Amplification of tumor hypoxic responses by MIF-dependent HIF stabilization

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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.