Abstract 1265: Mechanisms underlying genomic integrity in breast cancer-derived dormant circulating tumor cells
Authors: Boral Debasish , Liu Haowen N. , Yin Wei , Vishnoi Monika , Sprouse Marc L. , Chang Jenny C. , Thiery Jean Paul , Antonio Scamardo , Hong David S. , Marchetti Dario 
Published: July 2017

Abstract
Patients with metastatic breast cancer have a five-year survival rate of 22% compared to 99% in patients with cancer localized to the breast. This disparity in clinical outcome is at least in part caused by the lack of diagnostic tools for early detection of metastatic recurrence. This can be especially valid for breast cancer patients having brain metastases (BCBM), 10-20% of whom are diagnosed only at autopsy. The early detection of circulating tumor cells (CTCs) in peripheral blood during the metastatic cascade provides a window of opportunity to prevent BCBM onset. However, to target CTCs causing BCBM, one must dissect mechanisms that allow CTCs to retain their brain metastatic potency while remaining dormant in circulation. We hypothesized that hyperactive mechanisms of DNA repair preserve the genomic make-up of dormant CTCs allowing them to maintain over time the potential for BCBM competence. Comprehensive analyses of breast cancer patient-derived CTC transcriptomes showed that CTC gene signatures closely relate to basal-like breast cancer cell lines rather than the molecular subtype of the primary tumor. However, unlike actively cycling cell lines, a large proportion of CTCs were mitotically dormant (Ki67-/PCNA-) with low overall mRNA expression and transcriptional activity. Upon systemic injection of patient-derived common lineage-depleted cells into immuno-compromised mice, the majority of CTCs were found at metastatic sites in a state of mitotic/metabolic dormancy. Second, dormant CTCs had a lower incidence of double-strand DNA breaks (DSB) than proliferating cells as assessed by Serine139 phosphorylation status of gamma H2AX. Third, we discovered that the human telomere-associated protein RIF1, a mediator of alternative non-homologous end joining repair of DNA DSB interplaying with BRCA1/2, was significantly up-regulated in the Ki67-/PCNA- CTC population. RIF1/BRCA1 foci formation was confirmed in areas of DNA damage in these cells. Lastly, shRNA-mediated RIF1 knockdown promoted dormant cell populations to become more susceptible to UV and bleomycin-induced DNA damage while activating p38 and ERK stress-response pathways as well as their phosphorylation ratio. Moreover, RIF1 knockdown in MDA-MB231BR clone attenuated its metastatic competence to the brain. Collectively, these findings suggest that RIF1 regulates the genomic integrity of dormant CTCs allowing them to survive over time while in circulation. Further elucidation of RIF1-mediated CTC pathways and its dynamics with BRCA1/2 may provide novel strategies for therapeutic intervention. This can be clinically useful in breast cancer patients that are yet to develop overt BCBM.
Citation Format: Debasish Boral, Haowen N. Liu, Wei Yin, Monika Vishnoi, Marc L. Sprouse, Jenny C. Chang, Jean Paul Thiery, Scamardo Antonio, David S. Hong, Dario Marchetti. Mechanisms underlying genomic integrity in breast cancer-derived dormant circulating tumor cells [abstract]. In: Proceedings of the American Association for Cancer Research Annual Meeting 2017; 2017 Apr 1-5; Washington, DC. Philadelphia (PA): AACR; Cancer Res 2017;77(13 Suppl):Abstract nr 1265. doi:10.1158/1538-7445.AM2017-1265