Oleinik Natalia V, Atilgan Firdevs Cansu, Kassir Mohamed Faisal, Lee Han Gyul, Janneh Alhaji H, Wofford Wyatt, Walton Chase, Szulc Zdzislaw M, Hill Elizabeth G, Alekseyenko Alexander V, Cimen Huseyin, Hartman Jessica H, Voelkel-Johnson Christina, Lilly Michael B, Lemasters John J, Frizzell Norma, Yu Xue-Zhong, Mehrotra Shikhar, Ogretmen Besim
Medical University of South Carolina, Charleston, SC, United States.
Memorial Sloan Kettering Cancer Center, New York, NY, United States.
Cancer Res. 2025 Jun 20. doi: 10.1158/0008-5472.CAN-24-4042.
Bioactive ceramide induces cell death in part by promoting mitophagy. C18-ceramide levels are commonly reduced in head and neck squamous cell carcinoma (HNSCC), which correlates with poor prognosis, suggesting the potential of harnessing ceramide for cancer treatment. Here, we evaluated the ability of the ceramide analog LCL768 to induce mitophagy and metabolic stress in HNSCC. Mechanistically, LCL768 induced CerS1-mediated endogenous C18-ceramide accumulation in mitochondria to mediate mitophagy, which did not require the CerS1 transporter p17/PERMIT but was dependent on DRP1 activation via nitrosylation at C644. DRP1 facilitated anchoring of the endoplasmic reticulum (ER) and mitochondrial membranes by promoting the association between phosphatidylethanolamine in the ER and cardiolipin in mitochondrial membranes. Mutations of Drp1 that prevented its binding to ER and mitochondrial membranes blocked CerS1/C18-ceramide mitochondrial accumulation, inhibiting LCL768-mediated mitophagy. In addition, LCL768-driven mitophagy altered mitochondrial metabolism, resulting in fumarate depletion and leading to tumor suppression in vivo. Exogenous fumarate supplementation prevented LCL768-mediated mitophagy, mitochondrial trafficking of CerS1, ER-mitochondrial tethering, and tumor suppression in mice. Fumarate metabolism was associated with PARKIN succination at a catalytic cysteine (Cys431), inhibiting its association with PINK1 and ubiquitin and thereby preventing mitophagy. LCL768-induced fumarate depletion attenuated PARKIN succination to promote PARKIN activation and mitophagy, indicating a feed-forward mechanism that regulates mitophagy and fumarate metabolism through PARKIN succination. These data provide a mechanism whereby LCL768/CerS1-C18-ceramide-mediated mitophagy and tumor suppression are regulated by Drp1 nitrosylation, fumarate depletion, and PARKIN succination, providing a metabolic stress signature for lethal mitophagy.
生物活性神经酰胺部分通过促进线粒体自噬诱导细胞死亡。头颈部鳞状细胞癌（HNSCC）中C18 - 神经酰胺水平通常降低，这与预后不良相关，提示利用神经酰胺进行癌症治疗的潜力。在此，我们评估了神经酰胺类似物LCL768在HNSCC中诱导线粒体自噬和代谢应激的能力。机制上，LCL768诱导CerS1介导的内源性C18 - 神经酰胺在线粒体中积累以介导线粒体自噬，这不需要CerS1转运蛋白p17/PERMIT，但依赖于通过C644位点亚硝基化激活的DRP1。DRP1通过促进内质网（ER）中的磷脂酰乙醇胺与线粒体膜中的心磷脂之间的结合，促进内质网和线粒体膜的锚定。阻止Drp1与内质网和线粒体膜结合的突变会阻断CerS1/C18 - 神经酰胺在线粒体中的积累，抑制LCL768介导的线粒体自噬。此外，LCL768驱动的线粒体自噬改变了线粒体代谢，导致延胡索酸耗竭并在体内导致肿瘤抑制。外源性补充延胡索酸可预防LCL768介导的线粒体自噬、CerS1的线粒体转运、内质网 -线粒体连接以及小鼠中的肿瘤抑制。延胡索酸代谢与催化性半胱氨酸（Cys431）处的帕金森蛋白琥珀酰化有关，抑制其与PINK1和泛素的结合，从而阻止线粒体自噬。LCL768诱导的延胡索酸耗竭减弱了帕金森蛋白琥珀酰化以促进帕金森蛋白激活和线粒体自噬，表明一种通过帕金森蛋白琥珀酰化调节线粒体自噬和延胡索酸代谢的前馈机制。这些数据提供了一种机制，通过该机制LCL768/CerS1 - C18 - 神经酰胺介导的线粒体自噬和肿瘤抑制受Drp1亚硝基化、延胡索酸耗竭和帕金森蛋白琥珀酰化调节，为致死性线粒体自噬提供了一种代谢应激特征。
