Pyrimethamine
Pyrimethamine, a synthetic derivative of etheyl-pyrimidine, has been used as a antimalarial drug. It is a competetive inhibitor of dihydrofolate reductase (DHFR), interfering tetrahydrofolate synthesis. Mutation of PfDHFR codon 108 is an important predictor of pyrimethamine resistance in Plasmodium falciparum.
- Locus utilized: PF3D7_0417200 (dhfr)
- Codon: 108
- Filed utilized: geno_dhfr_108
| Amino acid change | Coding | Interpretation | Phenotype |
|---|---|---|---|
| S108 | geno_dhfr_108 == “S” | Wild-type | Sensitive |
| 108N | geno_dhfr_108 == “N” | Mutant | Resistant |
| * | geno_dhfr_108 == “*” | Heterozygous call | Undetermined |
| - | geno_dhfr_108 == “-” | Missing | Missing |
| 108-nonN | otherwise | Unknown mutant | Undetermined |
| Reference | Method | Location/Sample size/Genetic background | Finding summary |
|---|---|---|---|
| Sirawaraporn W, Sathitkul T, Sirawaraporn R, Yuthavong Y, Santi DV. Antifolate-resistant mutants of Plasmodium falciparum dihydrofolate reductase. Proc Natl Acad Sci U S A. 1997;94(4):1124-9. | - pf DHFR mutations at residues 16, 51, 59, 108 and 164 were generated to assess the correlation between these mutations and pyrimethamine and cycloguanil resistance. - Enzyme assay and kinetic analysis of DHFR mutants were used to determine drug susceptibility i.e. IC50 and Kinetic parameters. |
- The DHFR mutant fragment was inserted into pET expression vector and expressed in E. coli BL21(DE3) pLysS strain. | - Inhibitory constant (Ki) for pyrimethamine of the quadruple DHFR mutant fragment (Ki = 859 nM), with mutations at residues 51, 59, 108 and 164, was 500-fold higher than the Ki value of wild-type DHFR fragment (Ki = 1.5 nM). |
| Yuvaniyama J, Chitnumsub P, Kamchonwongpaisan S, Vanichtanankul J, Sirawaraporn W, Taylor P, et al. Insights into antifolate resistance from malarial DHFR-TS structures. Nat Struct Biol. 2003;10(5):357-65. | - Crystal structure of pf DHFR-TS wild-type and quadruple mutant were determined with NADPH, dUMP and WR99210. Whereas the double mutant is in complex with NADPH, dUMP and PYR. - Inhibition kinetics (Ki) and drug sensitivities (IC50) of wild-type and mutant were determined. |
- TM4/8.2 line (Wild type). - K1 and CB1 lines (C59R/S108N). - V1/S line (N51I/C59R/S108N/I164L). |
- The crystal structure suggested that the binding ability to pyrimenthamine of the quadruple mutant enzyme decreased and caused pyrimethamine resistance. - IC50 values of the parasites with wild-type pfphfr, 59+108 double mutation and 51+59+108+164 quadruple mutation were 0.08 uM 30.9 uM and >100 uM, respectively. |