Defining metabolic adaptations within the PDAC "arid" tumor microenvironment

项目摘要 我们建议的目标是确定与营养不良条件相关的治疗弱点， 肿瘤的胰腺导管腺癌（PDAC）仍然是一个主要的临床挑战，其特征在于： 间质致密血管稀少由此产生的缺氧和营养缺乏迫使癌症 细胞寻找生长所需的其他营养来源。奇怪的是，这些营养物质中的细胞- 由于药物扩散不良， 和减少的细胞增殖（使DNA损伤剂不那么有效）。因此，为了靶向这些细胞， 更好地了解它们科普营养缺乏的适应机制是迫切需要的。 needed. 在初步研究中，我们开发了一种培养方案-限制氧气、葡萄糖、氨基酸和血清 - 来模拟胰腺肿瘤的“干燥条件”。在这样的条件下，肿瘤细胞会减慢 增殖和代谢活性。使用该系统，我们进行了全基因组CRISPR筛选， 确定在干旱条件下需要但在营养丰富（“肥沃”）条件下不需要的基因。这 揭示了对TCA循环和氧化磷酸化的强烈依赖性，但也表明 参与许多生物合成活动（如翻译和细胞分裂）的基因在干旱条件下是有害的。 条件基于这些结果，我们假设细胞在严重的营养和氧气剥夺下存活 通过从有限的燃料供应中最大限度地产生能量，同时最小限度地使用这些资源， 大分子生产我们进一步假设，营养和氧气剥夺改变了动态 癌细胞与构成肿瘤微环境（TME）的其他细胞之间的串扰。重要的是， 个体肿瘤可能以不同的方式对这些代谢变化作出反应，从而导致肿瘤间的相互作用。 异质性我们的提案将探讨这些想法，以制定针对耐药患者的策略。 癌细胞在这些缺乏营养的小生境中生存。 目标1.描述干旱条件下代谢适应的异质性。 目标二。确定干燥条件如何影响TME中的蜂窝串扰。 目标3：在体外和体内测试代谢抑制剂在干燥条件下对细胞的活性。

PROJECT SUMMARY The goal of our proposal is to identify therapeutic vulnerabilities associated with nutrient-poor conditions in tumors. Pancreatic ductal adenocarcinoma (PDAC) remains a major clinical challenge and is characterized by a dense stroma and paucity of blood vessels. The resulting hypoxia and scarcity of nutrients forces cancer cells to seek alternative sources of nutrients for growth. Paradoxically, cells residing in these nutrient- deprived microenvironments are among the most resistant to therapy as a result of poor drug diffusion and reduced cell proliferation (rendering DNA-damaging agents less effective). To target these cells, therefore, a better understanding of the adaptive mechanisms by which they cope with nutrient deprivation is urgently needed. In preliminary studies, we developed a culture protocol – limiting for oxygen, glucose, amino acids, and serum – to model the “arid conditions” of pancreatic tumors. Under such conditions, tumor cells slow down proliferation and metabolic activity. Using this system, we performed a whole genome CRISPR screen to identify genes required under arid conditions but dispensable under nutrient-replete (“fertile”) conditions. This revealed a strong dependency on the TCA cycle and oxidative phosphorylation for survival, but also suggested that genes involved in many biosynthetic activities (e.g. translation and cell division) are detrimental under arid conditions. Based on these results, we hypothesize that cells survive severe nutrient and oxygen deprivation by maximizing energy generation from limited fuel supplies while minimizing the use of such resources for macromolecule production. We further hypothesize that nutrient and oxygen deprivation alters the dynamic crosstalk between cancer cells and other cells comprising the tumor microenvironment (TME). Importantly, individual tumors may respond to these metabolic changes in different ways, contributing to inter-tumoral heterogeneity. Our proposal will explore these ideas to develop strategies that target therapy-resistant cancer cells residing in these nutrient-deprived niches. Aim 1. Characterize the heterogeneity in metabolic adaptations to arid conditions. Aim 2. Determine how arid conditions influence cellular crosstalk in the TME. Aim 3. Test the activity of metabolic inhibitors against cells in arid conditions in vitro and in vivo.