients who test negative for a PV in BRCA1/2 (controls) may inform whether biologically more aggressive breast cancer is seen in BRCA1/2 carriers and optimal treatment approaches. Methods: A retrospective case control study was performed to compare RS results in women with breast cancer with an inherited BRCA1/2 PV versus patients who tested negative for an inherited BRCA1/2 PV. Female breast cancer patients seen between 2005-2020 at NorthShore University Health System with ER+Her2- early stage invasive breast cancer with 0-3 lymph nodes who completed genetic testing for BRCA1/2 were eligible for enrollment. BRCA1/2 cases were defined as individuals with an inherited PV in BRCA1/2 and controls were negative for BRCA1/2 or other known breast cancer risk gene PVs tested. Subjects were excluded if they had neoadjuvant therapy (hormonal or cytotoxic chemotherapy). Eligible cases were matched to control patients by age, grade, and stage. The Recurrence Score result was obtained by chart review; if not previously evaluated, Oncotype Dx assay was performed by Exact Sciences. Statistical analysis of the primary outcome used the paired t-test to determine mean difference in RS results between BRCA1/2 PV carriers and patients negative for a PV in BRCA1/2 using a 1:1 matched pairs design. Results: A total of 46 matched cases and controls were analyzed. Median age was 50 with a range of 28-74. Of the cases, 18 had a BRCA1 PV and 28 had a BRCA2 PV. Cases and controls were well matched for age (> 50 and ≤ 50); race, grade, stage, and progesterone receptor status. As expected, a higher number of BRCA1/2 carriers were treated with mastectomy while more of the controls received breast-conserving surgery. Chemotherapy was utilized more frequently in the cases (67.4%) versus the controls (54.4%). The average RS result was higher in the cases (27) than the controls (21.3) by a mean difference of 5.7 (p = 0.0195). Using Oncotype Dx cutoffs of low < 18, intermediate 18-30 and high ≥ 31, a statistically significant difference in RS result was noted in the cases versus controls. For cases in the highest risk group (Oncotype Dx ≥ 31), only 20% of their matches also had a score in the highest risk group while 35% had a score in the lowest risk group. Subgroup analysis showed that the cases had the largest difference in RS result from their controls in premenopausal women (age ≤ 50), BRCA1 carriers, and the node negative population. Conclusion We present one of the largest data sets available to date of a well-matched cohort of cases and controls which shows that BRCA1/2 PV carriers are more likely to have a higher Recurrence Score result than their matched controls when matched for age, grade, and stage. These findings suggest ER+ breast cancer in BRCA1/2 PV carriers is biologically more aggressive. Further investigation is warranted to evaluate how this important finding impacts adjuvant therapy recommendations for BRCA1/2 PV carriers. Citation Format: Poornima Saha, Ashley Aller, Amanda Deliere, Peter Hulick, Katharine Yao, Kristine Kuchta, Megan Sullivan, Allison DePersia. Application of 21-gene Breast Recurrence Score® assay to evaluate prognosis and benefit of adjuvant chemotherapy in BRCA1 and BRCA2 pathogenic variant carriers with early stage, estrogen receptor positive breast cancer [abstract]. In: Proceedings of the 2022 San Antonio Breast Cancer Symposium; 2022 Dec 6-10; San Antonio, TX. Philadelphia (PA): AACR; Cancer Res 2023;83(5 Suppl):Abstract nr P5-03-15.

OBJECTIVES/SPECIFIC AIMS: Prostate cancer is the second leading cause of cancer-related death among men in the U.S. and over half of all prostate cancer patients receive radiation therapy (RT). RT induces double-strand breaks (DSBs) in DNA which are lethal to cells if not repaired. While potentially curative, 10% of low-risk patients and 50% of high-risk patients treated with RT still experience tumor recurrence. Thus, identification of novel therapeutic targets to enhance RT will likely reduce prostate cancer mortality. The only clinical approach to enhance RT is androgen deprivation therapy, which targets androgen receptor (AR) signaling; however, its use is limited due to systemic side effects. We recently reported that PRMT5 epigenetically activates AR which led us to investigate if targeting PRMT5 sensitizes prostate cancer to RT. The goal of this project is to determine if PRMT5 is a therapeutic target for prostate cancer radiosensitization and analyze its mechanistic role in response to radiation. METHODS/STUDY POPULATION: To evaluate if targeting PRMT5 may sensitize prostate cancer cells to radiation, we performed a clonogenic assay of irradiated cells. To determine if PRMT5 is required for repair of radiation-induced DSBs, we performed foci analysis via immunocytochemistry. We then used RNA-seq, qPCR, western blot, and ChIP to evaluate a potential epigenetic role of PRMT5 in activating the expression of genes critical to DSB repair. To extend our findings, we analyzed clinical data from around 18,000 of cancer patients encompassing 43 cancer types to assess if PRMT5 expression correlates with the expression of its putative target genes. RESULTS/ANTICIPATED RESULTS: Targeting PRMT5 sensitizes prostate cancer cells to radiation independently of AR status. RNA-seq analysis revealed putative PRMT5 target genes including several involved in DSB repair and G2 arrest. Mechanistically, PRMT5 functions as a master epigenetic activator of DNA damage response (DDR) genes: PRMT5 maintains the basal expression of several DDR genes including BRCA1, BRCA2, and RAD51 and is recruited upon radiation to DDR gene promoters to activate their expression via histone methylation. Targeting PRMT5 decreases expression of these genes at the protein level and hinders repair of radiation-induced DSBs in multiple cancer and non-cancer cell types. Clinically, PRMT5 expression positively correlates with the expression of these DDR genes across all 43 cancer types analyzed. DISCUSSION/SIGNIFICANCE OF IMPACT: PRMT5 acts as a master epigenetic activator of genes involved in DDR and is critical for cells to survive radiation treatment. Importantly, PRMT5 epigenetically activates multiple genes that encode for well-characterized core repair proteins involved in HR (RAD51, RAD51AP1, RAD51D, BRCA1 and BRCA2) and NHEJ (NHEJ1, Ku80, XRCC4, and DNAPKcs), which may explain why PRMT5 is essential to repair IR-induced DSBs in several cell lines. As PRMT5 is overexpressed in many human cancers and its overexpression correlates with poor prognosis, our findings suggest that more efficient DSB repair via PRMT5 overexpression in these cancers may confer survival advantages particularly following DNA damaging treatments. Lastly, because targeting DSB repair is a clinically validated therapeutic approach for cancer treatment, our findings also suggest that PRMT5 targeting may be explored as a monotherapy or in combination therapy with radiation therapy or chemotherapy for cancer treatment.

SummaryBackgroundLynch syndrome is a rare familial cancer syndrome caused by pathogenic variants in the mismatch repair genes MLH1, MSH2, MSH6, or PMS2, that cause predisposition to various cancers, predominantly colorectal and endometrial cancer. Data are emerging that pathogenic variants in mismatch repair genes increase the risk of early-onset aggressive prostate cancer. The IMPACT study is prospectively assessing prostate-specific antigen (PSA) screening in men with germline mismatch repair pathogenic variants. Here, we report the usefulness of PSA screening, prostate cancer incidence, and tumour characteristics after the first screening round in men with and without these germline pathogenic variants.MethodsThe IMPACT study is an international, prospective study. Men aged 40–69 years without a previous prostate cancer diagnosis and with a known germline pathogenic variant in the MLH1, MSH2, or MSH6 gene, and age-matched male controls who tested negative for a familial pathogenic variant in these genes were recruited from 34 genetic and urology clinics in eight countries, and underwent a baseline PSA screening. Men who had a PSA level higher than 3·0 ng/mL were offered a transrectal, ultrasound-guided, prostate biopsy and a histopathological analysis was done. All participants are undergoing a minimum of 5 years' annual screening. The primary endpoint was to determine the incidence, stage, and pathology of screening-detected prostate cancer in carriers of pathogenic variants compared with non-carrier controls. We used Fisher's exact test to compare the number of cases, cancer incidence, and positive predictive values of the PSA cutoff and biopsy between carriers and non-carriers and the differences between disease types (ie, cancer vs no cancer, clinically significant cancer vs no cancer). We assessed screening outcomes and tumour characteristics by pathogenic variant status. Here we present results from the first round of PSA screening in the IMPACT study. This study is registered with ClinicalTrials.gov, NCT00261456, and is now closed to accrual.FindingsBetween Sept 28, 2012, and March 1, 2020, 828 men were recruited (644 carriers of mismatch repair pathogenic variants [204 carriers of MLH1, 305 carriers of MSH2, and 135 carriers of MSH6] and 184 non-carrier controls [65 non-carriers of MLH1, 76 non-carriers of MSH2, and 43 non-carriers of MSH6]), and in order to boost the sample size for the non-carrier control groups, we randomly selected 134 non-carriers from the BRCA1 and BRCA2 cohort of the IMPACT study, who were included in all three non-carrier cohorts. Men were predominantly of European ancestry (899 [93%] of 953 with available data), with a mean age of 52·8 years (SD 8·3). Within the first screening round, 56 (6%) men had a PSA concentration of more than 3·0 ng/mL and 35 (4%) biopsies were done. The overall incidence of prostate cancer was 1·9% (18 of 962; 95% CI 1·1–2·9). The incidence among MSH2 carriers was 4·3% (13 of 305; 95% CI 2·3–7·2), MSH2 non-carrier controls was 0·5% (one of 210; 0·0–2·6), MSH6 carriers was 3·0% (four of 135; 0·8–7·4), and none were detected among the MLH1 carriers, MLH1 non-carrier controls, and MSH6 non-carrier controls. Prostate cancer incidence, using a PSA threshold of higher than 3·0 ng/mL, was higher in MSH2 carriers than in MSH2 non-carrier controls (4·3% vs 0·5%; p=0·011) and MSH6 carriers than MSH6 non-carrier controls (3·0% vs 0%; p=0·034). The overall positive predictive value of biopsy using a PSA threshold of 3·0 ng/mL was 51·4% (95% CI 34·0–68·6), and the overall positive predictive value of a PSA threshold of 3·0 ng/mL was 32·1% (20·3–46·0).InterpretationAfter the first screening round, carriers of MSH2 and MSH6 pathogenic variants had a higher incidence of prostate cancer compared with age-matched non-carrier controls. These findings support the use of targeted PSA screening in these men to identify those with clinically significant prostate cancer. Further annual screening rounds will need to confirm these findings.FundingCancer Research UK, The Ronald and Rita McAulay Foundation, the National Institute for Health Research support to Biomedical Research Centres (The Institute of Cancer Research and Royal Marsden NHS Foundation Trust; Oxford; Manchester and the Cambridge Clinical Research Centre), Mr and Mrs Jack Baker, the Cancer Council of Tasmania, Cancer Australia, Prostate Cancer Foundation of Australia, Cancer Council of Victoria, Cancer Council of South Australia, the Victorian Cancer Agency, Cancer Australia, Prostate Cancer Foundation of Australia, Asociación Española Contra el Cáncer (AECC), the Instituto de Salud Carlos III, Fondo Europeo de Desarrollo Regional (FEDER), the Institut Català de la Salut, Autonomous Government of Catalonia, Fundação para a Ciência e a Tecnologia, National Institutes of Health National Cancer Institute, Swedish Cancer Society, General Hospital in Malmö Foundation for Combating Cancer.

Purpose: Androgen deprivation therapy (ADT) is the linchpin therapeutic for locally advanced and metastatic PCa. However, resistance to ADT is common and has been associated with a gain of function mutation (A1245C) in the 3β-HSD enzyme, which catalyzes intratumoral DHT synthesis. As androgen signaling is known to upregulate the DNA damage response (DDR), we investigated whether variant HSD3B1 modulates DDR and radiosensitivity in PCa via extragonadal androgen biosynthesis. Methods: We stably knocked down HSD3B1 in LNCaP, C42 and VCaP cell lines (which carry the protein stabilizing variant allele), and overexpressed the variant HSD3B1 allele in LAPC4 (that harbors WT allele, where the protein readily undergoes degradation). We examined the proliferative and clonogenic capacity of these cells in presence and absence of substrate, the extragonadal androgen precursors DHEA, followed by treatment with IR (200,400, 600 and 800 cGy, single fraction). We studied DNA DSB formation and resolution kinetics using phospho- γH2AX labelling and neutral COMET assay. We also measured 3β-HSD dependent changes in expression of DDR response genes pre- and post-radiation Results: Control shRNA transduced prostate cancer cell lines expressing the variant HSD3B1 allele had increased cell proliferation (3.1289-fold, p<0.001) and clonogenic survival (100 x more colonies after 800 cGY single fraction, p&lt 0.001) in the presence of DHEA compared to HSD3B1 knockdown LNCaP cells. We found variant HSD3B1 cell lines were more radioresistant to 400cGY single fraction IR and exhibited more efficient γH2AX foci resolution with significantly low no. of residual foci (1 foci/nucleus, p &lt 0.05) and smaller COMET Tail moment at 24 hrs in a DHEA dependent. We observe increased basal mRNA expression of DDR genes from specific repair networks including non-homologous end joining (PRKDC, XRCC4, XRCC5) and homologous recombination (RAD51, RAD54), but not mismatch repair (MSH2, MSH6), in variant HSD3B1 cells. Transcriptional induction of DDR gene expression following radiation and in presence of DHEA, was significantly more pronounced in HSD3B1 variant cells, suggesting AR establishes a regional chromatin environment which is more permissive to DDR gene transcriptional activation. Conclusion: Increased intracellular HSD3B1, which converts extra gonadal precursors to DHT, leads to increased expression of NHEJ and HR genes, more rapid resolution of γH2AX foci, and radioresistance in prostate cancer. This work has therapeutic implications for combined radiation and androgen directed therapy in localized and low volume metastatic prostate cancer. Prospective validation of personalized treatment strategies combining blockade of extragonadal androgen biosynthesis, conventional ADT, and radiotherapy in prostate cancer are warranted. Citation Format: Shinjini Ganguly, Aysegul Balyimez, Zaeem lone, Aimalie Hardaway, Monaben Patel, Elai Davicioni, Rahul Tendulkar, Eric Klein, Nima Sharifi, Omar Mian. Extragonadal and regional androgen biosynthesis associated with a common steroidogenic enzyme polymorphism promotes radioresistance in prostate cancer [abstract]. In: Proceedings of the Annual Meeting of the American Association for Cancer Research 2020; 2020 Apr 27-28 and Jun 22-24. Philadelphia (PA): AACR; Cancer Res 2020;80(16 Suppl):Abstract nr 2082.

Purpose: Radiotherapy (RT)-based bladder preservation is an alternative to perioperative chemotherapy and radical cystectomy (RC) for selected patients with muscle-invasive bladder cancer (MIBC). Recent data suggest that somatic DNA damage response (DDR) alterations are associated with improved outcome in MIBC patients receiving RC +/- perioperative chemotherapy. Improved understanding of the influence of DDR genetics on response to RT-based bladder preservation may aid patient selection. Methods: We performed deep, targeted capture sequencing of primary bladder tumors and paired/pooled normal specimens from 48 RT treated MIBC patients with a DDR gene-enriched panel. To test whether correlations of DDR alterations with outcomes were specific to a RT cohort, we also assessed a subset of these DDR genes that were sequenced in a previously published series of 89 patients who received RC +/- neoadjuvant platinum-based chemotherapy. Specimens were re-reviewed to confirm urothelial histology. Deleterious alterations were defined as somatic nonsense, frameshift, or splice site mutations, or missense mutations at or near functionally significant residues validated in the literature. Results: In the RT cohort, median RT dose was 66 Gy, and chemotherapy use was neoadjuvant and concurrent (46%), concurrent only (48%), or none (6%). Visibly complete TURBT was achieved in 71% of patients. Median surviving follow up was 28 months. There were 30 progression events (crude rate 63%), comprised of 22 metastases (crude 46%) and 24 local in-bladder recurrences (crude 50%). While 30 (63%) patients had alterations in DDR genes, only 13 (27%) patients had deleterious DDR alterations, specifically in ATM (2), BRCA1 (1), BRIP1 (1), ERCC2 (6), FANCD2 (1), and PALB2 (1). On multivariable Cox proportional hazards analysis, the presence of a deleterious DDR alteration was associated significantly with lower relapse risk (HR 0.28, 95% CI 0.08-0.95; p = 0.041), as well as with a trend towards lower risk for metastasis (HR 0.32, 95% CI 0.10-1.11; p = 0.07) and any disease progression (HR 0.35, 95% CI 0.12-1.03; p = 0.06). In the RC cohort, neoadjuvant chemotherapy was given in 39% of patients, and deleterious DDR gene alterations were noted only in 8% of the patients. On 2-sided log-rank testing, such alterations conferred a non-significant trend for improved recurrence-free survival (p = 0.20) and disease-specific survival (p = 0.10). Conclusion: Deleterious somatic alterations in DDR genes were associated with significantly improved outcomes in bladder cancer patients undergoing RT-based therapy and with a trend for improved outcomes in those treated with RC +/- platinum chemotherapy. Further research is warranted to validate these findings on an independent RT-treated dataset and to clarify the relationship in a larger chemotherapy-treated RC cohort. Citation Format: Neil B. Desai, Gopa Iyer, Eugene K. Cha, Sasinya N. Scott, Joseph Hreiki, John P. Sfakianos, Philip Kim, Aditya Bagroida, Bernard H. Bochner, Jonathan E. Rosenberg, Dean F. Bajorin, Michael F. Berger, Marisa A. Kollmeier, Hikmat Al-Ahmadie, David B. Solit. Deleterious alterations in DNA damage response genes are associated with improved outcome in muscle-invasive bladder cancer patients treated with radiation-based bladder preservation. [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 602. doi:10.1158/1538-7445.AM2015-602