title: Pfmdr1 copy number and arteminisin derivatives combination therapy failure in falciparum malaria in Cambodia.
abstract: BACKGROUND
  The combination of artesunate and mefloquine was introduced as the national first-line treatment for Plasmodium falciparum malaria in Cambodia in 2000. However, recent clinical trials performed at the Thai-Cambodian border have pointed to the declining efficacy of both artesunate-mefloquine and artemether-lumefantrine. Since pfmdr1 modulates susceptibility to mefloquine and artemisinin derivatives, the aim of this study was to assess the link between pfmdr1 copy number, in vitro susceptibility to individual drugs and treatment failure to combination therapy.
  
  
  METHODS
  Blood samples were collected from P. falciparum-infected patients enrolled in two in vivo efficacy studies in north-western Cambodia: 135 patients were treated with artemether-lumefantrine (AL group) in Sampovloun in 2002 and 2003, and 140 patients with artesunate-mefloquine (AM group) in Sampovloun and Veal Veng in 2003 and 2004. At enrollment, the in vitro IC50 was tested and the strains were genotyped for pfmdr1 copy number by real-time PCR.
  
  
  RESULTS
  The pfmdr1 copy number was analysed for 115 isolates in the AM group, and for 109 isolates in the AL group. Parasites with increased pfmdr1 copy number had significantly reduced in vitro susceptibility to mefloquine, lumefantrine and artesunate. There was no association between pfmdr1 polymorphisms and in vitro susceptibilities. In the patients treated with AM, the mean pfmdr1copy number was lower in subjects with adequate clinical and parasitological response compared to those who experienced late treatment failure (n = 112, p < 0.001). This was not observed in the patients treated with AL (n = 96, p = 0.364). The presence of three or more copies of pfmdr1 were associated with recrudescence in artesunate-mefloquine treated patients (hazard ratio (HR) = 7.80 [95%CI: 2.09-29.10], N = 115), p = 0.002) but not with recrudescence in artemether-lumefantrine treated patients (HR = 1.03 [95%CI: 0.24-4.44], N = 109, p = 0.969).
  
  
  CONCLUSIONS
  This study shows that pfmdr1 copy number is a molecular marker of AM treatment failure in falciparum malaria on the Thai-Cambodian border. However, while it is associated with increased IC50 for lumefantrine, pfmdr1 copy number is not associated with AL treatment failure in the area, suggesting involvement of other molecular mechanisms in AL treatment failures in Cambodia.
authors.affiliation: Institut Pasteur in Cambodia, Phnom Penh, Cambodia. pharath@pasteur-kh.org
authors.forename: Pharath
authors.initials: P
authors.lastname: Lim
authors.forename: Alisa P
authors.initials: AP
authors.lastname: Alker
authors.forename: Nimol
authors.initials: N
authors.lastname: Khim
authors.forename: Naman K
authors.initials: NK
authors.lastname: Shah
authors.forename: Sandra
authors.initials: S
authors.lastname: Incardona
authors.forename: Socheat
authors.initials: S
authors.lastname: Doung
authors.forename: Poravuth
authors.initials: P
authors.lastname: Yi
authors.forename: Denis Mey
authors.initials: DM
authors.lastname: Bouth
authors.forename: Christiane
authors.initials: C
authors.lastname: Bouchier
authors.forename: Odile Mercereau
authors.initials: OM
authors.lastname: Puijalon
authors.forename: Steven R
authors.initials: SR
authors.lastname: Meshnick
authors.forename: Chansuda
authors.initials: C
authors.lastname: Wongsrichanalai
authors.forename: Thierry
authors.initials: T
authors.lastname: Fandeur
authors.forename: Jacques
authors.initials: J
authors.lastname: Le Bras
authors.forename: Pascal
authors.initials: P
authors.lastname: Ringwald
authors.forename: Frédéric
authors.initials: F
authors.lastname: Ariey
mesh_terms: D000328:Adult; D000818:Animals; D000962:Antimalarials / Q000494:pharmacology / Q000627:therapeutic use*; D000077611:Artemether, Lumefantrine Drug Combination; D037621:Artemisinins / Q000494:pharmacology / Q000627:therapeutic use*; D000077332:Artesunate; D002160:Cambodia; D002648:Child; D002986:Clinical Trials as Topic; D004338:Drug Combinations; D004351:Drug Resistance / Q000235:genetics*; D004983:Ethanolamines / Q000494:pharmacology / Q000627:therapeutic use*; D005449:Fluorenes / Q000494:pharmacology / Q000627:therapeutic use*; D018628:Gene Dosage / Q000235:genetics*; D014644:Genetic Variation; D006801:Humans; D020128:Inhibitory Concentration 50; D016778:Malaria, Falciparum / Q000188:drug therapy*; D015767:Mefloquine / Q000494:pharmacology; D027425:ATP-Binding Cassette, Sub-Family C Proteins / Q000235:genetics*; D010963:Plasmodium falciparum / Q000187:drug effects / Q000235:genetics*; D011110:Polymorphism, Genetic; D016019:Survival Analysis; D013785:Thailand; D017211:Treatment Failure
chemical_list: D000962:Antimalarials; D000077611:Artemether, Lumefantrine Drug Combination; D037621:Artemisinins; D000077332:Artesunate; D004338:Drug Combinations; D004983:Ethanolamines; D005449:Fluorenes; D015767:Mefloquine; D027425:ATP-Binding Cassette, Sub-Family C Proteins; C513498:Mdr1 protein, Plasmodium falciparum