Optimising Controlled Human Malaria Infection Studies Using Cryopreserved P. falciparum Parasites Administered by Needle and Syringe

Abstract

Background: Controlled human malaria infection (CHMI) studies have become a routine tool to evaluate efficacy of candidate anti-malarial drugs and vaccines. To date, CHMI trials have mostly been conducted using the bite of infected mosquitoes, restricting the number of trial sites that can perform CHMI studies. Aseptic, cryopreserved P. falciparum sporozoites (PfSPZ Challenge) provide a potentially more accurate, reproducible and practical alternative, allowing a known number of sporozoites to be administered simply by injection.

Methodology: We sought to assess the infectivity of PfSPZ Challenge administered in different dosing regimens to malaria-naive healthy adults (n = 18). Six participants received 2,500 sporozoites intradermally (ID), six received 2,500 sporozoites intramuscularly (IM) and six received 25,000 sporozoites IM.

Findings: Five out of six participants receiving 2,500 sporozoites ID, 3/6 participants receiving 2,500 sporozoites IM and 6/6 participants receiving 25,000 sporozoites IM were successfully infected. The median time to diagnosis was 13.2, 17.8 and 12.7 days for 2,500 sporozoites ID, 2,500 sporozoites IM and 25,000 sporozoites IM respectively (Kaplan Meier method; p = 0.024 log rank test).

Conclusions: 2,500 sporozoites ID and 25,000 sporozoites IM have similar infectivities. Given the dose response in infectivity seen with IM administration, further work should evaluate increasing doses of PfSPZ Challenge IM to identify a dosing regimen that reliably infects 100% of participants.

Trial registration: ClinicalTrials.gov NCT01465048.

Conflict of interest statement

Competing Interests: BKLS, ERJ, PFB, AG, and SLH are employees of Sanaria Inc., which manufactures PfSPZ Challenge. Sanaria also manufactures other products in development based on aseptic, purified, cryopreserved PfSPZ, including PfSPZ Vaccine and PfSPZ-CVac. Sanaria sells parasites and mosquitoes to the research community, but none of its major products in development have been commercialized. BKLS and SLH are inventors on patents relevant to PfSPZ Challenge. None of the authors have had any consultancies relevant to this paper. This conflict of interest does not alter these authors’ adherence to all PLOS ONE policies on sharing data and materials, as detailed online in the guide for authors.

Figures

Figure 1. Flow chart of study design and volunteer recruitment.

20 participants were excluded following screening for the following reasons: significant psychiatric history (six individuals), consent withdrawn (four individuals), recruitment complete (2 individuals), lack of response from General Practitioner to medical screening letter (2 individuals), unexplained systolic murmur, elevated alanine transaminase, unexplained tachycardia and significant prior malaria exposure. In each group, the total dose of sporozoites was split between two injection sites and administered as two 50 µL injections, one in each deltoid.

Figure 2. PfSPZ Challenge Infectivity Data.

Kaplan-Meier analysis of time to patent parasitemia in days between injection and diagnosis (p = 0.024 logrank test). 2,500 ID = 2,500 sporozoites administered intradermally (ID). 2,500 IM = 2,500 sporozoites administered intramuscularly. 25,000 = 25,000 sporozoites administered intramuscularly. Median pre-patent period = 13.19 days for 2,500 sporozoites ID, 17.8 days for 2,500 sporozoites IM, 12.72 days for 25,000 sporozoites IM.

Figure 3. qPCR-measured parasite density for each individual subject grouped by dosing regimen.

Y axis = qPCR. X axis = days post injection of PfSPZ Challenge. Black subtitles indicate subject identification numbers; red subtitles indicate time to diagnosis by thick film microscopy in days. NI = not infected.

Figure 4. Estimation of Burden of Liver Infection and Liver-to-Blood Inoculum.

Data from participants successfully infected with malaria in groups 1, 2 and 3 compared to historical data from mosquito bite CHMI trials undertaken at our centre. SpZ = sporozoites. LLQ = lower limit of quantification by qPCR. ID = intradermal administration. IM = intramuscular administration. Mosquito bite = malaria naïve participants infected with P.falciparum by mosquito bite as infectivity control participants in vaccine efficacy studies undertaken recently at our centre (Ewer et al. submitted). (A) Peak qPCR-measured parasitaemia in first asexual cycle for each regimen. (B) Matrix scatterplot illustrating close correlation of different LBI measures with each other and with time to microscopic patency.

Figure 5. Parasite Multiplication Rate following PfSPZ Challenge is comparable to mosquito-bite subjects with similar LBIs.

Figure shows relationship Between PMR and LBI for participants successfully infected with malaria in groups 1–3 (green crosses), compared to historical data from mosquito bite CHMI trials undertaken at our centre involving malaria naïve unvaccinated subjects (blue dots), and malaria naïve ChAd63-MVA ME-TRAP vaccinated subjects (red dots). These data were obtained using the linear model; results were similar with the sine model (data not shown).

Figure 6. IFN-γ ELISPOT responses to individual malaria antigens post CHMI.

Median ex-vivo IFN-γ ELISPOT responses from the day before CHMI (C-1) and days 7, 11, 14, 35 and 90 post CHMI with cells stimulated against peptide pools to numerous malaria antigens. Each volunteer is represented as a single point. Lines represent the median response at each time point. (A) Individual responses to numerous malaria antigens over time (B) Sum responses to all peptides over time.