Tumor cell resistance to fluoropyrimidines and other inhibitors of
thymidylate synthase (TS) is a serious problem often associated with increased intracellular TS. Clinically, another problem that arises from the use of TS inhibitors is toxicity, which develops, in part, because normal cells may be adversely affected by doses of inhibitor that do not impact
tumor cells. To circumvent this problem, we have devised a new strategy called
enzyme-catalyzed therapeutic activation (ECTA), which takes advantage of overexpressed TS to enzymatically generate cytotoxic moieties preferentially in
tumor cells. We show herein that
tumor cells expressing elevated levels of TS are preferentially sensitive to
NB1011, a
phosphoramidate derivative of
(E)-5-(2-bromovinyl)-2'-deoxyuridine. We find support for the proposed mechanism of
NB1011 in the following results: 1) positive relationship between TS
protein level and sensitivity to
NB1011 in engineered HT1080
tumor cells, designed to express defined levels of TS
protein; 2)
NB1011 activity is enhanced on
tumor cells which express endogenous elevated TS; 3) cytotoxicity of
NB1011 is blocked by
raltitrexed (
Tomudex); 4)
NB1011 selection of TS-overexpressing MCF7TDX
tumor cells results in recovery of cell populations and clones with diminished TS levels (and restored sensitivity to
raltitrexed). A preliminary comparison of TS
mRNA levels in multiple normal tissues versus colon
tumor samples suggests that selective
tumor cytotoxicity of
NB1011 may be possible in the clinical setting. Because
NB1011 cytotoxicity is dependent upon activation by TS, its proposed mechanism of action is distinct from current TS-targeted drugs, which require inhibition of TS to be effective.