Prostate cancer is one of the most common cancers in men and is the second leading cause of cancer-related deaths in the USA. Many anti-tumor agents against prostate cancer cells have been developed, but their unacceptable systemic toxicity to normal tissues frequently limits their usage in clinics. Several previous studies have demonstrated that tumor necrosis factor-related apoptosis-inducing ligand (TRAIL) can induce cell death in a variety of transformed cells including prostate cancer cells, but not normal cells. Indole-3-carbinol (I3C), a phytochemical that is produced in fruits and vegetables, may play an important role in the prevention of many types of cancer, including hormone-related ones such as breast and prostate cancer. In this study, we examined the potential sensitizing effects of I3C on TRAIL-mediated apoptosis in a prostate cancer cell line, LNCaP. When LNCaP cells were incubated with I3C (either 30 or 90 microM) for 24 h and then treated with TRAIL (100 ng/ml), enhanced TRAIL-mediated apoptosis was observed. The enhanced apoptosis measured by poly(ADP-ribose) polymerase and caspase 3 cleavage. We also observed that loss of cell viability after treatment with I3C/TRAIL is greater compared with I3C and TRAIL alone. To determine the molecular mechanisms involved in the enhanced apoptosis, we examined the expression of two TRAIL death receptors (DR4 and DR5) and two TRAIL decoy receptors (DcR1 and DcR2). We found that treatment with I3C induced DR4 and DR5 expression at both transcriptional and translational levels. These findings suggest that I3C may be an effective sensitizer of TRAIL treatment against TRAIL-resistant prostate cancer cell lines such as LNCaP.