A novel CDK12 inhibitor induces homologous recombination deficiency to enhance PARP inhibitor efficacy in uterine serous carcinoma.

Comparing mutation frequencies for homologous recombination genes in uterine serous and high-grade serous ovarian carcinomas: A case for homologous recombination deficiency testing in uterine serous carcinoma

Pancreatic ductal adenocarcinoma (PDAC) is a devastating malignancy that affects 44,000 individuals annually in the US, with almost 90% lethality even when diagnosed prior to metastasis. There is an urgent unmet medical need both for new therapies as well as better matching of existing therapies to patients. To address this emergency, we are assessing the feasibility of implementing a strategy of using HRD (Homologous Recombination Deficiency) scores for better therapy matching in PDAC patients. Using a panel of 77 patient-derived xenograft (PDX) models that were developed from fresh surgical PDAC tumor samples, HRD scores were generated based on analysis of three biomarkers (LOH, TAI and LST) and mutational data for 45 genes. All 77 samples met inclusion criteria, 75 FFPE specimens generated mutation data. HRD analysis was successful for 71 specimens (range= 1 - 63 (median=22)), with the primary cause of failure identified as high non-tumor content. 53 PDX models had mutations in KRAS gene and 45 in TP53. We have also identified 4 PDX models with mutations in BRCA2, 3 models with mutations in ATM, 4 models with mutations in RAD51. We have also found frequent mutations in several other DNA repair genes (ATR, PALB2, MLH1, MSH2, MSH3, MSH6, FANCM), but most of these models retained one functional allele and were not associated with a high HRD score. Using this genomic analysis, all 71 PDX models were stratified into three clusters with high, medium and low HRD scores. Three PDX models with the highest and lowest HRD scores each were selected for an in vivo study with DNA-damaging platinum-based chemotherapeutic agents Cisplatin and Carboplatin, as well as with a clinically relevant PARP inhibitor Niraparib. The results of the PDX study will be reported and compared with responses to chemotherapy using RECIST V1.1 in patients. Note: This abstract was not presented at the meeting. Citation Format: Vladimir Khazak, Natalia Skobeleva, Anastasiia Vetkina, Ilya Serebriiskii, Kirsten M. Timms, Angela Davies, Igor Astsaturov. Assessment of HRD score as predictor of chemosensitivity of PDAC PDX xenograft models to DNA-damaging chemotherapy [abstract]. In: Proceedings of the American Association for Cancer Research Annual Meeting 2017; 2017 Apr 1-5; Washington, DC. Philadelphia (PA): AACR; Cancer Res 2017;77(13 Suppl):Abstract nr 1124. doi:10.1158/1538-7445.AM2017-1124

e17004 Background: PARP inhibitors (PARPi) have been approved in the treatment of metastatic castration-resistant prostate cancer with deleterious or suspected deleterious alterations in homologous recombination repair (HRR) genes. Patients carrying alterations of different HRR genes showed diverse response to PARPi, while the underlying mechanism is unrevealed. Homologous recombination deficiency (HRD) score serves as a promising way to interpret the genomic instability by targeted next-generation sequencing (NGS). In this study, we analyzed HRD score and its relationship to genomic characteristics in a prostate cancer (PC) cohort. Methods: Tumor tissue and matched white blood cells from 295 PC patients were analyzed by NGS-based assays, which target over 10,000 single-nucleotide polymorphisms distributed across human genome to characterize genomic instability, as well as the whole coding region of 733 or 233 genes. HRD score was derived by the sum of three indexes, including loss-of-heterozygosity, telomeric allelic imbalance, and large-scale state transitions. Deleterious or suspected deleterious mutations were included for analysis. Tumor mutation burden (TMB) was calculated by the number of somatic single nucleotide variants and indels in examined coding regions, with driver mutations excluded. An ovarian and fallopian tube cancer cohort, carrying deleterious or suspected deleterious mutations in BRCA1/2, was also included. Results: In PC cohort, 25 patients (8.47%) had BRCA1/2 mutations, with additional 39 patients (13.2%) carrying mutations in HRR genes other than BRCA1/2. HRD score was similar among tumors with germline BRCA1/2 mutation (n = 9), somatic BRCA1/2 mutation (n = 14) or both (n = 2) (median HRD score 38.0, 32.5 and 49.5; P = 0.281). In BRCA1/2 mutation carriers, HRD score in PC cohort was lower than ovarian cancer (n = 222) and fallopian tube cancer (n = 8) (median HRD score 36, 54 and 67; P < 0.001). PC samples with CDK12 (n = 35), ATM (n = 13) and PALB2 (n = 2) mutations had lower HRD score compared with BRCA1/2 mutants (median HRD score 24, 14, 11 and 36; P < 0.001). In PC patients without HRR mutation, HRD score was higher in TP53 mutants compared with TP53 wildtype (median HRD score 25 and 14; P < 0.001), while tumors with ERG fusion had lower HRD score than ERG fusion absent tumors (median HRD score 8 and 18.5; P = 0.029). In four TP53 mutated PC patients treated with PARPi in our center, three patients achieved PSA response. Furthermore, no linear correlation was discovered between TMB and HRD score, while tumors with higher HRD score (HRD score ≥ 30) had a bit lower TMB (median TMB 3.07 vs 3.91, P = 0.002). Conclusions: HRD score was affected by alterations in different HRR genes, TP53 mutation and ERG fusion, which should be taken into consideration of the future clinical trials and research on intrinsic mechanisms. Patients carrying alterations other than HRR genes might benefit from PARPi. Further studies are needed.

[Purpose]Homologous Recombination Deficiency (HRD) score has been developed to evaluate the DNA double-strand break repair function. BRCA1/2 germline mutation-associated breast cancers are known to show a high HRD score and a high sensitivity to platinum-based chemotherapy as well as PARP inhibitor. HRD can be theoretically induced by not only the dysfunction of BRA1/2 but also the dysfunction of the other molecules involved in homologous recombination, and, actually, high HRD score is reported to be seen in a significant proportion of sporadic breast tumors without BRCA1/2 mutation. The aim of the present study was to elucidate the clinicopathological characteristics of sporadic breast tumors with high HRD score as well as their sensitivity to sequential paclitaxel and FEC chemotherapy (P-FEC). [Methods]Tumor tissues obtained by Mammotome from 132 sporadic breast cancer patients (stage II-III) before neoadjuvant chemotherapy with P-FEC were subjected to assay for HRD score using Oncoscan CNV kit®. HRD score was a simple sum of NtAI, LOH, and LST (cutoff, 42), and were also subjected to the gene expression analysis using the Affymetrix microarray (U133 plus 2.0). BRCA1 promoter methylation was assayed by methylation specific real-time PCR. [Result]Of the 132 breast tumors, 32% showed a high HRD score which were significantly associated with high histological grade (P=0.001), negative progesterone receptor (P=0.023) and high Ki67 index (P=0.004). Triple negative breast cancer (TNBC)(n=19) showed a significantly (P<0.001) higher HRD score than the other subtypes (HR+/HER2-(n=73), HR+/HER2+(n=12), HR-/HER2+(n=18)). BRCA1 promoter methylation was significantly associated with a high HRD score. There was no significant correlation between HRD score and pCR to P-FEC when all tumors were considered but tumors with a high HRD score showed a significantly (P=0.020) lower pCR rate when only TNBC were considered. In TNBC, majority of tumors showed a high HRD score in five subtypes (BL1, BL2, IM, M, MSL) but no tumor showed a high HRD score in LAR (luminal androgen receptor) subtype. [Conclusion]Approximately one third of sporadic breast tumors show a high HRD score, indicating the presence of homologous recombination dysfunction, and they are most often seen in TNBC with BRCA1 promoter methylation and never in LAR subtype. Interestingly, breast tumors with a high HRD score were less sensitive to P-FEC. Citation Format: Imanishi S, Naoi Y, Shimazu K, Shimoda M, Kagara N, Tanei T, Miyake T, Kim SJ, Noguchi S. HRD (homologous recombination deficiency) score and its clinicopathological features in neoadjuvant chemotherapy treated sporadic breast cancers [abstract]. In: Proceedings of the 2018 San Antonio Breast Cancer Symposium; 2018 Dec 4-8; San Antonio, TX. Philadelphia (PA): AACR; Cancer Res 2019;79(4 Suppl):Abstract nr P3-11-06.

BACKGROUND AND OBJECTIVEUterine papillary serous cancer (UPSC) represents only 10% of all uterine cancers and is associated with a significantly worse prognosis compared with other histological types of endometrial cancers. It closely resembles the behavior of ovarian carcinoma.DESIGN AND SETTINGRetrospective study in a referral center covering period from February 1989 to January 2009.PATIENTS AND METHODSEighteen patients who underwent definitive surgery followed by adjuvant therapy—platinum-based chemotherapy, radiotherapy, or both—were reviewed. Median age was 62 years (range, 52–76 years). All patients underwent total abdominal hysterectomy and salpingo-oophorectomy. Positive lymph nodes were found in 4 of 7 patients who underwent lymph node sampling/dissection. Seven patients had stage I/II disease, whereas 11 patients had stage III disease. Six patients received chemotherapy, 5 patients received radiation therapy, while 7 patients received both chemotherapy and radiation therapy.RESULTMedian follow-up was 27 months. The median survival and relapse-free survival were 33 and 23 months, respectively. Eight patients were alive and free of disease, of whom 5 patients were stage I/II and 4 patients were stage III. Distant metastasis was the most common site of relapse. Early stage (I/II) was associated with significant improvement in relapse-free survival (RFS) and overall survival (OS) (P=.004 and P=.05, respectively). The combined-modality treatment including chemotherapy-radiotherapy showed statistically significant improvement in RFS (P=.012), while the improvement in OS did not reach statistical significance (P=.12).CONCLUSIONThis study indicates that postoperative combined treatment with chemotherapy and radiation therapy plays a role in the management of UPSC by improving RFS. Distant metastasis remains the major site of relapse. Future studies using combined-modality therapy are needed to improve the outcome in patients with UPSC.

e17634 Background: Rare gynecologic cancers, such as ovarian carcinosarcoma (OCS), uterine serous carcinoma (USC) and ovarian clear cell adenocarcinoma (OCCA), have limited treatment options and molecular alterations that drive cell proliferation and drug resistance, resulting in poor overall survival. The anti-microtubule agent (AMA) paclitaxel is used in first-line treatment of most gynecologic cancers; however, the AMA eribulin is known to have additional anticancer properties (Goto et al. Anticancer Res 38:2929, 2018; Ho et al. Cancer Res 82:4457, 2022). Eribulin ADCs have been developed for targeted delivery to limit toxicity and maximize response. FZEC is an anti-folate receptor α (FRA)-eribulin ADC (Shimizu et al. Clin Cancer Res 27:3905, 2021) and MORAb-109 targets eribulin to mesothelin (MSLN; Albone et al. Annals Oncol 31:S491, 2020). Here we provide preclinical data to support clinical trials of FZEC and MORAb-109 in rare gynecologic cancers. Methods: Fresh tumor tissue from patients consented to the WEHI Stafford Fox Rare Cancer Program was used to establish a cohort of patient-derived xenograft (PDX) models of high-grade serous ovarian carcinoma (HGSOC), OCS, USC and OCCA. PDX models were screened for FRA and MSLN expression by IHC and quantified by HALO software, and 15 models were selected for in vivo testing. Tumor-bearing mice were treated with eribulin (1 mg/kg TIW 3 wks), FZEC (12.5 mg/kg Q14Dx3) or MORAb-109 (25 mg/kg Q14Dx3) and compared to standard of care treatments. Results: About 80% of HGSOC PDX models were FRA positive with no difference between models from chemonaïve samples vs multiple lines of prior treatment. FRA expression was frequent in USC and OCCA models, with MSLN seen in a smaller subset. Despite most (80%) models being cisplatin refractory, 11/15 responded to eribulin with the 4 non-responders having high MDR1 expression and being from previously treated patients. Overall, responses to FZEC and MORAb-109 appeared to correlate with FRA and MSLN expression; importantly, some models with moderate to high target antigen expression showed deeper and more durable responses to FZEC and MORAb-109 compared to eribulin. Conclusions: Responses to FZEC and MORAb-109 correlated with FRA and MSLN expression, with equivalent or better activity compared to eribulin in target-positive PDX models of HGSOC, USC and OCCA. [Table: see text]

Serous carcinoma is the prototype of type 2 uterine carcinoma. In many cases, establishing a diagnosis is straightforward, but problems can arise in that papillary variants of endometrioid carcinoma may be mistaken for serous carcinoma, and glandular variants of serous carcinoma may be misdiagnosed as endometrioid carcinoma. Markers such as p53, oestrogen receptor and p16 may be of use in problematic cases, but there is overlap and these may not therefore be of value in an individual case. It has been shown recently that high-mobility group AT-hook 2 (HMGA2) is expressed by most ovarian serous carcinomas, and our aim was to ascertain whether it is also expressed in uterine serous carcinoma and of value in its distinction from endometrioid carcinoma. Whole tissue sections of uterine serous (n = 33) and endometrioid (n = 38) carcinoma were immunostained using HMGA2 antibody. As many of the diagnostic problems relate to the distinction between serous carcinoma and grade 3 endometrioid carcinoma, tissue microarrays (TMAs) containing uterine serous (n = 71) and uterine grade 3 endometrioid (n = 68) carcinomas were also stained. Staining was classified as negative (totally negative or occasional nuclei positive), 1+ (<10% of nuclei positive), 2+ (10-49% of nuclei positive), 3+ (50-74% of nuclei positive), or 4+ (≥75% of nuclei positive). On the whole tissue sections, positive staining was also classified as weak, moderate, or strong, and an immunohistochemical composite score, taking into account both extent and intensity of staining, was calculated. On whole tissue sections, there was a statistically significant difference between HMGA2 staining in serous and endometrioid carcinomas with regard to both extent and composite score, with higher expression in serous carcinomas (P < 0.0001). Thirty of 33 (91%) serous carcinomas were positive, usually with diffuse (3+ or 4+) staining. All five cases of serous endometrial intraepithelial carcinoma (EIC) (the postulated precursor of uterine serous carcinoma) were positive, as were 14 of 38 (37%) endometrioid carcinomas, usually with 1+ or 2+ staining. There was a statistically significant difference in HMGA2 staining in the TMAs between the serous and grade 3 endometrioid carcinomas, with higher expression in the former (P < 0.0001). Immunoreactivity for HMGA2 is diffusely positive in whole tissue sections in most uterine serous carcinomas and negative in most endometrioid carcinomas, although, as with other markers, there is overlap in individual cases. In conjunction with other markers, HMGA2 may be of value in problematic uterine carcinomas where the differential diagnosis includes serous and endometrioid carcinoma. As HMGA2 is expressed in serous EIC, this suggests that it may be implicated in the early development of uterine serous carcinoma.

Background: Uterine papillary serous carcinoma (UPSC) and uterine carcinosarcoma (UCS) are rare subtypes of endometrial cancer (EMCA) with poor prognosis. Both are high risk histotypes, though their distinct molecular and genomic profiles are undefined. Recent clinical trials such as RUBY, Keynote 775, DUO-E and NRG-GY018 have shown survival benefit in subsets of EMCA with mismatch repair deficiency (MMRd), however, small numbers of patients with UPSC and UCS were included. The role of MMRd or DNA damage &amp; response (DDR) mechanisms in general are under-explored in these EMCA subtypes. We therefore sought to characterize the immune, inflammatory, and DNA damage profiles of UPSCs and UCSs to better define the role of immunotherapy (IO) in these tumors. Methods: Total DNA damage was analyzed in 53 UPSC and 43 UCS FFPE samples collected at our institution using repair-assisted damage-detection (RADD). RNA was isolated from a subset of patients (n = 18/group) and expression (50 ng/per sample) was analyzed using the nCounter PanCancer IO 360 panel from Nanostring comprising 770 gene expression markers. Data was analyzed using the Rosalind Bioinformatics in conjunction with Nanostring’s IO360 analysis team. Results: We have previously shown a 3.6 fold decrease (p&amp;lt;0.0001) of unrepaired DNA damage between UCS samples compared to UPSC, suggesting greater DNA repair capacity in UCS than UPSC. Consistent with this data, transcriptional analysis revealed increased expression of DNA repair genes in UCS such as POLD1, MSH2, BRIP1, BIRC5, and CDK6, among others. Interestingly, decreased expression of DNA repair genes was noted in UPSC despite nearly 4 times the amount of unrepaired DNA damage. As expected, the immune system was activated in response to this increased DNA damage with higher levels of T cells, NK cells, IL-10, cytotoxic T cells, and Th1 cells. However, there was also increased expression of immune evasive genes such as PD-1, CD47, and VTCN1, among others. Additionally, expression of NK CD56dim was also upregulated, a known marker of immune exhaustion. This suggests DNA damage tolerance and immune exhaustion are hallmarks of UPSC. Conclusions: We present here a detailed, treatment-oriented molecular comparison between two histologically distinct EMCA subtypes, UPSC and UCS. Assessment of unrepaired DNA damage and, by proxy, DNA repair capacity, gives context to the immune transcriptomic landscape. Our data suggests that the immune exhausted molecular landscape of UPSC is more amenable to IO than that of UCS, while the DNA repair-robust UCS may be more vulnerable to agents targeting DNA Damage repair such as PARP inhibitors. Further, our data suggests that estimation of DNA repair capacity via RADD may be as treatment informative as molecular sequencing. Citation Format: Kevin J. Lee, Tanvi Joshi, Sagar Chokshi, Mackenzie Cummings, Jennifer Scalici, Nathaniel Jones. A molecular comparison reveals distinct transcriptomic differences between uterine carcinosarcoma and papillary serous carcinoma distinguishable by DNA damage [abstract]. In: Proceedings of the American Association for Cancer Research Annual Meeting 2025; Part 1 (Regular Abstracts); 2025 Apr 25-30; Chicago, IL. Philadelphia (PA): AACR; Cancer Res 2025;85(8_Suppl_1):Abstract nr 500.

Background: Platinum-containing chemotherapy is treatment with high efficacy in homologous recombination (HR) deficient cancers that arise in carriers of mutations in the BRCA1/2 genes. However, previous reports have shown that TNBC patients carrying no BRCA1/2 mutations also could benefit from platinum therapy. This study aimed to explore the association between HR-related gene mutations and genomic instability, and further evaluate the effectiveness of platinum-containing chemotherapy preliminarily. Methods: A total of 386 TNBC patients were screened from a surgical cohort (NCT01150513) and a metastatic cohort. Finally, 189 TNBC patients with clinical and tumor sequencing data availability were included, including 149 patients treated with radical surgery and adjuvant chemotherapy. All tumor samples were collected during the surgical operation and subjected to NGS for evaluating homologous recombination deficiency (HRD) score and 15 HR-related gene pathogenic or likely pathogenic mutations (including ATM, BARD1, BRCA1, BRCA2, BRIP1, CDK12, CHEK1, CHEK2, FANCL, PALB2, PPP2R2A, RAD51B, RAD51D, RAD51C, RAD54L). HRD score was an algorithmic assessment of three biomarkers of genomic instability as loss of heterozygosity, telomeric-allelic imbalance, large-scale state transitions and based on over 10,000 single-nucleotide polymorphisms in the human genome. Results: In 189 TNBC cohort, 40 patients (21.1%) had BRCA1/2 deleterious mutations, with additional 26 patients (13.8%) carrying mutations in HR-related genes other than BRCA1/2. Compared with BRCA1/2-intact patients, those carrying BRCA1/2m displayed a significantly higher HRD score (median, 35.5 vs 20.0; P=0.02). However, patients with HRm obtained a similar HRD score with HRwt patients (median, 28.0 vs 21.0, P=0.83). TNBC patients with PALB2m (n=8) had a small trend higher HRD score compared with BRCA1/2m (median HRD score 39.0 and 35.5), and TNBC patients with RAD51 family (n=7), ATM (n=5) and other HR-related gene mutations had a numerical trend lower HRD score compared with BRCA1/2m (median HRD score 22.0, 4.0, 14.5 and 35.5). Of the 149 patients in surgical cohort, 74 received platinum-containing, and 75 underwent platinum-free adjuvant chemotherapy. We analyzed the associations between HR-related gene mutations and disease-free survival (DFS), and found that patients with RAD51Bm (n=3), RAD51Cm (n=1) or PPP2R2Am (n=1) displayed worse DFS for patients treated with adjuvant chemotherapy (P =0.012, P&amp;lt;0.0001 or P&amp;lt;0.0001, respectively). Conclusions: In this study, we found that different HR-related genes mutation subtypes may exert distinct effects on genomic instability, and mutations in HR-related genes beyond the context of BRCA1/2 may serve as a biomarker for platinum-based adjuvant therapy for TNBC patients, which warrants further studies. Citation Format: Xue Wang, Yimeng Chen, Feng Du, Jian Yue, Yiran Si, Xiaochen Zhao, Lina Cui, Bei Zhang, Ting Bei, Peng Yuan, Binghe Xu. Effects of homologous recombination-related gene mutation subtypes on gene instability and the efficacy of platinum-containing regiments in triple-negative breast cancer [abstract]. In: Proceedings of the American Association for Cancer Research Annual Meeting 2022; 2022 Apr 8-13. Philadelphia (PA): AACR; Cancer Res 2022;82(12_Suppl):Abstract nr 5689.

Homologous recombination deficiency (HRD) score in germline BRCA2- versus ATM-altered prostate cancer.

e21121 Background: Homologous recombination (HR) gene mutations were reported to be associated with efficiency of immune checkpoint inhibitors (ICIs). However, up to date, no recognized standard has been set up for definition of HR mutation (+), which limited its clinical application. Homologous recombination deficiency (HRD) score reflects genomic scar induced by HR genes alteration and disfunction. Thus, it may be worth to explore whether HRD score could predict the efficacy of ICIs-based therapy. Methods: We collected tumor tissues from metastatic non-small cell lung cancer (NSCLC) patients who received first-line treatment of ICIs combined with platinum-based chemotherapy. HRD score testing were carried by BGI genomic scar analysis kit, which calculates the sum of loss of heterozygosity (LOH), telomeric allelic imbalance (TAI), large-scale state transitions (LST) scores, and corrected by ploidy of tumor samples, through next generation sequencing(NGS) data. The association of HRD score and response to ICIs plus chemotherapy were analyzed. Results: 22 EGFR/ALK wild-type metastatic NSCLC patients were included. The average HRD score was 24.57, varying from -26.37 to 92.34. Threshold traversal was performed based on analysis of the progression free survival (PFS) data from the included NSCLC patients. HRD score 31 or more were define as HRD (+). Kaplan-Meier PFS Survival analysis showed prolonged median PFS (mPFS) in HRD (+) versus HRD (-) NSCLC patients (N/A vs. 7.0 ms, Log-rank P = 0.029; HR 0.20, 95%CI 0.04-0.96, likelihood-ratio P = 0.03). The objective response rate (ORR) was 45.5% (5/11) in HRD(+) versus 27.2% (3/11) in HRD(-) patients. In patients with PD-L1 TPS≥1% (22C3), the mPFS was N/A vs. N/A, and the ORR was 75.0% (3/4) vs. 37.5% (3/8), in HRD (+) vs. HRD(-) patients. In patients with PD-L1 TPS &lt; 1% (22C3), the mPFS was 8.5 ms vs.2.0 ms (Log-rank P = 0.0085; HR 0.08, 95%CI 0.01-0.82, likelihood-ratio P = 0.02), and the ORR were 28.6% (2/7) vs. 0%(0/3), in HRD (+) vs. HRD(-) patients. Conclusions: In addition to PD-L1 expression level, HRD score would be a potential promising biomarker for predicting the efficiency of ICIs plus platinum-based chemotherapy in NSCLC patients.

In high-grade serous ovarian carcinoma (HGSC), deleterious mutations in DNA repair gene RAD51C are established drivers of defective homologous recombination and are emerging biomarkers of PARP inhibitor (PARPi) sensitivity. RAD51C promoter methylation (meRAD51C) is detected at similar frequencies to mutations, yet its effects on PARPi responses remain unresolved.In this study, three HGSC patient-derived xenograft (PDX) models with methylation at most or all examined CpG sites in the RAD51C promoter show responses to PARPi. Both complete and heterogeneous methylation patterns were associated with RAD51C gene silencing and homologous recombination deficiency (HRD). PDX models lost meRAD51C following treatment with PARPi rucaparib or niraparib, where a single unmethylated copy of RAD51C was sufficient to drive PARPi resistance. Genomic copy number profiling of one of the PDX models using SNP arrays revealed that this resistance was acquired independently in two genetically distinct lineages.In a cohort of 12 patients with RAD51C-methylated HGSC, various patterns of meRAD51C were associated with genomic "scarring," indicative of HRD history, but exhibited no clear correlations with clinical outcome. Differences in methylation stability under treatment pressure were also observed between patients, where one HGSC was found to maintain meRAD51C after six lines of therapy (four platinum-based), whereas another HGSC sample was found to have heterozygous meRAD51C and elevated RAD51C gene expression (relative to homozygous meRAD51C controls) after only neoadjuvant chemotherapy.As meRAD51C loss in a single gene copy was sufficient to cause PARPi resistance in PDX, methylation zygosity should be carefully assessed in previously treated patients when considering PARPi therapy. SIGNIFICANCE: Homozygous RAD51C methylation is a positive predictive biomarker for sensitivity to PARP inhibitors, whereas a single unmethylated gene copy is sufficient to confer resistance.

&lt;div&gt;Abstract&lt;p&gt;In high-grade serous ovarian carcinoma (HGSC), deleterious mutations in DNA repair gene &lt;i&gt;RAD51C&lt;/i&gt; are established drivers of defective homologous recombination and are emerging biomarkers of PARP inhibitor (PARPi) sensitivity. &lt;i&gt;RAD51C&lt;/i&gt; promoter methylation (me&lt;i&gt;RAD51C&lt;/i&gt;) is detected at similar frequencies to mutations, yet its effects on PARPi responses remain unresolved.&lt;/p&gt;&lt;p&gt;In this study, three HGSC patient-derived xenograft (PDX) models with methylation at most or all examined CpG sites in the &lt;i&gt;RAD51C&lt;/i&gt; promoter show responses to PARPi. Both complete and heterogeneous methylation patterns were associated with &lt;i&gt;RAD51C&lt;/i&gt; gene silencing and homologous recombination deficiency (HRD). PDX models lost me&lt;i&gt;RAD51C&lt;/i&gt; following treatment with PARPi rucaparib or niraparib, where a single unmethylated copy of &lt;i&gt;RAD51C&lt;/i&gt; was sufficient to drive PARPi resistance. Genomic copy number profiling of one of the PDX models using SNP arrays revealed that this resistance was acquired independently in two genetically distinct lineages.&lt;/p&gt;&lt;p&gt;In a cohort of 12 patients with &lt;i&gt;RAD51C&lt;/i&gt;-methylated HGSC, various patterns of me&lt;i&gt;RAD51C&lt;/i&gt; were associated with genomic “scarring,” indicative of HRD history, but exhibited no clear correlations with clinical outcome. Differences in methylation stability under treatment pressure were also observed between patients, where one HGSC was found to maintain me&lt;i&gt;RAD51C&lt;/i&gt; after six lines of therapy (four platinum-based), whereas another HGSC sample was found to have heterozygous me&lt;i&gt;RAD51C&lt;/i&gt; and elevated &lt;i&gt;RAD51C&lt;/i&gt; gene expression (relative to homozygous me&lt;i&gt;RAD51C&lt;/i&gt; controls) after only neoadjuvant chemotherapy.&lt;/p&gt;&lt;p&gt;As me&lt;i&gt;RAD51C&lt;/i&gt; loss in a single gene copy was sufficient to cause PARPi resistance in PDX, methylation zygosity should be carefully assessed in previously treated patients when considering PARPi therapy.&lt;/p&gt;Significance:&lt;p&gt;Homozygous &lt;i&gt;RAD51C&lt;/i&gt; methylation is a positive predictive biomarker for sensitivity to PARP inhibitors, whereas a single unmethylated gene copy is sufficient to confer resistance.&lt;/p&gt;&lt;/div&gt;

&lt;div&gt;Abstract&lt;p&gt;In high-grade serous ovarian carcinoma (HGSC), deleterious mutations in DNA repair gene &lt;i&gt;RAD51C&lt;/i&gt; are established drivers of defective homologous recombination and are emerging biomarkers of PARP inhibitor (PARPi) sensitivity. &lt;i&gt;RAD51C&lt;/i&gt; promoter methylation (me&lt;i&gt;RAD51C&lt;/i&gt;) is detected at similar frequencies to mutations, yet its effects on PARPi responses remain unresolved.&lt;/p&gt;&lt;p&gt;In this study, three HGSC patient-derived xenograft (PDX) models with methylation at most or all examined CpG sites in the &lt;i&gt;RAD51C&lt;/i&gt; promoter show responses to PARPi. Both complete and heterogeneous methylation patterns were associated with &lt;i&gt;RAD51C&lt;/i&gt; gene silencing and homologous recombination deficiency (HRD). PDX models lost me&lt;i&gt;RAD51C&lt;/i&gt; following treatment with PARPi rucaparib or niraparib, where a single unmethylated copy of &lt;i&gt;RAD51C&lt;/i&gt; was sufficient to drive PARPi resistance. Genomic copy number profiling of one of the PDX models using SNP arrays revealed that this resistance was acquired independently in two genetically distinct lineages.&lt;/p&gt;&lt;p&gt;In a cohort of 12 patients with &lt;i&gt;RAD51C&lt;/i&gt;-methylated HGSC, various patterns of me&lt;i&gt;RAD51C&lt;/i&gt; were associated with genomic “scarring,” indicative of HRD history, but exhibited no clear correlations with clinical outcome. Differences in methylation stability under treatment pressure were also observed between patients, where one HGSC was found to maintain me&lt;i&gt;RAD51C&lt;/i&gt; after six lines of therapy (four platinum-based), whereas another HGSC sample was found to have heterozygous me&lt;i&gt;RAD51C&lt;/i&gt; and elevated &lt;i&gt;RAD51C&lt;/i&gt; gene expression (relative to homozygous me&lt;i&gt;RAD51C&lt;/i&gt; controls) after only neoadjuvant chemotherapy.&lt;/p&gt;&lt;p&gt;As me&lt;i&gt;RAD51C&lt;/i&gt; loss in a single gene copy was sufficient to cause PARPi resistance in PDX, methylation zygosity should be carefully assessed in previously treated patients when considering PARPi therapy.&lt;/p&gt;Significance:&lt;p&gt;Homozygous &lt;i&gt;RAD51C&lt;/i&gt; methylation is a positive predictive biomarker for sensitivity to PARP inhibitors, whereas a single unmethylated gene copy is sufficient to confer resistance.&lt;/p&gt;&lt;/div&gt;

ObjectivesThis study aimed to investigate human epidermal growth factor receptor 2 (HER2) protein expression and gene amplification status in uterine serous carcinoma (USC) and ovarian high-grade serous carcinoma (HGSC), determine the frequencies of HER2 overexpression/amplification and HER2-low expression, evaluate two current HER2 interpretation criteria, and analyze the relationship between HER2 status and patient prognosis.MethodsA total of 51 patients with USC (including eight cases of mixed endometrial adenocarcinoma with predominant serous carcinoma components) and 165 patients with ovarian HGSC (including 43 patients who received neoadjuvant chemotherapy) were retrospectively recruited from the Department of Pathology, Peking University People’s Hospital between January 2019 and January 2025. Clinical and pathological characteristics were summarized. HER2 protein expression in whole-tissue sections was assessed using immunohistochemistry (IHC). HER2 scoring was performed according to two criteria: the 2023 American Society of Clinical Oncology/College of American Pathologists HER2 Testing Guidelines in Breast Cancer and the HER2 Immunohistochemistry Scoring System for Endometrial Serous Carcinoma proposed by the International Society of Gynecological Pathologists (ISGyP). Fluorescence in situ hybridization (FISH) was subsequently performed on cases with IHC scores of 1+, 2+, or 3+ for validation. Survival analysis was conducted based on HER2 status.ResultsHER2 gene amplification was detected in 15/51 (29.4%) tissues from patients with USC (including mixed carcinomas with predominant serous morphology). HER2 amplification was identified in 6/122 (4.9%) tissues from patients with primary ovarian HGSC without prior chemotherapy, and in 3/43 (7.0%) tissues from patients with ovarian HGSC with residual disease after neoadjuvant chemotherapy. Evaluation of HER2 IHC results using both interpretation criteria revealed no difference in HER2 3+ expression rates for both USC and ovarian HGSC. However, differences existed in the classification of HER2-low expression (defined as IHC 2+ with negative FISH or IHC 1+). The ISGyP criteria identified more cases with low HER2 expression. For patients with USC, HER2 positivity was not statistically significantly associated with overall survival (OS) or progression-free survival (PFS), although it showed a trend toward higher recurrence. In terms of prognosis, for patients with ovarian HGSC group, HER2 positivity was associated with worse OS (p = 0.049). Among patients with ovarian HGSC after neoadjuvant chemotherapy, HER2 positivity was a strong predictor for shorter PFS (p = 0.003).ConclusionA certain proportion of USC and ovarian HGSC cases exhibit HER2 positivity and HER2-low expression. Different interpretation criteria lead to variations in the assessment of HER2 IHC results. The ISGyP criteria can identify more cases with low HER2 expression. Moreover, our findings suggest that HER2 status could be a relevant prognostic marker in these malignancies. Traditional anti-HER2 targeted therapies are indicated for HER2-positive patients, while a broader population of patients with HER2-low expression may benefit from novel anti-HER2 antibody-drug conjugates.