Folate and antifolate cell binding and accumulation at physiologic folate concentrations.

Cellular transport of folate has been shown to be via either a carrier-mediated or -facilitated diffusion process. The transport constant (Kt) values have ranged from 1 to 20 microM in vitro. These studies have generally been done on cells grown in high concentrations of folic acid (10(-6) to 10(-5) M). When cells were grown in folate-depleted medium for 72 hours, growth was nearly equivalent to that observed in complete medium, but a significant quantity of a high-affinity membrane receptor for folate was detected. The dissociation constants for folic acid, 5-methyltetrahydrofolate and methotrexate were 0.4, 4, and 18 nM, respectively. Incubation of folate-depleted cells for 3 hours at 10(-8) M methyltetrahydrofolate restored the intracellular folate content to values equivalent to those found in cells grown in folate-complete medium and decreased membrane binding by greater than 95%. Binding of 5-methyltetrahydrofolate was inhibited by methotrexate, folic acid, and 5-formyltetrahydrofolate, but not by glutamic acid, pteroic acid, or p-aminobenzoylglutamic acid. Antisera against a folate-binding protein (FBP) blocked cell binding of folate and purified FBP also blocks cell binding of folate. These results suggest that there is a receptor-mediated uptake of folate, functional at physiologic concentrations of plasma folate. This receptor could potentially be a new membrane target for antifol, antineoplastic agents.