Persistent and multiclonal malaria parasite dynamics despite extended artemether-lumefantrine treatment in children.

Standard diagnostics used in longitudinal antimalarial studies are unable to characterize the complexity of submicroscopic parasite dynamics, particularly in high transmission settings. We use molecular markers and amplicon sequencing to characterize post-treatment stage-specific malaria parasite dynamics during a 42 day randomized trial of 3- versus 5 day artemether-lumefantrine in 303 children with and without HIV (ClinicalTrials.gov number NCT03453840). The prevalence of parasite-derived 18S rRNA is >70% in children throughout follow-up, and the ring-stage marker SBP1 is detectable in over 15% of children on day 14 despite effective treatment. We find that the extended regimen significantly lowers the risk of recurrent ring-stage parasitemia compared to the standard 3 day regimen, and that higher day 7 lumefantrine concentrations decrease the probability of ring-stage parasites in the early post-treatment period. Longitudinal amplicon sequencing reveals remarkably dynamic patterns of multiclonal infections that include new and persistent clones in both the early post-treatment and later time periods. Our data indicate that post-treatment parasite dynamics are highly complex despite efficacious therapy, findings that will inform strategies to optimize regimens in the face of emerging partial artemisinin resistance in Africa.

Mixed-method evaluation study of a targeted mass drug administration of long-acting anti-malarials among children aged 3 months to 15 years in the Bossangoa sub-prefecture, Ouham, Central African Republic, during the COVID-19 pandemic.

BACKGROUND: In 2020, during the COVID-19 pandemic, Médecins Sans Frontières (MSF) initiated three cycles of dihydroartemisin-piperaquine (DHA-PQ) mass drug administration (MDA) for children aged three months to 15 years within Bossangoa sub-prefecture, Central African Republic. Coverage, clinical impact, and community members perspectives were evaluated to inform the use of MDAs in humanitarian emergencies. METHODS: A household survey was undertaken after the MDA focusing on participation, recent illness among eligible children, and household satisfaction. Using routine surveillance data, the reduction during the MDA period compared to the same period of preceding two years in consultations, malaria diagnoses, malaria rapid diagnostic test (RDT) positivity in three MSF community healthcare facilities (HFs), and the reduction in severe malaria admissions at the regional hospital were estimated. Twenty-seven focus groups discussions (FGDs) with community members were conducted. RESULTS: Overall coverage based on the MDA card or verbal report was 94.3% (95% confidence interval (CI): 86.3-97.8%). Among participants of the household survey, 2.6% (95% CI 1.6-40.3%) of round 3 MDA participants experienced illness in the preceding four weeks compared to 30.6% (95% CI 22.1-40.8%) of MDA non-participants. One community HF experienced a 54.5% (95% CI 50.8-57.9) reduction in consultations, a 73.7% (95% CI 70.5-76.5) reduction in malaria diagnoses, and 42.9% (95% CI 36.0-49.0) reduction in the proportion of positive RDTs among children under five. A second community HF experienced an increase in consultations (+ 15.1% (- 23.3 to 7.5)) and stable malaria diagnoses (4.2% (3.9-11.6)). A third community HF experienced an increase in consultations (+ 41.1% (95% CI 51.2-31.8) and malaria diagnoses (+ 37.3% (95% CI 47.4-27.9)). There were a 25.2% (95% CI 2.0-42.8) reduction in hospital admissions with severe malaria among children under five from the MDA area. FGDs revealed community members perceived less illness among children because of the MDA, as well as fewer hospitalizations. Other indirect benefits such as reduced household expenditure on healthcare were also described. CONCLUSION: The MDA achieved high coverage and community acceptance. While some positive health impact was observed, it was resource intensive, particularly in this rural context. The priority for malaria control in humanitarian contexts should remain diagnosis and treatment. MDA may be additional tool where the context supports its implementation.

Safety and efficacy of dihydroartemisinin-piperaquine for intermittent preventive treatment of malaria in pregnant women with HIV from Gabon and Mozambique: a randomised, double-blind, placebo-controlled trial.

BACKGROUND: The cornerstone of malaria prevention in pregnancy, intermittent preventive treatment (IPTp) with sulfadoxine-pyrimethamine, is contraindicated in women with HIV who are receiving co-trimoxazole prophylaxis. We assessed whether IPTp with dihydroartemisinin-piperaquine is safe and effective in reducing the risk of malaria infection in women with HIV receiving co-trimoxazole prophylaxis and antiretroviral drugs. METHODS: For this randomised, double-blind, placebo-controlled clinical trial, women with HIV attending the first antenatal care clinic visit, resident in the study area, and with a gestational age up to 28 weeks were enrolled at five sites in Gabon and Mozambique. Participants were randomly assigned (1:1) to receive either IPTp with dihydroartemisinin-piperaquine at each scheduled antenatal care visit plus daily co-trimoxazole (intervention group) or placebo at each scheduled antenatal care visit plus daily co-trimoxazole (control group). Randomisation was done centrally via block randomisation (block sizes of eight), stratified by country. IPTp was given over 3 days under direct observation by masked study personnel. The number of daily IPTp tablets was based on bodyweight and according to the treatment guidelines set by WHO (target dose of 4 mg/kg per day [range 2-10 mg/kg per day] of dihydroartemisinin and 18 mg/kg per day [range 16-27 mg/kg per day] of piperaquine given once a day for 3 days). At enrolment, all participants received co-trimoxazole (fixed combination drug containing 800 mg trimethoprim and 160 mg sulfamethoxazole) for daily intake. The primary study outcome was prevalence of peripheral parasitaemia detected by microscopy at delivery. The modified intention-to-treat population included all randomly assigned women who had data for the primary outcome. Secondary outcomes included frequency of adverse events, incidence of clinical malaria during pregnancy, and frequency of poor pregnancy outcomes. All study personnel, investigators, outcome assessors, data analysts, and participants were masked to treatment assignment. This study is registered with ClinicalTrials.gov, NCT03671109. FINDINGS: From Sept 18, 2019, to Nov 26, 2021, 666 women (mean age 28·5 years [SD 6·4]) were enrolled and randomly assigned to the intervention (n=332) and control (n=334) groups. 294 women in the intervention group and 308 women in the control group had peripheral blood samples taken at delivery and were included in the primary analysis. Peripheral parasitaemia at delivery was detected in one (<1%) of 294 women in the intervention group and none of 308 women in the control group. The incidence of clinical malaria during pregnancy was lower in the intervention group than in the control group (one episode in the intervention group vs six in the control group; relative risk [RR] 0·12, 95% CI 0·03-0·52, p=0·045). In a post-hoc analysis, the composite outcome of overall malaria infection (detected by any diagnostic test during pregnancy or delivery) was lower in the intervention group than in the control group (14 [5%] of 311 women vs 31 [10%] of 320 women; RR 0·48, 95% CI 0·27-0·84, p=0·010). The frequency of serious adverse events and poor pregnancy outcomes (such as miscarriages, stillbirths, premature births, and congenital malformations) did not differ between groups. The most frequently reported drug-related adverse events were gastrointestinal disorder (reported in less than 4% of participants) and headache (reported in less than 2% of participants), with no differences between study groups. INTERPRETATION: In the context of low malaria transmission, the addition of IPTp with dihydroartemisinin-piperaquine to co-trimoxazole prophylaxis in pregnant women with HIV did not reduce peripheral parasitaemia at delivery. However, the intervention was safe and associated with a decreased risk of clinical malaria and overall Plasmodium falciparum infection, so it should be considered as a strategy to protect pregnant women with HIV from malaria. FUNDING: European and Developing Countries Clinical Trials Partnership 2 (EDCTP2) and Medicines for Malaria Venture. TRANSLATIONS: For the Portuguese and French translations of the abstract see Supplementary Materials section.

Co-delivery of artemisinin and metformin via PEGylated niosomal nanoparticles: potential anti-cancer effect in treatment of lung cancer cells.

PURPOSE: Despite the advances in treatment, lung cancer is a global concern and necessitates the development of new treatments. Biguanides like metformin (MET) and artemisinin (ART) have recently been discovered to have anti-cancer properties. As a consequence, in the current study, the anti-cancer effect of MET and ART co-encapsulated in niosomal nanoparticles on lung cancer cells was examined to establish an innovative therapy technique. METHODS: Niosomal nanoparticles (Nio-NPs) were synthesized by thin-film hydration method, and their physicochemical properties were assessed by FTIR. The morphology of Nio-NPs was evaluated with FE-SEM and AFM. The MTT assay was applied to evaluate the cytotoxic effects of free MET, free ART, their encapsulated form with Nio-NPs, as well as their combination, on A549 cells. Apoptosis assay was utilized to detect the biological processes involved with programmed cell death. The arrest of cell cycle in response to drugs was assessed using a cell cycle assay. Following a 48-h drug treatment, the expression level of hTERT, Cyclin D1, BAX, BCL-2, Caspase 3, and 7 genes were assessed using the qRT-PCR method. RESULTS: Both MET and ART reduced the survival rate of lung cancer cells in the dose-dependent manner. The IC50 values of pure ART and MET were 195.2 µM and 14.6 mM, respectively while in nano formulated form their IC50 values decreased to 56.7 µM and 78.3 µM, respectively. The combination of MET and ART synergistically decreased the proliferation of lung cancer cells, compared to the single treatments. Importantly, the combination of MET and ART had a higher anti-proliferative impact against A549 lung cancer cells, with lower IC50 values. According to the result of Real-time PCR, hTERT, Cyclin D1, BAX, BCL-2, Caspase 3, and Caspase 7 genes expression were considerably altered in treated with combination of nano formulated MET and ART compared to single therapies. CONCLUSION: The results of this study showed that the combination of MET and ART encapsulated in Nio-NPs could be useful for the treatment of lung cancer and can increase the efficiency of lung cancer treatment.

Increasing Prevalence of Artemisinin-Resistant HRP2-Negative Malaria in Eritrea.

BACKGROUND: Although the clinical efficacy of antimalarial artemisinin-based combination therapies in Africa remains high, the recent emergence of partial resistance to artemisinin in Plasmodium falciparum on the continent is troubling, given the lack of alternative treatments. METHODS: In this study, we used data from drug-efficacy studies conducted between 2016 and 2019 that evaluated 3-day courses of artemisinin-based combination therapy (artesunate-amodiaquine or artemether-lumefantrine) for uncomplicated malaria in Eritrea to estimate the percentage of patients with day-3 positivity (i.e., persistent P. falciparum parasitemia 3 days after the initiation of therapy). We also assayed parasites for mutations in Pfkelch13 as predictive markers of partial resistance to artemisinin and screened for deletions in hrp2 and hrp3 that result in variable performance of histidine rich protein 2 (HRP2)-based rapid diagnostic tests for malaria. RESULTS: We noted an increase in the percentage of patients with day-3 positivity from 0.4% (1 of 273) in 2016 to 1.9% (4 of 209) in 2017 and 4.2% (15 of 359) in 2019. An increase was also noted in the prevalence of the Pfkelch13 R622I mutation, which was detected in 109 of 818 isolates before treatment, from 8.6% (24 of 278) in 2016 to 21.0% (69 of 329) in 2019. The odds of day-3 positivity increased by a factor of 6.2 (95% confidence interval, 2.5 to 15.5) among the patients with Pfkelch13 622I variant parasites. Partial resistance to artemisinin, as defined by the World Health Organization, was observed in Eritrea. More than 5% of the patients younger than 15 years of age with day-3 positivity also had parasites that carried Pfkelch13 R622I. In vitro, the R622I mutation conferred a low level of resistance to artemisinin when edited into NF54 and Dd2 parasite lines. Deletions in both hrp2 and hrp3 were identified in 16.9% of the parasites that carried the Pfkelch13 R622I mutation, which made them potentially undetectable by HRP2-based rapid diagnostic tests. CONCLUSIONS: The emergence and spread of P. falciparum lineages with both Pfkelch13-mediated partial resistance to artemisinin and deletions in hrp2 and hrp3 in Eritrea threaten to compromise regional malaria control and elimination campaigns. (Funded by the Bill and Melinda Gates Foundation and others; Australian New Zealand Clinical Trials Registry numbers, ACTRN12618001223224, ACTRN12618000353291, and ACTRN12619000859189.).

Entomological impact of mass administration of ivermectin and dihydroartemisinin-piperaquine in The Gambia: a cluster-randomized controlled trial.

BACKGROUND: Vector control interventions in sub-Saharan Africa rely on insecticide-treated nets and indoor residual spraying. Insecticide resistance, poor coverage of interventions, poor quality nets and changes in vector behavior threaten the effectiveness of these interventions and, consequently, alternative tools are needed. Mosquitoes die after feeding on humans or animals treated with ivermectin (IVM). Mass drug administration (MDA) with IVM could reduce vector survival and decrease malaria transmission. The entomological impact of MDA of combined IVM and dihydroartemisinin-piperaquine was assessed in a community-based, cluster-randomized trial. METHODS: A cluster-randomized trial was implemented in 2018 and 2019 in 32 villages in the Upper River Region, The Gambia. The with the inhabitants of 16 intervention villages eligible to receive three monthly rounds of MDA at the beginning of the malaria transmission season. Entomological surveillance with light traps and human landing catches (HLC) was carried out during a 7- to 14-day period after each round of MDA, and then monthly until the end of the year. The mosquitocidal effect of IVM was determined by direct membrane feeding assays. RESULTS: Of the 15,017 mosquitoes collected during the study period, 99.65% (n = 14,965) were Anopheles gambiae sensu lato (An. gambiae s.l.), comprising Anopheles arabiensis (56.2%), Anopheles coluzzii (24.5%), Anopheles gambiae sensu stricto (An. gembiae s.s.; 16.0%) and Anopheles funestus sensu lato (An. funestus s.l.; 0.35%). No effect of the intervention on vector parity was observed. Vector density determined on light trap collections was significantly lower in the intervention villages in 2019 (adjusted incidence rate ratio: 0.39; 95% confidence interval [CI]: 0.20, 0.74; P = 0.005) but not in 2018. However, vector density determined in HLC collections was similar in both the intervention and control villages. The entomological inoculation rate was significantly lower in the intervention villages than in the control villages (odds ratio: 0.36, 95% CI: 0.19, 0.70; P = 0·003). Mosquito mortality was significantly higher when blood fed on IVM-treated individuals up to 21 days post-treatment, particularly in adults and individuals with a higher body mass index. CONCLUSION: Mass drug administration with IVM decreased vector density and the entomological inoculation rate while the effect on vector parity was less clear. Survival of mosquitoes fed on blood collected from IVM-treated individuals was significantly lower than that in mosquitoes which fed on controls. The influence of host characteristics on mosquito survivorship indicated that dose optimization could improve IVM efficacy. Future detailed entomological evaluation trials in which IVM is administered as stand-alone intervention may elucidate the contribution of this drug to the observed reduction in transmission.

Acute and subacute oral toxicity of artemisinin-hydroxychloroquine sulfate tablets in rats.

Artemisinin-hydroxychloroquine sulfate tablets (AH) are considered a relatively inexpensive and novel combination therapy for treating all forms of malaria, especially aminoquinoline drugs-resistant strains of P.falciparum. We aim to carry out acute and subacute oral toxicity studies in rats to acquire preclinical data on the safety of AH. Acute toxicity was evaluated in Sprague-Dawley (SD) rats at a single dose of 1980, 2970, 4450, 6670, and 10000 mg/kg. A 14-days subacute toxicity was assessed in SD rats at doses of 0, 146, 219, 328, and 429 mg/kg. The median lethal dose (LD50) of acute oral administration of AH in rats is found to be 3119 mg/kg, and toxic symptoms include decreased spontaneous activity, dyspnea, bristling, soft feces, spasticity, and convulsion. Repeated doses of AH have toxic effects on the nervous system, skin, blood system, liver, kidney, and spleen in rats. The main toxic reactions include epilation, emaciation, mental irritability, decreased body weight gain and food consumption, changes in the hematological and biochemical parameters, especially pathological lesions in the liver, kidney, and spleen. The no-observed-adverse-effect level (NOAEL) and lowest-observed-adverse-effect level (LOAEL) of AH are considered to be 219 mg/kg and 328 mg/kg, respectively.

Mass drug administration of ivermectin and dihydroartemisinin-piperaquine against malaria in settings with high coverage of standard control interventions: a cluster-randomised controlled trial in The Gambia.

BACKGROUND: Although the malaria burden has substantially decreased in sub-Saharan Africa, progress has stalled. We assessed whether mass administration of ivermectin (a mosquitocidal drug) and dihydroartemisinin-piperaquine (an antimalarial treatment) reduces malaria in The Gambia, an area with high coverage of standard control interventions. METHODS: This open-label, cluster-randomised controlled trial was done in the Upper River region of eastern Gambia. Villages with a baseline Plasmodium falciparum prevalence of 7-46% (all ages) and separated from each other by at least 3 km to reduce vector spillover were selected. Inclusion criteria were age and anthropometry (for ivermectin, weight of ≥15 kg; for dihydroartemisinin-piperaquine, participants older than 6 months); willingness to comply with trial procedures; and written informed consent. Villages were randomised (1:1) to either the intervention (ivermectin [orally at 300-400 µg/kg per day for 3 consecutive days] and dihydroartemisinin-piperaquine [orally depending on bodyweight] plus standard control interventions) or the control group (standard control interventions) using computer-based randomisation. Laboratory staff were masked to the origin of samples. In the intervention group, three rounds of mass drug administration once per month with ivermectin and dihydroartemisinin-piperaquine were given during two malaria transmission seasons from Aug 27 to Oct 31, 2018, and from July 15 to Sept 30, 2019. Primary outcomes were malaria prevalence by qPCR at the end of the second intervention year in November 2019, and Anopheles gambiae (s l) parous rate, analysed in the intention-to-treat population. This trial is registered with ClinicalTrials.gov, NCT03576313. FINDINGS: Between Nov 20 and Dec 7, 2017, 47 villages were screened for eligibility in the study. 15 were excluded because the baseline malaria prevalence was less than 7% (figure 1). 32 villages were enrolled and randomised to either the intervention or control group (n=16 in each group). The study population was 10 638, of which 4939 (46%) participants were in intervention villages. Coverage for dihydroartemisinin-piperaquine was between 49·0% and 58·4% in 2018, and between 76·1% and 86·0% in 2019; for ivermectin, coverage was between 46·9% and 52·2% in 2018, and between 71·7% and 82·9% in 2019. In November 2019, malaria prevalence was 12·8% (324 of 2529) in the control group and 5·1% (140 of 2722) in the intervention group (odds ratio [OR] 0·30, 95% CI 0·16-0·59; p<0·001). A gambiae (s l) parous rate was 83·1% (552 of 664) in the control group and 81·7% (441 of 540) in the intervention group (0·90, 0·66-1·25; p=0·537). In 2019, adverse events were recorded in 386 (9·7%) of 3991 participants in round one, 201 (5·4%) of 3750 in round two, and 168 (4·5%) of 3752 in round three. None of the 11 serious adverse events were related to the intervention. INTERPRETATION: The intervention was safe and well tolerated. In an area with high coverage of standard control interventions, mass drug administration of ivermectin and dihydroartemisinin-piperaquine significantly reduced malaria prevalence; however, no effect of ivermectin on vector parous rate was observed. FUNDING: Joint Global Health Trials Scheme. TRANSLATION: For the French translation of the abstract see Supplementary Materials section.

Enhanced Chemodynamic Therapy and Chemotherapy via Delivery of a Dual Threat ArtePt and Iodo-Click Reaction Mediated Glutathione Consumption.

Cisplatin has been used as standard regimen for hepatocellular carcinoma (HCC), but its therapeutic efficacy is greatly limited by the drug resistance. Cisplatin alone cannot achieve an ideal therapeutic outcome. Herein, a dual threat hybrid artemisinin platinum (ArtePt) is synthesized to combine chemodynamic therapy (CDT) with chemotherapy. On the one hand, artesunate can react with intracellular ferrous ion to generate reactive oxygen species (ROS) via Fenton reaction for CDT. On the other hand, cisplatin can target DNA for chemotherapy. However, GSH in cancer cells can effectively consume free radicals and detoxify cisplatin simultaneously, which compromised the efficacy of CDT and chemotherapy. Hence, an amphiphilic polymer with an iodine atom in the side chain is designed and encapsulated ArtePt to form NP(ArtePt). This iodine containing polymer NP(ArtePt) can effectively deplete intracellular GSH via an Iodo-Click reaction, thereby enhancing the effect of CDT as well as chemotherapy. Thereafter, a patient-derived xenograft model of hepatic carcinoma (PDXHCC ) is established to evaluate the therapeutic effect of NP(ArtePt), and a significant antitumor effect is achieved with NP(ArtePt). Overall, this study provides an effective strategy to combine CDT with chemotherapy to enhance the efficacy of cisplatin via Iodo-Click reaction, opening a new avenue for the cancer treatment.

Artemisinin Improves Acetylcholine-Induced Vasodilatation in Rats with Primary Hypertension.

PURPOSE: Endothelial dysfunction and the subsequent decrease in endothelium-dependent vascular relaxation of small arteries are major features of hypertension. Artemisinin, a well-known antimalarial drug, has been shown to exert protecting roles against endothelial cell injury in cardiac and pulmonary vascular diseases. The current study aimed to investigate the effects of artemisinin on endothelium-dependent vascular relaxation and arterial blood pressure, as well as the potential signalling pathways in spontaneously hypertensive rats (SHRs). METHODS: In this study, acetylcholine (ACh)-induced dose-dependent relaxation assays were performed to evaluate vascular endothelial function after treatment with artemisinin. Artemisinin was administered to the rats by intravenous injection or to arteries by incubation for the acute exposure experiments, and it was administered to rats by intraperitoneal injection for 28 days for the chronic experiments. RESULTS: Both acute and chronic administration of artemisinin decreased the heart rate and improved ACh-induced endothelium-dependent relaxation but negligibly affected the arterial blood pressure in SHRs. Incubation with artemisinin decreased basal vascular tension, NAD(P)H oxidase activity and reactive oxygen species (ROS) levels, but it also increased endothelial nitric oxide (NO) synthase (eNOS) activity and NO levels in the mesenteric artery, coronary artery, and pulmonary artery of SHRs. Artemisinin chronic administration to SHRs increased the protein expression of eNOS and decreased the protein expression of the NAD(P)H oxidase subunits NOX-2 and NOX-4 in the mesenteric artery. CONCLUSION: These results indicate that treatment with artemisinin has beneficial effects on reducing the heart rate and basal vascular tension and improving endothelium-dependent vascular relaxation in hypertension, which might occur by increasing eNOS activation and NO release and inhibiting NAD(P)H oxidase derived ROS production.

Identifying an optimal dihydroartemisinin-piperaquine dosing regimen for malaria prevention in young Ugandan children.

Intermittent preventive treatment (IPT) with dihydroartemisinin-piperaquine (DP) is highly protective against malaria in children, but is not standard in malaria-endemic countries. Optimal DP dosing regimens will maximize efficacy and reduce toxicity and resistance selection. We analyze piperaquine (PPQ) concentrations (n = 4573), malaria incidence data (n = 326), and P. falciparum drug resistance markers from a trial of children randomized to IPT with DP every 12 weeks (n = 184) or every 4 weeks (n = 96) from 2 to 24 months of age (NCT02163447). We use nonlinear mixed effects modeling to establish malaria protective PPQ levels and risk factors for suboptimal protection. Compared to DP every 12 weeks, DP every 4 weeks is associated with 95% protective efficacy (95% CI: 84-99%). A PPQ level of 15.4 ng/mL reduces the malaria hazard by 95%. Malnutrition reduces PPQ exposure. In simulations, we show that DP every 4 weeks is optimal across a range of transmission intensities, and age-based dosing improves malaria protection in young or malnourished children.

Metabolism is not a Major Contributor to the Toxicity of Piperaquine, a Long-acting Antimalarial Agent in Artemisinin-based Combination Therapy.

BACKGROUND: Hepatocellular damage has been reported for the antimalarial piperaquine (PQ) in the clinic after cumulative doses. OBJECTIVES: The role of metabolism in PQ toxicity was evaluated, and the mechanism mediating PQ hepatotoxicity was investigated. METHODS: The toxicity of PQ and its major metabolite (PQ N-oxide; M1) in mice was evaluated in terms of serum biochemical parameters. The role of metabolism in PQ toxicity was investigated in mice pretreated with an inhibitor of CYP450 (ABT) and/or FMO enzyme (MMI). The dose-dependent pharmacokinetics of PQ and M1 were studied in mice. Histopathological examination was performed to reveal the mechanism mediating PQ hepatotoxicity. RESULTS: Serum biochemical levels (ALT and BUN) increased significantly (P < 0.05) in mice after three-day oral doses of PQ (> 200 mg/kg/day), indicating hepatotoxicity and nephrotoxicity of PQ at a high dose. Weaker toxicity was observed for M1. Pretreatment with ABT and/or MMI did not increase PQ toxicity. PQ and M1 showed linear pharmacokinetics in mice after a single oral dose, and multiple oral doses led to their cumulative exposures. Histopathological examination showed that a high dose of PQ (> 200 mg/kg/day for three days) could induce hepatocyte apoptosis. The mRNA levels of targets in NF-κB and p53 pathways could be up-regulated by 2-30-fold in mice by PQ or M1. CONCLUSION: PQ metabolism led to detoxification of PQ, but there was a low possibility of altered toxicity induced by metabolism inhibition. The hepatotoxicity of PQ and its N-oxidation metabolite was partly mediated by NF-κB inflammatory pathway and p53 apoptosis pathway.

Ligand-modified homologous targeted cancer cell membrane biomimetic nanostructured lipid carriers for glioma therapy.

The main treatment measure currently used for glioma treatment is chemotherapy; the biological barrier of solid tumors hinders the deep penetration of nanomedicines and limits anticancer therapy. Furthermore, the poor solubility of many chemotherapeutic drugs limits the efficacy of antitumor drugs. Therefore, improving the solubility of chemotherapeutic agents and drug delivery to tumor tissues through the blood-brain barrier (BBB) and blood-brain tumor barrier (BBTB) are major challenges in glioma treatment. Nanostructured lipid carriers (NLCs) have high drug loading capacity, high stability, and high in vivo safety; moreover, they can effectively improve the solubility of insoluble drugs. Therefore, in this study, we used solvent volatilization and ultrasonic melting methods to prepare dihydroartemisinin nanostructured lipid carrier (DHA-NLC). We further used the glioma C6 cancer cell (CC) membrane to encapsulate DHA-NLC owing to the homologous targeting mechanism of the CC membrane; however, the targeting ability of the CC membrane was weak. We accordingly used targeting ligands for modification, and developed a bionanostructured lipid carrier with BBB and BBTB penetration and tumor targeting abilities. The results showed that DHA-loaded NGR/CCNLC (asparagine-glycine-arginine, NGR) was highly targeted, could penetrate the BBB and BBTB, and showed good anti-tumor effects both in vitro and in vivo, which could effectively prolong the survival time of tumor-bearing mice. Thus, the use of DHA-loaded NGR/CCNLC is an effective strategy for glioma treatment and has the potential to treat glioma.

Therapeutic Efficacy of Artemisinin-Based Combination Therapies in Democratic Republic of the Congo and Investigation of Molecular Markers of Antimalarial Resistance.

Routine assessment of the efficacy of artemisinin-based combination therapies (ACTs) is critical for the early detection of antimalarial resistance. We evaluated the efficacy of ACTs recommended for treatment of uncomplicated malaria in five sites in Democratic Republic of the Congo (DRC): artemether-lumefantrine (AL), artesunate-amodiaquine (ASAQ), and dihydroartemisinin-piperaquine (DP). Children aged 6-59 months with confirmed Plasmodium falciparum malaria were treated with one of the three ACTs and monitored. The primary endpoints were uncorrected and polymerase chain reaction (PCR)-corrected 28-day (AL and ASAQ) or 42-day (DP) cumulative efficacy. Molecular markers of resistance were investigated. Across the sites, uncorrected efficacy estimates ranged from 63% to 88% for AL, 73% to 100% for ASAQ, and 56% to 91% for DP. PCR-corrected efficacy estimates ranged from 86% to 98% for AL, 91% to 100% for ASAQ, and 84% to 100% for DP. No pfk13 mutations previously found to be associated with ACT resistance were observed. Statistically significant associations were found between certain pfmdr1 and pfcrt genotypes and treatment outcome. There is evidence of efficacy below the 90% cutoff recommended by WHO to consider a change in first-line treatment recommendations of two ACTs in one site not far from a monitoring site in Angola that has shown similar reduced efficacy for AL. Confirmation of these findings in future therapeutic efficacy monitoring in DRC is warranted.

Efficacy of dihydroartemisinin-piperaquine versus artemether-lumefantrine for the treatment of uncomplicated Plasmodium falciparum malaria among children in Africa: a systematic review and meta-analysis of randomized control trials.

BACKGROUND: Emergence of Plasmodium falciparum resistance to artemisinin and its derivatives poses a threat to the global effort to control malaria. The emergence of anti-malarial resistance has become a great public health challenge and continues to be a leading threat to ongoing malaria control efforts. The aim of this review was to synthesize available evidence on the efficacy of dihydroartemisinin-piperaquine (DHA-PQ) compared to artemether-lumefantrine (AL) for the treatment of uncomplicated falciparum malaria among children in Africa. METHODS: A systematic literature search was done to identify relevant articles from online databases PubMed/ MEDLINE, Embase, and Cochrane Central Register of Controlled Trials' database (CENTRAL) for retrieving randomized control trials comparing efficacy of DHA-PQ and AL for treatment of uncomplicated falciparum malaria in African children. The search was performed from August 2020 to April 2021. Using Rev-Man software (V5.4.1), R-studio and Comprehensive Meta-analysis software version 3, the extracted data from eligible studies were pooled as risk ratio (RR) with 95% confidence interval (CI). RESULTS: In this review, 25 studies which involved a total of 13,198 participants were included. PCR-unadjusted treatment failure in children aged between 6 months and 15 years was significantly lower in the DHA-PQ treatment arm on day 28 than that of AL (RR 0.14, 95% CI 0.08-0.26; participants = 1302; studies = 4; I2 = 0%, high quality of evidence). Consistently, the PCR-adjusted treatment failure was significantly lower with DHA-PQ treatment group on day 28 (RR 0.45, 95% CI 0.29-0.68; participants = 8508; studies = 16; I2 = 51%, high quality of evidence) and on day 42 (RR 0.60, 95% CI 0.47-0.78; participants = 5959; studies = 17; I2 = 0%, high quality of evidence). However, the efficacy was ≥ 95% in both treatment groups on day 28. CONCLUSION: From this review, it can be concluded that DHA-PQ reduces new infection and recrudescence on days 28 and 42 more than AL. This may trigger DHA-PQ to become a first-line treatment option.

Analysis of drug resistance marker genes of Plasmodium falciparum after implementation of artemisinin-based combination therapy in Pune district, India.

The global emergence and spread of malaria parasites resistant to antimalarial drugs is a major problem in malaria control and elimination. In this study, samples from Pune district were characterized to determine prevalence of molecular markers of resistance to chloroquine (pfcrt codons C72S, M74I, N75E, K76T and pfmdr-1 N86Y, Y184F), pyrimethamine (pfdhfr C50R, N51I, C59R, S108N), sulfadoxine (pfdhps, S436A, A437G, K540E, A581G), and artemisinin (pfkelch13, C580Y, R539T). The pfcrt K76T mutation was found in 78% samples as CVMNT, SVMNT and CVIET haplotype. The pfmdr-1 N86Y and Y184F mutations were found in 54% of samples. The pfdhfr double mutation C59R + S108N was present in 67% of samples, while the pfdhfr triple mutation (N51I + C59R + S108N) was not detected. The pfdhps mutations A437G and K540E were found in 67% of samples. Single mutants of pfdhps were rare, with K540E detected in only 6 patient samples. Similarly, pfdhps A581G was found in 13 of the isolates. The molecular markers associated with artemisinin resistance (mutations in pfkelch13 C580Y, R539T) were not detected in any of the isolates. These results suggest an emerging problem with multidrug-resistant P. falciparum. Though the genotype conventionally associated with artemisinin resistance was not observed, chloroquine-resistant genotype has reached complete fixation in the population. Moreover, the prevalence of mutations in both pfdhfr and pfdhps, with the presence of the quadruple mutant, indicates that continued monitoring is required to assess whether sulfadoxine-pyrimethamine can be used efficiently as a partner drug for artemisinin for the treatment of P. falciparum.

Selection of pfcrt K76 and pfmdr1 N86 Coding Alleles after Uncomplicated Malaria Treatment by Artemether-Lumefantrine in Mali.

BACKGROUND: Artemether-lumefantrine is a highly effective artemisinin-based combination therapy that was adopted in Mali as first-line treatment for uncomplicated Plasmodium falciparum malaria. This study was designed to measure the efficacy of artemether-lumefantrine and to assess the selection of the P. falciparum chloroquine resistance transporter (pfcrt) and P. falciparum multi-drug resistance 1 (pfmdr1) genotypes that have been associated with drug resistance. METHODS: A 28-day follow-up efficacy trial of artemether-lumefantrine was conducted in patients aged 6 months and older suffering from uncomplicated falciparum malaria in four different Malian areas during the 2009 malaria transmission season. The polymorphic genetic markers MSP2, MSP1, and Ca1 were used to distinguish between recrudescence and reinfection. Reinfection and recrudescence were then grouped as recurrent infections and analyzed together by PCR-restriction fragment length polymorphism (RFLP) to identify candidate markers for artemether-lumefantrine tolerance in the P. falciparum chloroquine resistance transporter (pfcrt) gene and the P. falciparum multi-drug resistance 1 (pfmdr1) gene. RESULTS: Clinical outcomes in 326 patients (96.7%) were analyzed and the 28-day uncorrected adequate clinical and parasitological response (ACPR) rate was 73.9%. The total PCR-corrected 28-day ACPR was 97.2%. The pfcrt 76T and pfmdr1 86Y population prevalence decreased from 49.3% and 11.0% at baseline (n = 337) to 38.8% and 0% in patients with recurrent infection (n = 85); p = 0.001), respectively. CONCLUSION: Parasite populations exposed to artemether-lumefantrine in this study were selected toward chloroquine-sensitivity and showed a promising trend that may warrant future targeted reintroduction of chloroquine or/and amodiaquine.

Reactive, self-administered malaria treatment against asymptomatic malaria infection: results of a cluster randomized controlled trial in The Gambia.

BACKGROUND: Selectively targeting and treating malaria-infected individuals may further decrease parasite carriage in low-burden settings. Using a trans-disciplinary approach, a reactive treatment strategy to reduce Plasmodium falciparum prevalence in participating communities was co-developed and tested. METHODS: This is a 2-arm, open-label, cluster-randomized trial involving villages in Central Gambia during the 2017 and 2018 malaria transmission season. Villages were randomized in a 1:1 ratio using a minimizing algorithm. In the intervention arm, trained village health workers delivered a full course of pre-packed dihydroartemisinin-piperaquine to all residents of compounds where clinical cases were reported while in the control arm, compound residents were screened for infection at the time of the index case reporting. All index cases were treated following national guidelines. The primary endpoint was malaria prevalence, determined by molecular methods, at the end of the intervention period. RESULTS: The trial was carried out in 50 villages: 34 in 2017 and 16 additional villages in 2018. At the end of the 2018 transmission season, malaria prevalence was 0.8% (16/1924, range 0-4%) and 1.1% (20/1814, range 0-17%) in the intervention and control arms, respectively. The odds of malaria infection were 29% lower in the intervention than in the control arm after adjustment for age (OR 0.71, 95% CI 0.27-1.84, p = 0.48). Adherence to treatment was high, with 98% (964/979) of those treated completing the 3-day treatment. Over the course of the study, only 37 villages, 20 in the intervention and 17 in the control arm, reported at least one clinical case. The distribution of clinical cases by month in both transmission seasons was similar and the odds of new clinical malaria cases during the trial period did not vary between arms (OR 1.04, 95% CI 0.57-1.91, p = 0.893). All adverse events were classified as mild to moderate and resolved completely. CONCLUSION: The systematic and timely administration of an anti-malarial treatment to residents of compounds with confirmed malaria cases did not significantly decrease malaria prevalence and incidence in communities where malaria prevalence was already low. Treatment coverage and adherence was very high. Results were strongly influenced by the lower-than-expected malaria prevalence, and by no clinical cases in villages with asymptomatic malaria-infected individuals. TRIAL REGISTRATION: This study is registered with ClinicalTrials.gov, NCT02878200. Registered 25 August 2016. https://clinicaltrials.gov/ct2/show/NCT02878200 .

Plasmodium falciparum phenotypic and genotypic resistance profile during the emergence of Piperaquine resistance in Northeastern Thailand.

Malaria remains a public health problem in Thailand, especially along its borders where highly mobile populations can contribute to persistent transmission. This study aimed to determine resistant genotypes and phenotypes of 112 Plasmodium falciparum isolates from patients along the Thai-Cambodia border during 2013-2015. The majority of parasites harbored a pfmdr1-Y184F mutation. A single pfmdr1 copy number had CVIET haplotype of amino acids 72-76 of pfcrt and no pfcytb mutations. All isolates had a single pfk13 point mutation (R539T, R539I, or C580Y), and increased % survival in the ring-stage survival assay (except for R539I). Multiple copies of pfpm2 and pfcrt-F145I were detected in 2014 (12.8%) and increased to 30.4% in 2015. Parasites containing either multiple pfpm2 copies with and without pfcrt-F145I or a single pfpm2 copy with pfcrt-F145I exhibited elevated IC90 values of piperaquine. Collectively, the emergence of these resistance patterns in Thailand near Cambodia border mirrored the reports of dihydroartemisinin-piperaquine treatment failures in the adjacent province of Cambodia, Oddar Meanchey, suggesting a migration of parasites across the border. As malaria elimination efforts ramp up in Southeast Asia, host nations militaries and other groups in border regions need to coordinate the proposed interventions.