Role of miR-193a as therapeutic agent and biomarker in cutaneous melanoma

Background. MicroRNAs (miRNAs) are promising diagnostic biomarkers in cancer and other diseases and may represent therapeutic targets or drug themselves. Their presence in blood and other physiological fluids may be a source of information useful for disease diagnosis, prognosis and treatment [1]. They are short evolutionarily-conserved regulatory RNAs whose main function is the down-regulation of proteins coded by target mRNAs (Ivkovic et al., 2017). Many miRNAs are involved in cancer development and progression. MiR-193a acts as potential tumour suppressor in malignant pleural mesothelioma, gastric and non-small-cell lung cancer (Williams et al., 2015; Jian et al., 2016; Yu et al., 2014) and it regulates drug and chemoradiation resistance in bladder and oesophageal cancer, respectively (Lv et al., 2016; Deng et al., 2014; Meng et al., 2016). As regards melanoma, actually a study evaluating the expression of miR-193a in cutaneous melanoma tissues and cell lines (Caramuta et al., 2010) and a pilot investigation from our laboratory on its levels in plasma of melanoma patients compared to healthy controls have been realized (Fogli et al., 2017). Nevertheless, no data are reported on the role of miR-193a on the control of melanoma cell proliferation and metastasis. Aim. In the present study, functional in vitro experiments on the effect of mir193a ectopic over-expression in melanoma cell lines was investigated in order to better understand its role in this disease. Parallely, its expression in plasma exosomes derived from stage IV melanoma patients was investigated in order to confirm its role as biomarker. Methods: A miR-193a mimic was transfected with lipofectamine in three different melanoma cell lines with different BRAF mutation status (A375, 501Mel and MeWo) and its influence on cell viability, clonogenicity, 3D spheroid formation and pAkt, pErk, BRaf protein levels were evaluated. Exosomes from plasma samples of melanoma patients and healthy donors, were isolated and their mir-193a levels were determined via quantitative real-time PCR using two types of normalization: versus endogenous miR-16 and exogenous C. elegans miR-39. Results. In vitro experiments on miR193a mimic-transfected cells showed a significant decrease of cell viability and clonogenicity. In addition, pAkt levels were reduced in miR193a overexpressing cells. A statistically significant decrease in the levels of miR-193a was observed in exosome-derived plasma of metastatic melanoma patients compared to healthy donors, with a greater decrease in patients with the BRAF V600E mutation. Conclusions. Our data suggest that miR193a represent a potential therapeutic agent reducing melanoma progression and acting prevalently on the Akt pathway. Future experiments will be aimed at investigating this therapeutic strategy in an in vivo melanoma model. Moreover, our results confirm the biomarker role of this miRNA and the influence exerted by BRAF mutation on its circulating levels. Keywords: miR-193a, cutaneous melanoma, exosome References Caramuta et al. (2010) J Invest Dermatol.;130(8):2062-70. Deng et al., (2014) Mol Cancer.;13:234. Fogli et al., (2017) Tumour Biol.;39(5):1010428317701646. Ivkovic et al., (2017) Cancer Lett.;12. pii: S0304-3835(17)30237-9. Jian et al., (2016) Tumour Biol.; Jul;37(7):8941-9. Liu et al., (2015) J Cancer Res Clin Oncol.;141:993–1006 Lv et al., (2014) Cell Death Dis.;5:e1402. Meng et al., (2016) Gene;579(2):139-45. [1] Schwarzenbachet al., (2014) Nat Rev Clin Oncol.,11(3):145-56 Williams et al., (2015) Oncotarget;6(27):23480-95. Yu et al., (2015) Oncogene; 34(4):413-23.