苦参碱通过BDK/BCKDC通路改善肿瘤恶病质骨骼肌萎缩的作用及机制

Cancer cachexia is a multi-factorial syndrome without effective therapeutic drug. The pathogenesis of cancer cachexia is complex, and ongoing skeletal muscle wasting is the most paradigmatic characteristic. Our research team found that skeletal muscle atrophy is accompanied by distinct metabolic disturbance of Branched Chain Amino Acids (BCAA) and up-regulation of Branched-Chain α-Keto Acid Dehydrogenase Complex (BCKDC) during the past decade of continuous research. BCKDC is the irreversible enzyme of BCAA catabolism and its activity is regulated by its kinase (Branched-Chain α-Keto Acid Dehydrogenase Kinase, BDK). Our previous study found that BDK interference would result in abnormal BDK / BCKDC pathway and dysfunction of muscle protein synthesis. Matrine is clinically approved to relieve symptoms of cancer cachexia. However, the mechanism is still unknown. Our recent studies showed that matrine could effectively alleviate the skeletal muscle atrophy and increase the expression of BDK from the in vivo cancer cachexia murine experiment. However, matrine could not reverse skeletal muscle atrophy when BDK was interfered from an in vitro experiment. Thus we hypothesize that the effect of matrine on alleviation of skeletal muscle atrophy during cancer cachexia is dependent on BDK and its downstream BDK / BCKDC pathway. To elucidate the hypothesis, we intend to design a series of in vitro and in vivo experiments, from molecular, cellular and tissue levels, to reveal the effect of BDK / BCKDC pathway on regulation of skeletal muscle protein synthesis, and clarify the mechanism of matrine in ameliorating skeletal muscle atrophy during cancer cachexia. This study can not only clarify the pharmacological mechanism of matrine on alleviation of skeletal muscle atrophy, but also reveal a new drug target and mechanism of skeletal muscle atrophy during cancer cachexia.

肿瘤恶病质（CC）发病机制复杂，临床缺乏有效治疗药物。骨骼肌萎缩是CC重要特征。我们团队近年持续研究发现骨骼肌萎缩伴随特异性支链氨基酸（BCAA）代谢改变和支链α酮酸脱氢酶复合物（BCKDC）的表达上调。BCKDC是BCAA分解代谢不可逆酶，受其激酶（BDK）调控。BDK介导的BDK/BCKDC通路异常会引起肌蛋白合成障碍和骨骼肌萎缩。苦参碱已被批准用于治疗CC，但作用机制不明。初步研究揭示苦参碱可有效改善CC骨骼肌萎缩，增加骨骼肌BDK表达，但不能逆转干扰BDK表达后所引起的骨骼肌萎缩。故推测：苦参碱通过BDK/BCKDC通路调控BCAA代谢，改善CC骨骼肌萎缩。本项目拟从分子、细胞和组织水平，通过体内外实验，研究BDK/BCKDC通路在骨骼肌蛋白合成中的作用，明确苦参碱对BDK/BCKDC通路的调控机制。本项目可揭示苦参碱抗CC的作用机制，也可为抗CC药物的开发研究提供新的靶点和思路。

肿瘤恶病质（cancer cachexia，CC）是一种代谢性多因素不可逆综合征。课题组前期进行了肿瘤恶病质生物标志物的临床研究，发现肿瘤恶病质骨骼肌萎缩与支链氨基酸（Branched chain amino acid，BCAA）代谢过程密切相关。BCAA的限制性不可逆代谢酶是支链α-酮酸脱氢酶复合物（Branched-Chain Alpha-keto Acid Dehydrogenase Complex, BCKDC）。BCKDC可被BDKDC激酶（BCKD kinase，BDK）磷酸化失活，被BCKDC磷酸酶（BCKD phosphatase，BDP）去磷酸化活化。干扰BDK表达或特异性抑制BDK活性可引起BCKDC活性升高，并伴随MHC、MyoD和MyoG表达减少。中国食品药品监督管理局(China Food and Drug Administration，CFDA)已经批准苦参碱注射液应用于临床预防和治疗癌症恶病质。但是作用机制目前仍不明确，苦参碱在改善地塞米松诱导的C2C12肌小管萎缩、增加MyoD、MHC表达同时，还可增加BDK mRNA表达。并且，敲减C2C12肌小管中的BDK基因后，苦参碱对地塞米松诱导的MyoD和MHC表达下调逆转作用减弱。苦参碱具有缓解CT26肿瘤恶病质症状的作用，并且这种作用主要与保存骨骼肌质量相关。作用特征分析表明，苦参碱一方面可抑制E3泛素连接酶表达而降低骨骼肌蛋白降解，另一方面可促进C2C12成肌细胞分化，改善肌小管萎缩，提高PI3K/Akt信号通路中的关键蛋白Akt、mTOR和FoxO3α磷酸化水平，改善骨骼肌蛋白合成。进一步机制研究表明BDK基因在骨骼肌萎缩中扮演重要的角色，肌萎缩可导致BDK表达下调；敲减BDK可降低骨骼肌质量，引起骨骼肌萎缩。而过表达BDK可改善恶病质所致骨骼肌萎缩。苦参碱可通过调控BDK表达增加骨骼肌质量和肌纤维尺寸，其机制与增加Akt、mTOR等蛋白磷酸化水平促进骨骼肌蛋白合成，抑制E3泛素化连接酶表达改善肌蛋白降解相关。