1,2‐Bis(methylsulfonyl)‐1‐(2‐chloroethyl)‐2‐[[1‐(4‐nitrophenyl)ethoxy]carbonyl]hydrazine (KS119): a Cytotoxic Prodrug with Two Stable Conformations Differing in Biological and Physical Properties

Abstract

The anticancer prodrug 1,2-bis(methylsulfonyl)-1-(2-chloroethyl)-2-[[1-(4-nitrophenyl)ethoxy]carbonyl]hydrazine (KS119) selectively releases a short-lived cytotoxin following enzymatic reduction in hypoxic environments found in solid tumors. KS119, in addition to two enantiomers, has two stable atropisomers (conformers differing in structure owing to hindered bond rotation) that interconvert at 37 °C in aqueous solution by first order kinetics with t1/2 values of ~50 and ~64 hours. The atropisomers differ in physical properties such as partition coefficients that allow their chromatographic separation on non-chiral columns. A striking difference in the rate of metabolism of the two atropisomers occurs in intact EMT6 murine mammary carcinoma cells under oxygen deficient conditions. A structurally related molecule, 1,2-bis(methylsulfonyl)-1-(2-chloroethyl)-2-[[1-(3-hydroxy-4-nitrophenyl)ethoxy]carbonyl]hydrazine (KS119WOH), was also found to exist in similar stable atropisomers. The ratio of the atropisomers of KS119 and structurally related agents has the potential to impact the bioavailability, activation and therapeutic activity. Thus, thermally stable atropisomers/conformers in small molecules can result in chemically and enantiomerically pure compounds having differences in biological activities.

Preliminary Studies with a New Hypoxia-Selective Cytotoxin, KS119W,In VitroandIn Vivo

“…EMT6 cells have been used extensively in the authors’ laboratories and their characteristics are well defined (1–4, 8–10, 16, 21–24, 26–40). Two features of EMT6 cells are important in these studies.…”

“…The bioreductive alkylating agent KS119, 1,2-bis(methylsulfonyl)-1-(2-chloroethyl)-2-[[1-(4-nitrophenyl)ethoxy]-carbonyl]hydrazine), developed in the Sartorelli laboratory at Yale, had an extremely large differential in its toxicity to severely hypoxic and aerobic cells in cell culture, and was effective in killing hypoxic cells of EMT6 mouse mammary tumors in vivo , as measured using a clonogenic assay (11, 20–24). However, the limited solubility of this compound precluded development as a potential anticancer drug.…”

“…Further studies in the Sartorelli laboratory and at Vion Pharmaceuticals resulted in the development of an analog, KS119W, that has increased aqueous solubility due to the addition of a phosphate group. The octanol:water partition coefficients were 151 for KS119 and 22.5 for KS119W (24). Both KS119 and KS119W are prodrugs that require enzymatic nitroreductive activation, which occurs preferentially in hypoxic cells, to generate 1,2-bis(methylsulfonyl)-1-(2-chloroethyl)hydrazine (90CE), which in turn, generates the alkylating species that produces DNA crosslinks (11, 20–24).…”

“…The octanol:water partition coefficients were 151 for KS119 and 22.5 for KS119W (24). Both KS119 and KS119W are prodrugs that require enzymatic nitroreductive activation, which occurs preferentially in hypoxic cells, to generate 1,2-bis(methylsulfonyl)-1-(2-chloroethyl)hydrazine (90CE), which in turn, generates the alkylating species that produces DNA crosslinks (11, 20–24). The molecules consist of three domains (25), a “trigger region” consisting of a nitro group, a benzyloxycarbonyl “linker” region, and a “therapeutic warhead” or “effector” region that consists of the cytotoxic crosslinking moiety 90CE (Fig.…”

“…1). The synthesis, chemistry, and biochemistry of KS119, KS119W, 90CE and some related compounds are described in publications from the Sartorelli laboratory (11, 20–24). The studies reported here compare the cytotoxicities of KS119 and KS119W to aerobic and hypoxic EMT6 cells in cell culture, and examine some environmental and cellular factors that influence the cytotoxicity of the newer analog, KS119W, to cells in vitro .…”