Estudio de mirnas como marcadores pronóstico en tumores gastrointestinales. Implicación en los mecanismos de resistencia a quimioterapia y radioterapia

Resumen

microRNAs (miRNA) are small RNAs that function as post-transcriptional regulators of gene expression. Recent evidence has shown that some miRNAs can act as oncogenes or tumor suppressors. This thesis was conducted to evaluate the potential role of miRNAs as prognosis markers and its implication in therapy resistance regulation in gastrointestinal tumors. First, we measured the expression of 250 human mature miRNAs in tumors of patients with gastric cancer. Our results revealed that downregulation of miR-451 was associated with worse prognosis. miR-451 was detected in normal epithelial cells by in situ hybridization and its expression levels decreased in gastric cancer and colorectal cancer tissues when compared to those observed in normal tissues. The above results prompted us to investigate the role of miR-451 in gastrointestinal tumors. In vitro experiments demonstrated that overexpression of miR-451 in gastric and CRC cells resulted in reduced cell proliferation. We identified the macrophage migration inhibitory factor (MIF) as a potential target of miR-451 by microarray and bioinformatic analysis. Overexpression of miR-451 induced a decreased in the MIF mRNA and protein levels. We further confirmed that MIF was a target of miR-451 by renilla-luciferase assays. Moreover, we found a significant inverse correlation between miR-451 and MIF expression in gastric tumors. Since, drug resistance remains a major clinical obstacle to the successful treatment of cancer and miRNAs have been implicated in the regulation of therapy resistance. We seek to elucidate whether miR-451 play a role in colorectal cancer chemoresistance. We proved that ectopic expression of miR-451 decreased the ability of CRC cells to grow under anchorage-independent conditions, reduced their in vivo tumorigenicity and increased their sensitivity to the irinotecan-active metabolite, SN38. By bioinformatic analysis and functional results we determined that ATP-binding cassette transporter protein 1 (ABCB1) was a target of miR-451 and responsible for increased sensitivity to SN38. Next, we sought to investigate whether miR-451 was also involved in the cancer stem cell (CSC) chemotherapy resistance. The phenotype of colon spheres was confirmed by the expression of several CSCs markers, by their high capacity to initiate xenograft tumors and by their chemoresistance. Interestingly, miR-451 was downregulated in colon spheres and its restoration reduced their number and size. Moreover, overexpression of miR-451 in colon spheres decreased their tumorigenicity and increased the sensitivity to SN38 through ABCB1 downregulation. These findings support the role of miR-451 as a regulator of cell proliferation, sensitivity to SN38 and CRC stem-like cell phenotype, and open new perspectives for the development of effective therapies for chemo-radioresistant cancers.