A Variant PfCRT Isoform Can Contribute to <italic toggle="yes">Plasmodium falciparum</italic> Resistance to the First-Line Partner Drug Piperaquine

ABSTRACT Current efforts to reduce the global burden of malaria are threatened by the rapid spread throughout Asia of Plasmodium falciparum resistance to artemisinin-based combination therapies, which includes increasing rates of clinical failure with dihydroartemisinin plus piperaquine (PPQ) in Cam...

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Autores principales:	Satish K. Dhingra, Devasha Redhi, Jill M. Combrinck, Tomas Yeo, John Okombo, Philipp P. Henrich, Annie N. Cowell, Purva Gupta, Matthew L. Stegman, Jonathan M. Hoke, Roland A. Cooper, Elizabeth Winzeler, Sachel Mok, Timothy J. Egan, David A. Fidock
Formato:	article
Lenguaje:	EN
Publicado:	American Society for Microbiology 2017
Materias:	malaria PfCRT Plasmodium falciparum artemisinin-based combination therapies digestive vacuole genome editing Microbiology QR1-502
Acceso en línea:	https://doaj.org/article/9c6d366c3a2a445c9fb90b0de038bd88
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spelling	oai:doaj.org-article:9c6d366c3a2a445c9fb90b0de038bd882021-11-15T15:51:30ZA Variant PfCRT Isoform Can Contribute to <italic toggle="yes">Plasmodium falciparum</italic> Resistance to the First-Line Partner Drug Piperaquine10.1128/mBio.00303-172150-7511https://doaj.org/article/9c6d366c3a2a445c9fb90b0de038bd882017-07-01T00:00:00Zhttps://journals.asm.org/doi/10.1128/mBio.00303-17https://doaj.org/toc/2150-7511ABSTRACT Current efforts to reduce the global burden of malaria are threatened by the rapid spread throughout Asia of Plasmodium falciparum resistance to artemisinin-based combination therapies, which includes increasing rates of clinical failure with dihydroartemisinin plus piperaquine (PPQ) in Cambodia. Using zinc finger nuclease-based gene editing, we report that addition of the C101F mutation to the chloroquine (CQ) resistance-conferring PfCRT Dd2 isoform common to Asia can confer PPQ resistance to cultured parasites. Resistance was demonstrated as significantly higher PPQ concentrations causing 90% inhibition of parasite growth (IC90) or 50% parasite killing (50% lethal dose [LD50]). This mutation also reversed Dd2-mediated CQ resistance, sensitized parasites to amodiaquine, quinine, and artemisinin, and conferred amantadine and blasticidin resistance. Using heme fractionation assays, we demonstrate that PPQ causes a buildup of reactive free heme and inhibits the formation of chemically inert hemozoin crystals. Our data evoke inhibition of heme detoxification in the parasite’s acidic digestive vacuole as the primary mode of both the bis-aminoquinoline PPQ and the related 4-aminoquinoline CQ. Both drugs also inhibit hemoglobin proteolysis at elevated concentrations, suggesting an additional mode of action. Isogenic lines differing in their pfmdr1 copy number showed equivalent PPQ susceptibilities. We propose that mutations in PfCRT could contribute to a multifactorial basis of PPQ resistance in field isolates. IMPORTANCE The global agenda to eliminate malaria depends on the continued success of artemisinin-based combination therapies (ACTs), which target the asexual blood stages of the intracellular parasite Plasmodium. Partial resistance to artemisinin, however, is now established in Southeast Asia, exposing the partner drugs to increased selective pressure. Plasmodium falciparum resistance to the first-line partner piperaquine (PPQ) is now spreading rapidly in Cambodia, resulting in clinical treatment failures. Here, we report that a variant form of the Plasmodium falciparum chloroquine resistance transporter, harboring a C101F mutation edited into the chloroquine (CQ)-resistant Dd2 isoform prevalent in Asia, can confer PPQ resistance in cultured parasites. This was accompanied by a loss of CQ resistance. Biochemical assays showed that PPQ, like CQ, inhibits the detoxification of reactive heme that is formed by parasite-mediated catabolism of host hemoglobin. We propose that novel PfCRT variants emerging in the field could contribute to a multigenic basis of PPQ resistance.Satish K. DhingraDevasha RedhiJill M. CombrinckTomas YeoJohn OkomboPhilipp P. HenrichAnnie N. CowellPurva GuptaMatthew L. StegmanJonathan M. HokeRoland A. CooperElizabeth WinzelerSachel MokTimothy J. EganDavid A. FidockAmerican Society for MicrobiologyarticlemalariaPfCRTPlasmodium falciparumartemisinin-based combination therapiesdigestive vacuolegenome editingMicrobiologyQR1-502ENmBio, Vol 8, Iss 3 (2017)
institution	DOAJ
collection	DOAJ
language	EN
topic	malaria PfCRT Plasmodium falciparum artemisinin-based combination therapies digestive vacuole genome editing Microbiology QR1-502
spellingShingle	malaria PfCRT Plasmodium falciparum artemisinin-based combination therapies digestive vacuole genome editing Microbiology QR1-502 Satish K. Dhingra Devasha Redhi Jill M. Combrinck Tomas Yeo John Okombo Philipp P. Henrich Annie N. Cowell Purva Gupta Matthew L. Stegman Jonathan M. Hoke Roland A. Cooper Elizabeth Winzeler Sachel Mok Timothy J. Egan David A. Fidock A Variant PfCRT Isoform Can Contribute to <italic toggle="yes">Plasmodium falciparum</italic> Resistance to the First-Line Partner Drug Piperaquine
description	ABSTRACT Current efforts to reduce the global burden of malaria are threatened by the rapid spread throughout Asia of Plasmodium falciparum resistance to artemisinin-based combination therapies, which includes increasing rates of clinical failure with dihydroartemisinin plus piperaquine (PPQ) in Cambodia. Using zinc finger nuclease-based gene editing, we report that addition of the C101F mutation to the chloroquine (CQ) resistance-conferring PfCRT Dd2 isoform common to Asia can confer PPQ resistance to cultured parasites. Resistance was demonstrated as significantly higher PPQ concentrations causing 90% inhibition of parasite growth (IC90) or 50% parasite killing (50% lethal dose [LD50]). This mutation also reversed Dd2-mediated CQ resistance, sensitized parasites to amodiaquine, quinine, and artemisinin, and conferred amantadine and blasticidin resistance. Using heme fractionation assays, we demonstrate that PPQ causes a buildup of reactive free heme and inhibits the formation of chemically inert hemozoin crystals. Our data evoke inhibition of heme detoxification in the parasite’s acidic digestive vacuole as the primary mode of both the bis-aminoquinoline PPQ and the related 4-aminoquinoline CQ. Both drugs also inhibit hemoglobin proteolysis at elevated concentrations, suggesting an additional mode of action. Isogenic lines differing in their pfmdr1 copy number showed equivalent PPQ susceptibilities. We propose that mutations in PfCRT could contribute to a multifactorial basis of PPQ resistance in field isolates. IMPORTANCE The global agenda to eliminate malaria depends on the continued success of artemisinin-based combination therapies (ACTs), which target the asexual blood stages of the intracellular parasite Plasmodium. Partial resistance to artemisinin, however, is now established in Southeast Asia, exposing the partner drugs to increased selective pressure. Plasmodium falciparum resistance to the first-line partner piperaquine (PPQ) is now spreading rapidly in Cambodia, resulting in clinical treatment failures. Here, we report that a variant form of the Plasmodium falciparum chloroquine resistance transporter, harboring a C101F mutation edited into the chloroquine (CQ)-resistant Dd2 isoform prevalent in Asia, can confer PPQ resistance in cultured parasites. This was accompanied by a loss of CQ resistance. Biochemical assays showed that PPQ, like CQ, inhibits the detoxification of reactive heme that is formed by parasite-mediated catabolism of host hemoglobin. We propose that novel PfCRT variants emerging in the field could contribute to a multigenic basis of PPQ resistance.
format	article
author	Satish K. Dhingra Devasha Redhi Jill M. Combrinck Tomas Yeo John Okombo Philipp P. Henrich Annie N. Cowell Purva Gupta Matthew L. Stegman Jonathan M. Hoke Roland A. Cooper Elizabeth Winzeler Sachel Mok Timothy J. Egan David A. Fidock
author_facet	Satish K. Dhingra Devasha Redhi Jill M. Combrinck Tomas Yeo John Okombo Philipp P. Henrich Annie N. Cowell Purva Gupta Matthew L. Stegman Jonathan M. Hoke Roland A. Cooper Elizabeth Winzeler Sachel Mok Timothy J. Egan David A. Fidock
author_sort	Satish K. Dhingra
title	A Variant PfCRT Isoform Can Contribute to <italic toggle="yes">Plasmodium falciparum</italic> Resistance to the First-Line Partner Drug Piperaquine
title_short	A Variant PfCRT Isoform Can Contribute to <italic toggle="yes">Plasmodium falciparum</italic> Resistance to the First-Line Partner Drug Piperaquine
title_full	A Variant PfCRT Isoform Can Contribute to <italic toggle="yes">Plasmodium falciparum</italic> Resistance to the First-Line Partner Drug Piperaquine
title_fullStr	A Variant PfCRT Isoform Can Contribute to <italic toggle="yes">Plasmodium falciparum</italic> Resistance to the First-Line Partner Drug Piperaquine
title_full_unstemmed	A Variant PfCRT Isoform Can Contribute to <italic toggle="yes">Plasmodium falciparum</italic> Resistance to the First-Line Partner Drug Piperaquine
title_sort	variant pfcrt isoform can contribute to <italic toggle="yes">plasmodium falciparum</italic> resistance to the first-line partner drug piperaquine
publisher	American Society for Microbiology
publishDate	2017
url	https://doaj.org/article/9c6d366c3a2a445c9fb90b0de038bd88
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A Variant PfCRT Isoform Can Contribute to <italic toggle="yes">Plasmodium falciparum</italic> Resistance to the First-Line Partner Drug Piperaquine

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