Recent findings suggest that over-expression of activated H-ras inhibited apoptotic cell death by blocking the activity of apoptotic
endonuclease(s). This study was designed using antisense H-ras
oligodeoxynucleotides (ODN) to evaluate whether alterations of H-ras expression in BEL-7402 human
hepatocellular carcinoma cells could influence the induction of apoptosis in vitro and in vivo. We found that, in vitro, continuous suppression of H-ras expression could decrease the proliferation of BEL-7402 cells and inhibit H-ras-induced entry into S phase. In situ end labeling showed that a large number of cells underwent apoptotic cell death
after treatment with antisense H-ras ODN (P < 0.01), and gel electrophoresis of
DNA extracted from these cells demonstrated a typical
DNA ladder, characteristic of apoptosis. In vivo study indicated that pretreatment with antisense H-ras significantly retarded
tumor growth in comparison with the untreated controls or
tumors treated with non-specific ODN (P < 0.01, P < 0.01). In situ end-labeling revealed that pronounced apoptotic nuclei were also present in the tissue treated with antisense H-ras ODN (P < 0.01). Immunocyto-histochemical study showed that expression of p21H-ras was significantly decreased
after treatment with antisense H-ras. These results indicate that suppression of H-ras over-expression by antisense ODN could effectively inhibit
tumor growth and revive the apoptotic pathway by releasing the activity of apoptotic
endonuclease(s). The data also suggest the need for further studies to elucidate molecular events involved in antisense H-ras-released apoptosis and evaluate its therapeutic implications.