Baseline In Vitro Activities of the Antimalarials Pyronaridine and Methylene Blue against Plasmodium falciparum Isolates from Kenya

Abstract

We have analyzed the in vitro activities of pyronaridine and methylene blue against 59 Plasmodium falciparum isolates from Kenya in association with polymorphisms in Pfcrt (codon 76), Pfmdr1 (codon 86), and Pfnhe (full sequence). The median inhibitory concentrations that kill 50% of parasites were 13.5 and 3.3 nM for pyronaridine and methylene blue, respectively. Their activities were not associated with polymorphisms in these genes. The drugs' high in vitro activities indicate that they would be efficacious against Kenyan isolates in vivo.C oartem (lumefantrine [LM] and artemether) and amodiaquine (AQ)-artesunate (ART) are currently the first lines of treatment of uncomplicated malaria (8,29). However, reports indicate that resistance to LM may arise relatively quickly (30). Likewise, evidence suggests that the efficacy of AQ, whose active in vivo metabolite is desethylamodiaquine (DEAQ), is reduced in areas of high chloroquine (CQ) resistance (12).The combinations piperaquine (PIQ)-dihydroartemisinin (DHA) and pyronaridine (PRN)-ART are being developed as alternative antimalarials (2). In spite of these alternatives, the search for new active compounds is being pursued. Methylene blue (MB) is an old antimalarial that was abandoned because of its side effect of turning urine blue. However, this drug has been extensively used for the treatment of methemoglobinemia (5). The burgeoning problem of drug resistance has led to a renewed interest in this drug (28). In this paper, we report on the in vitro activities of PRN and MB against Plasmodium falciparum field isolates in Kenya and on the change in their activities in relation to polymorphisms in Pfcrt at codon 76 (Pfcrt-76), in Pfmdr1 at codon 86 (Pfmdr1-86), and in Pfnhe.We analyzed fresh isolates of Plasmodium falciparum collected in the Kenyan district of Kilifi and adapted for long-term cultures as detailed previously (13,15). Antimalarial activity was measured in the presence of various concentrations of each compound, and results were expressed as the drug concentration required for 50% inhibition of [ 3 H]hypoxanthine incorporation into parasite nucleic acid (IC 50 ) (15). We employed two reference strains: V1S, the multidrug-resistant strain, and 3D7, the drug-sensitive strain. We analyzed the antimalarials chloroquine (CQ), MB, AQ, and quinine (QN) (purchased from Sigma Chemical Co., Poole, Dorset, United Kingdom) and LM, PIQ, DEAQ, and DHA (gifts from Steve Ward, Liverpool School of Tropical Medicine, Liverpool, United Kingdom).Blood samples (50 l) of in vitro-adapted isolates were spotted onto filter paper, and single-base changes at Pfcrt-76 and Pfmdr1-86 were detected as reported elsewhere (13). In this paper, we reanalyzed the sequencing of Pfnhe published previously (15).Statistical analyses were carried out using the Stata program (Stata version 11; College Station, TX). We compared differences between groups using the Wilcoxon rank-sum test and measured correlations using the nonparametric Spearman pairwise analysis. All statistical analysis was...

In Vitro Sensitivity of Pyronaridine in Thai Isolates of Plasmodium falciparum

“…This result is similar to previous studies conducted in Thailand 30 years ago and elsewhere at the present time. 16,29,35,36,[38][39][40][41][42][43] The clear standard cut-off point of pyronaridine is still unavailable. A related study suggested estimating using the geometric mean plus 2 SDs.…”

“…Pyronaridine IC 50 was also positively correlated with artesunate IC 50 which is similar to two related studies. 29,41 However, a statistically significant result (P < 0.05) can often be seen when comparing the drug activities against a large number of isolates. The r value > 0.7-0.8 or r 2 value > 0.5 is more suggestive of cross-resistance.…”

Machine learning models identify molecules active against the Ebola virus in vitro

“…Quinacrine is an old antimalarial drug now more widely used as an antiprotozoal for the treatment of giardiasis 66 and as an anthelmintic. Pyronaridine is a potent antimalarial (IC 50 13.5 nM) 67 , has activity against Babesi a spp. 68 , is active in vitro (EC 50 225 nM) and in vivo (85.2% efficacy 4 days treatment at 50 mg/kg) against T. cruzi 30 and is a P-glycoprotein inhibitor 69 .…”

Safety and efficacy of pyronaridine-artesunate in uncomplicated acute malaria: an integrated analysis of individual patient data from six randomized clinical trials

“…In vitro studies indicate potent activity against recent isolates of multidrug-resistant strains of both P. falciparum and P. vivax from Papua, Indonesia [26], against Kenyan P. falciparum isolates [27], and against around half of chloroquine-resistant P. falciparum strains from the China–Myanmar border area [28]. The PA clinical development programme included two Phase II trials and three Phase III trials in children and adults from Africa and Asia with uncomplicated P. falciparum malaria [15,22,24,25], as well as one comparative Phase III trial in children and adults with uncomplicated P. vivax infection [23].…”