The Impact of Extended Treatment With Artemether-lumefantrine on Antimalarial Exposure and Reinfection Risks in Ugandan Children With Uncomplicated Malaria: A Randomized Controlled Trial.

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.

Efavirenz-Based Antiretroviral Therapy but Not Pregnancy Increased Unbound Piperaquine Exposure in Women during Malaria Chemoprevention.

Dihydroartemisinin-piperaquine (DP) is highly effective for malaria chemoprevention during pregnancy, but the standard dosing of DP that is used for nonpregnant adults may not be optimal for pregnant women. We previously reported that the pharmacokinetic exposure of total piperaquine (PQ; both bound and unbound to plasma proteins) is reduced significantly in the context of pregnancy or efavirenz (EFV)-based antiretroviral therapy (ART). However, as PQ is >99% protein-bound, reduced protein binding during pregnancy may lead to an increase in the pharmacologically active unbound drug fraction (fu), relative to the total PQ. We investigated the impact of pregnancy and EFV use on the fu of PQ to inform the interpretation of pharmacokinetics. Plasma samples from 0 to 24 h after the third (final) DP dose were collected from pregnant women at 28 weeks gestation who were receiving or not receiving EFV-based ART as well as from women 34 to 54 weeks postpartum who were not receiving EFV-based ART, who served as controls. Unbound PQ was quantified via ultrafiltration and liquid chromatography-tandem mass spectrometry, with fu being calculated as PQunbound/PQtotal. The geometric mean fu did not differ between pregnant and postpartum women (P = 0.66), but it was 23% (P < 0.01) greater in pregnant women receiving EFV-based ART, compared to that in postpartum women who were not receiving EFV-based ART. The altered drug-protein binding, potentially due to the displacement of PQ from plasma proteins by EFV, resulted in only a 14% lower unbound PQ exposure (P = 0.13) in the presence of a 31% lower total PQ exposure (P < 0.01), as estimated by the area under the concentration time curve from 0 to 24 h post-last dose in pregnant women who were receiving EFV-based ART. The results suggest that the impact of pregnancy and EFV-based ART on the exposure and, in turn, the efficacy of PQ for malaria prevention may not be as significant as was suggested by the changes in the total PQ exposure. Further study during the terminal elimination phase (e.g., on day 28 post-dose) would help better characterize the unbound PQ exposure during the full dosing interval and, thus, the overall efficacy of PQ for malaria chemoprevention in this special population.

Piperaquine Exposure Is Altered by Pregnancy, HIV, and Nutritional Status in Ugandan Women.

Dihydroartemisinin-piperaquine (DHA-PQ) provides highly effective therapy and chemoprevention for malaria in pregnant African women. PQ concentrations of >10.3 ng/ml have been associated with reduced maternal parasitemia, placental malaria, and improved birth outcomes. We characterized the population pharmacokinetics (PK) of PQ in a post hoc analysis of human immunodeficiency virus (HIV)-infected and -uninfected pregnant women receiving DHA-PQ as chemoprevention every 4 or 8 weeks. The effects of covariates such as pregnancy, nutritional status (body mass index [BMI]), and efavirenz (EFV)-based antiretroviral therapy were investigated. PQ concentrations from two chemoprevention trials were pooled to create a population PK database from 274 women and 2,218 PK observations. A three-compartment model with an absorption lag best fit the data. Consistent with our prior intensive PK evaluation, pregnancy and EFV use resulted in a 72% and 61% increased PQ clearance, compared to postpartum and HIV-uninfected pregnant women, respectively. Low BMI at 28 weeks of gestation was associated with increased clearance (2% increase per unit decrease in BMI). Low-BMI women given DHA-PQ every 8 weeks had a higher prevalence of parasitemia, malaria infection, and placental malaria compared to women with higher BMIs. The reduced piperaquine exposure in women with low BMI as well as during EFV coadministration, compared to pregnant women with higher BMIs and not taking EFV, suggests that these populations could benefit from weekly instead of monthly dosing for prevention of malaria parasitemia. Simulations indicated that because of the BMI-clearance relationship, weight-based regimens would not improve protection compared to a 2,880 mg fixed-dose regimen when provided monthly. (The clinical trials described in this paper have been registered at ClinicalTrials.gov under identifiers NCT02163447 and NCT02282293.).

Pregnancy outcomes and risk of placental malaria after artemisinin-based and quinine-based treatment for uncomplicated falciparum malaria in pregnancy: a WorldWide Antimalarial Resistance Network systematic review and individual patient data meta-analysis.

BACKGROUND: Malaria in pregnancy, including asymptomatic infection, has a detrimental impact on foetal development. Individual patient data (IPD) meta-analysis was conducted to compare the association between antimalarial treatments and adverse pregnancy outcomes, including placental malaria, accompanied with the gestational age at diagnosis of uncomplicated falciparum malaria infection. METHODS: A systematic review and one-stage IPD meta-analysis of studies assessing the efficacy of artemisinin-based and quinine-based treatments for patent microscopic uncomplicated falciparum malaria infection (hereinafter uncomplicated falciparum malaria) in pregnancy was conducted. The risks of stillbirth (pregnancy loss at ≥ 28.0 weeks of gestation), moderate to late preterm birth (PTB, live birth between 32.0 and < 37.0 weeks), small for gestational age (SGA, birthweight of < 10th percentile), and placental malaria (defined as deposition of malaria pigment in the placenta with or without parasites) after different treatments of uncomplicated falciparum malaria were assessed by mixed-effects logistic regression, using artemether-lumefantrine, the most used antimalarial, as the reference standard. Registration PROSPERO: CRD42018104013. RESULTS: Of the 22 eligible studies (n = 5015), IPD from16 studies were shared, representing 95.0% (n = 4765) of the women enrolled in literature. Malaria treatment in this pooled analysis mostly occurred in the second (68.4%, 3064/4501) or third trimester (31.6%, 1421/4501), with gestational age confirmed by ultrasound in 91.5% (4120/4503). Quinine (n = 184) and five commonly used artemisinin-based combination therapies (ACTs) were included: artemether-lumefantrine (n = 1087), artesunate-amodiaquine (n = 775), artesunate-mefloquine (n = 965), and dihydroartemisinin-piperaquine (n = 837). The overall pooled proportion of stillbirth was 1.1% (84/4361), PTB 10.0% (619/4131), SGA 32.3% (1007/3707), and placental malaria 80.1% (2543/3035), and there were no significant differences of considered outcomes by ACT. Higher parasitaemia before treatment was associated with a higher risk of SGA (adjusted odds ratio [aOR] 1.14 per 10-fold increase, 95% confidence interval [CI] 1.03 to 1.26, p = 0.009) and deposition of malaria pigment in the placenta (aOR 1.67 per 10-fold increase, 95% CI 1.42 to 1.96, p < 0.001). CONCLUSIONS: The risks of stillbirth, PTB, SGA, and placental malaria were not different between the commonly used ACTs. The risk of SGA was high among pregnant women infected with falciparum malaria despite treatment with highly effective drugs. Reduction of malaria-associated adverse birth outcomes requires effective prevention in pregnant women.

Comparison on simultaneous caillary and venous parasite density and genotyping results from children and adults with uncomplicated malaria: a prospective observational study in Uganda.

BACKGROUND: Blood smear microscopy remains the gold-standard method to diagnose and quantify malaria parasite density. In addition, parasite genotyping of select loci is the most utilized method for distinguishing recrudescent and new infections and to determine the number of strains per sample. In research settings, blood may be obtained from capillary or venous compartments, and results from these matrices have been used interchangeably. Our aim was to compare quantitative results for parasite density and strain complexity from both compartments. METHODS: In a prospective observational study, children and adults presenting with uncomplicated Plasmodium falciparum malaria, simultaneous capillary and venous blood smears and dried blood spots were collected over 42-days following treatment with artemether-lumefantrine. Blood smears were read by two microscopists, any discrepancies resolved by a third reader. Parasite DNA fingerprinting was conducted using six microsatellites. Bland Altman analysis and paired t-test/McNemar's test were used to assess the difference in density readings and measurements. RESULTS: Two hundred twenty-three participants were included in the analysis (177 children (35 HIV-infected/142 HIV-uninfected), 21 HIV-uninfected pregnant women, and 25 HIV-uninfected non-pregnant adults). Parasite density measurements did not statistically differ between capillary and venous blood smears at the time of presentation, nor over the course of 42-day follow-up. Characterization of merozoite surface protein-2 (MSP-2) genetic polymorphism demonstrated a higher level of strain diversity at the time of presentation in venous samples, as compared with capillary specimens (p = 0.02). There was a high degree of variability in genotype-corrected outcomes when pairs of samples from each compartment were compared using MSP-2 alone, although the variability was reduced with the use of multiple markers. CONCLUSIONS: Parasite density measurements do not statistically differ between capillary and venous compartments in all studied demographic groups at the time of presentation with malaria, or over the course of follow-up. More strains were detected by MSP-2 genotyping in venous samples than in capillary samples at the time of malaria diagnosis. The use of multiple polymorphic markers reduces the impact of variability in strain detection on genotype-corrected outcomes. This study confirms that both capillary and venous compartments can be used for sampling with confidence in the clinical research setting. TRIAL REGISTRATION: The trial was registered at ClinicalTrials.gov under registration no. NCT01717885 .

Intermittent Preventive Treatment for Malaria in Pregnancy: Optimization of Target Concentrations of Dihydroartemisinin-Piperaquine.

Background: Dihydroartemisinin-piperaquine (DHA-PQ) is highly efficacious as intermittent preventive therapy for malaria during pregnancy (IPTp). Determining associations between piperaquine (PQ) exposure, malaria risk, and adverse birth outcomes informs optimal dosing strategies. Methods: Human immunodeficiency virus-uninfected pregnant women (n = 300) were enrolled in a placebo-controlled trial of IPTp at 12-20 weeks' gestation and randomized to sulfadoxine-pyrimethamine every 8 weeks, DHA-PQ every 8 weeks, or DHA-PQ every 4 weeks during pregnancy. Pharmacokinetic sampling for PQ was performed every 4 weeks, and an intensive pharmacokinetic substudy was performed in 30 women at 28 weeks' gestation. Concentration-effect relationships were assessed between exposure to PQ; the prevalence of Plasmodium falciparum infection during pregnancy; outcomes at delivery including placental malaria, low birth weight, and preterm birth; and risks for toxicity. Simulations of new dosing scenarios were performed. Results: Model-defined PQ target venous plasma concentrations of 13.9 ng/mL provided 99% protection from P. falciparum infection during pregnancy. Each 10-day increase in time above target PQ concentrations was associated with reduced odds of placental parasitemia, preterm birth, and low birth weight, though increases in PQ concentrations were associated with QT interval prolongation. Modeling suggests that daily or weekly administration of lower dosages of PQ, compared to standard dosing, will maintain PQ trough levels above target concentrations with reduced PQ peak levels, potentially limiting toxicity. Conclusions: The protective efficacy of IPTp with DHA-PQ was strongly associated with higher drug exposure. Studies of the efficacy and safety of alternative DHA-PQ IPTp dosing strategies are warranted. Clinical Trials Registration: NCT02163447.

Antiretroviral Choice for HIV Impacts Antimalarial Exposure and Treatment Outcomes in Ugandan Children.

BACKGROUND: The optimal treatment of malaria in human immunodeficiency virus (HIV)-infected children requires consideration of critical drug-drug interactions in coinfected children, as these may significantly impact drug exposure and clinical outcomes. METHODS: We conducted an intensive and sparse pharmacokinetic/pharmacodynamic study in Uganda of the most widely adopted artemisinin-based combination therapy, artemether-lumefantrine. HIV-infected children on 3 different first-line antiretroviral therapy (ART) regimens were compared to HIV-uninfected children not on ART, all of whom required treatment for Plasmodium falciparum malaria. Pharmacokinetic sampling for artemether, dihydroartemisinin, and lumefantrine exposure was conducted through day 21, and associations between drug exposure and outcomes through day 42 were investigated. RESULTS: One hundred forty-five and 225 children were included in the intensive and sparse pharmacokinetic analyses, respectively. Compared with no ART, efavirenz (EFV) reduced exposure to all antimalarial components by 2.1- to 3.4-fold; lopinavir/ritonavir (LPV/r) increased lumefantrine exposure by 2.1-fold; and nevirapine reduced artemether exposure only. Day 7 concentrations of lumefantrine were 10-fold lower in children on EFV vs LPV/r-based ART, changes that were associated with an approximate 4-fold higher odds of recurrent malaria by day 28 in those on EFV vs LPV/r-based ART. CONCLUSIONS: The choice of ART in children living in a malaria-endemic region has highly significant impacts on the pharmacokinetics and pharmacodynamics of artemether-lumefantrine treatment. EFV-based ART reduces all antimalarial components and is associated with the highest risk of recurrent malaria following treatment. For those on EFV, close clinical follow-up for recurrent malaria following artemether-lumefantrine treatment, along with the study of modified dosing regimens that provide higher exposure, is warranted.

Artemether-Lumefantrine Pharmacokinetics and Clinical Response Are Minimally Altered in Pregnant Ugandan Women Treated for Uncomplicated Falciparum Malaria.

Artemether-lumefantrine is a first-line regimen for the treatment of uncomplicated malaria during the second and third trimesters of pregnancy. Previous studies have reported changes in the pharmacokinetics and clinical outcomes following treatment with artemether-lumefantrine in pregnant women compared to nonpregnant adults; however, the results are inconclusive. We conducted a study in rural Uganda to compare the pharmacokinetics of artemether-lumefantrine and the treatment responses between 30 pregnant women and 30 nonpregnant adults with uncomplicated Plasmodium falciparum malaria. All participants were uninfected with HIV, treated with a six-dose regimen of artemether-lumefantrine, and monitored clinically for 42 days. The pharmacokinetics of artemether, its metabolite dihydroartemisinin, and lumefantrine were evaluated for 21 days following treatment. We found no significant differences in the overall pharmacokinetics of artemether, dihydroartemisinin, or lumefantrine in a direct comparison of pregnant women to nonpregnant adults, except for a statistically significant but small difference in the terminal elimination half-lives of both dihydroartemisinin and lumefantrine. There were seven PCR-confirmed reinfections (5 pregnant and 2 nonpregnant participants). The observation of a shorter terminal half-life for lumefantrine may have contributed to a higher frequency of reinfection or a shorter posttreatment prophylactic period in pregnant women than in nonpregnant adults. While the comparable overall pharmacokinetic exposure is reassuring, studies are needed to further optimize antimalarial efficacy in pregnant women, particularly in high-transmission settings and because of emerging drug resistance. (This study is registered at ClinicalTrials.gov under registration no. NCT01717885.).

Prevalence of sexual coercion and its association with unwanted pregnancies among young pregnant females in Kampala, Uganda: a facility based cross-sectional study.

BACKGROUND: Sexual coercion is associated with sexually transmitted infections and unwanted pregnancies with consequential unsafe abortions and increased maternal morbidity and mortality. Current literature focuses mainly on its risk factors but less on its resultant deleterious health effects. We conducted a study to determine the prevalence of sexual coercion and its association with unwanted pregnancies among young pregnant women. METHODS: In a cross-sectional study, four hundred and sixteen (416) consenting pregnant females aged 15-24 years attending antenatal clinics in Lubaga division Kampala district in Uganda were enrolled using systematic sampling. Quantitative and qualitative data on sexual coercion were collected by female interviewers. Adjusted Prevalence Proportion Ratios (Adj. PPRs) of unwanted pregnancy and associated 95 % confidence intervals were estimated by generalized linear models with log link function and Poisson family distribution using robust variance estimator. Quantitative data were analyzed using Stata version 10.0, while qualitative data were analyzed using manifest content analysis. RESULTS: Prevalence of sexual coercion was 24 % and was higher among those who had non consensual sexual debut (29.0 %) compared with those who had consensual sexual debut (22.6 %). The prevalence of unwanted pregnancy was 18.3 % and was higher among participants who had been sexually coerced relative to their counterparts (p < 0.001). History of sexual coercion in the past 12 months and non consensual sexual debut were associated with unwanted pregnancy [adj.PPR = 2.23, 95 % CI: (1.49-3.32)] and 1.72, 95 % CI: (1.16- 2.54)] respectively. Qualitative results indicated that different forms/contexts of sexual coercion, such as deception, transactional sex and physical force influenced unwanted pregnancies. DISCUSSION: This study highlights that a quarter of our participants in our quantitative study had experienced sexual coercion in the past twelve months and nearly a third of these, had history of non consensual sexual debut. Unwanted pregnancy was higher among the sexually coerced and those who had non consensual sexual debut. CONCLUSION: Sexual coercion among pregnant women aged 15-24 years in Kampala, Uganda is high and is significantly associated with unwanted pregnancy. Comprehensive sex education targeting young people (<25 years), along with availability and access to youth friendly centers may be useful in addressing sexual coercion and its negative outcomes.

Determination of sulfadoxine and pyrimethamine in microvolume human plasma using ultra high performance liquid chromatography-tandam mass spectrometry.

To support the pharmacokinetic study of sulfadoxine (SD) and pyrimethamine (PM) in pregnant women and children, sensitive methods with small sample volume are desirable. Here we report a method to determine SD and PM with microvolume plasma samples: 5 µL plasma samples were cleaned up by protein precipitation with acetonitrile. The deuterated analytes were used as the internal standards. The samples after cleanup were injected onto an ACE Excel SuperC18 column (50 × 2.1 mm, 1.7 µm, Hichrom Limited) connected to a Waters I class UPLC coupled with a Sciex Triple Quad 6500+ Mass Spectrometer and eluted with water and acetonitrile both containing 0.1% formic acid in a gradient mode at 0.8mL/min. Detection utilized ESI+ as the ion source and MRM as the quantification mode. The precursor-to-product ion transitions m/z 311→245 for SD and 249→233 for PM were selected for quantification. The ion transitions for the corresponding internal standards were 315→249 for SD-d4 and 254→235 for PM-d3. The simplest linear regression weighted by 1/x was used for the calibration curves. The calibration ranges were 1-200 µg/mL SD and 2 - 1000ng/mL PM. The mean (± standard deviation) recoveries were 94.3±3.2% (SD) and 97.0±1.5% (PM). The validated method was applied to analysis of 1719 clinical samples, demonstrating the method is suitable for the pharmacokinetic study with samples collected up to day 28 post-dose.

Day 3 parasitemia and Plasmodium falciparum Kelch 13 mutations among uncomplicated malaria patients treated with artemether-lumefantrine in Adjumani district, Uganda.

Artemisinin resistance threatens malaria control and elimination efforts globally. Recent studies have reported the emergence of Plasmodium falciparum parasites tolerant to artemisinin agents in sub-Saharan Africa, including Uganda. The current study assessed the day 3 parasite clearance and its correlation with P. falciparum K13 propeller gene (pfkelch13) mutations in P. falciparum parasites isolated from patients with uncomplicated malaria under artemether-lumefantrine (AL) treatment. This study enrolled 100 P. falciparum-positive patients to whom AL was prescribed between 09/September/2022 and 06/November/2022. Blood samples were collected in EDTA tubes before treatment initiation (day 0) and on day 3. Parasitemia was assessed by microscopy from blood smears and quantitative polymerase chain reaction (qPCR) from the DNA extracted. The day 0 parasite K13 gene was sequenced using Sanger sequencing. Sequence data were analysed using MEGA version 11 software. The data were analysed using STATA version 15, and the Mann‒Whitney U test was used to compare PCR parasite clearance on day 3 using the comparative CT value method and pfkelch13 mutations. The prevalence of day 3 parasitaemia was 24% (24/100) by microscopy and 63% (63/100) by qPCR from the AL-treated patients. P. falciparum K13-propeller gene polymorphism was detected in 18.8% (15/80) of the day 0 DNA samples. The K13 mutations found were C469Y, 12.5% (10/80); A675V, 2.5% (2/80); A569S, 1.25%, (1/80), A578S, 1.25%, (1/80) and; F491S, 1.25%, (1/80) a new allele not reported anywhere. The C469Y mutation, compared to the wild-type, was associated with delayed parasite clearance p=0.0278, Hodges-Lehmann estimation 3.2108 on the log scale, (95%CI 1.7076, 4.4730). There was a high prevalence of day 3 P. falciparum among malaria patients treated using artemether-lumefantrine. We conclude that the K13 mutation associated with artemisinin resistance by P. falciparum is present in Adjumani district, Uganda. This necessitates regular surveillance of the effectiveness and efficacy of artemether-lumefantrine in the country.

Day 3 parasitemia and Plasmodium falciparum Kelch 13 mutations among uncomplicated malaria patients treated with artemether-lumefantrine in Adjumani district, Uganda.

Artemisinin resistance threatens malaria control and elimination efforts globally. Recent studies have reported the emergence of Plasmodium falciparum parasites tolerant to artemisinin agents in sub-Saharan Africa, including Uganda. The current study assessed the day 3 parasite clearance and its correlation with P. falciparum K13 propeller gene (pfkelch13) mutations in P. falciparum parasites isolated from patients with uncomplicated malaria under artemether-lumefantrine (AL) treatment. This study enrolled 100 P. falciparum-positive patients to whom AL was prescribed between 09/September/2022 and 06/November/2022. Blood samples were collected in EDTA tubes before treatment initiation (day 0) and on day 3. Parasitemia was assessed by microscopy from blood smears and quantitative polymerase chain reaction (qPCR) from the DNA extracted. The day 0 parasite K13 gene was sequenced using Sanger sequencing. Sequence data were analysed using MEGA version 11 software. The data were analysed using STATA version 15, and the Mann‒Whitney U test was used to compare PCR parasite clearance on day 3 using the comparative CT value method and pfkelch13 mutations. The prevalence of day 3 parasitaemia was 24% (24/100) by microscopy and 63% (63/100) by qPCR from the AL-treated patients. P. falciparum K13-propeller gene polymorphism was detected in 18.8% (15/80) of the day 0 DNA samples. The K13 mutations found were C469Y, 12.5% (10/80); A675V, 2.5% (2/80); A569S, 1.25%, (1/80), A578S, 1.25%, (1/80) and; F491S, 1.25%, (1/80) a new allele not reported anywhere. The C469Y mutation, compared to the wild-type, was associated with delayed parasite clearance p = 0.0278, Hodges-Lehmann estimation 3.2108 on the log scale, (95%CI 1.7076, 4.4730). There was a high prevalence of day 3 P. falciparum among malaria patients treated using artemether-lumefantrine. We conclude the presence of the K13 mutation associated with artemisinin resistance by P. falciparum in Adjumani district, Uganda, necessitates regular surveillance of the effectiveness and efficacy of artemether-lumefantrine in the country.

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.

Drug-Drug Interaction Between Dihydroartemisinin-Piperaquine and Sulfadoxine-Pyrimethamine During Malaria Chemoprevention in Pregnant Women.

Co-administration of dihydroartemisinin-piperaquine (DP) and sulfadoxine-pyrimethamine (SP) for intermittent preventive treatment of malaria in pregnancy (IPTp) may be superior in preventing adverse birth outcomes compared with either therapy alone, but potential drug-drug interactions require investigation. We conducted intensive and sparse pharmacokinetic (PK) studies in a subset of Ugandan women participating in a randomized controlled trial of monthly IPTp with SP vs. DP vs. DP + SP. Intensive PK sampling was performed from day 0 to 23 after dosing at 28 weeks gestation in 87 participants across treatment arms. Sparse sampling was performed on day 28 (trough) after dosing at 20 and 28 gestational weeks in additional 196 participants receiving SP vs. DP + SP. Intensive PK analysis demonstrated that compared with SP alone, co-administration of DP + SP was associated with lower maximal concentrations, the area under the concentration-time curves (AUC), and day 23 concentrations of sulfadoxine (25%, 25%, and 27%) and pyrimethamine (26%, 34%, and 32%) (P < 0.05 for all comparisons). Sparse PK results demonstrated participants co-administered DP + SP had lower trough concentrations after dosing at 20 and 28 gestational weeks for sulfadoxine (6%, P = 0.68 and 31%, P = 0.023, respectively) and pyrimethamine (18%, P = 0.032 and 33%, P < 0.001, respectively) compared with SP alone. Co-administration of DP + SP was associated with a 19% reduction in piperaquine AUC (P = 0.046), but no significant difference in other PK parameters compared with DP alone. In summary, co-administration of DP + SP was associated with significantly reduced SP exposure, with a greater magnitude during the third vs. second trimester. The clinical consequences of this interaction are yet unknown.

Facilitators and Barriers to Uptake of the Med Safety Mobile App for Adverse Drug Reaction Reporting by Health Workers in Uganda: A Qualitative Study.

INTRODUCTION: Adverse drug reactions (ADRs) are an important public health challenge worldwide; however, pharmacovigilance systems are plagued by under-reporting. Mobile technologies, including mobile applications such as Med Safety, could strengthen ADR reporting. We explored the acceptability, and factors that could influence uptake of, Med Safety for ADR reporting by health workers in Uganda. METHODS: The study took place between July and September 2020 in 12 HIV clinics in Uganda and employed a qualitative exploratory research design. We conducted 22 in-depth interviews and 3 mixed-gender focus group discussions (49 participants) with a diverse range of health workers. We analysed the data using a thematic approach. RESULTS: There was goodwill among the health workers to adopt Med Safety for ADR reporting and the majority would recommend the app to other health workers. Training with practice increased acceptability of the app. Uptake of the app was favoured by the younger, technology proficient, health worker demographic; the app's offline and two-way risk communication functionalities; availability of free internet hotspots at some health facilities; goodwill and willingness of health workers to report ADRs; and the cumbersome nature of conventional ADR reporting tools. Potential barriers to the uptake of Med Safety were the perceived lengthy processes of initial app registration and completion of multiple screens during ADR reporting; challenges with health workers' smartphones (incompatibility with application, no space for more applications, low battery charge); high cost of internet data; poor internet connectivity; difficulty in recognising ADRs, language barrier and poor feedback to ADR reporters. CONCLUSION: There was goodwill among the health workers to adopt Med Safety for ADR reporting and the majority would recommend the app to other health workers. Training with practice increased acceptability of the app and should be integral in all future app roll-out campaigns. The identified facilitators and barriers could be used to appropriately guide future research and implementation to promote the uptake of Med Safety for pharmacovigilance in low- and middle-income countries.

Impact of Drug Exposure on Resistance Selection Following Artemether-Lumefantrine Treatment for Malaria in Children With and Without HIV in Uganda.

Artemisinin-based combination therapies (ACTs) are the primary treatment for malaria. It is essential to characterize the pharmacokinetics (PKs) and pharmacodynamics (PDs) of ACTs in vulnerable populations at risk of suboptimal dosing. We developed a population PK/PD model using data from our previous study of artemether-lumefantrine in HIV-uninfected and HIV-infected children living in a high-transmission region of Uganda. HIV-infected children were on efavirenz-, nevirapine-, or lopinavir-ritonavir-based antiretroviral regimens, with daily trimethoprim-sulfamethoxazole prophylaxis. We assessed selection for resistance in two key parasite transporters, pfcrt and pfmdr1, over 42-day follow-up and incorporated genotyping into a time-to-event model to ascertain how resistance genotype in relation to drug exposure impacts recurrence risk. Two hundred seventy-seven children contributed 364 episodes to the model (186 HIV-uninfected and 178 HIV-infected), with recurrent microscopy-detectable parasitemia detected in 176 episodes by day 42. The final model was a two-compartment model with first-order absorption and an estimated age effect on bioavailability. Systemic lumefantrine exposure was highest with lopinavir-ritonavir, lowest with efavirenz, and equivalent with nevirapine and HIV-uninfected children. HIV status and lumefantrine concentration were significant factors associated with recurrence risk. Significant selection was demonstrated for pfmdr1 N86 and pfcrt K76 in recurrent infections, with no evidence of selection for pfmdr1 Y184F. Less sensitive parasites were able to tolerate lumefantrine concentrations ~ 3.5-fold higher than more sensitive parasites. This is the first population PK model of lumefantrine in HIV-infected children and demonstrates selection for reduced lumefantrine susceptibility, a concern as we confront the threat to ACTs posed by emerging artemisinin resistance in Africa.

Effectiveness of the Med Safety mobile application in improving adverse drug reaction reporting by healthcare professionals in Uganda: a protocol for a pragmatic cluster-randomised controlled trial.

INTRODUCTION: Combination antiretroviral therapy (cART) has massively reduced HIV mortality. However, long-term cART increases the risk of adverse drug reactions (ADRs), which can lead to higher morbidity, mortality and healthcare costs for people living with HIV (PLHIV).Pharmacovigilance-monitoring the effects of medicines-is essential for understanding real-world drug safety. In Uganda, pharmacovigilance systems have only recently been developed, and rates of ADR reporting for cART are very low. Thus, the safety profile of medicines currently used to treat HIV and tuberculosis in our population is poorly understood.The Med Safety mobile application has been developed through the European Union's Innovative Medicines Initiative WEB-Recognising Adverse Drug Reactions project to promote digital pharmacovigilance. This mobile application has been approved for ADR-reporting by Uganda's National Drug Authority. However, the barriers and facilitators to Med Safety uptake, and its effectiveness in improving pharmacovigilance, are as yet unknown. METHODS AND ANALYSIS: A pragmatic cluster-randomised controlled trial will be implemented over 30 months at 191 intervention and 191 comparison cART sites to evaluate Med Safety. Using a randomisation sequence generated by the sealed envelope software, we shall randomly assign the 382 prescreened cART sites to the intervention and comparison arms. Each cART site is a cluster that consists of healthcare professionals and PLHIV receiving dolutegravir-based cART and/or isoniazid preventive therapy. Healthcare professionals enrolled in the intervention arm will be trained in the use of mobile-based, paper-based and web-based reporting, while those in the comparison arm will be trained in paper-based and web-based reporting only. ETHICS AND DISSEMINATION: Ethical approval was given by the School of Biomedical Sciences Research and Ethics Committee at Makerere University (SBS-REC-720), and administrative clearance was obtained from Uganda National Council for Science and Technology (HS1366ES). Study results will be shared with healthcare professionals, policymakers, the public and academia. TRIAL REGISTRATION NUMBER: PACTR202009822379650.

Home management of malaria with artemether-lumefantrine compared with standard care in urban Ugandan children: a randomised controlled trial.

BACKGROUND: Home management of malaria-the presumptive treatment of febrile children with antimalarial drugs-is advocated to ensure prompt effective treatment of the disease. We assessed the effect of home delivery of artemether-lumefantrine on the incidence of antimalarial treatment and on clinical outcomes in children from an urban setting with fairly low malaria transmission. METHODS: In Kampala, Uganda, 437 children aged between 1 and 6 years from 325 households were randomly assigned by a computer-generated sequence to receive home delivery of prepackaged artemether-lumefantrine for presumptive treatment of febrile illnesses (n=225) or current standard of care (n=212). Randomisation was done by household after a pilot period of 1 month. After randomisation, study participants were followed up for an additional 12 months and information on their health and treatment of illnesses was obtained by use of monthly questionnaires and household diaries, which were completed by the participants' carers. The primary outcome was treatment incidence density per person-year. Analysis of the primary outcome was done on the modified intention-to-treat population, which included all participants apart from those excluded before data collection. This trial is registered with ClinicalTrials.gov, number NCT00115921. FINDINGS: Eight participants in the home management group and four in the standard care group were excluded before data collection; therefore, the primary analysis was done in 217 and 208 participants, respectively. The home management group received nearly twice the number of antimalarial treatments as the standard care group (4.66 per person-year vs 2.53 per person-year; incidence rate ratio [IRR] 1.72, 95% CI 1.43-2.06, p<0.0001), and nearly five times the number given to children with microscopically confirmed malaria in a comparable cohort of children (4.66 per person-year vs 1.03 per person-year, IRR 5.19, 95% CI 4.24-6.35, p<0.0001). Clinical data were available for 189 children in the home management group and 176 in the control group at study end; the main reasons for exclusion were movement out of the study area or loss to follow-up. The proportion of participants with parasitaemia at final assessment in the intervention group was lower than in the control group (four [2%] vs 17 [10%], p=0.006), but there were no other differences in standard malariometric indices, including anaemia. Serious adverse events were captured retrospectively. One child died in each group (home management-severe pneumonia and possible septicaemia; standard care-presumed respiratory failure). INTERPRETATION: Although home management of malaria led to prompt treatment of fever, there was little effect on clinical outcomes. The substantial over-treatment suggests that artemether-lumefantrine provided in the home might not be appropriate for large urban areas or settings with fairly low malaria transmission. FUNDING: Gates Malaria Partnership.

Determination of piperaquine concentration in human plasma and the correlation of capillary versus venous plasma concentrations.

BACKGROUND: A considerable challenge in quantification of the antimalarial piperaquine in plasma is carryover of analyte signal between assays. Current intensive pharmacokinetic studies often rely on the merging of venous and capillary sampling. Drug levels in capillary plasma may be different from those in venous plasma, Thus, correlation between capillary and venous drug levels needs to be established. METHODS: Liquid chromatography coupled with tandem mass spectrometry (LC-MS/MS) was used to develop the method. Piperaquine was measured in 205 pairs of capillary and venous plasma samples collected simultaneously at ≥24hr post dose in children, pregnant women and non-pregnant women receiving dihydroartemisinin-piperaquine as malaria chemoprevention. Standard three-dose regimen over three days applied to all participants with three 40mg dihydroartemisinin/320mg PQ tablets per dose for adults and weight-based dose for children. Correlation analysis was performed using the program Stata® SE12.1. Linear regression models were built using concentrations or logarithm transformed concentrations and the final models were selected based on maximal coefficient of determination (R2) and visual check. RESULTS: An LC-MS/MS method was developed and validated, utilizing methanol as a protein precipitation agent, a Gemini C18 column (50x2.0mm, 5µm) eluted with basic mobile phase solvents (ammonium hydroxide as the additive), and ESI+ as the ion source. This method had a calibration range of 10-1000 ng/mL and carryover was negligible. Correlation analysis revealed a linear relationship: Ccap = 1.04×Cven+4.20 (R2 = 0.832) without transformation of data, and lnCcap = 1.01×lnCven+0.0125, (R2 = 0.945) with natural logarithm transformation. The mean ratio (±SD) of Ccap/Cven was 1.13±0.42, and median (IQR) was 1.08 (0.917, 1.33). CONCLUSIONS: Capillary and venous plasma PQ measures are nearly identical overall, but not readily exchangeable due to large variation. Further correlation study accounting for disposition phases may be necessary.

Efavirenz-Based Antiretroviral Therapy Reduces Artemether-Lumefantrine Exposure for Malaria Treatment in HIV-Infected Pregnant Women.

BACKGROUND: The choice of malaria treatment for HIV-infected pregnant women receiving efavirenz-based antiretroviral therapy must consider the potential impact of drug interactions on antimalarial exposure and clinical response. The aim of this study was to investigate the effects of efavirenz on artemether-lumefantrine (AL) because no studies have isolated the impact of efavirenz for HIV-infected pregnant women. METHODS: A prospective clinical pharmacokinetic (PK) study compared HIV-infected, efavirenz-treated pregnant women with HIV-uninfected pregnant women in Tororo, Uganda. All women received the standard 6-dose AL treatment regimen for Plasmodium falciparum malaria with intensive PK samples collected over 21 days and 42-days of clinical follow-up. PK exposure parameters were calculated for artemether, its active metabolite dihydroartemisinin (DHA), and lumefantrine to determine the impact of efavirenz. RESULTS: Nine HIV-infected and 30 HIV-uninfected pregnant women completed intensive PK evaluations. Relative to controls, concomitant efavirenz therapy lowered the 8-hour artemether concentration by 76% (P = 0.013), DHA peak concentration by 46% (P = 0.033), and day 7 and 14 lumefantrine concentration by 61% and 81% (P = 0.046 and 0.023), respectively. In addition, there were nonsignificant reductions in DHA area under the concentration-time curve0-8hr (35%, P = 0.057) and lumefantrine area under the concentration-time curve0-∞ (34%, P = 0.063) with efavirenz therapy. CONCLUSIONS: Pregnant HIV-infected women receiving efavirenz-based antiretroviral therapy during malaria treatment with AL showed reduced exposure to both the artemisinin and lumefantrine. These data suggest that malaria and HIV coinfected pregnant women may require adjustments in AL dosage or treatment duration to achieve exposure comparable with HIV-uninfected pregnant women.