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Purpose: The elevated levels of somatic copy-number alterations (SCNAs) in a subset of high-risk endometrial cancers are suggestive of defects in pathways governing genome integrity. We sought to assess the prevalence of homologous recombination deficiency (HRD) in endometrial cancers and its association with histopathologic and molecular characteristics.Experimental Design: Fresh tumor tissue was prospectively collected from 36 endometrial cancers, and functional HRD was examined by the ability of replicating tumor cells to accumulate RAD51 protein at DNA double-strand breaks (RAD51 foci) induced by ionizing radiation. Genomic alterations were determined by next-generation sequencing and array comparative genomic hybridization/SNP array. The prevalence of BRCA-associated genomic scars, a surrogate marker for HRD, was determined in the The Cancer Genome Atlas (TCGA) endometrial cancer cohort.Results: Most endometrial cancers included in the final analysis (n ¼ 25) were of non-endometrioid (52%), grade 3 (60%) histology, and FIGO stage I (72%). HRD was observed in 24% (n ¼ 6) of cases and was restricted to non-endometrioid endometrial cancers (NEEC), with 46% of NEECs being HRD compared with none of the endometrioid endometrial cancers (EEC, P ¼ 0.014). All but 1 of the HRD cases harbored either a pathogenic BRCA1 variant or high somatic copy-number (SCN) losses of HR genes. Analysis of TCGA cases supported these results, with BRCA-associated genomic scars present in up to 48% (63/132) of NEEC versus 12% (37/312) of EEC (P < 0.001).Conclusions: HRD occurs in endometrial cancers and is largely restricted to non-endometrioid, TP53-mutant endometrial cancers. Evaluation of HRD may help select patients that could benefit from treatments targeting this defect, including platinum compounds and PARP inhibitors. Analysis and interpretation of data (e.g., statistical analysis, biostatistics, computational analysis):

Recent studies have shown that the efficacy of PARP inhibitors in epithelial ovarian carcinoma (EOC) is related to tumor-specific defects in homologous recombination (HR) and extends beyond BRCA1/2 deficient EOC. A robust method with which to identify HR-deficient (HRD) carcinomas is therefore of utmost clinical importance. In this study, we investigated the proficiency of a functional HR assay based on the detection of RAD51 foci, the REcombination CAPacity (RECAP) test, in identifying HRD tumors in a cohort of prospectively collected epithelial ovarian carcinomas (EOCs). Of the 39 high-grade serous ovarian carcinomas (HGSOC), the RECAP test detected 26% (10/39) to be HRD, whereas ovarian carcinomas of other histologic subtypes (n = 10) were all HR-proficient (HRP). Of the HRD tumors that could be sequenced, 8/9 showed pathogenic BRCA1/2 variants or BRCA1 promoter hypermethylation, indicating that the RECAP test reliably identifies HRD, including but not limited to tumors related to BRCA1/2 deficiency. Furthermore, we found a trend towards better overall survival (OS) of HGSOC patients with RECAP-identified HRD tumors compared to patients with HRP tumors. This study shows that the RECAP test is an attractive alternative to DNA-based HRD tests, and further development of a clinical grade RECAP test is clearly warranted.

PARP inhibitor (PARPi) sensitivity is related to tumor-specific defects in homologous recombination (HR). Therefore, there is great clinical interest in tests that can rapidly and reliably identify HR deficiency (HRD). Functional HRD tests determine the actual HR status by using the (dis)ability to accumulate RAD51 protein at sites of DNA damage as read-out. In this study, we further improved and calibrated a previously described RAD51-based functional HRD test on 74 diagnostic formalin-fixed paraffin-embedded (FFPE) specimens (RAD51-FFPE test) from endometrial cancer (EC n = 25) and epithelial ovarian cancer (OC n = 49) patients. We established optimal parameters with regard to RAD51 foci cut-off (≥ 2) and HRD threshold (15%) using matched endometrial and ovarian carcinoma specimens for which HR status had been established using a RAD51-based test that required ex vivo irradiation of fresh tissue (RECAP test). The RAD51-FFPE test detected BRCA deficient tumors with 90% sensitivity and RECAP-HRD tumors with 87% sensitivity, indicating that it is an attractive alternative to DNA-based tests with the potential to be applied in routine diagnostic pathology.

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<p>Supplementary Material and Methods</p>

<p>Figure S2 shows that their is an unbiased natural devision between samples analyes by the oncoscan platform and the CGH Agilent platform.</p>

<p>Table S2 shows the clinicopathological characteristics of the homologous recombination intermediate cases.</p>