Homologous Recombination Deficiency Status Research Articles

Exploratory biomarker analysis in the phase III L-MOCA study of olaparib maintenance therapy in patients with platinum-sensitive relapsed ovarian cancer.

The prospective phase III multi-centre L-MOCA trial (NCT03534453) has demonstrated the encouraging efficacy and manageable safety profile of olaparib maintenance therapy in the Asian (mainly Chinese) patients with platinum-sensitive relapsed ovarian cancer (PSROC). In this study, we report the preplanned exploratory biomarker analysis of the L-MOCA trial, which investigated the effects of homologous recombination deficiency (HRD) and programmed cell death ligand 1 (PD-L1) expression on olaparib efficacy. HRD status was determined using the ACTHRD assay, an enrichment-based targeted next-generation sequencing assay. PD-L1 expression was assessed by SP263 immunohistochemistry assay. PD-L1 expression positivity was defined by the PD-L1 expression on ≥ 1% of immune cells. Kaplan-Meier method was utilised to analyse progression-free survival (PFS). This exploratory biomarker analysis included 225 patients and tested HRD status [N = 190; positive, N = 125 (65.8%)], PD-L1 expression [N = 196; positive, N = 56 (28.6%)], and BRCA1/2 mutation status (N = 219). The HRD-positive patients displayed greater median PFS than the HRD-negative patients [17.9 months (95% CI: 14.5-22.1) versus 9.2 months (95% CI: 7.5-13.8)]. PD-L1 was predominantly expressed on immune cells. Positive PD-L1 expression on immune cells was associated with shortened median PFS in the patients with germline BRCA1/2 mutations [14.5 months (95% CI: 7.4-18.2) versus 22.2 months (95% CI: 18.3-NA)]. Conversely, positive PD-L1 expression on immune cells wasassociated with prolonged median PFS in the patients with wild-type BRCA1/2 [20.9 months (95% CI: 13.9-NA) versus 8.3 months (95% CI: 6.7-13.8)]. HRD remained an effective biomarker for enhanced olaparib efficacy in the Asian patients with PSROC. Positive PD-L1 expression was associated with decreased olaparib efficacy in the patients with germline BRCA1/2 mutations but associated with improved olaparib efficacy in the patients with wild-type BRCA1/2. NCT03534453. Registered at May 23, 2018.

Abstract PO5-16-02: Investigating survivin as a novel target for immunotherapy in diverse breast cancer patients

Abstract Background: The BIRC5 gene encodes survivin, a tumor-specific antigen associated with aggressive tumor features, Black race and poor prognosis in breast cancer (BC). Recent studies have shown that anti-survivin vaccines stimulate a strong anti-tumor immune response in multiple tumor types. However, the relationship between BIRC5/survivin and immune response in BC is poorly understood, particularly among diverse patients. In this analysis, we sought to investigate the relationship between BIRC5/survivin and the BC immune microenvironment across clinical tumor features and RNA-based risk scores for genomic instability. Methods: We leveraged the Carolina Breast Cancer Study (CBCS), a large population-based study that oversampled young (≤50 years) and Black women with invasive BC. NanoString mRNA expression profiling was used to evaluate breast tumor expression for 10 major immune cell types, adaptive and innate immune response, TP53 and homologous recombination deficiency (HRD) pathways, and BIRC5/survivin from 1952 BC patients, including 1,030 (53%) Black and 922 (47%) non-Black women from the CBCS, with additional comparison in The Cancer genome Atlas (TCGA) (n=1,095 breast tumors). BIRC5 was evaluated both as a continuous and categorical variable, using the third expression quartile as a cut point for BIRC5-high, with the lower three quartiles categorized as BIRC5-low. We used generalized linear models to estimate beta values and adjusted p-values as the measure of association between BIRC5 status and immune-related expression in strata defined by estrogen receptor (ER) status, tumor stage, TP53 and HRD status. All analyses were adjusted for multiple comparisons with the Benjamini-Hochberg procedure. Results: In an analysis of continuous BIRC5 expression and global classes of immune response, BIRC5 mRNA expression was highest among tumors with an adaptive-enriched immune class (p< 0.001) in both CBCS and TCGA. However, when investigating immune gene expression according to BIRC5 status (i.e., BIRC5-high relative to BIRC5-low), multivariate differential expression analysis revealed significantly decreased immune cell-related expression scores among BIRC5-high tumors relative to BIRC5-low, and this relationship persisted in analysis stratified on stage (I/II vs III/IV), ER status, TP53 and HRD status. Across all strata, the strongest relationship between BIRC5-high and the immune microenvironment was observed for adaptive immune response (Beta: -0.14; p=0.005), with decreased immune gene expression related to B cells (Beta: -0.16; p=0.01), T cells (Beta: -0.14; p=0.007), Cytotoxic cells (Beta: -0.14; p=0.005), and PD-L1 (Beta: -0.15; p=0.005), relative to BIRC5-low. Conclusion: High BIRC5/survivin expression is associated with decreased immune-related gene expression in both ER-positive and ER-negative disease, and across all tumor stages. Anti-survivin therapies may be beneficial for boosting immune response in breast tumors with low immunogenicity, offering promise for BC subtypes previously ineligible for immunotherapy. Citation Format: Alina Hamilton, Qichen Wang, Sarah Soppe, Melissa Troester, Yara Abdou. Investigating survivin as a novel target for immunotherapy in diverse breast cancer patients [abstract]. In: Proceedings of the 2023 San Antonio Breast Cancer Symposium; 2023 Dec 5-9; San Antonio, TX. Philadelphia (PA): AACR; Cancer Res 2024;84(9 Suppl):Abstract nr PO5-16-02.

Abstract PO4-19-08: Randomized phase II study of talazoparib vs talazoparib plus atezolizumab for patients with premenopausal HR+/HER2- metastatic breast cancer harboring homologous recombination deficiency scar (Young-PALETTA, KCSG BR21-09)

Abstract Background Cyclin D kinase-4/6(CDK4/6) inhibitor combined with endocrine treatment have significantly increased progression free survival(PFS) and overall survival(OS) of patients with hormone receptor(HR)-positive/HER2-negative metastatic breast cancer, and currently is a standard 1st line endocrine treatment option. Homologous recombination deficiency(HRD) including germline(g) BRCA1/2 pathogenic mutation is known to be prevalent among young premenopausal patients. Recently several retrospective data suggested that patients with germline BRCA1/2 pathogenic mutation or HRD have poorer outcome with CDK4/6 inhibitor treatment, however there is no prospective evidence. Poly ADP-ribose polymerase(PARP) inhibitors have shown significant benefit over standard chemotherapy in patients with HER2-neg metastatic breast cancer and gBRCA1/2 pathogenic mutation in two Phase 3 trials. Since preclinical data suggested that PARP inhibitors play a role in anti-tumor immune modulation, immunotherapy combined with PARP inhibitors were explored and showed promising antitumor efficacy in several single arm trials. In this trial, we investigate talazoparib and atezolizumab combination compared with talazoparib monotherapy in premenopausal women with HR+/HER2- metastatic breast cancer with HRD-scar after failure with 1st line palbociclib and endocrine treatment combination. Methods This study is a multicenter, randomized, open-label phase II trial comparing talazoparib versus talazoparib with atezolizumab in premenopausal women with HR+/HER2- metastatic breast cancer harboring HRD scar whose disease have progressed with endocrine and palbociclib combination as the 1st-line endocrine treatment. Prior one line of cytotoxic chemotherapy other than platinum for metastatic setting is allowed, but prior palliative endocrine treatment is not allowed. Eligible patients will be pre-screened with tumor tissue biopsy and peripheral blood sampling before endocrine and palbociclib treatment commencement. HRD status is determined through targeted sequencing(SOLIDaccuTestTM by NGeneBio) of tumor tissue and germline HRD genes including BRCA1/2 variant test. Whole genome sequencing and RNA sequencing are performed for biomarker analyses. Patients harboring HRD scar will receive first line endocrine treatment with palbociclib plus aromatase inhibitor with ovarian suppression. After progression with 1L endocrine treatment, patients will be randomized to either talazoparib alone or talazoparib plus atezolizumab combination. Germline BRCA1/2 pathogenic mutation is a stratification factor. Primary endpoint is a progression free survival after randomization(PFS2). Secondary objectives are a composite of PFS1 and PFS2, PFS1, overall survival, patient reported outcome, and biomarkers for treatment response. A total of 178 subjects are required to detect hazard ratio of 0.62 (median PFS2 of at least 10.5 months with talazoparib and atezolizumab combination vs 6.5 months with talazoparib alone), to achieve 80% power at a two-sided a significance level at 10%, considering drop-out rate of 5%. The study period is expected to be 72 months overall(36 months for accrual and randomization and 24 months of follow-up for primary endpoint PFS2). Citation Format: Hee Kyung Ahn, Jieun Lee, Jee Hung Kim, Kyung-Hun Lee, Jee Hyun Kim, Min Hwan Kim, Minsuk Kwon, Han Jo Kim, Kyung Hae Jung, Yeon Hee Park. Randomized phase II study of talazoparib vs talazoparib plus atezolizumab for patients with premenopausal HR+/HER2- metastatic breast cancer harboring homologous recombination deficiency scar (Young-PALETTA, KCSG BR21-09) [abstract]. In: Proceedings of the 2023 San Antonio Breast Cancer Symposium; 2023 Dec 5-9; San Antonio, TX. Philadelphia (PA): AACR; Cancer Res 2024;84(9 Suppl):Abstract nr PO4-19-08.

Niraparib, Dostarlimab, and Bevacizumab as Combination Therapy in Pretreated, Advanced Platinum-Resistant Ovarian Cancer: Findings From Cohort A of the OPAL Phase II Trial.

To report the results of OPAL (ClinicalTrials.gov identifier: NCT03574779) cohort A, a single-arm substudy of niraparib plus dostarlimab and bevacizumab for the treatment of advanced, platinum-resistant ovarian cancer (PROC). Participants with PROC who received 1-2 previous lines of therapy were treated with niraparib (200 or 300 mg once daily), dostarlimab (500 mg once every 3 weeks for four 21-day cycles, followed by 1,000 mg once every 6 weeks), and bevacizumab (15 mg/kg once every 3 weeks). The primary end point was investigator-assessed objective response rate (ORR) per RECIST v1.1. Safety was also assessed. Exploratory biomarker end points included evaluation of changes in the tumor molecular profile and microenvironment using baseline and on-treatment tumor samples. Of 41 enrolled participants (median age, 66.0 years [range, 37-83 years]), 9.8% had tumors that were BRCA-mutated, 19.5% were homologous recombination (HR)-deficient, and 17.1% were HR repair (HRR)-mutated. As of the cutoff date, all participants discontinued treatment. The ORR was 17.1% (80% CI, 9.8 to 27.0), including one complete response (2.4%); the disease control rate was 73.2% (80% CI, 62.3 to 82.2). Two participants withdrew before first postbaseline scan because of adverse events (AEs). Grade ≥3 treatment-emergent AEs were reported in 92.7% of participants, with the most common being hypertension (26.8%). Response was not correlated with BRCA, HRR, HR deficiency (HRD), or PD-L1 status. Changes suggesting immune activation were observed in on-treatment samples after triplet therapy. Results demonstrated modest activity of niraparib, dostarlimab, and bevacizumab in participants with PROC, many of whom had prognostic factors for poor treatment response. Most participants with response were bevacizumab-naïve. No association was found with HRD, BRCA, or PD-L1 status. AEs were consistent with previous monotherapy reports, except that hypertension was reported more frequently.

Prevalence of Homologous Recombination Deficiency Among Patients With Germline RAD51C/D Breast or Ovarian Cancer

RAD51C and RAD51D are involved in DNA repair by homologous recombination. Germline pathogenic variants (PVs) in these genes are associated with an increased risk of ovarian and breast cancer. Understanding the homologous recombination deficiency (HRD) status of tumors from patients with germline PVs in RAD51C/D could guide therapeutic decision-making and improve survival. To characterize the clinical and tumor characteristics of germline RAD51C/D PV carriers, including the evaluation of HRD status. This retrospective cohort study included 91 index patients plus 90 relatives carrying germline RAD51C/D PV (n = 181) in Spanish hospitals from January 1, 2014, to December 31, 2021. Genomic and functional HRD biomarkers were assessed in untreated breast and ovarian tumor samples (n = 45) from June 2022 to February 2023. Clinical and pathologic characteristics were assessed using descriptive statistics. Genomic HRD by genomic instability scores, functional HRD by RAD51, and gene-specific loss of heterozygosity were analyzed. Associations between HRD status and tumor subtype, age at diagnosis, and gene-specific loss of heterozygosity in RAD51C/D were investigated using logistic regression or the t test. A total of 9507 index patients were reviewed, and 91 patients (1.0%) were found to carry a PV in RAD51C/D; 90 family members with a germline PV in RAD51C/D were also included. A total of 157 of carriers (86.7%) were women and 181 (55.8%) had received a diagnosis of cancer, mainly breast cancer or ovarian cancer. The most prevalent PVs were c.1026+5_1026+7del (11 of 56 [19.6%]) and c.709C>T (9 of 56 [16.1%]) in RAD51C and c.694C>T (20 of 35 [57.1%]) in RAD51D. In untreated breast cancer and ovarian cancer, the prevalence of functional and genomic HRD was 55.2% (16 of 29) and 61.1% (11 of 18) for RAD51C, respectively, and 66.7% (6 of 9) and 90.0% (9 of 10) for RAD51D. The concordance between HRD biomarkers was 91%. Tumors with the same PV displayed contrasting HRD status, and age at diagnosis did not correlate with the occurrence of HRD. All breast cancers retaining the wild-type allele were estrogen receptor positive and lacked HRD. In this cohort study of germline RAD51C/D breast cancer and ovarian cancer, less than 70% of tumors displayed functional HRD, and half of those that did not display HRD were explained by retention of the wild-type allele, which was more frequent among estrogen receptor-positive breast cancers. Understanding which tumors are associated with RAD51C/D and HRD is key to identify patients who can benefit from targeted therapies, such as PARP (poly [adenosine diphosphate-ribose] polymerase) inhibitors.

TP53-specific mutations serve as a potential biomarker for homologous recombination deficiency in breast cancer: a clinical next-generation sequencing study.

TP53 mutations and homologous recombination deficiency (HRD) occur frequently in breast cancer. However, the characteristics of TP53 pathogenic mutations in breast cancer patients with/without HRD are not clear. Clinical next-generation sequencing (NGS) of both tumor and paired blood DNA from 119 breast cancer patients (BRCA-119 cohort) was performed with a 520-gene panel. Mutations, tumor mutation burden (TMB), and genomic HRD scores were assessed from NGS data. NGS data from 47 breast cancer patients in the HRD test cohort were analyzed for further verification. All TP53 pathogenic mutations in patients had somatic origin, which was associated with the protein expression of estrogen receptor and progestogen receptor. Compared to patients without TP53 pathologic mutations, patients with TP53 pathologic mutations had higher levels of HRD scores and different genomic alterations. The frequency of TP53 pathologic mutation was higher in the HRD-high group (HRD score≥42) relative to that in the HRD-low group (HRD score<42). TP53 has different mutational characteristics between the HRD-low and HRD-high groups. TP53-specific mutation subgroups had diverse genomic features and TMB. Notably, TP53 pathogenic mutations predicted the HRD status of breast cancer patients with an area under the curve (AUC) of 0.61. TP53-specific mutations, namely HRD-low mutation, HRD-high mutation, and HRD common mutation, predicted the HRD status of breast cancer patients with AUC values of 0.32, 0.72, and 0.58, respectively. Interestingly, TP53 HRD-high mutation and HRD common mutation combinations showed the highest AUC values (0.80) in predicting HRD status. TP53-specific mutation combinations predict the HRD status of patients, indicating that TP53 pathogenic mutations could serve as a potential biomarker for poly-ADP-ribose polymerase (PARP) inhibitors in breast cancer patients .

Prediction of homologous recombination deficiency from Oncomine Comprehensive Assay Plus correlating with SOPHiA DDM HRD Solution.

Poly(ADP-ribose) polymerase (PARP) inhibitors are used for targeted therapy for ovarian cancer with homologous recombination deficiency (HRD). In this study, we aimed to develop a homologous recombination deficiency prediction model to predict the genomic integrity (GI) index of the SOPHiA DDM HRD Solution from the Oncomine Comprehensive Assay (OCA) Plus. We also tried to a find cut-off value of the genomic instability metric (GIM) of the OCA Plus that correlates with the GI index of the SOPHiA DDM HRD Solution. We included 87 cases with high-grade ovarian serous carcinoma from five tertiary referral hospitals in Republic of Korea. We developed an HRD prediction model to predict the GI index of the SOPHiA DDM HRD Solution. As predictor variables in the model, we used the HRD score, which included percent loss of heterozygosity (%LOH), percent telomeric allelic imbalance (%TAI), percent large-scale state transitions (%LST), and the genomic instability metric (GIM). To build the model, we employed a penalized logistic regression technique. The final model equation is -21.77 + 0.200 × GIM + 0.102 × %LOH + 0.037 × %TAI + 0.261 × %LST. To improve the performance of the prediction model, we added a borderline result category to the GI results. The accuracy of our HRD status prediction model was 0.958 for the test set. The accuracy of HRD status using GIM with a cut-off value of 16 was 0.911. The Oncomine Comprehensive Assay Plus provides a reliable biomarker for homologous recombination deficiency.

Predictive Value and Therapeutic Significance of Somatic BRCA Mutation in Solid Tumors.

Ten percent of patients with breast cancer, and probably somewhat more in patients with ovarian cancer, have inherited germline DNA mutations in the breast and ovarian cancer genes BRCA1 and BRCA2. In the remaining cases, the disease is caused by acquired somatic genetic and epigenetic alterations. Targeted therapeutic agents, such as poly ADP-ribose polymerases (PARP) inhibitors (PARPi), have emerged in treating cancers associated with germline BRCA mutations since 2014. The first PARPi was FDA-approved initially for ovarian cancer patients with germline BRCA mutations. Deleterious variants in the BRCA1/BRCA2 genes and homologous recombination deficiency status have been strong predictors of response to PARPi in a few solid tumors since then. However, the relevance of somatic BRCA mutations is less clear. Somatic BRCA-mutated tumors might also respond to this new class of therapeutics. Although the related literature is often controversial, recently published case reports and/or randomized studies demonstrated the effectiveness of PARPi in treating patients with somatic BRCA mutations. The aim of this review is to summarize the predictive role of somatic BRCA mutations and to provide further assistance for clinicians with the identification of patients who could potentially benefit from PARPi.

Abstract A076: T-cell exhaustion is an independent predictive biomarker of clinical outcome in high grade serous ovarian cancer regardless of homologous recombination deficiency status

Abstract Epithelial ovarian cancer is the most lethal gynecologic malignancy, with 5-year survival rates of 46%. High grade serous carcinoma (HGSC) is the most frequent subtype of epithelial ovarian cancer. Approximately 50% of cases are homologous recombination deficient (HRD). Whereas maintenance treatment by PARP inhibitors has significantly improved the prognosis of patients with HRD tumors, clinical outcome in this malignancy is still poor. Despite the immunogenic nature of HGSC, in which T-cell infiltration correlates to improved clinical outcome following surgery and chemotherapy, the disease exhibits low response rates to immune checkpoint blockade (ICB) monotherapy in advanced line settings. We have previously identified epithelial malignancies including HGSC, CD4 and CD8 populations of tumor-infiltrating and antigen-specific T cells that display features of terminal exhaustion. These populations were key players in the response to anti-PD-1 in patients with epithelial tumors. Here, we set out to characterize tumor-infiltrating T cells in a cohort of 80 HGSC patients treated at our institution from whom we collected tumor samples prior to therapy in which HRD status was assessed. We used multiplex immunofluorescence of FFPE tumor samples to assess infiltration by CD4 and CD8 T cells and multiparametric flow cytometry of frozen viable cell suspensions to assess CD4 and CD8 T-cell exhaustion. The link between biological parameters and clinical outcome was assessed by univariable and multivariable analyses. Assessment of the T-cell infiltrate by immunofluorescence showed no difference in the absolute numbers, per tumor or stroma mm2, of CD8 or CD4 T cells between HRD and homologous recombination proficient (HRP) tumors. Instead, analysis of exhausted CD8 and CD4 T cells showed an increased frequency of terminally exhausted PD-1HighTIM-3+ CD8 T cells in HRD tumors. This population, which expresses the ectonucleotidase CD39, was significantly correlated to the presence of T regulatory cells (Treg), which agreed with previous studies showing comigration of effector and regulatory T cells in ovarian cancer. In addition, terminally exhausted CD4 T cells, expressing the immune checkpoints PD-1 and TIM-3 as well as CD39, were enriched in HRD tumors. Among HRD tumors, no difference was observed between the exhaustion profile in tumors presenting deleterious BRCA1/2 mutations compared to those with other mutations in the homologous recombination pathway. Univariable analyses showed exhaustion parameters to be significant predictors of progression-free survival (PFS). The frequencies of exhausted CD4 and CD8 T cells stood out as predictors of PFS in multivariable analyses. Altogether, these results provide the first evidence of a link between T-cell exhaustion and HRD status and reveal T-cell exhaustion as an independent predictor of disease outcome. They also underline the major role played by the antitumor immune response in HGSC and warrant the development of combination immunotherapies despite the lack of response to ICB monotherapy. Citation Format: Anna Salvioni, Mathilde Del, Marie Michelas, Pierre Vuattoux, Clara-Maria Scarlata, Carlos Martinez-Gomez, Nathalie Van Acker, François-Xavier Frenois, Guillaume Bataillon, Jean-Pierre Delord, Alejandra Martinez, Maha Ayyoub. T-cell exhaustion is an independent predictive biomarker of clinical outcome in high grade serous ovarian cancer regardless of homologous recombination deficiency status [abstract]. In: Proceedings of the AACR Special Conference on Ovarian Cancer; 2023 Oct 5-7; Boston, Massachusetts. Philadelphia (PA): AACR; Cancer Res 2024;84(5 Suppl_2):Abstract nr A076.

Clinical evaluation of a low-coverage whole-genome test for detecting homologous recombination deficiency in ovarian cancer

BackgroundThe PAOLA-1/ENGOT-ov25 trial showed that maintenance olaparib plus bevacizumab increases survival of advanced ovarian cancer patients with homologous recombination deficiency (HRD). However, decentralized solutions to test for HRD in clinical routine are scarce. The goal of this study was to retrospectively validate on tumor samples from the PAOLA-1 trial, the decentralized SeqOne assay, which relies on shallow Whole Genome Sequencing (sWGS) to capture genomic instability and targeted sequencing to determine BRCA status. MethodsThe study comprised 368 patients from the PAOLA-1 trial. The SeqOne assay was compared to the Myriad MyChoice HRD test (Myriad Genetics), and results were analyzed with respect to Progression-Free Survival (PFS). ResultsWe found a 95% concordance between the HRD status of the two tests (95% Confidence Interval (CI); 92%−97%). The Positive Percentage Agreement (PPA) of the sWGS test was 95% (95% CI; 91%−97%) like its Negative Percentage Agreement (NPA) (95% CI; 89%−98%). In patients with HRD-positive tumors treated with olaparib plus bevacizumab, the PFS Hazard Ratio (HR) was 0.38 (95% CI; 0.26–0.54) with SeqOne assay and 0.32 (95% CI; 0.22–0.45) with the Myriad assay. In patients with HRD-negative tumors, HR was 0.99 (95% CI; 0.68–1.42) and 1.05 (95% CI; 0.70–1.57) with SeqOne and Myriad assays. Among patients with BRCA-wildtype tumors, those with HRD-positive tumors, benefited from olaparib plus bevacizumab maintenance, with HR of 0.48 (95% CI: 0.29–0.79) and of 0.38 (95% CI: 0.23 to 0.63) with the SeqOne and Myriad assay. ConclusionThe SeqOne assay offers a clinically validated approach to detect HRD.

272 (PB-088) Poster - Association between prognosis, carboplatin treatment response and homologous recombination deficiency status in early triple-negative breast cancer

272 (PB-088) Poster - Association between prognosis, carboplatin treatment response and homologous recombination deficiency status in early triple-negative breast cancer

Homologous Recombination Deficiency Unrelated to Platinum and PARP Inhibitor Response in Cell Line Libraries

While large publicly available cancer cell line databases are invaluable for preclinical drug discovery and biomarker development, the association between homologous recombination deficiency (HRD) and drug sensitivity in these resources remains unclear. In this study, we comprehensively analyzed molecular profiles and drug screening data from the Cancer Cell Line Encyclopedia. Unexpectedly, gene alterations in BRCA1/2 or homologous recombination-related genes, HRD scores, or mutational signature 3 were not positively correlated with sensitivity to platinum agents or PARP inhibitors. Rather, higher HRD scores and mutational signature 3 were significantly associated with resistance to these agents in multiple assays. These findings were consistent when analyzing exclusively breast and ovarian cancer cell lines and when using data from the COSMIC Cell Line Project. Collectively, the existing data from established cancer cell lines do not reflect the expected association between HRD status and drug response to platinum agents and PARP inhibitors in clinical tumors. This discrepancy may extend to other tumor characteristics, highlighting the importance of recognizing potential limitations in cell line data for researchers.

Evaluation of Homologous Recombination Deficiency in Ovarian Cancer.

Homologous recombination deficiency (HRD) is an important biomarker guiding selection of ovarian cancer patients who will derive the most benefit from poly(ADP-ribose) polymerase inhibitors (PARPi). HRD prevents cells from repairing double-stranded DNA damage with high fidelity, PARPis limit single-stranded repair, and together these deficits induce synthetic lethality. Germline or somatic BRCA mutations represent the narrowest definition of HRD, but do not reflect all patients who will have a durable PARPi response. HRD can also be defined by its downstream consequences, which are measured by different metrics depending on the test used. Ideally, all patients will undergo genetic counseling and germline testing shortly after diagnosis and have somatic testing sent once an adequate tumor sample is available. Should barriers to one test be higher, pursuing germline testing with reflex to somatic testing for BRCA wildtype patients or somatic testing first strategies are both evidence-based. Ultimately both tests offer complementary information, germline testing should be pursued for any patient with a history of ovarian cancer, and somatic testing is valuable at recurrence if not performed in the upfront setting. There is a paucity of data to suggest superiority of one germline or somatic assay; therefore, selection should optimize turnaround time, cost to patients, preferred result format, and logistical burden. Each clinic should implement a standard testing strategy for all ovarian cancer patients that ensures HRD status is known at the time of upfront chemotherapy completion to facilitate comprehensive counseling about anticipated maintenance PARPi benefit.

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&lt;0.03). Additionally, there was increased frequency in TP53 MT with HRD WT 54.8% versus HRD MT 28.1% (p&lt;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&lt;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]

Abstract IA008: Targeting homologous recombination deficiency (HRD) using optimized ionizing radiation and drug combinations

Abstract Homologous recombination deficiency (HRD) is found not only in germline mutation carriers of HR genes but is also observed in sporadic cancers when both copies of the HR gene are inactivated. Recognition of HRD is facilitated by using genomic landscape alterations in addition to target gene sequencing. In addition, HR function can be established by measuring RAD51 foci in tumor specimens, particularly if they are irradiated ex-vivo to reveal the HR pathway function more clearly. Triple negative breast cancer (TNBC) is highly enriched for HRD constituting ~45% of primary tumors and over 50% of locally recurrent or metastatic cancers. In a clinical trial of recurrent TNBC, we used the combination of ionizing radiation (IR) with low-dose weekly cisplatin (cis) to study the clinical response according to HRD status. The combination of IR + cis has been shown in HRD cells to be more sensitizing than either agent alone. In the clinical study of 49 TNBC patients, there were imaging responses in 22/29 (75%) HRD patients, but only in 4/20 (20%) HR-proficient patients. We have followed this study by investigating the combination of the PARP-inhibitor with radiotherapy in an ongoing clinical trial as well as with detailed cell-based studies. HRD cells have elevated PARP activity at baseline, as observed by the extent of PARylated proteins, plus the lower baseline levels of NAD. When PARP-inhibitors are combined with radiotherapy, there is demonstrable supra-additive sensitization at 6Gy of IR and up to 100nM talazoparib (TZ). However, when the dose of TZ is above 1 µM, there is paradoxical recovery from the combination therapy, due to rescue of NAD depletion in HRD cells. The threshold for seeing this paradoxical recovery effect in HR-proficient cells was not reached. One of the mechanisms of cell killing in HRD cells is enhanced parthanatos, which higher doses of TZ will prevent, particularly in conjunction with IR. The implication of this work is that the optimal combination of radiotherapy with PARP-inhibitor may not be the maximum tolerated dose, but rather a tuned level of the combination to maximize radiosensitization without exceeding the threshold for PARP-inhibitor mediated protection of cell death. It is already observed that the maximum tolerated dose of PARP-inhibitor in the setting of curative doses of IR have to be modified by 3-10-fold. Further optimization of the combination dosing to achieve the maximal therapeutic ratio will be discussed including the development of biomarkers to achieve this goal. Additional radiation combinations with synthetic lethal targets of HRD will be discussed. Citation Format: Simon N. Powell, Niamh McDermott, Robert Delsite. Targeting homologous recombination deficiency (HRD) using optimized ionizing radiation and drug combinations [abstract]. In: Proceedings of the AACR Special Conference in Cancer Research: DNA Damage Repair: From Basic Science to Future Clinical Application; 2024 Jan 9-11; Washington, DC. Philadelphia (PA): AACR; Cancer Res 2024;84(1 Suppl):Abstract nr IA008.

Biomarker Testing Journey Among Patients with Advanced Solid Tumors and Treatment Patterns by Homologous Recombination Repair Status: A Clinico-Genomic Database Study.

Defects in the homologous recombination repair (HRR) pathway can include mutations in BRCA1 and BRCA2 (BRCAm) and other HRR genes (HRRm). These mutations are associated with a homologous recombination deficiency (HRD) phenotype. We evaluated testing journey and treatment patterns by BRCAm, HRRm, and HRD status in a real-world dataset. Deidentified data for patients who had undergone comprehensive genomic profiling using FoundationOne®CDx were collected through December 31, 2020, from a real-world multi-tumor clinico-genomic database (CGDB) capturing data from clinics in the United States. Patients eligible for inclusion in this analysis had a confirmed diagnosis with advanced or metastatic disease between January 1, 2018, and December 31, 2019, for 1 of 15 solid tumor types. Objectives were to evaluate patient treatment patterns by BRCAm, HRRm, and HRD status and to describe the timing of when (throughout disease course) comprehensive genomic profiling was performed. Among 9457 patients included in the overall population with evaluable biomarker status, 7856 (83.1%) received≥1 systemic therapy. Among the 7856 patients who received systemic therapy, 2324 (30.0%) underwent testing before first-line therapy, 4114 (52.4%) were tested after receiving first-line therapy and before receiving subsequent therapy (if any), 970 (12.3%) were tested after second-line therapy and before receiving subsequent therapy (if any), and 447 (5.7%) patients underwent testing after receiving third-line therapy. A higher proportion of patients with BRCAm, HRRm, or HRD-positive status were treated with poly(ADP-ribose) polymerase (PARP) inhibitors across all lines of therapy. There was no evidence of a meaningful difference in the proportion of patients who received other treatment (including chemotherapy and immunotherapy) by BRCAm, HRRm, or HRD status. The majority of patients from this real-world dataset underwent FoundationOne®CDx testing after initiation of first-line treatment. Testing appeared to influence treatment patterns, with a higher proportion of patients with BRCAm, HRRm, and HRD-positive disease receiving PARP inhibitors.

Assessing the Phenotype of a Homologous Recombination Deficiency Using High Resolution Array-Based Comparative Genome Hybridization in Ovarian Cancer.

Ovarian cancer (OC) cells with homologous recombination deficiency (HRD) accumulate genomic scars (LST, TAI, and LOH) over a value of 42 in sum. PARP inhibitors can treat OC with HRD. The detection of HRD can be done directly by imaging these genomic scars, or indirectly by detecting mutations in the genes involved in HR. We show that HRD detection is also possible using high-resolution aCGH. A total of 30 OCs were analyzed retrospectively with high-resolution arrays as a test set and 19 OCs prospectively as a validation set. Mutation analysis was performed by HBOC TruRisk V2 panel to detect HR-relevant mutations. CNVs were clustered with respect to the involved HR genes versus the OC cases. In prospective validation, the HRD status determined by aCGH was compared with external HRD assessments. Two BRCA mutation carriers did not have HRD. OC could approximately differentiate into two groups with characteristic CNV patterns with different survival rates. Mutation frequencies have a linear regression on the HRD score. Mutations in individual HR-relevant genes do not always indicate HRD. This may depend on the mutation frequency in tumor cells. The aCGH shows the genomic scars of an HRD inexpensively and directly.

Homologous Recombination Deficiency (HRD) Scoring, by Means of Two Different Shallow Whole-Genome Sequencing Pipelines (sWGS), in Ovarian Cancer Patients: A Comparison with Myriad MyChoice Assay

High-grade serous ovarian cancer (HGSOC) patients carrying the BRCA1/2 mutation or deficient in the homologous recombination repair system (HRD) generally benefit from treatment with PARP inhibitors. Some international recommendations suggest that BRCA1/2 genetic testing should be offered for all newly diagnosed epithelial ovarian cancer, along with HRD assessment. Academic tests (ATs) are continuously under development, in order to break down the barriers patients encounter in accessing HRD testing. Two different methods for shallow whole-genome sequencing (sWGS) were compared to the reference assay, Myriad. All these three assays were performed on 20 retrospective HGSOC samples. Moreover, HRD results were correlated with the progression-free survival rate (PFS). Both sWGS chemistries showed good correlation with each other and a complete agreement, even when compared to the Myriad score. Our academic HRD assay categorized patients as HRD-Deficient, HRM-Mild and HRN-Negative. These three groups were matched with PFS, providing interesting findings in terms of HRD scoring and months of survival. Both our sWGS assays and the Myriad test correlated with the patient’s response to treatments. Finally, our AT confirms its capability of determining HRD status, with the advantage of being faster, cheaper, and easier to carry out. Our results showed a prognostic value for the HRD score.

Abstract B040: NCI 10129: A Phase 2 Study of the PARP inhibitor Olaparib (AZD2281) in IDH1 and IDH2 mutant advanced solid tumors

Abstract Introduction: Cholangiocarcinoma (CCA) is a highly aggressive malignancy with mutations in Isocitrate dehydrogenase (IDH) identified in approximately 15% of patients. Neomorphic mutations in IDH 1/2 result in accumulation of the oncometabolite 2-hydroxyglutarate (2G), which has been implicated in tumor progression via inhibitory effects on alpha ketoglutarate. Moreover, the mutant IDH dependent accumulation of 2HG results in homologous recombination deficiency (HRD) and renders these cells exquisitely sensitive to Poly (ADP-Ribose) polymerase (PARP) inhibitors. NCI 10129 evaluated the efficacy of olaparib for IDH mutated solid tumors, and here we present the CCA cohort. Methods: NCI 10129 is a 3 arm, open label phase 2 clinical trial performed in the NCI National Clinical Trials Network evaluating olaparib 300 mg twice daily for IDH mutated solid tumors refractory to standard treatment. Patients with CCA were enrolled in 2 cohorts 1) IDHi pre-treated CCA and 2) IDHi untreated CCA. The primary endpoint was overall response rate (ORR) and secondary endpoints includes progression-free survival (PFS), and overall survival (OS). An exploratory analysis measured the impact of circulating 2HG levels, circulating tumor DNA, and tumor genomics on patient outcomes. Results: Thirty patients with CCA enrolled on NCI10129 (15 each IDHi pre-treated and IDHi treatment naïve), 18 patients received ≥ 2 prior systemic treatments, and 23 patients had IDH1 mutations and 7 patients IDH2 mutations. There were no objective responses seen in the first 15 CCA patients in either cohort and enrollment was closed. The median PFS was 2.4 months 95% CI (1.9 - 6.5) and median OS 12.9 months 95% CI (6.3 – not reached). The IDHi pre-treated cohort had improved median OS compared to IDHi treatment naïve (15.3 vs 7.3, P = 0.04). There were 8/30 (27%) patients with clinical benefit (CB) as defined by PFS &amp;gt; 6 months. Patients with CB had lower baseline mean D-2HG levels compared to those without CB (1.4 umol/L vs 5.9 umol/L, p = 0.02). Whole exome sequencing revealed the study population to be enriched for HRD by sum scores and HRD mutational signatures without alternative HRD mutations other than IDH1/2. No improvement in survival was noted based on HRD status. RNA sequencing Gene Set Enrichment Analysis revealed that cell cycle and interferon alpha pathways were activated in patients with CB (p &amp;lt; 0.01). Olaparib was tolerable with the majority of adverse events grade 1-2. Conclusion: Olaparib did not result in objective responses in IDH mutated CCA. However, a subgroup of patients did demonstrate prolonged PFS comparable to historical controls for refractory CCA as well that seen with targeted agents such as IDHi. Exploratory analysis revealed this subgroup to be enriched for lower baseline 2HG levels. Serial ctDNA analysis is ongoing and will be presented. Given the CB subgroup and OS data, additional clinical trials leveraging the HRD properties of IDH mutations is warranted. Future studies may benefit from DNA damage combination therapies thereby enhancing potency. Citation Format: Michael Cecchini, Mary Jo Pilat, Nataliya Uboha, Nilofer S Azad, May Cho, Elizabeth J Davis, Jordi Rodon Ahnert, Gabriel Tinoco, Geoff I Shapiro, Simon Khagi, Benjamin Powers, Roman Groisberg, Jan Drappatz, Li Chen, Biswajit Das, Xun Bao, Jing Li, Abhijit Patel, Monica Niger, Deborah Doroshow, Diane Durecki, Scott Boerner, Ranjit Bindra, Percy Ivy, Yu Shyr, Patricia LoRusso. NCI 10129: A Phase 2 Study of the PARP inhibitor Olaparib (AZD2281) in IDH1 and IDH2 mutant advanced solid tumors [abstract]. In: Proceedings of the AACR-NCI-EORTC Virtual International Conference on Molecular Targets and Cancer Therapeutics; 2023 Oct 11-15; Boston, MA. Philadelphia (PA): AACR; Mol Cancer Ther 2023;22(12 Suppl):Abstract nr B040.

Concordance between Three Homologous Recombination Deficiency (HRD) Assays in Patients with High-Grade Epithelial Ovarian Cancer

Our aim was to evaluate the concordance between the Myriad MyChoice and two alternative homologous recombination deficiency (HRD) assays (AmoyDx HRD Focus NGS Panel and OncoScan™) in patients with epithelial ovarian cancer (EOC). Tissue samples from 50 patients with newly diagnosed EOC and known Myriad MyChoice HRD status were included. DNA aliquots from tumor samples, previously evaluated with Myriad MyChoice and centrally reassessed, were distributed to laboratories to assess their HRD status using the two platforms, after being blinded for the Myriad MyChoice CDx HRD status. The primary endpoint was the concordance between Myriad MyChoice and each alternative assay. Tumor samples were evaluated with an AmoyDx® HRD Focus Panel (n = 50) and with OncoScan™ (n = 43). Both platforms provided results for all tumors. Analysis showed that correlation was high for the Myriad MyChoice GI score and AmoyDx® HRD Focus Panel (r = 0.79) or OncoScan™ (r = 0.87) (continuous variable). The overall percent agreement (OPA) between Myriad MyChoice GI status (categorical variable) and each alternative assay was 83.3% (68.6–93.3%) with AmoyDx and 77.5% (61.5–89.2%) with OncoScan™. The OPA in HRD status between Myriad MyChoice and AmoyDx was 88.6% (75.4–96.2). False-positive rates were 31.6% (6/19) for AmoyDx GI status and 31.9% (7/22) for OncoScan™, while false-negative rates were 0% (0/28, AmoyDx) and 11.1% (2/18, OncoScan™) compared with the Myriad MyChoice GI status. While substantial concordance between Myriad MyChoice and alternative assays was demonstrated, prospective validation of the analytical performance and clinical relevance of these assays is warranted.