RTS,S/AS01E immunization increases antibody responses to vaccine-unrelated Plasmodium falciparum antigens associated with protection against clinical malaria in African children:a case-control study

BACKGROUND: Vaccination and naturally acquired immunity against microbial pathogens may have complex interactions that influence disease outcomes. To date, only vaccine-specific immune responses have routinely been investigated in malaria vaccine trials conducted in endemic areas. We hypothesized that RTS,S/A01E immunization affects acquisition of antibodies to Plasmodium falciparum antigens not included in the vaccine and that such responses have an impact on overall malaria protective immunity. METHODS: We evaluated IgM and IgG responses to 38 P. falciparum proteins putatively involved in naturally acquired immunity to malaria in 195 young children participating in a case-control study nested within the African phase 3 clinical trial of RTS,S/AS01E (MAL055 NCT00866619) in two sites of different transmission intensity (Kintampo high and Manhiça moderate/low). We measured antibody levels by quantitative suspension array technology and applied regression models, multimarker analysis, and machine learning techniques to analyze factors affecting their levels and correlates of protection. RESULTS: RTS,S/AS01E immunization decreased antibody responses to parasite antigens considered as markers of exposure (MSP142, AMA1) and levels correlated with risk of clinical malaria over 1-year follow-up. In addition, we show for the first time that RTS,S vaccination increased IgG levels to a specific group of pre-erythrocytic and blood-stage antigens (MSP5, MSP1 block 2, RH4.2, EBA140, and SSP2/TRAP) which levels correlated with protection against clinical malaria (odds ratio [95% confidence interval] 0.53 [0.3-0.93], p = 0.03, for MSP1; 0.52 [0.26-0.98], p = 0.05, for SSP2) in multivariable logistic regression analyses. CONCLUSIONS: Increased antibody responses to specific P. falciparum antigens in subjects immunized with this partially efficacious vaccine upon natural infection may contribute to overall protective immunity against malaria. Inclusion of such antigens in multivalent constructs could result in more efficacious second-generation multistage vaccines

Vaccine efficacy against all episodes of clinical malaria (primary case definition) during an 18-mo follow-up period after dose 3 in infants 6–12 wk of age at enrollment, ordered by increasing malaria incidence at each study site (intention-to-treat population).

<p>Interaction <i>p-</i>value = 0.1143. The size of each blue square reflects the relative number of participants enrolled at each study site; the horizontal bars show the lower limit (LL) and upper limit (UL) of the 95% confidence interval. Study sites are ordered from lowest (Kilifi) to highest (Siaya) incidence of clinical malaria, defined as a measured or reported fever within previous 24 h and parasite density >0 parasites/mm³ (i.e., clinical malaria secondary case definition), measured in control infants 6–12 wk of age at enrollment during 12 mo of follow-up. VE is VE against all episodes of clinical malaria meeting the primary case definition, unadjusted for covariates. Clinical malaria primary case definition: illness in a child brought to a study facility with a temperature of ≥37.5°C and <i>P. falciparum</i> asexual parasitemia at a density of >5,000 parasites/mm³ or a case of malaria meeting the primary case definition of severe malaria.</p

Number of cases of severe malaria (secondary case definition) averted per 1,000 participants vaccinated during a 20-mo follow-up period (intention-to-treat population).

<p>Severe malaria secondary case definition: <i>P. falciparum</i> asexual parasitemia at a density of >5,000 parasites/mm³ with one or more markers of disease severity, including cases in which a coexisting illness was present or could not be ruled out. Markers of severe disease were prostration, respiratory distress, a Blantyre coma score of ≤2 (on a scale of 0 to 5, with higher scores indicating a higher level of consciousness), two or more observed or reported seizures, hypoglycemia, acidosis, elevated lactate level, or a hemoglobin level of <5 g/dl. Coexisting illnesses were defined as radiographically proven pneumonia, meningitis established by analysis of cerebrospinal fluid, bacteremia, or gastroenteritis with severe dehydration.</p

Number of cases of severe malaria (secondary case definition) averted per 1,000 participants vaccinated during an 18-mo follow-up period (per-protocol population).

<p>Severe malaria secondary case definition: <i>P. falciparum</i> asexual parasitemia at a density of >5,000 parasites/mm³ with one or more markers of disease severity, including cases in which a coexisting illness was present or could not be ruled out. Markers of severe disease were prostration, respiratory distress, a Blantyre coma score of ≤2 (on a scale of 0 to 5, with higher scores indicating a higher level of consciousness), two or more observed or reported seizures, hypoglycemia, acidosis, elevated lactate level, or a hemoglobin level of <5 g/dl. Coexisting illnesses were defined as radiographically proven pneumonia, meningitis established by analysis of cerebrospinal fluid, bacteremia, or gastroenteritis with severe dehydration.</p

Vaccine efficacy against all episodes of clinical and severe malaria in children aged 5–17 mo at enrollment.

1<p>Event rate for clinical malaria; affected rate (percent) for severe malaria and hospitalization.</p>2<p>For clinical malaria: <i>p</i>-value from negative binomial regression. For severe malaria, malaria hospitalization, and all-cause hospitalization: <i>p</i>-value from two-sided Fisher's exact test.</p>3<p>Clinical malaria primary case definition: illness in a child brought to a study facility with a measured temperature of ≥37.5°C and <i>P. falciparum</i> asexual parasitemia at a density of >5,000 parasites/mm³ or a case of malaria meeting the primary case definition of severe malaria.</p>4<p>Clinical malaria secondary case definition: illness in a child brought to a study facility with a measured temperature of ≥37.5°C or reported fever within the last 24 h and <i>P. falciparum</i> asexual parasitemia at a density of >0 parasites/mm³.</p>5<p>Severe malaria primary case definition: <i>P. falciparum</i> asexual parasitemia at a density of >5,000 parasites/mm³ with one or more markers of disease severity and without diagnosis of a coexisting illness. Markers of severe disease were prostration, respiratory distress, a Blantyre coma score of ≤2 (on a scale of 0 to 5, with higher scores indicating a higher level of consciousness), two or more observed or reported seizures, hypoglycemia, acidosis, elevated lactate level, or hemoglobin level of <5 g/dl. Coexisting illnesses were defined as radiographically proven pneumonia, meningitis established by analysis of cerebrospinal fluid, bacteremia, or gastroenteritis with severe dehydration.</p>6<p>Malaria hospitalization case definition: a medical hospitalization with confirmed <i>P. falciparum</i> asexual parasitemia at a density of >5,000 parasites/mm³.</p>7<p>All-cause hospitalization primary case definition: a medical hospitalization of any cause, excluding planned admissions for medical investigation/care or elective surgery and admissions for trauma.</p

Study sites and malaria endemicity.

<p>Adapted from Hay et al. <a href="http://www.plosmedicine.org/article/info:doi/10.1371/journal.pmed.1001685#pmed.1001685-Hay1" target="_blank">[34]</a>. The location of each participating study site is shown on this previously published map showing the spatial distribution of <i>P. falciparum</i> (Pf) malaria endemicity. The data are the model-based geostatistical point estimates of the annual mean <i>P. falciparum</i> parasite rate (PR) age-standardized for 2–10 y for 2007 within the stable spatial limits of <i>P. falciparum</i> malaria transmission, displayed as a continuum of yellow to red from 0%–100% (see map legend). The rest of the land area was defined as unstable risk (medium grey) or no risk (light grey). Nanoro, Burkina Faso, has highly seasonal malaria transmission.</p