Increase of Dose Associated With Decrease in Protection Against Controlled Human Malaria Infection by PfSPZ Vaccine in Tanzanian Adults

Abstract

Background: A vaccine would be an ideal tool for reducing malaria's impact. PfSPZ Vaccine (radiation attenuated, aseptic, purified, cryopreserved Plasmodium falciparum [Pf] sporozoites [SPZ]) has been well tolerated and safe in >1526 malaria-naive and experienced 6-month to 65-year-olds in the United States, Europe, and Africa. When vaccine efficacy (VE) of 5 doses of 2.7 × 105 PfSPZ of PfSPZ Vaccine was assessed in adults against controlled human malaria infection (CHMI) in the United States and Tanzania and intense field transmission of heterogeneous Pf in Mali, Tanzanians had the lowest VE (20%).

Methods: To increase VE in Tanzania, we increased PfSPZ/dose (9 × 105 or 1.8 × 106) and decreased numbers of doses to 3 at 8-week intervals in a double blind, placebo-controlled trial.

Results: All 22 CHMIs in controls resulted in parasitemia by quantitative polymerase chain reaction. For the 9 × 105 PfSPZ group, VE was 100% (5/5) at 3 or 11 weeks (P < .000l, Barnard test, 2-tailed). For 1.8 × 106 PfSPZ, VE was 33% (2/6) at 7.5 weeks (P = .028). VE of dosage groups (100% vs 33%) was significantly different (P = .022). Volunteers underwent repeat CHMI at 37-40 weeks after last dose. 6/6 and 5/6 volunteers developed parasitemia, but time to first parasitemia was significantly longer than controls in the 9 × 105 PfSPZ group (10.89 vs 7.80 days) (P = .039), indicating a significant reduction in parasites in the liver. Antibody and T-cell responses were higher in the 1.8 × 106 PfSPZ group.

Conclusions: In Tanzania, increasing the dose from 2.7 × 105 to 9 × 105 PfSPZ increased VE from 20% to 100%, but increasing to 1.8 × 106 PfSPZ significantly reduced VE.

Clinical trials registration: NCT02613520.

Keywords: Plasmodium falciparum; PfSPZ; controlled human malaria infection; malaria; vaccine efficacy.

Figure 1.

Volunteer participation (CONSORT 2010 Diagram). Abbreviations: CONSORT, Consolidated Standards of Reporting Trials; CHMI, controlled human malaria infection; NS, normal saline; PfSPZ, Plasmodium falciparum sporozoite.

Figure 2.

Kaplan-Meier survival curves in immunized volunteers vs controls as assessed by qPCR. Kaplan-Meier curves in volunteers undergoing: A, CHMI 23 days after the last of 3 doses with 9.0 × 10⁵ PfSPZ (n = 3) vs NS and infectivity controls (n = 6); B, CHMI 79 days after the last of 3 doses with 9.0 × 10⁵ PfSPZ (n = 2) vs CHMI 52 days after the last of 3 doses of 1.8 × 10⁶ PfSPZ (n = 6) vs NS and infectivity controls (n = 6); C, volunteers undergoing a second CHMI 259 to 286 days after the last dose of 9.0 × 10⁵ PfSPZ (n = 6) or 1.8 × 10⁶ PfSPZ (n = 6) vs NS and infectivity controls (n = 10). Abbreviations: CHMI, controlled human malaria infection; NS, normal saline; PCR, polymerase chain reaction; PfSPZ, Plasmodium falciparum sporozoite.

Figure 3.

IgG antibodies to PfCSP by ELISA at time of first CHMI (CHMIs no. 1 and no. 2). A, IgG antibodies to PfSPZ by aIFA (C) and automated inhibition of PfSPZ invasion of hepatoma cells (aISI) (E) and at time of second CHMI (CHMI no.3) by ELISA (B), aIFA (D) and ISI (F) in subjects who received 9 × 10⁵ PfSPZ or 1.8 × 10⁶ PfSPZ doses of PfSPZ Vaccine. Filled circles (●) represent volunteers remaining uninfected after CHMI; open circles (○) represent volunteers infected after CHMI. For the PfCSP ELISA vaccinees were considered to have a positive antibody response if their net OD 1.0 and OD 1.0 ratio, calculated, respectively, by subtracting or dividing the OD 1.0 by the prevaccination OD 1.0, were ≥ 50 and ≥ 3.0. By these criteria, in the 9.0 × 10⁵ PfSPZ group, 5/6, 6/6, and 1/6 were positive 2 weeks after third dose, and before their first and second CHMIs. In the 1.8 × 10⁶ PfSPZ group 6/6, 6/6, and 5/6 were positive 2 weeks after third dose, and before their first and second CHMIs. No control volunteers were positive at any time point. For preimmunization anti-PfCSP levels, there were no significant differences between infected and noninfected vaccinees (Table S6). In the aIFA, volunteers with a net arbitrary fluorescence units (AFU) 2 × 10⁵ of ≥150 and a ratio of post to pre AFU 2 × 10⁵ of ≥ 3.0 were considered positive (Table S5). By these criteria, in the 9.0 × 10⁵ PfSPZ group, 5/6, 5/6, and 4/6 were positive 2 weeks after the third dose, and before their first and second CHMIs (Table S5). In the 1.8 × 10⁶ PfSPZ group, 5/6, 5/6, and 4/6 were positive 2 weeks after the third dose, and before their first and second CHMIs (Table S5). Antibodies to PfSPZ at time of first and second CHMIs by aIFA are shown in Figure 3C and 3DTable S5. At the time of the first CHMI the 5 uninfected (protected) subjects in the 9 × 10⁵ PfSPZ group had a lower median net AFU 2 × 10⁵ (421) than did the 2 uninfected (2710) subjects in the 1.8 × 10⁶ group and higher than the 4 infected (301) subjects in the 1.8 × 10⁶ group, but the differences were not significant (P = .38 and .73). At the time of the second CHMI the 6 infected subjects in the 9 × 10⁵ PfSPZ group had a lower median net AFU 2 × 10⁵ (361) than did the one uninfected (3567) subject in the 1.8 × 10⁶ group and higher than the 5 infected (209) subjects in the 1.8 × 10⁶ group (P = .57 and .43). For the aISI, volunteers with a net ISI reciprocal serum dilution for 80% inhibition of ≥10 and ratio of post to preimmune ISI reciprocal serum dilution for 80% inhibition of ≥ 3.0 were considered positive. By these criteria, in the 9.0 × 10⁵ PfSPZ group, 0/6, 2/6, and 2/6 were positive 2 weeks after the third dose and before their first and second CHMIs (Table S5). In the 1.8 × 10⁶ PfSPZ group, 2/6, 4/6, and 3/6 were positive 2 weeks after the third dose, and before their first and second CHMIs (Table S5). Antibodies to PfSPZ at time of first and second CHMIs by ISI are shown in Figure 3E and 3F and Table S5. At time of the first CHMI the 5 uninfected (protected) subjects in the 9 × 10⁵ PfSPZ group had a nonsignificant lower median net reciprocal serum dilution for 80% inhibition (22.57) than did the 2 uninfected (95.19) and 4 infected (34.12) subjects in the 1.8 × 10⁶ group, but the differences did not reach the level of statistical significance (P = .095 and .90). At the time of the second CHMI the 6 infected subjects in the 9 × 10⁵ PfSPZ group had a nonsignificant lower median net reciprocal serum dilution for 80% inhibition (10.91) than did the one uninfected (70.38) subject in the 1.8 × 10⁶ group and the 5 infected (14.78) subjects in the 1.8 × 10⁶ group (P = .29 and .54). Abbreviations: aIFA, automated immunofluorescence assay; CHMI, controlled human malaria infection; ELISA, enzyme-linked immunosorbent assay; IgG, immunoglobulin G; ISI, inhibition of sporozoite invasion assay; PfCSP, Pf circumsporozoite protein; PfSPZ, Plasmodium falciparum sporozoite.

Figure 4.

Pf-specific memory CD4 T-cell responses following different PfSPZ Vaccine doses. T-cell responses were assessed by flow cytometry 2 weeks after first and third doses of vaccine, and prior to the first and second CHMIs. We incubated PBMCs from vaccinees and controls with radiation attenuated (150 Gy) aseptic, purified, cryopreserved PfSPZ for ~18 hours and then assessed the percent of T cells that specifically responded to the PfSPZ. The percent of memory CD4 T cells in the blood expressing IFNγ, IL-2, or TNFα preimmunization, 2 weeks after the first and third doses of 9.0 × 10⁵ PfSPZ Vaccine (left) or 1.8 × 10⁶ PfSPZ Vaccine (right), and before each CHMI time point is shown. Results are the percentage of cytokine-producing cells after incubation with PfSPZ minus the percentage of cytokine-producing cells after incubation with vaccine diluent (medium with 1% human serum albumin). Colored symbols indicate vaccine groups, whereas open symbols represent placebo controls. Red symbols represent individuals challenged at the first CHMI time point, whereas blue and green symbols indicate individuals challenged at the second CHMI time point. Bars indicate median values within each group. Differences within each age group between pre- and post-vaccination groups were assessed by 2-way ANOVA with Dunnett’s correction for multiple comparisons. *P < .05, **P < .01. For preimmunization T-cell responses, there were no significant differences between infected and noninfected vaccinees (Table S7). Abbreviations: ANOVA, analysis of variance; CHMI, controlled human malaria infection; IFNγ, interferon γ; IL, interleukin; PBMC, peripheral blood mononuclear cell; Pf, Plasmodium falciparum; PfSPZ, Plasmodium falciparum sporozoite; TNFα, tumor necrosis factor α.