Influence of a Basic Side Chain on the Properties of Hypoxia-Selective Nitro Analogues of the Duocarmycins: Demonstration of Substantial Anticancer Activity in Combination with Irradiation or Chemotherapy

A new series of nitro analogues of the duocarmycins was prepared and evaluated for hypoxia-selective anticancer activity. The compounds incorporate 13 different amine-containing side chains designed to bind in the minor groove of DNA while spanning a wide range of base strength from p<i>K</i>_a 9.64 to 5.24. The most favorable in vitro properties were associated with strongly basic side chains, but the greatest in vivo antitumor activity was found for compounds containing a weakly basic morpholine. This applies to single-agent activity and for activity in combination with irradiation or chemotherapy (gemcitabine or docetaxel). In combination with a single dose of γ irradiation <b>50</b> at 42 μmol/kg eliminated detectable clonogens in some SiHa cervical carcinoma xenografts, and in combination with gemcitabine using a well-tolerated multidose schedule, the same compound caused regression of all treated A2780 ovarian tumor xenografts. In the latter experiment, three of seven animals receiving the combination treatment were completely tumor free at day 100

Reductive Metabolism Influences the Toxicity and Pharmacokinetics of the Hypoxia-Targeted Benzotriazine Di-Oxide Anticancer Agent SN30000 in Mice

3-(3-Morpholinopropyl)-7,8-dihydro-6H-indeno[5,6-e][1,2,4]triazine 1,4-dioxide (SN30- 000), an analog of the well-studied bioreductive prodrug tirapazamine (TPZ), has improved activity against hypoxic cells in tumor xenografts. However, little is known about its biotransformation in normal tissues. Here, we evaluate implications of biotransformation of SN30000 for its toxicokinetics in NIH-III mice. The metabolite profile demonstrated reduction to the 1-N-oxide (M14), oxidation of the morpholine side-chain (predominantly to the alkanoic acid M18) and chromophore, and subsequent glucuronidation. Plasma pharmacokinetics of SN30000 and its reduced metabolites was unaffected by the presence of HT29 tumor xenografts, indicating extensive reduction in normal tissues. This bioreductive metabolism, as modeled by hepatic S9 preparations, was strongly inhibited by oxygen indicating that it proceeds via the one-electron (radical) intermediate previously implicated in induction of DNA double strand breaks and cytotoxicity by SN30000. Plasma pharmacokinetics of SN30000 and M14 (but not M18) corresponded closely to the timing of reversible acute clinical signs (reduced mobility) and marked hypothermia (rectal temperature drop of ∼8°C at nadir following the maximum tolerated dose). Similar acute toxicity was elicited by dosing with TPZ or M14, although M14 did not induce the kidney and lung histopathology caused by SN30000. M14 also lacked antiproliferative potency in hypoxic cell cultures. In addition M14 showed much slower redox cycling than SN30000 in oxic cultures. Thus a non-bioreductive mechanism, mediated through M14, appears to be responsible for the acute toxicity of SN30000 while late toxicities are consistent with DNA damage resulting from its one-electron reduction. A two-compartment pharmacokinetic model, in which clearance of SN30000 is determined by temperature-dependent bioreductive metabolism to M14, was shown to describe the non-linear PK of SN30000 in mice. This study demonstrates the importance of non-tumor bioreductive metabolism in the toxicology and pharmacokinetics of benzotriazine di-oxides designed to target tumor hypoxia