BACR 14: MicroRNA-31 mediates chemoresistance in malignant pleural mesothelioma via reduced cisplatin intranuclear accumulation
1School of Biological, Biomedical and Environmental Sciences, University of Hull, Hull, UK,2Hull York Medical School, Hull, UK,3Centre for Oncology and Haemotology, Castle Hill Hospital, Hull and East Yorkshire NHS Trust, Hull, UK
Malignant pleural mesothelioma (MPM) is associated with extremely poor prognosis and many patients are unresponsive to treatment. MicroRNA (miR) are small non-coding RNA that regulate gene expression and have been demonstrated to alter cellular sensitivity to cytotoxic agents in various cancers. Deletion of miR-31 has previously been associated with poor prognosis in MPM. Here, we aim to investigate the role of miR-31 in modulating chemosensitivity in MPM.
The miR-31-deficient NCI-H2452 and miR-31 positive P31 cell lines were used as in vitro models of MPM. Stable miR-31 overexpression and zip down was achieved via liposomal-based transfection. Clonogenic assay was employed as a measure of cellular sensitivity to cisplatin. 3T3 assay was used to assess proliferation. Inductively coupled plasma mass spectrometry (ICP-MS) was utilised to quantify cellular cisplatin flux. Subcellular fractionation was applied to isolate intracellular organelles. Gene expression was assessed by qPCR, and protein expression by Western Blot.
Reintroduction of miR-31 into cisplatin treated NCI-H2452 cells significantly increased cell survival; conversely, zipping down of miR-31 in P31 increased cellular sensitivity to cisplatin. Additionally, miR-31 reintroduction mediated a delay in the cytotoxic activity of cisplatin. The expression of the lysosomal transporter ABCB9 was increased upon miR-31 re-expression. Interestingly, a higher relative intracellular concentration of platinum was observed in miR-31 transfected cells. However, a significantly decreased intranuclear concentration of platinum was determined in miR-31 expressing cells, suggesting a mechanism underpinning resistance involving altered nuclear transport.
Here, miR-31 expression was found to significantly enhance chemoresistance in MPM cells in vitro. While deletions in the genomic location encoding miR-31, 9p21.3, may be associated with an overall poor prognosis, the loss of miR-31 may not actually contribute to the chemoresistance observed in MPM patients. Our current work examines the impact of how miR-31 functionally modulates intracellular spatial distribution of cisplatin.