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Two hookworm vaccine candidates, Na-GST-1 and Na-APR-1, formulated with Glucopyranosyl Lipid A (GLA-AF) adjuvant, have been shown to be safe, well tolerated, and to induce antibody responses in a Phase 1 clinical trial (Clinicaltrials.gov NCT02126462) conducted in Gabon. Here, we characterized T cell responses in 24 Gabonese volunteers randomized to get vaccinated three times with Na-GST-1 and Na-APR-1 at doses of 30μg (n = 8) or 100μg (n = 10) and as control Hepatitis B (n = 6). Blood was collected pre- and post-vaccination on days 0, 28, and 180 as well as 2-weeks after each vaccine dose on days 14, 42, and 194 for PBMCs isolation. PBMCs were stimulated with recombinant Na-GST-1 or Na-APR-1, before (days 0, 28 and 180) and two weeks after (days 14, 42 and 194) each vaccination and used to characterize T cell responses by flow and mass cytometry. A significant increase in Na-GST-1 -specific CD4+ T cells producing IL-2 and TNF, correlated with specific IgG antibody levels, after the third vaccination (day 194) was observed. In contrast, no increase in Na-APR-1 specific T cell responses were induced by the vaccine. Mass cytometry revealed that, Na-GST-1 cytokine producing CD4+ T cells were CD161+ memory cells expressing CTLA-4 and CD40-L. Blocking CTLA-4 enhanced the cytokine response to Na-GST-1. In Gabonese volunteers, hookworm vaccine candidate, Na-GST-1, induces detectable CD4+ T cell responses that correlate with specific antibody levels. As these CD4+ T cells express CTLA-4, and blocking this inhibitory molecules resulted in enhanced cytokine production, the question arises whether this pathway can be targeted to enhance vaccine immunogenicity., Author summary Two hookworm vaccine candidate (Na-GST-1 and Na-APR-1) have been tested in Gabonese and found to be safe and to induce antibody response. We aimed to study the cellular immune responses among vaccinated and unvaccinated volunteers. We found that Na-GST-1 induced CD4+ T cell responses (IL-2, TNF) among the vaccinated volunteers that received the high vaccine dose (100 ug). Furthermore Na-GST-1 specific memory T cells were found to express the inhibitory molecule CTLA-4. These responses was not observed in those who received the low dose of the Na-GST-1 vaccine, or those who received Na-APR-1 or HBV. By blocking CTLA-4, we observed an increase in TNF production. Our data suggest that an intervention involving blockage of the CTLA-4 molecule in the vaccinated could be beneficial in endemic settings where vaccine responses have been shown to be lower compared to non-endemic settings.

Background Hookworms cause substantial morbidity in children and women of reproductive age. The control strategy of mass drug administration is suboptimal, hence the need for a vaccine. Necator americanus aspartic protease-1 (Na-APR-1) and N americanus glutathione S-transferase-1 (Na-GST-1) are involved in the digestion and detoxification of haemoglobin in the hookworm digestive tract. In animal models, vaccination against these antigens resulted in protection from challenge infection. Both vaccine candidates were shown to be safe and well tolerated when administered separately to healthy adults. We assessed the safety and immunogenicity of co-administered Na-GST-1 and Na-APR-1 (M74) vaccines in healthy Gabonese adults.Methods This randomised, controlled, double-blind, phase 1, dose-escalation trial was done at the Centre de Recherches Medicales de Lambarene, in a region of Gabon where N americanus and other helminths are prevalent. Healthy adults aged 18-50 years and living in Lambarene or the surrounding areas were recruited to the study. Participants were enrolled consecutively into two dose cohorts (30 mu g or 100 mu g of the experimental vaccines) and randomly assigned in blocks (block size four) to receive three doses of either co-administered Na-GST-1 plus Na-APR-1 (M74; 30 mu g or 100 mu g of each), adjuvanted with Alhydrogel (aluminium hydroxide gel suspension) together with an aqueous formulation of glucopyranosyl lipid A, or hepatitis B vaccine plus saline (control group). Vaccines were administered intramuscularly on days 0, 28, and 180. The primary endpoint was safety, with immunogenicity a secondary endpoint. The intention-to-treat population was used for safety analyses, whereas for immunogenicity analyses, the per-protocol population was used (participants who received all scheduled vaccinations). Control vaccine recipients for both dose cohorts were combined for the analyses. The trial is registered with ClinicalTrials.gov, NCT02126462.Findings Between Oct 27, 2014, and Jan 31, 2015, 56 individuals were screened for eligibility, of whom 32 were enrolled and randomly assigned to one of the three study groups (12 each in the 30 mu g and 100 mu g experimental vaccine groups and eight in the control group). Both study vaccines were well tolerated in both dose groups. The most common adverse events were mild-to-moderate injection-site pain, headache, myalgia, and nausea. No severe or serious adverse events related to the vaccines were recorded. 52 unsolicited vaccine-related adverse events occurred during the study, but there was no difference in frequency between vaccine groups. IgG antibodies were induced to each of the vaccine antigens, with mean IgG levels increasing after each vaccination. Vaccination with 100 mu g of each vaccine antigen consistently induced IgG seroconversion (IgG levels above the reactivity threshold). Peak IgG responses were observed 2 weeks after the third vaccine dose for both antigens, with all participants who received the 100 mu g doses seroconverting at that timepoint. IgG levels steadily declined until the final study visit 6 months after the third vaccination, although they remained significantly higher than baseline in the 100 mu g dose group.Interpretation Vaccination with recombinant Na-GST-1 and Na-APR-1 (M74) in healthy adults living in N americanusendemic areas of Gabon was safe and induced IgG to each antigen. To our knowledge, this study is the first to report results of Na-APR-1 (M74) co-administered with Alhydrogel in participants from an N americanus-endemic area. Further clinical development of these vaccines should involve efficacy studies. Funding European Union Seventh Framework Programme. Copyright (C) 2020 Elsevier Ltd. All rights reserved.

Background Hookworm disease is one of the most prevalent of the neglected tropical diseases. To date, the control of hookworm infection has been limited to mass-administration of anthelminthic drugs. Despite this, the global hookworm prevalence does not decrease, thus there is a need for a vaccine. We evaluated the Na-GST-1 and Na-APR-1 hookworm vaccine candidates simultaneously in a hookworm endemic Gabonese population. Methods Eligible healthy Gabonese adults aged 18–50 years were enrolled in a randomised, double blind, controlled phase I trial. The first cohort received 30 µg Na-GST-1 co-administered with 30 µg Na-APR-1. The second cohort received 100 µg Na-GST-1 and 100 µg Na-APR-1. All doses were administered after mixing with 5 µg of an aqueous formulation of glucopyranosyl Lipid A (GLA-AF), a Toll-like Receptor-4 agonist. Hepatitis B vaccination (HBV) was administered as a comparator. Study subjects were vaccinated on days 0, 28 and 180 by intramuscular injection. IgG antibody levels were measured by qualified ELISA. This study evaluated the safety, reactogenicity, and immunogenicity of Na- GST-1/Alhydrogel® co-administered with Na-APR-1/Alhydrogel®. Results Thirty-two study participants were enrolled. No serious adverse events or significant changes in haematological, renal or liver function parameters were observed. Mild-to-moderate injection-site pain, headache and fever were common adverse events. Elevated Na-GST-1 and Na-APR-1 IgG antibody levels were detected on day 194. Significant differences in mean antibody levels were observed between dose groups for Na-APR-1 [30 µg: 18 (50–86.9) vs 100 µg: 197 (131– 264); p vs 100 µg: 402.54 (283.65–521); p=0.5]. Conclusions Co-administration of the hookworm vaccine candidates (Na-GST-1 and Na-APR-1) was safe and well tolerated. In order to achieve optimal antibody levels, a series of three high doses needs to be administered. Additional investigations are necessary to consider this combination as a potential bivalent vaccine candidate.

Background Characterising the effect of natural acquired immunity and sickle cell anaemia on the pattern of Plasmodium falciparum parasitaemia may be useful to understand the pathophysiological mechanisms of protection against malaria. Controlled human malaria infection (CHMI) by direct venous inoculation of aseptic, purified, cryopreserved sporozoites (Pf-SPZ challenge) is a new tool which can be used to investigate the pathophysiology of malaria. Methods The study was performed in Lambarene, Gabon, one of seven African partners in the EDCTP-funded CHMI platform. Adults aged 18–35 from three groups NI: 5 non-immune (NI), 11 semi-immunes with haemoglobin AA (IA), and 9 semi-immunes with haemoglobin AS (IS) received 3,200 sporozoites after a curative treatment course with clindamycin. Capillary blood samples were taken daily up to Day 28 to determine parasitaemia by real-time quantitative polymerase chain reaction (RT-qPCR). Treatment was administered for a malaria episode or at Day 28, whichever came first. Results Parasitaemia was detected in 5 (100%) subjects in the NI group, 9 (82%) in the IA group and 7 (78%) in the IS group. All volunteers in the NI group showed similar patterns with parasitaemia starting around Day 8 and rising quickly. Patterns for parasitaemia in the immune groups (IA and IS) were highly heterogeneous. Although time points of initial parasitaemia and duration of parasitaemia varied, all semi-immunes managed to control parasitaemia for at least several days. There were no discernible differences in patterns between the IS and IA group, including the area under curve of parasitaemia over time. Conclusions No parasitaemia was detected in 20% of the semi-immunes, likely due to liver stage immunity. The highly variable patterns of parasitaemia did not allow us to discern immune mechanisms against blood stages. Haemoglobin AS had no visible effect on parasite dynamics at the low parasitaemia encountered.

Background Malaria remains the first cause of death in Africa. In endemic area, it overlaps with other infections including helminths infections. It has been shown that there are interactions between the two parasites infection. Lambarene is the endemic area for urogenital schistosomiasis, which co-exist with P. falciparum malaria. Therefore, we decide to assess for the first time the effect of schistosomiasis infection on malaria infection burden. Methods In order to assess the effect of S. haematobium on malaria infection burden, a cross-sectional study was conducted in school children aged 6–16 years old. One blood smear was performed and 3 urine samples were obtained to assess the presence of infections. Chi-square test and generalised linear model were used to compare the risk to be infected by P. falciparum parasite and Mann-Whitney-Wilcoxon test to compare the parasitaemia of P. falciparum . Demographic data was also collected. Results A total of 741 children were included. The overall prevalence was 20% and 31% for P. falciparum microscopic carriage and S. haematobium infection, respectively. Co-infection of both was found in 65 (9%) participants. S. haematobium and P. falciparum are highly prevalent in PK compared to Bindo and Makouke areas. At univariable analysis, schistosomiasis-infected subjects have an odd of 2.11 [1.46–3.07] to be infected by P. falciparum parasite compared to non-infected subjects. Locality was found to confound the association which remains significant after adjustment for age, gender and locality (aOR=1.69, [1.13–2.59]). The effect of S. haematobium on the P. falciparum parasitaemia outcome was also assessed. There is no effect of Schistosoma infection on malaria parasite density (p-value=0.92). Conclusions S. haematobium infection increases the risk of being infected with P. falciparum but doesn9t affect the parasitaemia density of P. falciparum malaria in our study population.