miR-126-3p down-regulation contributes to dabrafenib acquired resistance in melanoma by up-regulating ADAM9 and VEGF-A

Journal of Experimental & Clinical Cancer Research

Alessandra Carè¹  Simona Mastroeni²  Gian Carlo Antonini Cappellini³  Nadia Felli⁴  Ester Alvino⁵  Stefania D’Atri⁶  Pedro Miguel Lacal⁶  Federica Ruffini⁶  Simona Caporali⁶  Lauretta Levati⁶  Laura Bonmassar⁶  Ulrich Pfeffer⁷  Adriana Amaro⁷ 

[1] Center of Gender Medicine, Istituto Superiore di Sanità;Clinical Epidemiology Unit, IDI-IRCCS;Department of Oncology and Dermatological Oncology, IDI-IRCCS;Department of Oncology and Molecular Medicine, Istituto Superiore di Sanità;Institute of Translational Pharmacology, National Council of Research;Laboratory of Molecular Oncology, Istituto Dermopatico dell’Immacolata, IDI-IRCCS;Molecular Pathology, IRCCS-Ospedale Policlinico San Martino;

关键词: miR-126-3p;    melanoma;    BRAF inhibitors;    acquired resistance;    proliferation;    invasiveness;   

DOI  :  10.1186/s13046-019-1238-4

来源: DOAJ

【 摘 要 】

Abstract Background Development of resistance to inhibitors of BRAF (BRAFi) and MEK (MEKi) remains a great challenge for targeted therapy in patients with BRAF-mutant melanoma. Here, we explored the role of miRNAs in melanoma acquired resistance to BRAFi. Methods miRNA expression in two BRAF-mutant melanoma cell lines and their dabrafenib-resistant sublines was determined using Affymetrix GeneChip® miRNA 3.1 microarrays and/or qRT-PCR. The effects of miR-126-3p re-expression on proliferation, apoptosis, cell cycle, ERK1/2 and AKT phosphorylation, dabrafenib sensitivity, invasiveness and VEGF-A secretion were evaluated in the dabrafenib-resistant sublines using MTT assays, flow cytometry, immunoblotting, invasion assays in Boyden chambers and ELISA. ADAM9, PIK3R2, MMP7 and CXCR4 expression in the sensitive and dabrafenib-resistant cells was determined by immunoblotting. Small RNA interference was performed to investigate the consequence of VEGFA or ADAM9 silencing on proliferation, invasiveness or dabrafenib sensitivity of the resistant sublines. Long-term proliferation assays were carried out in dabrafenib-sensitive cells to assess the effects of enforced miR-126-3p expression or ADAM9 silencing on resistance development. VEGF-A serum levels in melanoma patients treated with BRAFi or BRAFi+MEKi were evaluated at baseline (T0), after two months of treatment (T2) and at progression (TP) by ELISA. Results miR-126-3p was significantly down-regulated in the dabrafenib-resistant sublines as compared with their parental counterparts. miR-126-3p replacement in the drug-resistant cells inhibited proliferation, cell cycle progression, phosphorylation of ERK1/2 and/or AKT, invasiveness, VEGF-A and ADAM9 expression, and increased dabrafenib sensitivity. VEGFA or ADAM9 silencing impaired proliferation and invasiveness of the drug-resistant sublines. ADAM9 knock-down in the resistant cells increased dabrafenib sensitivity, whereas miR-126-3p enforced expression or ADAM9 silencing in the drug-sensitive cells delayed the development of resistance. At T0 and T2, statistically significant differences were observed in VEGF-A serum levels between patients who responded to therapy and patients who did not. In responder patients, a significant increase of VEGF-A levels was observed at TP versus T2. Conclusions Strategies restoring miR-126-3p expression or targeting VEGF-A or ADAM9 could restrain growth and metastasis of dabrafenib-resistant melanomas and increase their drug sensitivity. Circulating VEGF-A is a promising biomarker for predicting patients’ response to BRAFi or BRAFi+MEKi and for monitoring the onset of resistance.

【 授权许可】

Unknown