Abstract P2-05-02: Low molecular weight cyclin E facilitate replication stress tolerance in breast cancer development

Abstract

Abstract Low Molecular Weight Cyclin E (LMW-E) are the tumor specific, oncogenic forms of cyclin E that are post translationally generated by neutrophil elastase (NE) mediated cleavage of the 50 KDa full-length cyclin E1 (FL-cycE, encoded by CCNE1 gene). While FL-cycE localizes mainly to the cell nucleus, LMW-E lack the N-terminus nuclear localization signal and are detected in both the nucleus and cytoplasm. Compared to FL-cycE, LMW-E exhibit longer half-life and higher affinity to their kinase partner CDK2 and are resistant to natural CDK inhibitors such as p21 and p27. It is currently assumed that LMW-E drive the tumorigenic process by promoting G1/S cell cycle transition and accelerating mitotic exit. Here we report that LMW-E overexpression also promotes genomic instability by deregulating DNA replication in a CDC6 dependent manner. To this end, we developed an immunohistochemistry (IHC) assay with a cyclin E antibody that can identify LMW-E expressing tumors and examined the association of genetic instability of each tumor with LMW-E status in 2 different cohorts of breast cancer patients. Cohort 1 is a retrospective cohort of 725 patients with stage I-II breast cancer treated at MD Anderson (Houston, TX) between 1985 and 1999. Cohort 2 is a prospective cohort of 85 patients with stage I-II breast cancer who enrolled in our study at MD Anderson between January 2000 and June 2010. Our results show that positive LMW-E status in stage 1 or 2 breast cancer patients correlates with increasing copy number variations, as identified by Molecular Inversion Probe (MIP) in cohort 1 and somatic mutations, as identified by Whole Exome Sequencing (WES) in cohort 2. Second, using immortalized human mammary epithelial cells (hMECs) engineered to express doxycycline inducible LMW-E or FL-cycE in CCNE1 knock-out background, we found that FL-cycE over-expression leads to DNA damage, cell cycle arrest and cell death. LMW-E overexpression, on the other hand, facilitates cell proliferation with damaged DNA, resulting in multi-nuclei and micro-nuclei formation in daughter cells. Third, overexpression of FL-cycE reduces chromatin bound MCM complex, while LMW-E overexpression promotes the chromatin loading of pre-replication complex including MCMs. Lastly, we show that LMW-E but not FL-cycE is the major form of cyclin E that binds to the chromatin. Specifically, in both LMW-Einducible hMECs and LMW-Ehighbreast tumor cell lines, CDC6 is required for the nuclear translocation and chromatin loading of LMW-E. Our findings have revealed the unique oncogenic function of LMW-E in deregulating replication licensing, promoting replication stress tolerance and genomic instability that fuels tumor development. These findings also provide potential novel therapeutic strategies for treating LMW-Ehigh breast tumors, who do not respond to the current standard of care therapies. Citation Format: Mi Li, Kwang Huei Low, Tuyen Bui, Kelly K Hunt, Khandan Keyomarsi. Low molecular weight cyclin E facilitate replication stress tolerance in breast cancer development [abstract]. In: Proceedings of the 2021 San Antonio Breast Cancer Symposium; 2021 Dec 7-10; San Antonio, TX. Philadelphia (PA): AACR; Cancer Res 2022;82(4 Suppl):Abstract nr P2-05-02.