Binding of cytotoxin P4 from Naja nigricollis nigricollis to B16F10 melanoma and WEHI-3B leukemia cells.

Cobra venoms cause irreversible destruction of cells cultured in vitro [1,2]. The venom of Naja nigricollis nigricollis possessed the most potent cytotoxic activity towards B16F10 melanoma cells among various examined venoms [2]. The main cytotoxic factor (P4) isolated from this venom showed preferential activity on tumor cell lines and caused lysis at concentrations of 10(-7) M (0.8-1 micrograms/ml) [3]. The present study examined the binding of cytotoxin P4 to melanoma B16F10 and WEHI-3B leukemia cell lines and found that, like cytotoxicity, it depended on concentration, temperature and incubation time. Cytotoxin concentrations that elicited no apparent damage to cells during the first hour of incubation caused lysis after a longer period of incubation, suggesting that a critical number of bound molecules is required in order to cause cell death. Bivalent ions, such as Mg2+, Ca2+ or Sr2+, which decreased binding to the cells also inhibited cytotoxicity. Competition experiments as well as the displacement of 75% of the bound radiolabelled cytotoxin with 'cold' cytotoxin, suggest the presence of specific binding sites for the toxin in the examined tumor cells. The non-specific binding of the cytotoxin P4 to sea urchin ova and sperm cells without affecting their fertility, even at high concentrations of 10(-5) M, indicates that the specific binding to cells is probably a necessary condition for cell lysis.