Targeting Mitochondrial Redox Capacity to Overcome Cancer Subtype that Regrowth After Radiation

摘要(R01CA251663) 放射治疗(RT)被广泛用于治疗局限性前列腺癌(PCa)。然而，癌细胞往往 通过未知的机制产生对RT的耐药性，导致癌症复发。为了改善RT，有以下几点 迫切需要发现导致细胞产生抵抗力的细胞事件。我们之前已经证明了 前列腺癌的异质性，特别是在具有丰富线粒体亚群的前列腺癌中，通常会存活下来 放疗后再生(称为耐辐射前列腺癌，或RR-PCA)。线粒体质量增加， 在RR-PCA细胞中获得数量、活性氧物种(ROS)和生物发生标志物。我们进一步 证明了线粒体生物发生调控因子TFAM(转录因子A， 线粒体)，可显著恢复RR-PCA细胞对RT的敏感性。因此，我们的首要假设是 RT激活的线粒体生物发生，通过ROS，是一种获取机制，推动PCa存活后- RT，这一前提将在拟议的研究中经过严格的审查。已知RO可直接和 通过融合、分裂、有丝分裂和生物发生来间接调节线粒体的动态平衡。我们放映了 FDA批准的药物，寻找对正常细胞无毒并有能力提高 阻断线粒体蛋白翻译时PCa细胞线粒体过氧化氢(MtH_2O_2)水平。 我们发现阿奇霉素(Azm)，一种大环内酯类抗生素，是一种有效的原型化合物，具有 两处房产都有。我们进一步证明，AZM与RT联合使用可通过 与Azm或RT相比，线粒体亚群丰富。因此，我们建议将我们的发现 并找出能有效抑制照射后癌细胞存活的机制(S)，提高RT 功效。其具体目的是：1)确定RT激活线粒体的分子机制(S) 生物发生促进线粒体丰富的前列腺癌细胞存活和代谢适应 体外和体内；2)确定mtH_2O_2超载是否针对固有线粒体和RT-获得性 逆转录获得的线粒体在阻断线粒体蛋白翻译的同时增强 RR-PCA细胞的辐射敏感性，以及3)使用线粒体过氧化氢生成器和线粒体蛋白改善RT 以AZM为原型的翻译抑制剂在小鼠原位异种移植模型和患者来源的异种移植中的应用 激活线粒体生物发生的前列腺癌模型。这项研究使用了最先进的平台，包括 反相蛋白质阵列、稳定同位素代谢组学、全内反射荧光 使用Imaris软件的显微成像、tempol增强的MRI成像和高分辨率O2k-FluoRespiroeter。 拟议的研究有望揭示同时靶向的新的分子洞察力 线粒体氧化还原能力和线粒体生物发生提高了RT治疗RR-PCa的疗效。

ABSTRACT (R01CA251663) Radiation therapy (RT) is widely used to treat localized prostate cancer (PCa). However, cancer cells often develop resistance to RT through unknown mechanisms, resulting in cancer recurrence. To improve RT, there is a dire need to uncover cellular events that cause cells to become resistant. We previously demonstrated that PCa heterogeneity, particularly in prostate cancers with an abundant mitochondria subpopulation, often survive and regrow after RT (termed radiation resistant prostate cancer, or RR-PCa). Elevation of mitochondrial mass, number, reactive oxygen species (ROS), and biogenesis markers is acquired in RR-PCa cells. We further demonstrated that knockdown of the mitochondrial biogenesis regulator, TFAM (transcription factor A, mitochondrial), significantly restored the sensitivity of RR-PCa cells to RT. Hence, our overarching hypothesis is that RT-activated mitochondrial biogenesis, via ROS, is an acquisition mechanism that drives PCa survival post- RT, a premise that will undergo stringent examination in the proposed studies. ROS are known to directly and indirectly regulate mitochondrial homeostasis through fusion, fission, mitophagy, and biogenesis. We screened FDA-approved drugs in search of compounds that are nontoxic to normal cells and have the ability to raise the level of mitochondrial hydrogen peroxide (mtH2O2) in PCa cells while blocking mitochondrial protein translation. We found azithromycin (AZM), a macrolide antibiotic, to be an effective prototype compound that possesses both properties. We further demonstrated that AZM combined with RT enhances the death of PCa cells with an abundant mitochondrial subpopulation, compared to AZM or RT alone. Thus, we propose to advance our findings and identify the mechanism(s) that effectively inhibit the survival of post-irradiated cancer cells, to improve RT efficacy. The specific aims are: 1) to define the molecular mechanism(s) by which RT-activated mitochondrial biogenesis promotes cell survival and metabolic adaptations of PCa cells with abundant mitochondria, both in vitro and in vivo; 2) to determine if overloading mtH2O2 to target inherent mitochondria and RT-acquired mitochondria while blocking mitochondrial protein translation in RT-acquired mitochondria enhances radiosensitivity of RR-PCa cells, and 3) to improve RT using a mtH2O2 generator and a mitochondrial protein translation inhibitor, AZM as prototype, in an orthotopic mouse xenograft model and a patient-derived xenograft model of PCa with activated mitochondrial biogenesis. This study uses state-of-the-art platforms including reverse phase protein array, stable isotope-resolved metabolomics, total internal reflection fluorescence microscopy with Imaris software, TEMPOL-enhanced MRI imaging, and a high resolution O2k-FluoRespirometer. The proposed studies are expected to uncover novel molecular insights by which concurrently targeting mitochondrial redox capacity and mitochondrial biogenesis improve RT efficacy in the treatment of RR-PCa.