title: CRISPR-Cas9-modified pfmdr1 protects Plasmodium falciparum asexual blood stages and gametocytes against a class of piperazine-containing compounds but potentiates artemisinin-based combination therapy partner drugs.
abstract: Emerging resistance to first-line antimalarial combination therapies threatens malaria treatment and the global elimination campaign. Improved therapeutic strategies are required to protect existing drugs and enhance treatment efficacy. We report that the piperazine-containing compound ACT-451840 exhibits single-digit nanomolar inhibition of the Plasmodium falciparum asexual blood stages and transmissible gametocyte forms. Genome sequence analyses of in vitro-derived ACT-451840-resistant parasites revealed single nucleotide polymorphisms in pfmdr1, which encodes a digestive vacuole membrane-bound ATP-binding cassette transporter known to alter P. falciparum susceptibility to multiple first-line antimalarials. CRISPR-Cas9 based gene editing confirmed that PfMDR1 point mutations mediated ACT-451840 resistance. Resistant parasites demonstrated increased susceptibility to the clinical drugs lumefantrine, mefloquine, quinine and amodiaquine. Stage V gametocytes harboring Cas9-introduced pfmdr1 mutations also acquired ACT-451840 resistance. These findings reveal that PfMDR1 mutations can impart resistance to compounds active against asexual blood stages and mature gametocytes. Exploiting PfMDR1 resistance mechanisms provides new opportunities for developing disease-relieving and transmission-blocking antimalarials.
authors.affiliation: Department of Microbiology and Immunology, Columbia University Medical Center, New York, NY, 10032, USA.
authors.forename: Caroline L
authors.initials: CL
authors.lastname: Ng
authors.affiliation: Dipartimento di Malattie Infettive, Parassitarie ed Immunomediate, Istituto Superiore di Sanità, 00161 Rome, Italy.
authors.forename: Giulia
authors.initials: G
authors.lastname: Siciliano
authors.affiliation: Department of Microbiology and Immunology, Columbia University Medical Center, New York, NY, 10032, USA.
authors.forename: Marcus C S
authors.initials: MC
authors.lastname: Lee
authors.affiliation: Department of Microbiology and Immunology, Columbia University Medical Center, New York, NY, 10032, USA.
authors.forename: Mariana J
authors.initials: MJ
authors.lastname: de Almeida
authors.affiliation: Division of Host-Microbe Systems & Therapeutics, Department of Pediatrics, University of California San Diego, La Jolla, CA, 92093, USA.
authors.forename: Victoria C
authors.initials: VC
authors.lastname: Corey
authors.affiliation: Division of Host-Microbe Systems & Therapeutics, Department of Pediatrics, University of California San Diego, La Jolla, CA, 92093, USA.
authors.forename: Selina E
authors.initials: SE
authors.lastname: Bopp
authors.affiliation: Dipartimento Tecnologie e Salute, Istituto Superiore di Sanità, 00161 Rome, Italy.
authors.forename: Lucia
authors.initials: L
authors.lastname: Bertuccini
authors.affiliation: Department of Medical Parasitology and Infection Biology, Swiss Tropical and Public Health Institute, CH-4002 Basel, Switzerland.
authors.forename: Sergio
authors.initials: S
authors.lastname: Wittlin
authors.affiliation: Department of Microbiology and Immunology, Columbia University Medical Center, New York, NY, 10032, USA.
authors.forename: Rachel G
authors.initials: RG
authors.lastname: Kasdin
authors.affiliation: Department of Drug Discovery, Actelion Pharmaceuticals Ltd., CH-4123 Allschwil, Switzerland.
authors.forename: Amélie
authors.initials: A
authors.lastname: Le Bihan
authors.affiliation: Department of Drug Discovery, Actelion Pharmaceuticals Ltd., CH-4123 Allschwil, Switzerland.
authors.forename: Martine
authors.initials: M
authors.lastname: Clozel
authors.affiliation: Division of Host-Microbe Systems & Therapeutics, Department of Pediatrics, University of California San Diego, La Jolla, CA, 92093, USA.
authors.forename: Elizabeth A
authors.initials: EA
authors.lastname: Winzeler
authors.affiliation: Dipartimento di Malattie Infettive, Parassitarie ed Immunomediate, Istituto Superiore di Sanità, 00161 Rome, Italy.
authors.forename: Pietro
authors.initials: P
authors.lastname: Alano
authors.affiliation: Department of Microbiology and Immunology, Columbia University Medical Center, New York, NY, 10032, USA.|Division of Infectious Diseases, Department of Medicine, Columbia University Medical Center, New York, NY, 10032, USA.
authors.forename: David A
authors.initials: DA
authors.lastname: Fidock
mesh_terms: D018528:ATP-Binding Cassette Transporters / Q000235:genetics / Q000378:metabolism; D000178:Acrylamides / Q000494:pharmacology*; D000962:Antimalarials / Q000494:pharmacology*; D037621:Artemisinins / Q000494:pharmacology*; D064112:Clustered Regularly Interspaced Short Palindromic Repeats; D016054:DNA, Protozoan / Q000235:genetics / Q000378:metabolism; D004351:Drug Resistance; D004357:Drug Synergism; D006801:Humans; D016778:Malaria, Falciparum / Q000188:drug therapy / Q000469:parasitology; D027425:ATP-Binding Cassette, Sub-Family C Proteins / Q000235:genetics / Q000378:metabolism*; D010879:Piperazines / Q000494:pharmacology*; D010963:Plasmodium falciparum / Q000187:drug effects* / Q000235:genetics* / Q000378:metabolism; D017354:Point Mutation; D020641:Polymorphism, Single Nucleotide
chemical_list: D018528:ATP-Binding Cassette Transporters; D000178:Acrylamides; D000962:Antimalarials; D037621:Artemisinins; D016054:DNA, Protozoan; D027425:ATP-Binding Cassette, Sub-Family C Proteins; D010879:Piperazines; C000619473:ACT-451840; C513498:Mdr1 protein, Plasmodium falciparum; C031327:artemisinin