Apoptosis, angiogenesis and proliferation: trifunctional measure of tumour response to radiotherapy for oral cancer

Abstract

Local recurrence is a significant problem following radiotherapy in oral carcinoma and hence there is a paramount need for predictive markers. This study therefore analysed the predictive value of pre-treatment status of angiogenesis, apoptosis, expression of apoptosis regulatory p53, bax and bcl-2 proteins as well as tissue proliferation in relation to tumour response to radiotherapy. Sixty-nine histologically defined invasive carcinoma lesions were included in the study. Extent of apoptosis was defined morphologically and by the TUNEL (Tdt-mediated dUTP biotin nick end labelling) assay. Expression of apoptosis regulatory p53, bax and bcl-2 proteins were evaluated by immunocytochemistry. Mutant p53 protein was detected using a mutant p53-specific ELISA. The extent of tissue proliferation was evaluated by cyclin D1 expression. Angiogenesis was evaluated by CD34 antigen expression. All patients were treated with radical radiotherapy and followed up for 36 months. High levels of p53 protein detected by immunocytochemistry were found to be associated with poor response to treatment or disease relapse. Detection of mutant p53 protein also showed significant association with poor prognosis. Low levels of angiogenesis had a correlation with recurrence status. Tumours showing less vascularisation as well as increased apoptosis had a poor prognosis. Expression of p53 and bcl-2 proteins showed direct correlation with angiogenesis. There was no correlation between clinical status and any of the experimental parameters with histopathological grades of invasive lesions. Presence of mutant p53 protein is suggestive of poor tumour response to radiotherapy. Expression of p53 and increased apoptosis in less vascularised tumours is associated with treatment resistance. A predictive assay based on these results designed to analyse individual tumour samples showed presence of apoptotic cells near the vasculature to be indicative of good prognosis, while absence of apoptotic cells or highly proliferative cells and/or expression of bcl-2 protein in cells around the vasculature to be an indicator of poor prognosis.