Tumor necrosis factor-alpha sensitizes prostate cancer cells to gamma-irradiation-induced apoptosis.
LNCaP prostate cancer cells are highly resistant to induction of programmed cell death by y-irradiation and somewhat sensitive to the death-inducing effects of tumor necrosis factor (TNF)-alpha. Simultaneous exposure of LNCaP cells to TNF-alpha and 8 Gy of irradiation was synergistic and resulted in a 3-fold increase of apoptotic cells within 72 h compared to TNF-alpha alone. It appeared that TNF-alpha sensitized the cells to irradiation because, when cells were irradiated 24 h after exposure to TNF-alpha, increased cell death was observed. In contrast, irradiation delivered 24 h prior to TNF-alpha exposure did not result in more cell death than after TNF-alpha alone. TNF-alpha induced expression of its own mRNA, but TNF-alpha mRNA induction was neither induced nor enhanced by irradiation. Activation of the transcription factor nuclear factor kappaB can be induced by TNF-alpha and has a modulating antiapoptotic effect. But enhancement of TNF-alpha-induced cell death by irradiation did not result from altered activation of nuclear factor kappaB. TNF-alpha treatment of LNCaP cells resulted in partial activation of caspase-8 and -6 but not caspase-3. There was only minimal poly(ADP-ribose) polymerase cleavage seen in LNCaP cells after exposure to both TNF-alpha and irradiation at 72 h, a time when 60% of the cells were apoptotic. Experiments with peptide inhibitors of cysteine and serine proteases suggested that caspases were the predominant mediators of apoptosis induced by TNF-alpha alone but that serine proteases contributed significantly to cell death induced by TNF-alpha plus irradiation. TNF-alpha increased production of ceramide in LNCaP cells 48 h after exposure. Although irradiation alone had no effect on ceramide production in LNCaP cells, TNF-alpha plus irradiation induced significantly more ceramide than TNF-alpha alone. Ceramide production did not occur immediately after exposure to TNF-alpha, but rather was delayed such that ceramide levels were increased only 24 h after exposure to apoptotic stimuli. Moreover, non-toxic levels of exogenous C2-ceramide sensitized LNCaP cells to irradiation similarly to TNF-alpha, suggesting that one mechanism by which LNCaP cells were sensitized to irradiation was by increased intracellular ceramide. Hence, ceramide generation is a critical component in radiation-induced apoptosis in human prostate cancer cells. Inhibition of ceramide generation may provide a selective advantage in the development of radioresistance in prostate cancer.
Kimura
K
K
Department of Medicine, Lombardi Cancer Center, Georgetown University, Washington, DC 20007-2007, USA.
Bowen
C
C
Spiegel
S
S
Gelmann
E P
EP
D017209:Apoptosis / Q000187:drug effects / Q000528:radiation effects*; D020169:Caspases / Q000502:physiology; D002518:Ceramides / Q000096:biosynthesis; D005720:Gamma Rays; D006801:Humans; D008297:Male; D016328:NF-kappa B / Q000378:metabolism; D011471:Prostatic Neoplasms / Q000473:pathology* / Q000532:radiotherapy; D011836:Radiation Tolerance; D014409:Tumor Necrosis Factor-alpha / Q000096:biosynthesis / Q000494:pharmacology*; D019014:fas Receptor / Q000502:physiology
D002518:Ceramides; D016328:NF-kappa B; D014409:Tumor Necrosis Factor-alpha; D019014:fas Receptor; D020169:Caspases