Artemether-lumefantrine (AL) is the most widely used antimalarial drug for treating uncomplicated falciparum malaria. This study evaluated whether the K65Q mutation in the Plasmodium falciparum cysteine desulfurase IscS (Pfnfs1) gene was associated with alternated susceptibility to lumefantrine using clinical parasite samples from Ghana and the China-Myanmar border area. Parasite isolates from the China-Myanmar border had significantly higher IC50 values to lumefantrine than parasites from Ghana. In addition, the K65 allele was significantly more prevalent in the Ghanaian parasites (34.5%) than in the China-Myanmar border samples (6.8%). However, no difference was observed in the lumefantrine IC50 value between the Pfnfs1 reference K65 allele and the non reference 65Q allele in parasites from the two regions. These data suggest that the Pfnfs1 K65Q mutation may not be a reliable marker for reduced susceptibility to lumefantrine.

Artemether-lumefantrine is an artemisinin-based combination therapy for the treatment of malaria, which are primarily metabolized by cytochrome P450 3A4. Therapeutic difference caused by gene polymorphisms of CYP3A4 may lead to uncertain adverse side effects or treatment failure. The aim of this study was to evaluate the effect of CYP3A4 gene polymorphism on artemether-lumefantrine metabolism in vitro. Enzyme kinetics assay was performed using recombinant human CYP3A4 cell microsomes. The analytes, dihydroartimisinin and desbutyl-lumefantrine, were detected by ultra-performance liquid chromatography tandem mass spectrometry. The results demonstrated that compared to CYP3A4.1, the intrinsic clearance of CYP3A4.4, 5, 9, 16, 18, 23, 24, 28, 31-34 significantly reduced for artemether (58.5%-93.3%), and CYP3A4.17 almost loss catalytic activity. Simultaneously, CYP3A4.5, 14, 17, 24 for lumefantrine were decreased by 56.1%-99.6%, and CYP3A4.11, 15, 18, 19, 23, 28, 29, 31-34 for lumefantrine was increased by 51.7%-296%. The variation in clearance rate indicated by molecular docking could be attributed to the disparity in the binding affinity of artemether and lumefantrine with CYP3A4. The data presented here have enriched our understanding of the effect of CYP3A4 gene polymorphism on artemether-lumefantrine metabolizing. These findings serve as a valuable reference and provide insights for guiding the treatment strategy involving artemether-lumefantrine.

Drug resistance in Plasmodium falciparum compromises the effectiveness of antimalarial therapy. This study aimed to evaluate the extent of drug resistance in parasites obtained from international travelers returning from Ghana to guide the management of malaria cases. Eighty-two clinical parasite isolates were obtained from patients returning from Ghana in 2016-2018, of which 29 were adapted to continuous in vitro culture. Their geometric mean IC50 values to a panel of 11 antimalarial drugs, assessed using the standard SYBR Green-I drug sensitivity assay, were 2.1, 3.8, 1.0, 2.7, 17.2, 4.6, 8.3, 8.3, 19.6, 55.1, and 11,555 nM for artemether, artesunate, dihydroartemisinin, lumefantrine, mefloquine, piperaquine, naphthoquine, pyronaridine, chloroquine, quinine, and pyrimethamine, respectively. Except for chloroquine and pyrimethamine, the IC50 values for other tested drugs were below the resistance threshold. The mean ring-stage survival assay value was 0.8%, with four isolates exceeding 1%. The mean piperaquine survival assay value was 2.1%, all below 10%. Mutations associated with chloroquine resistance (pfcrt K76T and pfmdr1 N86Y) were scarce, consistent with the discontinuation of chloroquine a decade ago. Instead, the pfmdr1 86N-184F-1246D haplotype was predominant, suggesting selection by the extensive use of artemether-lumefantrine. No mutations in the pfk13 propeller domain were detected. The pfdhfr/pfdhps quadruple mutant IRNGK associated with resistance to sulfadoxine-pyrimethamine reached an 82% prevalence. In addition, five isolates had pfgch1 gene amplification but, intriguingly, increased susceptibilities to pyrimethamine. This study showed that parasites originating from Ghana were susceptible to artemisinins and the partner drugs of artemisinin-based combination therapies. Genotyping drug resistance genes identified the signature of selection by artemether-lumefantrine. Parasites showed substantial levels of resistance to the antifolate drugs. Continuous resistance surveillance is necessary to guide timely changes in drug policy.

Artemether, an artemisinin derivative, is a component of the commonly used artemisinin-based combination therapy, artemether-lumefantrine. In this study, we cloned the VH and VL genes of a cell line (mAb 2G12E1) producing a monoclonal antibody specific to artemether, and used to construct a recombinant DNA of single-chain variable fragment (scFv). The scFv was constructed into prokaryotic expression vectors pET32a (+), pET22b (+), pGEX-2T, and pMAL-p5x, respectively. However, only the pMAL-p5x/scFv could be induced to express soluble scFv with comparable sensitivity and specificity to that of mAb 2G12E1. Based on the anti-artemether scFv, an indirect competitive enzyme-linked immunosorbent assay (icELISA) was developed. The 50% of inhibition concentration (IC50) value and the working range based on IC20 to IC80 were 4.33 ng mL-1 and 1.05-22.65 ng mL-1, respectively. The artemether content in different drugs were determined by the developed icELISA, and the results were consistent to those determined by ultra performance liquid chromatography (UPLC). The anti-artemether scFv prepared in the current study could be a valuable genetically engineered antibody applied for artemether monitoring and specific binding mechanism studying.

BACKGROUND: Malaria significantly rebounded in 2018 in the Comoros; this created an urgent need to conduct clinical trials to investigate the effectiveness of artemisinin and its derivatives.
METHODS: An open-label, non-randomised controlled trial of artemisinin-piperaquine (AP) and artemether-lumefantrine (AL) was conducted in Grande Comore island from June 2019 to January 2020. A total of 238 uncomplicated falciparum malaria cases were enrolled and divided 1:1 into two treatments. The primary endpoint was the 42-day adequate clinical and parasitological responses (ACPR). Secondary endpoints were parasitaemia and fever clearance at day 3, gametocytes and tolerability.
RESULTS: The 42-day ACPR before and after PCR correction were 91.43% (95% CI 83.93-95.76%) and 98.06% (95% CI 92.48-99.66%) for AP treatment, respectively, and 96.00% (95% CI 88.17-98.14%) and 98.97% (95% CI 93.58-99.95%) for AL treatment, respectively. Complete clearance of the parasitaemia and fever for both groups was detected on day 3. Gametocytes disappeared on day 21 in the AP group and on day 2 in AL group. Specifically, the adverse reactions were mild in both groups.
CONCLUSIONS: It was found that AP and AL maintained their high efficacy and tolerance in the Comoros. Nonetheless, asymptomatic malaria infections bring new challenges to malaria control.

BACKGROUND: Artemisinin-based combination therapy (ACT), such as artemisinin-piperaquine (AP), dihydroartemisinin-piperaquine (DP), and artemether-lumefantrine (AL), is the first-line treatment for malaria in many malaria-endemic areas. However, we lack a detailed evaluation of the cardiotoxicity of these ACTs. This study aimed to analyze the electrocardiographic effects of these three ACTs in malaria patients.
METHODS: We analyzed the clinical data of 89 hospitalized patients with falciparum malaria who had received oral doses of three different ACTs. According to the ACTs administered, these patients were divided into three treatment groups: 27 treated with AP (Artequick), 31 with DP (Artekin), and 31 with AL (Coartem). Electrocardiograms and other indicators were recorded before and after the treatment. The QT interval was calculated using Fridericia\'s formula (QTcF) and Bazett\'s formula (QTcB).
RESULTS: Both QTcF and QTcB interval prolongation occurred in all three groups. The incidence of such prolongation between the three groups was not significantly different. The incidence of both moderate and severe prolongation was not significantly different between the three groups. The ΔQTcF and ΔQTcB of the three groups were not significantly different. The intra-group comparison showed significant prolongation of QTcF after AL treatment.
CONCLUSIONS: Clinically recommended doses of DP, AL, and AP may cause QT prolongation in some malaria patients but do not cause torsades de pointes ventricular tachycardia or other arrhythmias.

BACKGROUND: Currently, artemisinin-based combination therapy (ACT) is the first-line anti-malarial treatment in malaria-endemic areas. However, resistance in Plasmodium falciparum to artemisinin-based combinations emerging in the Greater Mekong Sub-region is a major problem hindering malaria elimination. To continuously monitor the potential spread of ACT-resistant parasites, this study assessed the efficacy of artemether-lumefantrine (AL) for falciparum malaria in western Myanmar.
METHODS: Ninety-five patients with malaria symptoms from Paletwa Township, Chin State, Myanmar were screened for P. falciparum infections in 2015. After excluding six patients with a parasite density below 100 or over 150,000/µL, 41 P. falciparum patients were treated with AL and followed for 28 days. Molecular markers associated with resistance to 4-amino-quinoline drugs (pfcrt and pfmdr1), antifolate drugs (pfdhps and pfdhfr) and artemisinin (pfk13) were genotyped to determine the prevalence of mutations associated with anti-malarial drug resistance.
RESULTS: For the 41 P. falciparum patients (27 children and 14 adults), the 28-day AL therapeutic efficacy was 100%, but five cases (12.2%) were parasite positive on day 3 by microscopy. For the pfk13 gene, the frequency of NN insert after the position 136 was 100% in the day-3 parasite-positive group as compared to 50.0% in the day-3 parasite-negative group, albeit the difference was not statistically significant (P = 0.113). The pfk13 K189T mutation (10.0%) was found in Myanmar for the first time. The pfcrt K76T and A220S mutations were all fixed in the parasite population. In pfmdr1, the Y184F mutation was present in 23.3% of the parasite population, and found in both day-3 parasite-positive and -negative parasites. The G968A mutation of pfmdr1 gene was first reported in Myanmar. Prevalence of all the mutations in pfdhfr and pfdhps genes assessed was over 70%, with the exception of the pfdhps A581G mutation, which was 3.3%.
CONCLUSIONS: AL remained highly efficacious in western Myanmar. Pfk13 mutations associated with artemisinin resistance were not found. The high prevalence of mutations in pfcrt, pfdhfr and pfdhps suggests high-degree resistance to chloroquine and antifolate drugs. The pfmdr1 N86/184F/D1246 haplotype associated with selection by AL in Africa reached > 20% in this study. The detection of > 10% patients who were day-3 parasite-positive after AL treatment emphasizes the necessity of continuously monitoring ACT efficacy in western Myanmar.

The artemether-lumefantrine combination requires food intake for the optimal absorption of lumefantrine. In an attempt to enhance the bioavailability of lumefantrine, new solid dispersion formulations (SDF) were developed, and the pharmacokinetics of two SDF variants were assessed in a randomized, open-label, sequential two-part study in healthy volunteers. In part 1, the relative bioavailability of the two SDF variants was compared with that of the conventional formulation after administration of a single dose of 480 mg under fasted conditions in three parallel cohorts. In part 2, the pharmacokinetics of lumefantrine from both SDF variants were evaluated after a single dose of 480 mg under fed conditions and a single dose of 960 mg under fasted conditions. The bioavailability of lumefantrine from SDF variant 1 and variant 2 increased up to ∼48-fold and ∼24-fold, respectively, relative to that of the conventional formulation. Both variants demonstrated a positive food effect and a less than proportional increase in exposure between the 480-mg and 960-mg doses. Most adverse events (AEs) were mild to moderate in severity and not suspected to be related to the study drug. All five drug-related AEs occurred in subjects taking SDF variant 2. No clinically significant treatment-emergent changes in vital signs, electrocardiograms, or laboratory blood assessments were noted. The solid dispersion formulation enhances the lumefantrine bioavailability to a significant extent, and SDF variant 1 is superior to SDF variant 2.

From 2001 to 2003, anti-malarial combination coartem and fansimef, recommended by WHO, were used to treat falciparum malaria in Mali, 28 cases in each group. The mean fever clearance time, mean asexual parasite clearance time and the cure rate in 28 days were 35.3 +/- 6. 4, 34.7 +/- 6.9 hours and 100% respectively in coartem group, and 32. 6 +/- 5.8, 36.8 +/- 5.3 hours and 96.4% respectively in the fansimef group, with no significant difference between the two groups (P > 0.05).

One hundred and two Chinese out-patients with naturally acquired, previously untreated, falciparum malaria were selected to evaluate the efficacy of a new combination anti-malaria therapy, CGP 56697 (artemether plus benflumetol). In this open non-comparative trial each patient received a combination of 80 mg artemether and 480 mg benflumetol given orally at 0, 8, 24 and 48 hours (total: 320 mg artemether, 1,920 mg benflumetol). Patients were kept for 28 days in a transmission-free hospital in an area with chloroquine resistant falciparum malaria to prevent reinfection and to aid diagnosis of recrudescence. Progress and possible adverse effects were monitored by blood film parasitology, blood biochemistry assays, urinalysis, ECG and X-ray. Ninety-eight of the 102 patients were shown to be free of infection at 28 days, a 96.1% cure rate. Parasite reduction at 24 hours was 99.4%. Time to effect complete parasite clearance ranged from 24 to 54 hours (median 30 hours). Time for fever clearance ranged from 6 to 78 hours (median 18 hours). Recrudescence was low (3.9%). No significant adverse side-effects were encountered. It is concluded that CGP 56697, a combination anti-malaria therapy of artemether with benflumetol, offered a rapid and highly effective treatment for acute uncomplicated falciparum malaria in an area of chloroquine-resistant malaria in China.