Extracellular vesicle-mediated transfer of long non-coding RNA ROR modulates chemosensitivity in human hepatocellular cancer

Hepatocellular cancers (HCC) are highly resistant to chemotherapy. TGF beta has been associated with chemoresistance in some human cancers but the mechanisms involved are unknown. We explored how TGF beta might contribute to altered responses to therapy by assessing the involvement and mechanistic contribution of extracellular vesicle long non-coding RNA (lncRNA) in mediating TGF beta-dependent chemoresistance. TGF beta reduced the sensitivity of HCC cells to sorafenib or doxorubicin and altered the release of both extracellular vesicles and of selected lncRNA within these vesicles. Amongst these, lincRNA-ROR (linc-ROR), a stress-responsive lncRNA was highly expressed in HCC cells and enriched within extracellular vesicles derived from tumor cells. Incubation with HCC-derived extracellular vesicles increased linc-ROR expression and reduced chemotherapy-induced cell death in recipient cells. Sorafenib increased linc-ROR expression in both tumor cells and extracellular vesicles, whereas siRNA to linc-ROR increased chemotherapy-induced apoptosis and cytotoxicity. Tumor-initiating cells that express CD133 have an increased resistance to therapy. TGF beta increased expression of CD133+ cells and colony growth in limiting dilution assays, both of which were attenuated by linc-ROR knockdown. These data provide mechanistic insights into primary chemoresistance in HCC by showing that: (a) TGF beta selectively enriches linc-RoR within extracellular vesicles, which has a potential role in intercellular signaling in response to TGF beta; (b) expression and enrichment of linc-ROR during chemotherapeutic stress plays a functional role in chemoresistance; and (c) the effects of TGF beta on chemoresistance in HCC may involve linc-RoR-dependent effects on tumor-initiating cells. These findings implicate extracellular vesicle lncRNA as mediators of the chemotherapeutic response, and support targeting linc-ROR to enhance chemosensitivity in HCC. (C) 2014 The Authors. Published by Elsevier B.V. on behalf of the Federation of European Biochemical Societies.