Antifolate activity of isoaminopterin in HeLa cells.

The antifolate activity and the transport characteristics of isoaminopterin (IA) in HeLa cells were studied and compared with those of methotrexate (MTX). Both IA and MTX inhibited the incorporation of [2-(14)C]deoxyuridine into HeLa cell DNA by 50% at a concentration of 0.09 muM by 8 h postaddition of the compounds. Unlike MTX, the inhibition of DNA synthesis by IA was time dependent and reached a maximum at 24 h. IA-induced inhibition was due to interference with folate metabolism, since it could be completely reversed with N(5)-formyl-tetrahydrofolate. Competitive transport experiments between IA and either radiolabeled MTX or radiolabeled folate showed that IA preferentially uses the reduced folate/MTX transport system. IA inhibited MTX uptake by 50% at a concentration of 6.8 muM but had a negligible effect on folate transport.