Amplification of tumor hypoxic responses by MIF-dependent HIF stabilization

MIF 依赖性 HIF 稳定作用放大肿瘤缺氧反应

• 批准号: 8230775
• 负责人: ROBERT A MITCHELL
• 金额: $ 29.79万
• 依托单位: UNIVERSITY OF LOUISVILLE
• 依托单位国家: 美国
• 财政年份: 2009
• 资助国家: 美国
• 起止时间: 2009-05-01 至 2014-02-28
• 项目状态: 已结题
• 来源: https://reporter.nih.gov/project-details/8230775
• 关键词: Ablation; Adenocarcinoma Cell; Apoptosis; Binding; Biochemical; COPS5 gene; Cancer Patient; Cell Line; Cells; Complement; Complex; Cullin Proteins; Data; Development; Disease Progression; Exhibits; Fibrinogen; Functional disorder; Gene Targeting; Genetic; Genetic Transcription; Growth; Growth Factor; Health; Human; Hypoxia; Hypoxia Inducible Factor; Image; Immigration; In Situ Nick-End Labeling; In Vitro; Injection of therapeutic agent; Laboratories; Lesion; Maintenance; Malignant Neoplasms; Malignant neoplasm of pancreas; Masks; Mediating; Metabolic; Migration Inhibitory Factor; Molecular; Mus; Mutant Strains Mice; Neoplasm Metastasis; Nude Mice; Oxygen; Oxygen measurement, partial pressure, arterial; PECAM1 gene; Pancreatic Adenocarcinoma; Pancreatic Ductal Adenocarcinoma; Pathology; Pathway interactions; Patients; Pimonidazole; Plasma; Post-Translational Protein Processing; Proteins; Relative (related person); Resistance; Role; SLC2A1 gene; Signal Transduction; Small Interfering RNA; Solid Neoplasm; Staining method; Stains; Subgroup; Therapeutic; Transgenes; Transgenic Mice; Tumor Burden; Vascular Endothelial Growth Factors; Work; Xenograft procedure; angiogenesis; bHLH-PAS factor HLF; base; cell motility; cytokine; density; environmental adaptation; extracellular; fibroblast migration inhibitory factor; human cancer mouse model; hypoxia inducible factor 1; in vivo; inhibitor/antagonist; mouse model; neovascularization; novel; outcome forecast; overexpression; pancreatic cancer cells; pancreatic neoplasm; phenylpyruvate tautomerase; prevent; protein degradation; response; transcription factor; tumor; tumor progression; tumor xenograft; tumorigenesis; ubiquitin ligase; ubiquitin-protein ligase

DESCRIPTION (provided by applicant): Pancreatic ductal adenocarcinoma (PDAC) has the grim distinction of being one of the most prognostically unfavorable human cancers. It has been suggested that low oxygen tensions within PDAC tumors contribute significantly to its metastatic potential and chemotherapeutic resistance. Hypoxia-inducible factors (HIFs) transcribe gene products that contribute to metabolic adaptation, neo-angiogenic potential, metastatic spread and therapeutic resistance. We recently discovered that macrophage migration inhibitory factor (MIF), one of the oldest cytokines ever described, is elevated in PDAC patient's plasma and induced by hypoxia-induced, HIF-1a-dependent transcription in pancreatic cancer cells. Intriguingly, MIF was found to reciprocally modulate hypoxia- induced HIF-1a stabilization. Several human PDAC cell lines transfected with siRNA against MIF recapitulate MIF-/- fibroblasts in defective hypoxia-induced HIF-1a stabilization and subsequent HIF-dependent transcription. Moreover, MIF-deficient PDAC cells display defective xenograft tumor outgrowth, less HIF-1a and significantly reduced microvascular density than MIF-containing xenografts. Finally, our results indicate that MIF promotes HIF-1a stability by functionally regulating a subunit of the COP9 signalosome, CSN5, in repressing a unique, HIF-1-specific ubiquitin E3 ligase. Our central hypothesis is that MIF overexpression in pancreatic adenocarcinoma lesions acts to modulate either the levels or activity of free CSN5 that, in turn, serves to repress or mask a unique, oxygen-independent ubiquitin E3 ligase specific for HIF-1a. Studies outlined in this application seek to identify the contribution and requirements for MIF in PDAC growth, pathophysiology and hypoxic adaptation. We will additionally delineate the precise mechanisms and molecular determinants involved in MIF-dependent HIF-1 stabilization. In order to fulfill the stated objectives of this application the following experimental aims are proposed: 1) Elucidate the basic mechanism by which MIF contributes to hypoxia-induced HIF-1a stabilization, 2) Define the molecular determinants and effectors of HIF-1 degradation in MIF-deficient cells, and 3) Determine if MIF contributes to pancreatic ductal adenocarcinoma growth, pathophysiology and hypoxic adaptation. PUBLIC HEALTH RELEVANCE: All solid tumors require microenvironmental adaptation throughout tumorigenesis. One of the hallmarks of this adaptive response is the development of intratumoral hypoxia that stimulates HIF-directed expression of pro-angiogenic/metastatic gene products. Our findings establish that one of these gene products, MIF, is elevated in the plasma of a subset of pancreatic cancer patients. We further describe a unique functional inter-relationship between the extracellular cytokine/growth factor MIF and the transcription factor, HIF-1a in pancreatic ductal adenocarcinoma (PDAC) cells. This point is demonstrated by our data showing that cells lacking MIF exhibit defective hypoxia and PHD inhibitor-induced HIF-1a stabilization and subsequent transcription of metabolic and angiogenic gene products. More importantly, xenograft mouse models of PDAC tumorigenesis reveal that MIF-deficient tumor outgrowth, HIF-1a expression and tumor-associated angiogenesis are severely impaired when compared to cells containing MIF. Because HIF expression contributes directly to tumor aggressiveness and cancers of the pancreas are reportedly very hypoxic, it is likely that MIF functionally contributes to tumor maintenance, environmental adaptation and ultimately prognosis of pancreatic adenocarcinoma patients.

描述（由申请人提供）：胰腺导管腺癌（PDAC）是预后最不利的人类癌症之一。有人认为，PDAC 肿瘤内的低氧张力对其转移潜力和化疗耐药性有显着影响。缺氧诱导因子（HIF）转录的基因产物有助于代谢适应、新血管生成潜力、转移扩散和治疗耐药。我们最近发现，巨噬细胞迁移抑制因子 (MIF) 是有史以来描述的最古老的细胞因子之一，在 PDAC 患者血浆中升高，并由胰腺癌细胞中缺氧诱导的 HIF-1a 依赖性转录诱导。有趣的是，MIF被发现可以相互调节缺氧诱导的HIF-1a稳定性。几种用针对 MIF 的 siRNA 转染的人 PDAC 细胞系在缺陷性缺氧诱导的 HIF-1a 稳定和随后的 HIF 依赖性转录中重现了 MIF-/- 成纤维细胞。此外，与含有 MIF 的异种移植物相比，MIF 缺陷的 PDAC 细胞表现出有缺陷的异种移植肿瘤生长、HIF-1a 减少以及微血管密度显着降低。最后，我们的结果表明，MIF 通过功能性调节 COP9 信号体 CSN5 的亚基来抑制独特的 HIF-1 特异性泛素 E3 连接酶，从而促进 HIF-1a 的稳定性。我们的中心假设是，胰腺腺癌病灶中的 MIF 过表达可以调节游离 CSN5 的水平或活性，而游离 CSN5 反过来又可以抑制或掩盖一种独特的、不依赖于氧的 HIF-1a 特异性泛素 E3 连接酶。本申请中概述的研究旨在确定 MIF 在 PDAC 生长、病理生理学和缺氧适应中的贡献和要求。我们还将进一步描述 MIF 依赖性 HIF-1 稳定作用的精确机制和分子决定因素。为了实现本申请的既定目标，提出以下实验目标：1) 阐明 MIF 促进缺氧诱导的 HIF-1a 稳定的基本机制，2) 定义 MIF 缺陷细胞中 HIF-1 降解的分子决定因素和效应因素，以及 3) 确定 MIF 是否有助于胰腺导管腺癌的生长、病理生理学和 缺氧适应。公共健康相关性：所有实体瘤在整个肿瘤发生过程中都需要微环境适应。这种适应性反应的标志之一是肿瘤内缺氧的发展，刺激 HIF 定向表达促血管生成/转移基因产物。我们的研究结果表明，其中一种基因产物 MIF 在部分胰腺癌患者的血浆中升高。我们进一步描述了胰腺导管腺癌细胞 (PDAC) 中细胞外细胞因子/生长因子 MIF 和转录因子 HIF-1a 之间独特的功能相互关系。我们的数据证明了这一点，即缺乏 MIF 的细胞表现出缺氧缺陷和 PHD 抑制剂诱导的 HIF-1a 稳定以及随后代谢和血管生成基因产物的转录。更重要的是，PDAC肿瘤发生的异种移植小鼠模型表明，与含有MIF的细胞相比，MIF缺陷的肿瘤生长、HIF-1a表达和肿瘤相关血管生成严重受损。由于 HIF 表达直接影响肿瘤的侵袭性，并且据报道胰腺癌的缺氧程度很高，因此 MIF 可能在功能上有助于胰腺腺癌患者的肿瘤维持、环境适应和最终预后。