Studies of the biochemical pharmacology of the fermentation-derived antitumor agent, (αS, 5S) -α-amino-3-chloro-4, 5-dihydro-5-isoxazoleacetic acid (at-125)

Abstract

(αS, 5S)-α-Amino-3-chloro-4,5-dihydro-5-isoxazoleacetic acid (AT-125) isolated from fermentation broths of Streptomyces sviceus was found to have significant activity against a number of tumors in experimental animals. It is currently being developed by the U.S. National Cancer Institute for clinical evaluation. AT-125 was shown to be a potent inhibitor of growth of L1210 and KB cells in culture. These effects were markedly dependent on the time of exposure to the agent. Similar results were observed when survival (residual proliferative capacity) of L1210 cells was measured. With L1210 cells (in culture and in vivo) AT-125 inhibited the incorporation of 3H-TdR into macromolecules to a greater extent than was observed when 3H-UR was the precursor. The activity of AT-125 in crude fermentation broths was detected in an in vitro antimicrobial prescreen designed to specifically detect materials with antimetabolite activity. Its activity in such a system was rather specifically antagonized by the amino acid, L-histidine. Such was not the case, however, in mammalian cells; L-histidine was without effect on AT-125 activity. In mammalian cells, the effects of AT-125 on growth inhibition or inhibition of radioactive precursors into macromolecules were rather specifically antagonized by L-glutamine. AT-125 was found to have significantly greater toxicity towards female than male mice, and sensitivity appeared to decrease with animal age. Further studies showed that AT-125 toxicity could be alleviated by coadministration of testosterone. In this respect, the biological activity of AT-125 was quite similar to that of a nucleoside antitumor agent, deaza UR. The primary locus of activity of deaza UR (after conversion to the triphosphate) has been shown to be inhibition of CTP synthetase. In studies with rat liver homogenates, AT-125 has been shown to inhibit CTP synthetase as well as other enzymes which catalyze the transfer of the amido group of L-glutamine. Further investigation of the effects of AT-125 have shown it to be a very potent inhibitor (K i = 2 × 10 −6 M) of purified rat liver CTP synthetase. AT-125 also causes effects on ribonucleotide pools which are consistent with inhibition of this enzyme and with the inhibition of another L-glutamine-dependent enzyme, XMP aminase. The importance of these two enzymes as loci for the biochemical action of AT-125 was supported when it was found that a combination of CR and GR significantly antagonized the growth inhibitory activity of AT-125.