Patel Vikas, Szász István, Koroknai Viktória, Kiss Tímea, Balázs Margit
Doctoral School of Health Sciences, University of Debrecen, 4032 Debrecen, Hungary.
MTA-DE Public Health Research Group, University of Debrecen, 4032 Debrecen, Hungary.
Cancers (Basel). 2021 Dec 1;13(23):6058. doi: 10.3390/cancers13236058.
Combination treatment using BRAF/MEK inhibitors is a promising therapy for patients with advanced mutant melanoma. However, acquired resistance largely limits the clinical efficacy of this drug combination. Identifying resistance mechanisms is essential to reach long-term, durable responses. During this study, we developed six melanoma cell lines with acquired resistance for BRAFi/MEKi treatment and defined the molecular alterations associated with drug resistance. We observed that the invasion of three resistant cell lines increased significantly compared to the sensitive cells. RNA-sequencing analysis revealed differentially expressed genes that were functionally linked to a variety of biological functions including epithelial-mesenchymal transition, the ROS pathway, and KRAS-signalling. Using proteome profiler array, several differentially expressed proteins were detected, which clustered into a unique pattern. Galectin showed increased expression in four resistant cell lines, being the highest in the WM1617 cells. We also observed that the resistant cells behaved differently after the withdrawal of the inhibitors, five were not drug addicted at all and did not exhibit significantly increased lethality; however, the viability of one resistant cell line (WM1617) decreased significantly. We have selected three resistant cell lines to investigate the protein expression changes after drug withdrawal. The expression patterns of CapG, Enolase 2, and osteopontin were similar in the resistant cells after ten days of "drug holiday", but the Snail protein was only expressed in the WM1617 cells, which showed a drug-dependent phenotype, and this might be associated with drug addiction. Our results highlight that melanoma cells use several types of resistance mechanisms involving the altered expression of different proteins to bypass drug treatment.
使用BRAF/MEK抑制剂的联合治疗对晚期突变型黑色素瘤患者来说是一种很有前景的疗法。然而，获得性耐药在很大程度上限制了这种药物联合疗法的临床疗效。识别耐药机制对于实现长期、持久的反应至关重要。在本研究中，我们建立了六种对BRAFi/MEKi治疗产生获得性耐药的黑色素瘤细胞系，并确定了与耐药相关的分子改变。我们观察到，与敏感细胞相比，三种耐药细胞系的侵袭能力显著增强。RNA测序分析揭示了差异表达基因，这些基因在功能上与多种生物学功能相关，包括上皮-间质转化、ROS途径和KRAS信号传导。使用蛋白质组分析阵列，检测到了几种差异表达的蛋白质，它们聚集成一种独特的模式。半乳糖凝集素在四种耐药细胞系中表达增加，在WM1617细胞中表达最高。我们还观察到，在撤除抑制剂后，耐药细胞的行为有所不同，其中五种完全没有药物成瘾性，并且没有表现出明显增加的致死率；然而，一种耐药细胞系（WM1617）的活力显著下降。我们选择了三种耐药细胞系来研究撤除药物后蛋白质表达的变化。在“停药期”十天后，耐药细胞中CapG、烯醇化酶2和骨桥蛋白的表达模式相似，但Snail蛋白仅在表现出药物依赖表型的WM1617细胞中表达，这可能与药物成瘾有关。我们的结果强调，黑色素瘤细胞利用多种耐药机制，包括不同蛋白质表达的改变来绕过药物治疗。
