SITE OF ACTION OF 2 NOVEL PYRIMIDINE BIOSYNTHESIS INHIBITORS ACCURATELY PREDICTED BY THE COMPARE PROGRAM

The computer algorithm COMPARE provides information regarding the biol ogical mechanism of action of a compound. In this study, excellent cor relations were obtained for thoxy[1,1'-biphenyl]-4,4'-diyl)diimino]bis -benzoic acid (redoxal) and enyl)-2-methyl-1H-naphth[2,3-d]imidazole-4 ,9-dione (BNID) and two well-studied dihydroorotate dehydrogenase (DHO D) inhibitors, dichloroallyl lawsone and brequinar, in terms of antipr oliferative activity against tumor cell lines in vitro. When redoxal a nd BNID were incubated with MOLT-4 cells for 72 hr, 50% growth inhibit ion was achieved at 0.7 and 3.5 mu M, respectively. After 24 hr of inc ubation, pyrimidine triphosphate pools were shown to be decreased by 5 0% by redoxal (1 mu M) and BNID (0.25 mu M) Addition of either uridine (50 mu M) or cytidine (100 mu M) antagonized the cellular cytotoxicit y caused by either drug; uridine corrected the UTP and CTP deficit, wh ereas cytidine corrected only the CTP deficit. Exposure of MOLT-4 cell s to a 1 mu M concentration of either drug for 18 hr followed by a 1-h r exposure to [C-14]bicarbonate showed a 97% decrease of incorporation of [C-14] into pyrimidine triphosphates accompanied by a 91- and 82-f old increase in radioactive incorporation into L-dihydroorotate and N- carbamyl-L-aspartate, respectively. By direct exposure of DHOD prepare d from MOLT-4 cell mitochondria to a range of concentrations of the tw o drugs, apparent K-i values of 0.33 mu M (redoxal) and 0.53 mu M (BNI D) were determined. These data provide direct evidence for inhibition of DHOD by redoxal and BNID in MOLT-4 lymphoblasts.