Combining EHMT and PARP Inhibition: A Strategy to Diminish Therapy-Resistant Ovarian Cancer Tumor Growth while Stimulating Immune Activation.

Despite the success of poly-ADP-ribose polymerase inhibitors (PARPi) in the clinic, high rates of resistance to PARPi presents a challenge in the treatment of ovarian cancer, thus it is imperative to find therapeutic strategies to combat PARPi resistance. Here, we demonstrate that inhibition of epigenetic modifiers euchromatic histone lysine methyltransferases 1/2 (EHMT1/2) reduces the growth of multiple PARPi-resistant ovarian cancer cell lines and tumor growth in a PARPi-resistant mouse model of ovarian cancer. We found that combinatory EHMT and PARP inhibition increases immunostimulatory double-stranded RNA formation and elicits several immune signaling pathways in vitro. Using epigenomic profiling and transcriptomics, we found that EHMT2 is bound to transposable elements, and that EHMT inhibition leads to genome-wide epigenetic and transcriptional derepression of transposable elements. We validated EHMT-mediated activation of immune signaling and upregulation of transposable element transcripts in patient-derived, therapy-naïve, primary ovarian tumors, suggesting potential efficacy in PARPi-sensitive disease as well. Importantly, using multispectral immunohistochemistry, we discovered that combinatory therapy increased CD8 T-cell activity in the tumor microenvironment of the same patient-derived tissues. In a PARPi-resistant syngeneic murine model, EHMT and PARP inhibition combination inhibited tumor progression and increased Granzyme B+ cells in the tumor. Together, our results provide evidence that combinatory EHMT and PARP inhibition stimulates a cell autologous immune response in vitro, is an effective therapy to reduce PARPi-resistant ovarian tumor growth in vivo, and promotes antitumor immunity activity in the tumor microenvironment of patient-derived ex vivo tissues of ovarian cancer.

Combining EHMT and PARP Inhibition: A Strategy to Diminish Therapy-Resistant Ovarian Cancer Tumor Growth while Stimulating Immune Activation.

Despite the success of Poly-ADP-ribose polymerase inhibitors (PARPi) in the clinic, high rates of resistance to PARPi presents a challenge in the treatment of ovarian cancer, thus it is imperative to find therapeutic strategies to combat PARPi resistance. Here, we demonstrate that inhibition of epigenetic modifiers Euchromatic histone lysine methyltransferases 1/2 (EHMT1/2) reduces the growth of multiple PARPi-resistant ovarian cancer cell lines and tumor growth in a PARPi-resistant mouse model of ovarian cancer. We found that combinatory EHMT and PARP inhibition increases immunostimulatory dsRNA formation and elicits several immune signaling pathways in vitro. Using epigenomic profiling and transcriptomics, we found that EHMT2 is bound to transposable elements, and that EHMT inhibition leads to genome-wide epigenetic and transcriptional derepression of transposable elements. We validated EHMT-mediated activation of immune signaling and upregulation of transposable element transcripts in patient-derived, therapy-naïve, primary ovarian tumors, suggesting potential efficacy in PARPi-sensitive disease as well. Importantly, using multispectral immunohistochemistry, we discovered that combinatory therapy increased CD8 T cell activity in the tumor microenvironment of the same patient-derived tissues. In a PARPi-resistant syngeneic murine model, EHMT and PARP inhibition combination inhibited tumor progression and increased Granzyme B+ cells in the tumor. Together, our results provide evidence that combinatory EHMT and PARP inhibition stimulates a cell autologous immune response in vitro, is an effective therapy to reduce PARPi resistant ovarian tumor growth in vivo, and promotes anti-tumor immunity activity in the tumor microenvironment of patient-derived ex vivo tissues of ovarian cancer.

Multi-Institutional Study of Referral Patterns for Gynecologic Oncology Consultation.

PURPOSE: Evaluation by a gynecologic oncologist (GO) is associated with improved clinical outcomes for patients with gynecologic cancers, yet little is known about health care factors that influence patients' referrals to GO. METHODS: Medical records of 50 consecutive new patients seen in GO clinics at each of six referral centers across the United States were reviewed. Patient and disease characteristics were collected along with referral indication, evaluation and referral dates, diagnostic procedures, provider specialties, and zone improvement plan (ZIP) code of up to three referring providers per patient. The primary outcome was interval between first evaluation and referral. Univariate associations were evaluated with Chi-square and Wilcoxon rank-sum tests and multivariable associations with negative binomial regression models. Secondary outcome was prolonged time to GO referral, defined as greater than the 75th percentile. Logistic regression was used for multivariable modeling. RESULTS: Three hundred patient records were analyzed. The median time from first health care encounter to referral was 15 days (IQR, 5-43). The mean distance from residence to GO was 39.8 miles (standard deviation, 53.8). Seventy-one percent of GO referrals were initiated by obstetrician-gynecologists, 9% by family physicians, and 6% internists. Presentation-to-referral interval was 76% shorter for patients evaluated by an emergency medicine clinician (exp(Beta), 0.24; 95% CI, 0.11 to 0.53; P < .001). Public insurance was associated with 1.47 times longer time to referral compared with private insurance (exp(Beta), 1.47; 95% CI, 1.05 to 2.04; P = .001). Residents of nonmetropolitan ZIP codes were less likely to have prolonged time to referral (odds ratio [OR], 0.288; P = .017). Distance from residence to GO (per 10 miles) increased the likelihood of prolonged time to referral (OR, 1.10; P = .010). CONCLUSION: Interventions are needed to improve recognition and referral of patients for gynecologic oncology evaluation. Community outreach and engagement with obstetrician-gynecologists should be prioritized to improve times to referral.

Targeting BRPF3 moderately reverses olaparib resistance in high grade serous ovarian carcinoma.

PARP inhibitors (PARPi) kill cancer cells by stalling DNA replication and preventing DNA repair, resulting in a critical accumulation of DNA damage. Resistance to PARPi is a growing clinical problem in the treatment of high grade serous ovarian carcinoma (HGSOC). Acetylation of histone H3 lysine 14 (H3K14ac) and associated histone acetyltransferases (HATs) and epigenetic readers have known functions in DNA repair and replication. Our objectives are to examine their expression and activities in the context of PARPi-resistant HGSOC, and to determine if targeting H3K14ac or associated proteins has therapeutic potential. Using mass spectrometry profiling of histone modifications, we observed increased H3K14ac enrichment in PARPi-resistant HGSOC cells relative to isogenic PARPi-sensitive lines. By reverse-transcriptase quantitative PCR and RNA-seq, we also observed altered expression of numerous HATs in PARPi-resistant HGSOC cells and a PARPi-resistant PDX model. Knockdown of HATs only modestly altered PARPi response, although knockdown and inhibition of PCAF significantly increased resistance. Pharmacologic inhibition of HBO1 depleted H3K14ac but did not affect PARPi response. However, knockdown and inhibition of BRPF3, a bromodomain and PHD-finger containing protein that is known to interact in a complex with HBO1, did reduce PARPi resistance. This study demonstrates that depletion of H3K14ac does not affect PARPi response in HGSOC. Our data suggest that the bromodomain function of HAT proteins, such as PCAF, or accessory proteins, such as BRPF3, may play a more direct role compared to direct HATs function in PARPi response.

Combination CDC-like kinase inhibition (CLK)/Dual-specificity tyrosine-regulated kinase (DYRK) and taxane therapy in CTNNB1-mutated endometrial cancer.

SM08502 (cirtuvivint) is a novel pan CDC-like kinase (CLK) and Dual specificity tyrosine kinase (DYRK) inhibitor that targets mRNA splicing and is optimized for Wnt pathway inhibition. Previous evaluation of single agent CLK/DYRK inhibition (SM04690) demonstrated inhibition of tumor progression and ß-catenin/TCF transcriptional activity in CTNNB1-mutant endometrial cancer (EC). In-vitro analysis of SM08502 similarly decreases Wnt transcriptional activity and cellular proliferation while increasing cellular apoptosis. SM08502 is an active single-agent therapy with IC50's in the nanomolar range for all EC cell lines evaluated. Combination of SM08502 with paclitaxel has synergistic effect in vitro, as demonstrated by Combination Index <1, and inhibits tumor progression in four endometrial cancer models (HEC265, Ishikawa, Ishikawa-S33Y, and SNGM). In our in vivo mouse models, Ishikawa demonstrated significantly lower tumor volumes of combination vs SM08502 alone (Repeated Measures one-way ANOVA, p = 0.04), but not vs paclitaxel alone. HEC265, SNGM, and Ishikawa-S33Y tumors all had significantly lower tumor volumes with combination SM08502 and paclitaxel compared to single-agent paclitaxel (Repeated Measures one-way ANOVA, p = 0.01, 0.004, and 0.0008, respectively) or single-agent SM08502 (Repeated Measures one-way ANOVA, p = 0.002, 0.005, and 0.01, respectively) alone. Mechanistically, treatment with SM08502 increases alternative splicing (AS) events compared to treatment with paclitaxel. AS regulation is an important post-transcriptional mechanism associated with the oncogenic process in many cancers, including EC. Results from these studies have led to a Phase I evaluation of this combination in recurrent EC.

Combinatory EHMT and PARP inhibition induces an interferon response and a CD8 T cell-dependent tumor regression in PARP inhibitor-resistant models.

Euchromatic histone lysine methyltransferases 1 and 2 (EHMT1/2), which catalyze demethylation of histone H3 lysine 9 (H3K9me2), contribute to tumorigenesis and therapy resistance through unknown mechanisms of action. In ovarian cancer, EHMT1/2 and H3K9me2 are directly linked to acquired resistance to poly-ADP-ribose polymerase (PARP) inhibitors and are correlated with poor clinical outcomes. Using a combination of experimental and bioinformatic analyses in several PARP inhibitor resistant ovarian cancer models, we demonstrate that combinatory inhibition of EHMT and PARP is effective in treating PARP inhibitor resistant ovarian cancers. Our in vitro studies show that combinatory therapy reactivates transposable elements, increases immunostimulatory dsRNA formation, and elicits several immune signaling pathways. Our in vivo studies show that both single inhibition of EHMT and combinatory inhibition of EHMT and PARP reduces tumor burden, and that this reduction is dependent on CD8 T cells. Together, our results uncover a direct mechanism by which EHMT inhibition helps to overcome PARP inhibitor resistance and shows how an epigenetic therapy can be used to enhance anti-tumor immunity and address therapy resistance.

Pathologic distribution at the time of interval tumor reductive surgery informs personalized surgery for high-grade ovarian cancer.

INTRODUCTION: The surgical approach for interval debulking surgery after neoadjuvant chemotherapy has been extrapolated from primary tumor reductive surgery for high-grade ovarian cancer. The study objective was to compare pathologic distribution of malignancy at interval debulking surgery versus primary tumor reductive surgery. METHODS: Patients with a diagnosis of high-grade serous or mixed, non-mucinous, epithelial ovarian, fallopian tube or primary peritoneal cancer who underwent neoadjuvant chemotherapy or primary tumor reductive surgery and had at least 6 months of follow-up were identified through tumor registry at a single institution from January 1995 to April 2016. Pathologic involvement of organs was categorized as macroscopic, microscopic, or no tumor. Statistical analyses included Mann-Whitney and Fisher's exact tests. RESULTS: Of 918 patients identified, 366 (39.9%) patients underwent interval debulking surgery and 552 (60.1%) patients underwent primary tumor reductive surgery. Median age was 62.3 years (range 25.3-92.5). The majority of patients in the interval debulking surgery group were unstaged (261, 71.5%). In the patients who had a primary tumor reductive surgery, 406 (74.6%) had stage III disease. In both groups, the majority of patients had serous histology: 325 (90%) and 435 (78.8%) in the interval debulking and primary tumor reductive surgery groups, respectively. There was a statistically significant difference between disease distribution on the uterus between the groups; 31.4% of the patients undergoing interval debulking surgery had no evidence of uterine disease compared with 22.1% of primary tumor reductive surgery specimens (p<0.001). In the adnexa, there was macroscopic disease present in 253 (69.2%) and 482 (87.4%) of cases in the interval vs primary surgery groups, respectively (p<0.001). Within the omentum, no tumor was present in the omentum in 52 (14.2%) in the interval surgery group versus 91 (16.5%) in the primary surgery group (p<0.001). In the interval surgery group, there was no tumor involving the small and large bowel in 49 (13.4%) and 28 (7.7%) pathologic specimens, respectively. This was statistically significantly different from the small and large bowel in the primary surgery group, of which there was no tumor in 20 (3.6%, p<0.001) and 16 (2.9%, p<0.001) of cases, respectively. CONCLUSION: In patients undergoing interval debulking surgery, there was less macroscopic involvement of tumor in the uterus, adnexa and bowel compared with patients undergoing primary cytoreductive surgery.