Mechanistic insights into lysosomal nutrient efflux in cancer

癌症中溶酶体营养物流出的机制见解

• 批准号: 10341120
• 负责人: Kacper Rogala
• 金额: $ 11.51万
• 依托单位: WHITEHEAD INSTITUTE FOR BIOMEDICAL RES
• 依托单位国家: 美国
• 财政年份: 2021
• 资助国家: 美国
• 起止时间: 2021-02-04 至 2022-08-31
• 项目状态: 已结题
• 来源: https://reporter.nih.gov/project-details/10341120
• 关键词: Affect; Amino Acids; Area; Arginine; Autophagocytosis; Binding; Biochemical; Biological Assay; Biology; Cell physiology; Chemicals; Choking; Coupled; Cryoelectron Microscopy; Cytosol; Cytotoxic Chemotherapy; DNA; Development; Eating; Environment; Essential Amino Acids; Extracellular Protein; Face; Food; Future; Gatekeeping; Goals; Growth; Human; Impairment; In Vitro; Institutes; Integral Membrane Protein; Laboratories; Lead; Learning; Leucine; Libraries; Lipids; Lysosomes; Malignant Neoplasms; Malignant neoplasm of pancreas; Mammalian Cell; Mediating; Medicine; Membrane Proteins; Membrane Transport Proteins; Mentors; Metabolic; Molecular; Molecular Conformation; Molecular Machines; Motivation; Mutation; Normal Cell; Nutrient; Oncogenic; Outcome; Pathway interactions; Pharmaceutical Chemistry; Pharmaceutical Preparations; Phase; Phosphotransferases; Positioning Attribute; Process; Proteins; Protocols documentation; Recording of previous events; Research; Research Personnel; Role; Scientist; Signal Transduction; Starvation; Structure; Testing; Training; Transport Process; Validation; Work; X-Ray Crystallography; analog; base; cancer cell; cancer type; career; chemotherapy; diagnostic tool; drug development; drug discovery; effective therapy; expectation; experimental study; extracellular; fighting; improved; innovation; insight; lysosomal proteins; mTOR protein; metabolomics; mutant; nanobodies; nanodisk; novel; novel therapeutics; pancreatic cancer cells; prevent; programs; proteoliposomes; rational design; reconstitution; screening; small molecule; small molecule inhibitor; small molecule libraries; structural biology; success; therapeutic target; tumor; tumor metabolism; tumor microenvironment

ABSTRACT Many RAS-transformed aggressive cancer cells are able to escape cytotoxic chemotherapy and survive in near-starvation conditions. One adaptation making them hard to kill is their ability to scavenge extracellular proteins and recycle the cellular components using autophagy, both of which are then digested in lysosomes to recover free amino acids. The process of scavenging and internalization is known as macropinocytosis, and cancer cells aquire mutations to upregulate it when faced with nutrient-poor conditions. To fight such cancers, researchers are currently targeting the macropinocytosis machinery; however, because this process is not very well studied, and likely involves hundreds of proteins with redundant functions, such therapy might prove challenging to exectute without involving multiple drugs. We propose a better way of targeting nutrient-scavenging cancers by focusing on a downstream process of releasing digested nutrients from lysosomes to cytosol. The Sabatini Lab showed that the release of digested amino acids from lysosomes is orchestrated by the mTORC1 pathway, and specifically by SLC38A9. This lysosomal membrane protein senses the rising levels of digested amino acids in lysosomes by directly binding arginine. Our lab found that this sensing is coupled to activation of the transporter function, and results in the efflux of essential non-polar amino-acids, such as leucine, from lysosomes to cytosol. Importantly, RAS-transformed pancreatic cancer cells that feed on extracellular protein were unable to efficiently form tumors in the absence of SLC38A9. These results present a novel therapeutic idea of targeting a metabolic vulnerability in cancers transformed by oncogenic RAS signaling. In this five-year project we will elucidate the molecular mechanism of releasing digested amino acids from lysosomes to cytosol via SLC38A9, and therefore provide a rational approach to drug discovery. In parallel to that, we will screen for small molecules that specifically bind to SLC38A9, and develop them into chemical probes that modulate its transport activity. Impaired efflux function of SLC38A9 will lead to entrapment of macropinocytosis-derived amino-acids within the lysosomes, and our expectation is that this treatment will impair the growth of RAS-mutant and other tumors addicted to protein scavenging, while sparing normal cells that lack this requirement. Over the first two years of the mentored phase, I will be based at the Whitehead Institute, where I will learn cell signaling and metabolomics approaches from the experts in the field. I will also venture into a completely new research area to me, chemical biology, working with experts at the Broad Institute. After the completion of my K99 training, my aspiration is to lead a laboratory that combines cell signaling, structural biology, and chemical biology to study membrane transporters and their role in cancer metabolism. In parallel to understanding basic biology, I want my lab to develop specific small-molecule modulators that adjust transport activities of those proteins, facilitating further research in the field, and in long term – new medicines.

摘要 许多RAS转化的侵袭性癌细胞能够逃脱细胞毒性化疗并在体内存活。 近乎饥饿的状态使它们难以被杀死的一个适应是它们能够在细胞外生长 蛋白质，并利用自噬回收细胞成分，然后两者都在溶酶体中消化， 回收游离氨基酸。清除和内化的过程被称为巨胞饮， 癌细胞在面对营养不良的条件时会发生突变以上调其表达。为了对抗这些癌症， 研究人员目前的目标是巨胞饮机制;然而，由于这一过程不是很 经过充分研究，可能涉及数百种具有冗余功能的蛋白质，这种疗法可能会证明 在不涉及多种药物的情况下执行具有挑战性。我们提出了一个更好的定位方法 营养素清除癌症通过集中在下游过程中释放消化的营养素， 溶酶体到胞质溶胶。萨巴蒂尼实验室表明，从溶酶体中释放消化的氨基酸是 由mTORC 1途径，特别是SLC 38 A9协调。这种溶酶体膜蛋白 通过直接结合精氨酸来感知溶酶体中被消化的氨基酸水平的上升。我们的实验室发现 这种感觉与转运蛋白功能的激活相耦合，并导致必需的非极性 氨基酸，如亮氨酸，从溶酶体到胞质溶胶。重要的是，RAS转化的胰腺癌 以细胞外蛋白质为食的细胞在不存在SLC 38 A9的情况下不能有效地形成肿瘤。 这些结果提出了一种新的治疗思路，即靶向通过代谢途径转化的癌症中的代谢脆弱性。 致癌RAS信号传导。在这个为期五年的项目中，我们将阐明释放的分子机制， 通过SLC 38 A9将氨基酸从溶酶体消化到胞质溶胶，因此提供了一种合理的方法， 药物发现与此同时，我们将筛选特异性结合SLC 38 A9的小分子， 将其发展成化学探针来调节其运输活动。SLC 38 A9的外排功能受损将 导致大胞饮作用衍生的氨基酸包埋在溶酶体内，我们的预期是， 这种治疗将损害RAS突变体和其他依赖蛋白质清除的肿瘤的生长， 而不伤害缺乏这种需要的正常细胞。在指导阶段的前两年，我将根据 在Whitehead研究所，我将从专家那里学习细胞信号和代谢组学方法， 外地我也将冒险进入一个对我来说完全新的研究领域，化学生物学，与专家一起工作 在布罗德研究所在完成K99培训后，我的愿望是领导一个实验室， 结合细胞信号，结构生物学和化学生物学研究膜转运蛋白及其作用 在癌症代谢中的作用。在理解基础生物学的同时，我希望我的实验室能开发出 调节这些蛋白质转运活性的小分子调节剂，促进了对 从长远来看，新药。