Exploitation of folate and antifolate polyglutamylation to achieve selective anticancer chemotherapy

Abstract

Synthesis of poly(gamma-glutamate) metabolites of natural folates and antifolates is a critical process. Folypolyglutamates are essential for cell proliferation. Polyglutamates of glutamate (Glu)-containing antifolates are often critical for their cytotoxic action and are relevant to antifolate resistance. However, the role of polyglutamate synthesis in selectivity is less clear. We have undertaken a research program to further define the significance of polyglutamate metabolism and to devise ways to exploit this metabolism to achieve greater therapeutic selectivity in cancer chemotherapy. This article briefly reviews several approaches tested thus far. Inhibition of folypolyglutamate synthesis should lead to cell death. Current ornithine (Orn)-containing folate-based inhibitors of the enzyme responsible for their synthesis, folypolyglutamate synthetase (FPGS), are poorly transported, apparently because of interference by the protonated delta-amine. Replacement of Orn with 4, 4-difluoroOrn, the delta-amine of which has a much lower pKa and is thus less protonated at physiological pH, was explored. Since it is unclear how polyglutamylation contributes to selectivity, we explored generic means either to eliminate or to enhance polyglutamylation. The data indicate that substitution for Glu in an antifolate by some Glu analogs in which the gamma-COOH is either altered or replaced (e.g., gamma-tetrazole-Glu) leads to loss of both FPGS substrate activity and binding; antifolate target specificity is unchanged, while uptake is actually enhanced. Substitution of 3,3-difluoroGlu for Glu leads to enhanced polyglutamylation (although probably only to the diglutamate), retention of target specificity, and at least equal uptake. Comparative studies of the same antifolate containing different replacements for Glu, such as gamma-tetrazole-Glu (no polyglutamylation) or 3,3-difluoroGlu (enhanced polyglutamylation), will be useful in exploring the role and significance of polyglutamylation.

Design and Synthesis of Histidine Analogues of Folic Acid and Methotrexate as Potential Folylpolyglutamate Synthetase Inhibitors

“…It would be of interest to directly compare the rate and pH dependence of the cellular uptake of radioactive isotope-labeled analogues, once they become available. Folic acid derivatve 6 did not show significant cell growth inhibition, as expected, since side chain-modified folic acid derivatives generally lack significant cytotoxicity. , The results suggest that by optimizing the p K a of potential FPGS inhibitors bearing basic side chains, the opposite ionization requirements of cellular uptake and enzyme inhibitory activity can be satisfied …”

“…16,21 The results suggest that by optimizing the pK a of potential FPGS inhibitors bearing basic side chains, the opposite ionization requirements of cellular uptake and enzyme inhibitory activity can be satisfied. 23 Inhibition of DHFR. It is reasonable to assume that the cytotoxicity of side chain-modified MTX analogues is primarily due to inhibition of DHFR, since structural variations at this region have relatively little effect on enzyme inhibitory potency.…”

A Review of Small-Molecule Inhibitors of One-Carbon Enzymes: SHMT2 and MTHFD2 in the Spotlight

“…However, there is still interest in pursuing pro-drug strategies for the inhibitors in which there was not dose-limiting toxicity because of promising preclinical data . Folate analogs are reported to inhibit FPGS, albeit only modestly in the midmicromolar range …”

“…16 Folate analogs are reported to inhibit FPGS, albeit only modestly in the midmicromolar range. 48 2.4.3. Folate and Serine Transporter Inhibitors.…”