Abstract 2347: MicroRNA mediated CDK4/6 inhibitor resistance via extracellular signaling

Abstract

Abstract Multiple potent and highly selective inhibitors of the cell cycle kinases - CDK4 and CDK6 are in development. One such inhibitor, palbociclib, was recently approved for use in combination with letrozole for the treatment of estrogen receptor positive (ER+) breast cancer. Cyclin D-dependent kinase activity is a driving factor for ER+ breast carcinogenesis, irrespective of CCND1 amplification, making CDK4/6 inhibition a promising approach for this breast cancer subset. However, as with all cancer treatments, resistance will be a major issue limiting the efficacy of this approach. To date, mechanisms of palbociclib resistance have not been extensively investigated. Through interrogation of the generated CDK4/6 inhibition resistant cells we discovered overexpression of CDK6, specifically, mediates resistance. CDK6 depletion reversed resistance and overexpression caused resistance, whereas the same was not true when CDK4 or D cyclin protein levels were manipulated. Interestingly, when generating and analyzing resistant cell populations, we observed a “bystander effect”, by which resistance was transmissible between cells. The “resistant bystander” cells display all characteristics of the drug exposure induced resistant cells, however became resistant in just 48 hours as oppose to 14 weeks. Analysis of conditioned medium revealed that the transmission of resistance is dependent on exosomes, but not protein or DNA. We identified a specific miRNA, present in the exosomes of resistant cells, by microarray which caused resistance when excreted. Additionally, overexpression and inhibition of the miRNA confirmed that it is responsible for causing resistance. miRNA overexpressing cells exhibited the same phenotype as drug induced resistant cells, and furthermore, could cause resistance in neighboring cell populations by exosome mediated signaling. Using Biotin labelled miRNA-mRNA pulldown followed by RNA-seq, we identified the TGFβ pathway as the miRNA target. Downregulation of the TGFβ caused a decrease in the CDK inhibitors, p15 and p21, resulting in an increased CDK6 protein level and palbociclib resistance. We subsequently confirmed these data in patient samples by comparing before treatment and post relapse biopsies. These findings highlight a novel mechanism of conferred drug resistance as well as new insights into acquired CDK4/6 inhibitor resistance. Citation Format: Liam Cornell, Geoffrey I. Shapiro. MicroRNA mediated CDK4/6 inhibitor resistance via extracellular signaling [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 2347. doi:10.1158/1538-7445.AM2017-2347