Child Health and Infection with Low Density (CHILD) malaria: a protocol for a randomised controlled trial to assess the long-term health and socioeconomic impacts of testing and treating low-density malaria infection among children in Tanzania.

INTRODUCTION: As malaria declines, low-density malaria infections (LMIs) represent an increasing proportion of infections and may have negative impacts on child health and cognition, necessitating development of targeted and effective solutions. This trial assesses the health, cognitive and socioeconomic impact of two strategies for detecting and treating LMI in a low transmission setting. METHODS AND ANALYSIS: The study is a 3-arm open-label individually randomised controlled trial enrolling 600 children aged 6 months to 10 years in Bagamoyo district, Tanzania. Children are randomised to one of three arms: active case detection with molecular (ACDm) testing by high volume quantitative PCR (qPCR), passive case detection also with molecular testing (PCDm) and a control of standard PCD using rapid diagnostics tests (RDTs). Over the 2-year trial, ACDm participants receive malaria testing using RDT and qPCR three times annually, and malaria testing by RDT only when presenting with fever. PCDm and PCD participants receive malaria testing by RDT and qPCR or RDT only, respectively, when presenting with fever. RDT or qPCR positive participants with uncomplicated malaria are treated with artemether lumefantrine. The primary outcome is cumulative incidence of all-cause sick visits. Secondary outcomes include fever episodes, clinical failure after fever episodes, adverse events, malaria, non-malarial infection, antibiotic use, anaemia, growth faltering, cognition and attention, school outcomes, immune responses, and socioeconomic effects. Outcomes are assessed through monthly clinical assessments and testing, and baseline and endline neurodevelopmental testing. The trial is expected to provide key evidence and inform policy on health, cognitive and socioeconomic impact of interventions targeting LMI in children. ETHICS AND DISSEMINATION: Study is approved by Tanzania NatHREC and institutional review boards at University of California San Francisco and Ifakara Health Institute. Findings will be reported on ClinicalTrials.gov, in peer-reviewed journals and through stakeholder meetings. TRIAL REGISTRATION NUMBER: NCT05567016.

Safety and efficacy of malaria vaccine candidate R21/Matrix-M in African children: a multicentre, double-blind, randomised, phase 3 trial.

BACKGROUND: Recently, we found that a new malaria vaccine, R21/Matrix-M, had over 75% efficacy against clinical malaria with seasonal administration in a phase 2b trial in Burkina Faso. Here, we report on safety and efficacy of the vaccine in a phase 3 trial enrolling over 4800 children across four countries followed for up to 18 months at seasonal sites and 12 months at standard sites. METHODS: We did a double-blind, randomised, phase 3 trial of the R21/Matrix-M malaria vaccine across five sites in four African countries with differing malaria transmission intensities and seasonality. Children (aged 5-36 months) were enrolled and randomly assigned (2:1) to receive 5 µg R21 plus 50 µg Matrix-M or a control vaccine (licensed rabies vaccine [Abhayrab]). Participants, their families, investigators, laboratory teams, and the local study team were masked to treatment. Vaccines were administered as three doses, 4 weeks apart, with a booster administered 12 months after the third dose. Half of the children were recruited at two sites with seasonal malaria transmission and the remainder at standard sites with perennial malaria transmission using age-based immunisation. The primary objective was protective efficacy of R21/Matrix-M from 14 days after third vaccination to 12 months after completion of the primary series at seasonal and standard sites separately as co-primary endpoints. Vaccine efficacy against multiple malaria episodes and severe malaria, as well as safety and immunogenicity, were also assessed. This trial is registered on ClinicalTrials.gov, NCT04704830, and is ongoing. FINDINGS: From April 26, 2021, to Jan 12, 2022, 5477 children consented to be screened, of whom 1705 were randomly assigned to control vaccine and 3434 to R21/Matrix-M; 4878 participants received the first dose of vaccine. 3103 participants in the R21/Matrix-M group and 1541 participants in the control group were included in the modified per-protocol analysis (2412 [51·9%] male and 2232 [48·1%] female). R21/Matrix-M vaccine was well tolerated, with injection site pain (301 [18·6%] of 1615 participants) and fever (754 [46·7%] of 1615 participants) as the most frequent adverse events. Number of adverse events of special interest and serious adverse events did not significantly differ between the vaccine groups. There were no treatment-related deaths. 12-month vaccine efficacy was 75% (95% CI 71-79; p<0·0001) at the seasonal sites and 68% (61-74; p<0·0001) at the standard sites for time to first clinical malaria episode. Similarly, vaccine efficacy against multiple clinical malaria episodes was 75% (71-78; p<0·0001) at the seasonal sites and 67% (59-73; p<0·0001) at standard sites. A modest reduction in vaccine efficacy was observed over the first 12 months of follow-up, of similar size at seasonal and standard sites. A rate reduction of 868 (95% CI 762-974) cases per 1000 children-years at seasonal sites and 296 (231-362) at standard sites occurred over 12 months. Vaccine-induced antibodies against the conserved central Asn-Ala-Asn-Pro (NANP) repeat sequence of circumsporozoite protein correlated with vaccine efficacy. Higher NANP-specific antibody titres were observed in the 5-17 month age group compared with 18-36 month age group, and the younger age group had the highest 12-month vaccine efficacy on time to first clinical malaria episode at seasonal (79% [95% CI 73-84]; p<0·001) and standard (75% [65-83]; p<0·001) sites. INTERPRETATION: R21/Matrix-M was well tolerated and offered high efficacy against clinical malaria in African children. This low-cost, high-efficacy vaccine is already licensed by several African countries, and recently received a WHO policy recommendation and prequalification, offering large-scale supply to help reduce the great burden of malaria in sub-Saharan Africa. FUNDING: The Serum Institute of India, the Wellcome Trust, the UK National Institute for Health Research Oxford Biomedical Research Centre, and Open Philanthropy.

Safety and Immunogenicity of Radiation-Attenuated PfSPZ Vaccine in Equatoguinean Infants, Children, and Adults.

The radiation-attenuated Plasmodium falciparum sporozoites (PfSPZ) Vaccine has demonstrated safety and immunogenicity in 5-month-old to 50-year-old Africans in multiple trials. Except for one, each trial has restricted enrollment to either infants and children or adults < 50 years old. This trial was conducted in Equatorial Guinea and assessed the safety, tolerability, and immunogenicity of three direct venous inoculations of 1.8 × 106 or 2.7 × 106 PfSPZ, of PfSPZ Vaccine, or normal saline administered at 8-week intervals in a randomized, double-blind, placebo-controlled trial stratified by age (6-11 months and 1-5, 6-10, 11-17, 18-35, and 36-61 years). All doses were successfully administered. In all, 192/207 injections (93%) in those aged 6-61 years were rated as causing no or mild pain. There were no significant differences in solicited adverse events (AEs) between vaccinees and controls in any age group (P ≥ 0.17). There were no significant differences between vaccinees and controls with respect to the rates or severity of unsolicited AEs or laboratory abnormalities. Development of antibodies to P. falciparum circumsporozoite protein occurred in 67/69 vaccinees (97%) and 0/15 controls. Median antibody levels were highest in infants and 1-5-year-olds and declined progressively with age. Antibody responses in children were greater than in adults protected against controlled human malaria infection. Robust immunogenicity, combined with a benign AE profile, indicates children are an ideal target for immunization with PfSPZ Vaccine.

Role of human Pegivirus infections in whole Plasmodium falciparum sporozoite vaccination and controlled human malaria infection in African volunteers.

BACKGROUND: Diverse vaccination outcomes and protection levels among different populations pose a serious challenge to the development of an effective malaria vaccine. Co-infections are among many factors associated with immune dysfunction and sub-optimal vaccination outcomes. Chronic, asymptomatic viral infections can contribute to the modulation of vaccine efficacy through various mechanisms. Human Pegivirus-1 (HPgV-1) persists in immune cells thereby potentially modulating immune responses. We investigated whether Pegivirus infection influences vaccine-induced responses and protection in African volunteers undergoing whole P. falciparum sporozoites-based malaria vaccination and controlled human malaria infections (CHMI). METHODS: HPgV-1 prevalence was quantified by RT-qPCR in plasma samples of 96 individuals before, post vaccination with PfSPZ Vaccine and after CHMI in cohorts from Tanzania and Equatorial Guinea. The impact of HPgV-1 infection was evaluated on (1) systemic cytokine and chemokine levels measured by Luminex, (2) PfCSP-specific antibody titers quantified by ELISA, (3) asexual blood-stage parasitemia pre-patent periods and parasite multiplication rates, (4) HPgV-1 RNA levels upon asexual blood-stage parasitemia induced by CHMI. RESULTS: The prevalence of HPgV-1 was 29.2% (28/96) and sequence analysis of the 5' UTR and E2 regions revealed the predominance of genotypes 1, 2 and 5. HPgV-1 infection was associated with elevated systemic levels of IL-2 and IL-17A. Comparable vaccine-induced anti-PfCSP antibody titers, asexual blood-stage multiplication rates and pre-patent periods were observed in HPgV-1 positive and negative individuals. However, a tendency for higher protection levels was detected in the HPgV-1 positive group (62.5%) compared to the negative one (51.6%) following CHMI. HPgV-1 viremia levels were not significantly altered after CHMI. CONCLUSIONS: HPgV-1 infection did not alter PfSPZ Vaccine elicited levels of PfCSP-specific antibody responses and parasite multiplication rates. Ongoing HPgV-1 infection appears to improve to some degree protection against CHMI in PfSPZ-vaccinated individuals. This is likely through modulation of immune system activation and systemic cytokines as higher levels of IL-2 and IL17A were observed in HPgV-1 infected individuals. CHMI is safe and well tolerated in HPgV-1 infected individuals. Identification of cell types and mechanisms of both silent and productive infection in individuals will help to unravel the biology of this widely present but largely under-researched virus.

Immunogenicity and Protective Efficacy of Radiation-Attenuated and Chemo-Attenuated PfSPZ Vaccines in Equatoguinean Adults.

Plasmodium falciparum sporozoite (PfSPZ) Vaccine (radiation-attenuated, aseptic, purified, cryopreserved PfSPZ) and PfSPZ-CVac (infectious, aseptic, purified, cryopreserved PfSPZ administered to subjects taking weekly chloroquine chemoprophylaxis) have shown vaccine efficacies (VEs) of 100% against homologous controlled human malaria infection (CHMI) in nonimmune adults. Plasmodium falciparum sporozoite-CVac has never been assessed against CHMI in African vaccinees. We assessed the safety, immunogenicity, and VE against homologous CHMI of three doses of 2.7 × 106 PfSPZ of PfSPZ Vaccine at 8-week intervals and three doses of 1.0 × 105 PfSPZ of PfSPZ-CVac at 4-week intervals with each arm randomized, double-blind, placebo-controlled, and conducted in parallel. There were no differences in solicited adverse events between vaccinees and normal saline controls, or between PfSPZ Vaccine and PfSPZ-CVac recipients during the 6 days after administration of investigational product. However, from days 7-13, PfSPZ-CVac recipients had significantly more AEs, probably because of Pf parasitemia. Antibody responses were 2.9 times higher in PfSPZ Vaccine recipients than PfSPZ-CVac recipients at time of CHMI. Vaccine efficacy at a median of 14 weeks after last PfSPZ-CVac dose was 55% (8 of 13, P = 0.051) and at a median of 15 weeks after last PfSPZ Vaccine dose was 27% (5 of 15, P = 0.32). The higher VE in PfSPZ-CVac recipients of 55% with a 27-fold lower dose was likely a result of later stage parasite maturation in the liver, leading to induction of cellular immunity against a greater quantity and broader array of antigens.

Providing Ancillary Care in Clinical Research: A Case of Diffuse Large B-Cell Lymphoma during a Malaria Vaccine Trial in Equatorial Guinea.

Providing medical care for participants in clinical trials in resource-limited settings can be challenging and costly. Evaluation and treatment of a young man who developed cervical lymphadenopathy during a malaria vaccine trial in Equatorial Guinea required concerted efforts of a multinational, multidisciplinary team. Once a diagnosis of diffuse large B-cell lymphoma was made, the patient was taken to India to receive immunochemotherapy. This case demonstrates how high-quality medical care was provided for a serious illness that occurred during a trial that was conducted in a setting in which positron emission tomography for diagnostic staging, an oncologist for supervision of treatment, and an optimal therapeutic intervention were not available. Clinical researchers should anticipate the occurrence of medical conditions among study subjects, clearly delineate the extent to which health care will be provided, and set aside funds commensurate with those commitments.

The Equatoguinean Malaria Vaccine Initiative: From the Launching of a Clinical Research Platform to Malaria Elimination Planning in Central West Africa.

Fifteen years of investment in malaria control on Bioko Island, Equatorial Guinea (EG), dramatically reduced malaria-associated morbidity and mortality, but the impact has plateaued. To progress toward elimination, EG is investing in the development of a malaria vaccine. We assessed the unique public-private partnership that has had such a significant impact on malaria on Bioko Island and now added a major effort on malaria vaccine development. As part of a $79M commitment, the EG government (75%) and three American energy companies (25%) have invested since 2012 greater than $55M in the Equatoguinean Malaria Vaccine Initiative (EGMVI) to support clinical development of Sanaria® PfSPZ vaccines (Sanaria Inc., Rockville, MD). In turn, the vaccine development program is building human capital and physical capacity. The EGMVI established regulatory and ethical oversight to ensure compliance with the International Conference on Harmonization and Good Clinical Practices for the first importation of investigational product, ethical approval, and conduct of a clinical trial in Equatoguinean history. The EGMVI has completed three vaccine trials in EG, two vaccine trials in Tanzania, and a malaria incidence study, and initiated preparations for a 2,100-volunteer clinical trial. Personnel are training for advanced degrees abroad and have been trained in Good Clinical Practices and protocol-specific methods. A new facility has established the foundation for a national research institute. Biomedical research and development within this visionary, ambitious public-private partnership is fostering major improvements in EG. The EGMVI plans to use a PfSPZ Vaccine alongside standard malaria control interventions to eliminate Pf malaria from Bioko, becoming a potential model for elimination campaigns elsewhere.

Whole blood transcriptome changes following controlled human malaria infection in malaria pre-exposed volunteers correlate with parasite prepatent period.

Malaria continues to be one of mankind's most devastating diseases despite the many and varied efforts to combat it. Indispensable for malaria elimination and eventual eradication is the development of effective vaccines. Controlled human malaria infection (CHMI) is an invaluable tool for vaccine efficacy assessment and investigation of early immunological and molecular responses against Plasmodium falciparum infection. Here, we investigated gene expression changes following CHMI using RNA-Seq. Peripheral blood samples were collected in Bagamoyo, Tanzania, from ten adults who were injected intradermally (ID) with 2.5x104 aseptic, purified, cryopreserved P. falciparum sporozoites (Sanaria® PfSPZ Challenge). A total of 2,758 genes were identified as differentially expressed following CHMI. Transcriptional changes were most pronounced on day 5 after inoculation, during the clinically silent liver phase. A secondary analysis, grouping the volunteers according to their prepatent period duration, identified 265 genes whose expression levels were linked to time of blood stage parasitemia detection. Gene modules associated with these 265 genes were linked to regulation of transcription, cell cycle, phosphatidylinositol signaling and erythrocyte development. Our study showed that in malaria pre-exposed volunteers, parasite prepatent period in each individual is linked to magnitude and timing of early gene expression changes after ID CHMI.

Advancing Global Health through Development and Clinical Trials Partnerships: A Randomized, Placebo-Controlled, Double-Blind Assessment of Safety, Tolerability, and Immunogenicity of PfSPZ Vaccine for Malaria in Healthy Equatoguinean Men.

Equatorial Guinea (EG) has implemented a successful malaria control program on Bioko Island. A highly effective vaccine would be an ideal complement to this effort and could lead to halting transmission and eliminating malaria. Sanaria® PfSPZ Vaccine (Plasmodium falciparum sporozoite Vaccine) is being developed for this purpose. To begin the process of establishing the efficacy of and implementing a PfSPZ Vaccine mass vaccination program in EG, we decided to conduct a series of clinical trials of PfSPZ Vaccine on Bioko Island. Because no clinical trial had ever been conducted in EG, we first successfully established the ethical, regulatory, quality, and clinical foundation for conducting trials. We now report the safety, tolerability, and immunogenicity results of the first clinical trial in the history of the country. Thirty adult males were randomized in the ratio 2:1 to receive three doses of 2.7 × 105 PfSPZ of PfSPZ Vaccine (N = 20) or normal saline placebo (N = 10) by direct venous inoculation at 8-week intervals. The vaccine was safe and well tolerated. Seventy percent, 65%, and 45% of vaccinees developed antibodies to Plasmodium falciparum (Pf) circumsporozoite protein (PfCSP) by enzyme-linked immunosorbent assay, PfSPZ by automated immunofluorescence assay, and PfSPZ by inhibition of sporozoite invasion assay, respectively. Antibody responses were significantly lower than responses in U.S. adults who received the same dosage regimen, but not significantly different than responses in young adult Malians. Based on these results, a clinical trial enrolling 135 subjects aged 6 months to 65 years has been initiated in EG; it includes PfSPZ Vaccine and first assessment in Africa of PfSPZ-CVac. ClinicalTrials.gov identifier: NCT02418962.

Controlled human malaria infection of Tanzanians by intradermal injection of aseptic, purified, cryopreserved Plasmodium falciparum sporozoites.

Controlled human malaria infection (CHMI) by mosquito bite has been used to assess anti-malaria interventions in > 1,500 volunteers since development of methods for infecting mosquitoes by feeding on Plasmodium falciparum (Pf) gametocyte cultures. Such CHMIs have never been used in Africa. Aseptic, purified, cryopreserved Pf sporozoites, PfSPZ Challenge, were used to infect Dutch volunteers by intradermal injection. We conducted a double-blind, placebo-controlled trial to assess safety and infectivity of PfSPZ Challenge in adult male Tanzanians. Volunteers were injected intradermally with 10,000 (N = 12) or 25,000 (N = 12) PfSPZ or normal saline (N = 6). PfSPZ Challenge was well tolerated and safe. Eleven of 12 and 10 of 11 subjects, who received 10,000 and 25,000 PfSPZ respectively, developed parasitemia. In 10,000 versus 25,000 PfSPZ groups geometric mean days from injection to Pf positivity by thick blood film was 15.4 versus 13.5 (P = 0.023). Alpha-thalassemia heterozygosity had no apparent effect on infectivity. PfSPZ Challenge was safe, well tolerated, and infectious.

Comparison of bioavailability between the most available generic tablet formulation containing artemether and lumefantrine on the Tanzanian market and the innovator's product.

BACKGROUND: Existence of anti-malarial generic drugs with low bioavailability marketed on sub-Saharan Africa raises a concern on patients achieving therapeutic concentrations after intake of such products. This work compared bioavailability of one generic tablet formulation with innovator's product. Both were fixed dose combination tablet formulations containing artemether and lumefantrine. METHODOLOGY: The study was conducted in Dar Es Salaam, Tanzania, in which a survey of the most abundant generic containing artemether-lumefantrine tablet formulation was carried out in retail pharmacies. The most widely available generic (Artefan®, Ajanta Pharma Ltd, Maharashtra, India) was sampled for bioavailability comparison with Coartem® (Novartis Pharma, Basel, Switzerland)--the innovator's product. A randomized, two-treatment cross-over study was conducted in 18 healthy Tanzanian black male volunteers. Each volunteer received Artefan® (test) and Coartem® (as reference) formulation separated by 42 days of drug-free washout period. Serial blood samples were collected up to 168 hours after oral administration of a single dose of each treatment. Quantitation of lumefantrine plasma levels was done using HPLC with UV detection. Bioequivalence of the two products was assessed in accordance with the US Food and Drug Authority (FDA) guidelines. RESULTS: The most widely available generic in pharmacies was Artefan® from India. All eighteen enrolled volunteers completed the study and both test and reference tablet formulations were well tolerated. It was possible to quantify lumefantrine alone, therefore, the pharmacokinetic parameters reported herein are for lumefantrine. The geometric mean ratios for Cmax, AUC0-t and AUC0-∞ were 84% in all cases and within FDA recommended bioequivalence limits of 80%-125%, but the 90% confidence intervals were outside FDA recommended limits (CI 49-143%, 53-137%, 52-135% respectively). There were no statistical significant differences between the two formulations with regard to PK parameters (P > 0.05). CONCLUSIONS: Although the ratios of AUCs and Cmax were within the acceptable FDA range, bioequivalence between Artefan® and Coartem® tablet formulations was not demonstrated due to failure to comply with the FDA 90% confidence interval criteria. Based on the observed total drug exposure (AUCs), Artefan® is likely to produce a similar therapeutic response as Coartem®.

Evaluating the effectiveness of IPTi on malaria using routine health information from sentinel health centres in southern Tanzania.

BACKGROUND: Intermittent preventive treatment of malaria in infants (IPTi) consists of the administration of a treatment dose of sulphadoxine-pyrimethamine (SP) at the time of routine vaccinations. The use of routine Health Management and Information Services (HMIS) data to investigate the effect of IPTi on malaria, anaemia, and all-cause attendance in children aged 2-11 months presenting to 11 health centres in southern Tanzania is described. METHODS: Clinical diagnosis of malaria was confirmed with a positive blood slide reading from a quality assurance laboratory. Anaemia was defined using two thresholds (mild [Hb<11 g/dL], severe [Hb<8 g/dL]). Incidence rates between IPTi and non-implementing health centres were calculated using Poisson regression, and all statistical testing was based on the t test due to the clustered nature of the data. RESULTS: Seventy two per cent of infants presenting in intervention areas received at least one dose of IPTi--22% received all three. During March 2006-April 2007, the incidence of all cause attendance was two attendances per person, per year (pppy), including 0.2 episodes pppy of malaria, 0.7 episodes of mild and 0.13 episodes of severe anaemia. Point estimates for the effect of IPTi on malaria varied between 18% and 52%, depending on the scope of the analysis, although adjustment for clustering rendered these not statistically significant. CONCLUSIONS: The point estimate of the effect of IPTi on malaria is consistent with that from a large pooled analysis of randomized control trials. As such, it is plausible that the difference seen in health centre data is due to IPTi, even thought the effect did not reach statistical significance. Findings draw attention to the challenges of robust inference of effects of interventions based on routine health centre data. Analysis of routine health information can reassure that interventions are being made available and having desired effects, but unanticipated effects should trigger data collection from representative samples of the target population.

Safety of sulfadoxine/pyrimethamine for intermittent preventive treatment of malaria in infants: evidence from large-scale operational research in southern Tanzania.

Intermittent preventive treatment with sulfadoxine/pyrimethamine (SP) is recommended for malaria prevention in infants (IPTi-SP). Serious adverse events, including Stevens-Johnson syndrome (SJS), have been reported following exposure to SP, but few infant-specific data exist. The safety of IPTi-SP was evaluated as part of a pilot implementation programme in southern Tanzania using three methods: spontaneous adverse event reporting to capture suspected adverse drug reactions (ADR); a census survey documenting rash-related hospital admissions among children < 2 years of age; and verbal autopsies (VA) completed for rash-related deaths in 2-11-month-olds. Approximately 82 000 IPTi-SP doses were administered to approximately 29 000 children. In total, 119 suspected ADRs were reported, 13 in children aged <2 years, only one of whom had received IPTi-SP. The census involved 243 612 households. Only one rash-related admission was reported amongst 1292 children aged 2-11 months, but this child had no history of exposure to SP. Moreover, 30 of 699 deaths in 2-11-month-olds were said to have been associated with a skin rash. The rates of rash-associated death were 0.59/1000 person-years at risk (PYAR) and 1.17/1000 PYAR in intervention and comparison areas, respectively (P = 0.79). VAs did not suggest SJS or any other ADR. We conclude that IPTi-SP is associated with a very low incidence of severe skin reactions. [ClinicalTrials.gov identifier: NCT00152204].