Effects of methotrexate and other antifolates on the growth and dihydrofolate reductase activity of leishmania promastigotes

Scott, David A.; Coombs, Graham H.; Sanderson, Barbara E.

doi:10.1016/0006-2952(87)90508-9

Cited by 30 publications

(20 citation statements)

References 15 publications

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“…Antiamastigote activity of these 2,4-diaminoaquinazoline analogs of dihydrofolate was not well correlated with inhibitory activity against L. mexicana promastigote DHFR. This lack of correlation is consistent with the data of Scott et al (16), who found that two 2,4-diaminopyrimidines had minimum lethal concentrations against L. mexicana promastigotes that were equal (12 and 13 ,uM), but had 50% inhibitory concentrations against their reductases that differed (2 and 0.3 ,uM). On the other hand, a modest correlation between ED50 against L. donovani amastigotes in mouse peritoneal macrophages and 50% inhibition of L. major DHFR was seen by Sirawaraporn et al (17).…”

supporting

confidence: 91%

Antileishmanial activities of 2,4-diaminoquinazoline putative dihydrofolate reductase inhibitors

Berman

King

Edwards

1989

Antimicrob Agents Chemother

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2,4-Diaminoquinazoline analogs of folate were assessed as antileishmanial agents and as dihydrofolate reductase inhibitors. Against Leishmania major in human macrophages in vitro, two compounds with tertiary amines attached directly to the quinazoline ring were remarkably active. The 50% effective doses were in the picogram per milliliter range (12 to 91 pg/ml), and the in vitro therapeutic indices were -105. These compounds were 1,000 times more active on an absolute basis and had a 100 times more favorable therapeutic index than any compound previously tested in this model. Antileishmanial activity was not correlated with activity against Leishmania mexicana promastigote reductase, which suggests that folate utilization in general, rather than reductase activity specifically, was being inhibited.Tetrahydrofolate (THF) is an important cellular cofactor that acts as an acceptor and donor of 1-carbon fragments. For example, serine hydroxymethyl transferase removes a methyl group from serine, creating glycine, and forms N5,N10-methylene THF from THF. After conversion of N5,N10-methylene THF to N5-methyl THF by methylene THF reductase, N5-methyl THF is utilized by methionine synthetase, through which homocysteine is methylated to methionine and THF is regenerated. N5,N1O-THF also is a cofactor in the formation of thymidylate from deoxyuridylate by thymidylate synthetase. In this reaction, however, the cofactor donates both a 1-carbon group and hydrogen and is oxidized to dihydrofolate in the process. The ability of the cell to continue to synthesize thymidylate and DNA requires rereduction of dihydrofolate to THF by dihydrofolate reductase (DHFR). Structural analogs of THF such as methotrexate, trimethoprim, trimetrexate, and pyrimethamine and cycloguanil specifically inhibit DHFRs of tumor cells, bacteria, Pneumocystis species, and plasmodia, respectively, and are widely used clinical agents.Leishmania species and some other parasitic protozoa contain a bifunctional polypeptide possessing the enzymatic activity of both thymidylate synthetase and DHFR, in contradistinction to mammalian cells, in which the enzyme activities reside on separate polypeptides (12). Since DHFR is a target for rational chemotherapy and is structurally distinct in Leishmania species, intense work on its detailed structure is now being pursued. Thus, Grumont and coworkers (10, 11) have cloned, expressed, and compared the Leishmania major enzyme with that of monofunctional enzymes with the aim of rational design of a specifically antileishmanial agent. In addition, the activity of standard DHFR inhibitors against Leishmania amastigotes (mammalian forms) in murine macrophages (13), of novel 2,4-diaminopyrimidines or triazines against promastigotes (insect vector forms) (5, 16), and of novel 5-(substituted benzyl)-2,4-diaminopyrimidines against promastigotes and amastigotes in mouse peritoneal macrophages (17) is being investigated in the hope that agents devised for other diseases might have significant antileishmanial activity.We determined ...

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supporting

confidence: 91%

Antileishmanial activities of 2,4-diaminoquinazoline putative dihydrofolate reductase inhibitors

Berman

King

Edwards

1989

Antimicrob Agents Chemother

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“…SLD promastigote Ag was prepared according to the method described by Scott et al (36) and modified by Choudhry et al (37). Briefly, late logphase promastigotes (10 9 ) were harvested from 3-to 4-day cultures, washed four times in cold PBS, resuspended in PBS containing protease inhibitors mixture (Sigma-Aldrich), and sonicated (Soniprep-150) for two periods of 1.5 min each in ice, (separated by an interval of 3 min) at medium amplitude.…”

Section: Preparation Of Soluble L Donovani (Sld) Promastigote Agmentioning

confidence: 99%

Immunization with the DNA-Encoding N-Terminal Domain of Proteophosphoglycan of Leishmania donovani Generates Th1-Type Immunoprotective Response against Experimental Visceral Leishmaniasis

Samant

Gupta

Kumari

et al. 2009

The Journal of Immunology

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Leishmania produce several types of mucin-like glycoproteins called proteophosphoglycans (PPGs) which exist as secretory as well as surface-bound forms in both promastigotes and amastigotes. The structure and function of PPGs have been reported to be species and stage specific as in the case of Leishmania major and Leishmania mexicana; there has been no such information available for Leishmania donovani. We have recently demonstrated that PPG is differentially expressed in sodium stibogluconate-sensitive and -resistant clinical isolates of L. donovani. To further elucidate the structure and function of the ppg gene of L. donovani, a partial sequence of its N-terminal domain of 1.6 kb containing the majority of antigenic determinants, was successfully cloned and expressed in prokaryotic as well as mammalian cells. We further evaluated the DNA-encoding N-terminal domain of the ppg gene as a vaccine in golden hamsters (Mesocricetus auratus) against the L. donovani challenge. The prophylactic efficacy to the tune of ∼80% was observed in vaccinated hamsters and all of them could survive beyond 6 mo after challenge. The efficacy was supported by a surge in inducible NO synthase, IFN-γ, TNF-α, and IL-12 mRNA levels along with extreme down-regulation of TGF-β, IL-4, and IL-10. A rise in the level of Leishmania-specific IgG2 was also observed which was indicative of enhanced cellular immune response. The results suggest the N-terminal domain of L. donovani ppg as a potential DNA vaccine against visceral leishmaniasis.

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“…These data suggest that even when gene amplification is a potential resistance mechanism, it is not necessarily the favored event. (64,100), nor did MTX-resistant Leishmania tarentolae selected from a line lacking pre-existing H amplification (117). Thus, any given resistant line may fail to include amplification in its spectrum of resistance mechanisms.…”

Section: Multiple Mechanisms Of Drug Resistancementioning

confidence: 99%

GENE AMPLIFICATION IN LEISHMANIA

Beverley

1991

Annu. Rev. Microbiol.

216

128

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Effects of methotrexate and other antifolates on the growth and dihydrofolate reductase activity of leishmania promastigotes

Cited by 30 publications

References 15 publications

Antileishmanial activities of 2,4-diaminoquinazoline putative dihydrofolate reductase inhibitors

Antileishmanial activities of 2,4-diaminoquinazoline putative dihydrofolate reductase inhibitors

Immunization with the DNA-Encoding N-Terminal Domain of Proteophosphoglycan of Leishmania donovani Generates Th1-Type Immunoprotective Response against Experimental Visceral Leishmaniasis

GENE AMPLIFICATION IN LEISHMANIA

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