Amodiaquine

Amodiaquine (AQ), a 4-aminoquinoline derivative, is currently being used as a partner drug in combination with artesunate for the treatment of uncomplicated malaria in several African countries. Associations between reduced amodiaquine susceptibility in Plasmodium falciparum and polymorphisms in pfmdr1 and pfcrt have been reported (Folarin OA, et al, 2011, Acta Trop).

• The N86Y pfmdr1 mutation confers 1.5-1.8-fold increase in amodiaquine IC₅₀ (Veiga MI, et al, 2016, Nat Commun).
• Parasites with wild-type pfcrt haplotype (CVMNK at codons 72-76) showed similar amodiaquine IC₅₀ to the parasites with CVIET and SVMNT haplotypes (Lakshmanan V, et al, 2005, EMBO).
• Introducing the 76K mutation into Dd2 parasite line (CVIET haplotype at codons 72-76) and introducing CVIET and CVIEK haplotypes into 7G8 parasite line demonstrated that pfcrt K76T mutations did not cause a significant change in amodiaquine IC₅₀ (Lakshmanan V, et al, 2005, EMBO).

Since strong evidence supporting amodiaquine resistance is still lacking, we do not apply phenotype predicting rules for amodiaquine.

Evidences of amodiaquine resistance

Association study

Reference	Method	Location/Sample size/Genetic background	Finding summary
Folarin OA, Bustamante C, Gbotosho GO, Sowunmi A, Zalis MG, Oduola AM, et al. In vitro amodiaquine resistance and its association with mutations in pfcrt and pfmdr1 genes of Plasmodium falciparum isolates from Nigeria. Acta Trop. 2011;120(3):224-30	- In vitro amodiaquine susceptibility was assessed by using a modified schizont inhibition assay. - The potential molecular markers of amodiaquine resistance, polymorphisms in pfcrt codons 72-76 and pfmdr1 codon 86, were assessed by nested PCR followed by sequencing.	- 97 Plasmodium falciparum isolates from Nigerian patients (Aged 6 months to 12 years).	- Geometric mean amodiaquine IC50 of parasites with N86Y pfmdr1 mutations was approximately 2.5 times higher than that of wild-type pfmdr1 wild-type. - Parasites with pfcrt CVIET haplotype had approximately 9 and 4 times higher amodiaquine IC50 values than those of CVMNK and CVMNT haplotypes, respectively.

Experimental study

Reference	Method	Location/Sample size/Genetic background	Finding summary
Lakshmanan V, Bray PG, Verdier-Pinard D, Johnson DJ, Horrocks P, Muhle RA, et al. A critical role for PfCRT K76T in Plasmodium falciparum verapamil-reversible chloroquine resistance. EMBO J. 2005;24(13):2294-305.	- Transgenic parasites were created by allelic exchange approach. - Drug susceptibility was evaluated by hypoxanthine incorporation assay.	- 7G8 (SVMNT), Dd2 (CVIET), and 3D7 (CVNMK) lines	- 3D7 (CVMNK; IC50 37.2 nM) shows the same level of amodiaquine IC50 to those of 7G8 (SVMNT; IC50 36.7 nM) and Dd2 (CVIET; IC50 42.6 nM). - Introducing the 76T mutation in the parasites with CVIEK background did not alter amodiaquine IC50 (CVIEK; IC50 22.6 nM, CVIET; IC50 26.8 nM). - Introducing the K76 mutation in the CVIET haplotype did not significantly reduce amodiaquine IC50 (CVIET; IC50 50.3 nM, CVIEK; IC50 32.7 nM).
Veiga MI, Dhingra SK, Henrich PP, Straimer J, Gnadig N, Uhlemann AC, et al. Globally prevalent PfMDR1 mutations modulate Plasmodium falciparum susceptibility to artemisinin-based combination therapies. Nat Commun. 2016;7:11553.	- Genetically modified PfMDR1 at amino acid residues 86 and 184 by using zinc-finger nuclease assay. - In vitro drug susceptibility was assessed by using a flow cytometry-based method	- 2 progeny lines of 7G8 (Brazil) and GB4 (Ghana) were used. (1) NF10 line (CVIET pfcrt haplotype at codons 72-76; YF pfmdr1 haplotype at codons 86 and 184). (2) KC5 line (SVMNT pfcrt haplotype at codons 72-76; YF pfmdr1 haplotype at codons 86 and 184).	- Replacing 86Y mutant allele in NF10 and KC5 lines with N86 wild-type allele conferred approximately 1.5-1.8-time decrease in amodiaquine IC50 compared to the parental lines.

See also

• pfmdr1 (PF3D7_0523000)
• Artemisinin