Mutant Ovarian Cancer Research Articles

Loss of CHD1 causes DNA repair defects and enhances prostate cancer therapeutic responsiveness.

The CHD1 gene, encoding the chromo‐domain helicase DNA‐binding protein‐1, is one of the most frequently deleted genes in prostate cancer. Here, we examined the role of CHD1 in DNA double‐strand break (DSB) repair in prostate cancer cells. We show that CHD1 is required for the recruitment of CtIP to chromatin and subsequent end resection during DNA DSB repair. Our data support a role for CHD1 in opening the chromatin around the DSB to facilitate the recruitment of homologous recombination (HR) proteins. Consequently, depletion of CHD1 specifically affects HR‐mediated DNA repair but not non‐homologous end joining. Together, we provide evidence for a previously unknown role of CHD1 in DNA DSB repair via HR and show that CHD1 depletion sensitizes cells to PARP inhibitors, which has potential therapeutic relevance. Our findings suggest that CHD1 deletion, like BRCA1/2 mutation in ovarian cancer, may serve as a marker for prostate cancer patient stratification and the utilization of targeted therapies such as PARP inhibitors, which specifically target tumors with HR defects. ![][1] Chromo‐domain helicase DNA‐binding protein‐1 (CHD1) promotes HR‐mediated DSB repair. The depletion of CHD1 enhances cellular sensitivity to PARP inhibitors, which has potential therapeutic implications for CHD1‐depleted prostate cancers. [1]: /embed/graphic-1.gif

Olaparib in the maintenance treatment of platinum-sensitive relapse of BRCA mutant ovarian cancer in routine clinical practice: first results of observational study in Russian patients

Background and aims. There is no data on olaparib efficacy and safety in Russian routine clinical practice. Methods. We analysed the 30 consecutive patients who received maintenance olaparib treatment for platinum-sensitive relapse (PSR) of ovarian or fallopian tube cancer in Russian Cancer Centers. Patients were prescribed olaparib capsules 400 mg twice daily. Radiographic assessments were done every 8 weeks. Patient characteristics. Age median 55 (range 39-68); 26 (86,6%) patients had gBRCA1, 2 (6,6%) patients had sBRCA1, 2 (6,6%) patients had gBRCA2. Number of relapse: median 1 (range 1-10), number of lines of chemotherapy: median 2 (range 2-11). Last regimen of chemotherapy: taxane + platinum (± bevacizumab) 90% (27/30), platinum monotherapy 10% (3/30). Best response to the last chemotherapy complete response 43,3% (13/30), partial response 36,7% (11/30), stable disease 20% (6/30). Results. Median follow-up in 13 CR patients was 12 mos. 1 CR patient progressed after 9 mos of maintenance olaparib. Median follow-up in 11 PR patients was 7 mos. 3 PR patients achieved CR on olaparib. 1 PR patient progressed after 6 mos of olaparib maintenance. Median follow-up in SD patients was 12 mos. 1 SD patient achieved PR on olaparib, there were no progressions. 10 (30%) patients had adverse events (AEs). 1 patient had grade 3 AE and 2 patients had AEs leading to dose reduction. There were no grade 4 AEs. Conclusions. Olaparib is safe and effective maintenance treatment of PSR ovarian cancer in routine clinical settings.

Abstract IA20: Therapeutic targeting of ARID1A mutation in ovarian cancer

Abstract SWI/SNF chromatin remodeling complexes regulate gene transcription by changing chromatin structure through hydrolyzing ATP. Cancer genome sequencing found that mutations in genes encoding for the subunits of the SWI/SNF complexes collectively occur in ~20% of all human cancers. For example, saturation analysis of The Cancer Genome Atlas (TCGA) cancer mutational profile reveals that the ARID1A subunit of the SWI/SNF complex shows one of the highest mutation rates among epigenetic regulators. Most notably, ARID1A is mutated in over 50% of ovarian clear cell carcinomas and 30% of ovarian endometrioid carcinomas. ARID1A mutation is a known genetic driver of ovarian cancer. Over 90% of the ARID1A mutations observed in ovarian cancer are frame-shift or nonsense mutations that result in loss of ARID1A protein expression. Loss of ARID1A correlates with late-stage disease and predicts early recurrence of ovarian clear cell carcinoma. Ovarian clear cell carcinoma ranks second as the cause of death from epithelial ovarian cancer and is associated with the worst prognosis among the major ovarian cancer subtypes when diagnosed at advanced stages. Additionally, for advanced-stage disease, there is currently no effective therapy. Here we will discuss emerging opportunities for therapeutic targeting of ARID1A mutation based on the genetic makeup of ARID1-mutated ovarian cancer with precision. They include synthetic lethality, the mutual exclusivity between ARID1A and TP53 mutations in ovarian cancer, and mechanism-guided combinatory approaches. Citation Format: Rugang Zhang. Therapeutic targeting of ARID1A mutation in ovarian cancer. [abstract]. In: Proceedings of the AACR Conference: Addressing Critical Questions in Ovarian Cancer Research and Treatment; Oct 1-4, 2017; Pittsburgh, PA. Philadelphia (PA): AACR; Clin Cancer Res 2018;24(15_Suppl):Abstract nr IA20.

Abstract 395: Identifying the functional drivers of noncoding somatic mutations in ovarian cancer

Abstract Epithelial ovarian cancer (OC) is a heterogeneous disease that has been stratified into different histologic subtypes: high-grade serous (HGSOC), clear cell (CCOC), endometrioid (EnOC) and mucinous (MOC). HGSOC is the most common, but every histotype is characterized by largely distinct germline genetics, somatic alterations and clinical biomarkers. Recently, whole-genome sequencing (WGS) studies have catalogued genome-wide somatic variation for most OC histotypes. These data demonstrate that OC histotypes harbor thousands of noncoding somatic mutations and our next major challenge is to distinguish the few important noncoding somatic drivers from the thousands of passenger mutations. We hypothesized that driver noncoding somatic mutations impact disease development and progression through altering the sequence of regulatory elements (REs), such as enhancers and promoters, eventually resulting in perturbation of the expression of target genes. To systematically address this hypothesis, we established genome-wide H3K27ac epigenomic profiles, annotating active REs for the different ovarian cancer histotypes using chromatin immunoprecipitation sequencing (ChIP-seq) in 20 fresh frozen primary OC tissue samples—five tumors for each major histotype. In parallel, we performed transcriptional profiling using RNA sequencing (RNA-seq). Together, these two datasets enabled us to evaluate epigenetic alterations and the transcriptome. We identified histotype-specific active REs, and common active REs across all histotypes. We used the RNA-seq data to assess the effect of the histotype-specific REs in gene expression, and to find target genes of cis-REs and novel histotype-specific biomarkers. Next, we integrated these unique profiles with WGS data from 232 OCs (169 HGSOCs, 35 CCOCs and 28 EnOCs). The number of somatic single-nucleotide variants per sample range from 481 to 40,764 (mean=7199, sd = 5751). Of these, 9.4% were noncoding and overlapped active REs in OC. Using a Poisson binomial distribution, we tested the significance of the observed number of mutated samples for any given active RE. Importantly, the method adjusted for interpatient mutation rate heterogeneity. Using the Benjamini-Hochberg procedure for p-value correction, we identified several significantly mutated active REs, including the promoters of POLR3E and WDR74. In conclusion, we have used an integrative method to identify functional, driver noncoding somatic mutations for ovarian cancer based on their interaction with disease-specific regulatory elements, and their putative target genes. This represents a powerful way to distinguish important noncoding somatic drivers from a much larger number of passenger mutations that accumulate during tumor development. The approach can be applied to other cancer types if sufficient depth and quality of genetic, transcriptomic and epigenomic datasets are available. Citation Format: Rosario I. Corona, Ji-Heui Seo, Dennis J. Hazelett, Xianzhi Lin, Paulette Y. Mhawech-Fauceglia, Jenny Lester, Sohrab Shah, David G. Huntsman, Beth Y. Karlan, Benjamin P. Berman, Matthew L. Freedman, Simon A. Gayther, Kate Lawrenson. Identifying the functional drivers of noncoding somatic mutations in ovarian cancer [abstract]. In: Proceedings of the American Association for Cancer Research Annual Meeting 2018; 2018 Apr 14-18; Chicago, IL. Philadelphia (PA): AACR; Cancer Res 2018;78(13 Suppl):Abstract nr 395.

Abstract CT159: Olaparib combinations (OLAPCO)for homology-directed DNA repairdefects (HDR)

Abstract INTRODUCTION: Genetic profiling of tumors used to identify treatment choices for cancer patients focus on kinase mutations and well-defined translocations; multiple clinical trials have been initiated for specific kinase-targeted agents. About 15-20% of patients have actionable mutations. HDR have been found in ~ 20 % of patient tumors molecularly profiled at the Yale Cancer Center. Mutations in HDR related genes often occur in addition to other mutations in signaling pathways. Specific targeted treatment options for these patients are limited, as breast and ovarian cancer mutations are the only indication for the HDR pathway targeted PARP inhibitors. Additional drugs are in development, but few clinical studies based on HDR other than BRCA1/2 mutations are open. METHODS: The OLAPCO study targets HDR in patients irrespective of tumor origin. The protocol includes four arms to which patients can be enrolled based on tumor profiling results: olaparib monotherapy (PARP inhibitor) for HDR; olaparib & AZD5363 ( AKT inhibitor) for PIK3CA/AKT/PTEN pathway dysregulation and ARID1A mutations; olaparib & AZD1775 ( wee1 inhibitor) for TP53 mutations and olaparib & AZD6738 (ATR inhibitor) for HDR and olaparib pre-treated patients. The clinical endpoints are to determine tumor overall response rate (ORR) and clinical benefit rate (CBR) by RECIST after 16 weeks of treatment. RESULTS: Arms a and b are open to patient accrual. Together, 34 pts have been treated, 12 remain on treatment, with no new or > Gr 2 (CTCAE4) treatment-related toxicities. TABLE 1Olaparib total = 23; median # cycles= 2, range 1-10+ ; 7 ongoingMutationATMBRCA1IDH1/2FHPALB2CHK2BAP1ARID 1A#52821212ORR1 PRCBR10201000TABLE 2Olaparib & AZD 5363 total = 12; median # cycles = 2, range 1-8+; 5 ongoingMutationAKT1AKT3AKT2 & PALB2ARID 1AARID 1A & PIK3CAPTEN#321411ORR1CBR10101 DISCUSSION: HDR defects are actionable mutations, increasing the number of patients for whom tumor profiling can identify a therapeutic option. This exploratory trial is expanding the therapeutic spectrum of olaparib alone and in combination, to include heretofore non-canonical alterations in HDR, beyond BRCA1/2. This trial design allows additional molecular targets, agents & combinations to be rapidly evaluated. AstraZeneca support is gratefully acknowledged. Citation Format: Joseph P. Eder, Geoffrey I. Shapiro, James M. Cleary, Vickie L. Keedy, Vickie L. Keedy, Ranjit Bindra, Brian Shuch, Jeffrey Sklar, Khanh Do, Juliane Juergensmeier. Olaparib combinations (OLAPCO)for homology-directed DNA repairdefects (HDR) [abstract]. In: Proceedings of the American Association for Cancer Research Annual Meeting 2018; 2018 Apr 14-18; Chicago, IL. Philadelphia (PA): AACR; Cancer Res 2018;78(13 Suppl):Abstract nr CT159.

Abstract 874: Pharmacologically induced RAS-GTP levels and CRAF-BRAF hetero-dimerization drive sensitization to Type II pan-RAF inhibitors in KRAS mutant cancer

Abstract Although effective in BRAF V600 mutant melanoma, Type 1.5 RAF inhibitors such as vemurafinib and dabrafenib have not proven to be successful in KRAS mutant cancers, neither as single agent nor in combination with MEK inhibitors. Through large-scale cellular compound combination screening we found that Type II RAF inhibitors such as AZ-628 do show synergistic activity with MEK inhibitors in multiple KRAS mutant indications, including NSCLC, colorectal cancer and ovarian cancer. The combination of Type II RAF inhibitors and MEK inhibitors demonstrates robust and durable abrogation of MAPK signaling both on canonical markers of MAPK activity such as pERK and pRSC as well as transcriptional output. We also observe synergistic in vivo tumor growth inhibition in two independent models of KRAS mutant cancer by this combination treatment. We found that treatment with MEK inhibitors alone drives the increase of active RAS-GTP levels and induces CRAF:BRAF hetero-dimerization. These induced dimers are active and able to phosphorylate MEK in vitro. This increased dimerization renders cells sensitive to Type 2 RAF inhibitors. We find that this effect is not limited to KRAS mutant cells, as a subset of KRAS wild-type cells show increased RAS-GTP levels upon MEK inhibitor treatment. These cells also show synergistic sensitivity to Type 2 RAF inhibition. Additionally, we observed significantly higher synergy and higher RAS-GTP levels in KRAS G13D mutant cells, which have intrinsically high GDP exchange and low intrinsic GTP hydrolysis. Finally, we show that GDC-0941 and GDC-0032, two broad PI3K inhibitors, also induce RAS-GTP levels in cells independent of PIK3CA or KRAS mutation status. We subsequently observed a synergistic sensitivity to Type 2 RAF inhibitors in these PI3K inhibitor-treated cells. Overall, we demonstrate that pharmacologic inhibition of MEK or PI3K increases RAS-GTP levels and drives increased CRAF:BRAF hetero-dimerization. This in turn sensitizes cells to Type 2 RAF inhibition, leading to a synergistic drug effect. Combination of these inhibitors may be a viable therapeutic approach in KRAS mutant cancer, and may be especially effective in KRAS G13D-carrying tumors. Citation Format: Christiaan N. Klijn, Ivana Yen, Frances Shanahan, Mark Merchant, Christine Orr, Thomas Hunsaker, Matthew Durk, Hank La, Xiaoling Zhang, Scott Martin, Eva Lin, John Chan, Yihong Yu, Dhara Amin, Amy Gustafson, Scott Foster, Joachim Rudolph, Shiva Malek. Pharmacologically induced RAS-GTP levels and CRAF-BRAF hetero-dimerization drive sensitization to Type II pan-RAF inhibitors in KRAS mutant cancer [abstract]. In: Proceedings of the American Association for Cancer Research Annual Meeting 2018; 2018 Apr 14-18; Chicago, IL. Philadelphia (PA): AACR; Cancer Res 2018;78(13 Suppl):Abstract nr 874.

Abstract 2344: Identification and validation of synthetic lethal cancer pairs from genome-wide CRISPR screens

Abstract Although most canonical somatic cancer mutations have now been cataloged, only a fraction are amenable to drug discovery efforts. These are limited to targets with gain of function mutations and druggable protein domains. Examples include kinases such as FLT3 and metabolic enzymes such as IDH1/2. However, excluded from this list are undruggable protein targets with gain of function mutations, such as KRAS, or tumor suppressors which acquire loss of function mutations, such as APC. The concept of synthetic lethality is being used to address this gap in cancer discovery. Cells which carry these directly undruggable somatic mutations become susceptible to inhibition of secondary targets. This susceptibility is specific in context to cells carrying the cancer associated mutation, offering both a novel approach for establishing efficacy as well as a potential therapeutic window for normal cells. The successful development of PARP inhibitors for BRCA mutant ovarian and breast cancers has provided the first clinical validation of this idea. Pooled, genome-wide knock out experiments are being pursued to systematically discover more of these types of synthetic lethalities. Typically, a sgRNA library is used to transfect a CAS9 expressing cancer cell line, and sequencing of the guides over time identifies lethalities specific to the cell line. Once this experiment is repeated over hundreds of cell lines, one can use the genomic information of the cell lines, such as mutations or copy number aberrations, to identify potential new combinations of molecular lethality. We have analyzed pooled CRISPR knockout data in a large panel of cancer cell lines generated from two independent sources (Broad Institute and Open Targets) to identify these types of lethality. Besides identifying targets for prevalent cancer mutations, we have also identified lethal combinations based on genes aberrantly deleted/silenced in cancers, termed “collateral lethality.” These include pairs of paralogs, many of which code enzymes with redundant functions. In this presentation we will outline our analysis strategies and methods, and describe the development of a downstream experimental validation pipeline, using a specific target/predictor pair to exemplify our approach. Citation Format: Benjamin Schwartz, Junping Jing, Euan Stronach, David Cooper, Boris Wilson, Sarah Middleton, Mugdha Khaladkar, Coco Dong, Usman Shabon, Yanhua Rao, Stefanie Riesenberg. Identification and validation of synthetic lethal cancer pairs from genome-wide CRISPR screens [abstract]. In: Proceedings of the American Association for Cancer Research Annual Meeting 2018; 2018 Apr 14-18; Chicago, IL. Philadelphia (PA): AACR; Cancer Res 2018;78(13 Suppl):Abstract nr 2344.

Abstract 2322: Ferroptosis induced by erastin in RAS mutant ovarian cancer cells

Abstract Although apoptosis has been thought as the principal mechanism of programed cell death in chemotherapy, other forms of non-apoptotic death also has been noted. The oncogenic RAS-selective lethal small molecule erastin triggers a unique iron-dependent form of cell death termed ferroptosis. This mechanism is expected to induce eradication of chemotherapeutic (e.g. platinum) resistant cancer cells. In this study we demonstrate this new form of cell death in ovarian cancer cells. The clear cell epithelial ovarian cancer cell lines, TOV-21G (KRAS_G13C) and KOC-7C (RAS wild type) were used in this study. Erastin dose-dependently inhibited growth in TOV-21G cells, but not KOC-7C cells. Treatment of TOV-21G cells with erastin resulted in increase of ROS. Iron chelator deferoxamine (DFO) rescued growth inhibition induced by erastin in TOV-21G cells. Although cisplatin inhibited growth in TOV-21G cells, this inhibition was not rescued by DFO. The GSH level was reduced by erastin, but this reduction was not rescued by DFO. In addition, the basal GSH level was higher and the basal ROS level was lower in KOC-7C cells than those in TOV-21G cells. Erastin induced ferroptosis by reducing the GSH level in RAS mutant ovarian cancer cells. Although DFO rescued growth inhibition induced by erastin, the pathway was not on System Xc-. The sensitivity to erastin might depends on the basal level of GSH in cells. Ferroptosis had great potential to become a new approach in anti-tumor therapies in ovarian cancer. Citation Format: Motoki Takenaka, Noriko Suzuki, Minako Mori, Hitomi Aoki, Tasuku Hirayama, Hideko Nagasawa, Ken-ichirou Morishige. Ferroptosis induced by erastin in RAS mutant ovarian cancer cells [abstract]. In: Proceedings of the American Association for Cancer Research Annual Meeting 2018; 2018 Apr 14-18; Chicago, IL. Philadelphia (PA): AACR; Cancer Res 2018;78(13 Suppl):Abstract nr 2322.

Beyond BRCA1 and BRCA2: Implementation of a multigene panel for the upfront testing of germline mutations in ovarian cancer.

1542Background: Germline mutations impacting homologous recombination repair have been associated with predisposition to breast (BC) and ovarian cancer (OC). OC, even if no family history (FH) conf...

Abstract P4-07-01: Assessment of the hereditary component in 94 cancer predisposition genes to triple negative breast cancer

Abstract Background: In women with triple negative breast cancer (TNBC) unselected for age or family history, 8-14% and 5% of patients harbor germline mutations in BRCA1 and BRCA2, respectively. Diagnosis of TNBC <60 years of age is one of the NCCN criterion for genetic testing of BRCA1 and BRCA2. The contribution of germline mutations in other cancer predisposition genes to TNBC, is, however, not well-studied. Methods: TNBC was classified as tumors with <1% positively staining cells for ER and PR and HER2 = 0+, 1+ or 2+/not amplified. Genomic DNA was isolated from blood samples and targeted sequencing was performed using the TruSight Cancer panel (Illumina). Pathogenic mutations were identified using VariantStudio and classified as pathogenic, uncertain significance (VUS) or benign using ClinVar. Results: 196 female patients diagnosed with TNBC 2001-2014 had genomic DNA available. Average age at diagnosis was 52.8 years (range 34.1-83.4 years). The majority of patients were of European (66%) or African (31%) American ancestry; 26% had a family history and 13% had died of disease with an average time to death of 2.81 years. Twenty-three (12%) of women with TNBC had pathogenic mutations in breast cancer genes BRCA1 (n=14), BRCA2 (n=5), PALB2 (n=1) and CHEK2 (n=3), two women had mutations in the colon cancer genes MUTYH, one had a mutation in the ovarian cancer gene BRIP1, and an additional three women had pathogenic mutations in cancer predisposition genes FANCD2, SDHB and XPC. An additional 42 women had VUS in 20 genes, including one in BRCA1 and 5 in BRCA2. Discussion: Although the majority of pathogenic mutations in this cohort of women with TNBC were in the BRCA1 and BRCA2 genes (10%), panel testing allowed for the detection of mutations in other breast (2%), colon (1%), ovarian (1%) and other cancer (2%) predisposition genes. Panel testing thus identifies genes other than BRCA1/2 associated with increased risk of TNBC and may incidentally identify women who would benefit from enhanced surveillance for other cancers. The opinions or assertions contained herein are the private ones of the author/speaker and are not to be construed as official or reflecting the views of the Department of Defense, the Uniformed Services University of the Health Sciences or any other agency of the U.S. Government. Citation Format: Ellsworth RE, Lovejoy LA, Shriver CD. Assessment of the hereditary component in 94 cancer predisposition genes to triple negative breast cancer [abstract]. In: Proceedings of the 2017 San Antonio Breast Cancer Symposium; 2017 Dec 5-9; San Antonio, TX. Philadelphia (PA): AACR; Cancer Res 2018;78(4 Suppl):Abstract nr P4-07-01.

Tumour profiling for treatment of patients with ovarian cancers

With the introduction of Poly(ADP-ribose) polymerase inhibitors, a novel and effective targeted treatment has become available for women with recurrent ovarian cancers and inactivating BRCA1 or BRCA2 variants. The utility of diagnostic testing for germline BRCA1/2 mutations in ovarian cancer is well established; testing assists in establishing causation, prognosis, and treatment options, future cancer risk and cancer risk reduction in affected family members.1 There is increasing evidence supporting the clinical importance of testing for somatic BRCA mutation testing in ovarian cancers,2 but there are no Australian publications quantifying additional yield or clinical utility in comparison to germline testing. In order to assess utility of BRCA1/2 somatic mutation testing over and above germline testing, we tested 18 FFPE samples from patients with high-grade serous ovarian cancer to estimate the prevalence of low allele frequency, actionable, somatic variants in a small cohort. We detected somatic variants of significance in BRCA1/2 and other homologous recombination genes which were actionable and would not have been detected through germline testing. Our results were consistent with other published reports,3 confirming that somatic BRCA1/2 testing in patients with advanced-stage epithelial ovarian cancer may provide clinicians and patients with additional information that could influence clinical management.

Chemokine Network and Overall Survival in TP53 Wild-Type and Mutant Ovarian Cancer.

Ovarian cancer (OC) has the highest mortality rate among gynecological malignancies. Because chemokine network is involved in OC progression, we evaluated associations between chemokine expression and survival in tumor suppressor protein p53 (TP53) wild-type (TP53WT) and mutant (TP53m) OC datasets. TP53 was highly mutated in OC compared to other cancer types. Among OC subtypes, CXCL14 was predominantly expressed in clear cell OC, and CCL15 and CCL20 in mucinous OC. TP53WT endometrioid OC highly expressed CXCL14 compared to TP53m, showing better progression-free survival but no difference in overall survival (OS). TP53m serous OC highly expressed CCL8, CCL20, CXCL10 and CXCL11 compared to TP53WT. CXCL12 and CCL21 were associated with poor OS in TP53WT serous OC. CXCR2 was associated with poor OS in TP53m serous OC, while CXCL9, CCL5, CXCR4, CXCL11, and CXCL13 were associated with better OS. Taken together, specific chemokine signatures may differentially influence OS in TP53WT and TP53m OC.

BRCA1 and BRCA2 mutations in ovarian cancer

BRCA1 and BRCA2 mutations in ovarian cancer

Prognosis Analysis of Histone Deacetylases mRNA Expression in Ovarian Cancer Patients.

Histone deacetylases modulate the dynamic balance of histone acetylation and deacetylation in cells, which participate in epigenetic regulations. Accumulated evidence has demonstrated that histone deacetylases are associated with angiogenesis, cell proliferation and survival in a variety of human cancers. However, the expression and distinct prognostic value of histone deacetylases in ovarian cancer have not been well elucidated. In the present study, we collected the overall survival (OS), progress free survival (PFS), and histone deacetylases (HDAC1-11) mRNA expression in ovarian cancer from the Kaplan-Meier plotter online database. We investigated the relationship between histone deacetylases mRNA level and the clinicopathological parameters of the ovarian cancer patients, such as histology subtypes, clinical stages, grades and TP53 mutation. Our analysis data showed that over-expression of HDAC1, HDAC2, HDAC4, HDAC5 and HDAC11 were correlated to poor overall survival and unfavorable progress free survival in all ovarian cancer patients. Notably, the higher level of HDAC11 was associated with the worse OS and PFS for serous/ stage III+IV/ grade III/ TP53 mutation ovarian cancer patients. In conclusion, HDACs may play a crucial role in the prognosis of ovarian cancer, but it is worth noting that HDAC11 may be a biomarker for poor prognosis in ovarian cancer patients.

Chromatin accessibility underlies synthetic lethality of SWI/SNF subunits in ARID1A-mutant cancers.

ARID1A, a subunit of the SWI/SNF chromatin remodeling complex, is frequently mutated in cancer. Deficiency in its homolog ARID1B is synthetically lethal with ARID1A mutation. However, the functional relationship between these homologs has not been explored. Here, we use ATAC-seq, genome-wide histone modification mapping, and expression analysis to examine colorectal cancer cells lacking one or both ARID proteins. We find that ARID1A has a dominant role in maintaining chromatin accessibility at enhancers, while the contribution of ARID1B is evident only in the context of ARID1A mutation. Changes in accessibility are predictive of changes in expression and correlate with loss of H3K4me and H3K27ac marks, nucleosome spacing, and transcription factor binding, particularly at growth pathway genes including MET. We find that ARID1B knockdown in ARID1A mutant ovarian cancer cells causes similar loss of enhancer architecture, suggesting that this is a conserved function underlying the synthetic lethality between ARID1A and ARID1B.

Abstract 2421: The role of p53 mutational status and SOX9 suppression in chemotherapy response of ovarian cancer

Abstract Ovarian cancers are highly heterogeneous where platinum based chemotherapy which induces DNA crosslinking resulting in apoptosis of the cell is the preferred treatment. However, many patients are intrinsically resistant or quickly develop resistance. The Sox factors are a large family of transcription factors that play important roles in tumor development and progression in a variety of human malignancies and diverse developmental processes, but their impact in clinical tumorigenesis is still unclear. An analysis of genomic changes in ovarian cancer has provided the most comprehensive and integrated view of cancer genes for any cancer type to date. Ovarian serous adenocarcinoma tumors from 500 patients were examined by The Cancer Genome Atlas (TCGA) Research Network and analyses are reported in a recent issue of Nature. This evidence suggests that epigenetic deregulation, such as methylation, may be a key factor in the onset and maintenance of chemoresistance. Previous microarray analysis results in our lab correctly identified a subset of about 300 genes that when methylated altered the chemoresistance of the ovarian epithelium cells in culture. Of the genes identified in the analysis we further set out to characterize oncogenes and tumor suppressor genes that interact with the guardian of the genome, TP53, to determine if we could elucidate the mechanism by which it increased resistance. Using several in-vitro assays, we determined that the loss of p53 in conjunction with SOX9 decreased the level of apoptosis in response to carboplatin. Furthermore, in cells with mutated p53/SOX9 show an increase in tumorigenesis. Regulation of several pathways with p53 mutations in ovarian cancer might represent a therapy response prediction and could be a future therapeutic target for ovarian cancer. In addition, the crosstalk between p53/SOX9 and epigenetic regulators may present a valid treatment option for increasing carboplatin sensitivity in resistant patients. Note: This abstract was not presented at the meeting. Citation Format: Noelle L. Cutter, Matthew Lucito, Ryan Frank, Kimberly Doyle. The role of p53 mutational status and SOX9 suppression in chemotherapy response of ovarian cancer [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 2421. doi:10.1158/1538-7445.AM2017-2421

Abstract 4035: A novel combi-molecule engineered to target the putative synthetic lethal interactions between the epidermal growth factor receptor (EGFR) and poly(ADP-ribose)polymerase (PARP)

Abstract Over the past decade, PARP inhibition has been actively pursued as a novel approach for the selective therapy of tumors with BRCA1/2 mutations. The therapeutic benefits of PARP inhibitors have now been proven in the clinic against BRCA1/2 mutant ovarian cancers. This is hitherto limited to BRCA1/2 mutations, which only accounts for 5-10% of all cancers with hereditary mutations in the homologous recombination pathway. Therefore, new strategies are not only required to enhance the potency of PARP inhibitors but also to expand their use beyond BRCA mutation. While several combination modalities have been reported for PARP inhibitors, the concept of targeted PARP inhibitor has not yet been explored. Here using our novel combi-targeting approach, we report on the design of PARP inhibitors targeted to EGFR, a tyrosine kinase receptor overexpressed in several solid tumors. Recently, reports on the relationship between EGFR, PARP and BRCA have begun to emerge, one of which described a contextual synthetic lethality between EGFR and PARP (PloS one 7.10 (2012)). Here we report on the design and synthesis of novel PARP-EGFR combi-molecule based on structural modification of olaparib as a PARP inhibitor warhead and the quinazoline moiety for targeting EGFR. The results showed that: (a) it is capable of inducing a dose-dependent inhibition of PARP in isolated enzyme assay, (b) it induced a dose-dependent inhibition of EGFR in an isolated kinase assay, (c) it showed a dose-dependent inhibition of EGFR phosphorylation and downstream signaling in whole-cell assay, (d) it was selectively potent towards BRCA2 mutant and also EGFR-overexpressing cell lines, (d) it was extremely potent with activities superior to that of olaparib or gefitinib alone and their corresponding equimolar combination in three established triple negative breast cancer cell lines, (e) subcellular distribution analysis showed that it was abundantly localized in the perinuclear region. These results in toto suggest that this new combi-molecule could be developed as a single drug modality emulating the combination of PARP and EGFR inhibitors with the added benefit of being targeted to EGFR-expressing tumor cells. Citation Format: Zhor Senhaji Mouhri, Martin Rupp, Bertrand J. Jean-Claude. A novel combi-molecule engineered to target the putative synthetic lethal interactions between the epidermal growth factor receptor (EGFR) and poly(ADP-ribose)polymerase (PARP) [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 4035. doi:10.1158/1538-7445.AM2017-4035

Abstract 2475: In vitro and in vivo assessment of the mechanism of action of the PARP inhibitor rucaparib

Abstract Background: Rucaparib (CO-338) is an oral small molecule inhibitor of poly (ADP-ribose) polymerase (PARP) that is being developed for patients with BRCA1 and BRCA2 mutated and homologous recombination deficient ovarian cancer (Swisher et al., 2016; Lancet Oncol. Epub). The mechanism of action (MOA) of rucaparib was characterized in the studies reported here. Methods: Enzymatic and cellular reporter assay profiling were performed by BPS Biosciences. Rucaparib potency was assessed using 6-day cell viability assays (CellTiter-Glo). In vivo PK/PD and efficacy studies in the MDA-MB-436 (BRCA1 mutant) and the HBCx-17 (BRCA2 mutant) TNBC models were performed at Crown Biosciences and XenTech, respectively. Results: In biochemical assays rucaparib inhibited PARP-1, PARP-2 and PARP-3, with IC50 values of 0.8, 0.5, and 28 nM, respectively. Rucaparib weakly inhibited PARP5a (tankyrase 1[TNKS1]) and PARP5b (tankyrase 2[TNKS2]) with an IC50 of 796 and 486 nM; however rucaparib did not inhibit TNKS1/2 in a cellular reporter assay evaluating Wnt pathway signaling (IC50 >10 µM). Rucaparib cytotoxicity was evaluated in the BRCA1 mutant (UWB1.289) and wild-type (UWB1.289+BRCA1) isogenic pair, and a panel (n=26) of breast and ovarian cancer cell lines. The UWB1.289 cell line was more sensitive to rucaparib (IC50 = 375 nM) than the UWB1.289+BRCA1 cell line (IC50 = 5430 nM). Rucaparib also decreased the cell viability of several BRCA1 and BRCA2 mutant ovarian and breast cancer cell lines. Mechanistic analyses in the UWB1.289 cell line demonstrated a dose dependent decrease in poly (ADP) ribosylation (PAR; IC50 = 2.8 nM), and an increase in DNA damage and apoptosis. Taken together, these results are consistent with the concept of synthetic lethality, where simultaneous inhibition of multiple DNA repair pathways results in irreparable DNA damage and cell death. In vivo PK/PD and efficacy studies assessed the MOA and activity of rucaparib in the MDA-MD-436 (BRCA1 mutant) xenograft model. In a PK/PD study, dose dependent inhibition of PAR was observed in the tumors of mice treated with 15, 50 and 150 mg/kg BID rucaparib, resulting in statistically significant (p < 0.05) inhibition of 45%, 86% and 96%, respectively. An inverse and dose dependent correlation between PAR and rucaparib levels in the plasma and tumor was observed, and the levels of rucaparib in the tumor were consistently higher (> 2-fold) than the levels of rucaparib measured in plasma. Dose dependent and statistically significant tumor growth inhibition was observed in both subcutaneous and orthotopic xenograft studies evaluating the anti-tumor activity of rucaparib in the MDA-MB-436 model. Rucaparib also demonstrated potent activity in the BRCA2 mutant HBCx-17 patient-derived xenograft model. Conclusion: These results demonstrate that rucaparib is a potent and selective inhibitor of PARP1, PARP-2, and PARP-3 with in vitro and in vivo activity in BRCA1/2 deficient tumors. Citation Format: Liliane Robillard, Minh Nguyen, Thomas C. Harding, Andrew D. Simmons. In vitro and in vivo assessment of the mechanism of action of the PARP inhibitor rucaparib [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 2475. doi:10.1158/1538-7445.AM2017-2475

Abstract 2807: Developing a variety of primary mouse tumor allograft models for evaluating efficacy of PARP inhibitor

Abstract Recent success of multiple poly (adenosine diphosphate [ADP]) ribose polymerase inhibitors (PARPi) in treating BRCA mutant ovarian cancers has fueled the development of PARPi in several areas, including expanding its use in other BRCA mutant or wildtype cancers; exploring resistant mechanisms and overcoming resistance; seeking for its combination with chemotherapy, radiotherapy and emerging immunotherapy to achieve persistent efficacy. Especially considering the mechanism of action of PARPi, its use can increase the mutational loads in tumors with defects in homologous recombination and DNA double strand break repair, which potentially would enhance the immunogenicity of the tumors. Therefore it would be very interesting to test the effectiveness of combining PARPi with anti-PD1/PD-L1 or anti-CTLA4. However, relevant animal models are lacking for such studies.Syngeneic mouse tumors offer about 30 viable in vivo models for proof of concept studies. However, these models suffer from several limitations: 1) only a few available and responsive to the current checkpoint inhibitors – for given strains of mouse, there are only very few choices of cells/disease types; 2) not mimicking patient tumors since they are in vitro immortalized cell lines; 3) not reflective of patient disease pathways thus unfit for common target agents, either in mono- or combination therapy. We have established allografts of spontaneous mouse tumors (MuPrime®) derived from genetically engineered mouse models (GEMMs) as a new type of immuno-oncology models with following advantages: 1) its primary nature of “stem cell diseases” and relevant tumor microenvironment; 2) its diverse cancer types/strains of mice; 3) engineered oncogenic drivers as seen in human diseases, deriving from a wide range of available GEMMs, suitable for targeted agents. Thus far, we have built a small library of allografts and are testing them to facilitate pharmacological investigation, particularly those of immuno-oncology.We have found a few models, including an APCmin/+ skin squamous cell carcinoma and a P53 null sarcoma responded well to PARPi Niraparib. Continuous treatment with Niraparib led to development of partial or complete resistance. We are now profiling the resistant models to find out their general mutation loads, as well as specific biomarkers render resistant mechanisms. We are also treating the tumors with mouse PD1 antibody to test whether presumably enhanced mutation loads would sensitize these tumors to immunotherapy. The data of these studies will be presented at the meeting. Citation Format: Gavin Jiagui Qu, Davy Xuesong Ouyang, Li Chen, Likun Zhang, Annie Xiaoyu An, Jie Cai, Henry Q. X. Li. Developing a variety of primary mouse tumor allograft models for evaluating efficacy of PARP inhibitor [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 2807. doi:10.1158/1538-7445.AM2017-2807

Abstract 2580: Ceramide-rubusoside nano-micelles restores p53-mediated tumor suppression in p53R248W mutant ovarian cancer cells

Abstract Ceramide (Cer), a sphingolipid metabolite, has crucial functions in anti-proliferation, cell differentiation and apoptosis. Cellular Cer levels highly correlates to therapeutic efficacy of anticancer drugs, however, the hydrophobicity Cer has limits to develop it as a therapeutic agent. Here, we report that Cer-rubusoside (Cer-RUB) nanomicelles enhanced Cer bioavailability and restored p53-mediated tumor suppression. These nanomicelles significantly increased the levels of NBD-C6-Cer by more than 50-fold in serum, 40-fold in tumor and 70-fold in lung of tumor-bearing mice (1 mg/kg, i.p., 24 hr), respectively. Cer-RUB (3 µM) significantly increased OVCAR-3 (p53 R248W) cell sensitivity to cisplatin, and the IC50 values decreased by 3-fold, even though these values were no significant changes in A2780 (wt p53) cells. Furthermore, Cer-RUB treatments (1 mg/kg, i.p. every 3 days for 24 days) significantly increased cisplatin efficacy in tumor growth-arrest in OVCAR-3 tumor-mice, and the tumor volumes reduced by 52% (1653 vs. 805 mm3) in combination with cisplatin, compared to cisplatin alone. We found that ovarian cancer stem cells (CD44+/CD133+) decreased by 10-fold (0.85 vs. 0.08% of total cells) in OVCAR-3 tumors treated with Cer-RUB combined with cisplatin, as compared to cisplatin alone, respectively. Western blotting showed that Cer-RUB treatments selectively increased the expression levels of phosphorylated p53 (Ser15), BAX and p21 in OVCAR-3 cancer cells and tumors, respectively. These studies indicates that the gain-of-function (GOF) of p53 R248W leads OVCAR-3 cell resistance to cisplatin and tumor growth. Cer-RUB can restore p53 and eliminates the GOF of p53 R248W in cancers. Citation Format: sachin K. khiste, Zhijun Liu, Yong-Yu Liu. Ceramide-rubusoside nano-micelles restores p53-mediated tumor suppression in p53R248W mutant ovarian cancer cells [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 2580. doi:10.1158/1538-7445.AM2017-2580