Abstract 5104: The RNA binding protein HuR enhances exosome secretion in colorectal cancer

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Abstract Enhanced secretion of extracellular vesicles such as microvesicles and exosomes by cancer cells has been recently recognized as a new means of transferring oncogenic information within the tumor microenvironment. Through their ability to carry specific RNA and protein cargo, tumor-derived exosomes can directly impact neighboring cells to support tumor progression. However, it remains unclear what regulates exosome production in tumor cells and the factors influencing loading of tumor-promoting RNA cargo. Our prior work has established that colorectal cancer (CRC) cells and tumors overexpress the key RNA-binding protein HuR early in GI tumor development. When overexpressed and present in the cytoplasm, HuR can promote mRNA stabilization of tumor-promoting genes through binding of 3’UTR AU-rich elements (ARE). While the ability of HuR overexpression to promote gene expression is well recognized, our understanding of how HuR communicates this information within the tumor microenvironment is limited. To test if HuR overexpression could impact secreted exosomes, Tet-regulated HuR-inducible HeLa cells were used to demonstrate that cytoplasmic HuR overexpression promoted a 4-fold increase in exosomes produced. Furthermore, HuR was detected in exosomes produced only from HuR-overexpressing cells. The presence of HuR in exosomes directly impacted mRNA cargo, as selective uptake of ARE-containing mRNAs was observed in exosomes derived from HuR overexpressing cells. To test if these effects were seen in CRC cells that endogenously overexpress HuR, exosome levels from CRC cells were compared to normal primary human intestinal epithelial and myofibroblast cells. When normalized to total cell numbers, CRC cells secrete ∼3-fold greater exosome levels than normal cells and enhanced exosome production was dependent upon HuR. Knockdown of HuR in CRC cells directly impacted exosome secretion to levels observed in normal cells. HuR was also only detected in exosomes produced from CRC cells. Using an inducible model of RasV12-mediated transformation of intestinal epithelial cells, endogenous HuR overexpression was observed associated with 4-fold greater levels of exosomes containing HuR as cargo. These findings were reflected in vivo where GI-tumor bearing APCMin/+ mice produced ∼3-fold more serum exosomes, with HuR as exosomal cargo in APCMin/+ mice where as no expression of exosomal HuR was detected in wild-type mice. This work has identified a novel connection between HuR-mediated post-transcriptional regulation and tumor-derived exosome production, along with providing the first evidence indicating the presence of exosomal HuR as a serum-based CRC biomarker. Citation Format: Ranjan Preet, Shufei Zhuang, Wei-ting Hung, Lane K. Christenson, Dan A. Dixon. The RNA binding protein HuR enhances exosome secretion in colorectal cancer. [abstract]. In: Proceedings of the 107th Annual Meeting of the American Association for Cancer Research; 2016 Apr 16-20; New Orleans, LA. Philadelphia (PA): AACR; Cancer Res 2016;76(14 Suppl):Abstract nr 5104.