Antimycobacterial Activities of 2,4-Diamino-5-Deazapteridine Derivatives and Effects on Mycobacterial Dihydrofolate Reductase

Abstract

Development of new antimycobacterial agents for Mycobacterium avium complex (MAC) infections is important particularly for persons coinfected with human immunodeficiency virus. The objectives of this study were to evaluate the in vitro activity of 2,4-diamino-5-methyl-5-deazapteridines (DMDPs) against MAC and to assess their activities against MAC dihydrofolate reductase recombinant enzyme (rDHFR). Seventy-seven DMDP derivatives were evaluated initially for in vitro activity against one to three strains of MAC (NJ168, NJ211, and/or NJ3404). MICs were determined with 10-fold dilutions of drug and a colorimetric (Alamar Blue) microdilution broth assay. MAC rDHFR 50% inhibitory concentrations versus those of human rDHFR were also determined. Substitutions at position 5 of the pteridine moiety included ™CH 3 , ™CH 2 CH 3 , and ™CH 2 OCH 3 groups. Additionally, different substituted and unsubstituted aryl groups were linked at position 6 through a two-atom bridge of either ™CH 2 NH, ™CH 2 N(CH 3 ), ™CH 2 CH 2 , or ™CH 2 S. All but 4 of the 77 derivatives were active against MAC NJ168 at concentrations of <13 g/ml. Depending on the MAC strain used, 81 to 87% had MICs of <1.3 g/ml. Twenty-one derivatives were >100-fold more active against MAC rDHFR than against human rDHFR. In general, selectivity was dependent on the composition of the two-atom bridge at position 6 and the attached aryl group with substitutions at the 2 and 5 positions on the phenyl ring. Using this assessment, a rational synthetic approach was implemented that resulted in a DMDP derivative that had significant intracellular activity against a MAC-infected Mono Mac 6 monocytic cell line. These results demonstrate that it is possible to synthesize pteridine derivatives that have selective activity against MAC.

Identification and Characterization of a Unique Adenosine Kinase from Mycobacterium tuberculosis

“…The MIC of methyl-Ado was evaluated for M. tuberculosis strains H37Ra, SRICK1, and SRICK1-pVV16/Rv2202c using a colorimetric broth microdilution assay as previously described (29). Dilutions of methylAdo, ethambutol, and dimethyl sulfoxide (DMSO) were prepared in Middlebrook 7H9 medium supplemented with 0.2% glycerol and OADC.…”

New Mycobacterium avium Antifolate Shows Synergistic Effect when Used in Combination with Dihydropteroate Synthase Inhibitors

“…This is significant because MAC is intrinsically resistant to trimethoprim, a commonly used drug that targets prokaryotic DHFRs but not human DHFR. We have shown that the 50% inhibitory concentration of trimethoprim for the MAC DHFR is 4,100 nM, in comparison to the new DMDPs, which have 50% inhibitory concentrations around 1.0 nM (7). DHFR is a key enzyme in the folate biosynthetic pathway that catalyzes the reduction of dihydrofolate to tetrahydrofolate, derivatives of which function in single carbon transfers at various oxidation states for the synthesis of purines, methionine, glycine, pantothenate, thymidylate, and Nformylmethionyl-tRNA (3,5).…”

“…Demonstration of the antimycobacterial activity of these new antifolates was initially aided by efforts of the National Institutes of Health-sponsored Tuberculosis Antimicrobial Acquisition and Coordinating Facility. Continued efforts have resulted in a specific group of DMDPs that have selective activity for MAC dihydrofolate reductase (DHFR) but not human DHFR, which makes them good candidates for further development (7). This is significant because MAC is intrinsically resistant to trimethoprim, a commonly used drug that targets prokaryotic DHFRs but not human DHFR.…”

“…These strains were used previously by us to show selective activity against MAC of the DMDP derivatives (7).…”

“…pyrimidine} is the hydrochloride salt form of SRI-8686, a DMDP derivative whose synthesis has been described previously (7). Sulfamethoxazole and dapsone were purchased from Sigma.…”