Repeat controlled human malaria infection of healthy UK adults with blood-stage Plasmodium falciparum: Safety and parasite growth dynamics

Abstract

In endemic settings it is known that natural malaria immunity is gradually acquired following repeated exposures. Here we sought to assess whether similar acquisition of blood-stage malaria immunity would occur following repeated parasite exposure by controlled human malaria infection (CHMI). We report the findings of repeat homologous blood-stage Plasmodium falciparum (3D7 clone) CHMI studies VAC063C (ClinicalTrials.gov NCT03906474) and VAC063 (ClinicalTrials.gov NCT02927145). In total, 24 healthy, unvaccinated, malaria-naïve UK adult participants underwent primary CHMI followed by drug treatment. Ten of these then underwent secondary CHMI in the same manner, and then six of these underwent a final tertiary CHMI. As with primary CHMI, malaria symptoms were common following secondary and tertiary infection, however, most resolved within a few days of treatment and there were no long term sequelae or serious adverse events related to CHMI. Despite detectable induction and boosting of anti-merozoite serum IgG antibody responses following each round of CHMI, there was no clear evidence of anti-parasite immunity (manifest as reduced parasite growth in vivo) conferred by repeated challenge with the homologous parasite in the majority of volunteers. However, three volunteers showed some variation in parasite growth dynamics in vivo following repeat CHMI that were either modest or short-lived. We also observed no major differences in clinical symptoms or laboratory markers of infection across the primary, secondary and tertiary challenges. However, there was a trend to more severe pyrexia after primary CHMI and the absence of a detectable transaminitis post-treatment following secondary and tertiary infection. We hypothesize that this could represent the initial induction of clinical immunity. Repeat homologous blood-stage CHMI is thus safe and provides a model with the potential to further the understanding of naturally acquired immunity to blood-stage infection in a highly controlled setting.

Clinical trial registration: ClinicalTrials.gov, identifier NCT03906474, NCT02927145.

Keywords: Plasmodium falciparum; controlled human malaria infection (CHMI); human challenge model; malaria; malaria immunology.

Figure 1

VAC063C flow chart of study design and volunteer recruitment. (A) Enrolment into the VAC063C study began in October 2018. Twenty five volunteers were screened for eligibility. Eleven eligible volunteers were identified, six of whom were VAC063 participants returning for a tertiary controlled human malaria infection (CHMI) (Group 1), two of whom were VAC063 participants returning for a secondary CHMI (Group 2) and three of whom were newly enrolled participants for primary CHMI (Group 3). Clinical follow-up continued until 90 days after challenge (C+90) and was completed by 14^th January 2019. Volunteer demographics are summarized in Table S1 . (B) In total (across three CHMI studies, VAC063A, B and C) 24 participants underwent primary CHMI, 10 of whom subsequently underwent secondary CHMI and six of those participants returned for a tertiary CHMI. The first CHMI (VAC063A) was on 14^th November 2017, the second (VAC063B) was on 6^th March 2018 and the third (VAC063C) was on 6^th November 2018.

Figure 2

Safety of repeated P. falciparum blood-stage CHMI. Participants were asked about the presence of 13 solicited systemic adverse events (AE) at each study visit following CHMI. (A) The maximum severity of any solicited AE reported by each participant in the 48 hours before and after diagnosis during primary (n=24), secondary (n=10) and tertiary (n=6) CHMI is shown as a proportion of the total number of participants. (B) VAC063C: The solicited AEs recorded during the CHMI period are shown as the maximum severity reported by each participant and as a proportion of the participants reporting each individual AE for primary (n=3), secondary (n=2) and tertiary (n=6) CHMI. AE data was collected until 90 days after challenge. Colour coding as per panel (A, C) The solicited AEs recorded during the CHMI period are shown as the maximum severity reported by each participant and as a proportion of the participants reporting each individual AE for primary (n=24), secondary (n=10) and tertiary (n=6) CHMI for all participants across the three CHMI studies. Groups refer to specific study group numbers in VAC063A, B and C. Colour coding as per panel (A).

Figure 3

Symptom evolution during repeated P. falciparum blood-stage CHMI. The maximum severity of any solicited systemic adverse events (AE) recorded at the indicated time points during the CHMI period for each participant for primary (n=24), secondary (n=10) and tertiary (n=6) CHMI.

Figure 4

Objective measurements and laboratory AEs during repeated P. falciparum blood-stage CHMI. (A) Frequency and severity of pyrexia, anaemia, thrombocytopaenia and lymphopaenia during primary (n=24), secondary (n=10) and tertiary (n=6) CHMI from screening until 90 days post-CHMI. (B) Alanine aminotransferase (ALT) for each participant in VAC063C at each timepoint for primary (n=3), secondary (n=2) and tertiary (n=6) CHMI showing significant transaminitis after treatment of the primary infection (P = 0.03 as calculated by Kruskal-Wallis test of peak ALT values). Dashed lines show the local cut off for Grade 1, 2 and 3 abnormalities. Time is number of days post-CHMI, except for day of diagnosis (DoD) and day 6 post-treatment (T+6) which vary by participant. The same information for VAC063 A and B (which did not include a T+6 assessment) is presented in Figure S2 . (C) Maximum recorded temperature during primary (n=24), secondary (n=10) and tertiary (n=6) CHMI. Individual data points and the median are shown. Dashed lines show the local cut off for Grade 1, 2 and 3 abnormalities. Kruskal-Wallis test showed no significant difference between primary, secondary and tertiary CHMI (P = 0.31).​ (D) Median haemoglobin concentration and lymphocyte count for all participants across VAC063A, B and C over time for primary (n=24), secondary (n=10) and tertiary (n=6) CHMI (of which n=18 were female and n=22 male). Time points are the same as in panel (B) Kruskal-Wallis test showed no significant difference between primary, secondary and tertiary CHMI (minimum haemoglobin concentration P = 0.93; minimum lymphocyte count P = 0.19).

Figure 5

Parasite growth dynamics following repeated P. falciparum blood-stage CHMI. (A) qPCR data for primary (n=24), secondary (n=10) and tertiary (n=6) CHMI across the VAC063A, B and C studies. Parasitaemia measured by qPCR over time in parasites/mL (p/mL) blood is shown for each volunteer. CHMI was initiated by intravenous inoculation of P. falciparum infected erythrocytes on day 0. The lower limit of quantification is indicated by the dotted line at 20 p/mL; values below this level are plotted for information only. (B) Median qPCR data are shown for primary, secondary and tertiary CHMI. Colours as in panel A. (C) Parasitaemia at the time of diagnosis. Individual data points and the median ± inter-quartile range for primary (n=24), secondary (n=9) and tertiary (n=6) CHMI are shown. Note one participant requested antimalarial treatment at day C+20 during secondary CHMI without reaching the diagnostic criteria so no data point is shown for this participant. P value as calculated by Kruskal-Wallis test showed no significant difference between groups. (D) Kaplan-Meier plot of time to diagnosis in days. Note one secondary CHMI participant requested treatment and was censored at day C+20. Log-rank (Mantel-Cox) test showed no significant difference in time to diagnosis between primary, secondary and tertiary CHMI. (E) The parasite multiplication rate (PMR) per 48 hours was modelled from the qPCR data up until the time point of diagnosis. Individual data points and median ± inter-quartile range for primary, secondary and tertiary CHMI PMR are shown. P value as calculated by Kruskal-Wallis test showed no significant difference.

Figure 6

Individual variation in parasite growth dynamics of repeated P. falciparum blood-stage CHMI. qPCR data for primary (blue), secondary (pink) and tertiary (purple) CHMI. Parasitaemia is shown over time for each volunteer who underwent more than one CHMI (total n=10). The PMR per 48 hours was modelled from the qPCR data up until the time point of diagnosis for each participant for each CHMI and is shown below the relevant parasite growth graph. (A) Data from participants who showed minimal variation in parasite growth dynamics between each CHMI (n=7 for primary and secondary CHMI, of whom n=4 also underwent tertiary CHMI). (B) One participant showed consistently slower parasite growth with each subsequent CHMI. (C) One participant showed slower parasite growth during secondary CHMI compared to primary but did not undergo a tertiary CHMI. (D) One participant showed completely suppressed parasite growth on secondary CHMI until day C+19 but no change compared to primary upon tertiary CHMI.

Figure 7

Induction of serum antibody responses to merozoite antigens during CHMI. (A) Serum anti-MSP1₁₉ total IgG ELISA was conducted on samples from participants for primary (n=19 at C-1; n=21 at C+28), secondary (n=8) and tertiary (n=6) CHMI. Sera from the pre-CHMI (C-1) and 28 days post-CHMI (C+28) time points were tested. (B) Serum anti-AMA1 total IgG as measured by standardized ELISA on samples from participants for primary (n=6 at C-1; n=24 at C+28), secondary (n=9 at C-1; n=10 at C+28) and tertiary (n=6) CHMI. Samples colour coded as per previous figures. Individual data points and median are shown.