Pathogenic ATM Mutations Research Articles

Survival impact of homologous recombination deficiency in veterans with cholangiocarcinoma including mutual exclusivity with pathogenic KRAS and TP53.

546 Background: Cholangiocarcinoma (CCA) has poor prognosis and limited treatment options. The US Veteran Health Administration’s (VHA) National Precision Oncology Program (NPOP) was established to characterize pathogenic drivers across the integrated VA network. Here, we summarize an important subgroup of CCA with homologous recombination deficiency (HRD) and its impact on overall survival (OS). Methods: Next-generation sequencing (NGS) was performed using FoundationOne CDx Analysis included clinicopathological, molecular alterations including, single nucleotide variants, gene amplification/loss, and gene fusion/rearrangements obtained from centralized data of the VHA’s Corporate Data Warehouse. Homologous recombination deficiency was defined by pathogenic variant mutations (MT) in ARID1A, ATM, ATR, BAP1, BRCA1, BRCA2, CDK12, CHEK2, FANC, NBN, PALB2, and RAD51. Statistical analysis was performed using Chi-squared test for concurrent gene enrichment and log-rank Kaplan-Meier analysis. Results: 483 CCA samples underwent tissue NGS as standard of care. Pathogenic variants in HRD genes included ARID1A (15.5%, n=75), BAP1 (9.5%, n=46), ATM (2.7%, n=13), PALB2 (2.3%, n=11) and BRCA (2.0%, n=10), representing 31.7% (n=153) of the population. HRD mutations were found in similar proportions of localized (31.8%) and metastatic (31.5%) clinical presentations at diagnosis. OS was not significantly different between CCA with HRD mutations (12.3 mo WT vs. 13.8 months with HRD MT, p=64). KRAS MT (n=77, 15.9% of population) did not impact OS (WT 12.3 mo v. MT 13.1 mo) including subgroup analysis stratified by HRD status. There was increased frequency in KRAS MT with HRD WT 18.5% versus HRD MT10.4% (p<0.03). Additionally, there was increased frequency in TP53 MT with HRD WT 54.8% versus HRD MT 28.1% (p<0.00001). TP53 MT (n=224, 46.3% of population) conferred inferior OS (WT 12.3 mo v. MT 13.1 mo) including subgroup analysis stratified by HRD status. Subgroup analysis of TP53 MT had worsened OS with concurrent HRD MT (n=39, 4.1 mo) v. HRD WT (n=158, 9.0 mo; p<0.04). Conclusions: Across a diverse integrated healthcare system of Veterans, mutations associated with HRD were found to be common, however, not prognostic in OS across a diverse patient population of CCA. Further work is needed to clarify exposure to platinum containing chemotherapy. TP53 MT CCA remains a critical unmet need that drives worsened outcomes including here in patients with mutations in HRD. Future analysis should consider mutation signatures of HRD, loss in heterozygosity of HRD, and platinum exposure to further define the subgroups of CCA with mutations in HRD. [Table: see text]

Identification of a Predictive Genomic Biomarker for Prostate Directed Therapy in Synchronous Low-Volume Metastatic Castration Sensitive Prostate Cancer.

Standard of care management for metastatic castration sensitive prostate cancer (mCSPC) includes androgen deprivation therapy (ADT) with docetaxel or second-generation anti-androgen therapy. Recently, randomized data has demonstrated radiotherapy to the prostate is associated with an improvement in overall survival among patients with low-volume metastatic disease. Tumor genomics represents an additional dimension to understand the clinical trajectory of patients with mCSPC. Herein we aim to evaluate a high-risk genomic signature for its ability to predict response to prostate directed therapy (PDT). We performed a single institution retrospective review of men with low-volume mCSPC who underwent next-generation sequencing of their tumor. Patients were classified according to the presence of high-risk (HiRi) mutation including pathogenic mutations in either TP53, ATM, BRCA1/2, or Rb1. Our primary endpoint was to determine the effect of PDT on overall survival (OS) in patients with and without a HiRi mutation. Survival analysis was performed with the Kaplan-Meier method compared with log-rank test and multivariable cox regression. Interaction between HiRi mutation and PDT was evaluated. A total of 101 patients with synchronous low-volume CSPC were included in our analysis with a median follow-up of 44 months. Approximately half of patients were found to have a HiRi pathogenic mutation (48.5%) with TP53 mutations accounting for 75.5% of HiRi mutations. On multivariable cox regression PDT was associated with improvement in OS (HR = 0.37, 95% CI 0.16-0.88; p = 0.03). When stratified by presence of HiRi mutation, PDT was not associated with any clinical outcome. Patients with HiRi mutations demonstrated a median OS of 73 vs 66.8 months (p = 0.28) for no PDT and PDT, respectively. Conversely, patients without a HiRi mutation demonstrated a significant improvement in median OS of 60 vs 105.3 months (p<0.01) for no PDT and PDT, respectively. The p-value for interaction for OS between PDT and HiRi mutation was statistically significant (p<0.01). Here we have identified a high-risk genomic biomarker that appears predictive for response to PDT in men with synchronous low-volume mCSPC. Further work validating these results with prospective randomized data is warranted.

Identification of a Predictive Genomic Biomarker for Prostate-directed Therapy in Synchronous Low-volume Metastatic Castration-sensitive Prostate Cancer

BackgroundStandard of care management for synchronous metastatic castration-sensitive prostate cancer (mCSPC) includes androgen deprivation therapy with a second-generation antiandrogen therapy and/or docetaxel. Recently, randomized data have demonstrated that prostate-directed therapy (PDT) is associated with an improvement in overall survival (OS) among patients with low-volume metastatic disease. Tumor genomics represents an additional dimension to define the clinical trajectory of patients with mCSPC. ObjectiveTo evaluate a high-risk (HiRi) genomic signature to predict the benefit from PDT. Design, setting, and participantsWe performed a single-institution retrospective review of men with synchronous low-volume mCSPC who underwent DNA panel sequencing of their tumor. Patients were classified according to the presence of HiRi mutation including pathogenic mutations in TP53, ATM, BRCA1, BRCA2, or Rb1. Outcome measurements and statistical analysisThe primary endpoint was to determine the effect of PDT on OS in patients with and without a HiRi mutation. A survival analysis was performed with the Kaplan-Meier method compared with log-rank test and multivariable Cox regression. The interaction between HiRi mutation and PDT was evaluated. Results and limitationsA total of 101 patients with synchronous low-volume CSPC were included with a median follow-up of 44 mo. Approximately half of patients were found to have a HiRi pathogenic mutation (49%). Patients with HiRi mutations demonstrated median OS of 73 versus 66.8 mo (p = 0.3) for no PDT versus PDT. Conversely, patients without a HiRi mutation demonstrated a significant improvement in OS of 60 versus 105.3 mo (p < 0.001) for no PDT versus PDT. The p value for interaction for OS between PDT and HiRi mutation was statistically significant (p < 0.001). Limitations include the retrospective nature of the study. ConclusionsHere, we have identified a HiRi genomic biomarker that appears predictive for the lack of benefit from PDT in men with synchronous low-volume mCSPC. Further work validating these results is warranted. Patient summaryIn this report, we evaluated a high-risk genomic biomarker to predict the benefit from prostate-directed therapy for men with synchronous low-volume metastatic castration-sensitive prostate cancer. We found that men without a high-risk mutation appear to experience a greater clinical benefit from prostate-directed therapy than those with a high-risk mutation.

Abstract 6603: BRCA1 promoter methylation in sporadic breast cancer patients detected by liquid biopsy

Abstract Background: BRCA1 promoter methylation (PM) is an early initiating event in cancer, occurring in 3 to 65.2% of all breast tumors, and 30 to 65% of triple negative tumors. BRCA1 PM has been associated with defective homologous recombination repair (HRR), early onset of breast and ovarian cancer, and improved clinical response to adjuvant chemotherapy. Historically, there has been no diagnostic assay that comprehensively evaluates both BRCA1 PM and genomic alterations in cell-free circulating tumor DNA (ctDNA). Here, we describe the novel detection of BRCA1 PM and genomic alterations in a cohort of patients with breast cancer using GuardantINFINITY, a liquid biopsy assay interrogating 800+ genes and genome-wide methylation detection. Method: We assessed for BRCA1 PM in ctDNA from 274 patients with late-stage breast cancer. Genomic sequencing of 800+ genes and PM profiling of 398 genes was performed by GuardantINFINITY. The positive calling threshold for PM was established by comparing cell-free DNA derived from patients with cancer and cancer-free donors. The limit of detection (LoD) was determined through in silico and experimental titrations of ctDNA from clinical samples and cell lines with known gene PM into the plasma of cancer-free donors. Results: Among the 274 patients with advanced breast cancer, 8 (2.9%) had germline pathogenic mutations in BRCA1, BRCA2, or ATM. BRCA1 PM was detected in 11/274 (4.0%) patients at the predefined threshold of &amp;gt;99% specificity. BRCA1 PM detection in this cohort was 8.9% (8/90) when excluding samples with low tumor shedding (&amp;lt;1% epigenomic tumor fraction in cfDNA). Among the 11 patients with BRCA1 PM detected in ctDNA, one had a co-occurring somatic BRCA1 nonsense variant (p.S361*); none of the remaining patients with BRCA1 PM had another HRR-related mutation detected in cfDNA. Among patients without BRCA1 PM detected, pathogenic somatic alterations were detected in BRCA2, ATM, and CHEK2 in 25 (9.4%) patients. In silico simulations using clinical samples with BRCA1 PM indicated an LoD of 0.0408%. BRCA1 PM was not detected in 3210 individual and mixed cancer-free clinical samples, indicating a high specificity for BRCA1 PM calls. Conclusion: GuardantINFINITY, a plasma-based diagnostic assay, detected both BRCA1 PM and genomic alterations in this unspecified advanced breast cancer cohort. The BRCA1 PM detection rates of 4.0-8.9% are consistent with values previously reported in the literature. As BRCA1 PM has important prognostic and therapeutic implications for the management of breast (as well as ovarian) cancers, additional studies are warranted to further describe the PM patterns across breast cancer subtypes and how these patterns both influence and are influenced by disease evolution and therapeutic response. Liquid biopsy thus serves as a suitable method to noninvasively identify and monitor changes in both genomics and epigenomics. Citation Format: Jennifer Yen, Sai Chen, Colby Jenkins, Brooke Overstreet, Yu Fu, Jun Zhao, Tingting Jiang, Leylah Drusbosky, Stephen Pettitt, Michael Dorschner, Lauren Lawrence, Han-Yu Chuang, Andrew Tutt. BRCA1 promoter methylation in sporadic breast cancer patients detected by liquid biopsy [abstract]. In: Proceedings of the American Association for Cancer Research Annual Meeting 2023; Part 1 (Regular and Invited Abstracts); 2023 Apr 14-19; Orlando, FL. Philadelphia (PA): AACR; Cancer Res 2023;83(7_Suppl):Abstract nr 6603.

Impact of clonal hematopoiesis on tumor control following radiation therapy.

3145 Background: Clonal hematopoiesis (CH) has well established associations with adverse clinical outcomes including all-cause mortality, cardiovascular disease, and progression to hematologic malignancy. The presence of CH has also been demonstrated to adversely impact survival from non-hematologic cancers, however whether CH may modulate response to radiation therapy (RT) in solid tumors is not known. Here we investigate the potential impact of CH mutations on radiation outcomes. Methods: We analyzed data from two previously well annotated cohorts of patients with tumors harboring somatic ATM mutations (n = 358) and FAT1 mutations (n = 365) who received RT and underwent prospective tumor and matched WBC sequencing utilizing the MSK-IMPACT assay. CH variants were detected in the blood samples utilizing a well-validated variant detection and filtration pipeline. Given that pathogenic mutations in ATM have been shown to be strongly associated with improved response to RT, these patients were excluded to avoid confounding. Additionally, patients with blood sampling for CH assessment that occurred more than 6 months after RT were excluded to address the possibility of therapy-related CH. We compared outcomes including irradiated tumor progression in patients with and without CH. Results: The final analysis consisted of 412 patients who underwent 811 total courses of radiation. A wide spectrum of solid tumor types were represented, most commonly non-small cell lung cancer (32.5%) and breast cancer (11.9%). A total of 161 patients (39.0%) had CH, with the most commonly mutated genes being DNMT3A (25.6%), PPM1D (6.2%), TET2 (5.8%), and TP53 (5.0%), consistent with prior studies of CH. Patients with CH were older at blood sample collection (67.6 vs 60.2 years, p &lt; 0.001), reflecting an expected increase in CH burden with age. Fine Gray competing risks analysis, with death treated as a competing event and with clustering around patient identifier, showed no difference in irradiated tumor progression between patients with and without CH (HR 1.03, 95% CI 0.69 – 1.53, p = 0.896). Similarly, subanalyses by CH variant allele frequency and putative CH-driver mutations did not reveal an association between CH and response to RT. A hypothesis generating subgroup analysis by common cancer types, however, suggested that CH was associated with increased risk of progression post-radiation in prostate (HR 4.68, 1.14 – 19.1) and thyroid (HR 3.13, 1.55 – 6.34) cancer cohorts, warranting further investigation. Conclusions: We found no difference in irradiated tumor progression among patients who did and did not have CH. There may be an association between CH and poor radiation outcomes in certain cancer types, and further studies are needed to clarify the specific clinical and genomic factors that may influence radiation response.

Germline mutations in penetrant cancer predisposition genes are rare in men with prostate cancer selecting active surveillance.

Pathogenic germline mutations in several rare penetrant cancer predisposition genes are associated with an increased risk of aggressive prostate cancer (PC). Our objectives were to determine the prevalence of pathogenic germline mutations in men with low-risk PC on active surveillance, and assess whether pathogenic germline mutations associate with grade reclassification or adverse pathology, recurrence, or metastases, in men treated after initial surveillance. Men prospectively enrolled in the Canary Prostate Active Surveillance Study (PASS) were retrospectively sampled for the study. Germline DNA was sequenced utilizing a hereditary cancer gene panel. Mutations were classified according to the American College of Clinical Genetics and Genomics' guidelines. The association of pathogenic germline mutations with grade reclassification and adverse characteristics was evaluated by weighted Cox proportional hazards modeling and conditional logistic regression, respectively. Overall, 29 of 437 (6.6%) study participants harbored a pathogenic germline mutation of which 19 occurred in a gene involved in DNA repair (4.3%). Eight participants (1.8%) had pathogenic germline mutations in three genes associated with aggressive PC: ATM, BRCA1, and BRCA2. The presence of pathogenic germline mutations in DNA repair genes did not associate with adverse characteristics (univariate analysis HR=0.87, 95% CI: 0.36-2.06, p=0.7). The carrier rates of pathogenic germline mutations in ATM, BRCA1, and BRCA2did not differ in men with or without grade reclassification (1.9% vs. 1.8%). The frequency of pathogenic germline mutations in penetrant cancer predisposition genes is extremely low in men with PC undergoing active surveillance and pathogenic germline mutations had no apparent association with grade reclassification or adverse characteristics.

Association between pathogenic germline mutations in BRCA2 and ATM and tumor-infiltrating lymphocytes in primary prostate cancer.

Pathogenic mutations in homologous recombination (HR) DNA repair genes may be associated with increased tumor mutational burden and numbers of tumor-infiltrating lymphocytes (TIL). Though HR-deficient prostate tumors have been anecdotally associated with improved responses to immunotherapy, it is unclear whether HR mutations or HR deficiency (HRD) scores predict for increased T-cell densities in this cancer. We evaluated 17 primary prostate tumors from patients with pathogenic germline BRCA2 mutations (gBRCA2) and 21 primary prostate tumors from patients with pathogenic germline ATM (gATM) mutations, which were compared to 19 control tumors lacking HR gene mutations, as well as the TCGA prostate cancer cohort. HRD score was estimated by targeted sequencing (gBRCA2 and gATM) or by SNP microarray (TCGA). Tumor-associated T-cell densities were assessed using validated automated digital image analysis of CD8 and FOXP3 immunostaining (gBRCA2 or gATM) or by methylCIBERSORT (TCGA). CD8 + and FOXP3 + T-cell densities were significantly correlated with each other in gBRCA2 and gATM cases. There was no significant difference between CD8 + or FOXP3 + TIL densities in gBRCA2 or gATM cases compared to controls. In the TCGA cohort, HRD score was associated with predicted CD8 + and FOXP3 + TILs. Associations were also seen for HRD score and TIL density among the germline-mutated cases. In contrast to mismatch repair-deficient primary prostate tumors, cancers from germline BRCA2 or ATM mutation carriers do not appear to be associated with elevated TIL density. However, measures of genomic scarring, such as HRD score, may be associated with increased tumor-infiltrating T-cells.

Association of ATM mutations in metastatic prostate cancer with differential genomic alteration profiles from homologous recombination deficient and proficient tumors.

5063 Background: ATM mutations, one of a family of DNA repair defects prevalent in prostate cancer, have been included in a list of actionable mutations for PARP inhibitor (PARPi) therapeutic trials. Despite preclinical evidence, PARPi have shown minimal clinical activity in ATM mutant prostate cancer (ATMmPCa). The present analysis explores co-occurring genomic alterations that may drive outcomes of metastatic PCa (mPCa) patients with tumors harboring ATM mutations and provide clues for understanding therapy resistance and potential targets. Methods: This study included molecular profiling analysis of 1375 cases of mPCa. Tumors were analyzed using next-generation sequencing (NGS), whole transcriptome sequencing (WTS), and immunohistochemistry (IHC) (Caris Life Sciences, Phoenix, AZ). dMMR/MSI-H status was determined by IHC, NGS, and fragment analysis and tumor mutational burden (TMB) was calculated based on somatic nonsynonymous missense mutations. We performed differential gene expression analysis of HR-associated transcripts such as ATR, PARP1-3, RAD50, RAD51A/B/C/D and RAD54. Significance was determined using the ꭓ2 test and Benjamini-Hochberg method. Results: Fifty-nine (4.2%) cases harbored pathogenic ATM mutations, 84 (6.2%) harbored BRCA2 mutations. 1018 tumors (74%) were deemed homologous recombination proficient (HRP) and 155 tumors (11.3%) were HR Deficient (HRD); harboring one or more mutation in HR-related genes excluding ATM and BRCA2. The mutation rate of TP53 was significantly lower in ATMmPCa (12.0%) compared to BRCA2mPCa (35%), HRD (35%) and HRP (46.6%) tumors. ATMmPCa showed higher rates of SMAD2 (3.7%/1%) and FLCN (5.2%/0.3%) alterations compared to HRP cases. PARP1 and RAD51D gene expression was reduced in ATMmPCa compared to HRP (p &lt; 0.05) and BRCA2mPCa ( p &lt; 0.05) tumors, respectively. No differences in gene expression levels were detected for ATR, PARP2, PARP3, RAD50, and RAD54. Chromosomal segments demonstrating differential CNA in ATMmPCa vs HRP, HRD, or BRCA2mPCa included FGF19, FGF4, PTPN11, ALDH2, DAXX, BCL7A, CCND1, BMPR1A and MEF2B (Q-value &lt; 0.05 determined by ꭓ2). The most common CNA in ATMmPCa was CCND1, present in approximately 13% (7/55) of cases. Compared to BRCA2mPCa and HRD cases, ATMmPCa cases are less likely to display markers of immunotherapy response such as dMMR/MSI-H or TMB ≥10 mutations/MB. Conclusions: ATMmPCa demonstrated several differences in co-occurring alterations compared to BRCA2mPCa, HRD and HRP mPCa. ATMmPCa tumors were less likely to harbor alterations in TP53 compared to BRCA2, HRD or HRP tumors. CNA in ATMmPCa occurred in 9 genes across distinct mPCa molecular subtypes and were enriched for those associated with the 11q13 amplicon harboring Cyclin D1. The FGF and PTPN11 related pathways are potentially targetable pathways in ATMmPC and may merit further study.

Vulnerability to low-dose combination of irinotecan and niraparib in ATM-mutated colorectal cancer

BackgroundDespite the advancements in new therapies for colorectal cancer (CRC), chemotherapy still constitutes the mainstay of the medical treatment. For this reason, new strategies to increase the efficacy of chemotherapy are desirable. Poly-ADP-Ribose Polymerase inhibitors (PARPi) have shown to increase the activity of DNA damaging chemotherapeutics used in the treatment of CRC, however previous clinical trials failed to validate these results and pointed out dose-limiting toxicities that hamper the use of such combinations in unselected CRC patients. Nevertheless, in these studies little attention was paid to the mutational status of homologous recombination repair (HRR) genes.MethodsWe tested the combination of the PARPi niraparib with either 5-fluorouracil, oxaliplatin or irinotecan (SN38) in a panel of 12 molecularly annotated CRC cell lines, encompassing the 4 consensus molecular subtypes (CMSs). Synergism was calculated using the Chou-Talalay method for drug interaction. A correlation between synergism and genetic alterations in genes involved in homologous recombination (HR) repair was performed. We used clonogenic assays, mice xenograft models and patient-derived 3D spheroids to validate the results. The induction of DNA damage was studied by immunofluorescence.ResultsWe showed that human CRC cell lines, as well as patient-derived 3D spheroids, harboring pathogenic ATM mutations are significantly vulnerable to PARPi/chemotherapy combination at low doses, regardless of consensus molecular subtypes (CMS) and microsatellite status. The strongest synergism was shown for the combination of niraparib with irinotecan, and the presence of ATM mutations was associated to a delay in the resolution of double strand breaks (DSBs) through HRR and DNA damage persistence.ConclusionsThis work demonstrates that a numerically relevant subset of CRCs carrying heterozygous ATM mutations may benefit from the combination treatment with low doses of niraparib and irinotecan, suggesting a new potential approach in the treatment of ATM-mutated CRC, that deserves to be prospectively validated in clinical trials.

Pathogenic germline variants are associated with poor survival in stage III/IV melanoma patients

Patients with late stage resected cutaneous melanoma have poor overall survival (OS) and experience irreversible adverse events from systemic therapy. There is a clinical need to identify biomarkers to predict outcome. Performing germline/tumour whole-exome sequencing of 44 stage III/IV melanoma patients we identified pathogenic germline mutations in CDKN2A, CDK4, ATM, POLH, MRE11A, RECQL4 and XPC, affecting 7/44 patients. These mutations were associated with poor OS (p = 0.0082). We confirmed our findings in The Cancer Genome Atlas (TCGA) human skin cutaneous melanoma cohort where we identified pathogenic variants in 40/455 patients (p = 0.0203). Combining these cohorts (n = 499) further strengthened these findings showing germline carriers had worse OS (p = 0.0009). Additionally, we determined whether tumour mutation burden (TMB) or BRAF status were prognostic markers of survival. Low TMB rate (< 20 Mut/Mb; p = 0.0034) and BRAF p.V600 mutation (p = 0.0355) were associated with worse progression-free survival. Combining these biomarkers indicated that V600 mutant patients had significantly lower TMB (p = 0.0155). This was confirmed in the TCGA (n = 443, p = 0.0007). Integrative analysis showed germline mutation status conferred the highest risk (HR 5.2, 95% CI 1.72–15.7). Stage IV (HR 2.5, 0.74–8.6) and low TMB (HR 2.3, 0.57–9.4) were similar, whereas BRAF V600 status was the weakest prognostic biomarker (HR 1.5, 95% CI 0.44–5.2).

Abstract 3536: Genetic validation of an ATM immunohistochemistry assay for genomic and clinical-pathologic characterization of primary prostate cancers with ATM loss

Abstract Background: ATM, a protein kinase involved in double-strand DNA break repair, is mutated in a significant fraction of prostate cancers, and targeted therapies such as ATR inhibitors appear promising for ATM-deficient tumors. However, few ATM-deficient primary prostate cancers have been clinically and genomically characterized. Design: We validated an automated and dichotomously-scored clinical-grade immunohistochemistry (IHC) assay to detect ATM protein loss using prostate cancer cell lines and 52 very high-grade prostate tumors (Gleason score 5+4=9 or 5+5=10) with known ATM genomic status. We examined the frequency of ATM protein loss among 29 tumors with known pathogenic germline ATM mutations or variants of unknown significance (VUS). Finally, we screened more than 800 additional primary prostate carcinomas using tissue microarrays (TMA) for ATM protein loss. Targeted somatic genomic sequencing using HRD Plus (Myriad Genetics) or UW-OncoPlex was conducted in a subset of immunostained tumors in each cohort. Results: ATM protein loss by IHC was found in 13% (7/52) of very high-grade (primary Gleason pattern 5) tumors with somatic genomic sequencing data. Of 7 cases with ATM protein loss, 4 had a deleterious ATM mutation and 3 had apparent single copy ATM genomic loss. Of the remaining 45 cases without ATM protein loss, none had ATM genomic alterations. In the cohort of tumors with pathogenic germline ATM mutations, 74% (17/23) had ATM protein loss, but only 45% (9/20) with evaluable sequencing had a second pathogenic alteration in ATM detected (all of these had protein loss). Of those with germline mutations and ATM protein loss, the majority (76% or 13/17) had an apparent clonal alteration with homogeneous protein loss in all tumor cells sampled. In contrast, among 6 cases with germline ATM VUS, only one had protein loss. On TMA analysis, ATM loss was present in 3% (25/824) of evaluable tumors overall, and was significantly more common in tumors from Grade Group 5 (12/130; 9.2%) compared to all other Grade Groups (13/694; 1.9%) (P&amp;lt;0.0001). Of tumors found by TMA screening with available sequencing, 80% (4/5) with homogeneous ATM protein loss and 58% (7/12) with heterogeneous ATM protein loss had detectable pathogenic alterations in ATM. Of these, 82% (9/11) were inferred to have somatic ATM variants. Conclusions: Validated ATM IHC is a sensitive assay for detecting underlying ATM alterations and may be useful to evaluate VUS in ATM. However, not all tumors with ATM protein loss have detectable genomic alterations. ATM protein loss is likely an early clonal event in tumors with germline mutations and is significantly enriched in high-grade prostate cancers. Citation Format: Harsimar Kaur, Jessica Hicks, Colin Pritchard, Angelo De Marzo, William Isaacs, Jerry Lanchbury, Kirsten Timms, Emmanuel Antonarakis, Tamara L. Lotan. Genetic validation of an ATM immunohistochemistry assay for genomic and clinical-pathologic characterization of primary prostate cancers with ATM loss [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 3536.

Pathogenic ATM Mutations in Cancer and a Genetic Basis for Radiotherapeutic Efficacy.

Radiation therapy is one of the most commonly used cancer therapeutics but genetic determinants of clinical benefit are poorly characterized. Pathogenic germline variants in ATM are known to cause ataxia-telangiectasia, a rare hereditary syndrome notable for marked radiosensitivity. In contrast, somatic inactivation of ATM is a common event in a wide variety of cancers, but its clinical actionability remains obscure. We analyzed 20 107 consecutively treated advanced cancer patients who underwent targeted genomic sequencing as part of an institutional genomic profiling initiative and identified 1085 harboring a somatic or germline ATM mutation, including 357 who received radiotherapy (RT). Outcomes of irradiated tumors harboring ATM loss-of-function (LoF) mutations were compared with those harboring variants of unknown significance. All statistical tests were 2-sided. Among 357 pan-cancer patients who received 727 courses of RT, genetic inactivation of ATM was associated with improved radiotherapeutic efficacy. The 2-year cumulative incidence of irradiated tumor progression was 13.2% vs 27.5% for tumors harboring an ATM LoF vs variant of unknown significance allele, respectively (hazard ratio [HR] = 0.51, 95% confidence interval [CI] = 0.34 to 0.77, P = .001). The greatest clinical benefit was seen in tumors harboring biallelic ATM inactivation (HR = 0.19, 95% CI = 0.06 to 0.60, P = .005), with statistically significant benefit also observed in tumors with monoallelic ATM inactivation (HR = 0.57, 95% CI = 0.35 to 0.92, P = .02). Notably, ATM LoF was highly predictive of outcome in TP53 wild-type tumors but not among TP53-mutant tumors. We demonstrate that somatic ATM inactivation is associated with markedly improved tumor control following RT. The identification of a radio-sensitive tumor phenotype across multiple cancer types offers potential clinical opportunities for genomically guided RT.

ATM mutation carriers and family history of pancreatic cancer.

1529 Background: Multigene panel testing (MGT) is commonly utilized in patients with a personal or family history of cancer. One of the more common gene mutations identified is in the ATM gene, associated with a moderately increased risk of breast and other cancers. There are reports of an association with pancreatic cancer, however the exact risks are unclear. The aim of this study is to describe the family history of pancreatic cancer in a cohort of ATM mutation carriers, and to evaluate possible genotype/phenotype correlation. Methods: Patients who underwent MGT, between ‘13 and ‘19, and tested positive for a pathogenic/likely pathogenic ATM mutation were included in this study. Family history, with a focus on pancreatic cancer, and genetic testing results were analyzed. Results: A total of 114 patients were identified to carry an ATM mutation. Twenty-two (19.3%) individuals had a family history of pancreatic cancer in a close relative, and of those, 13 (11.4%) had an affected first degree relative, and 11 (9.6%) had an affected second degree relative. Among the families with pancreatic cancer, 20 close relatives had a personal history of pancreatic cancer, with the youngest diagnosed at age 40, the oldest diagnosed at age 91, and a mean age of diagnosis of 66.5 years. Thirteen unique variants were identified: 4 splice site, 3 missense, 3 frameshift, 1 nonsense, and 1 silent. Two families had the known high-penetrance ATM mutation, c.7271T &gt; C (p.V2424G). Conclusions: This study describes the association of pancreatic cancer in individuals found to carry pathogenic ATM mutations. A significant proportion (19.3%) of patients had a family history of pancreatic cancer in a close relative, diagnosed as young as age 40. The mean age of diagnosis was slightly younger than the average age in the general population (age 70). As pancreatic cancer screening continues to improve, this information will be an important component to help guide cancer risk assessment and future screening recommendations for ATM mutation carriers. Additional larger studies are needed to further characterize pancreatic cancer risks in patients with ATM gene mutations.

Mutation Status and Epithelial Differentiation Stratify Recurrence Risk in Chordoid Meningioma-A Multicenter Study with High Prognostic Relevance.

Chordoid meningioma is a rare WHO grade II histologic variant. Its molecular alterations or their impact on patient risk stratification have not been fully explored. We performed a multicenter, clinical, histological, and genomic analysis of chordoid meningiomas from 30 patients (34 tumors), representing the largest integrated study to date. By NHERF1 microlumen immunohistochemical detection, three epithelial differentiation (ED) groups emerged: #1/fibroblastic-like, #2/epithelial-poorly-differentiated and #3/epithelial-well-differentiated. These ED groups correlated with tumor location and genetic profiling, with NF2 and chromatin remodeling gene mutations clustering in ED group #2, and TRAF7 mutations segregating in ED group #3. Mutations in LRP1B were found in the largest number of cases (36%) across ED groups #2 and #3. Pathogenic ATM and VHL germline mutations occurred in ED group #3 patients, conferring an aggressive or benign course, respectively. The recurrence rate significantly correlated with mutations in NF2, as single gene, and with mutations in chromatin remodeling and DNA damage response genes, as groups. The recurrence rate was very high in ED group #2, moderate in ED group #3, and absent in ED group #1. This study proposes guidelines for tumor recurrence risk stratification and practical considerations for patient management.

Abstract 2134: Niraparib combined with abiraterone acetate inhibits the growth of BRCA2wt prostate tumors

Abstract Patients with metastatic prostate cancer may initially respond well to first- and second-generation androgen receptor (AR)-targeted therapies, but many will experience disease progression, indicating a need for alternative therapies. A subset of these patients bears prostate tumors that harbor deleterious mutations in DNA damage repair (DDR) pathway genes, such as BRCA2. These DDR mutations render the tumors sensitive to drugs that inhibit poly (ADP-ribose) [PAR] polymerase-1 (PARP1), inducing synthetic lethality. Studies indicate that there are interactions between AR signaling pathways and DDR pathways in prostate cancer cells, supporting the hypothesis that dual AR/PARP inhibition could inhibit prostate tumor growth more potently than single agents, regardless of the DDR status of the tumor. This hypothesis was tested using well-known human and mouse prostate tumor models that are sensitive to androgen deprivation. Genotyping of the models confirmed that none of them carried homozygous pathogenic mutations in DDR genes, including BRCA1, BRCA2, and ATM. When the cell lines were incubated in vitro with niraparib, a potent and highly selective PARP1/2 inhibitor, PAR formation was significantly inhibited, and there was induction of γH2AX in treated cells, indicating the drug induced DNA double-stranded breaks. In addition, the cytotoxicity IC50 values for niraparib were in the range of 1-10 µM, as expected for cells that are DDRwt. In vivo efficacy studies demonstrated that the VCaP and Myc-CaP tumor models were insensitive or modestly sensitive to single agent treatment with niraparib or abiraterone acetate (AA). In contrast, the combination treatment of niraparib plus AA significantly reduced tumor growth and increased survival (p&amp;lt;0.05) as compared with control or single agent arms. Studies are ongoing to understand the mechanisms of tumor growth control mediated by the combination of these two agents. The results support the hypothesis that prostate tumors may respond to a combination treatment with niraparib, a PARP inhibitor, and AA, an androgen synthesis inhibitor, even in the absence of DDR anomalies. A Phase 3 study is planned to evaluate this hypothesis and the safety and efficacy of this combination in humans. Citation Format: Rajendra N. Damle, Rebecca Hawkins, Jennifer Hosbach, Georges Habineza Ndikuyeze, Jenny Driscoll, Natalie L. Fulton, David Derosa, Anna Hughes, Gerald Chu, Karl Calara-Nielsen, Denis Smirnov, Dong Shen, Linda A. Snyder. Niraparib combined with abiraterone acetate inhibits the growth of BRCA2wt prostate tumors [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 2134.

Frequency and phenotypic spectrum of KMT2B dystonia in childhood: A single-center cohort study.

Childhood-onset dystonia is often genetically determined. Recently, KMT2B variants have been recognized as an important cause of childhood-onset dystonia. To define the frequency of KMT2B mutations in a cohort of dystonic patients aged <18 years at onset, the associated clinical and radiological phenotype, and the natural history of disease. Whole-exome sequencing or customized gene panels were used to screen a cohort of 65 patients who had previously tested negative for all other known dystonia-associated genes. We identified 14 patients (21.5%) carrying KMT2B variants, of which 1 was classified as a variant of unknown significance. We also identified 2 additional patients carrying pathogenic mutations in GNAO1 and ATM. Overall, we established a definitive genetic diagnosis in 23% of cases. We observed a spectrum of clinical manifestations in KMT2B variant carriers, ranging from generalized dystonia to short stature or intellectual disability alone, even within the same family. In 78.5% of cases, dystonia involved the lower limbs at onset, with later caudocranial generalization. Eight patients underwent pallidal DBS with a median decrease of Burke-Fahn-Marsden Dystonia Rating Scale-Motor score of 38.5% in the long term. We also report on 4 asymptomatic carriers, suggesting that some KMT2B mutations may be associated with incomplete disease penetrance. KMT2B mutations are frequent in childhood-onset dystonia and cause a complex neurodevelopmental syndrome, often featuring growth retardation and intellectual disability as additional phenotypic features. A dramatic and long-lasting response to DBS is characteristic of DYT-KMT2B dystonia. © 2019 International Parkinson and Movement Disorder Society.

PS1132 INHERITED DNA REPAIR AND CELL CYCLE GENE DEFECTS IN CHRONIC LYMPHOCYTIC LEUKEMIA

Background:Chronic lymphocytic leukemia (CLL) is among the most heritable cancers, with 60% of disease risk genetically determined. However, most of the genetic heritability of CLL remains unexplained. Previously, we identified ATM as the first CLL risk gene.Aims:Here we leverage a deep‐learning‐based germline variant calling algorithm to explore germline mutational enrichment in DNA repair and cell cycle genes in CLL.Methods:A two‐stage case‐control analysis was conducted using gene‐based mutational enrichment analysis of 50 established cancer predisposition DNA repair and cell cycle genes. In the discovery phase, a total of 285 Spanish patients and 5,608 ancestry‐matched controls were evaluated. In the validation stage, an independent cohort of 514 European patients and 27,173 ancestry‐matched controls were analyzed. An FDR correction was applied to both datasets and genes with a q‐value &lt;0.2 in both cohorts were considered significant.Results:Our joint analysis of 799 CLL patients from 2 genetically distinct cohorts and 32,781 ancestry‐matched cancer‐free controls identified ATM and CHEK2 as significantly enriched in both CLL datasets. First, our analysis recaptured the previously reported finding of ATM variant enrichment in CLL patients. Carriers of pathogenic ATM mutations in our cohorts (n = 9 patients, discovery: 1.05%, validation: 1.17%) were 2.8–3.7 times more likely to develop CLL compared to cancer‐free individuals (discovery: OR = 2.8, 95%CI = 0.7–9.0, q‐value = 0.181; validation: OR = 3.7, 95%CI = 1.6–8.3, q‐value = 0.0454). In addition, our analysis identified 21 CLL patients carrying pathogenic CHEK2 alterations (discovery: 1.40%, validation: 3.31%), making CLL patients 4.4–8.0 times more likely to carry such alterations compared to controls (discovery: OR = 8.0, 95%CI = 2.3–27.0, q‐value = 0.026; validation: OR = 4.4, 95%CI = 2.5–7.3, q‐value &lt;0.001).Summary/Conclusion:Our analysis of genetically distinct CLL cohorts, using a high‐sensitivity variant calling algorithm, supports CHEK2 as a potentially novel CLL predisposition gene that may explain a portion of the missing monogenic heritability of CLL. In addition, this study highlights the DNA repair and cell cycle regulation pathways as potential drivers of CLL susceptibility.

PS1133 GENOMIC ARRAYS IDENTIFY HIGH-RISK CHRONIC LYMPHOCYTIC LEUKEMIA WITH GENOMIC COMPLEXITY: A MULTI-CENTER STUDY

Background: Chronic lymphocytic leukemia (CLL) is among the most heritable cancers, with 60% of disease risk genetically determined. However, most of the genetic heritability of CLL remains unexplained. Previously, we identified ATM as the first CLL risk gene. Aims: Here we leverage a deep-learning-based germline variant calling algorithm to explore germline mutational enrichment in DNA repair and cell cycle genes in CLL. Methods: A two-stage case-control analysis was conducted using gene-based mutational enrichment analysis of 50 established cancer predisposition DNA repair and cell cycle genes. In the discovery phase, a total of 285 Spanish patients and 5,608 ancestry-matched controls were evaluated. In the validation stage, an independent cohort of 514 European patients and 27,173 ancestry-matched controls were analyzed. An FDR correction was applied to both datasets and genes with a q-value <0.2 in both cohorts were considered significant. Results: Our joint analysis of 799 CLL patients from 2 genetically distinct cohorts and 32,781 ancestry-matched cancer-free controls identified ATM and CHEK2 as significantly enriched in both CLL datasets. First, our analysis recaptured the previously reported finding of ATM variant enrichment in CLL patients. Carriers of pathogenic ATM mutations in our cohorts (n = 9 patients, discovery: 1.05%, validation: 1.17%) were 2.8–3.7 times more likely to develop CLL compared to cancer-free individuals (discovery: OR = 2.8, 95%CI = 0.7–9.0, q-value = 0.181; validation: OR = 3.7, 95%CI = 1.6–8.3, q-value = 0.0454). In addition, our analysis identified 21 CLL patients carrying pathogenic CHEK2 alterations (discovery: 1.40%, validation: 3.31%), making CLL patients 4.4–8.0 times more likely to carry such alterations compared to controls (discovery: OR = 8.0, 95%CI = 2.3–27.0, q-value = 0.026; validation: OR = 4.4, 95%CI = 2.5–7.3, q-value <0.001). Summary/Conclusion: Our analysis of genetically distinct CLL cohorts, using a high-sensitivity variant calling algorithm, supports CHEK2 as a potentially novel CLL predisposition gene that may explain a portion of the missing monogenic heritability of CLL. In addition, this study highlights the DNA repair and cell cycle regulation pathways as potential drivers of CLL susceptibility.

ATM loss in primary prostate cancer: Analysis of &gt;1000 cases using a validated clinical-grade immunohistochemistry (IHC) assay.

5069 Background: ATM is a protein kinase acting as the main signal transducer of double-strand DNA break repair, in addition to mediating other cellular functions. Germline and somatic pathogenic mutations in ATM occur in a significant fraction of prostate cancers, and targeted therapies for ATM-deficient tumors ( e.g. ATR inhibitors) appear promising. Because DNA sequencing assays frequently cannot distinguish mono-allelic from bi-allelic ATM alterations, a clinical-grade protein IHC assay for ATM loss is needed to select men for these trials and better characterize ATM-deficient tumors. Methods: We validated an automated dichotomously-scored IHC assay to detect ATM protein loss in primary prostate cancer using prostate cancer cell lines with and without bi-allelic ATM inactivation and 49 high-grade (primary Gleason pattern 5) prostate tumors with known ATM genomic status. We then examined the frequency of ATM loss among 23 tumors with pathogenic germline ATM mutations, as well as &gt; 1000 additional primary prostate carcinomas using tissue microarrays (TMA). Results: ATM loss by IHC was found in 14% (7/49) of primary Gleason pattern 5 (5+4 = 9 and 5+5 = 10) tumors. Of these, all cases with adequate tumor content and DNA yield had underlying pathogenic ATM mutations. Of the remaining 42 cases without ATM protein loss, none had ATM alterations. Among men with pathogenic germline ATM mutations, 74% (17/23) had ATM loss by IHC. Of these, 76% (13/17) had homogeneous loss of ATM protein in all tumor cells within a dominant tumor nodule, suggesting that ATM loss was an early clonal event. On TMA analysis, 90% (944/1044) of tumors were evaluable for ATM status by IHC. Among these, ATM loss was seen in 3.3% (31/944), and was significantly more common in tumors with Gleason scores 9-10 (20/198; 10.1%) than in those of all other Gleason grades (11/746; 1.5%) ( P&lt; 0.0001). Conclusions: Validated ATM IHC is a sensitive assay for detecting underlying genomic ATM alterations. ATM protein loss appears to be an early event occurring in the majority of tumors with underlying germline pathogenic ATM mutations, and is significantly enriched in high-grade prostate cancers (especially Gleason grades 9-10).

Breast radiotherapy among ATM-mutation carriers.

1504 Background: Syndromes of DNA repair deficiency may confer both cancer predisposition and increased sensitivity to DNA damaging agents, such as ionizing radiation. Whereas homozygous deficiency of ATM causes the ataxia telangiectasia syndrome, heterozygous ATM mutation carriers exhibit increased rates of breast, pancreas and prostate cancers. ATM repairs DNA double-strand breaks; consequently, mutation carriers may exhibit excessive radiotherapeutic (RT) toxicity. We evaluated the tolerability of adjuvant breast radiation in ATM mutation carriers. Methods: We identified 167 ATM mutation carriers presenting to our institution with breast cancer; of these, 91 received RT and records were reviewed for RT-related morbidity. Toxicities were graded per CTCAE v5. Associations of clinicopathologic features with toxicity were evaluated by multivariate regression. Results: Of 91 ATM mutation carriers receiving breast RT, 31% (n = 28) harbored a pathogenic mutation whereas 69% (n = 63) harbored variants of uncertain significance (VUS). 71% (n = 65) underwent lumpectomy and adjuvant whole-breast RT; 29% (n = 26) had mastectomy and PMRT. Nine patients underwent bilateral RT for a total of 100 RT courses. 86% of RT courses comprised whole-breast/chest-wall tangents; 14% were VMAT or protons. Median tangent dose was 50Gy; 62% included an additional boost (median 10Gy) and 48% used a bolus (median thickness 0.3cm). Lymph nodes were treated in 43% (n = 39). At last on-treatment visit, 31% had grade 2 dermatitis, 4% had other grade 2 events (fatigue, seroma, decreased range of motion, or pain), and 1% had grade 3 dermatitis. At 1 year, 13% had grade ≥2 toxicity: grade 2a lymphedema (n = 3), and grade 2 (n = 1) or 3 (n = 2) capsular contracture. At last follow-up, 4 PMRT patients had capsular contracture (3 with grade 2, 1 with grade 3); 1 patient had grade 2b lymphedema. Overall, no patients had significant (grade ≥2) telangiectasias, fibrosis, or fat necrosis; no grade 4 or 5 toxicities were seen. Neither pathogenic ATM mutations nor VUS were associated with acute or late RT toxicities. Conclusions: We found no evidence of excess breast radiation toxicity among ATM mutation carriers, either pathogenic or VUS. Breast conserving therapy can be safely considered in this population.