Hendrickson, Andrea E. Wahner

Abstract Background: The combination of cediranib (ced) and olaparib (olap) improves progression-free survival and overall response rates (ORR) in women with recurrent plat sensitive high-grade serous (HGS) ovarian cancer compared to olap alone. In this Phase 2 study of ced/olap in women with relapsed ovarian cancer (NCT02345265), we observed an ORR of 74% with ced/olap in plat sensitive relapsed patients (pts) and an ORR of 20% in plat resistant pts. BROCA-HR was performed to characterize tumor HRD status and to correlate this with the activity of ced/olap in the plat sensitive and resistant settings. Methods: Pts with relapsed ovarian cancer across 11 centers were enrolled to 2 cohorts, either plat sensitive or resistant. All pts received ced 30mg daily and olap tablets 200mg BID. Archival, pre-treatment, and on-treatment biopsies were collected on all pts where safely feasible; pts also had the option of undergoing a post-progression biopsy. A baseline blood sample was collected for germline testing and matched control. Available samples were sequenced with BROCA-HR, and HRD status was determined by (1) presence of an HR-related mutation or (2) an increase in loss of heterozygosity (LOH) score. Results: 72 pts were enrolled, with 70 pts receiving study treatment (35 plat sensitive and 35 plat resistant). BROCA-HR analysis is complete on 60 samples, with 10 samples in final analysis. The concordance of HRD status by presence of an HR-related mutation or by LOH score was relatively low (agreement=36/60=60%, Cohen's Kappa=0.28). In 19 pts with a germline BRCA mutation, 16 were HRD+ by LOH (LOH high), 2 were HRD- by LOH (LOH low), and 1 not quantifiable. 3 pts had a somatic BRCA mutation; 1 was LOH high, 1 LOH low, and 1 not quantifiable. 4 pts had non-BRCA somatic HR-related mutations (3 CDK12; 1 ATR); 3 were LOH high and 1 LOH low. Of the 34 pts without HR-related mutations, 15 were LOH high, 16 were LOH low, and 3 not quantifiable. In 29 plat sensitive pts with BROCA-HR results, 18 were HRD+ by mutation (12 germline BRCA, 3 somatic BRCA, 3 somatic other), and 16 were HRD+ by LOH (LOH high). The ORR in plat sensitive pts by HRD status with available HRD results was: 22/29=75.9% (13/18=72% HRDmut+; 9/11=82% HRDmut-); (12/16=75% LOH high; 6/9= 67% LOH low). In 31 plat resistant pts with BROCA-HR results, 8 were HRD+ by mutation (7 germline BRCA, 0 somatic BRCA, 1 somatic other), and 19 were HRD+ by LOH. The ORR in plat resistant pts by HRD status with available HRD results was: 7/31=22.6% (4/8=50% HRDmut+; 3/23=13% HRDmut-, P=0.05); (7/19=37% LOH high; 0/11= 0% LOH low, P=0.03). Updated results with the full data set are pending. Conclusion: Ced/olap has clinical activity in women with relapsed ovarian cancer, both in the plat sensitive and plat resistant settings. In women with plat sensitive disease, where response rates are high, HRD status by presence of an HR-related mutation or LOH status does not appear to provide significant additional information regarding likelihood of response. In women with plat resistant disease, the presence of HRD by an HR-related mutation or LOH status is correlated with a higher likelihood of response. Citation Format: Joyce F Liu, Su-Chun Cheng, Robert M Wenham, Andrea E Wahner Hendrickson, Deborah K Armstrong, Nancy Chan, David E Cohn, Jung-Min Lee, Richard T Penson, Mihaela Cristea, James Abbruzzese, Koji Matsuo, Alexander B Olawaiye, Jennifer Curtis, Geoffrey Shapiro, Ursula A Matulonis, Elise Kohn, Percy Ivy, Elizabeth M Swisher. Correlation of homologous recombination deficiency (HRD) status by BROCA-HR sequencing with response to combination cediranib and olaparib in relapsed ovarian cancer [abstract]. In: Proceedings of the AACR-NCI-EORTC International Conference on Molecular Targets and Cancer Therapeutics; 2019 Oct 26-30; Boston, MA. Philadelphia (PA): AACR; Mol Cancer Ther 2019;18(12 Suppl):Abstract nr A074. doi:10.1158/1535-7163.TARG-19-A074

5546 Background: CHFR, an E3 ubiquitin ligase that regulates Aurora A and Pololike Kinase 1, plays a critical role in the cellular response to mitotic stress. In particular, cells containing CHFR arrest in G2 when microtubule dynamics are disrupted. Conversely, low CHFR expression is associated with hypersensitivity to mitotic spindle poisons in breast cancer cell lines. Despite the extensive use of taxanes in front line treatment of epithelial ovarian cancer (EOC), little is known about CHFR expression in this disease. We examined CHFR expression and its relationship with clinical characteristics and outcome in women with EOC. Methods: Pretreatment EOC samples from women enrolled in the Mayo Clinic Biospecimen Resource for Ovarian Cancer Research who underwent initial surgical debulking between 1999-2009 were stained in triplicate with anti-CHFR antibodies. Samples were scored in a blinded fashion for CHFR expression. Associations between CHFR staining and histology, grade, and stage were assessed using chi-squared tests; associations between CHFR and overall survival (OS) or time to disease recurrence (TTR) were assessed using Cox proportional hazards models. Results: Samples from 354 women were stained; median age at diagnosis was 60.5 (range 21-93). At a median follow-up of 67 months, 221 women (62%) had died. Across all patients, moderate/strong CHFR staining was associated with poor OS (HR = 1.39, 95% CI: 1.07-1.81, p = 0.015). Moreover, moderate/strong CHFR expression was strongly associated with serous histology (p < 0.0001), high grade (p < 0.0001) and advanced stage (p = 0.0002). To assess whether the association between CHFR expression and OS was related to taxane sensitivity, a subgroup analysis was performed in the subset of patients (N=131) with high-grade serous EOC who received initial platinum/taxane chemotherapy. Moderate/strong CHFR expression failed to correlate with OS (HR=0.91, 95% CI: 0.57-1.45, p = 0.68) or TTR from the start of chemotherapy (HR = 0.85, 95% CI: 0.55-1.31, p = 0.55) in this subset. Conclusions: CHFR expression is associated with poor OS and adverse clinical characteristics in patients with EOC but does not appear to be related to taxane sensitivity in high-grade serous tumors.

Abstract BRCA1 plays a key role in homologous recombination (HR) DNA repair. Accordingly, changes that downregulate BRCA1, including BRCA1 mutations and reduced BRCA1 transcription, due to promoter hypermethylation or loss of the BRCA1 transcriptional regulator CDK12, disrupt HR in multiple cancers. In addition, BRCA1 has also been implicated in the regulation of metabolism. Here, we show that reducing BRCA1 expression, either by CDK12 or BRCA1 depletion, led to metabolic reprogramming of ovarian cancer cells, causing decreased mitochondrial respiration and reduced ATP levels. BRCA1 depletion drove this reprogramming by upregulating nicotinamide N-methyltransferase (NNMT). Notably, the metabolic alterations caused by BRCA1 depletion and NNMT upregulation sensitized ovarian cancer cells to agents that inhibit mitochondrial metabolism (VLX600 and tigecycline) and to agents that inhibit glucose import (WZB117). These observations suggest that inhibition of energy metabolism may be a potential strategy to selectively target BRCA1-deficient high-grade serous ovarian cancer, which is characterized by frequent BRCA1 loss and NNMT overexpression. Significance: Loss of BRCA1 reprograms metabolism, creating a therapeutically targetable vulnerability in ovarian cancer.

Abstract Abstract 970 The mammalian target of rapamycin, mTOR, is a highly conserved serine/threonine kinase known to play a role in regulating mRNA translation, cell cycle progression, cell proliferation and apoptosis. As a downstream effector of the phosphatidylinositol 3-kinase (PI3K)/Akt signaling pathway, mTOR is a component of two distinct complexes, TORC1 and TORC2. While TORC1 facilitates cell cycle progression from G1 into S phase by phosphorylating p70S6 kinase and eukaryotic initiation factor 4E binding protein 1 (4E-BP1), TORC2 catalyzes the activating phosphorylation of Akt on Ser473, providing a feedback loop for further activation of mTOR. Phase II trials have shown activity of the TORC1-selective inhibitor rapamycin and its analogs in a wide range of lymphoma subtypes. The purpose of this study was to evaluate the anti-proliferative and pro-apoptotic effects of the dual TORC1/TORC2 inhibitor OSI-027 in human neoplastic lymphoid cells in vitro. MTS assays demonstrated that OSI-027 inhibited proliferation in a wide range of lymphoid lines, including SeAx (Sezary syndrome), DoHH2 (large cell lymphoma), RL (follicular lymphoma) and Jurkat (T cell ALL), as well as clinical lymphoma and T cell ALL samples, with IC50 values ranging from 0.078 to 10 μM. Propidium iodide staining followed by flow cytometry for subdiploid cells revealed induction of apoptosis within 48 h of treatment with OSI-027 (but not rapamycin) in SeAx, DoHH2, and Jurkat cells. Examination of Jurkat variants with alterations in key proteins involved in the death receptor versus mitochondrial pathway revealed diminished apoptotic responses to OSI-027 when Bcl-2 was overexpressed or caspase 9 was silenced, indicating involvement of the mitochondrial pathway. Immunoblotting for Bcl-2 family members revealed upregulation of Bim and Puma after a 48-hour exposure to OSI-027 but not rapamycin. This upregulation was also seen at the mRNA level, with a 12- to 20-fold increase in Puma mRNA and 4- to 12-fold induction of Bim mRNA. Small interfering RNA (siRNA)-mediated knockdown of Bim and Puma significantly diminished the apoptotic response to OSI-027. Because the Foxo3a transcription factor has been implicated in Bim and Puma expression and is known to be activated when Akt is inhibited, we next examined whether Bim and Puma induction was Foxo3a-dependent. Luciferase reporter assays showed that OSI-027 activated the full-length Puma and Bim promoters and that this activation was diminished when the Foxo3a binding sites were deleted or mutated. In addition, OSI-027 induced nuclear translocation of Foxo3a, while Foxo3a siRNA diminished OSI-027-induced apoptosis in Jurkat cells. Collectively, these results indicate that OSI-027 inhibits proliferation and induces apoptosis in a wide range of neoplastic lymphoid cells through a process that involves Foxo3a-mediated upregulation of Bim and Puma. These results also suggest that dual inhibition of TORC1 and TORC2 may be an effective treatment strategy in lymphoid malignancy. Disclosures: Barr: OSI Pharmaceuticals: Employment. Witzig:Novartis and Celgene: Patents & Royalties, Research Funding, Served on advisory boards with Novartis and Celgene – both uncompensated with compensation to Mayo Clinic.

5518 Background: Platinum resistant ovarian cancer (OC) remains a therapeutic challenge. High grade serous OC (HGSOC) harbors TP53 mutations leading to increased dependency on S- and G2-phase checkpoints. Wee1 inhibition with Adavosertib (AZD 1775) (A) induces G2 checkpoint escape. Gemcitabine (G) is an antimetabolite therapy and blocks the progression of cells through the G1/S phase. We hypothesized that combining G+A would be synergistic and overcome resistance. Methods: We conducted a multicentre double-blind 2:1 randomized phase 2 trial to assess the progression free survival (PFS) in women with recurrent platinum-resistant/refractory HGSOC receiving G+A or G+placebo (P) (NCT02151292). Eligibility required measurable disease and feasibility of paired tumor biopsies; no limitation in prior lines of therapy. Non HGSOC histologic subtypes were enrolled in a separate non-randomized exploratory cohort. A/P was given orally at 175mg QD on D1-2, D8-9 and D15-16 with G 1000mg/m² IV D1, D8 and D15 in a 28-day cycle until progression or unacceptable AE. Tumor staging was scheduled every 8 weeks. TP53 mutations were analyzed on archival tissue with Sanger sequencing, TAm-Seq and IHC. TP53 mutation will be also assessed in circulating tumor DNA (ctDNA). Whole exome and RNA sequencing were performed on paired tumor tissues. Results: 124 patients (pts) with median of 3 prior lines of therapy (range 1-10) from 12 centres across Canada and US were enrolled between Sep 2014 to May 2018, with 99 pts randomized (65 in Arm G+A and 34 in G+P). 5 pts were ineligible; 64 pts have died. The median follow-up was 14.3 months. Main related AE was hematologic toxicity (Anemia G≥3: 31% in G+A vs 18% in G+P; Thrombocytopenia G≥3: 31% vs 6%; Neutropenia G≥3: 62% vs 30%). PFS was significantly improved from 3.0 to 4.6 months (HR 0.56 (95%CI: 0.35-0.90, p=0.015 Log rank). There was a significant improvement in overall survival (OS) from 7.2 to 11.5 months (HR 0.56 (95%CI: 0.34-0.92, P=0.022). Partial response by RECIST 1.1 was observed in 13 (21%) and 1 (3%) pts for Arms G+A and G+P, respectively (p=0.02). From the 25 pts in the exploratory cohort, 3 (12%) partial responses were observed. Final results will be reported at the meeting. Conclusion: Addition of adavosertib to gemcitabine in women with platinum resistant/refractory OC improved response rate, PFS and OS with manageable toxicity. Clinical trial information: NCT02151292.

5510 Background: Single agent aromatase inhibitor (AI) therapy is associated with limited clinical activity in ovarian cancer (OC) and endometrial cancers (EC). AI therapy was associated with a progression free survival (PFS) at 12 weeks of only 20% in relapsed OC (Bowman et al, 2002) and a median PFS of 1 month in relapsed EC (Rose et al, 2000). In Estrogen Receptor (ER) positive metastatic breast cancer, clinical studies had shown a significant prolongation of PFS with the addition of the cyclin kinase 4/6 inhibitor ribociclib to AI (Hortobagyi et al, 2016). Here, we report the results of a phase 2 clinical trial of the combination of ribociclib and letrozole in patients with relapsed ER positive OC or EC. Objectives: Primary endpoint was the proportion of patients with relapsed ER positive OC or EC alive and progression-free after 12 weeks of therapy (PFS12) with the combination of ribociclib given at a dose of 400 mg orally daily and letrozole 2.5 mg orally daily. A PFS of 45% was considered a favorable result based on the data referenced above from Bowman et al. Methods: Eligibility criteria included patients with relapsed ER positive OC or EC, with measurable disease, not previously treated with ribociclib or AIs. Xenografts were created from CT guided tumor biopsies at baseline to assess feasibility. Results: A total of 40 patients were enrolled (20 with OC and 20 with EC) ) with a median age of 61 years (range: 30-82) and 64.5 (range: 52-75) in the OC and EC groups respectively. Among the OC patients, 17 had high grade serous carcinomas and 3 had low grade serous carcinomas. 11 EC patients had endometrioid cancers (3 with grade 1 tumors) and 9 had high grade serous tumors. Ten out of 20 OC patients and 11/20 EC patients were alive and progression-free at 12 weeks (PFS12 of 50 and 55%, respectively). The most common grade 3 or higher adverse events (occurring in at least 5 pts) were leukopenia (18%), lymphopenia (18%), neutropenia (13%), and fatigue (13%). 34 tumor biopsies were suitable for injection into mice and 44% engrafted. ER expression persisted through multiple passages in mice. Two of three EC PDX models exhibited improved PFS with letrozole/ribociclib compared to letrozole alone. Conclusions: The combination of ribociclib and letrozole is associated with a promising 50% and 55% PFS12 in patients with ER positive relapsed OC or EC respectively. Creation of xenograft tumor models from CT guided biopsies of OC and EC tumors was feasible. Clinical trial information: NCT02657928.