Implicació del miRNorma tumoral en l´expressió del transportador concentratiu de nucleòsids hCNT1

Abstract

Human Concentrative Nucleoside Transporters (hCNTs) are encoded by SLC28 gene family. It has been demonstrated that hCNT1 is lost in some cancers. Furthermore, hCNTs have the ability of translocating selected nucleoside- derivatives, currently used in anticancer therapies, for that reason, a change in their expression profile could affect drug bioavailability and drug chemoresistance. Recent studies demonstrate that hCNT1 has also additional functions, being a transceptor and it is likely to be relevant in tumor biology and during the carcinogenic process. Although mechanisms responsible for hCNT1 loss during carcinogenesis are unknown, it seems likely that epigenetic modifications might be able to affect hCNT1 expression and functional activity. miRNAs are small, non-codding transcripts which play a key role during the carcinogenic process with a discriminatory expression profiles significantly different to healthy individuals. miRNAs pair to mRNA by complementarity to 3’UTR extreme of the target gene, modulating its expression. The objective of this work aims to elucidate miRNAs associated with hCNT1 loss of expression in CRC, PDAC and HCC. The analysis of paired clinical samples of CRC, PDAC and HCC confirm the decrease of hCNT1 compared to the adjacent non-tumoral tissue. Also, increased expression of miR-106a, miR-17 and miR- 18a is correlated with hCNT1 loss of expression in the same clinical samples. The luciferase assay validates hCNT1 as a direct target of miR-106a and miR-17. Furthermore, the modulation of these miRNAs could affect hCNT1 mRNA and protein levels. A new 3D cell culture model (spheroids) is presented to increase basal levels of endogenous hCNT1 compared to monolayer cell culture. This new model allows to evaluate hCNT1 modulation by miRNAs as an easy and complete way. The results obtained in spheroid culture confirm the ability of miR-106a and miR-17 to regulate hCNT1 expression. Moreover, miRNAs silencing increase gemcitabine effect in spheroids. All these results propose miR-106a and miR-17 as possible new therapeutic targets to improve nucleoside analogs treatment by increasing hCNT1 and also the possibility of using them as biomarkers.