Why functional pre-erythrocytic and bloodstage malaria vaccines fail: a meta-analysis of fully protective immunizations and novel immunological model

Abstract

Background: Clinically protective malaria vaccines consistently fail to protect adults and children in endemic settings, and at best only partially protect infants.

Methodology/principal findings: We identify and evaluate 1916 immunization studies between 1965-February 2010, and exclude partially or nonprotective results to find 177 completely protective immunization experiments. Detailed reexamination reveals an unexpectedly mundane basis for selective vaccine failure: live malaria parasites in the skin inhibit vaccine function. We next show published molecular and cellular data support a testable, novel model where parasite-host interactions in the skin induce malaria-specific regulatory T cells, and subvert early antigen-specific immunity to parasite-specific immunotolerance. This ensures infection and tolerance to reinfection. Exposure to Plasmodium-infected mosquito bites therefore systematically triggers immunosuppression of endemic vaccine-elicited responses. The extensive vaccine trial data solidly substantiate this model experimentally.

Conclusions/significance: We conclude skinstage-initiated immunosuppression, unassociated with bloodstage parasites, systematically blocks vaccine function in the field. Our model exposes novel molecular and procedural strategies to significantly and quickly increase protective efficacy in both pipeline and currently ineffective malaria vaccines, and forces fundamental reassessment of central precepts determining vaccine development. This has major implications for accelerated local eliminations of malaria, and significantly increases potential for eradication.

Figure 1. PRISMA Flow Diagram showing inclusion/exclusion criteria for studies documenting complete protection 1965-February 2010.

Figure 2. Protective vaccination physically bypasses skin at immunization or challenge (90%) or involves skin immunomodulation (10%).

A. Exposure to parasites in the skin coincides closely with vaccine failure. Green background- immunization procedures. Lilac background- challenge procedures and percent of total experiments showing complete protection (% total) formed by a subset of studies (category) using a given experimental procedure (categories a-i; supporting data in references listed below). Inclined syringe- administration route is intravenous (i.v.) for live parasites, or, the method does not involve live parasites, but uses dead parasites or purified antigen, antibody, or recombinant DNA (dead/Ag/Ab/subunit) and therefore bypasses parasite interactions with host skin. Multiple mosquitoes- live parasites administered by multiple simultaneous mosquito bites. Single mosquito- live parasites naturally transmitted by 4–15 bites. Mosquito in aromatic ring- live parasites administered by 12–15 bites/session with chloroquine. Vertical syringe- live parasites delivered subcutaneously (s.c) or intradermally (i.d.) or intramuscularly (i.m.); (i), uncontrolled exposure to endemic mosquitos. B. Protective immunization physically bypasses the skin at either immunization or challenge in 90% of cases (a,b,c,e,h). Protective immunization which transits skin during immunization (c,d,e,f,g,h) either: bypasses the skin physically at challenge (c,e,h); or, involves skin immunomodulation during immunization (d,g, 10% of cases). Within A and B: Red lines and numbers- experiment bypasses parasite-skin interactions at stage indicated by red lines; black lines and numbers- parasites interact with skin at stage indicated by black lines. Asterisk (*)- immunization via unmodified skin, limited to less-virulent P. berghei (f). Skin bypass- method physically avoids live parasite interactions in the host skin. Malaria exposure- skin exposure to infected mosquito bite before first immunization; naïve- no pre-exposure; exposed (endemic)- chronic exposure. x- this study shows complete protection of 40 of 41 mice challenged. y: one person in one study was infected one time via the skin prior to protective immunization and was therefore moderately tolerized. Data pertaining to experimental categories (a–h): a:, , , , , , , , , , , , , , , , , , , , , , , , , , , , , , , , , , , , , , , , , , , , , , , , , , , , , , , , , , , , , , , , , , , , , , , , , , , , , , , , , , , , , , , , , , , , , , , , , , , , , , , , , , , , , , , , , , , , , , , , , , , , , , , , , , , , . b: , , , , , , , , , . c: . d: , , , , , , , , , , , , , , , . e: , , . f: . g: . h: , , , , . Studies containing data for multiple relevant experimental conditions are referenced accordingly in each appropriate category. Multiple experiments contributed by a single study are indicated beneath study reference number (eg. reference 124 X2) in Supplementary Table S1. (Meta-analysis data extended reference list).

Figure 3. All protective immunization circumvents initial malaria-specific Treg activation in the skin: a functional model.

A: Natural transmission (mosquito bite) allows skinstage parasites (sporozoites, spz, green fragments) to migrate (dotted gray arrows) through skin cells to both lymph nodes (LN) and liver (via blood vessels, bv), and induce malaria-specific regulatory T cells in the skin and LN that suppress (red blocker lines) local and systemic protective immune responses (green arrows), resulting in bloodstage infections (gray fragments below liver), and amplification cycles within red blood cells (rbc). B, C, D: Protective immunity develops where immunization and/or challenge avoids parasite-skin interaction and Treg activation/induction; B: Physical bypass of the skin, (by intravenous (i.v.) attenuated parasites, purified antigen, antibody or intramuscular or intranasal subunit vaccine) at immunization, or i.v. challenge with unattenuated infective parasites, avoids all induction or activation of skin and liverstage-specific Tregs. C: Chloroquine (CQ) accumulates and blocks all MHC II and rapid, (but not classical, slow) MHC I malaria-antigen presentation in the skin, which would otherwise immediately induce/activate antigen-specific Tregs. D: Multiple simultaneous mosquito bites create a strongly pro-inflammatory local skin and/or systemic milieu which inhibit Treg activation and induction processes. LN Inset: Inside LN, metamorphosing skinstage sporozoite parasites (spz) are in close contact with, or invade, host antigen presenting dendritic cells (DC).