Abstract PR01: Cyclin E: Targeting cell cycle dependencies in CCNE1-amplified tumors

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 harbor a defect in the homologous recombination (HR) pathway of DNA repair. In contrast, the 20% that harbor CCNE1 amplifications 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 from G1 to S phase. 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. This leads to an accumulation of DNA damage and altered gene expression of genes involved in DNA replication and fork protection. 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. One of those identified hits was the RAD51 paralog XRCC2, which is known to be involved in the homologous recombination DNA repair pathway and in fork protection. We could show that XRCC2 expression is upregulated in response to Cyclin E overexpression in FTSECs and when analyzing the TCGA patient cohorts, we found a strong correlation between RNAseq expression of XRCC2 and Cyclin E. We could further demonstrate that the knockdown of XRCC2 is synthetic lethal in CCNE1 amplified ovarian cancer cell lines but not in cells that harbor no CCNE1 amplification, indicating that the upregulation of XRCC2 creates a dependency in CCNE1-amplified tumors. Since overexpression of Cyclin E leads to unscheduled S-phase entry and stress on the replication fork, we speculated that one of the roles of XRCC2 might be to stabilize the replication fork in CCNE1-overexpressing cells. We found that the knockdown of XRCC2 in Cyclin E-overexpressing cells leads to a strong reduction in fork speed and fork recovery. To further understand this mechanism we analyzed the binding partners of XRCC2 in CCNE1 amplified cells by mass spectrometry. Interestingly, we found that XRCC2 interacts with the minichromosome maintenance deficient 7 (MCM7) protein. MCM7 is part of the MCM complex that unwinds the DNA during replication. Surprisingly, the downregulation XRCC2 also led to a strong reduction in MCM7 protein expression, indicating that XRCC2 may play an important role in stabilization of the MCM complex. This is especially interesting since CCNE1-amplified cells are more dependent on active MCM complexes and are more sensitive to MCM complex reduction compared to normal cells. Further defining the factors that contribute to the XRCC2-MCM7 interaction at the replication fork may define novel vulnerabilities in CCNE1-amplified tumors. This abstract is also being presented as Poster A03. Citation Format: Kai Doberstein, Alison Karst, Paul T. Kroeger, Jr., Paul Jones, William Hahn, Ronny Drapkin. Cyclin E: Targeting cell cycle dependencies in CCNE1-amplified tumors. [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 PR01.