Targeting Stromal Influences on BCKA Addiction in PDAC Tumors

靶向基质对 PDAC 肿瘤 BCKA 成瘾的影响

• 批准号: 10453984
• 负责人: THEODORE S LAWRENCE
• 金额: $ 58.14万
• 依托单位: UNIVERSITY OF MICHIGAN AT ANN ARBOR
• 依托单位国家: 美国
• 财政年份: 2022
• 资助国家: 美国
• 起止时间: 2022-03-01 至 2027-02-28
• 项目状态: 未结题
• 来源: https://reporter.nih.gov/project-details/10453984
• 关键词: Allografting; Anabolism; Branched-Chain Amino Acids; Cell Line; Cells; Cessation of life; Citric Acid Cycle; Clustered Regularly Interspaced Short Palindromic Repeats; Communication; Complex; Coupling; Cytosol; Data; Dependence; Ecosystem; Epithelial; Event; Extracellular Matrix; Extracellular Matrix Degradation; Failure; Fibroblasts; Foundations; Genes; Genetic; Goals; Growth; In Vitro; Isotopes; Keto Acids; Knockout Mice; Knowledge; Lead; Lipids; MADH4 gene; Malignant Neoplasms; Malignant neoplasm of pancreas; Metabolic; Metabolic Pathway; Methods; Mitochondria; Modeling; Nature; Neoplasm Circulating Cells; Neoplasm Metastasis; Nucleotides; Nutrient; Organoids; Oxidoreductase; Pancreas; Pancreatic Ductal Adenocarcinoma; Pathway interactions; Patients; Play; Production; Proteolysis; Public Health; Reaction; Recurrence; Regulation; Role; Sampling; Slice; Source; Stromal Cells; Symbiosis; System; Testing; Time; Tissues; Tumor Volume; addiction; amino acid metabolism; base; branched-chain-amino-acid transaminase; cancer cell; cancer therapy; cell growth; cell transformation; conditional knockout; drug development; efficacy testing; gabapentin; in vivo; in vivo Model; inhibitor; insight; loss of function; new therapeutic target; novel; oxidation; pancreatic cancer cells; pancreatic ductal adenocarcinoma cell; pancreatic ductal adenocarcinoma model; tool; traditional therapy; tumor; tumor metabolism; tumor microenvironment

Failure of traditional therapy in pancreatic ductal adenocarcinoma (PDAC) is due to our limited understanding of how the tumor microenvironment (TME) can facilitate the rapid progression or recurrence of PDAC. In PDAC, the stromal cells identified as cancer associated fibroblasts (CAFs) account for up to 90% of tumor volume. Recent studies have revealed the significance of branched chain amino acids (BCAAs) in cancer. However, the role of stromal cells in support of BCAA metabolism in tumors is still poorly understood. Understanding cancer-stromal ecosystem requires insight into the intersection of cancer- associated transformations in the stroma with reprogramming of their BCAA metabolism. SMAD4 deletion is a frequent event in PDAC. However, the metabolic role of SMAD4-deleted PDAC cells in modulating activation of stromal cell transformation is unknown. Our proposed aims here have aimed to bridge this knowledge gap. We propose a differential BCAA metabolism in cancer and stromal compartments of PDAC tumors and uncover a heavy reliance of PDAC cells on CAF-secreted branched chain keto acids (BCKAs) in stromal- rich tumors. We hypothesize that reactive stromal BCAA metabolism is altered from quiescent stroma and is the driver for regulating PDAC cell growth by secreting BCKAs. Second, SMAD4-deleted PDAC cells induces metabolic alterations in stromal cells. First, we will validate stromal BCKA synthesis metabolic pathway as a lethal target in patient-derived circulating tumor cell (CTC)-organoids and patient-derived tissue slices, using 13C-based isotope tracing and genetic tools and unravel that CAF-secreted extracellular matrix (ECM) is a source for BCKA. To understand the mechanism behind ECM degradation, we will regulate proteasomal proteolysis of ECM in CTC-organoids and patient-derived tissue slices. Second, to elucidate SMAD4-deletion in PDAC cells induced metabolic rewiring of CAFs we have proposed loss-of-function studies and use CTC- organoids and tissue slices to demonstrate increased stromal-reliance of SMAD4-deleted PDAC cells on stromal BCAT1 and elucidate temporal metabolic reprogramming in SMAD4-deleted CTC-organoids over time course of organoid formation. Third, we will test the efficacy of targeting BCKA anabolism in CAFs with conditional knockout mice and perform in vivo studies targeting stromal BCKA anabolic pathway using patient- derived low-passage cell lines and the syngeneic allograft model. Importantly, we will inhibit stromal BCAT1 in the proposed Aims using Gabapentin, Erg 240 (BCAT1 inhibitor from Ergon Pharma), ERG 245 (BCAT1/2 inhibitor from Ergon Pharma), & CRISPR targeting of BCAT1 in CAFs. Notably, our proposed aims will reveal a novel BCAA metabolism-centric regulatory role of reactive stroma in cancers and will uncover the underlying mode of action of this regulation. These findings can lead to novel therapeutics targeting communication between cancer cells and their microenvironment.

胰腺导管腺癌(PDAC)的传统治疗失败是由于我们的有限 了解肿瘤微环境(TME)如何促进快速进展或复发 PDAC的性能。在PDAC中，被鉴定为癌症相关成纤维细胞(CAF)的基质细胞占高达 占肿瘤体积的90%。最近的研究揭示了支链氨基酸的意义 (BCAAs)在癌症中。然而，间质细胞在肿瘤中支持支链氨基酸代谢的作用仍然很差。 明白了。理解癌症-间质生态系统需要洞察癌症的交叉点- 间质中的转化与其支链氨基酸代谢的重新编程有关。删除Smad4是一种 PDAC中的频繁事件。然而，Smad4缺失的PDAC细胞在调节细胞活化中的代谢作用 间质细胞转化尚不清楚。我们在这里提出的目标旨在弥合这一知识鸿沟。 我们提出了在PDAC肿瘤的癌和间质间质中BCAA的不同代谢和 揭示了PDAC细胞对基质中分泌的支链酮酸(BCKA)的严重依赖。 丰富的肿瘤。我们假设反应性间质BCAA代谢是从静止的间质改变而来的 通过分泌BCKA调节PDAC细胞生长的驱动力。第二，Smad4缺失的PDAC细胞诱导 基质细胞的代谢改变。首先，我们将验证基质BCKA合成代谢途径作为一种 病人来源的循环肿瘤细胞(CTC)有机物和患者来源的组织切片中的致死靶，使用 基于13C的同位素示踪和遗传工具以及解开CAF分泌的细胞外基质(ECM)是 BCKA的来源。为了了解细胞外基质降解的机制，我们将调节蛋白酶体。 细胞外基质在CTC-有机物和患者来源的组织切片中的蛋白分解。第二，阐明Smad4-缺失 在PDAC细胞诱导的CAF代谢重排中，我们提出了功能丧失研究，并使用CTC- 器官和组织切片显示Smad4缺失的PDAC细胞对基质的依赖增加 基质BCAT1和阐明Smad4缺失的CTC-有机物随时间的时间代谢重编程 有机体形成的过程。第三，我们将测试在CAF中靶向BCKA合成代谢的效果 条件基因敲除小鼠，并使用患者-BCKA合成代谢途径进行体内靶向研究 来源的低传代细胞系和同种异体移植模型。重要的是，我们将抑制基质BCAT1在 建议的AIMS使用Gabapentin，ERG 240(Ergon Pharma的BCAT1抑制剂)，ERG 245(BCAT1/2 Ergon Pharma的抑制剂)，以及在CAF中针对BCAT1的CRISPR靶向。值得注意的是，我们提出的目标将揭示 反应性间质在癌症中以支链氨基酸代谢为中心的新调节作用，并将揭示潜在的 这一规定的行动方式。这些发现可能导致针对沟通的新疗法 癌细胞与其微环境之间的关系。