Ripretinib是一种多激酶抑制剂药物，于2020年获得FDA批准，于2021年获得EMA批准，用于治疗先前使用激酶抑制剂无效的晚期胃肠道间质瘤（GIST）。该药物最常见的副作用是肌肉疼痛和疲劳，这可能导致治疗中断或剂量降低。骨骼肌细胞高度依赖ATP来执行其功能，线粒体损伤可能在由激酶抑制剂引起的骨骼肌毒性中发挥作用。然而，分子机制尚未在文献中明确确定。本研究旨在利用小鼠C2C12肌母细胞源性肌管，阐明线粒体在ripreinib对骨骼肌毒性的作用。肌管暴露于1-20μM浓度的ripreinib中，处理时间为24h。为了确定线粒体受损在ripreinib诱导的骨骼肌毒性中的潜在作用，ripreinib处理后检测细胞内ATP水平、线粒体膜电位（MMP）、线粒体ROS产生（mtROS）、线粒体DNA（mtDNA）拷贝数和线粒体质量。此外，还研究了在线粒体生物合成和线粒体自噬中发挥作用的PGC1α / NRF1 / NRF2表达水平变化。另外，还评估了线粒体电子传递链（ETC）酶活性。最后，进行了分子对接研究，以查看ripreinib与线粒体中的DNA聚合酶伽玛（POLG）的可能相互作用。根据研究发现，ripreinib降低了ATP水平和mtDNA拷贝数，引起MMP丢失，降低了线粒体质量。随着ripreinib暴露，ETC复合物的活性被抑制，这与观察到的ATP耗竭和MMP损失相一致。分子对接研究揭示ripreinib具有抑制POLG的潜力，这支持了观察到的mtDNA抑制。在核部分，PGC1α的表达减少，表明PGC-1α未被激活，因为NRF1表达减少，而NRF2水平未显示出显著变化。因此，所有治疗组的mtROS产生增加，与线粒体自噬相关的基因表达和Parkin蛋白表达水平在高剂量下上调。总之，线粒体损伤/丧失可能是ripreinib诱导的骨骼肌毒性的潜在原因之一。然而，需要进一步研究以确认体内的结果。版权所有©2023 Elsevier B.V.发行。

Ripretinib is a multikinase inhibitor drug approved in 2020 by the FDA and in 2021 by EMA for use in the treatment of advanced gastrointestinal stromal tumors (GIST) which have not adequately responded to previous treatments with kinase inhibitors. The most common side effects of the drug are myalgia and fatigue, which likely causes interruption of the treatment or reduction of the dose. Skeletal muscle cells highly depend on ATP to perform their functions and mitochondrial damage may play a role in skeletal muscle toxicity induced by kinase inhibitors. However, the molecular mechanism has not been clearly identified in the literature yet. In this study, it has been aimed to elucidate the role of mitochondria in the toxic effect of ripretinib on skeletal muscle using the mouse C2C12 myoblast-derived myotubes. The myotubes were exposed to ripretinib at the range of 1-20μM concentrations for 24h. To determine the potential role of mitochondrial impairment in ripretinib-induced skeletal muscle toxicity, intracellular ATP level, mitochondrial membrane potential (MMP), mitochondrial ROS production (mtROS), mitochondrial DNA (mtDNA) copy number, and mitochondrial mass were examined after ripretinib treatment. Furthermore, changes in PGC 1α/NRF 1/NRF 2 expression levels that play a role in mitochondrial biogenesis and mitophagy were investigated. Additionally, the mitochondrial electron transport chain (ETC) enzyme activities were evaluated. Lastly, a molecular docking study was done to see ripretinib's possible interaction with DNA polymerase gamma (POLG) which is important for DNA replication in the mitochondria. According to the findings, ripretinib decreases the ATP level and mtDNA copy number, induces loss of MMP, and reduces mitochondrial mass. The activities of the ETC complexes were inhibited with ripretinib exposure which is in line with the observed ATP depletion and MMP loss. The molecular docking study revealed that ripretinib has inhibitory potential against POLG which supports the observed inhibition of mtDNA. The expression of PGC 1α was reduced in the nuclear fraction indicating that PGC-1α was not activated since the NRF 1 expression was reduced and NRF 2 level did not show significant change. Consequently, mtROS production increased in all treatment groups and mitophagy-related gene expressions and Parkin protein expression level were up-regulated at high doses. In conclusion, mitochondrial damage/loss can be one of the underlying causes of ripretinib-induced skeletal muscle toxicity. However, further studies are needed to confirm the results in vivo.Copyright © 2023. Published by Elsevier B.V.