Modest heterologous protection after Plasmodium falciparum sporozoite immunization: a double-blind randomized controlled clinical trial

Abstract

Background: A highly efficacious vaccine is needed for malaria control and eradication. Immunization with Plasmodium falciparum NF54 parasites under chemoprophylaxis (chemoprophylaxis and sporozoite (CPS)-immunization) induces the most efficient long-lasting protection against a homologous parasite. However, parasite genetic diversity is a major hurdle for protection against heterologous strains.

Methods: We conducted a double-blind, randomized controlled trial in 39 healthy participants of NF54-CPS immunization by bites of 45 NF54-infected (n = 24 volunteers) or uninfected mosquitoes (placebo; n = 15 volunteers) against a controlled human malaria infection with the homologous NF54 or the genetically distinct NF135.C10 and NF166.C8 clones. Cellular and humoral immune assays were performed as well as genetic characterization of the parasite clones.

Results: NF54-CPS immunization induced complete protection in 5/5 volunteers against NF54 challenge infection at 14 weeks post-immunization, but sterilely protected only 2/10 and 1/9 volunteers against NF135.C10 and NF166.C8 challenge infection, respectively. Post-immunization plasma showed a significantly lower capacity to block heterologous parasite development in primary human hepatocytes compared to NF54. Whole genome sequencing showed that NF135.C10 and NF166.C8 have amino acid changes in multiple antigens targeted by CPS-induced antibodies. Volunteers protected against heterologous challenge were among the stronger immune responders to in vitro parasite stimulation.

Conclusions: Although highly protective against homologous parasites, NF54-CPS-induced immunity is less effective against heterologous parasite clones both in vivo and in vitro. Our data indicate that whole sporozoite-based vaccine approaches require more potent immune responses for heterologous protection.

Trial registration: This trial is registered in clinicaltrials.gov, under identifier NCT02098590 .

Keywords: Controlled human malaria infection; Heterologous protection; Immune responses; Malaria; Plasmodium falciparum; Sporozoite; Vaccine.

Fig. 1

Clinical trial profile

Fig. 2

Parasitemia following homologous and heterologous challenge infection. The percentage of volunteers remaining qPCR negative (Kaplan–Meier survival proportions) after challenge infection with a homologous NF54 (n = 5 immunized; n = 5 controls) (a) or heterologous NF135.C10 (n = 10 immunized; n = 5 controls) (b) or NF166.C8 (n = 9 immunized; n = 5 controls) (c) strain is shown. Solid lines represent CPS-immunized volunteers and dotted lines represent placebo control-immunized volunteers. ** P < 0.01 as determined by Log-rank (Mantel Cox) test

Fig. 3

Neutralizing effect of CPS-induced antibodies on in vitro sporozoite functionality of homologous and heterologous P. falciparum strains. (a) The number of primary human hepatocytes infected by homologous NF54 sporozoites in the presence of pre- or post-immunization plasma in all (n = 24) CPS-immunized volunteers was determined by microscopy. (b) P. falciparum NF54, NF135.C10, or NF166.C8 sporozoites were pre-incubated with pre- or post-immunization plasma from CPS-immunized volunteers and the percent inhibition of intra-hepatic development of NF54, NF135.C10, or NF166.C8 was calculated for post- compared to pre-immunization plasma for each individual volunteer and presented as squares (NF135.C10), triangles (NF166.C8), or circles (NF54). Data are shown as the mean of triplicate measurements for each individual volunteer (a) or the median of all data points with an interquartile range (b). Differences in the percent inhibition of intra-hepatic development between parasite strains were tested using one-way ANOVA with Bonferroni’s multiple comparison correction

Fig. 4

Whole-genome sequencing shows genetic variations between study strains. Phylogenetic positions of the three P. falciparum strains (NF54, NF135.C10 and NF166.C8) used in the study relative to other known P. falciparum strains. Whole-genome sequencing was used to infer relatedness to P. falciparum strains from different areas [31]. Asian strains: THA Thailand (dark red), VIE Vietnam (light red), CAM Cambodia (orange); East African strains: KEN Kenya; West African strains: GUI Guinea (light green), GHA Ghana (dark green); NF strains and 7G8 (blue)

Fig. 5

Analysis of in vitro intra-hepatic sporozoite development inhibition by CPS-induced antibodies, cellular responses and protection status in vivo. CPS-induced antibody-mediated inhibition of in vitro challenge strain intra-hepatic development and cytotoxic and cytokine-producing T cell responses to NF54-infected erythrocytes are shown. The 10th and 90th percentile of each response in all (n = 19) CPS-immunized volunteers that received a heterologous challenge infection are shown as grey box-and-whisker plots. The green triangle represents 1 out of 9 CPS-immunized volunteers sterilely protected against NF166.C8 challenge infection, while the orange square and upside down triangle represent the 2 out of 10 CPS-immunized volunteers with sterile protection against NF135.C10 challenge infection