Jeanette Saskowski

Suberoylanilide hydroxamic acid (SAHA) enhances olaparib activity by targeting homologous recombination DNA repair in ovarian cancer

OBJECTIVES Approximately 50% of serous epithelial ovarian cancers (EOC) contain molecular defects in homologous recombination (HR) DNA repair pathways. Poly(ADP-ribose) polymerase inhibitors (PARPi) have efficacy in HR-deficient, but not in HR-proficient, EOC tumors as a single agent. Our goal was to determine whether the histone deacetylase inhibitor, suberoylanilide hydroxamic acid (SAHA), can sensitize HR-proficient ovarian cancer cells to the PARPi AZD-2281 (olaparib). METHODS Ovarian cancer cell lines (SKOV-3, OVCAR-8, NCI/ADR-Res, UWB1.289 BRCA1null and UWB1.289+BRCA1 wild-type) were treated with saline vehicle, olaparib, SAHA or olaparib/SAHA. Sulforhodamine B (SRB) assessed cytotoxicity and immunofluorescence and Western blot assays assessed markers of apoptosis (cleaved PARP) and DNA damage (pH2AX and RAD51). Drug effects were also tested in SKOV-3 xenografts in Nude mice. Affymetrix microarray experiments were performed in vehicle and SAHA-treated SKOV-3 cells. RESULTS In a microarray analysis, SAHA induced coordinated down-regulation of HR pathway genes, including RAD51 and BRCA1. Nuclear co-expression of RAD51 and pH2AX, a marker of efficient HR repair, was reduced approximately 40% by SAHA treatment alone and combined with olaparib. SAHA combined with olaparib induced apoptosis and pH2AX expression to a greater extent than either drug alone. Olaparib reduced cell viability at increasing concentrations and SAHA enhanced these effects in 4 of 5 cell lines, including BRCA1 null and wild-type cells, in vitro and in SKOV-3 xenografts in vivo. CONCLUSIONS These results provide preclinical rationale for targeting DNA damage response pathways by combining small molecule PARPi with HDACi as a mechanism for reducing HR efficiency in ovarian cancer.

Low ascorbic acid and increased oxidative stress in gulo(-/-) mice during development.

Vitamin C (ascorbic acid, AA) depletion during prenatal and postnatal development can lead to oxidative stress in the developing brain and other organs. Such damage may lead to irreversible effects on later brain function. We studied the relationship between AA deficiency and oxidative stress during development in gulonolactone oxidase (gulo) knockout mice that are unable to synthesize their own ascorbic acid. Heterozygous gulo(+/-) mice can synthesize AA and typically have similar tissue levels to wild-type mice. Gulo(+/-) dams were mated with gulo(+/-) males to provide offspring of each possible genotype. Overall, embryonic day 20 (E20) and postnatal day 1 (P1) pups were protected against oxidative stress by sufficient AA transfer during pregnancy. On postnatal day 10 (P10) AA levels were dramatically lower in liver and cerebellum in gulo(-/-) mice and malondialdehyde (MDA) levels were significantly increased. In postnatal day 18 pups (P18) AA levels decreased further in gulo(-/-) mice and oxidative stress was observed in the accompanying elevations in MDA in liver, and F(2)-isoprostanes in cortex. Further, total glutathione levels were higher in gulo(-/-) mice in cortex, cerebellum and liver, indicating that a compensatory antioxidant system was activated. These data show a direct relationship between AA level and oxidative stress in the gulo(-/-) mice. They reinforce the critical role of ascorbic acid in preventing oxidative stress in the developing brain in animals that, like humans, cannot synthesize their own AA.

Romidepsin (FK228) combined with cisplatin stimulates DNA damage-induced cell death in ovarian cancer

OBJECTIVE Romidepsin (FK228) was recently approved by the FDA for the treatment of cutaneous and peripheral T cell lymphoma. We have shown in vitro efficacy of FK228 in ovarian cancer. Here, our goal was to evaluate FK228 combined with cisplatin in ovarian cancer in vitro and in vivo. METHODS Ovarian cancer cell lines were treated with cisplatin, FK228 or the combination of drugs. Colorimetric assays were used to determine cytotoxicity in vitro. Mice engrafted with 5×10(6) SKOV-3 ovarian cancer cells were treated with cisplatin, FK228 or the combination, and tumor weights and volumes were measured. We assessed molecular markers of proliferation (mib-1), apoptosis (cleaved PARP and cleaved caspase 3) and DNA damage (pH2AX, RAD51 and 53BP1). RESULTS FK228 enhanced the cytotoxic effects of cisplatin in ovarian cells compared to vehicle-treated controls or each drug alone. The combination of FK228 and cisplatin-induced apoptosis and activated aberrant DNA damage responses demonstrated by increased expression of pH2AX, RAD51 and 53BP1. Mice treated with FK228, cisplatin and both drugs showed reduced tumor weights and volumes. Drug-treated tumors showed decreased mib-1 and increased cleaved-caspase 3 expression levels. The number and intensity of pH2AX stained cells was greatest in tumors exposed to the combination of FK228 and cisplatin. CONCLUSION FK228 causes DNA damage-induced apoptosis and enhances the anti-tumor effects of cisplatin. The DNA damage mark pH2AX is activated by FK228 and cisplatin and may be a useful pharmacodynamic mark of these effects.

TR3 Modulates Platinum Resistance in Ovarian Cancer

In metastatic ovarian cancer, resistance to platinum chemotherapy is common. Although the orphan nuclear receptor TR3 (nur77/NR4A1) is implicated in mediating chemotherapy-induced apoptosis in cancer cells, its role in ovarian cancer has not been determined. In an ovarian cancer tissue microarray, TR3 protein expression was elevated in stage I tumors, but downregulated in a significant subset of metastatic tumors. Moreover, TR3 expression was significantly lower in platinum-resistant tumors in patients with metastatic disease, and low TR3 staining was associated with poorer overall and progression-free survival. We have identified a direct role for TR3 in cisplatin-induced apoptosis in ovarian cancer cells. Nucleus-to-cytoplasm translocation of TR3 was observed in cisplatin-sensitive (OVCAR8, OVCAR3, and A2780PAR) but not cisplatin-resistant (NCI/ADR-RES and A2780CP20) ovarian cancer cells. Immunofluorescent analyses showed clear overlap between TR3 and mitochondrial Hsp60 in cisplatin-treated cells, which was associated with cytochrome c release. Ovarian cancer cells with stable shRNA- or transient siRNA-mediated TR3 downregulation displayed substantial reduction in cisplatin effects on apoptotic markers and cell growth in vitro and in vivo. Mechanistic studies showed that the cisplatin-induced cytoplasmic TR3 translocation required for apoptosis induction was regulated by JNK activation and inhibition of Akt. Finally, cisplatin resistance was partially overcome by ectopic TR3 overexpression and by treatment with the JNK activator anisomycin and Akt pathway inhibitor, wortmannin. Our results suggest that disruption of TR3 activity, via downregulation or nuclear sequestration, likely contributes to platinum resistance in ovarian cancer. Moreover, we have described a treatment strategy aimed at overcoming platinum resistance by targeting TR3.

Tracking NF-κB activity in tumor cells during ovarian cancer progression in a syngeneic mouse model

BackgroundNuclear factor-kappa B (NF-kappaB) signaling is an important link between inflammation and peritoneal carcinomatosis in human ovarian cancer. Our objective was to track NF-kappaB signaling during ovarian cancer progression in a syngeneic mouse model using tumor cells stably expressing an NF-kappaB reporter.MethodsID8 mouse ovarian cancer cells stably expressing an NF-kappaB-dependent GFP/luciferase (NGL) fusion reporter transgene (ID8-NGL) were generated, and injected intra-peritoneally into C57BL/6 mice. NGL reporter activity in tumors was non-invasively monitored by bioluminescence imaging and measured in luciferase assays in harvested tumors. Ascites fluid or peritoneal lavages were analyzed for inflammatory cell and macrophage content, and for mRNA expression of M1 and M2 macrophage markers by quantitative real-time RT-PCR. 2-tailed Mann-Whitney tests were used for measuring differences between groups in in vivo experiments.ResultsIn ID8-NGL cells, responsiveness of the reporter to NF-kappaB activators and inhibitors was confirmed in vitro and in vivo. ID8-NGL tumors in C57BL/6 mice bore histopathological resemblance to human high-grade serous ovarian cancer and exhibited similar peritoneal disease spread. Tumor NF-kappaB activity, measured by the NGL reporter and by western blot of nuclear p65 expression, was markedly elevated at late stages of ovarian cancer progression. In ascites fluid, macrophages were the predominant inflammatory cell population. There were elevated levels of the M2-like pro-tumor macrophage marker, mannose-receptor, during tumor progression, and reduced levels following NF-kappaB inhibition with thymoquinone.ConclusionsOur ID8-NGL reporter syngeneic model is suitable for investigating changes in tumor NF-kappaB activity during ovarian cancer progression, how NF-kappaB activity influences immune cells in the tumor microenvironment, and effects of NF-kappaB-targeted treatments in future studies.

Panobinostat sensitizes cyclin E high, homologous recombination-proficient ovarian cancer to olaparib

OBJECTIVE Homologous recombination (HR) proficient ovarian cancers, including CCNE1 (cyclin E)-amplified tumors, are resistant to poly (ADP-ribose) polymerase inhibitors (PARPi). Histone deacetylase inhibitors (HDACi) are effective in overcoming tumor resistance to DNA damaging drugs. Our goal was to determine whether panobinostat, a newly FDA-approved HDACi, can sensitize cyclin E, HR-proficient ovarian cancer cells to the PARPi olaparib. METHODS Expression levels of CCNE1 (cyclin E), BRCA1, RAD51 and E2F1 in ovarian tumors and cell lines were extracted from The Cancer Genome Atlas (TCGA) and Broad-Novartis Cancer Cell Line Encyclopedia (CCLE). In HR-proficient ovarian cancer cell line models (OVCAR-3, OVCAR-4, SKOV-3, and UWB1.289+BRCA1 wild-type), cell growth and viability were assessed by sulforhodamine B and xenograft assays. DNA damage and repair (pH2AX and RAD51 co-localization and DRGFP reporter activity) and apoptosis (cleaved PARP and cleaved caspase-3) were assessed by immunofluorescence and Western blot assays. RESULTS TCGA and CCLE data revealed positive correlations (Spearman) between cyclin E E2F1, and E2F1 gene targets related to DNA repair (BRCA1 and RAD51). Panobinostat downregulated cyclin E and HR repair pathway genes, and reduced HR efficiency in cyclin E-amplified OVCAR-3 cells. Further, panobinostat synergized with olaparib in reducing cell growth and viability in HR-proficient cells. Similar co-operative effects were observed in xenografts, and on pharmacodynamic markers of HR repair, DNA damage and apoptosis. CONCLUSIONS These results provide preclinical rationale for using HDACi to reduce HR in cyclin E-overexpressing and other types of HR-proficient ovarian cancer as a means of enhancing PARPi activity.

Abstract 5056: The bromodomain inhibitor JQ1 sensitizes homologous recombination proficient ovarian cancer cells to the PARP inhibitor olaparib

Introduction: Lack of effective treatment options for high-grade serous ovarian cancers (HGSOC) retaining functional homologous recombination (HR) DNA repair pathways is a significant clinical problem. HR-proficient HGSOC tumors, for example those harboring cyclin E amplifications, have poorer clinical outcomes and show relative resistance to DNA-damaging platinum agents and newer poly ADP ribose polymerase inhibitors (PARPi). We and others have shown that using epigenetic drugs to reduce HR efficiency in HR-proficient HGSOC sensitizes these cancer cells to DNA damaging agents. One mechanism by which these drugs reduce HR efficiency is by transcriptional down-regulation of HR pathway components. An emerging class of epigenetic mediators of pro-tumorigenic transcription is the bromodomain (BRD) family of proteins, and BRD inhibitors (BRDi) have shown promising preclinical anti-tumor efficacy. However, it is unknown whether BRDi sensitize HR-proficient HGSOC to DNA damaging agents. Aims: To test the hypothesis that BRDi decrease efficiency of HR DNA repair in HR-proficient ovarian cancer cells, thereby sensitizing them to PARPi. Methods: The HR-proficient ovarian cancer cell lines, OVCAR-3 (cyclin E-amplified) and SKOV3, were treated with 0.01% DMSO vehicle, the PARPi olaparib (Astra Zeneca), the BRDi JQ1 or with the olaparib/JQ1 combination. Sulforhodamine B (SRB) assays assessed cell growth and viability (72 hours treatment). Immunofluorescence (IF) assays assessed markers of DNA damage (pH2AX), apoptosis (cleaved caspase-3), and HR efficiency (RAD51 foci, and GFP expression in cells co-transfected with I-Sce1 endonuclease and DRGFP HR reporter plasmids) (24 hours treatment). Steady state levels of the HR protein BRCA1, pH2AX and cleaved caspase-3 were assessed by western blot (24 hours treatment). Results: The combination of JQ1 and olaparib synergistically reduced cancer cell viability following isobologram analyses of SRB experiments. Compared to either drug alone, the JQ1/olaparib combination also significantly reduced BRCA1 expression and increased protein levels of cleaved caspase-3 and pH2AX in western blots, and also increased the number of cells displaying DNA damage and apoptosis in IF assays. Finally, JQ1 and olaparib combined to significantly reduce HR efficiency in our RAD51 foci formation and DRGFP assays compared to olaparib alone. Conclusions: Our results suggest that BRDi sensitize HR-proficient cells to DNA damaging drugs, in part by reducing efficiency of HR DNA repair. These findings have important implications for expanding the use of PARPi in HR-proficient HGSOC through rational combinations with epigenetic drugs such as BRDi that target the HR pathway. Citation Format: Andrew J. Wilson, Janese Thompson, Abdirahman Osman, Jeanette Saskowski, Dineo Khabele. The bromodomain inhibitor JQ1 sensitizes homologous recombination proficient ovarian cancer cells to the PARP inhibitor olaparib [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 5056. doi:10.1158/1538-7445.AM2017-5056

Abstract A15: The histone deacetylase inhibitor panobinostat sensitizes cyclin E-amplified ovarian cancer cells to poly ADP ribose polymerase inhibitors via E2F1 downregulation.

Background: The lack of treatment options for chemoresistant ovarian cancer, the deadliest gynecologic malignancy, is a central clinical problem. Approximately 50% of high-grade serous ovarian tumors with defects in DNA repair pathways including germline BRCA1 and 2 mutations (the so-called BRCAness phenotype), and are sensitive to DNA damaging agents such as cisplatin and poly ADP ribose polymerase inhibitors (PARPi). In contrast, non-BRCAness tumors such as those exhibiting CCNE1 (cyclin E) amplification are chemoresistant, show upregulated DNA repair processes and cell cycle progression via activation of the CDK2/cyclin E/E2F1 axis, and have poor clinical outcomes. Targeting the cyclin E axis with CDK inhibitors has been unsuccessful in part due to chemoresistance through upregulation of oncogenic E2F1. An area of urgent need is to develop new treatments for these intractable subtypes of non-BRCAness ovarian cancer. To that end, our group has shown that epigenetic histone deacetylase inhibitor (HDACi) therapy sensitizes ovarian cancer cells to various chemotherapeutic drugs, including PARPi, through reduction of expression of DNA repair genes. Aims: The objective of this study was to (i) determine expression patterns of cyclin E axis components in publically available databases of high-grade serous ovarian tumors and established cell lines; and (ii) test an epigenetic drug strategy using HDACi to convert non-BRCAness ovarian cancer cells to a BRCAness phenotype as a means of sensitizing chemoresistant cells to PARPi. Methods: RNA-seq V2 RSEM expression levels of cyclin E, CDK2, BRCA1 and E2F1 in 265 ovarian tumors were extracted from The Cancer Genome Atlas (TCGA) via cbioportal.org. mRNA expression of these genes in ovarian cancer cell lines was extracted from the Broad-Novartis Cancer Cell Line Encyclopedia (CCLE). Preliminary western blot studies indicated that of the FDA-approved HDACi (vorinostat, romidepsin, panobinostat), panobinostat was most effective at reducing protein expression of cyclin E axis components (cyclin E, BRCA1 and E2F1). We therefore treated cultured cyclin E overexpressing ovarian cancer cells via CCNE1 amplification (OVCAR-3) or high-level gain (OVCAR-4) with panobinostat alone and in combination with the PARPi, olaparib. The following endpoints were examined: cell viability in sulforhodamine B (SRB) assays, with drug interactions tested for synergism by isobologram analysis; DNA damage by immunofluorescence analysis of the double-strand DNA break marker, pH2AX(ser139); apoptosis by immunofluorescence and western blot analysis of cleaved PARP and cleaved caspase-3. Two-tailed Student9s t tests were used for measuring differences between groups in drug treatment experiments. Results: There was significant correlation (Spearman) between cyclin E expression and that of CDK2, E2F1 and BRCA1 in ovarian tumors and in serous-like ovarian cancer cell lines. These results are consistent with the relationship between cyclin E overexpression and enhanced E2F1-dependent transcription of established E2F1 targets, including cyclin E itself, and DNA repair genes such as BRCA1. In SRB assays, the combination of panobinostat and olaparib synergized in both OVCAR-3 and OVCAR-4 cells. When combined, the drugs also induced co-operative effects on expression of pH2AX(ser139) and apoptosis markers. Enhanced cytotoxicity of the combination was accompanied by a panobinostat-induced reduction in expression of cyclin E, E2F1 and BRCA1. A key role for E2F1 downregulation in mediating this BRCAness shift was suggested by the observation that E2F1 knockdown with siRNA synergized with panobinostat in reducing BRCA1 expression and inducing apoptosis. Conclusions: The development of a new PARPi combination therapy with panobinostat has immediate prospects for rapid translation to the clinic and great potential for improving clinical outcomes for non-BRCAness, chemoresistant ovarian cancer. Citation Format: Andrew J. Wilson, Jeanette Saskowski, Dineo Khabele. The histone deacetylase inhibitor panobinostat sensitizes cyclin E-amplified ovarian cancer cells to poly ADP ribose polymerase inhibitors via E2F1 downregulation. [abstract]. In: Proceedings of the AACR Special Conference on Advances in Ovarian Cancer Research: Exploiting Vulnerabilities; Oct 17-20, 2015; Orlando, FL. Philadelphia (PA): AACR; Clin Cancer Res 2016;22(2 Suppl):Abstract nr A15.

Abstract B81: The histone deacetylase inhibitor panobinostat sensitizes cyclin E-amplified ovarian cancer cells to the cyclin-dependent kinase 2 inhibitor dinaciclib.

Background: The lack of treatment options for chemoresistant ovarian cancer, the deadliest gynecologic malignancy, is a central clinical problem. Approximately 50% of high-grade serous ovarian tumors harbor defects in DNA repair pathways including germline BRCA1 and 2 mutations (a BRCAness phenotype), and are sensitive to DNA damaging agents such as cisplatin and poly ADP ribose polymerase inhibitors (PARPi). In contrast, non-BRCAness tumors such as those exhibiting CCNE1 (cyclin E) amplification are chemoresistant, show upregulated DNA repair processes and cell cycle progression via activation of the CDK2/cyclin E/E2F1 axis, and have poor clinical outcomes. Targeting the cyclin E axis with CDK2 inhibitors such as dinaciclib is a promising clinical strategy. However, their efficacy in cyclin E-overexpressing ovarian cancer cells is limited by acquired chemoresistance mediated in part through upregulation of CDK2, cyclin E and E2F1, and toxicity. Therefore, new strategies are needed to improve the efficacy of CDK2-targeting therapy in ovarian cancer. Our group has shown that epigenetic histone deacetylase inhibitor (HDACi) therapy sensitizes ovarian cancer cells to multiple chemotherapeutic drugs. Aims: The objectives of this study were to (i) determine expression patterns of cyclin E axis components in publically available databases of high-grade serous ovarian tumors and established cell lines; and (ii) test an epigenetic drug strategy using HDACi to convert non-BRCAness ovarian cancer cells to a BRCAness phenotype as a means of sensitizing chemoresistant cells to dinaciclib. Methods: RNA-seq V2 RSEM expression levels of cyclin E, CDK2, BRCA1 and E2F1 in 265 ovarian tumors were extracted from The Cancer Genome Atlas (TCGA) via cbioportal.org. mRNA expression of these genes in ovarian cancer cell lines was extracted from the Broad-Novartis Cancer Cell Line Encyclopedia (CCLE). Preliminary western blot studies indicated that of the FDA-approved HDACi (vorinostat, romidepsin, panobinostat), panobinostat was most effective at reducing protein expression of cyclin E axis components (cyclin E, BRCA1 and E2F1). We therefore treated cultured cyclin E overexpressing ovarian cancer cells via CCNE1 amplification (OVCAR-3) or high-level gain (OVCAR-4) with panobinostat alone and in combination with the CDK2 inhibitor, dinaciclib. The following endpoints were examined: cell viability in sulforhodamine B (SRB) assays; apoptosis by western blot analysis of cleaved PARP; and protein expression of cyclin E, E2F1 and BRCA1 by western blot. Two-tailed Student9s t tests were used for measuring differences between groups in drug treatment experiments. Results: There was significant correlation (Spearman) between cyclin E expression and that of CDK2, E2F1 and BRCA1 in ovarian tumors and in serous-like ovarian cancer cell lines. These results are consistent with the relationship between cyclin E overexpression and enhanced E2F1-dependent transcription of established E2F1 targets, including cyclin E itself, and DNA repair genes such as BRCA1. In SRB assays, the combination of panobinostat and dinaciclib synergized in both OVCAR-3 and OVCAR-4 cells. When combined, the drugs also induced co-operative effects on expression of apoptosis markers. Enhanced cytotoxicity of the combination was accompanied by panobinostat-induced reduction in expression of cyclin E, E2F1 and BRCA1. Conclusions: The development of a new CDK2 inhibitor combination therapy with panobinostat has immediate prospects for rapid translation to the clinic and great potential for improving clinical outcomes for non-BRCAness, chemoresistant ovarian cancer. Citation Format: Andrew J. Wilson, Jeanette Saskowski, Dineo Khabele. The histone deacetylase inhibitor panobinostat sensitizes cyclin E-amplified ovarian cancer cells to the cyclin-dependent kinase 2 inhibitor dinaciclib. [abstract]. In: Proceedings of the AACR Special Conference on Advances in Ovarian Cancer Research: Exploiting Vulnerabilities; Oct 17-20, 2015; Orlando, FL. Philadelphia (PA): AACR; Clin Cancer Res 2016;22(2 Suppl):Abstract nr B81.

Abstract 4730: The histone deacetylase inhibitor panobinostat sensitizes cyclin E-amplified ovarian cancer cells to poly ADP-ribose polymerase inhibitor olaparib

Objective: Ovarian cancer is the second most common gynecological cancer and the number one cause of cancer death in gynecological cancers. In the United States alone, there are about 21,290 expected new cases in 2015 and about 14,180 expected deaths. Approximately 95% of ovarian malignancies are epithelial ovarian cancer (EOC). Most (50%) of these EOC tumors have BRCAness features - contain molecular defect in homologous recombination (HR) DNA repair pathways. About 20% of the non-BRACness subtypes of EOC exhibit increased expression of cyclin E (CCNE1) which is associated with resistance to DNA damaging drugs and increased mortality. A first-in-class drug, a poly(ADP-ribose) polymerase inhibitor (PARPi) olaparib was approved by the FDA for the treatment of advanced ovarian cancer patients with BRCA mutations who have had three or more lines chemotherapy. PARPi efficacy is lower in non-BRCAness tumors but may be enhanced by other drug combinations. Our group has shown that epigenetic histone deacetylase inhibitor (HDACi) therapy sensitizes ovarian cancer cells to various chemotherapeutic drugs. Our goal is to expand olaparib therapy to patients with chemoresistant tumors by sensitizing CCNE1 amplified EOC with the potent HDACi, panobinostat. Method: Ovarian cancer cell lines used were OVCAR-3 (CCNE1-amplified) and SKOV3 (CCNE1-non-amplified). They were co-treated with 0.01% DMSO vehicle, olaparib, panobinostat or olaparib/panobinostat combination. Sulforhodamine B (SRB) assays assessed cytotoxicity and immunofluorescence (IF) assessed markers of apoptosis (cleaved caspase-3). Images were analyzed using the Adobe Photoshop counting tool. Data analysis was performed via the Microsoft Excel average and percentage functions. Figures and p-value analysis were created using GraphPad Prism. Results: Our previous studies have demonstrated through western blot analysis of cleaved PARP expression that panobinostat enhances the pro-apoptotic effect of olaparib in CCNE1 amplified OVCAR-3 ovarian cancer cells. In this current study, IF was used to assess the established marker of apoptosis, cleaved caspase-3, in OVCAR3 and SKOV3 cells. Compared to vehicle-treated cells, olaparib alone induced no difference in the number of cleaved caspase-3 expression in both SKOV3 and OVCAR3 cells. However, there was significant upregulation in the percentage of cells positive for cleaved caspase-3 expression in combination treatments compared to single drug treatments in both cell lines. Conclusion: The response of ovarian cancer cells to the PARPi olaparib is enhanced by co-treatment with HDACi panobinostat in vitro. This indicates that, responses to olaparib in HR-proficient EOC can be improved by combination therapy with panobinostat. Thus we have identified a novel way to expand the use of olaparib for the treatment of advanced ovarian cancer to patients with non-BRCA tumors. Citation Format: Kofi Sarfo-Kantanka, Andrew J. Wilson, Alexandra Steck, Jeanette Saskowski, Dineo Khabele. The histone deacetylase inhibitor panobinostat sensitizes cyclin E-amplified ovarian cancer cells to poly ADP-ribose polymerase inhibitor olaparib. [abstract]. In: Proceedings of the 107th Annual Meeting of the American Association for Cancer Research; 2016 Apr 16-20; New Orleans, LA. Philadelphia (PA): AACR; Cancer Res 2016;76(14 Suppl):Abstract nr 4730.