Feasibility, safety, and impact of the RTS,S/AS01E malaria vaccine when implemented through national immunisation programmes: evaluation of cluster-randomised introduction of the vaccine in Ghana, Kenya, and Malawi.

BACKGROUND: The RTS,S/AS01E malaria vaccine (RTS,S) was introduced by national immunisation programmes in Ghana, Kenya, and Malawi in 2019 in large-scale pilot schemes. We aimed to address questions about feasibility and impact, and to assess safety signals that had been observed in the phase 3 trial that included an excess of meningitis and cerebral malaria cases in RTS,S recipients, and the possibility of an excess of deaths among girls who received RTS,S than in controls, to inform decisions about wider use. METHODS: In this prospective evaluation, 158 geographical clusters (66 districts in Ghana; 46 sub-counties in Kenya; and 46 groups of immunisation clinic catchment areas in Malawi) were randomly assigned to early or delayed introduction of RTS,S, with three doses to be administered between the ages of 5 months and 9 months and a fourth dose at the age of approximately 2 years. Primary outcomes of the evaluation, planned over 4 years, were mortality from all causes except injury (impact), hospital admission with severe malaria (impact), hospital admission with meningitis or cerebral malaria (safety), deaths in girls compared with boys (safety), and vaccination coverage (feasibility). Mortality was monitored in children aged 1-59 months throughout the pilot areas. Surveillance for meningitis and severe malaria was established in eight sentinel hospitals in Ghana, six in Kenya, and four in Malawi. Vaccine uptake was measured in surveys of children aged 12-23 months about 18 months after vaccine introduction. We estimated that sufficient data would have accrued after 24 months to evaluate each of the safety signals and the impact on severe malaria in a pooled analysis of the data from the three countries. We estimated incidence rate ratios (IRRs) by comparing the ratio of the number of events in children age-eligible to have received at least one dose of the vaccine (for safety outcomes), or age-eligible to have received three doses (for impact outcomes), to that in non-eligible age groups in implementation areas with the equivalent ratio in comparison areas. To establish whether there was evidence of a difference between girls and boys in the vaccine's impact on mortality, the female-to-male mortality ratio in age groups eligible to receive the vaccine (relative to the ratio in non-eligible children) was compared between implementation and comparison areas. Preliminary findings contributed to WHO's recommendation in 2021 for widespread use of RTS,S in areas of moderate-to-high malaria transmission. FINDINGS: By April 30, 2021, 652 673 children had received at least one dose of RTS,S and 494 745 children had received three doses. Coverage of the first dose was 76% in Ghana, 79% in Kenya, and 73% in Malawi, and coverage of the third dose was 66% in Ghana, 62% in Kenya, and 62% in Malawi. 26 285 children aged 1-59 months were admitted to sentinel hospitals and 13 198 deaths were reported through mortality surveillance. Among children eligible to have received at least one dose of RTS,S, there was no evidence of an excess of meningitis or cerebral malaria cases in implementation areas compared with comparison areas (hospital admission with meningitis: IRR 0·63 [95% CI 0·22-1·79]; hospital admission with cerebral malaria: IRR 1·03 [95% CI 0·61-1·74]). The impact of RTS,S introduction on mortality was similar for girls and boys (relative mortality ratio 1·03 [95% CI 0·88-1·21]). Among children eligible for three vaccine doses, RTS,S introduction was associated with a 32% reduction (95% CI 5-51%) in hospital admission with severe malaria, and a 9% reduction (95% CI 0-18%) in all-cause mortality (excluding injury). INTERPRETATION: In the first 2 years of implementation of RTS,S, the three primary doses were effectively deployed through national immunisation programmes. There was no evidence of the safety signals that had been observed in the phase 3 trial, and introduction of the vaccine was associated with substantial reductions in hospital admission with severe malaria. Evaluation continues to assess the impact of four doses of RTS,S. FUNDING: Gavi, the Vaccine Alliance; the Global Fund to Fight AIDS, Tuberculosis and Malaria; and Unitaid.

Seasonal Malaria Vaccination with or without Seasonal Malaria Chemoprevention.

BACKGROUND: Malaria control remains a challenge in many parts of the Sahel and sub-Sahel regions of Africa. METHODS: We conducted an individually randomized, controlled trial to assess whether seasonal vaccination with RTS,S/AS01E was noninferior to chemoprevention in preventing uncomplicated malaria and whether the two interventions combined were superior to either one alone in preventing uncomplicated malaria and severe malaria-related outcomes. RESULTS: We randomly assigned 6861 children 5 to 17 months of age to receive sulfadoxine-pyrimethamine and amodiaquine (2287 children [chemoprevention-alone group]), RTS,S/AS01E (2288 children [vaccine-alone group]), or chemoprevention and RTS,S/AS01E (2286 children [combination group]). Of these, 1965, 1988, and 1967 children in the three groups, respectively, received the first dose of the assigned intervention and were followed for 3 years. Febrile seizure developed in 5 children the day after receipt of the vaccine, but the children recovered and had no sequelae. There were 305 events of uncomplicated clinical malaria per 1000 person-years at risk in the chemoprevention-alone group, 278 events per 1000 person-years in the vaccine-alone group, and 113 events per 1000 person-years in the combination group. The hazard ratio for the protective efficacy of RTS,S/AS01E as compared with chemoprevention was 0.92 (95% confidence interval [CI], 0.84 to 1.01), which excluded the prespecified noninferiority margin of 1.20. The protective efficacy of the combination as compared with chemoprevention alone was 62.8% (95% CI, 58.4 to 66.8) against clinical malaria, 70.5% (95% CI, 41.9 to 85.0) against hospital admission with severe malaria according to the World Health Organization definition, and 72.9% (95% CI, 2.9 to 92.4) against death from malaria. The protective efficacy of the combination as compared with the vaccine alone against these outcomes was 59.6% (95% CI, 54.7 to 64.0), 70.6% (95% CI, 42.3 to 85.0), and 75.3% (95% CI, 12.5 to 93.0), respectively. CONCLUSIONS: Administration of RTS,S/AS01E was noninferior to chemoprevention in preventing uncomplicated malaria. The combination of these interventions resulted in a substantially lower incidence of uncomplicated malaria, severe malaria, and death from malaria than either intervention alone. (Funded by the Joint Global Health Trials and PATH; ClinicalTrials.gov number, NCT03143218.).

Estimation of trajectory of protective efficacy in infectious disease prevention trials using recurrent event times.

In studies of infectious disease prevention, the level of protective efficacy of medicinal products such as vaccines and prophylactic drugs tends to vary over time. Many products require administration of multiple doses at scheduled times, as opposed to one-off or continual intervention. Accurate information on the trajectory of the level of protective efficacy over time facilitates informed clinical recommendations and implementation strategies, for example, with respect to the timing of administration of the doses. Based on concepts from pharmacokinetic and pharmacodynamic modeling, we propose a non-linear function for modeling the trajectory after each dose. The cumulative effect of multiple doses of the products is captured by an additive series of the function. The model has the advantages of parsimony and interpretability, while remaining flexible in capturing features of the trajectories. We incorporate this series into the Andersen-Gill model for analysis of recurrent event time data and compare it with alternative parametric and non-parametric functions. We use data on clinical malaria disease episodes from a trial of four doses of an anti-malarial drug combination for chemoprevention to illustrate, and evaluate the performance of the methods using simulation. The proposed method out-performed the alternatives in the analysis of real data in terms of Akaike and Bayesian Information Criterion. It also accurately captured the features of the protective efficacy trajectory such as the area under curve in simulations. The proposed method has strong potential to enhance the evaluation of disease prevention measures and improve their implementation strategies.

Facilitators and barriers to seasonal malaria chemoprevention (SMC) uptake in Nigeria: a qualitative approach.

BACKGROUND: SMC was adopted in Nigeria in 2014 and by 2021 was being implemented in 18 states, over four months between June and October by 143000 community drug distributors (CDDs) to a target population of 23million children. Further expansion of SMC is planned, extending to 21 states with four or five monthly cycles. In view of this massive scale-up, the National Malaria Elimination Programme undertook qualitative research in five states shortly after the 2021 campaign to understand community attitudes to SMC so that these perspectives inform future planning of SMC delivery in Nigeria. METHODS: In 20 wards representing urban and rural areas with low and high SMC coverage in five states, focus group discussions were held with caregivers, and in-depth interviews conducted with community leaders and community drug distributors. Interviews were also held with local government area and State malaria focal persons and at national level with the NMEP coordinator, and representatives of partners working on SMC in Nigeria. Interviews were recorded and transcribed, those in local languages translated into English, and transcripts analysed using NVivo software. RESULTS: In total, 84 focus groups and 106 interviews were completed. Malaria was seen as a major health concern, SMC was widely accepted as a key preventive measure, and community drug distributors (CDDs) were generally trusted. Caregivers preferred SMC delivered door-to-door to the fixed-point approach, because it allowed them to continue daily tasks, and allowed time for the CDD to answer questions. Barriers to SMC uptake included perceived side-effects of SMC drugs, a lack of understanding of the purpose of SMC, mistrust and suspicions that medicines provided free may be unsafe or ineffective, and local shortages of drugs. CONCLUSIONS: Recommendations from this study were shared with all community drug distributors and others involved in SMC campaigns during cascade training in 2022, including the need to strengthen communication about the safety and effectiveness of SMC, recruiting distributors from the local community, greater involvement of state and national level pharmacovigilance coordinators, and stricter adherence to the planned medicine allocations to avoid local shortages. The findings reinforce the importance of retaining door-to-door delivery of SMC.

Modelling non-linear patterns of time-varying intervention effects on recurrent events in infectious disease prevention studies.

Protective efficacy of vaccines and pharmaceutical products for prevention of infectious diseases usually vary over time. Information on the trajectory of the level of protection is valuable. We consider a parsimonious, non-linear and non-monotonic function for modelling time-varying intervention effects and compare it with several alternatives. The cumulative effects of multiple doses of intervention over time can be captured by an additive series of the function. We apply it to the Andersen-Gill model for analysis of recurrent time-to-event data. We re-analyze data from a trial of intermittent preventive treatment for malaria to illustrate and evaluate the method by simulation.

The Anti-Circumsporozoite Antibody Response of Children to Seasonal Vaccination With the RTS,S/AS01E Malaria Vaccine.

BACKGROUND: A trial in African children showed that combining seasonal vaccination with the RTS,S/AS01E vaccine with seasonal malaria chemoprevention reduced the incidence of uncomplicated and severe malaria compared with either intervention given alone. Here, we report on the anti-circumsporozoite antibody response to seasonal RTS,S/AS01E vaccination in children in this trial. METHODS: Sera from a randomly selected subset of children collected before and 1 month after 3 priming doses of RTS,S/AS01E and before and 1 month after 2 seasonal booster doses were tested for anti-circumsporozoite antibodies using enzyme-linked immunosorbent assay. The association between post-vaccination antibody titer and incidence of malaria was explored. RESULTS: A strong anti-circumsporozoite antibody response to 3 priming doses of RTS,S/AS01E was seen (geometric mean titer, 368.9 enzyme-linked immunosorbent assay units/mL), but titers fell prior to the first booster dose. A strong antibody response to an annual, pre-malaria transmission season booster dose was observed, but this was lower than after the primary vaccination series and lower after the second than after the first booster dose (ratio of geometric mean rise, 0.66; 95% confidence interval [CI], .57-.77). Children whose antibody response was in the upper tercile post-vaccination had a lower incidence of malaria during the following year than children in the lowest tercile (hazard ratio, 0.43; 95% CI, .28-.66). CONCLUSIONS: Seasonal vaccination with RTS,S/AS01E induced a strong booster antibody response that was lower after the second than after the first booster dose. The diminished antibody response to the second booster dose was not associated with diminished efficacy. CLINICAL TRIALS REGISTRATION: NCT03143218.

Effect of Adding Azithromycin to Seasonal Malaria Chemoprevention.

BACKGROUND: Mass administration of azithromycin for trachoma control led to a sustained reduction in all-cause mortality among Ethiopian children. Whether the addition of azithromycin to the monthly sulfadoxine-pyrimethamine plus amodiaquine used for seasonal malaria chemoprevention could reduce mortality and morbidity among African children was unclear. METHODS: We randomly assigned children 3 to 59 months of age, according to household, to receive either azithromycin or placebo, together with sulfadoxine-pyrimethamine plus amodiaquine, during the annual malaria-transmission season in Burkina Faso and Mali. The drug combinations were administered in four 3-day cycles, at monthly intervals, for three successive seasons. The primary end point was death or hospital admission for at least 24 hours that was not due to trauma or elective surgery. Data were recorded by means of active and passive surveillance. RESULTS: In July 2014, a total of 19,578 children were randomly assigned to receive seasonal malaria chemoprevention plus either azithromycin (9735 children) or placebo (9843 children); each year, children who reached 5 years of age exited the trial and new children were enrolled. In the intention-to-treat analysis, the overall number of deaths and hospital admissions during three malaria-transmission seasons was 250 in the azithromycin group and 238 in the placebo group (events per 1000 child-years at risk, 24.8 vs. 23.5; incidence rate ratio, 1.1; 95% confidence interval [CI], 0.88 to 1.3). Results were similar in the per-protocol analysis. The following events occurred less frequently with azithromycin than with placebo: gastrointestinal infections (1647 vs. 1985 episodes; incidence rate ratio, 0.85; 95% CI, 0.79 to 0.91), upper respiratory tract infections (4893 vs. 5763 episodes; incidence rate ratio, 0.85; 95% CI, 0.81 to 0.90), and nonmalarial febrile illnesses (1122 vs. 1424 episodes; incidence rate ratio, 0.79; 95% CI, 0.73 to 0.87). The prevalence of malaria parasitemia and incidence of adverse events were similar in the two groups. CONCLUSIONS: Among children in Burkina Faso and Mali, the addition of azithromycin to the antimalarial agents used for seasonal malaria chemoprevention did not result in a lower incidence of death or hospital admission that was not due to trauma or surgery than antimalarial agents plus placebo, although a lower disease burden was noted with azithromycin than with placebo. (Funded by the Joint Global Health Trials scheme; ClinicalTrials.gov number, NCT02211729.).

The duration of protection against clinical malaria provided by the combination of seasonal RTS,S/AS01E vaccination and seasonal malaria chemoprevention versus either intervention given alone.

BACKGROUND: A recent trial of 5920 children in Burkina Faso and Mali showed that the combination of seasonal vaccination with the RTS,S/AS01E malaria vaccine (primary series and two seasonal boosters) and seasonal malaria chemoprevention (four monthly cycles per year) was markedly more effective than either intervention given alone in preventing clinical malaria, severe malaria, and deaths from malaria. METHODS: In order to help optimise the timing of these two interventions, trial data were reanalysed to estimate the duration of protection against clinical malaria provided by RTS,S/AS01E when deployed seasonally, by comparing the group who received the combination of SMC and RTS,S/AS01E with the group who received SMC alone. The duration of protection from SMC was also estimated comparing the combined intervention group with the group who received RTS,S/AS01E alone. Three methods were used: Piecewise Cox regression, Flexible parametric survival models and Smoothed Schoenfeld residuals from Cox models, stratifying on the study area and using robust standard errors to control for within-child clustering of multiple episodes. RESULTS: The overall protective efficacy from RTS,S/AS01E over 6 months was at least 60% following the primary series and the two seasonal booster doses and remained at a high level over the full malaria transmission season. Beyond 6 months, protective efficacy appeared to wane more rapidly, but the uncertainty around the estimates increases due to the lower number of cases during this period (coinciding with the onset of the dry season). Protection from SMC exceeded 90% in the first 2-3 weeks post-administration after several cycles, but was not 100%, even immediately post-administration. Efficacy begins to decline from approximately day 21 and then declines more sharply after day 28, indicating the importance of preserving the delivery interval for SMC cycles at a maximum of four weeks. CONCLUSIONS: The efficacy of both interventions was highest immediately post-administration. Understanding differences between these interventions in their peak efficacy and how rapidly efficacy declines over time will help to optimise the scheduling of SMC, malaria vaccination and the combination in areas of seasonal transmission with differing epidemiology, and using different vaccine delivery systems. TRIAL REGISTRATION: The RTS,S-SMC trial in which these data were collected was registered at clinicaltrials.gov: NCT03143218.

Impact of seasonal RTS,S/AS01E vaccination plus seasonal malaria chemoprevention on the nutritional status of children in Burkina Faso and Mali.

BACKGROUND: A recent trial in Burkina Faso and Mali showed that combining seasonal RTS,S/AS01E malaria vaccination with seasonal malaria chemoprevention (SMC) substantially reduced the incidence of uncomplicated and severe malaria in young children compared to either intervention alone. Given the possible negative effect of malaria on nutrition, the study investigated whether these children also experienced lower prevalence of acute and chronic malnutrition. METHODS: In Burkina Faso and Mali 5920 children were randomized to receive either SMC alone, RTS,S/AS01E alone, or SMC combined with RTS,S/AS01E for three malaria transmission seasons (2017-2019). After each transmission season, anthropometric measurements were collected from all study children at a cross-sectional survey and used to derive nutritional status indicators, including the binary variables wasted and stunted (weight-for-height and height-for-age z-scores below - 2, respectively). Binary and continuous outcomes between treatment groups were compared by Poisson and linear regression. RESULTS: In 2017, compared to SMC alone, the combined intervention reduced the prevalence of wasting by approximately 12% [prevalence ratio (PR) = 0.88 (95% CI 0.75, 1.03)], and approximately 21% in 2018 [PR = 0.79 (95% CI 0.62, 1.01)]. Point estimates were similar for comparisons with RTS,S/AS01E, but there was stronger evidence of a difference. There was at least a 30% reduction in the point estimates for the prevalence of severe wasting in the combined group compared to the other two groups in 2017 and 2018. There was no difference in the prevalence of moderate or severe wasting between the groups in 2019. The prevalence of stunting, low-MUAC-for-age or being underweight did not differ between groups for any of the three years. The prevalence of severe stunting was higher in the combined group compared to both other groups in 2018, and compared to RTS,S/AS01E alone in 2017; this observation does not have an obvious explanation and may be a chance finding. Overall, malnutrition was very common in this cohort, but declined over the study as the children became older. CONCLUSIONS: Despite a high burden of malnutrition and malaria in the study populations, and a major reduction in the incidence of malaria in children receiving both interventions, this had only a modest impact on nutritional status. Therefore, other interventions are needed to reduce the high burden of malnutrition in these areas. TRIAL REGISTRATION: https://www.clinicaltrials.gov/ct2/show/NCT03143218 , registered 8th May 2017.

Ratio estimators of intervention effects on event rates in cluster randomized trials.

We consider five asymptotically unbiased estimators of intervention effects on event rates in non-matched and matched-pair cluster randomized trials, including ratio of mean counts r1 , ratio of mean cluster-level event rates r2 , ratio of event rates r3 , double ratio of counts r4 , and double ratio of event rates r5 . In the absence of an indirect effect, they all estimate the direct effect of the intervention. Otherwise, r1 , r2, and r3 estimate the total effect, which comprises the direct and indirect effects, whereas r4 and r5 estimate the direct effect only. We derive the conditions under which each estimator is more precise or powerful than its alternatives. To control bias in studies with a small number of clusters, we propose a set of approximately unbiased estimators. We evaluate their properties by simulation and apply the methods to a trial of seasonal malaria chemoprevention. The approximately unbiased estimators are practically unbiased and their confidence intervals usually have coverage probability close to the nominal level; the asymptotically unbiased estimators perform well when the number of clusters is approximately 32 or more per trial arm. Despite its simplicity, r1 performs comparably with r2 and r3 in trials with a large but realistic number of clusters. When the variability of baseline event rate is large and there is no indirect effect, r4 and r5 tend to offer higher power than r1 , r2, and r3 . We discuss the implications of these findings to the planning and analysis of cluster randomized trials.

The Duration of Protection from Azithromycin Against Malaria, Acute Respiratory, Gastrointestinal, and Skin Infections When Given Alongside Seasonal Malaria Chemoprevention: Secondary Analyses of Data from a Clinical Trial in Houndé, Burkina Faso, and Bougouni, Mali.

BACKGROUND: Mass drug administration (MDA) with azithromycin (AZ) is being considered as a strategy to promote child survival in sub-Saharan Africa, but the mechanism by which AZ reduces mortality is unclear. To better understand the nature and extent of protection provided by AZ, we explored the profile of protection by time since administration, using data from a household-randomized, placebo-controlled trial in Burkina Faso and Mali. METHODS: Between 2014 and 2016, 30 977 children aged 3-59 months received seasonal malaria chemoprevention (SMC) with sulfadoxine-pyrimethamine plus amodiaquine and either AZ or placebo monthly, on 4 occasions each year. Poisson regression with gamma-distributed random effects, accounting for the household randomization and within-individual clustering of illness episodes, was used to compare incidence of prespecified outcomes between SMC+AZ versus SMC+placebo groups in fixed time strata post-treatment. The likelihood ratio test was used to assess evidence for a time-treatment group interaction. RESULTS: Relative to SMC+placebo, there was no evidence of protection from SMC+AZ against hospital admissions and deaths. Additional protection from SMC+AZ against malaria was confined to the first 2 weeks post-administration (protective efficacy (PE): 24.2% [95% CI: 17.8%, 30.1%]). Gastroenteritis and pneumonia were reduced by 29.9% [21.7; 37.3%], and 34.3% [14.9; 49.3%], respectively, in the first 2 weeks postadministration. Protection against nonmalaria fevers with a skin condition persisted up to 28 days: PE: 46.3% [35.1; 55.6%]. CONCLUSIONS: The benefits of AZ-MDA are broad-ranging but short-lived. To maximize impact, timing of AZ-MDA must address the challenge of targeting asynchronous morbidity and mortality peaks from different causes.

Effectiveness of seasonal malaria chemoprevention (SMC) treatments when SMC is implemented at scale: Case-control studies in 5 countries.

BACKGROUND: Seasonal malaria chemoprevention (SMC) has shown high protective efficacy against clinical malaria and severe malaria in a series of clinical trials. We evaluated the effectiveness of SMC treatments against clinical malaria when delivered at scale through national malaria control programmes in 2015 and 2016. METHODS AND FINDINGS: Case-control studies were carried out in Mali and The Gambia in 2015, and in Burkina Faso, Chad, Mali, Nigeria, and The Gambia in 2016. Children aged 3-59 months presenting at selected health facilities with microscopically confirmed clinical malaria were recruited as cases. Two controls per case were recruited concurrently (on or shortly after the day the case was detected) from the neighbourhood in which the case lived. The primary exposure was the time since the most recent course of SMC treatment, determined from SMC recipient cards, caregiver recall, and administrative records. Conditional logistic regression was used to estimate the odds ratio (OR) associated with receipt of SMC within the previous 28 days, and SMC 29 to 42 days ago, compared with no SMC in the past 42 days. These ORs, which are equivalent to incidence rate ratios, were used to calculate the percentage reduction in clinical malaria incidence in the corresponding time periods. Results from individual countries were pooled in a random-effects meta-analysis. In total, 2,126 cases and 4,252 controls were included in the analysis. Across the 7 studies, the mean age ranged from 1.7 to 2.4 years and from 2.1 to 2.8 years among controls and cases, respectively; 42.2%-50.9% and 38.9%-46.9% of controls and cases, respectively, were male. In all 7 individual case-control studies, a high degree of personal protection from SMC against clinical malaria was observed, ranging from 73% in Mali in 2016 to 98% in Mali in 2015. The overall OR for SMC within 28 days was 0.12 (95% CI: 0.06, 0.21; p < 0.001), indicating a protective effectiveness of 88% (95% CI: 79%, 94%). Effectiveness against clinical malaria for SMC 29-42 days ago was 61% (95% CI: 47%, 72%). Similar results were obtained when the analysis was restricted to cases with parasite density in excess of 5,000 parasites per microlitre: Protective effectiveness 90% (95% CI: 79%, 96%; P<0.001), and 59% (95% CI: 34%, 74%; P<0.001) for SMC 0-28 days and 29-42 days ago, respectively. Potential limitations include the possibility of residual confounding due to an association between exposure to malaria and access to SMC, or differences in access to SMC between patients attending a clinic and community controls; however, neighbourhood matching of cases and controls, and covariate adjustment, attempted to control for these aspects, and the observed decline in protection over time, consistent with expected trends, argues against a major bias from these sources. CONCLUSIONS: SMC administered as part of routine national malaria control activities provided a very high level of personal protection against clinical malaria over 28 days post-treatment, similar to the efficacy observed in clinical trials. The case-control design used in this study can be used at intervals to ensure SMC treatments remain effective.

Evaluation of seasonal malaria chemoprevention in two areas of intense seasonal malaria transmission: Secondary analysis of a household-randomised, placebo-controlled trial in Houndé District, Burkina Faso and Bougouni District, Mali.

BACKGROUND: Seasonal malaria chemoprevention (SMC) is now widely deployed in the Sahel, including several countries that are major contributors to the global burden of malaria. Consequently, it is important to understand whether SMC continues to provide a high level of protection and how SMC might be improved. SMC was evaluated using data from a large, household-randomised trial in Houndé, Burkina Faso and Bougouni, Mali. METHODS AND FINDINGS: The parent trial evaluated monthly SMC plus either azithromycin (AZ) or placebo, administered as directly observed therapy 4 times per year between August and November (2014-2016). In July 2014, 19,578 children aged 3-59 months were randomised by household to study group. Children who remained within the age range 3-59 months in August each year, plus children born into study households or who moved into the study area, received study drugs in 2015 and 2016. These analyses focus on the approximately 10,000 children (5,000 per country) under observation each year in the SMC plus placebo group. Despite high coverage and high adherence to SMC, the incidence of hospitalisations or deaths due to malaria and uncomplicated clinical malaria remained high in the study areas (overall incidence rates 12.5 [95% confidence interval (CI): 11.2, 14.1] and 871.1 [95% CI: 852.3, 890.6] cases per 1,000 person-years, respectively) and peaked in July each year, before SMC delivery began in August. The incidence rate ratio comparing SMC within the past 28 days with SMC more than 35 days ago-adjusted for age, country, and household clustering-was 0.13 (95% CI: 0.08, 0.20), P < 0.001 for malaria hospitalisations and deaths from malaria and 0.21 (95% CI 0.20, 0.23), P < 0.001 for uncomplicated malaria, indicating protective efficacy of 87.4% (95% CI: 79.6%, 92.2%) and 78.3% (95% CI: 76.8%, 79.6%), respectively. The prevalence of malaria parasitaemia at weekly surveys during the rainy season and at the end of the transmission season was several times higher in children who missed the SMC course preceding the survey contact, and the smallest prevalence ratio observed was 2.98 (95% CI: 1.95, 4.54), P < 0.001. The frequency of molecular markers of sulfadoxine-pyrimethamine (SP) and amodiaquine (AQ) resistance did not increase markedly over the study period either amongst study children or amongst school-age children resident in the study areas. After 3 years of SMC deployment, the day 28 PCR-unadjusted adequate clinical and parasitological response rate of the SP + AQ regimen in children with asymptomatic malaria was 98.3% (95% CI: 88.6%, 99.8%) in Burkina Faso and 96.1% (95% CI: 91.5%, 98.2%) in Mali. Key limitations of this study are the potential overdiagnosis of uncomplicated malaria by rapid diagnostic tests and the potential for residual confounding from factors related to adherence to the monthly SMC schedule. CONCLUSION: Despite strong evidence that SMC is providing a high level of protection, the burden of malaria remains substantial in the 2 study areas. These results emphasise the need for continuing support of SMC programmes. A fifth monthly SMC course is needed to adequately cover the whole transmission season in the study areas and in settings with similar epidemiology. TRIAL REGISTRATION: The AZ-SMC trial in which these data were collected was registered at clinicaltrials.gov: NCT02211729.

Maximal dose-response of vitamin-K2 (menaquinone-4) on undercarboxylated osteocalcin in women with osteoporosis.

Low concentrations of serum vitamin K accompany high concentrations of undercarboxylated osteocalcin (ucOC) and osteoporotic fractures. Although vitamin K2 (MK-4) is approved as a therapeutic agent for the treatment of osteoporosis in some countries, the dose-response is unknown. The objective of this study was to assess the improvement in carboxylation of osteocalcin (OC) in response to escalating doses of MK-4 supplementation. A nine-week, open-labeled, prospective cohort study was conducted in 29 postmenopausal women who suffered hip or vertebral compression fractures. Participants took low-dose MK-4 (0.5 mg) for 3 weeks (until the second visit), then medium-dose MK-4 (5 mg) for 3 weeks (until the third visit), then high-dose MK-4 (45 mg) for 3 weeks. The mean ± SD age of the participants was 69 ± 9 years. MK-4 dose (p < 0.0001), but neither age nor other relevant medications (e.g. bisphosphonates) correlated with improvement in %ucOC. As compared to baseline concentrations (geometric mean ± SD) of 16.8 ± 2.4, 0.5 mg supplementation halved %ucOC to 8.7 ± 2.2 (p < 0.0001) and the 5-mg dose halved %ucOC again (to 3.9 ± 2.2; p = 0.0002 compared to 0.5-mg dose). However, compared to 5 mg/day, there was no additional benefit of 45 mg/day (%ucOC 4.6; p = NS vs. 5-mg dose). MK-4 supplementation resulted in borderline increases in γ-carboxylated osteocalcin (glaOC; p = 0.07). There were no major side effects of MK-4 supplementation. In postmenopausal women with osteoporotic fractures, supplementation with either 5 or 45 mg/day of MK-4 reduces ucOC to concentrations typical of healthy, pre-menopausal women.

Seasonal malaria chemoprevention combined with community case management of malaria in children under 10 years of age, over 5 months, in south-east Senegal: A cluster-randomised trial.

BACKGROUND: Seasonal malaria chemoprevention (SMC) is recommended in the Sahel region of Africa for children under 5 years of age, for up to 4 months of the year. It may be appropriate to include older children, and to provide protection for more than 4 months. We evaluated the effectiveness of SMC using sulfadoxine-pyrimethamine plus amodiaquine given over 5 months to children under 10 years of age in Saraya district in south-east Senegal in 2011. METHODS AND FINDINGS: Twenty-four villages, including 2,301 children aged 3-59 months and 2,245 aged 5-9 years, were randomised to receive SMC with community case management (CCM) (SMC villages) or CCM alone (control villages). In all villages, community health workers (CHWs) were trained to treat malaria cases with artemisinin combination therapy after testing with a rapid diagnostic test (RDT). In SMC villages, CHWs administered SMC to children aged 3 months to 9 years once a month for 5 months. The study was conducted from 27 July to 31 December 2011. The primary outcome was malaria (fever or history of fever with a positive RDT). The prevalence of anaemia and parasitaemia was measured in a survey at the end of the transmission season. Molecular markers associated with resistance to SMC drugs were analysed in samples from incident malaria cases and from children with parasitaemia in the survey. SMC was well tolerated with no serious adverse reactions. There were 1,472 RDT-confirmed malaria cases in the control villages and 270 in the SMC villages. Among children under 5 years of age, the rate difference was 110.8/1,000/month (95% CI 64.7, 156.8; p < 0.001) and among children 5-9 years of age, 101.3/1,000/month (95% CI 66.7, 136.0; p < 0.001). The mean haemoglobin concentration at the end of the transmission season was higher in SMC than control villages, by 6.5 g/l (95% CI 2.0, 11; p = 0.007) among children under 5 years of age, and by 5.2 g/l (95% CI 0.4, 9.9; p = 0.035) among children 5-9 years of age. The prevalence of parasitaemia was 18% in children under 5 years of age and 25% in children 5-9 years of age in the control villages, and 5.7% and 5.8%, respectively, in these 2 age groups in the SMC villages, with prevalence differences of 12.5% (95% CI 6.8%, 18.2%; p < 0.001) in children under 5 years of age and 19.3% (95% CI 8.3%, 30.2%; p < 0.001) in children 5-9 years of age. The pfdhps-540E mutation associated with clinical resistance to sulfadoxine-pyrimethamine was found in 0.8% of samples from malaria cases but not in the final survey. Twelve children died in the control group and 14 in the SMC group, a rate difference of 0.096/1,000 child-months (95% CI 0.99, 1.18; p = 0.895). Limitations of this study include that we were not able to obtain blood smears for microscopy for all suspected malaria cases, such that we had to rely on RDTs for confirmation, which may have included false positives. CONCLUSIONS: In this study SMC for children under 10 years of age given over 5 months was feasible, well tolerated, and effective in preventing malaria episodes, and reduced the prevalence of parasitaemia and anaemia. SMC with CCM achieved high coverage and ensured children with malaria were promptly treated with artemether-lumefantrine. TRIAL REGISTRATION: www.clinicaltrials.gov NCT01449045.

Sphingolipid changes do not underlie fatty acid-evoked GLUT4 insulin resistance nor inflammation signals in muscle cells.

Ceramides contribute to obesity-linked insulin resistance and inflammation in vivo, but whether this is a cell-autonomous phenomenon is debated, particularly in muscle, which dictates whole-body glucose uptake. We comprehensively analyzed lipid species produced in response to fatty acids and examined the consequence to insulin resistance and pro-inflammatory pathways. L6 myotubes were incubated with BSA-adsorbed palmitate or palmitoleate in the presence of myriocin, fenretinide, or fumonisin B1. Lipid species were determined by lipidomic analysis. Insulin sensitivity was scored by Akt phosphorylation and glucose transporter 4 (GLUT4) translocation, while pro-inflammatory indices were estimated by IκBα degradation and cytokine expression. Palmitate, but not palmitoleate, had mild effects on Akt phosphorylation but significantly inhibited insulin-stimulated GLUT4 translocation and increased expression of pro-inflammatory cytokines Il6 and Ccl2 Ceramides, hexosylceramides, and sphingosine-1-phosphate significantly heightened by palmitate correlated negatively with insulin sensitivity and positively with pro-inflammatory indices. Inhibition of sphingolipid pathways led to marked changes in cellular lipids, but did not prevent palmitate-induced impairment of insulin-stimulated GLUT4 translocation, suggesting that palmitate-induced accumulation of deleterious lipids and insulin resistance are correlated but independent events in myotubes. We propose that muscle cell-endogenous ceramide production does not evoke insulin resistance and that deleterious effects of ceramides in vivo may arise through ancillary cell communication.

Safety and Efficacy of Adding a Single Low Dose of Primaquine to the Treatment of Adult Patients With Plasmodium falciparum Malaria in Senegal, to Reduce Gametocyte Carriage: A Randomized Controlled Trial.

Introduction: More information is needed about the safety of low-dose primaquine in populations where G6PD deficiency is common. Methods: Adults with Plasmodium falciparum malaria were randomized to receive 1 of 3 artemisinin combination therapies (ACTs) with or without primaquine (0.25 mg/kg). Glucose-6-phosphate dehydrogenase (G6PD) status was determined using a rapid test. Patients were followed for 28 days to record hemoglobin concentration, adverse events, and gametocyte carriage. The primary end point was the change in Hb at day 7. Results: In sum, 274 patients were randomized, 139 received an ACT alone, and 135 received an ACT + primaquine. The mean reduction in Hb at day 7 was similar in each group, a difference in the ACT + PQ versus the ACT alone group of -0.04 g/dL (95% confidence interval [CI] -0.23, 0.31), but the effect of primaquine differed according to G6PD status. In G6PD-deficient patients the drop in Hb was 0.63 g/dL (95% CI 0.03, 1.24) greater in those who received primaquine than in those who received an ACT alone. In G6PD-normal patients, the reduction in Hb was 0.22 g/dL (95% CI -0.08, 0.52) less in those who received primaquine (interaction P = .01). One G6PD normal patient who received primaquine developed moderately severe anaemia (Hb < 8 g/dL). Dark urine was more frequent in patients who received primaquine. Primaquine was associated with a 73% (95% CI 24-90) reduction in gametocyte carriage (P = .013). Conclusion: Primaquine substantially reduced gametocyte carriage. However, the fall in Hb concentration at day 7 was greater in G6PD-deficient patients who received primaquine than in those who did not and one patient who received primaquine developed moderately severe anemia. Clinical Trial registration: PACTR201411000937373 (www.pactr.org).

Identification of 4-Aminopyrazolopyrimidine Metabolite That May Contribute to the Hypolipidemic Effects of LY2584702 in Long Evans Diet-Induced Obese Rats.

LY2584702 is an inhibitor of p70 S6 kinase-1 previously developed for the treatment of cancer. In two phase 1 trials in oncology patients, significant reductions of total cholesterol, low-density lipoprotein cholesterol (LDL-C), and triglyceride were observed. In the current study, we sought to understand the potential mechanism of action of this compound in regulating lipid metabolism. In Long Evans diet-induced obese (DIO) rats, oral administration of LY2584702 for 3-4 weeks led to robust reduction of LDL-C up to 60%. An unexpected finding of liver triglyceride (TG) increase implicated a metabolite of LY2584702, 4-aminopyrazolo[3,4-day]pyrimidine (4-APP), in modulation of lipid metabolism in these rats. We showed that low-dose 4-APP, when administered orally for 3-4 weeks to Long Evans DIO rats, produced lipoprotein profile changes that were strikingly similar to LY2584702. Kinetic studies suggested that both LY2584702 and 4-APP had no effect on chylomicron-TG secretion and only exerted a modest effect on hepatic very low-density lipoprotein (VLDL)-TG secretion. In human hepatoma HepG2 cells, 4-APP, but not LY2584702, increased LDL uptake. We hypothesize that generation of the 4-APP metabolite may contribute to the efficacy of LY2584702 in lowering LDL-C in rats and potentially in humans as well. This mechanism of LDL-C lowering may include inhibition of VLDL production and increase in LDL clearance.

Seasonal vaccination against malaria: a potential use for an imperfect malaria vaccine.

In many parts of the African Sahel and sub-Sahel, where malaria remains a major cause of mortality and morbidity, transmission of the infection is highly seasonal. Seasonal malaria chemoprevention (SMC), which involves administration of a full course of malaria treatment to young children at monthly intervals during the high transmission season, is proving to be an effective malaria control measure in these areas. However, SMC does not provide complete protection and it is demanding to deliver for both families and healthcare givers. Furthermore, there is a risk of the emergence in the future of resistance to the drugs, sulfadoxine-pyrimethamine and amodiaquine, that are currently being used for SMC. Substantial progress has been made in the development of malaria vaccines during the past decade and one malaria vaccine, RTS,S/AS01, has received a positive opinion from the European Medicines Authority and will soon be deployed in large-scale, pilot implementation projects in sub-Saharan Africa. A characteristic feature of this vaccine, and potentially of some of the other malaria vaccines under development, is that they provide a high level of efficacy during the period immediately after vaccination, but that this wanes rapidly, perhaps because it is difficult to develop effective immunological memory to malaria antigens in subjects exposed previously to malaria infection. A potentially effective way of using malaria vaccines with high initial efficacy but which provide only a short period of protection could be annual, mass vaccination campaigns shortly before each malaria transmission season in areas where malaria transmission is confined largely to a few months of the year.

Semiparametric estimation of time-varying intervention effects using recurrent event data.

We consider the estimation of the optimal interval between doses for interventions such as malaria chemoprevention and vaccine booster doses that are applied intermittently in infectious disease control. A flexible exponential-like function to model the time-varying intervention effect in the framework of Andersen-Gill model for recurrent event time data is considered. The partial likelihood estimation approach is adopted, and a large scale simulation study is carried out to evaluate the performance of the proposed method. A simple guideline for the choice of the optimal interval between successive doses is proposed. The methodology is illustrated with the analysis of data from a malaria chemoprevention trial. Copyright © 2017 John Wiley & Sons, Ltd.