Abstract PR05: Identifying potential targets in Cyclin E-amplified tumors.
Authors: Kai Doberstein, Alison Karst, Dariush Etemadmoghadam, Paul Jones, Gavin Dunn, William Hahn, David Bowtell, Ronny Drapkin
Journal: Clinical Cancer Research (2016)

Abstract Genomic instability is a hallmark of high grade serous ovarian carcinoma (HGSOC). Based on The Cancer Genome Atlas (TCGA), it is estimated that approximately 50% of HGSOCs harbour a defect in the homologous recombination (HR) pathway of DNA repair. A majority of these cases are due to the germline and/or somatic inactivation of the BRCA1 or BRCA2 genes. In contrast, the 20% of HGSOC that harbor CCNE1 amplifications are mutually exclusive with BRCA mutations and appear to have an intact HR pathway. These tumors are associated with shorter overall survival and resistance to platinum-based chemotherapy. Cyclin E is the activating partner of cyclin-dependent kinase 2 (CDK2) which controls cell cycle progression through phosphorylation of pRB and induction of E2F transcriptional activity. Our previous data showed that CCNE1 amplification and overexpression occurs early in serous tumorigenesis. Importantly, in immortalized human fallopian tube secretory epithelial cells (FTSEC), constitutive cyclin E overexpression imparts malignant characteristics to these cells, including increased proliferation, loss of contact inhibition and clonogenicity. This leads to an accumulation of DNA damage and altered gene expression of genes involved in DNA replication fork protection and the BRCA-Fanconi Anemia pathway. However, in the setting of hTERT expression and a p53 mutant, Cyclin E overexpression alone was not capable of fully transforming the FTSECs. Therefore, in order to identify cooperating genetic alterations, we performed an in vitro gain-of-function (GOF) screen using an open reading frame (ORF) library of approximately 800 genes that are recurrently amplified in HGSOC. The transduced cells were then tested for anchorage independent colony formation on ultra-low attachment (ULA) plates and soft agar. A total of 92 genes were identified between the two assays, with 28 genes registering as a “hit” in both assays. A subset was then retested for in vivo growth in immunocompromised mice using the empty vector as the negative control. Positive hits included CHD2, GAB2, AKT, PITRM1, PTPRB, XRCC2, PSME4, and SLC38A1. Interestingly, by using RNA interference we found that knock-down of some of the genes found in the GOF screen is synthetic lethal in cancer cells that overexpress Cyclin E or have a CCNE1 amplification. The identified hits included genes involved in the DNA damage response pathways and the Fanconi Anemia pathway. Underlining the importance of these genes, analysis of the TCGA revealed that both pathways are strongly upregulated in Cyclin E overexpressing tumors. These results suggest that targeting cooperating genetic dependencies in CCNE1 amplified tumors may be a novel therapeutic avenue. We are currently using our panel of patient-derived tumor xenograft and cell lines to address this possibility. This abstract is also presented as Poster A06. Citation Format: Kai Doberstein, Alison Karst, Dariush Etemadmoghadam, Paul Jones, Gavin Dunn, William Hahn, David Bowtell, Ronny Drapkin. Identifying potential targets in Cyclin E-amplified tumors. [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 PR05.