Concentration and avidity of antibodies to different circumsporozoite epitopes correlate with RTS,S/AS01E malaria vaccine efficacy

Abstract

RTS,S/AS01E has been tested in a phase 3 malaria vaccine study with partial efficacy in African children and infants. In a cohort of 1028 subjects from one low (Bagomoyo) and two high (Nanoro, Kintampo) malaria transmission sites, we analysed IgG plasma/serum concentration and avidity to CSP (NANP-repeat and C-terminal domains) after a 3-dose vaccination against time to clinical malaria events during 12-months. Here we report that RTS,S/AS01E induces substantial increases in IgG levels from pre- to post-vaccination (p < 0.001), higher in NANP than C-terminus (2855 vs 1297 proportional change between means), and higher concentrations and avidities in children than infants (p < 0.001). Baseline CSP IgG levels are elevated in malaria cases than controls (p < 0.001). Both, IgG magnitude to NANP (hazard ratio [95% confidence interval] 0.61 [0.48-0.76]) and avidity to C-terminus (0.07 [0.05-0.90]) post-vaccination are significantly associated with vaccine efficacy. IgG avidity to the C-terminus emerges as a significant contributor to RTS,S/AS01E-mediated protection.

Fig. 1

Effect of RTS,S/AS01E vaccination on anti-CSP IgG responses. a Ratios of mean concentrations (log₁₀[EU mL⁻¹]) between RTS,S/AS01E and comparators, before (month [M]0) and after (M3) vaccination, adjusted by site, with adjusted-p-values obtained in linear regression coefficients comparing for significance of logarithm of ratios estimated through mixed models. Error bars represent 95% confidence intervals (CI) estimated fitting the mixed model. b Correlations between CSP NANP and C-terminus (C-term) IgG concentrations. Regression lines with scatterplots show the association among: (i) all subjects pre-vaccination and post-vaccination in a dashed orange line (r = 0.93; 95% CI = 0.93, 0.94); (ii) comparator vaccinees pre-vaccination (black circles) and post-vaccination (black triangles) and RTS,S/AS01E pre-vaccination (green circles) (r = 0.48; 95% CI = 0.42, 0.54) in a solid black line; (iii) RTS,S/AS01E post-vaccination (green triangles; r = 0.66; 95% CI = 0.62, 0.70) in a solid green colour line. c Correlations between CSP NANP and C-term IgG avidity index (AI). Only samples with IgG concentration above the assay lower limit of quantification (LLOQ) (NANP >1.43 EU mL⁻¹ and C-term > 2.79 EU mL⁻¹) are shown, and those were mostly referring to post-vaccination RTS,S/AS01E vaccinees (r = 0.36; 95% CI = 0.29, 0.42). The strength of associations represented by the slope of each regression line is also shown (all slopes with p value < 0.05, obtained in linear regression coefficient of the slope, i.e. a t-test of the slope )

Fig. 2

Effect of age cohort and site on RTS,S/AS01E immunogenicity. Comparison of the impact of RTS,S/AS01E vaccination on anti-CSP responses across age cohorts (a IgG; c AI) and sites (b IgG; d AI). a Estimates of changes over time of infants (aged 6–12 weeks at vaccination) and children (aged 5–17 weeks at vaccination) and statistical significance of those were estimated through multivariate mixed models. Trajectories of NANP of infants and children were significantly different within vaccination group (all p values < 0.001). b Estimates of changes over time in each study site. Trajectories were statistically significantly different across sites (p adjusted for multiple testing for all pairwise comparisons < 0.001). Post-vaccination AI is compared across age cohorts (c) and sites (d) through t-tests and ANOVA, respectively. Boxplots illustrate the medians and the 25th and 75th quartiles, diamonds show the geometric mean, whiskers display the 1.5 interquartile ranges, and dots the outliers. M0: pre-vaccination, M3: post-vaccination

Fig. 3

Effect of CSP antibody concentration on protection against the first malaria episode. Kaplan–Meier estimates of time to the first clinical malaria episode during the 12-month follow-up period after the third vaccine dose by the three tertiles (high in grey; medium in blue; low in yellow) of IgG concentrations (EU mL⁻¹) to NANP (a) and to C-terminus (b) at month 3 in RTS,S vaccinees. The median survival time (days), i.e., time after which 50% of the cohort has not yet had an event, is shown for each antibody tertile with the corresponding 95% confidence intervals. p-values assessing differences in the distribution of survival time across the three strata in each of the subgroups were estimated through the Log-rank test. The number of subjects in each tertile are also indicated next to the plots, stratified by age group

Fig. 4

Effect of CSP antibody avidity on protection against the first malaria episode. Kaplan–Meier estimates of time to the first clinical malaria episode during the 12-month follow-up period after the third vaccine dose by the three tertiles (high in grey; medium in blue; low in yellow) of avidity index values at month 3 in RTS,S/AS01E vaccinees. Only plasma/serum samples which IgG concentrations were above the lower limit of quantification (LLOQ) (1.43 EU mL⁻¹ and 2.79 EU mL⁻¹ in NANP (a) and C-terminus [C-term] (b), respectively) were included. The median value for NANP was 0.43 and for C-term 0.11 and the number of subjects included was n = 691 for NANP and n = 698 for C-term. The median survival time (days), i.e., time at which the survivorship function equals 0.5, are shown next to the plots for each strata with the corresponding 95% confidence intervals. p-values assessing differences in the distribution of survival time between the two strata were significant and have been estimated through the Log-rank test. The number of subjects in each tertile are also indicated next to the plots, stratified by age group

Fig. 5

Effect of CSP antibody concentration on protection against multiple malaria episodes. Association between continuous anti-CSP IgG responses and the hazard of recurrent clinical malaria events within a 12-month follow-up period. a One unit increase in IgG NANP protects from clinical malaria and is significant (95% confidence interval [CI] excludes 1) in RTS,S/AS01E vaccinees in the beginning of the follow-up period. As times progresses, increased NANP IgG is less protective and becomes not significant (95% CI includes 1) at approximately 120 post-vaccination days. b The hazard ratio (HR) of C-terminus (C-term) IgG and recurrent clinical malaria episodes is shown suggesting that increased C-term IgG is significantly protective from clinical malaria until day 90 in RTS,S/AS01E vaccinees. c Increased avidity index (AI) to NANP and, in d increased AI to C-term, is constantly and significantly associated with a lower risk of clinical malaria in RTS,S/AS01E vaccinees. HRs were estimated in a time-varying proportional hazard model with a HR exponentially varying over time (p-value of Schonfeld residual tests for NANP and C-term IgG was <0.001, thus, rejecting a constant HR). Models for IgG concentrations and AIs also included study sites and age cohort with the following HRs: NANP IgG—HR_Kintampo = 25 [14,46], HR_Nanoro = 31 [17,57], HR_{612 weeks, <180 days} = 2.40 (1.39; 4.14) * [0.99 (0.99; 0.99)]^Days; HR_{612 weeks, >180 days} = 0.05 (0.02; 0.11) * [1.01 (1.00; 1.01)]^Days; C-term IgG—HR_Kintampo = 20 [11, 35], HR_Nanoro = 25 [14, 43], HR_{612 weeks, <180 days} = 3.21 (1.90; 5.41) * [0.99 (0.98; 0.99)]^Days, HR_{612 weeks, >180 days} = 0.06 (0.03; 0.14) * [1.01 (1.01; 1.01)]^Days; NANP AI - HR_Kintampo = 29 [16, 52], HR_Nanoro = 37 [21, 68], HR_{612 weeks} = 1.03 [0.91, 1.18]; C-term AI—HR_Kintampo = 23 [14, 40], HR_Nanoro = 31 [18, 52], HR_{612 weeks} = 0.98 [0.85, 1.13]

Fig. 6

Effect of HBsAg antibody concentration on protection against malaria episodes. Association between categorized anti-HBsAg IgG responses and time to the first clinical malaria episode (a), and between continuous anti-HBsAg IgG responses and the hazard of first (b) and recurrent (c) clinical malaria events within a 12-month follow-up period. a Kaplan–Meier curves of time to first clinical malaria for subjects with anti-HBsAg IgG concentration in the upper (grey), intermediate (blue) and lower (yellow) tertiles. Median survival time (and 95% confidence interval [CI]) of subjects in each tertile and sample size stratified by age group are shown, as well as the p-value of the log-rank test comparing survival time across tertile-based categories. b One unit increase in continuous anti-HBsAg IgG protects from clinical malaria in analysis unadjusted and adjusted for anti-NANP IgG concentration. c One unit increase in continuous anti-HBsAg IgG protects from clinical malaria until day 180 and is significantly protective of recurrent events of malaria (95% CI excludes 1) but 180 days after vaccination associations become not statistically significant. Comparable results are shown in d when repeating analysis of recurrent malaria events but adjusting for anti-NANP IgG concentration (p-value of Schonfeld residual tests for analysis of recurrent events unadjusted for anti-NANP was 0.01 and adjusted for anti-NANP was 0.02, thus, rejecting a constant hazard ratio). All models were adjusted by study site and age cohort