Analysis of the dose-dependent stage-specific in vitro efficacy of a multi-stage malaria vaccine candidate cocktail

Abstract

BackgroundThe high incidence and mortality rate of malaria remains a serious burden for many developing countries, and a vaccine that induces durable and highly effective immune responses is, therefore, desirable. An earlier analysis of the stage-specific in vitro efficacy of a malaria vaccine candidate cocktail (VAMAX) considered the general properties of complex multi-component, multi-stage combination vaccines in rabbit immunization experiments using a hyper-immunization protocol featuring six consecutive boosts and a strong, lipopolysaccharide-based adjuvant. This follow-up study investigates the effect of antigen dose on the in vitro efficacy of the malaria vaccine cocktail using a conventional vaccination scheme (one prime and two boosts) and a human-compatible adjuvant (Alhydrogel®).ResultsIgG purified from rabbits immunized with 0.1, 1, 10 or 50 µg doses of the VAMAX vaccine candidate cocktail was analysed for total IgG and antigen-cocktail-specific titers. An increase in cocktail-specific titers was observed between 0.1 and 1 µg and between 10 and 50 µg, whereas no significant difference in titers was observed between 1 and 10 µg. Antigen component-specific antibody titers and stage-specific in vitro efficacy assays were performed with pooled IgG from animals immunized with 1 and 50 µg of the VAMAX cocktail. Here, the component-specific antibody levels showed clear dose dependency whereas the determined stage-specific in vitro IC50 values (as a correlate of efficacy) were only dependent on the titer amounts of stage-specific antibodies.ConclusionsThe stage-specific in vitro efficacy of the VAMAX cocktail strongly correlates with the corresponding antigen-specific titers, which for their part depend on the antigen dose, but there is no indication that the dose has an effect on the in vitro efficacy of the induced antibodies. A comparison of these results with those obtained in the previous hyper-immunization study (where higher levels of antigen-specific IgG were observed) suggests that there is a significant need to induce an immune response matching efficacy requirements, especially for a PfAMA1-based blood stage vaccine, by using higher doses, better adjuvants and/or better formulations.

A polyclonal antibody against a recombinantly expressed Triticum aestivum RHT-D1A protein

“…The usual dose of a soluble protein administered with Freund's adjuvant to rabbits is in the range of 50 to 1000 μg, and for mice, it is 10-200 μg; for goats or sheep, the typical dose is 250-5000 μg [28]. Nevertheless, for primary injection, most investigators use doses 100-200 μg [29,30] or even less than 25-50 μg [31,32]. It should be stressed that in some studies concerning the polyclonal antibody production in rabbits, for primary immunization, researchers have used doses of 400 to 500 μg [33,34] even up to 1.0 mg [35,36].…”

Functional antibodies against Plasmodium falciparum sporozoites are associated with a longer time to qPCR-detected infection among schoolchildren in Burkina Faso

“…A significant challenge in these studies is that both protective immunity and cumulative exposure increase with age, and so it is often unclear whether measured responses mediate protection or are merely a marker of past exposure 26, 27 . In an attempt to move beyond indirect epidemiological associations, we explored functional anti-sporozoite immunity by assessing the ability of plasma to inhibit sporozoite gliding motility 28 and hepatocyte invasion 29 and related these in vitro phenotypes to field findings. We observed that CSP IgG antibodies showed a strong positive association with sporozoite gliding motility inhibition.…”

“…Subsequently, wells were washed thrice with 100 μl/well PBS and stored in 150 μl/well PBS at 4°C in the dark until analysis. Gliding trails were imaged automatically with the BioTek Cytation cell imager (25 images per well at 200x magnification) and images were analysed automatically by FIJI software (under ImageJ version 2.0.0-rc-68/1.52h) with Otsu’s thresholding 28 . Results were plotted in GraphPad Prism version 5.03.The number of pixels present on a stitched image made from 25 individual pictures taken per well is a measure of the amount of shed CSP in that particular well and therefore, differences in the number of pixels can be interpreted as differences in sporozoite gliding trail surface 28 .…”

Antimalarial pantothenamide metabolites target acetyl-CoA synthesis inPlasmodium falciparum

“…From day 4 to day 9 after inoculation, cultures were treated with 50 mM N-acetylglucosamine to eliminate asexual bloodstage parasites. At day 11 post inoculation, gametocytes (predominantly stage IV) were isolated by Percoll density gradient centrifugation as described previously (30 liver stages in human primary hepatocytes were analyzed essentially as described before (31).…”

“…Briefly, 50.000 human primary hepatocytes were seeded in collagen-coated 96 plates according to the supplier's protocol (Tebu-bio) and combined with compounds serially diluted in DMSO and culture medium to achieve a final DMSO concentration of 0.1%. Hepatocytes were infected with 50,000 P. falciparum NF54 sporozoites per well and developing liver schizonts were visualized 4 days after infection by a DAPI nuclear stain and αhsp70 immunostaining as described before (31).…”