Regulation of the Hypoxia Inducible Factor (HIF)

描述(申请人提供)：哺乳动物细胞利用低氧反应途径来感知和响应氧气供应的变化。这一途径既有助于对低氧应激的生理适应，也有助于许多疾病状态的进展，在这些疾病中，氧气供应受到影响，包括缺血和癌症。低氧诱导转录因子(HIF)位于该途径的核心，参与调节多种肿瘤相关靶点的表达，包括糖酵解、细胞存活/增殖、血管生成和侵袭/转移相关基因。最近，我们的实验室和其他人已经证明，缺氧诱导因子的稳定性和辅助激活因子的募集分别受到氧依赖的脯氨基和天冬酰胺羟基酶的强烈调控。虽然HIF可以作为肿瘤内缺氧的功能而被诱导，但在癌症中，HIF活性经常被各种氧不依赖的机制上调，这些机制是由激活癌基因或灭活抑癌基因的基因改变引起的。这项提案的目标有两个。(1)我们将研究我们已经确定的HIF活性与候选肿瘤抑制基因生长抑制家族成员4(ING4)之间的关系，据报道，ING4可抑制胶质母细胞瘤的血管生成和肿瘤生长。为此目的提出了一种研究ING4介导的HIF抑制的组合方法(B)ING4通过HIF Pro羟基酶被招募到HIF的机制(C)ING4相关因子被招募到HIF和可能的其他转录因子以及(D)在人类肿瘤中ING4表达、HIF关联和HIF活性之间的关系。(2)HIF由两个亚基组成，每个亚基含有两个PAS结构域，它们在HIF功能中的作用尚未完全阐明。利用最近确定的HIF PAS-B结构域的溶液结构，我们将(A)通过体外分析和细胞培养模型研究HIF PAS结构域的结构/功能关系；(B)展示在细胞培养和小鼠肿瘤模型中通过改变PAS结构域来干扰HIF功能的潜在治疗功效；以及(C)开发直接与PAS结构域结合的小分子HIF拮抗剂。总之，这些研究将增加我们对HIF调控的理解，并可能为治疗干预提供新的途径。

DESCRIPTION (provided by applicant): Mammalian cells employ a hypoxic response pathway to sense and respond to changes in oxygen availability. This pathway contributes to both the physiological adaptation to hypoxic stress as well as the progression of a number of disease states in which oxygen supply is compromised, including ischemia and cancer. The Hypoxia Inducible transcription Factor (HIF) lies at the heart of this pathway and mediates expression of several targets relevant to tumorigenesis including genes involved in glycolysis, cell survival/proliferation, angiogenesis, and invasion/metastasis. Recently, our laboratory and others have shown that both HIF stability and coactivator recruitment are acutely regulated by oxygen-dependent prolyl and asparaginyl hydroxylases, respectively. While HIF can be so induced as a function of intratumoral hypoxia, HIF activity is frequently upregulated in cancers by a variety of oxygen-independent mechanisms resulting from genetic alterations that activate oncogenes or inactivate tumor suppressors. The objectives of this proposal are twofold. (1) We will investigate a relationship we've identified between HIF activity and a candidate tumor suppressor gene, Inhibitor of Growth family member 4 (ING4), reported to repress angiogenesis and tumor growth in glioblastomas. Proposed in this Aim is a combination approaches for the study of (a) ING4-mediated repression of HIF (b) the mechanism by which ING4 is recruited to HIF via the HIF prolyl hydroxylase (c) the identification of ING4-associated factors recruited to HIF and perhaps other transcription factors and (d) the relationship between ING4 expression, HIF association, and HIF activity in human tumors. (2) HIF is composed of two subunits, each of which contains two PAS domains whose roles in HIF function have not yet been fully elucidated. Using insights derived from the recently determined solution structures of the HIF PAS-B domains as a starting point we will (a) investigate the structure/function relationships of the HIF PAS domains via in vitro assays and cell culture models (b) demonstrate the potential therapeutic utility of disrupting HIF function through alterations to the PAS domains both in cell culture and in mouse tumor models and (c) pursue the development of small molecule HIF antagonists that directly bind the PAS domains. Together, these studies will increase our understanding of HIF regulation and may provide new avenues for therapeutic intervention.