RESUMEN
Polymorphisms in the Plasmodium falciparum multidrug resistance protein 1 (pfmdr1) gene and the Plasmodium falciparum chloroquine resistance transporter (pfcrt) gene alter the malaria parasite's susceptibility to most of the current antimalarial drugs. However, the precise mechanisms by which PfMDR1 contributes to multidrug resistance have not yet been fully elucidated, nor is it understood why polymorphisms in pfmdr1 and pfcrt that cause chloroquine resistance simultaneously increase the parasite's susceptibility to lumefantrine and mefloquine-a phenomenon known as collateral drug sensitivity. Here, we present a robust expression system for PfMDR1 in Xenopus oocytes that enables direct and high-resolution biochemical characterizations of the protein. We show that wild-type PfMDR1 transports diverse pharmacons, including lumefantrine, mefloquine, dihydroartemisinin, piperaquine, amodiaquine, methylene blue, and chloroquine (but not the antiviral drug amantadine). Field-derived mutant isoforms of PfMDR1 differ from the wild-type protein, and each other, in their capacities to transport these drugs, indicating that PfMDR1-induced changes in the distribution of drugs between the parasite's digestive vacuole (DV) and the cytosol are a key driver of both antimalarial resistance and the variability between multidrug resistance phenotypes. Of note, the PfMDR1 isoforms prevalent in chloroquine-resistant isolates exhibit reduced capacities for chloroquine, lumefantrine, and mefloquine transport. We observe the opposite relationship between chloroquine resistance-conferring mutations in PfCRT and drug transport activity. Using our established assays for characterizing PfCRT in the Xenopus oocyte system and in live parasite assays, we demonstrate that these PfCRT isoforms transport all 3 drugs, whereas wild-type PfCRT does not. We present a mechanistic model for collateral drug sensitivity in which mutant isoforms of PfMDR1 and PfCRT cause chloroquine, lumefantrine, and mefloquine to remain in the cytosol instead of sequestering within the DV. This change in drug distribution increases the access of lumefantrine and mefloquine to their primary targets (thought to be located outside of the DV), while simultaneously decreasing chloroquine's access to its target within the DV. The mechanistic insights presented here provide a basis for developing approaches that extend the useful life span of antimalarials by exploiting the opposing selection forces they exert upon PfCRT and PfMDR1.
Asunto(s)
Antimaláricos , Malaria Falciparum , Parásitos , Animales , Antimaláricos/farmacología , Antimaláricos/uso terapéutico , Cloroquina/metabolismo , Cloroquina/farmacología , Cloroquina/uso terapéutico , Resistencia a Medicamentos/genética , Resistencia a Múltiples Medicamentos , Lumefantrina/farmacología , Lumefantrina/uso terapéutico , Malaria Falciparum/parasitología , Mefloquina/metabolismo , Mefloquina/farmacología , Mefloquina/uso terapéutico , Proteínas Asociadas a Resistencia a Múltiples Medicamentos/genética , Proteínas Asociadas a Resistencia a Múltiples Medicamentos/metabolismo , Proteínas Asociadas a Resistencia a Múltiples Medicamentos/uso terapéutico , Parásitos/metabolismo , Plasmodium falciparum/genética , Plasmodium falciparum/metabolismo , Isoformas de Proteínas/metabolismo , Proteínas Protozoarias/genética , Proteínas Protozoarias/metabolismoRESUMEN
BACKGROUND: Recent cases of clinical failure in malaria patients in the United Kingdom (UK) treated with artemether-lumefantrine have implications for malaria chemotherapy worldwide. METHODS: Parasites were isolated from an index case of confirmed Plasmodium falciparum treatment failure after standard treatment, and from comparable travel-acquired UK malaria cases. Drug susceptibility in vitro and genotypes at 6 resistance-associated loci were determined for all parasite isolates and compared with clinical outcomes for each parasite donor. RESULTS: A traveler, who returned to the UK from Uganda in 2022 with Plasmodium falciparum malaria, twice failed treatment with full courses of artemether-lumefantrine. Parasites from the patient exhibited significantly reduced susceptibility to artemisinin (ring-stage survival, 17.3% [95% confidence interval {CI}, 13.6%-21.1%]; P < .0001) and lumefantrine (effective concentration preventing 50% of growth = 259.4â nM [95% CI, 130.6-388.2â nM]; P = .001). Parasite genotyping identified an allele of pfk13 encoding both the A675V variant in the Pfk13 propeller domain and a novel L145V nonpropeller variant. In vitro susceptibility testing of 6 other P. falciparum lines of Ugandan origin identified reduced susceptibility to artemisinin and lumefantrine in 1 additional line, also from a 2022 treatment failure case. These parasites did not harbor a pfk13 propeller domain variant but rather the novel nonpropeller variant T349I. Variant alleles of pfubp1, pfap2mu, and pfcoronin were also identified among the 7 parasite lines. CONCLUSIONS: We confirm, in a documented case of artemether-lumefantrine treatment failure imported from Uganda, the presence of pfk13 mutations encoding L145V and A675V. Parasites with reduced susceptibility to both artemisinin and lumefantrine may be emerging in Uganda.
Asunto(s)
Antimaláricos , Artemisininas , Malaria Falciparum , Malaria , Humanos , Lumefantrina/farmacología , Lumefantrina/uso terapéutico , Plasmodium falciparum , Antimaláricos/farmacología , Antimaláricos/uso terapéutico , Combinación Arteméter y Lumefantrina/farmacología , Combinación Arteméter y Lumefantrina/uso terapéutico , Uganda , Resistencia a Medicamentos , Arteméter/farmacología , Arteméter/uso terapéutico , Artemisininas/farmacología , Artemisininas/uso terapéutico , Malaria/tratamiento farmacológico , Malaria Falciparum/tratamiento farmacológico , Malaria Falciparum/parasitología , Insuficiencia del Tratamiento , Reino Unido , Proteínas Protozoarias/genéticaRESUMEN
Capillary samples offer practical benefits compared with venous samples for the measurement of drug concentrations, but the relationship between the two measures varies between different drugs. We measured the concentrations of lumefantrine, mefloquine, piperaquine in 270 pairs of venous plasma and concurrent capillary plasma samples collected from 270 pregnant women with uncomplicated falciparum or vivax malaria. The median and range of venous plasma concentrations included in this study were 447.5 ng/mL (8.81-3,370) for lumefantrine (day 7, n = 76, median total dose received 96.0 mg/kg), 17.9 ng/mL (1.72-181) for desbutyl-lumefantrine, 1,885 ng/mL (762-4,830) for mefloquine (days 3-21, n = 90, median total dose 24.9 mg/kg), 641 ng/mL (79.9-1,950) for carboxy-mefloquine, and 51.8 ng/mL (3.57-851) for piperaquine (days 3-21, n = 89, median total dose 52.2 mg/kg). Although venous and capillary plasma concentrations showed a high correlation (Pearson's correlation coefficient: 0.90-0.99) for all antimalarials and their primary metabolites, they were not directly interchangeable. Using the concurrent capillary plasma concentrations and other variables, the proportions of venous plasma samples predicted within a ±10% precision range was 34% (26/76) for lumefantrine, 36% (32/89) for desbutyl-lumefantrine, 74% (67/90) for mefloquine, 82% (74/90) for carboxy-mefloquine, and 24% (21/89) for piperaquine. Venous plasma concentrations of mefloquine, but not lumefantrine and piperaquine, could be predicted by capillary plasma samples with an acceptable level of agreement. Capillary plasma samples can be utilized for pharmacokinetic and clinical studies, but caution surrounding cut-off values is required at the individual level.CLINICAL TRIALSThis study is registered with ClinicalTrials.gov as NCT01054248.
Asunto(s)
Antimaláricos , Lumefantrina , Malaria Falciparum , Malaria Vivax , Mefloquina , Piperazinas , Quinolinas , Humanos , Femenino , Mefloquina/sangre , Mefloquina/uso terapéutico , Mefloquina/farmacocinética , Antimaláricos/sangre , Antimaláricos/uso terapéutico , Antimaláricos/farmacocinética , Embarazo , Quinolinas/sangre , Quinolinas/farmacocinética , Quinolinas/uso terapéutico , Lumefantrina/uso terapéutico , Lumefantrina/sangre , Malaria Falciparum/tratamiento farmacológico , Malaria Falciparum/sangre , Adulto , Malaria Vivax/tratamiento farmacológico , Malaria Vivax/sangre , Adulto Joven , Etanolaminas/sangre , Etanolaminas/farmacocinética , Etanolaminas/uso terapéutico , Fluorenos/sangre , Fluorenos/uso terapéutico , Fluorenos/farmacocinética , AdolescenteRESUMEN
Malaria remains a leading cause of morbidity and mortality in Burkina Faso, which utilizes artemether-lumefantrine as the principal therapy to treat uncomplicated malaria and seasonal malaria chemoprevention with monthly sulfadoxine-pyrimethamine plus amodiaquine in children during the transmission season. Monitoring the activities of available antimalarial drugs is a high priority. We assessed the ex vivo susceptibility of Plasmodium falciparum to 11 drugs in isolates from patients presenting with uncomplicated malaria in Bobo-Dioulasso in 2021 and 2022. IC50 values were derived using a standard 72 h growth inhibition assay. Parasite DNA was sequenced to characterize known drug resistance-mediating polymorphisms. Isolates were generally susceptible, with IC50 values in the low-nM range, to chloroquine (median IC5010 nM, IQR 7.9-24), monodesethylamodiaquine (22, 14-46) piperaquine (6.1, 3.6-9.2), pyronaridine (3.0, 1.3-5.5), quinine (50, 30-75), mefloquine (7.1, 3.7-10), lumefantrine (7.1, 4.5-12), dihydroartemisinin (3.7, 2.2-5.5), and atovaquone (0.2, 0.1-0.3) and mostly resistant to cycloguanil (850, 543-1,290) and pyrimethamine (33,200, 18,400-54,200), although a small number of outliers were seen. Considering genetic markers of resistance to aminoquinolines, most samples had wild-type PfCRT K76T (87%) and PfMDR1 N86Y (95%) sequences. For markers of resistance to antifolates, established PfDHFR and PfDHPS mutations were highly prevalent, the PfDHPS A613S mutation was seen in 19% of samples, and key markers of high-level resistance (PfDHFR I164L; PfDHPS K540E) were absent or rare (A581G). Mutations in the PfK13 propeller domain known to mediate artemisinin partial resistance were not detected. Overall, our results suggest excellent susceptibilities to drugs now used to treat malaria and moderate, but stable, resistance to antifolates used to prevent malaria.
Asunto(s)
Antimaláricos , Antagonistas del Ácido Fólico , Malaria Falciparum , Malaria , Niño , Humanos , Antimaláricos/farmacología , Antimaláricos/uso terapéutico , Plasmodium falciparum , Malaria Falciparum/tratamiento farmacológico , Malaria Falciparum/parasitología , Combinación Arteméter y Lumefantrina/uso terapéutico , Antagonistas del Ácido Fólico/farmacología , Burkina Faso , Arteméter/uso terapéutico , Pirimetamina/farmacología , Pirimetamina/uso terapéutico , Malaria/tratamiento farmacológico , Lumefantrina/farmacología , Lumefantrina/uso terapéutico , Combinación de Medicamentos , Polimorfismo Genético/genética , Resistencia a Medicamentos/genética , Proteínas Protozoarias/genética , Proteínas Protozoarias/uso terapéuticoRESUMEN
In 2023, we updated data collected since 2010 on Plasmodium falciparum K13 and MDR1 drug resistance markers in Huye district, southern Rwanda. Artemisinin resistance-associated PfK13 markers occurred in 17.5% of 212 malaria patients (561H, 9.0%; 675V, 5.7%; and 469F, 2.8%), nearly double the frequency from 2019. PfMDR1 N86, linked with lumefantrine tolerance, was close to fixation at 98%. In southern Rwanda, markers signaling resistance to artemisinin and lumefantrine are increasing, albeit at a relatively slow rate.
Asunto(s)
Antimaláricos , Artemisininas , Malaria Falciparum , Humanos , Plasmodium falciparum/genética , Antimaláricos/farmacología , Antimaláricos/uso terapéutico , Malaria Falciparum/tratamiento farmacológico , Malaria Falciparum/epidemiología , Rwanda/epidemiología , Prevalencia , Artemisininas/farmacología , Artemisininas/uso terapéutico , Lumefantrina/uso terapéutico , Resistencia a Medicamentos/genética , Proteínas Protozoarias/genética , Proteínas Protozoarias/uso terapéuticoRESUMEN
While the Plasmodium falciparum malaria parasite continues to cause severe disease globally, Mozambique is disproportionally represented in malaria case totals. Acquisition of copy number variations (CNVs) in the parasite genome contributes to antimalarial drug resistance through overexpression of drug targets. Of interest, piperaquine resistance is associated with plasmepsin 2 and 3 CNVs (pfpmp2 and pfpmp3, respectively), while CNVs in the multidrug efflux pump, multidrug resistance-1 (pfmdr1), increase resistance to amodiaquine and lumefantrine. These antimalarials are partner drugs in artemisinin combination therapies (ACTs) and therefore, CNV detection with accurate and efficient tools is necessary to track ACT resistance risk. Here, we evaluated ~300 clinically derived samples collected from three sites in Mozambique for resistance-associated CNVs. We developed a novel, medium-throughput, quadruplex droplet digital PCR (ddPCR) assay to simultaneously quantify the copy number of pfpmp3, pfpmp2, and pfmdr1 loci in these clinical samples. By using DNA from laboratory parasite lines, we show that this nanodroplet-based method is capable of detecting picogram levels of parasite DNA, which facilitates its application for low yield and human host-contaminated clinical surveillance samples. Following ddPCR and the application of quality control standards, we detected CNVs in 13 of 229 high-quality samples (prevalence of 5.7%). Overall, our study revealed a low number of resistance CNVs present in the parasite population across all three collection sites, including various combinations of pfmdr1, pfpmp2, and pfpmp3 CNVs. The potential for future ACT resistance across Mozambique emphasizes the need for continued molecular surveillance across the region.
Asunto(s)
Antimaláricos , Variaciones en el Número de Copia de ADN , Resistencia a Medicamentos , Malaria Falciparum , Plasmodium falciparum , Proteínas Protozoarias , Antimaláricos/farmacología , Mozambique , Plasmodium falciparum/genética , Plasmodium falciparum/efectos de los fármacos , Humanos , Resistencia a Medicamentos/genética , Variaciones en el Número de Copia de ADN/genética , Malaria Falciparum/parasitología , Malaria Falciparum/tratamiento farmacológico , Proteínas Protozoarias/genética , Reacción en Cadena de la Polimerasa/métodos , Quinolinas/farmacología , Amodiaquina/farmacología , Proteínas Asociadas a Resistencia a Múltiples Medicamentos/genética , Ácido Aspártico Endopeptidasas/genética , Artemisininas/farmacología , Lumefantrina/farmacología , PiperazinasRESUMEN
BACKGROUND: Therapeutic efficacy studies (TESs) and detection of molecular markers of drug resistance are recommended by the World Health Organization (WHO) to monitor the efficacy of artemisinin-based combination therapy (ACT). This study assessed the trends of molecular markers of artemisinin resistance and/or reduced susceptibility to lumefantrine using samples collected in TES conducted in Mainland Tanzania from 2016 to 2021. METHODS: A total of 2,015 samples were collected during TES of artemether-lumefantrine at eight sentinel sites (in Kigoma, Mbeya, Morogoro, Mtwara, Mwanza, Pwani, Tabora, and Tanga regions) between 2016 and 2021. Photo-induced electron transfer polymerase chain reaction (PET-PCR) was used to confirm presence of malaria parasites before capillary sequencing, which targeted two genes: Plasmodium falciparum kelch 13 propeller domain (k13) and P. falciparum multidrug resistance 1 (pfmdr1). RESULTS: Sequencing success was ≥ 87.8%, and 1,724/1,769 (97.5%) k13 wild-type samples were detected. Thirty-seven (2.1%) samples had synonymous mutations and only eight (0.4%) had non-synonymous mutations in the k13 gene; seven of these were not validated by the WHO as molecular markers of resistance. One sample from Morogoro in 2020 had a k13 R622I mutation, which is a validated marker of artemisinin partial resistance. For pfmdr1, all except two samples carried N86 (wild-type), while mutations at Y184F increased from 33.9% in 2016 to about 60.5% in 2021, and only four samples (0.2%) had D1246Y mutations. pfmdr1 haplotypes were reported in 1,711 samples, with 985 (57.6%) NYD, 720 (42.1%) NFD, and six (0.4%) carrying minor haplotypes (three with NYY, 0.2%; YFD in two, 0.1%; and NFY in one sample, 0.1%). Between 2016 and 2021, NYD decreased from 66.1% to 45.2%, while NFD increased from 38.5% to 54.7%. CONCLUSION: This is the first report of the R622I (k13 validated mutation) in Tanzania. N86 and D1246 were nearly fixed, while increases in Y184F mutations and NFD haplotype were observed between 2016 and 2021. Despite the reports of artemisinin partial resistance in Rwanda and Uganda, this study did not report any other validated mutations in these study sites in Tanzania apart from R622I suggesting that intensified surveillance is urgently needed to monitor trends of drug resistance markers and their impact on the performance of ACT.
Asunto(s)
Antimaláricos , Artemisininas , Carubicina/análogos & derivados , Malaria Falciparum , Humanos , Lumefantrina/farmacología , Lumefantrina/uso terapéutico , Plasmodium falciparum/genética , Antimaláricos/farmacología , Antimaláricos/uso terapéutico , Tanzanía , Artemisininas/farmacología , Artemisininas/uso terapéutico , Arteméter/uso terapéutico , Proteínas Asociadas a Resistencia a Múltiples Medicamentos/genética , Combinación Arteméter y Lumefantrina/farmacología , Combinación Arteméter y Lumefantrina/uso terapéutico , Malaria Falciparum/epidemiología , Biomarcadores , Resistencia a Medicamentos/genética , Proteínas Protozoarias/genética , Proteínas Protozoarias/uso terapéuticoRESUMEN
A simple external calibration liquid chromatography-diode array detector method was developed, validated, and applied for the determination of lumefantrine (Lum) in dried blood spot (DBS) samples collected from malaria patients in Botswana. The samples were validated in accordance with the United States Food and Drug Administration guidelines for bioanalytical methods after sample preparation using solid-liquid extraction. Separation was achieved using an XTerra C18 column (50 × 4.6 mm, 5 µm), and a binary solvent system of acetonitrile and water adjusted to pH 2.3 was used as the mobile phase. The validated method was applied for the determination of Lum in DBS samples collected from malaria patients infected with Plasmodium falciparum in Botswana. The calibration curve was linear between 0.5 and 12 µg/mL with a coefficient of determination (R2 ) of 0.9996. The limit of detection and the lower limit of quantification were 0.5 and 1.4 µg/mL, respectively. The efficiency of extraction measured as percentage recovery ranged between 84.2% and 107.8% at the three quality control (QC) levels, that is, low QC, mid QC, and high QC. In conclusion, data suggest that the method is suitable for the determination of trace Lum in biofluids and can also be used for therapeutic drug monitoring and pharmacokinetic profiling.
Asunto(s)
Malaria , Humanos , Lumefantrina , Cromatografía Líquida de Alta Presión/métodos , Calibración , BotswanaRESUMEN
BACKGROUND: Artemether-lumefantrine (AL) is the most widely used artemisinin-based combination therapy in Sub-Saharan Africa and is threatened by the emergence of artemisinin resistance. Dosing is suboptimal in young children. We hypothesized that extending AL duration will improve exposure and reduce reinfection risks. METHODS: We conducted a prospective, randomized, open-label pharmacokinetic/pharmacodynamic study of extended duration AL in children with malaria in high-transmission rural Uganda. Children received 3-day (standard 6-dose) or 5-day (10-dose) AL with sampling for artemether, dihydroartemisinin, and lumefantrine over 42-day clinical follow-up. Primary outcomes were (1) comparative pharmacokinetic parameters between regimens and (2) recurrent parasitemia analyzed as intention-to-treat. RESULTS: A total of 177 children aged 16 months to 16 years were randomized, contributing 227 total episodes. Terminal median lumefantrine concentrations were significantly increased in the 5-day versus 3-day regimen on days 7, 14, and 21 (P < .001). A predefined day 7 lumefantrine threshold of 280 ng/mL was strongly predictive of recurrence risk at 28 and 42 days (P < .001). Kaplan-Meier estimated 28-day (51% vs 40%) and 42-day risk (75% vs 68%) did not significantly differ between 3- and 5-day regimens. No significant toxicity was seen with the extended regimen. CONCLUSIONS: Extending the duration of AL was safe and significantly enhanced overall drug exposure in young children but did not lead to significant reductions in recurrent parasitemia risk in our high-transmission setting. However, day 7 levels were strongly predictive of recurrent parasitemia risk, and those in the lowest weight-band were at higher risk of underdosing with the standard 3-day regimen. CLINICAL TRIAL REGISTRATION: ClinicalTrials.gov number NCT03453840.
Asunto(s)
Antimaláricos , Artemisininas , Malaria Falciparum , Malaria , Niño , Humanos , Lactante , Preescolar , Antimaláricos/efectos adversos , Combinación Arteméter y Lumefantrina/uso terapéutico , Uganda , Arteméter/uso terapéutico , Reinfección , Parasitemia/tratamiento farmacológico , Estudios Prospectivos , Malaria Falciparum/tratamiento farmacológico , Fluorenos/efectos adversos , Artemisininas/efectos adversos , Malaria/tratamiento farmacológico , Lumefantrina/uso terapéutico , Etanolaminas/efectos adversos , Combinación de MedicamentosRESUMEN
An amorphous drug-polymer salt (ADPS) can be remarkably stable against crystallization at high temperature and humidity (e.g., 40°C/75% RH) and provide fast release. Here, we report that process conditions strongly influence the degree of proton transfer (salt formation) between a drug and a polymer and in turn the product's stability and release. For lumefantrine (LMF) formulated with poly(acrylic acid) (PAA), we first show that the amorphous materials prepared by slurry conversion and antisolvent precipitation produce a single trend in which the degree of drug protonation increases with PAA concentration from 0% for pure LMF to â¼100% above 70 wt % PAA, independent of PAA's molecular weight (1.8, 450, and 4000 kg/mol). This profile describes the equilibrium for salt formation and can be modeled as a chemical equilibrium in which the basic molecules compete for the acidic groups on the polymer chain. Relative to this equilibrium, the literature methods of hot-melt extrusion (HME) and rotary evaporation (RE) reached much lower degrees of salt formation. For example, at 40 wt % drug loading, HME reached 5% salt formation and RE 15%, both well below the equilibrium value of 85%. This is noteworthy given the common use of HME and RE in manufacturing amorphous formulations, indicating a need for careful control of process conditions to ensure the full interaction between the drug and the polymer. This need arises due to the low mobility of macromolecules and the mutual hindrance of adjacent reaction sites. We find that a high degree of salt formation enhances drug stability and release. For example, at 50% drug loading, an HME-like formulation with 19% salt formation crystallized faster and released only 20% of the drug relative to a slurry-prepared formulation with 70% salt formation. Based on this work, we recommend slurry conversion as the method for preparing ADPS for its ability to enhance salt formation and continuously adjust drug loading. While this work focused on salt formation, the impact of process conditions on the molecular-level interactions between a drug and a polymer is likely a general issue for amorphous solid dispersions, with consequences on product stability and drug release.
Asunto(s)
Polímeros , Protones , Polímeros/química , Sales (Química) , Química Farmacéutica/métodos , Solubilidad , Lumefantrina , Estabilidad de Medicamentos , Composición de Medicamentos/métodosRESUMEN
BACKGROUND: The emergence of resistance to artemisinin derivatives in Southeast Asia constitutes a serious threat for other malaria endemic areas, particularly in Côte d'Ivoire. To delay this resistance, the application of the control measures recommended by the National Malaria Control Programme (NMCP) for a correct management, in the private pharmacies, is a necessity. The purpose of this study was, therefore, to assess the level of knowledge and practices of private pharmacy auxiliary in Abidjan about the management of malaria. METHODS: A descriptive cross-sectional study was conducted from April to November 2015. It included auxiliaries of private pharmacies in Abidjan. Data collection material was a structured an open pretested questionnaire. Data analysis was carried out using Package for Social Science (SPSS) software version 21.1. Chi square test was used to compare proportions for a significance threshold of 0.05 for the p value. RESULTS: A total, 447 auxiliaries from 163 private pharmacies were interviewed. It was noted that the auxiliaries had a good knowledge of clinical signs of uncomplicated malaria (99.1%), biological examinations (54.6% for the thick film and 40.7% for rapid diagnostic tests (RDTs) and anti-malarial drugs (99.3% for artemether + lumefantrine, AL). The strategies of vector control (long-lasting insecticide-treated mosquito nets (LLITNs, Repellent ointments, cleaning gutters, elimination of larvae breeding site and intermittent preventive treatment with sulfadoxine-pyrimethamine (IPTp-SP) in pregnant women were also known by the auxiliaries, respectively 99.8% and 77.4%. However, the malaria pathogen (25.1%) and the NMCP recommendations (e.g. use of AL or AS + AQ as first-line treatment for uncomplicated malaria and IPTp-SP in pregnant women) were not well known by the auxiliaries (28.2% and 26.9% for uncomplicated and severe malaria). Concerning the practices of the auxiliaries, 91.1% offered anti-malarial drugs to patients without a prescription and 47.3% mentioned incorrect dosages. The combination artemether + lumefantrine was the most recommended (91.3%). The delivery of anti-malarial drugs was rarely accompanied by advice on malaria prevention, neither was it carried out on the result of an RDT. CONCLUSION: The epidemiology and the NMCP recommendations for the diagnostic and therapeutic management of malaria, are not well known to auxiliaries, which may have implications for their practices. These results show the need to sensitize and train private pharmacy auxiliaries, and also to involve them in NMCP activities.
Asunto(s)
Antimaláricos , Malaria , Farmacias , Farmacia , Humanos , Femenino , Embarazo , Antimaláricos/uso terapéutico , Côte d'Ivoire , Estudios Transversales , Malaria/epidemiología , Combinación de Medicamentos , Encuestas y Cuestionarios , Lumefantrina/uso terapéutico , Arteméter/uso terapéuticoRESUMEN
BACKGROUND: Substandard anti-malarial agents pose a significant challenge to effective malaria control and elimination efforts especially in sub-Saharan Africa. The quality of anti-malarials in most low-and-middle income countries (LMICs) is affected by several factors including inadequate regulation and limited resources. In this study, the pharmacopeial quality of artemether-lumefantrine (AL) in low and high malaria transmission settings in Uganda was assessed. METHODS: This was a cross-sectional study conducted among randomly selected private drug outlets. The AL anti-malarials available in drug outlets were purchased using overt method. The samples were screened for quality using visual inspection, weight uniformity, content assay and dissolution tests. The assay test was done using liquid chromatography-mass spectrometry (LC-MS). The samples were considered substandard if the active pharmaceutical ingredient (API) content was outside 90-110% range of the label claim. Dissolution test was conducted following United States Pharmacopoeia (USP) method. Data was analysed using descriptive statistics and presented as means with standard deviations, frequencies, and proportions. Correlation between medicine quality and independent variables was determined using Fisher's exact test of independence at 95% level of significance. RESULTS: A total of 74 AL anti-malarial samples were purchased from high (49/74; 66.2%) and low (25/74; 33.8%) malaria transmission settings. The most common batch of AL was LONART, 32.4% (24/74), with 33.8% (25/74) being 'Green leaf'. Overall prevalence of substandard quality artemether-lumefantrine was 18.9% (14/74; 95% CI: 11.4-29.7). Substandard quality AL was significantly associated with setting (p = 0.002). A total of 10 samples (13.5%) failed artemether content assay test while, 4 samples (5.4%, 4/74) failed the lumefantrine assay test. One sample from a high malaria transmission setting failed both artemether and lumefantrine assay content test. Of the samples that failed artemether assay test, 90% had low (< 90%) artemether content. All the samples passed visual inspection and dissolution tests. CONCLUSION: Artemether-lumefantrine agents, the recommended first-line treatment for uncomplicated malaria with APIs outside the recommended pharmacopeial content assay limit is common especially in high malaria transmission settings. There is need for continuous surveillance and monitoring of the quality of artemisinin-based anti-malarials across the country by the drug regulatory agency.
Asunto(s)
Antimaláricos , Malaria Falciparum , Malaria , Humanos , Antimaláricos/análisis , Combinación Arteméter y Lumefantrina/uso terapéutico , Combinación Arteméter y Lumefantrina/análisis , Arteméter/uso terapéutico , Uganda , Estudios Transversales , Malaria/prevención & control , Lumefantrina/uso terapéutico , Malaria Falciparum/tratamiento farmacológicoRESUMEN
BACKGROUND: When people with human immunodeficiency virus (HIV) infection (PWH) develop malaria, they are at risk of poor anti-malarial treatment efficacy resulting from impairment in the immune response and/or drug-drug interactions that alter anti-malarial metabolism. The therapeutic efficacy of artemether-lumefantrine was evaluated in a cohort of PWH on antiretroviral therapy (ART) and included measurement of day 7 lumefantrine levels in a subset to evaluate for associations between lumefantrine exposure and treatment response. METHODS: Adults living with HIV (≥ 18 years), on ART for ≥ 6 months with undetectable HIV RNA viral load and CD4 count ≥ 250/mm3 were randomized to daily trimethoprim-sulfamethoxazole (TS), weekly chloroquine (CQ) or no prophylaxis. After diagnosis of uncomplicated Plasmodium falciparum malaria, a therapeutic efficacy monitoring was conducted with PCR-correction according to WHO guidelines. The plasma lumefantrine levels on day 7 in 100 episodes of uncomplicated malaria was measured. A frailty proportional hazards model with random effects models to account for clustering examined the relationship between participant characteristics and malaria treatment failure within 28 days. Pearson's Chi-squared test was used to compare lumefantrine concentrations among patients with treatment failure and adequate clinical and parasitological response (ACPR). RESULTS: 411 malaria episodes were observed among 186 participants over 5 years. The unadjusted ACPR rate was 81% (95% CI 77-86). However, after PCR correction to exclude new infections, ACPR rate was 94% (95% CI 92-97). Increasing age and living in Ndirande were associated with decreased hazard of treatment failure. In this population of adults with HIV on ART, 54% (51/94) had levels below a previously defined optimal day 7 lumefantrine level of 200 ng/ml. This occurred more commonly among participants who were receiving an efavirenz-based ART compared to other ART regimens (OR 5.09 [95% CI 1.52-7.9]). Participants who experienced treatment failure had lower day 7 median lumefantrine levels (91 ng/ml [95% CI 48-231]) than participants who experienced ACPR (190 ng/ml [95% CI 101-378], p-value < 0.008). CONCLUSION: Recurrent malaria infections are frequent in this population of PWH on ART. The PCR-adjusted efficacy of AL meets the WHO criteria for acceptable treatment efficacy. Nevertheless, lumefantrine levels tend to be low in this population, particularly in those on efavirenz-based regimens, with lower concentrations associated with more frequent malaria infections following treatment. These results highlight the importance of understanding drug-drug interactions when diseases commonly co-occur.
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Antimaláricos , Artemisininas , Infecciones por VIH , Malaria Falciparum , Malaria , Humanos , Adulto , Antimaláricos/uso terapéutico , Malaui , Artemisininas/uso terapéutico , Arteméter/uso terapéutico , Combinación de Medicamentos , Combinación Arteméter y Lumefantrina/uso terapéutico , Malaria/tratamiento farmacológico , Malaria Falciparum/tratamiento farmacológico , Malaria Falciparum/prevención & control , Lumefantrina/uso terapéutico , Infecciones por VIH/tratamiento farmacológico , Resultado del Tratamiento , Etanolaminas/uso terapéutico , Fluorenos/uso terapéuticoRESUMEN
BACKGROUND: Artemisinin-based combination therapy (ACT) is the first-line treatment for uncomplicated malaria in Ghana. Artemisinin (ART) tolerance in Plasmodium falciparum has arisen in Southeast Asia and recently, in parts of East Africa. This is ascribed to the survival of ring-stage parasites post treatment. The present study sought to assess and characterize correlates of potential ART tolerance based on post-treatment parasite clearance, ex vivo and in vitro drug sensitivity, and molecular markers of drug resistance in P. falciparum isolates from children with uncomplicated malaria in Ghana. METHODS: Six months to fourteen years old children presenting with acute uncomplicated malaria (n = 115) were enrolled in two hospitals and a Health Centre in Ghana's Greater Accra region and treated with artemether-lumefantrine (AL) according to body weight. Pre- and post-treatment parasitaemia (day 0 and day 3) was confirmed by microscopy. The ex vivo ring-stage survival assay (RSA) was used to detect percent ring survival while the 72 h SYBR Green I assay was used to measure the 50% inhibition concentration (IC50s) of ART and its derivatives and partner drugs. Genetic markers of drug tolerance /resistance were evaluated using selective whole genome sequencing. RESULTS: Of the total of 115 participants, 85 were successfully followed up on day 3 post-treatment and 2/85 (2.4%) had parasitaemia. The IC50 values of ART, artesunate (AS), artemether (AM), dihydroartemisinin (DHA), amodiaquine (AQ), and lumefantrine (LUM) were not indicative of drug tolerance. However, 7/90 (7.8%) pre-treatment isolates had > 10% ring survival rates against DHA. Of the four isolates (2 RSA positive and 2 RSA negative) with high genomic coverage, P. falciparum (Pf) kelch 13 K188* and Pfcoronin V424I mutations were only present in the two RSA positive isolates with > 10% ring survival rates. CONCLUSIONS: The observed low proportion of participants with day-3 post-treatment parasitaemia is consistent with rapid ART clearance. However, the increased rates of survival observed in the ex vivo RSA against DHA, maybe a pointer of an early start of ART tolerance. Furthermore, the role of two novel mutations in PfK13 and Pfcoronin genes, harboured by the two RSA positive isolates that had high ring survival in the present study, remains to be elucidated.
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Antimaláricos , Artemisininas , Malaria Falciparum , Malaria , Humanos , Niño , Antimaláricos/farmacología , Antimaláricos/uso terapéutico , Plasmodium falciparum/genética , Combinación Arteméter y Lumefantrina/uso terapéutico , Ghana , Combinación de Medicamentos , Arteméter/uso terapéutico , Artemisininas/farmacología , Artemisininas/uso terapéutico , Malaria/tratamiento farmacológico , Lumefantrina/uso terapéutico , Malaria Falciparum/tratamiento farmacológico , Tolerancia a MedicamentosRESUMEN
BACKGROUND: Pfcrt gene has been associated with chloroquine resistance and the pfmdr1 gene can alter malaria parasite susceptibility to lumefantrine, mefloquine, and chloroquine. In the absence of chloroquine (CQ) and extensive use of artemether-lumefantrine (AL) from 2004 to 2020 to treat uncomplicated falciparum malaria, pfcrt haplotype, and pfmdr1 single nucleotide polymorphisms (SNPs) were determined in two sites of West Ethiopia with a gradient of malaria transmission. METHODS: 230 microscopically confirmed P. falciparum isolates were collected from Assosa (high transmission area) and Gida Ayana (low transmission area) sites, of which 225 of them tested positive by PCR. High-Resolution Melting Assay (HRM) was used to determine the prevalence of pfcrt haplotypes and pfmdr1 SNPs. Furthermore, the pfmdr1 gene copy number (CNV) was determined using real-time PCR. A P-value of less or equal to 0.05 was considered significant. RESULTS: Of the 225 samples, 95.5%, 94.4%, 86.7%, 91.1%, and 94.2% were successfully genotyped with HRM for pfcrt haplotype, pfmdr1-86, pfmdr1-184, pfmdr1-1042 and pfmdr1-1246, respectively. The mutant pfcrt haplotypes were detected among 33.5% (52/155) and 80% (48/60) of isolates collected from the Assosa and Gida Ayana sites, respectively. Plasmodium falciparum with chloroquine-resistant haplotypes was more prevalent in the Gida Ayana area compared with the Assosa area (COR = 8.4, P = 0.00). Pfmdr1-N86Y wild type and 184F mutations were found in 79.8% (166/208) and 73.4% (146/199) samples, respectively. No single mutation was observed at the pfmdr1-1042 locus; however, 89.6% (190/212) of parasites in West Ethiopia carry the wild-type D1246Y variants. Eight pfmdr1 haplotypes at codons N86Y-Y184F-D1246Y were identified with the dominant NFD 61% (122/200). There was no difference in the distribution of pfmdr1 SNPs, haplotypes, and CNV between the two study sites (P > 0.05). CONCLUSION: Plasmodium falciparum with the pfcrt wild-type haplotype was prevalent in high malaria transmission site than in low transmission area. The NFD haplotype was the predominant haplotype of the N86Y-Y184F-D1246Y. A continuous investigation is needed to closely monitor the changes in the pfmdr1 SNPs, which are associated with the selection of parasite populations by ACT.
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Antimaláricos , Malaria Falciparum , Malaria , Humanos , Antimaláricos/farmacología , Antimaláricos/uso terapéutico , Etiopía/epidemiología , Combinación Arteméter y Lumefantrina/uso terapéutico , Arteméter/uso terapéutico , Malaria Falciparum/parasitología , Cloroquina/farmacología , Cloroquina/uso terapéutico , Malaria/tratamiento farmacológico , Lumefantrina/uso terapéutico , Plasmodium falciparum , Polimorfismo de Nucleótido Simple , Proteínas Asociadas a Resistencia a Múltiples Medicamentos/genética , Proteínas Protozoarias/genética , Proteínas Protozoarias/uso terapéutico , Resistencia a Medicamentos/genéticaRESUMEN
BACKGROUND: Malaria remains a public health concern globally. Resistance to anti-malarial drugs has consistently threatened the gains in controlling the malaria parasites. Currently, artemether-lumefantrine (AL) and dihydroartemisinin-piperaquine (DP) are the treatment regimens against Plasmodium falciparum infections in many African countries, including Kenya. Recurrent infections have been reported in patients treated with AL or DP, suggesting the possibility of reinfection or parasite recrudescence associated with the development of resistance against the two therapies. The Plasmodium falciparum cysteine desulfurase IscS (Pfnfs1) K65 selection marker has previously been associated with decreased lumefantrine susceptibility. This study evaluated the frequency of the Pfnfs1 K65 resistance marker and associated K65Q resistant allele in recurrent infections collected from P. falciparum-infected individuals living in Matayos, Busia County, in western Kenya. METHODS: Archived dried blood spots (DBS) of patients with recurrent malaria infection on clinical follow-up days after treatment with either AL or DP were used in the study. After extraction of genomic DNA, PCR amplification and sequencing analysis were employed to determine the frequencies of the Pfnfs1 K65 resistance marker and K65Q mutant allele in the recurrent infections. Plasmodium falciparum msp1 and P. falciparum msp2 genetic markers were used to distinguish recrudescent infections from new infections. RESULTS: The K65 wild-type allele was detected at a frequency of 41% while the K65Q mutant allele was detected at a frequency of 22% in the recurrent samples. 58% of the samples containing the K65 wild-type allele were AL treated samples and while 42% were DP treated samples. 79% of the samples with the K65Q mutation were AL treated samples and 21% were DP treated samples. The K65 wild-type allele was detected in three recrudescent infections (100%) identified from the AL treated samples. The K65 wild-type allele was detected in two recrudescent DP treated samples (67%) while the K65Q mutant allele was identified in one DP treated (33%) recrudescent sample. CONCLUSIONS: The data demonstrate a higher frequency of the K65 resistance marker in patients with recurrent infection during the study period. The study underscores the need for consistent monitoring of molecular markers of resistance in regions of high malaria transmission.
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Antimaláricos , Malaria Falciparum , Malaria , Quinolinas , Humanos , Combinación Arteméter y Lumefantrina/uso terapéutico , Antimaláricos/uso terapéutico , Plasmodium falciparum/genética , Kenia/epidemiología , Reinfección/inducido químicamente , Reinfección/tratamiento farmacológico , Prevalencia , Combinación de Medicamentos , Arteméter/uso terapéutico , Malaria Falciparum/tratamiento farmacológico , Malaria Falciparum/epidemiología , Malaria Falciparum/parasitología , Quinolinas/uso terapéutico , Lumefantrina/uso terapéutico , Malaria/tratamiento farmacológico , MutaciónRESUMEN
Glioblastoma multiforme (GBM) is an aggressive cancer without currently effective therapies. Radiation and temozolomide (radio/TMZ) resistance are major contributors to cancer recurrence and failed GBM therapy. Heat shock proteins (HSPs), through regulation of extracellular matrix (ECM) remodeling and epithelial mesenchymal transition (EMT), provide mechanistic pathways contributing to the development of GBM and radio/TMZ-resistant GBM. The Friend leukemia integration 1 (Fli-1) signaling network has been implicated in oncogenesis in GBM, making it an appealing target for advancing novel therapeutics. Fli-1 is linked to oncogenic transformation with up-regulation in radio/TMZ-resistant GBM, transcriptionally regulating HSPB1. This link led us to search for targeted molecules that inhibit Fli-1. Expression screening for Fli-1 inhibitors identified lumefantrine, an antimalarial drug, as a probable Fli-1 inhibitor. Docking and isothermal calorimetry titration confirmed interaction between lumefantrine and Fli-1. Lumefantrine promoted growth suppression and apoptosis in vitro in parental and radio/TMZ-resistant GBM and inhibited tumor growth without toxicity in vivo in U87MG GBM and radio/TMZ-resistant GBM orthotopic tumor models. These data reveal that lumefantrine, an FDA-approved drug, represents a potential GBM therapeutic that functions through inhibition of the Fli-1/HSPB1/EMT/ECM remodeling protein networks.
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Antimaláricos/administración & dosificación , Antineoplásicos Alquilantes/administración & dosificación , Neoplasias Encefálicas/tratamiento farmacológico , Glioblastoma/tratamiento farmacológico , Lumefantrina/administración & dosificación , Temozolomida/administración & dosificación , Neoplasias Encefálicas/radioterapia , Línea Celular Tumoral , Resistencia a Antineoplásicos , Transición Epitelial-Mesenquimal/efectos de los fármacos , Regulación Neoplásica de la Expresión Génica/efectos de los fármacos , Glioblastoma/genética , Glioblastoma/metabolismo , Glioblastoma/radioterapia , Proteínas de Choque Térmico/genética , Proteínas de Choque Térmico/metabolismo , Humanos , Proteínas de Microfilamentos/genética , Proteínas de Microfilamentos/metabolismo , Chaperonas Moleculares/genética , Chaperonas Moleculares/metabolismo , Transactivadores/genética , Transactivadores/metabolismoRESUMEN
The global spread of multi-drug resistant P. falciparum, P. vivax, and P. malariae strains and absence of long-term effective vaccine makes chemotherapy the mainstay of malaria control strategies in endemic settings. The Mossman's assay and the Organization for Economic Co-operation and Development (OECD), 2001 guideline 423, were used to determine the cytotoxicity and acute oral toxicity of a novel hybrid drug, artesunate-3-Chloro-4(4-chlorophenoxy) aniline (ATSA), in vitro and in vivo, respectively. A modified Desjardins method was used to screen for antiplasmodial activity using P. falciparum (3D7 and W2) strains in vitro. The Peter's 4-day suppressive tests (4DTs) was used to evaluate the in vivo antimalaria activity using P. berghei ANKA strain, lumefantrine resistant (LuR), and piperaquine resistant (PQR) P. berghei lines. In silico prediction of absorption, distribution, metabolism, excretion, and toxicity (ADMET) profiles was assayed using PreADMET online prediction tool. The reference drug in all experiments was artesunate (ATS). Statistical significance between ATSA's activities in treated and control mice was evaluated by one-way analysis of variance (ANOVA). Results show that inhibitory concentrations-50 (IC50) of ATSA is 11.47 ± 1.3 (3D7) and 1.45 ± 0.26 (W2) against 4.66 ± 0.93 (3D7) and 0.60 ± 0.15 (W2) ng/ml of ATS with a selective index of 2180.91(3D7) and a therapeutic index (TI) of > 71). No mortalities were observed in acute oral toxicity assays and mean weight differences for test and controls were statistically insignificant (P > 0.05). The in vivo activity of ATSA was above 40% with effective dosage-50 (ED50) of 4.211, 2.601, and 3.875 mg/kg body weight against P. berghei ANKA, LuR, and PQR lines, respectively. The difference between treated and control mice was statistically significant (P < 0.05). ATSA has high intestinal absorption (HIA) > 95% and has medium human ether-a-go-go related gene (hERG) K+ channel inhibition risks. Preclinical and clinical studies on ATSA are recommended to evaluate its value in developing novel drugs for future management of multi-drug resistant malaria parasites.
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Antimaláricos , Malaria Falciparum , Malaria Vivax , Malaria , Humanos , Animales , Ratones , Antimaláricos/farmacología , Artesunato/uso terapéutico , Plasmodium falciparum , Malaria/parasitología , Malaria Falciparum/parasitología , Lumefantrina/farmacología , Lumefantrina/uso terapéutico , Compuestos de Anilina/farmacología , Compuestos de Anilina/uso terapéutico , Plasmodium bergheiRESUMEN
Toxoplasma gondii is an obligate protozoon that can infect all warm-blooded animals including humans. T. gondii afflicts one-third of the human population and is a detriment to the health of livestock and wildlife. Thus far, traditional drugs such as pyrimethamine and sulfadiazine used to treat T. gondii infection are inadequate as therapeutics due to relapse, long treatment period, and low efficacy in parasite clearance. Novel, efficacious drugs have not been available. Lumefantrine, as an antimalarial, is effective in killing T. gondii but has no known mechanism of action. We combined metabolomics with transcriptomics to investigate how lumefantrine inhibits T. gondii growth. We identified significant alternations in transcripts and metabolites and their associated functional pathways that are attributed to lumefantrine treatment. RH tachyzoites were used to infect Vero cells for three hours and subsequently treated with 900 ng/mL lumefantrine. Twenty-four hours post-drug treatment, we observed significant changes in transcripts associated with five DNA replication and repair pathways. Metabolomic data acquired through liquid chromatography-tandem mass spectrometry (LC-MS) showed that lumefantrine mainly affected sugar and amino acid metabolism, especially galactose and arginine. To investigate whether lumefantrine damages T. gondii DNA, we conducted a terminal transferase assay (TUNEL). TUNEL results showed that lumefantrine significantly induced apoptosis in a dose-dependent manner. Taken together, lumefantrine effectively inhibited T. gondii growth by damaging DNA, interfering with DNA replication and repair, and altering energy and amino acid metabolisms.
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Toxoplasma , Animales , Chlorocebus aethiops , Humanos , Toxoplasma/metabolismo , Células Vero , Transcriptoma , Lumefantrina/farmacología , Aminoácidos/metabolismoRESUMEN
Despite repeated malaria infection, individuals living in areas where malaria is endemic remain vulnerable to reinfection. The Janus kinase (JAK1/2) inhibitor ruxolitinib could potentially disrupt the parasite-induced dysfunctional immune response when administered with antimalarial therapy. This randomized, single-blind, placebo-controlled, single-center phase 1 trial investigated the safety, tolerability, and pharmacokinetic and pharmacodynamic profile of ruxolitinib and the approved antimalarial artemether-lumefantrine in combination. Ruxolitinib pharmacodynamics were assessed by inhibition of phosphorylation of signal transducer and activator of transcription 3 (pSTAT3). Eight healthy male and female participants ages 18 to 55 years were randomized to either ruxolitinib (20 mg) (n = 6) or placebo (n = 2) administered 2 h after artemether-lumefantrine (80/480 mg) twice daily for 3 days. Mild adverse events occurred in six participants (four ruxolitinib; two placebo). The combination of artemether-lumefantrine and ruxolitinib was well tolerated, with adverse events and pharmacokinetics consistent with the known profiles of both drugs. The incidence of adverse events and artemether, dihydroartemisinin (the major active metabolite of artemether), and lumefantrine exposure were not affected by ruxolitinib coadministration. Ruxolitinib coadministration resulted in a 3-fold-greater pSTAT3 inhibition compared to placebo (geometric mean ratio = 3.01 [90% confidence interval = 2.14 to 4.24]), with a direct and predictable relationship between ruxolitinib plasma concentrations and %pSTAT3 inhibition. This study supports the investigation of the combination of artemether-lumefantrine and ruxolitinib in healthy volunteers infected with Plasmodium falciparum malaria. (This study has been registered at ClinicalTrials.gov under registration no. NCT04456634.).