Multigenic architecture of piperaquine resistance trait in Plasmodium falciparum

Silva, Miguel; Calçada, Carla; Teixeira, Miguel G; Veiga, Maria Isabel; Ferreira, Pedro Eduardo

doi:10.1016/s1473-3099(19)30689-9

Cited by 21 publications

(23 citation statements)

References 5 publications

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“…Based on the limited combinations and parasites screened as part of these initial efforts, we were able to identify combinations of PYR and PQP that have an antagonistic interactions in parasite lines with amplified plasmepsin II/III copy number variations (CNVs), a highly prevalent genotype in the Greater Mekong Subregion (11,29,30). These isolates were collected as part of a DHA-PQP efficacy study that demonstrated decreased clinical efficacy of the ACT (19) (35). Further investigation to validate and elucidate the molecular mechanism mediating the induction of an antagonistic interaction between PQP and PYR in the context of plasmepsin II/III CNV will be required, which may also aid in understanding the specific mechanism of action of these two antimalarial compounds.…”

Section: Discussionmentioning

confidence: 99%

Modulation of triple artemisinin-based combination therapy pharmacodynamics byPlasmodium falciparumgenotype

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Itkin

Chen

et al. 2020

Preprint

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AbstractThe first-line treatments for uncomplicated Plasmodium falciparum malaria are artemisinin-based combination therapies (ACTs), consisting of an artemisinin derivative combined with a longer acting partner drug. However, the spread of P. falciparum with decreased susceptibility to artemisinin and partner drugs presents a significant challenge to malaria control efforts. To stem the spread of drug resistant parasites, novel chemotherapeutic strategies are being evaluated, including the implementation of triple artemisinin-based combination therapies (TACTs). Currently, there is limited knowledge on the pharmacodynamics and pharmacogenetic interactions of proposed TACT drug combinations. To evaluate these interactions, we established an in vitro high-throughput process for measuring the drug dose-response to three distinct antimalarial drugs present in a TACT. Sixteen different TACT combinations were screened against fifteen parasite lines from Cambodia, with a focus on parasites with differential susceptibilities to piperaquine and artemisinins. Analysis revealed drug-drug interactions unique to specific genetic backgrounds, including antagonism between piperaquine and pyronaridine associated with gene amplification of plasmepsin II/III, two aspartic proteases that localize to the parasite digestive vacuole. From this initial study, we identified parasite genotypes with decreased susceptibility to specific TACTs, as well as potential TACTs that display antagonism in a genotype-dependent manner. Our assay and analysis platform can be further leveraged to inform drug implementation decisions and evaluate next-generation TACTs.One Sentence SummaryIn vitro process to evaluate triple-drug combinations for prioritizing antimalarial combinations for in vivo evaluation.

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Section: Discussionmentioning

confidence: 99%

Modulation of triple artemisinin-based combination therapy pharmacodynamics byPlasmodium falciparumgenotype

Ansbro

Itkin

Chen

et al. 2020

Preprint

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“…However, the overexpression of pfpm2 and pfpm3 in the 3D7 genetic background did not alter the sensitivity of P. falciparum to PPQ, suggesting that the increase in pfpm2 copy number alone is not the sole modulator of PPQ resistance [ 31 ]. Silva et al [ 32 ] have utilized gene editing and chemical inhibition to demonstrate that pfpm2 amplification contributed to PPQ resistance and that the background of the engineered parasites was necessary to gain a bimodal dose–response, in which a second peak of survival for a subset of the parasite is detected at higher PPQ concentrations [ 22 ]. It was also suggested that the initial selection of plasmepsin and pfmdr1 copy number variations developed a genetic background important for novel pfcrt mutations to emerge [ 32 ].…”

Section: Introductionmentioning

confidence: 99%

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

Boonyalai

Vesely

Thamnurak

et al. 2020

Malar J

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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 atovaquone-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

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“…Silva et al [32] have utilized gene editing and chemical inhibition to demonstrate that pfpm2 ampli cation contributed to PPQ resistance and that the background of the engineered parasites was necessary to gain a bimodal dose-response, in which a second peak of survival for a subset of the parasite is detected at higher PPQ concentrations [22]. It was also suggested that the initial selection of plasmepsin and pfmdr1 copy number variations developed a genetic background important for novel pfcrt mutations to emerge [32].…”

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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Multigenic architecture of piperaquine resistance trait in Plasmodium falciparum

Abstract: for regulatory and ethical submission. We declare no competing interests.

Cited by 21 publications

References 5 publications

Modulation of triple artemisinin-based combination therapy pharmacodynamics byPlasmodium falciparumgenotype

Modulation of triple artemisinin-based combination therapy pharmacodynamics byPlasmodium falciparumgenotype

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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