Secretory Mitophagy in Cancer Metabolic Reprogramming

癌症代谢重编程中的分泌性线粒体自噬

• 批准号: 10743163
• 负责人: Marissa Ashton Howard
• 金额: $ 20.45万
• 依托单位: GEORGE MASON UNIVERSITY
• 依托单位国家: 美国
• 财政年份: 2023
• 资助国家: 美国
• 起止时间: 2023-07-11 至 2025-06-30
• 项目状态: 未结题
• 来源: https://reporter.nih.gov/project-details/10743163
• 关键词: 4T1; Autophagocytosis; Autophagosome; Biogenesis; Breast Cancer cell line; Cell Survival; Cells; Chloroquine; Culture Media; DNA Damage; Docking; Fluorescence; Goals; Harvest; Human; Hypoxia; Immune; Immunotherapy; In Vitro; Intercellular Fluid; Knock-out; Knowledge; Length; Link; Lysosomes; MDA MB 231; Malignant Neoplasms; Mammary Neoplasms; Measurement; Metabolic; Methods; Mitochondria; Mitochondrial DNA; Mitochondrial Proteins; Molecular; Molecular Analysis; Monitor; Morphology; Mus; Needles; Organelles; Outcome; Oxidative Stress; Oxidative Stress Induction; PINK1 gene; Pathway interactions; Peripheral; Phenotype; Population; Process; Protein Export Pathway; Proteins; Publishing; Radiation therapy; Role; Sampling; Sirolimus; Solid Neoplasm; Stress; System; Testing; Therapeutic; aspirate; bafilomycin A1; cancer cell; cancer therapy; chemotherapy; clinically relevant; extracellular; extracellular vesicles; improved; in vivo; in vivo monitoring; knock-down; neoplastic cell; new therapeutic target; particle; pressure; prevent; real time monitoring; recruit; sensor; therapeutic target; therapy resistant; tool; triple-negative invasive breast carcinoma; tumor; tumor microenvironment; tumor progression

Tumor metabolic reprogramming is a hallmark of cancer progression, survival, and therapeutic resistance. A targetable class of cancer metabolic adaptation exploits mitophagy, a specialized autophagy pathway known to be linked to the cancer phenotype. Mitophagy selectively eliminates dysfunctional mitochondria by targeting them, via autophagosome shuttling, to the lysosome for degradation. Cancer cell mitophagy is triggered by elevated oxidative stress and mitochondria DNA damage caused by hypoxia, radiotherapy, molecular therapy, and immunotherapy. A high mitophagy demand can overwhelm the lysosome capacity resulting in the accumulation of damaged mitochondria that is harmful to the cell, and can suppress biogenesis of healthy mitochondria. We hypothesize that the newly discovered process of secretory mitophagy exports damaged mitochondrial fission-released segments to reduce the overload pressure on the lysosomal system, and thereby sustains cancer cell survival in the face of therapeutic mitochondrial stress. We have discovered a form of secretory mitophagy occurring in vivo in a growing solid tumor. Our molecular analysis of the full repertoire of extracellular vesicles (EV) shed into the resident tumor interstitial fluid (IF) in vivo yielded a rich set of information about the functional state of mitochondria within the tumor cells, and the host cells. Within tumor interstitial fluid, and within the culture media of cancer cells undergoing oxidative stress, we identified an EV-packaged full set of mitochondria molecules comprising the peripheral fission pinched-off segment of the mitochondrial organelle. It has recently been found that peripheral mitophagy fission, regulated by mitochondrial fission 1 protein (FIS1), is a key essential regulator of mitophagy, and is distinct from mid-zone mitochondria fission associated with mitochondria biogenesis. We also found that mitophagy inducer PTEN induced kinase 1 (PINK1) cleavage status (full length versus cleaved), is prominently reflected in the set of mitochondrial proteins exported within IF EVs, and may constitute a new quantitative measurement tool to monitor the real-time state of tumor intracellular mitophagy. Our findings raise important mechanistic questions, that we will explore in the Aims, concerning the unknown intracellular steps of secretory mitophagy and how the content of the exported EVs reflects the internal mitophagy state. Under Aim 1 we will test the hypothesis that peripheral fission secretory mitophagy is induced by high mitophagy demands that overwhelm lysosomal engulfment, and explore if secretory mitophagy is distinct from secretory autophagy within the murine 4T1 and human MDA-MB-231 triple negative breast cancer cell lines. Under Aim 2 we will test the hypothesis that the ratio of full length to cleaved PINK1 is elevated (mitochondria damage sensor) within the exported murine syngeneic breast tumor interstitial fluid EVs, following treatment with mitophagy inducers Mito-CP and Rapamycin. The outcome is new understanding of the importance of secretory mitophagy that can constitute an important therapeutic target, and a new clinically relevant means of monitoring the in vivo state of mitophagic flux within the tumor microenvironment.

肿瘤代谢重编程是癌症进展、存活和治疗抗性的标志。一 一种靶向的癌症代谢适应利用线粒体自噬，一种已知的专门的自噬途径， 与癌症表型有关。线粒体自噬通过靶向选择性消除功能障碍的线粒体 它们通过自噬体穿梭到达溶酶体进行降解。癌细胞的线粒体自噬是由 缺氧、放疗、分子治疗等引起的氧化应激和线粒体DNA损伤， 和免疫疗法。高线粒体自噬需求可压倒溶酶体能力，导致线粒体自噬。 积累受损的线粒体，这是有害的细胞，并可以抑制生物合成的健康 线粒体我们推测，新发现的分泌性线粒体自噬输出过程损害了 线粒体分裂释放的片段，以减少对溶酶体系统的过载压力，从而 维持癌细胞在治疗性线粒体压力下的存活。我们发现了一种 在体内生长的实体瘤中发生的分泌性线粒体自噬。我们对所有基因组的分子分析 细胞外囊泡（EV）脱落到体内驻留的肿瘤间质液（IF）中产生了丰富的信息 关于肿瘤细胞和宿主细胞内线粒体的功能状态。在肿瘤间质液中， 在经历氧化应激的癌细胞的培养基中，我们发现了一个EV包装的全套 包括线粒体细胞器的外周分裂夹断部分的线粒体分子。 最近发现，由线粒体分裂1蛋白（FIS 1）调节的外周线粒体自噬分裂， 是线粒体自噬的关键重要调节因子，不同于与线粒体自噬相关的中区线粒体裂变 线粒体生物发生我们还发现，线粒体自噬诱导剂PTEN诱导的激酶1（PINK 1）切割状态 (full长度与切割），显著地反映在IFEV内输出的线粒体蛋白质组中， 有望成为一种新的实时监测肿瘤细胞内状态的定量测量工具 线粒体自噬我们的研究结果提出了重要的机械问题，我们将在目标中探讨， 分泌性线粒体自噬的未知细胞内步骤以及输出EV的含量如何反映细胞内的线粒体自噬。 线粒体自噬状态在目标1下，我们将检验外周分裂分泌型线粒体自噬被诱导的假设。 通过高线粒体自噬的需求压倒溶酶体吞噬，并探讨分泌性线粒体自噬是否是独特的 来自鼠4 T1和人MDA-MB-231三阴性乳腺癌细胞内的分泌性自噬 线在目标2下，我们将检验全长与切割的PINK 1的比率升高的假设 （线粒体损伤传感器）在输出的鼠同基因乳腺肿瘤间质液EV中， 用线粒体自噬诱导剂Mito-CP和雷帕霉素处理。其结果是， 分泌性线粒体自噬的重要性，可以构成一个重要的治疗靶点，和一个新的临床 监测肿瘤微环境内的线粒体吞噬通量的体内状态的相关手段。