Endowing human pancreatic ribonuclease with toxicity for cancer cells

Onconase(R) is an amphibian protein that is now in Phase IH clinical trials as a cancer chemotherapeutic. Human pancreatic ribonuclease (RNase 1) is h omologous to Onconase(R) but is not cytotoxic. Here, ERDD RNase 1, which is the L86E/N88R/G89D/R91D variant of RNase 1, is shown to have conformationa l. stability and ribonucleolytic activity similar to that of the wild-type enzyme but >10(3)-fold less affinity for the endogenous cytosolic ribonucle ase inhibitor protein. Most significantly, ERDD RNase 1 is toxic to hum an leukemia cells. The addition of a non-native disulfide bond to ERDD RNase I not only increases the conformational stability of the enzyme but also inc reases its cytotoxicity such that its IC50 value is only 8-fold greater tha n that of Onconase(R). Thus, only a few amino acid substitutions are necess ary to make a human protein toxic to human cancer cells. This finding has s ignificant implications for human cancer chemotherapy.