Abstract C38: Small molecule targeting of the mRNA stability factor HuR impacts COX-2 expression in colon cancer
Authors: Blanco Fernando F. , Li Yuan , Meisner-Kober Nicole , Anant Shrikant , Dixon Dan A. 
Published: May 2019

Background: Colorectal cancer (CRC) is the third most common cancer among American adults and a leading cause of cancer-related mortality. The initiation and progression of colorectal tumorigenesis is characterized by genetic alterations that promote expression of tumor-promoting genes such as COX-2. In non-transformed cells, the mRNAs of these genes are often targeted for rapid decay through AU-rich elements (ARE) present in their 3'UTR. However, ARE-mediated mRNA decay is compromised during colorectal tumorigenesis, allowing for pathogenic gene overexpression. Our prior work identified the RNA-binding protein HuR to be overexpressed during colon tumorigenesis. In this capacity, HuR promotes COX-2 expression by interfering with rapid mRNA degradation and is associated with poor clinical prognosis. In the present study, we examined the therapeutic potential of inhibiting HuR using the small molecule MS-444.

Methods: HCT116, HCA-7, SW480, RKO, and HT-29 colon cancer cells along with non-transformed intestinal epithelial (RIE-1) and colonocytes (YAMC) were used. MS-444 was obtained from Novartis. Cell proliferation and apoptosis were assayed by MTT assay and flow cytometry. HuR trafficking was visualized by immunofluorescence. qPCR, western blot, and immunohistochemistry were performed to assess changes in gene expression. In vivo studies were performed to xenograft tumor models of colon cancer cells using IVIS-based imaging for COX-2.

Results: Treatment of the CRC cells lines HCT116, HCA-7, SW480, RKO, and HT-29 with MS-444 resulted in growth inhibition with IC50 values ranging between 5.6-12.8 μM, and this observed growth inhibition resulted from increased cell apoptosis. Importantly, treatment of non-transformed intestinal and colonic epithelial cells with MS-444 exhibited IC50 values of 40.7μM and 28.1 μM, respectively, and were resistant to MS-444-induced apoptosis. The expression of HuR was not impacted by MS-444, whereas MS-444 altered HuR's presence in the cytoplasm. HuR performs its mRNA stabilization function when present in the cytoplasm, and CRC cells treated with MS-444 displayed nuclear HuR as a result inhibition of HuR cytoplasmic trafficking. Consistent with growth-inhibitory IC50 values, MS-444 inhibited COX-2 mRNA and protein expression in CRC cells. This inhibition required the COX-2 ARE-containing 3'UTR and occurred on a post-transcriptional level. In vivo, delivery of MS-444 resulted in xenograft CRC tumor growth inhibition and prevented HuR cytoplasmic localization. Furthermore, treatment of MS-444 inhibited tumor COX-2 overexpression.

Conclusions: These findings indicate that small molecule-based inhibition of HuR results in selective CRC cell death in part by promoting COX-2 mRNA degradation, while also indicating that HuR is a necessary component for colorectal tumor cell survival.

Citation Format: Fernando F. Blanco, Yuan Li, Nicole Meisner-Kober, Shrikant Anant, Dan A. Dixon. Small molecule targeting of the mRNA stability factor HuR impacts COX-2 expression in colon cancer. [abstract]. In: Proceedings of the Twelfth Annual AACR International Conference on Frontiers in Cancer Prevention Research; 2013 Oct 27-30; National Harbor, MD. Philadelphia (PA): AACR; Can Prev Res 2013;6(11 Suppl): Abstract nr C38.