Immunohistochemical evaluation of tumor hypoxia and angiogenesis: Pathological significance and prognostic role in head and neck squamous cell carcinomas

Abstract

Objectives: Tumor hypoxia and angiogenesis have been implicated in therapeutic resistance of head and neck squamous cell carcinomas (HNSCCs). Immunohistochemical evaluation of hypoxia-inducible factor-1 alpha (HIF-1 α), a hypoxia transcription factor, and vascular endothelial growth factor (VEGF), a hypoxia-responsive pro-angiogenic factor, can be exploited for prognostication and guiding treatment intensification or de-escalation decisions in HNSCC patients. The purpose of the study is to evaluate the immunohistochemical expression patterns of HIF-1 α and VEGF and the microvessel density (MVD) for angiogenesis in HNSCC and assess their pathological significance and prognostic role. Materials and Methods: In this cross-sectional study, immunohistochemical expression of HIF-1 α, VEGF, and MVD through Cluster of Differentiation (CD31) was evaluated in paraffin-embedded tumor resection tissue of 44 patients with HNSCC. Associations among HIF-1 α, VEGF, and MVD with clinicopathological variables were assessed. Statistical Analysis: For assessment of association between HIF-1α, VEGF and MVD by CD 31 immunohistochemical markers and other clinicopathological variables Pearson’s chi-square test and Fisher’s exact tests were used. Analysis of survival was done using Kaplan-Meier statistics. Also, the univariate and multivariate analysis were performed using the Cox proportional hazard regression model for the calculation of hazard ratios. Results: Nuclear expression of HIF-1 α showed significant association with MVD (P = 0.007) and cytoplasmic expression of HIF-1 α with histologic grade (P = 0.03). Overexpression of HIF-1 α was more frequent in T3/T4 stage. In addition to cytoplasmic staining, VEGF showed a unique nuclear expression pattern in four cases of advanced disease with nodal metastasis. Logistic regression analysis showed tumors with nuclear overexpression of HIF-1 α to have increased MVD (P = 0.05), and tumors with higher MVD to have a presence of lymphovascular invasion (P = 0.014). Multivariate analysis showed HIF-1 α nuclear overexpression to be significantly associated with decreased survival of patients (P = 0.05). Conclusions: Immunohistochemical overexpression of HIF-1 α and MVD quantification can serve as cost-effective tools for prognostication and treatment modification of HNSCC patients in resourcelimited settings.