Plasmodium falciparum resistance to piperaquine driven by PfCRT

Dhingra, Satish K.; Small-Saunders, Jennifer L.; Ménard, Didier; Fidock, David A.

doi:10.1016/s1473-3099(19)30543-2

Cited by 59 publications

(99 citation statements)

References 5 publications

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“…All isolates carrying two copies of pfpm2 or pfmdr1 were WT for Pfkelch13 (Supplementary Table 3). We also tested 14 of the 20 Pfkelch13 561H mutants for mutations in the chloroquine resistance transporter gene (pfcrt; PF3D7_0709000), whose variants can confer resistance to chloroquine or piperaquine 31,32 . All 561H mutants carried WT pfcrt ( Supplementary Table 4).…”

Section: Letters Nature Medicinementioning

confidence: 99%

Emergence and clonal expansion of in vitro artemisinin-resistant Plasmodium falciparum kelch13 R561H mutant parasites in Rwanda

Uwimana

Legrand

Stokes

et al. 2020

Nat Med

558

560

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Artemisinin resistance (delayed P. falciparum clearance following artemisinin-based combination therapy), is widespread across Southeast Asia but to date has not been reported in Africa1–4. Here we genotyped the P. falciparum K13 (Pfkelch13) propeller domain, mutations in which can mediate artemisinin resistance5,6, in pretreatment samples collected from recent dihydroarteminisin-piperaquine and artemether-lumefantrine efficacy trials in Rwanda7. While cure rates were >95% in both treatment arms, the Pfkelch13 R561H mutation was identified in 19 of 257 (7.4%) patients at Masaka. Phylogenetic analysis revealed the expansion of an indigenous R561H lineage. Gene editing confirmed that this mutation can drive artemisinin resistance in vitro. This study provides evidence for the de novo emergence of Pfkelch13-mediated artemisinin resistance in Rwanda, potentially compromising the continued success of antimalarial chemotherapy in Africa.

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Section: Letters Nature Medicinementioning

confidence: 99%

Emergence and clonal expansion of in vitro artemisinin-resistant Plasmodium falciparum kelch13 R561H mutant parasites in Rwanda

Uwimana

Legrand

Stokes

et al. 2020

Nat Med

558

560

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“…The PPQ survival assay (PSA) has been developed, with a survival rate of more than 10% de ning a PPQ resistant phenotype [18]. Genetic markers proposed as modulators of PPQ resistance include P. falciparum multidrug resistance 1 (pfmdr1) (PF3D7_0523000) [19][20][21], P. falciparum plasmepsin 2 (pfpm2) (PF3D7_1408000) [20][21][22][23], P. falciparum exonuclease (pfexo) (PF3D7_1362500) [20,23,24], and speci c mutations on P. falciparum chloroquine resistance transporter (pfcrt) (PF3D7_0709000) [18,[25][26][27]. An association between in vitro PPQ-resistant isolates and single-copy pfmdr1 was found [6,7,18,28]; however, not all single copy pfmdr1 isolates demonstrate in vitro PPQ resistance.…”

Section: Introductionmentioning

confidence: 99%

“…However, the role of E415G-Exo in mediating PPQ resistance, in the absence of other markers of resistance, is unclear. Speci c novel mutations of the pfcrt gene have been shown to be associated with PPQ resistance since parasites with a variant of the Dd2 pfcrt allele, either T93S, H97Y, F145I, I218F, M343L, or G353V, have higher median PSA survival rates than those harboring the wildtype Dd2 allele [18,26,27], and a C350R substitution in the pfcrt gene resulted in decreased susceptibility to PPQ [33]. Dhingra et al [27] showed that T93S and I218F-PfCRT mutations have increased in the past 5 years in Southeast Asia although showing an insigni cant tness cost compared to F145I-parasites.…”

Section: Introductionmentioning

confidence: 99%

Piperaquine resistant Cambodian Plasmodium falciparum clinical isolates: in vitro genotypic and phenotypic characterization

Boonyalai

Vesely

Thamnurak

et al. 2020

Preprint

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Abstract Background High rates of dihydroartemisinin-piperaquine (DHA-PPQ) treatment failures have been documented for uncomplicated Plasmodium falciparum in Cambodia. The genetic markers plasmepsin 2 ( pfpm2 ), exonuclease ( pfexo ) and chloroquine resistance transporter ( pfcrt ) genes are associated with PPQ resistance and are used for monitoring the prevalence of drug resistance and guiding malaria drug treatment policy.Methods To examine the relative contribution of each marker to PPQ resistance, in vitro culture and the PPQ survival assay were performed on seventeen P. falciparum isolates from northern Cambodia, and the presence of E415G-Exo and pfcrt mutations (T93S, H97Y, F145I, I218F, M343L, C350R, and G353V) as well as pfpm2 copy number polymorphisms were determined. Parasites were then cloned by limiting dilution and the cloned parasites were tested for drug susceptibility. Isobolographic analysis of several drug combinations for standard clones and newly cloned P. falciparum Cambodian isolates was also determined.Results The characterization of culture-adapted isolates revealed that the presence of novel pfcrt mutations (T93S, H97Y, F145I, and I218F) with E415G-Exo mutation can confer PPQ-resistance, in the absence of pfpm2 amplification. In vitro testing of PPQ resistant parasites demonstrated a bimodal dose-response, the existence of a swollen digestive vacuole phenotype, and an increased susceptibility to quinine, chloroquine, mefloquine and lumefantrine. To further characterize drug sensitivity, parental parasites were cloned in which a clonal line, 14-B5, was identified as sensitive to artemisinin and piperaquine, but resistant to chloroquine. Assessment of the clone against a panel of drug combinations revealed antagonistic activity for six different drug combinations. However, mefloquine-proguanil and atovoquone-proguanil combinations revealed synergistic antimalarial activity.Conclusions Surveillance for PPQ resistance in regions relying on DHA-PPQ as the first-line treatment is dependent on the monitoring of molecular markers of drug resistance. P. falciparum harbouring novel pfcrt mutations with E415G-exo mutations displayed PPQ resistant phenotype. The presence of pfpm2 amplification was not required to render parasites PPQ resistant suggesting that the increase in pfpm2 copy number alone is not the sole modulator of PPQ resistance. Genetic background of circulating field isolates appear to play a role in drug susceptibility and biological responses induced by drug combinations. The use of latest field isolates may be necessary for assessment of relevant drug combinations against P. falciparum strains and when down-selecting novel drug candidates.

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“…The PPQ survival assay (PSA) has been developed, with a survival rate of more than 10% defining a PPQ resistant phenotype [17]. Several genetic markers have been proposed as modulators of PPQ resistance, such as P. falciparum multidrug resistance 1 (pfmdr1) (PF3D7_0523000) [18][19][20], plasmepsin 2 (PM2) (PF3D7_1408000) [19][20][21][22], E415G exonuclease (Exo) (PF3D7_1362500) [19,22,23], and specific mutations on P. falciparum chloroquine resistance transporter (PfCRT) (PF3D7_0709000) [17,[24][25][26]. An association between in vitro PPQ-resistant isolates and single-copy pfmdr1 was found [6,7,17,27]; however, not all single copy pfmdr1 isolates demonstrated in vitro PPQ resistance and the association of single copy number of pfmdr1 with decreased sensitivity may be circumstantial [19], highlighting the need to characterize more specific PPQ markers of drug resistance.…”

Section: Introductionmentioning

confidence: 99%

“…However, the role of exo-E415G in mediating PPQ resistance, in the absence of other markers of resistance, is unclear. The specific novel mutations of the pfcrt gene are believed to be associated with PPQ resistance since parasites with a variant of the Dd2 pfcrt allele, either T93S, H97Y, F145I, I218F, M343L, or G353V, have higher median PSA survival rates than those harboring the Dd2 allele [17,25,26], and a C350R substitution in the pfcrt gene decreased susceptibility to PPQ [31]. Dhingra,et.…”

Section: Introductionmentioning

confidence: 99%

Piperaquine resistant Cambodian Plasmodium falciparum clinical isolates: in vitro genotypic and phenotypic characterization

Boonyalai

Vesely

Thamnurak

et al. 2020

Preprint

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Background: High rates of dihydroartemisinin-piperaquine (DHA-PPQ) treatment failures have been documented for uncomplicated Plasmodium falciparum in Cambodia. It is essential to establish sensitive and reliable markers of PPQ resistance which can be adopted for monitoring the prevalence of drug resistance and guide drug policy change. Several genetic markers have been proposed such as copy numbers of P. falciparum multidrug resistance 1 ( pfmdr1 ) and plasmepsin 2 (PM2), and mutations in exonuclease (exo-E415G) and P. falciparum chloroquine resistance transporter (PfCRT) genes.Methods: To examine the relative contribution of each marker to PPQ resistance, in vitro culture and the PPQ survival assay were performed on seventeen P. falciparum isolates from northern Cambodia and the presence of exo-E415G and PfCRT mutations as well as PM2 copy number polymorphisms were determined. Parasites were then cloned by limiting dilution and the cloned parasites were tested for drug susceptibility. The efficacy of several drug combinations between standard clones and newly cloned P. falciparum Cambodian isolates was also measured.Results: The characterization of culture-adapted isolates revealed that exo-E415G and novel PfCRT mutations can confer PPQ-resistance, in the absence of PM2 amplification. In vitro testing of PPQ resistant parasites showed bimodal dose-response phenotype as previously reported in both Cambodian isolates and genome-edited Dd2 strains. Our finding is the first PPQ resistant clinical isolate with documented swollen digestive vacuole phenotype. From the field isolates, we were able to clone a new parasite line, 14-B5, which is sensitive to arteminsinin and piperaquine but resistant to chloroquine. Most of the drug combinations tested revealed antagonistic interaction against the 14-B5 line, indicating its different genetic background from other P. falciparum laboratory reference lines.Conclusions: Surveillance for PPQ resistance in regions that rely on DHA-PPQ as the first line treatment should depend on the monitoring of the reflective molecular markers. Bimodal dose response curves and swelling of the food vacuole can be used as PPQ resistant phenotype. The use of currently circulating parasite isolates is necessary for relevant drug combination development against current P. falciparum resistance profiles.

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Plasmodium falciparum resistance to piperaquine driven by PfCRT

Cited by 59 publications

References 5 publications

Emergence and clonal expansion of in vitro artemisinin-resistant Plasmodium falciparum kelch13 R561H mutant parasites in Rwanda

Emergence and clonal expansion of in vitro artemisinin-resistant Plasmodium falciparum kelch13 R561H mutant parasites in Rwanda

Piperaquine resistant Cambodian Plasmodium falciparum clinical isolates: in vitro genotypic and phenotypic characterization

Piperaquine resistant Cambodian Plasmodium falciparum clinical isolates: in vitro genotypic and phenotypic characterization

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