The postulation that the activity of key
enzymes that reveal marked increases should be potential targets for anticancer
chemotherapy (47) was supported by new evidence on the alterations of
CDP reductase,
CTP synthetase and
OMP decarboxylase in
hepatoma 3924A cell cultures. Inhibitors of these
enzymes (
VF-122,
acivicin,
pyrazofurin) and that of
IMP dehydrogenase (
tiazofurin) efficiently killed
hepatoma 3924A cells in culture, as demonstrated by the clonogenic assay.
Acivicin,
pyrazofurin,
tiazofurin and
VF-122 were lethal against
tumor cells in the exponential phase of growth with IC50 of 1.5, 5, 10 and 4.5 microM, respectively. All these
antimetabolites exhibited cytotoxicity preponderantly against exponential-phase cultures, indicating that all the four drugs belong to Class II (phase-specific agents) in the Kinetic Classification of
Anticancer Agents (38).
Dibromodulcitol, a bifunctional
alkylating agent, revealed cycle-specific cytotoxicity (Class III agent) against
hepatoma 3924A, yielding IC50 values of 2.3 and 5.5 microM for exponentially and stationary growing cells, respectively. Using isobologram analysis on the survival data of 3924A cells, synergistic interaction was observed when DBD in combination with
acivicin,
pyrazofurin and
tiazofurin was examined. DBD in combination with
VF-122 exhibited additive lethality against
hepatoma cells in culture. The synergistic and additive cytotoxicity in combinations of DBD with these
antimetabolites was accompanied by the concurrent depletion of
ribonucleotide and/or
deoxyribonucleotide pools. The synergistic
biological results of
drug combinations of
acivicin with DBD can be accounted for by the action of
acivicin in inhibiting
CTP synthetase, resulting in a synergistic decrease in
CTP content, and by inhibition of
DNA synthesis caused by DBD. The synergistic and additive depletion of
UTP,
CTP,
dTTP and
dCTP pools in the combinations of DBD with
pyrazofurin may be responsible for the synergistic lethality of these combinations. Synergism, in terms of pool depletion, was observed for
GTP and
dCTP; summation was detected for
dGTP when DBD and
tiazofurin were given concurrently. The synergistic cytotoxicity of this
drug combination may be a consequence of these alterations. The additive lethality of DBD-VF-122
drug combinations was reflected in the additive elevations of the ribonucleoside
diphosphate concentrations. These observations indicate that treatments based on the Kinetic Classification and on the biochemical targeting of the
drug should have an impact on the design of in vivo
chemotherapy.