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Antimicrob Agents Chemother. 1993 August; 37(8): 1674-1678

Development and characterization of a rapid screening assay for identifying antipneumocystis agents.

A Martinez and J A Kovacs

Critical Care Medicine Department, National Institutes of Health, Bethesda, Maryland 20892.

ABSTRACT

We developed a rapid assay for screening of compounds with potential antipneumocystis activity on the basis of incorporation of [35S]methionine into proteins newly synthesized by Pneumocystis carinii. Unambiguous evidence that P. carinii synthesizes proteins in vitro was provided by immunoprecipitation studies demonstrating the incorporation of [35S]methionine into the major surface glycoprotein. Treatment with two clinically active antipneumocystis agents, atovaquone (10(-4) M) or pentamidine (10(-4) M), prevented this incorporation. Total [35S]methionine incorporation paralleled incorporation into the major surface glycoprotein, permitting rapid assessment of anti-P. carinii activity by scintillation counting. Treatment with pentamidine (1 x 10(-4) M), atovaquone, trimethoprim (1 x 10(-4) M)-sulfamethoxazole (7.9 x 10(-4) M), piritrexim (1 x 10(-7) M), RO11-8958 (1 x 10(-4) M), and amphotericin B (1 microgram/ml) resulted in a greater than 67% inhibition (P < 0.05) of [35S]methionine incorporation. No decrease in [35S]methionine incorporation was seen with dapsone (10(-5) M), trimethoprim (10(-4) M), recombinant mouse tumor necrosis factor (500 ng/ml), or gamma interferon. This rapid in vitro assay should be a useful adjunct in the development of new antipneumocystis agents.

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Antimicrob Agents Chemother. 1993 August; 37(8): 1674-1678

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