Regulation of Hypoxia-inducible Factor 1 by Cyclin Dependent Kinases

Pennsylvania State Univ Hershey Med Ctr, Hershey PA

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Abstract

DESCRIPTION (provided by applicant): The hypoxia-inducible factor 1 (HIF-1) transcription factor is a critical mediator of the cellular response to oxygen deprivation. The ? subunits of HIF-1 are rapidly degraded in the presence of oxygen, but are stabilized under hypoxic conditions and activate the transcription of genes that promote angiogenesis, survival, invasion, and glucose metabolism. HIF-1 can be activated by genetic alterations that enhance the transcription or inhibit the degradation of HIF-1?. For example, von Hippel-Lindau (VHL) disease predisposes sufferers to aggressive tumors of the renal system, which are characterized by the stabilization of HIF-1?. The constitutive expression of HIF-1? drives the development of these tumors, which are highly angiogenic and resistant to therapy. Thus, HIF-1 inhibition represents a promising strategy for the treatment of patients with hypoxic and VHL-deficient tumors. While the oxygen-dependent regulation of HIF-1? has been well studied, the oxygen- independent regulation of HIF-1? is not completely understood. We have evidence that the stability of HIF-1? is regulated by cyclin dependent kinases (CDKs), specifically CDK1 and CDK4. The central hypothesis driving this research is that HIF-1? expression is regulated by CDKs, and identification of the molecular mechanisms underlying this regulation will produce novel strategies to inhibit tumor survival, angiogenesis, and therapeutic resistance associated with HIF-1 activation. We will address this hypothesis via the following aims: 1] Identify the mechanism by which CDKs regulate HIF-1? expression. 2] Test the anti-tumor efficacy of CDK inhibition on HIF-1? expressing tumors, and identify synergistic combinations with approved anticancer therapies. This proposal relies on three major approaches: 1] monitoring the expression levels and activity of HIF- 1 in response to CDK inhibition, 2] the use of mass spectrometry to identify of novel post-translational modifications that alter HIF-1? stability, and 3] utilizing CDK inhibitors in HIF-1? expressing tumors in combination with standard therapies. Preliminary data indicate that inhibition of CDK kinase activity significantly decreases the stability of HIF-1? at the protein level. Therefore, we plan to perform mass spectrometry and biochemical analyses to identify novel post-translational modifications on HIF-1? and determine the mechanism through which CDKs regulate HIF-1a expression. Next, we will investigate the efficacy of CDK inhibition as a novel therapeutic approach to target renal tumors that constitutively express HIF-1? due to the loss of VHL. First, in vitro assays will be used to assess the efficacy of CDK inhibitors and identify promising combinations with standard anticancer therapies. Then, in vivo tumor models will serve to determine if CDK-mediated repression of HIF-1? inhibits tumor growth and angiogenesis, and sensitizes HIF-1?-driven tumors to undergo cell death in response to standard therapies. Ultimately, the goal of this proposal is to facilitate the clinical translation of CDK inhibitors for use in tumors with constitutively active IF-1 while also yielding biological information about how HIF-1? stability is regulated by CDKs.