Abstract 829: MicroRNA regulation of radiation sensitivity in colorectal cancer

Abstract

Abstract Background Patients with locally advanced rectal cancer receive neoadjuvant chemoradiation therapy (CRT) and subsequent surgery. While 10-25% of patients have complete response to CRT, the remaining patients undergo extensive tumor excision resulting in significant quality of life issues. Response to CRT is an independent predictor of overall survival highlighting the importance of improving CRT response rates. Several tumor intrinsic factors govern responses to CRT including specific gene expression programs. Emerging evidence suggests that microRNAs (miRs) modulate gene expression programs in response to radiation and have been implicated in several pathological processes associated with colorectal cancer progression. In this context, we hypothesized that differential expression of miRs regulates colorectal cancer radiation sensitivity and can be used as a biomarker to predict radiation efficacy. Methods To investigate the differences in miR profiles between rectal cancer patients that had either a pathological partial response (PR) or no response (NR), we isolated RNA from FFPE biopsies using the miRvana microRNA isolation kit (Life Technologies). We used the Nanostring miR profiling platform and obtained absolute counts for >700 human miRs. We performed in vitro gain and loss of studies with candidate miR transfections in human CRC cell lines and used a luminescence-based assay for proliferation (Cell titer glo, Promega). Surviving fraction assays were performed by seeding cells and counting colonies stained with Crystal Violet. Eight distinct miR bioinformatic target prediction algorithms initiated miR target identification, and a RISC-Trap assay was implemented to confirm miR-target interactions. The Cancer Genome Atlas (TCGA) datasets were accessed to acquire provisional survival data on 639 colorectal adenocarcinoma patients. Results We identified 17 miRs that were differentially expressed, and miR-451a, among the most upregulated miRs, inhibited proliferation and colony formation in 2D and 3D assays in the presence of radiation. Target prediction algorithms highlighted CAB39, EMSY, EREG, and MEX3C as prominent miR-451a targets in colorectal cancer and/or radiation. TCGA subset anaylsis found CAB39 and EMSY protein levels were found to be upregulated in 14% and 6% of cases, respectively, and upregulated co-expression of these genes significantly reduced 3 year overall survival (69% vs 78%, p < 0.05). miR-target interaction was confirmed via the RISC-Trap assay with miR-451a mimic transfection resulting in robust 2.4-, 1.2-, and 2.8-fold enrichment of EREG, CAB39, and EMSY, respectively. In our partial responders patient cohort possessing upregulated miR-451a, there was significant downregulation of CAB39 and EMSY mRNA and protein compared to non-responders. Conclusions miRs alter cell survival networks affecting radiation sensitivity and serve to identify pathways amenable to alternative therapeutic modulation Citation Format: Shushan Rana, Katherine Kelley, Rebecca Ruhl, Charles Thomas, Liana Tsikitis, Sudarshan Anand. MicroRNA regulation of radiation sensitivity in colorectal cancer [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 829. doi:10.1158/1538-7445.AM2017-829