Inhibition of Pneumocystis carinii dihydropteroate synthetase by para-acetamidobenzoic acid: possible mechanism of action of isoprinosine in human immunodeficiency virus infection

Abstract

Isoprinosine has been reported to decrease progression to AIDS, primarily by preventing Pneumocystis carinii pneumonia (PCP), in human immunodeficiency virus-infected patients, but the mechanism of action is unknown. para-Acetamidobenzoic acid (PAcBA), one component of isoprinosine, is structurally related to para-aminobenzoic acid (PABA), a precursor of de novo folate synthesis. This pathway is known to be important for P. carinii because sulfonamides, which are effective anti-P. carinii agents, inhibit incorporation of PABA into folate precursors by the enzyme dihydropteroate synthetase (DHPS). Inhibition of P. carinii DHPS by PAcBA was investigated by using two assays. In short-term cultures of P. carinii from rats, [3H]PABA incorporation into reduced folates was inhibited by both isoprinosine (mean +/- standard error 50% inhibitory concentration [IC50], 20 +/- 8.4 microM) and PAcBA free acid (IC50, 240 +/- 100 microM); a soluble PAcBA salt was more potent than PAcBA free acid alone (IC50, 29 +/- 48 microM). The activity of PAcBA free acid was confirmed in a cell-free DHPS inhibition assay (IC50, 120 +/- 120 microM). Inosine and dimethylaminopropanol, two other components of isoprinosine, were poor inhibitors of PABA incorporation (IC50, > 1,000 microM). PAcBA free acid also showed activity in inhibiting the DHPS of Toxoplasma gondii, but was a poor inhibitor of the DHPSs of Escherichia coli and Saccharomyces cerevisiae. In a rat model of PCP, the PAcBA salt administered intraperitoneally demonstrated no activity against established PCP either alone or when used in combination with trimethoprim; the lack of efficacy in this model may be due to the rapid metabolism of the drug. Prevention of PCP by PaCBA through inhibition of P. carinii DHPS may explain the activity of isoprinosine in decreasing the progression to AIDS in human immunodeficiency virus-infected patients.