Walter Otieno

Genetic diversity and protective efficacy of the RTS,S/AS01 malaria vaccine

BACKGROUND The RTS,S/AS01 vaccine targets the circumsporozoite protein of Plasmodium falciparum and has partial protective efficacy against clinical and severe malaria disease in infants and children. We investigated whether the vaccine efficacy was specific to certain parasite genotypes at the circumsporozoite protein locus. METHODS We used polymerase chain reaction-based next-generation sequencing of DNA extracted from samples from 4985 participants to survey circumsporozoite protein polymorphisms. We evaluated the effect that polymorphic positions and haplotypic regions within the circumsporozoite protein had on vaccine efficacy against first episodes of clinical malaria within 1 year after vaccination. RESULTS In the per-protocol group of 4577 RTS,S/AS01-vaccinated participants and 2335 control-vaccinated participants who were 5 to 17 months of age, the 1-year cumulative vaccine efficacy was 50.3% (95% confidence interval [CI], 34.6 to 62.3) against clinical malaria in which parasites matched the vaccine in the entire circumsporozoite protein C-terminal (139 infections), as compared with 33.4% (95% CI, 29.3 to 37.2) against mismatched malaria (1951 infections) (P=0.04 for differential vaccine efficacy). The vaccine efficacy based on the hazard ratio was 62.7% (95% CI, 51.6 to 71.3) against matched infections versus 54.2% (95% CI, 49.9 to 58.1) against mismatched infections (P=0.06). In the group of infants 6 to 12 weeks of age, there was no evidence of differential allele-specific vaccine efficacy. CONCLUSIONS These results suggest that among children 5 to 17 months of age, the RTS,S vaccine has greater activity against malaria parasites with the matched circumsporozoite protein allele than against mismatched malaria. The overall vaccine efficacy in this age category will depend on the proportion of matched alleles in the local parasite population; in this trial, less than 10% of parasites had matched alleles. (Funded by the National Institutes of Health and others.).

Immunogenicity of the RTS,S/AS01 malaria vaccine and implications for duration of vaccine efficacy: secondary analysis of data from a phase 3 randomised controlled trial

Summary Background The RTS,S/AS01 malaria vaccine targets the circumsporozoite protein, inducing antibodies associated with the prevention of Plasmodium falciparum infection. We assessed the association between anti-circumsporozoite antibody titres and the magnitude and duration of vaccine efficacy using data from a phase 3 trial done between 2009 and 2014. Methods Using data from 8922 African children aged 5–17 months and 6537 African infants aged 6–12 weeks at first vaccination, we analysed the determinants of immunogenicity after RTS,S/AS01 vaccination with or without a booster dose. We assessed the association between the incidence of clinical malaria and anti-circumsporozoite antibody titres using a model of anti-circumsporozoite antibody dynamics and the natural acquisition of protective immunity over time. Findings RTS,S/AS01-induced anti-circumsporozoite antibody titres were greater in children aged 5–17 months than in those aged 6–12 weeks. Pre-vaccination anti-circumsporozoite titres were associated with lower immunogenicity in children aged 6–12 weeks and higher immunogenicity in those aged 5–17 months. The immunogenicity of the booster dose was strongly associated with immunogenicity after primary vaccination. Anti-circumsporozoite titres wane according to a biphasic exponential distribution. In participants aged 5–17 months, the half-life of the short-lived component of the antibody response was 45 days (95% credible interval 42–48) and that of the long-lived component was 591 days (557–632). After primary vaccination 12% (11–13) of the response was estimated to be long-lived, rising to 30% (28–32%) after a booster dose. An anti-circumsporozoite antibody titre of 121 EU/mL (98–153) was estimated to prevent 50% of infections. Waning anti-circumsporozoite antibody titres predict the duration of efficacy against clinical malaria across different age categories and transmission intensities, and efficacy wanes more rapidly at higher transmission intensity. Interpretation Anti-circumsporozoite antibody titres are a surrogate of protection for the magnitude and duration of RTS,S/AS01 efficacy, with or without a booster dose, providing a valuable surrogate of effectiveness for new RTS,S formulations in the age groups considered. Funding UK Medical Research Council.

Complement receptor 1 polymorphisms associated with resistance to severe malaria in Kenya.

BackgroundIt has been hypothesized that the African alleles Sl2 and McCbof the Swain-Langley (Sl) and McCoy (McC) blood group antigens of the complement receptor 1 (CR1) may confer a survival advantage in the setting of Plasmodium falciparum malaria, but this has not been demonstrated.MethodsTo test this hypothesis, children in western Kenya with severe malaria-associated anaemia or cerebral malaria were matched to symptomatic uncomplicated malaria controls by age and gender. Swain-Langley and McCoy blood group alleles were determined by restriction fragment length polymorphism and conditional logistic regression was carried out.ResultsNo significant association was found between the African alleles and severe malaria-associated anaemia. However, children with Sl2/2 genotype were less likely to have cerebral malaria (OR = 0.17, 95% CI 0.04 to 0.72, P = 0.02) than children with Sl1/1. In particular, individuals with Sl2/2 McCa/bgenotype were less likely to have cerebral malaria (OR = 0.18, 95% CI 0.04 to 0.77, P = 0.02) than individuals with Sl1/1 McCa/a.ConclusionThese results support the hypothesis that the Sl2 allele and, possibly, the McCballele evolved in the context of malaria transmission and that in certain combinations probably confer a survival advantage on these populations.

Reduced immune complex binding capacity and increased complement susceptibility of red cells from children with severe malaria-associated anemia.

Plasmodium falciparum malaria causes 1–2 million deaths per year. Most deaths occur as a result of complications such as severe anemia and cerebral malaria (CM) (coma). Red cells of children with severe malaria-associated anemia (SMA) have acquired deficiencies in the complement regulatory proteins complement receptor 1 (CR1, CD35) and decay accelerating factor (DAF, CD55). We investigated whether these deficiencies affect the ability of erythrocytes to bind immune complexes (ICs) and regulate complement activation. We recruited 75 children with SMA (Hb ≤ 6 g/dL) from the holoendemic malaria region of the Lake Victoria basin, western Kenya, and 74 age- and gender-matched uncomplicated malaria controls. In addition, we recruited 32 children with CM and 52 age- and gender-matched controls. Deficiencies in red cell CR1 and CD55 in children with SMA were accompanied by a marked decline in IC binding capacity and increased C3b deposition in vivo and ex vivo. Importantly, these changes were specific because they were not seen in red cells of children with CM or their controls. These data suggest that the declines in red cell CR1 and CD55 seen in children with SMA are of physiologic significance and may predispose erythrocytes to complement-mediated damage and phagocytosis in vivo.

Results from tandem Phase 1 studies evaluating the safety, reactogenicity and immunogenicity of the vaccine candidate antigen Plasmodium falciparum FVO merozoite surface protein-1 (MSP142) administered intramuscularly with adjuvant system AS01

BackgroundThe development of an asexual blood stage vaccine against Plasmodium falciparum malaria based on the major merozoite surface protein-1 (MSP1) antigen is founded on the protective efficacy observed in preclinical studies and induction of invasion and growth inhibitory antibody responses. The 42 kDa C-terminus of MSP1 has been developed as the recombinant protein vaccine antigen, and the 3D7 allotype, formulated with the Adjuvant System AS02A, has been evaluated extensively in human clinical trials. In preclinical rabbit studies, the FVO allele of MSP142 has been shown to have improved immunogenicity over the 3D7 allele, in terms of antibody titres as well as growth inhibitory activity of antibodies against both the heterologous 3D7 and homologous FVO parasites.MethodsTwo Phase 1 clinical studies were conducted to examine the safety, reactogenicity and immunogenicity of the FVO allele of MSP142 in the adjuvant system AS01 administered intramuscularly at 0-, 1-, and 2-months: one in the USA and, after evaluation of safety data results, one in Western Kenya. The US study was an open-label, dose escalation study of 10 and 50 μg doses of MSP142 in 26 adults, while the Kenya study, evaluating 30 volunteers, was a double-blind, randomized study of only the 50 μg dose with a rabies vaccine comparator.ResultsIn these studies it was demonstrated that this vaccine formulation has an acceptable safety profile and is immunogenic in malaria-naïve and malaria-experienced populations. High titres of anti-MSP1 antibodies were induced in both study populations, although there was a limited number of volunteers whose serum demonstrated significant inhibition of blood-stage parasites as measured by growth inhibition assay. In the US volunteers, the antibodies generated exhibited better cross-reactivity to heterologous MSP1 alleles than a MSP1-based vaccine (3D7 allele) previously tested at both study sites.ConclusionsGiven that the primary effector mechanism for blood stage vaccine targets is humoral, the antibody responses demonstrated to this vaccine candidate, both quantitative (total antibody titres) and qualitative (functional antibodies inhibiting parasite growth) warrant further consideration of its application in endemic settings.Trial registrationsClinical Trials NCT00666380

Assessment of severe malaria in a multicenter, phase III, RTS, S/AS01 malaria candidate vaccine trial: case definition, standardization of data collection and patient care

BackgroundAn effective malaria vaccine, deployed in conjunction with other malaria interventions, is likely to substantially reduce the malaria burden. Efficacy against severe malaria will be a key driver for decisions on implementation. An initial study of an RTS, S vaccine candidate showed promising efficacy against severe malaria in children in Mozambique. Further evidence of its protective efficacy will be gained in a pivotal, multi-centre, phase III study. This paper describes the case definitions of severe malaria used in this study and the programme for standardized assessment of severe malaria according to the case definition.MethodsCase definitions of severe malaria were developed from a literature review and a consensus meeting of expert consultants and the RTS, S Clinical Trial Partnership Committee, in collaboration with the World Health Organization and the Malaria Clinical Trials Alliance. The same groups, with input from an Independent Data Monitoring Committee, developed and implemented a programme for standardized data collection.The case definitions developed reflect the typical presentations of severe malaria in African hospitals. Markers of disease severity were chosen on the basis of their association with poor outcome, occurrence in a significant proportion of cases and on an ability to standardize their measurement across research centres. For the primary case definition, one or more clinical and/or laboratory markers of disease severity have to be present, four major co-morbidities (pneumonia, meningitis, bacteraemia or gastroenteritis with severe dehydration) are excluded, and a Plasmodium falciparum parasite density threshold is introduced, in order to maximize the specificity of the case definition. Secondary case definitions allow inclusion of co-morbidities and/or allow for the presence of parasitaemia at any density. The programmatic implementation of standardized case assessment included a clinical algorithm for evaluating seriously sick children, improvements to care delivery and a robust training and evaluation programme for clinicians.ConclusionsThe case definition developed for the pivotal phase III RTS, S vaccine study is consistent with WHO recommendations, is locally applicable and appropriately balances sensitivity and specificity in the diagnosis of severe malaria. Processes set up to standardize severe malaria data collection will allow robust assessment of the efficacy of the RTS, S vaccine against severe malaria, strengthen local capacity and benefit patient care for subjects in the trial.Trial registrationClinicaltrials.gov NCT00866619

Distinct pattern of class and subclass antibodies in immune complexes of children with cerebral malaria and severe malarial anaemia.

Plasmodium falciparum infection can lead to deadly complications such as severe malaria‐associated anaemia (SMA) and cerebral malaria (CM). Children with severe malaria have elevated levels of circulating immune complexes (ICs). To further investigate the quantitative differences in antibody class/subclass components of ICs in SMA and CM, we enrolled 75 children with SMA and 32 children with CM from hospitals in western Kenya and matched them to 74 and 52 control children, respectively, with uncomplicated symptomatic malaria. Total IgG IC levels were always elevated in children with malaria upon enrolment, but children with CM had the highest levels of any group. Conditional logistic regression showed a borderline association between IgG4‐containing IC levels and increased risk of SMA (OR = 3·11, 95% CI 1·01–9·56, P = 0·05). Total IgG ICs (OR = 2·84, 95% CI 1·08–7·46, P = 0·03) and IgE‐containing ICs (OR = 6·82, OR 1·88–24·73, P ≤ 0·01) were associated with increased risk of CM. These results point to differences in the contribution of the different antibody class and subclass components of ICs to the pathogenesis of SMA and CM and give insight into potential mechanisms of disease.

The Levels of CD16/Fcγ Receptor IIIA on CD14+ CD16+ Monocytes Are Higher in Children with Severe Plasmodium falciparum Anemia than in Children with Cerebral or Uncomplicated Malaria

ABSTRACT Fc gamma receptor IIIA (CD16/FcγRIIIA) on monocytes/macrophages may play an important role in the pathogenesis of severe malarial anemia (SMA) by promoting phagocytosis of IgG-coated uninfected red cells and by allowing the production of tumor necrosis factor alpha (TNF-α) upon cross-linking by immune complexes (ICs). However, not much is known about the differential expression of this receptor on monocytes of children with severe malaria and uncomplicated malaria. Therefore, we investigated the expression of CD16/FcγRIIIA on monocytes of children with SMA, cerebral malaria (CM), and their age-matched uncomplicated malaria controls by flow cytometry. Since CD14low (CD14+) monocytes are considered more mature and macrophage-like than CD14high (CD14++) monocytes, we also compared the level of expression of CD16/FcγRIIIA according to the CD14 level and studied the relationship between CD16/FcγRIIIA expression and intracellular TNF-α production upon stimulation by ICs. CD16/FcγRIIIA expression was the highest overall on CD14+ CD16+ monocytes of children with SMA at enrollment. At convalescence, SMA children were the only ones to show a significant decline in the same parameter. In contrast, there were no significant differences among groups in the expression of CD16/FcγRIIIA on CD14++ CD16+ monocytes. A greater percentage of CD14+ CD16+ monocytes produced TNF-α upon stimulation than any other monocyte subset, and the amount of intracellular TNF-α correlated positively with CD16/FcγRIIIA expression. Furthermore, there was an inverse correlation between hemoglobin levels and CD16/FcγRIIIA expression in children with SMA and their controls. These data suggest that monocytes of children with SMA respond differently to Plasmodium falciparum infection by overexpressing CD16/FcγRIIIA as they mature, which could enhance erythrophagocytosis and TNF-α production.

Safety and immunogenicity of RTS,S/AS01 malaria vaccine in infants and children with WHO stage 1 or 2 HIV disease: a randomised, double-blind, controlled trial

BACKGROUND Malaria remains a major global public health concern, especially in sub-Saharan Africa. The RTS,S/AS01 malaria candidate vaccine was reviewed by the European Medicines Agency and received a positive scientific opinion; WHO subsequently recommended pilot implementation in sub-Saharan African countries. Because malaria and HIV overlap geographically, HIV-infected children should be considered for RTS,S/AS01 vaccination. We therefore aimed to assess the safety of RTS,S/AS01 in HIV-infected children at two sites in western Kenya. METHODS We did a randomised, double-blind, controlled trial at the clinical trial sites of the Kenya Medical Research Institute (KEMRI)-Walter Reed Army Institute of research in Kisumu and the KEMRI/US Centers for Disease Control and Prevention in Siaya. Eligible participants were infants and children aged from 6 weeks to 17 months with WHO stage 1 or 2 HIV disease (documented positive by DNA PCR), whether or not they were receiving antiretroviral therapy (ART). We randomly assigned participants (1:1) to receive three doses of either RTS,S/AS01 or rabies vaccine (both 0·5 mL per dose by intramuscular injection), given once per month at 0, 1, and 2 months. We did the treatment allocation using a web-based central randomisation system stratified by age (6 weeks-4 months, 5-17 months), and by baseline CD4% (<10, 10-14, 15-19, and ≥20). Data were obtained in an observer-blind manner, and the vaccine recipient, their parent or carer, the funder, and investigators responsible for the assessment of endpoints were all masked to treatment allocation (only staff responsible for the preparation and administration of the vaccines were aware of the assignment and these individuals played no other role in the study). We provided ART, even if the participants were not receiving ART before the study, and daily co-trimoxazole for prevention of opportunistic infections. The primary outcome was the occurrence of serious adverse events until 14 months after dose 1 of the vaccine, assessed in the intention-to-treat population. This trial was registered at ClinicalTrials.gov, number NCT01148459. FINDINGS Between July 30, 2010, and May 24, 2013, we enrolled 200 children to our study and randomly assigned 99 to receive RTS,S/AS01 and 101 to receive rabies vaccine. 177 (89%) of the 200 children enrolled completed 14 months of follow-up. Serious adverse events were noted in 41 (41·4%, 95% CI 31·6-51·8) of 99 RTS,S/AS01 recipients and 37 (36·6%, 27·3-46·8) of 101 rabies-vaccine recipients (relative risk 1·1, 95% CI 0·8-1·6). 20 (20·2%, 95% CI 12·8-29·5) of 99 RTS,S/AS01 recipients and 12 (11·9%, 6·3-19·8) of 101 rabies-vaccine recipients had at least one serious adverse event within 30 days after vaccination, mainly pneumonia, febrile convulsions, and salmonella sepsis. Five (5·1%, 95% CI 1·7-11·4) of 99 RTS,S/AS01 recipients and four (4·0%, 1·1-9·8) of 101 rabies-vaccine recipients died, but no deaths were deemed related to vaccination. Mortality was associated with five cases of pneumonia (1% RTS,S/AS01 recipients vs 3% rabies-vaccine recipients), five cases of gastroenteritis (3% RTS,S/AS01 recipients vs 2% rabies-vaccine recipients), five cases of malnutrition (2% RTS,S/AS01 recipients vs 3% rabies-vaccine recipients), one case of sepsis (1% rabies-vaccine recipients), one case of Haemophilus influenza meningitis (1% rabies-vaccine recipients), and one case of tuberculosis (1% RTS,S/AS01 recipients). INTERPRETATION RTS, S/AS01 was well tolerated when given to children with WHO clinical stage 1 or 2 HIV disease along with high antiretroviral and co-trimoxazole use. Children with HIV disease could be included in future RTS,S/AS01 vaccination programmes. FUNDING GlaxoSmithKline Biologicals SA and PATH Malaria Vaccine Initiative.

Clinical and Molecular Evidence for a Case of Buruli Ulcer (Mycobacterium ulcerans Infection) in Kenya

Mycobacterium ulcerans infection is an emerging disease that causes indolent, necrotizing skin lesions known as Buruli ulcer (BU) and occasional contiguous or metastatic bone lesions. Buruli ulcer is named after Buruli County in Uganda (east Africa), where an epidemic occurred in the 1960s. Today, BU is most common in central and west Africa. We describe clinical and molecular evidence for a case of BU in Kenya.