Targeting Stromal Influences on BCKA Addiction in PDAC Tumors

靶向基质对 PDAC 肿瘤 BCKA 成瘾的影响

基本信息

批准号：
10581670
负责人：
THEODORE S LAWRENCE
金额：
$ 57.97万
依托单位：
UNIVERSITY OF MICHIGAN AT ANN ARBOR
依托单位国家：
美国
项目类别：
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/10581670
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 Deamination Dependence Ecosystem Epithelium Event Extracellular Matrix Extracellular Matrix Degradation Failure Fibroblasts Foundations Genes Genetic Goals Growth In Vitro Isotopes Keto Acids Knockout Mice Knowledge 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 Starvation Stromal Cells Symbiosis System Testing Time Tissues Tumor Volume addiction amino acid metabolism branched-chain-amino-acid transaminase cancer cell cancer therapy carcinogenesis cell growth cell transformation conditional knockout drug development efficacy evaluation 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

项目摘要

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局限性了解肿瘤微环境 (TME) 如何促进快速进展或复发的 PDAC。在 PDAC 中，被确定为癌症相关成纤维细胞 (CAF) 的基质细胞占肿瘤体积的90%。最近的研究揭示了支链氨基酸的重要性（支链氨基酸）在癌症中的作用。然而，基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。了解癌症-间质生态系统需要深入了解癌症-间质的交叉点基质中的转化与其支链氨基酸代谢的重新编程相关。 SMAD4 缺失是 PDAC 中频繁发生的事件。然而，SMAD4 缺失的 PDAC 细胞在调节基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差距。我们提出了 PDAC 肿瘤的癌症和基质室中支链氨基酸代谢的差异揭示 PDAC 细胞对基质中 CAF 分泌的支链酮酸 (BCKA) 的严重依赖丰富的肿瘤。我们假设反应性基质 BCAA 代谢从静止基质中发生改变，并且是通过分泌 BCKA 调节 PDAC 细胞生长的驱动力。其次，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 合成代谢途径作为患者来源的循环肿瘤细胞（CTC）类器官和患者来源的组织切片中的致命靶标，使用基于 13C 的同位素示踪和遗传工具，并揭示了 CAF 分泌的细胞外基质 (ECM) 是一种 BCKA 的来源。为了了解 ECM 降解背后的机制，我们将调节蛋白酶体 CTC 类器官和患者来源的组织切片中 ECM 的蛋白水解。其次，阐明 SMAD4 缺失在 PDAC 细胞诱导的 CAF 代谢重连中，我们提出了功能丧失研究并使用 CTC- 类器官和组织切片证明 SMAD4 缺失的 PDAC 细胞对基质的依赖性增加基质 BCAT1 并阐明 SMAD4 缺失的 CTC 类器官随时间的时间代谢重编程类器官形成的过程。第三，我们将测试 CAF 中针对 BCKA 合成代谢的功效条件敲除小鼠并使用患者进行针对基质 BCKA 合成代谢途径的体内研究衍生的低传代细胞系和同基因同种异体移植模型。重要的是，我们将抑制基质 BCAT1 使用加巴喷丁、Erg 240（来自 Ergon Pharma 的 BCAT1 抑制剂）、ERG 245 (BCAT1/2 来自 Ergon Pharma 的抑制剂），以及 CAF 中 BCAT1 的 CRISPR 靶向。值得注意的是，我们提出的目标将揭示活性基质在癌症中以支链氨基酸代谢为中心的新型调节作用，并将揭示其潜在的机制本法规的作用方式。这些发现可以带来针对沟通的新疗法癌细胞与其微环境之间的关系。