Hyperthermia improves the antitumour effect of metronomic cyclophosphamide in a rat transplantable brain tumour.

Abstract

As low-dose metronomic cyclophosphamide (CTX) and hyperthermia (HT) both exert antitumour effects in part through antiangiogenic mechanisms, interactive effects of the two modalities were explored. Subcutaneously implanted rat tumours (BT4An) were treated with CTX 35 mg/kg i.p. three doses a week for two weeks, local water-bath HT yielding mean tumour temperature of 43 degrees C for one hour at day 0, both modalities combined (CTX-HT(0)), or saline. TUNEL assays, immunohistochemical staining of thrombospondin 1 (TSP-1) and real time RT-PCR of TSP-1 mRNA were analysed the first three hours after completed treatment day 0. Metronomic dosed CTX (p=0.006) and HT (p<0.001) both delayed tumour growth. The combined regimen was superior to either modality alone (p<0.001). Complete tumour regressions were observed in CTX (6%), HT (12%), and CTX-HT(0) (41%) treated rats. TSP-1 protein was specifically upregulated in the vascular matrix of tumours receiving CTX (weak), HT (moderate) and CTX-HT(0) (strong). In contrast, reduced expression of TSP-1 protein was observed in tumour cells after HT alone and CTX-HT(0). TUNEL assays indicated induction of apoptosis by HT and CTX-HT(0) 90 minutes after end of the first treatment. A single session of local HT enhances the effects of low-dose metronomic CTX, possibly in part mediated through a differential effect on TSP-1 protein levels in tumour cells and tumour vasculature.