Anti-sporozoite monoclonal antibody for malaria prevention: secondary efficacy outcome of a phase 2 randomized trial

Abstract

CIS43LS is a long-acting monoclonal antibody specific for the Plasmodium falciparum circumsporozoite protein expressed on sporozoites. We previously reported that CIS43LS is protective against P. falciparum infection as detected by thick blood smear (TBS; primary endpoint) in a phase 2 double-blind randomized trial involving 330 healthy Malian adults receiving placebo or a single intravenous infusion of 10 mg kg^-1 or 40 mg kg^-1 of CIS43LS (1:1:1). At enrollment, all participants received artemether-lumefantrine to clear possible P. falciparum infection. Although TBS examination is the standard assay to assess efficacy in malaria vaccines trials in endemic areas, it has poor analytical sensitivity; therefore, it remained unknown whether CIS43LS had achieved sterile protection against infection. Here we report the prespecified secondary efficacy endpoint that used a Plasmodium 18S rRNA quantitative reverse transcription-PCR (qRT-PCR) assay that is ~2,000-fold more sensitive than TBS. We analyzed 5,015 dried blood spots collected before CIS43LS or placebo administration and biweekly thereafter over a 6-month malaria season. At 6 months, efficacy of CIS43LS against qRT-PCR-detected infection assessed in a time-to-event analysis was 87.4% for 40 mg kg^-1 (adjusted 95% confidence interval (CI), 79.5-92.3; P < 0.001) and 77.0% for 10 mg kg^-1 (adjusted 95% CI, 65.0-84.0; P < 0.001) versus placebo. A post hoc analysis with a gametocyte mRNA-specific qRT-PCR assay showed 6-month efficacy against gametocytemia of 87.7% for 40 mg kg^-1 (adjusted 95% CI, 75.6-93.8; P < 0.001) and 73.0% for 10 mg kg^-1 (adjusted 95% CI, 54.0-84.0; P < 0.001), versus placebo. These data indicate that a single dose of anti-sporozoite monoclonal antibodies can achieve durable, sterile protection against P. falciparum infection, underscoring their potential to reduce malaria disease burden and transmission. ClinicalTrials.gov identifier: NCT04329104 .

Fig. 1. Efficacy trial design and CONSORT diagram.

a, In the efficacy trial that began before the 6-month malaria transmission season, participants received a standard 3-day treatment course of AL at enrollment (day −14 ± 7). On day 0 they were randomized and received a single IV infusion of normal saline placebo (n = 110), or CIS43LS at a dose of 10 mg kg⁻¹ (n = 110) or 40 mg kg⁻¹ (n = 110). DBS and TBS were obtained for 18S qRT–PCR and microscopic examination, respectively, at enrollment and on days 0, 1, 3, 7, 14, 21 and 28 and every 2 weeks thereafter for 24 weeks and during unscheduled illness visits when malaria was suspected. DBS from study day 1 were not analyzed by 18S qRT–PCR. b, Screening, enrollment, randomization and follow-up for the initial safety study and efficacy trial.

Fig. 2. Distribution of P. falciparum parasitemia as detected by 18S qRT–PCR and TBS examination and their correlation.

a,b, A total of 5,015 paired DBS and TBS collected from 330 participants over the 6-month study period were analyzed by 18S qRT–PCR and TBS microscopy, respectively. 18S qRT–PCR results were plotted as not detected (ND), low positive (LP) or quantitatively positive with a numeric value of ≥2.0 log₁₀ parasites ml⁻¹; histograms show the qualitative and quantitative results of 18S qRT–PCR (a) and TBS microscopy (b). c, Pearson correlation of P. falciparum parasite density by 18S qRT–PCR versus TBS among 214 paired samples that were quantitatively positive by both assays. P value for the Pearson correlation was determined by two-sided t-test. The solid red line shows a linear regression fit. d, Distribution of LP and quantitatively positive 18S qRT–PCR results among 486 samples that were negative by TBS. The red dashed vertical line indicates the approximate LoD of TBS microscopy (~40,000 parasites ml⁻¹). Pf, P. falciparum.

Fig. 3. Time to clearance of baseline P. falciparum infection after AL treatment.

Among all study participants (n = 330), the percentage who had baseline P. falciparum at enrollment before AL administration (study day −14 ± 7) is shown, as detected by 18S qRT–PCR (red line) or TBS microscopy (blue line), followed by the percentage of participants who failed to clear their baseline infections by 18S qRT–PCR or TBS by the day of CIS43LS/placebo administration (day 0) or by scheduled visits thereafter through day 70. For each participant who had infection detected by 18S qRT–PCR at enrollment, subsequent clearance of infection was defined as two consecutive negative 18S qRT–PCR assay results. For seven participants (two in the placebo arm, three in the 10 mg kg⁻¹ arm and two in the 40 mg kg⁻¹ arm), DBS at enrollment were not available for 18S qRT–PCR analysis.

Fig. 4. CIS43LS efficacy against P. falciparum infection as determined by Plasmodium 18S qRT–PCR.

Cumulative incidence of P. falciparum blood-stage infection during a 6-month malaria season (irrespective of symptoms being present) after a single IV infusion of 10 mg kg⁻¹ of CIS43LS (n = 110), 40 mg kg⁻¹ of CIS43LS (n = 110) or placebo (n = 110). P. falciparum infections were detected by 18S qRT–PCR analysis of DBS collected during scheduled trial visits and unscheduled illness visits when malaria was suspected. DBS were collected before the administration of CIS43LS or placebo on day 0 and then on days 1, 3, 7, 14, 21 and 28 and every 2 weeks thereafter for a total of 24 weeks (day 168). Only DBS collected between days 21 and 168 were included. Estimates of the cumulative risk of infection are plotted along with the 95% CIs (shaded areas).

Fig. 5. Time to clearance of baseline P. falciparum gametocytemia after AL administration and subsequent risk of gametocytemia after CIS43LS or placebo administration.

a, The percentage of participants infected with P. falciparum gametocytes as detected by a gametocyte mRNA-specific qRT–PCR assay at enrollment before AL administration (study day −14 ± 7), on day 0 when CIS43LS or placebo was administered and at scheduled visits thereafter through day 70, is shown for each trial arm (n = 110 for each arm). b, Cumulative incidence of P. falciparum gametocytemia during a 6-month malaria season after a single IV infusion of 10 mg kg⁻¹ of CIS43LS (n = 110), 40 mg kg⁻¹ of CIS43LS (n = 110) or placebo (n = 110) during scheduled and unscheduled illness visits when malaria was suspected. Only DBS collected between days 7 and 168 were included in this analysis. Estimates of the cumulative risk of gametocytemia are plotted along with the 95% CIs (shaded areas).

Extended Data Fig. 1. Assessment of bias between 18S qRT-PCR- and TBS-based parasite densities.

A Bland-Altman plot demonstrates minimal bias (0.21 log₁₀ parasites/mL) between log₁₀ transformed Plasmodium 18S qRT-PCR- and TBS-based parasite densities. There was no density dependence to the bias (P = 0.19), as determined by simple linear regression with a two-sided P value. The solid black line represents the difference between log₁₀ measurements (bias) and the dashed red lines represent mean bias ± 1 SD.

Extended Data Fig. 2. Comparison of Plasmodium 18S qRT-PCR and TBS results, including discrepant results.

A total of 5015 paired DBS and TBS collected from 330 participants over the 6-month study period were analyzed by 18S qRT-PCR and microscopy, respectively. P. falciparum was detected quantitatively by both tests in 214 (4.3%) paired samples, and not detected by either test in 4159 (82.9%) paired samples. Among the remaining 642 discrepant samples, 624 (97.2%) were TBS-negative and either qualitatively low positive (LP; n = 139) or quantitatively positive by 18S qRT-PCR (n = 485). The remaining 18 discrepant samples were 18 s qRT-PCR-negative and TBS-positive (2.8% of discrepant samples and 0.4% of samples overall). Discrepant samples are highlighted by red boxes and the number of such samples is shown.

Extended Data Fig. 3. Relationship between enrollment date and time to clearance of baseline P. falciparum infection.

The y axis represents the number of days from enrollment (when the antimalarial drug AL was given) to the first qualitatively negative P. falciparum 18S qRT-PCR, shown by calendar date of enrollment (x axis). Among those who were infected at enrollment, clearance was defined as two consecutive negative 18S qRT-PCR results after enrollment. Each dot represents one participant. The 16 participants who remained 18S qRT-PCR positive through study day 21 (28 – 36 days after enrollment) are shown in shades of red. Participants who were uninfected at enrollment are shown in shades of gray/black (n = 196), and those who were infected at enrollment and cleared infections by study day 21 are shown in shades of blue (n = 118). Diamonds (n = 12) represent participants who were censored (that is participants who were still infected with P. falciparum when the last recorded study visit occurred).

Extended Data Fig. 4. Longitudinal P. falciparum genotype analysis of participants who were infected at baseline and remained 18S qRT-PCR positive by study day 21.

Panels a-l show allelic diversity for 6 P. falciparum gene fragments determined via amplicon sequencing (AmpSeq) of DBS from 12 participants who had P. falciparum infection detected by 18S qRT-PCR at enrollment (when AL was administered) and who remained infected through at least study day 21 after CIS43LS/placebo administration (study day 0). Clearance of infection was defined as two consecutive negative 18S qRT-PCR results after enrollment. Associated 18S qRT-PCR and thick blood smear (TBS) records are presented at the top of each participant plot. Timepoints ascend left to right from enrollment (8–13 days before study day 0) through 24 weeks for most participants. Circles represent alleles detected by AmpSeq and are grouped according to the analyzed locus (boxes). Circles are ordered vertically within boxes according to timepoint of first detection (earliest at bottom). Matching circle colors represent repeated observations of the same allele, either longitudinally (that is, left to right, see also colored edges) or between participants. The simultaneous detection of multiple alleles within a single locus indicates a polyclonal infection state. A total of 147 unique alleles were observed across all loci for the 12-participant dataset.