Identification of Hypoxia-Regulated Proteins in Head and Neck Cancer by Proteomic and Tissue Array Profiling

Abstract

Hypoxia within solid tumors decreases therapeutic efficacy, and identification of hypoxia markers may influence the choice of therapeutic modality. Here, we used a proteomic approach to identify hypoxia-regulated proteins and validated their use as endogenous indicators of tumor hypoxia. Using two-dimensional gel electrophoresis and PowerBlot (antibody-based array), we identified a group of 20 proteins that are increased >1.5-fold during hypoxia. The majority of these proteins such as IB kinase ␤ (IKK␤), MKK3b, highly expressed in cancer (HEC), density-regulated protein 1, P150 glued , nuclear transport factor 2, binder of ARL 2, Paxillin, and transcription termination factor I have not been previously reported to be hypoxia inducible. The increase in these proteins under hypoxia was mediated through posttranscriptional mechanisms. We additionally characterized the role of IKK␤, a regulator of the nuclear factor-B transcription factor, during hypoxia. We demonstrated that IKK␤ mediates cell survival during hypoxia and is induced in a variety of squamous cell carcinoma cell lines. Furthermore, we showed that IKK␤ expression from tumor specimens correlated with tumor oxygenation in patients with head and neck squamous cell carcinomas. These data suggest that IKK␤ is a novel endogenous marker of tumor hypoxia and may represent a new target for anticancer therapy.

Prolyl hydroxylase-1 negatively regulates IκB kinase-β, giving insight into hypoxia-induced NFκB activity

“…The presence of an HIF-␣ LxxLAP consensus motif in IKK␤ led us to examine the expression of these proteins under hypoxia. In line with previous studies (15), HeLa cells exposed to graded hypoxia for 24 h demonstrated increased IKK␤ protein levels, whereas IKK␥/ NEMO (which does not contain the hydroxylation motif) did not increase (Fig. 5A).…”

The Unfolded Protein Response: A Novel Component of the Hypoxic Stress Response in Tumors

“…Although HIF controls the expression of many hypoxiaregulated genes, expression profiling studies conducted under hypoxia (21)(22)(23)(24)(25)(26)(27)(28) reveal that HIF activation alone cannot account for the full repertoire of changes that occur intracellularly as oxygen becomes limiting. Hypoxic cells elicit additional HIF-1-independent adaptive responses that contribute to increased survival under low oxygen conditions.…”

Head and neck cancer

“…Chen et al identified the hypoxia-regulated proteins in HNC by proteomic and tissue array profiling. 81 Hypoxia is a major determinant of local, regional, and distant failure after anticancer therapy. 82 At the molecular level, hypoxia selects tumors with an increased malignant phenotype, 83 resulting in resistance to apoptosis and greater propensity for distant metastases.…”

“…In human HNSCC, expression of this protein appeared to be a good indicator of tumor hypoxia and may represent a novel anticancer therapeutic target. 81 Studying the normal to tumor transition of esophageal SCC, Chung et al quantified the change in 7 proteins in this tissue transition using multiplex tissue immunoblotting. 84 Their data suggest that decreased expression of pan-cytokeratin (CK)-4, CK-14, and annexin 1 are early events.…”