Hypoxic regulation of macrophage migration and function via fibroblast reprogramming in pancreatic cancer

通过胰腺癌成纤维细胞重编程对巨噬细胞迁移和功能的缺氧调节

• 批准号: 10677376
• 负责人: Sean Matthew Hannifin
• 金额: $ 4.08万
• 依托单位: UNIVERSITY OF MICHIGAN AT ANN ARBOR
• 依托单位国家: 美国
• 财政年份: 2023
• 资助国家: 美国
• 起止时间: 2023-08-01 至 2027-07-31
• 项目状态: 未结题
• 来源: https://reporter.nih.gov/project-details/10677376
• 关键词: Area; Blood Vessels; CCL2 gene; Cancer Etiology; Cells; Cessation of life; Coculture Techniques; Communication; Data; Development; Disease; Disease Progression; Environment; Exclusion; Fibrinogen; Fibroblasts; Gene Expression; Goals; HIF1A gene; Human; Hypoxia; Hypoxia Inducible Factor; Immune; Immune System Diseases; Immunosuppression; Immunotherapeutic agent; Immunotherapy; Infiltration; Inflammatory; Macrophage; Malignant Neoplasms; Malignant neoplasm of pancreas; Mediating; Modeling; Mus; Myelogenous; Outcome; Oxygen; Pancreas; Pancreatic Ductal Adenocarcinoma; Patients; Phenotype; Play; Population; Regulation; Resistance; Role; Signal Transduction; Stress; Stromal Cells; Survival Rate; T-Lymphocyte; Testing; United States; cell type; chemokine; cytokine; density; effective therapy; experience; immunosuppressive macrophages; insight; migration; monocyte; neoplastic cell; neutralizing antibody; normoxia; pancreatic cancer cells; pancreatic neoplasm; pancreatic tumorigenesis; recruit; response; therapy resistant; transcription factor; tumor; tumor hypoxia; tumor progression; tumor-immune system interactions

PROJECT SUMMARY Pancreatic ductal adenocarcinoma (PDAC) is the third leading cause of cancer-related deaths in the United States despite representing only 2% of all cancer cases. PDAC is characterized by an extensive fibroinflammatory stroma and a hypoxic microenvironment, which contributes to disease progression and therapeutic resistance. Macrophages and cancer-associated fibroblasts (CAFs) are the predominant cell types within the PDAC stroma. Macrophages are a major immunosuppressive population in PDAC. These cells are highly plastic and, as a result, their environment plays an important role in regulating their function. Macrophages adapt to hypoxia, a condition of low oxygen availability, primarily through the stabilization of oxygen-liable transcription factors called hypoxia-inducible factors (HIFs). Although PDAC is extremely hypoxic and both the tumor cells and stromal cells experience hypoxia, the effects of hypoxia and HIFs on macrophages and their communication with other stromal cells in PDAC remain largely unknown. Our group has recently found that hypoxia promotes the acquisition of inflammatory cancer-associated fibroblasts (inflammatory CAFs), a recently defined PDAC fibroblast subtype that produces high levels of inflammatory cytokines and chemokines. By injecting a hypoxia probe into mice bearing PDAC, we have observed that inflammatory CAFs and macrophages predominantly reside in hypoxic tumor regions compared with normoxic regions whereas T cells are largely excluded from hypoxic tumor areas. Based on these preliminary data, I hypothesize that inflammatory fibroblasts induced by hypoxia promote macrophage infiltration into hypoxic tumor regions and facilitate an immunosuppressive macrophage phenotype. This hypothesis will be investigated with the following two Aims: (1) to determine the role of hypoxia-induced fibroblast-secreted factors in regulating macrophage migration and function in PDAC and (2) to define the role of macrophage HIFs in regulating macrophage migration and function in PDAC. To complete Aim 1, I will culture macrophages with conditioned media derived from mono- and co- cultures of pancreatic tumor cells and fibroblasts under either hypoxia or normoxia, and then assess the expression of immunosuppressive macrophage markers as well as their migration. To complete Aim 2, I will culture HIF1α-deficient and HIF2α-deficient macrophages under hypoxia with fresh media or conditioned media from co-cultures of pancreatic tumor cells and fibroblasts under hypoxia. Then I will assess the expression of immunosuppressive markers in these macrophages and evaluate their migration. Finally, I will define the role of macrophage HIFs in mediating pancreatic tumorigenesis using orthotopic tumors from mice lacking myeloid expression of HIF1ɑ or HIF2ɑ. My proposal will provide new insights into how hypoxia promotes an immunosuppressive PDAC microenvironment by modulating the macrophage-fibroblast crosstalk, and ultimately inform strategies to overcome resistance to immunotherapy.

项目总结 胰腺导管腺癌是美国癌症相关死亡的第三大原因。 尽管只占所有癌症病例的2%，但美国各州的癌症发病率仍然很高。PDAC的特点是具有广泛的 纤维炎症间质和低氧微环境，这有助于疾病的进展和 治疗抵抗力。巨噬细胞和肿瘤相关成纤维细胞(CAF)是主要的细胞类型 在PDAC基质内。巨噬细胞是PDAC的主要免疫抑制细胞群。这些细胞是 高度可塑性，因此，它们的环境在调节它们的功能方面发挥着重要作用。巨噬细胞 适应低氧，一种氧气利用率低的状态，主要是通过稳定易氧 转录因子称为缺氧诱导因子(HIF)。尽管PDAC极度缺氧，而且 肿瘤细胞和间质细胞经历低氧，低氧和低氧对巨噬细胞及其功能的影响 与PDAC中其他基质细胞的通讯在很大程度上仍不清楚。我们小组最近发现， 低氧促进炎性癌症相关成纤维细胞(炎性CAF)的获得，这是最近的一种 明确的PDAC成纤维细胞亚型可产生高水平的炎性细胞因子和趋化因子。通过 将低氧探针注射到携带PDAC的小鼠体内，我们观察到炎性CAF和巨噬细胞 与正常氧区相比，T细胞主要驻留在缺氧区，而T细胞主要分布在缺氧区 被排除在低氧肿瘤区域之外。根据这些初步数据，我推测炎性成纤维细胞 低氧诱导巨噬细胞向低氧肿瘤区侵袭 免疫抑制型巨噬细胞表型。对这一假设的研究将有以下两个目的： (1)确定低氧诱导的成纤维细胞分泌因子在调节巨噬细胞迁移和 在PDAC中的功能和(2)确定巨噬细胞HIFs在调节巨噬细胞移动和功能中的作用 在PDAC中。为了完成目标1，我将用来自单细胞和联合细胞的条件培养液培养巨噬细胞。 在低氧和常氧条件下培养胰腺肿瘤细胞和成纤维细胞，然后评估 免疫抑制巨噬细胞标志物的表达及其迁移。为了完成目标2，我会 用新鲜培养液和条件培养液在低氧条件下培养HIF1α缺陷和HIF2α缺陷巨噬细胞 来自胰腺肿瘤细胞和成纤维细胞在低氧条件下的共培养。然后我会评估一下 这些巨噬细胞中的免疫抑制标志物，并评估它们的迁移。最后，我将定义 巨噬细胞缺氧诱导因子在小鼠缺髓系原位肿瘤中的作用 HIF1ɑ或HIF2ɑ的表达。我的提案将提供新的见解，了解缺氧如何促进 通过调节巨噬细胞-成纤维细胞串扰的免疫抑制PDAC微环境，最终 告知克服免疫疗法耐药性的策略。