Garbarino Ombretta, Valenti Giulia Elda, Monteleone Lorenzo, Pietra Gabriella, Mingari Maria Cristina, Benzi Andrea, Bruzzone Santina, Ravera Silvia, Leardi Riccardo, Farinini Emanuele, Vernazza Stefania, Grottoli Melania, Marengo Barbara, Domenicotti Cinzia
Department of Experimental Medicine, General Pathology Section, University of Genoa, Genoa, Italy.
IRCCS Ospedale Policlinico San Martino, Genova, Italy.
Front Oncol. 2023 Jul 18;13:1210130. doi: 10.3389/fonc.2023.1210130. eCollection 2023.
Malignant melanoma is the most lethal form of skin cancer which shows BRAF mutation in 50% of patients. In this context, the identification of BRAF mutation led to the development of specific inhibitors like PLX4032. Nevertheless, although its initial success, its clinical efficacy is reduced after six-months of therapy leading to cancer relapse due to the onset of drug resistance. Therefore, investigating the mechanisms underlying PLX4032 resistance is fundamental to improve therapy efficacy. In this context, several models of PLX4032 resistance have been developed, but the discrepancy between and results often limits their clinical translation.
The herein reported model has been realized by treating with PLX4032, for six months, patient-derived BRAF-mutated melanoma cells in order to obtain a reliable model of acquired PLX4032 resistance that could be predictive of patient's treatment responses. Metabolic analyses were performed by evaluating glucose consumption, ATP synthesis, oxygen consumption rate, P/O ratio, ATP/AMP ratio, lactate release, lactate dehydrogenase activity, NAD/NADH ratio and pyruvate dehydrogenase activity in parental and drug resistant melanoma cells. The intracellular oxidative state was analyzed in terms of reactive oxygen species production, glutathione levels and NADPH/NADP ratio. In addition, a principal component analysis was conducted in order to identify the variables responsible for the acquisition of targeted therapy resistance.
Collectively, our results demonstrate, for the first time in patient-derived melanoma cells, that the rewiring of oxidative phosphorylation and the maintenance of pyruvate dehydrogenase activity and of high glutathione levels contribute to trigger the onset of PLX4032 resistance.
Therefore, it is possible to hypothesize that inhibitors of glutathione biosynthesis and/or pyruvate dehydrogenase activity could be used in combination with PLX4032 to overcome drug resistance of BRAF-mutated melanoma patients. However, the identification of new adjuvant targets related to drug-induced metabolic reprogramming could be crucial to counteract the failure of targeted therapy in metastatic melanoma.
恶性黑色素瘤是最致命的皮肤癌形式，50%的患者存在BRAF突变。在此背景下，BRAF突变的鉴定促使了如PLX4032等特异性抑制剂的研发。然而，尽管其初期取得了成功，但治疗六个月后临床疗效会降低，由于耐药性的出现导致癌症复发。因此，研究PLX4032耐药的潜在机制对于提高治疗效果至关重要。在此背景下，已开发出多种PLX4032耐药模型，但不同研究结果之间的差异常常限制了它们的临床转化。
本文报道的模型是通过用PLX4032处理患者来源的BRAF突变黑色素瘤细胞六个月来实现的，以获得一个可靠的获得性PLX4032耐药模型，该模型可预测患者的治疗反应。通过评估亲本和耐药黑色素瘤细胞中的葡萄糖消耗、ATP合成、耗氧率、P/O比值、ATP/AMP比值、乳酸释放、乳酸脱氢酶活性、NAD/NADH比值和丙酮酸脱氢酶活性来进行代谢分析。从活性氧产生、谷胱甘肽水平和NADPH/NADP比值方面分析细胞内氧化状态。此外，进行主成分分析以确定导致获得靶向治疗耐药性的变量。
总体而言，我们的结果首次在患者来源的黑色素瘤细胞中表明，氧化磷酸化的重新布线以及丙酮酸脱氢酶活性和高谷胱甘肽水平的维持有助于引发PLX4032耐药的发生。
因此，可以推测谷胱甘肽生物合成抑制剂和/或丙酮酸脱氢酶活性抑制剂可与PLX4032联合使用，以克服BRAF突变黑色素瘤患者的耐药性。然而，确定与药物诱导的代谢重编程相关的新辅助靶点对于对抗转移性黑色素瘤靶向治疗的失败可能至关重要。
