Cytokine-mediated antitumor effect of OK-432 on urinary bladder tumor cells in vitro

Abstract

Fatal complications from the intravesical instillation of bacillus Calmette-Guérin (BCG) for the treatment of superficial urinary bladder tumors have been reported. OK-432, an immunomodulating agent like BCG, may be an effective and safe agent for the treatment of urinary bladder tumors. We investigated the cytokine-mediated antitumor effect of OK-432 on established human bladder cancer cell lines (T24 and KK-47) in vitro. Peripheral blood mononuclear cells (PBMCs) from a healthy volunteer were cultured with OK-432 for various periods, and the culture supernatants were used as conditioned media. Cytokines in the culture supernatants were quantified. The antitumor effect of OK-432 was evaluated by colony-forming assays, using the conditioned media as the culture media. The colony survival of T24 and KK-47 cells was significantly inhibited by conditioned media from 24-h cultures of PBMCs incubated with OK-432 at concentrations of 0.05 and 0.1 Klinische Einheit (KE)/ml. Conditioned media from PBMCs cultivated with OK-432 for 7 days at 0.01 and 0.05 KE/ml also significantly inhibited the colony survival of both cell lines. Higher concentrations of interleukin-1 beta (IL-1 beta) and tumor necrosis factor alpha (TNF alpha) were detected in conditioned media cultivated with OK-432 for 24 h than in media from PBMCs alone. However, higher concentrations of interferon gamma (IFN gamma) were detected in conditioned media cultivated with OK-432 for 7 days. Approximately 90% of the inhibition of KK-47 cells by the 24-h conditioned media was neutralized by an anti-TNF monoclonal antibody. The inhibition of T24 cells was neutralized approximately 50% by the same antibody. The inhibition of T24 and KK-47 cells by 7-day conditioned media was completely neutralized by an anti-IFN gamma monoclonal antibody. The cultivation of PBMCs with OK-432 inhibited the production of granulocyte-colony-stimulating factor (G-CSF) by PBMCs. The inhibition may play a role in the mechanism of the antitumor effect of OK-432. Urinary bladder tumor cell lines have different sensitivities to cytokines. The cytokines induced by OK-432 vary with the concentration of OK-432 and the culture period. It is suggested that in intravesical instillation of OK-432 for treatment of urinary bladder tumor, the optimal dose and interval of instillation should be considered.