To target malignant cells residing in hypoxic regions of solid
tumors, we have designed and synthesized
prodrugs generating the cytotoxic alkylating species
1,2-bis(methylsulfonyl)-1-(2-chloroethyl)hydrazine (90CE) after bioreductive activation. We postulate that one of these agents, 1,2-bis(methylsulfonyl)-1-(2-chloroethyl)-2-[[1-(4-nitrophenyl)ethoxy]carbonyl]
hydrazine (
KS119), requires enzymatic nitro reduction to produce 90CE, whereas another agent, 1,2-bis(methylsulfonyl)-1-(2-chloroethyl)-2-[(4-nitrobenzyloxy)carbonyl]
hydrazine (PNBC), can also be activated by nucleophilic attack by
thiols such as
glutathione (GSH)/GST. We demonstrated that these agents selectively kill hypoxic EMT6 mouse mammary
carcinoma and CHO cells. In
hypoxia, 50 microM
KS119 produced 5 logs of kill of EMT6 cells without discernable cytotoxicity in air; similar effects were observed with CHO cells. PNBC was less efficacious against hypoxic
tumor cells and also had some toxicity to aerobic cells, presumably because of GST/
thiol activation, making PNBC less interesting as a selective hypoxic-cell
cytotoxin. BALB/c mice with established EMT6 solid
tumors were used to demonstrate that
KS119 could reach and kill hypoxic cells in solid
tumors. To gain information on bioreductive
enzymes involved in the activation of
KS119, cytotoxicity was measured in CHO cell lines overexpressing
NADH:cytochrome b5 reductase (NBR),
NADPH:cytochrome P450 reductase (NPR), or
NADPH:
quinone oxidoreductase 1 (NQO1). Increased cytotoxicity occurred in cells overexpressing NBR and NPR, whereas overexpressed NQO1 had no effect. These findings were supported by enzymatic studies using purified NPR and
xanthine oxidase to activate
KS119.
KS119 has significant potential as a
hypoxia-selective
tumor-cell
cytotoxin and is unlikely to cause major toxicity to well oxygenated normal tissues.