Multifunctional IgG/IgM antibodies and cellular cytotoxicity are elicited by the full-length MSP1 SumayaVac-1 malaria vaccine.

"the clinical trial protocol and the ethical votes are available at dryad 10 5061/dryad kwh70rz0f. purified igg from msp1 fl vaccinees collected at day 85 four weeks after the third immunization (fig 1 ) induced phagocytosis of msp1 fl -coated fluorescent microspheres by both the monocyte cell line thp1 ( p < 0 0001) and neutrophils ( p < 0 0001) in the opsonic phagocytosis assay (fig 2a supplementary data 1 ) msp1 fl -coated microspheres yielded results comparable to those of merozoites (supplementary fig 1 ).; purified igg from msp1 fl vaccinees could also activate neutrophils to release a respiratory burst in the adrb assay ( p < 0 0001) (fig 2b supplementary data 1 ) consistent with previous reports .; production as compared with pre-immunization controls ( p = 0 001 and p = 0 001 respectively) (fig 2c supplementary data 1 ).; furthermore purified igg from msp1 fl vaccines activated the classical complement pathway as indicated by c1q fixation and c3b deposition ( p < 0 0001 and p = 0 0002 respectively) and the formation of the membrane attack complex (c5-c9 deposition) ( p = 0 0002) (fig 2d supplementary data 1 ).; we observed that the readouts of all functional assays peaked at day 85 sometimes exceeding the activity levels of semi-immune kenyan adults (who control) (fig 3a-f supplementary data 2 ) and then declined over a 3-month period but peaked again at day 210 four weeks after a booster immunization on day 182.; the increase in functional activity after the booster was significant for c1q fixation ( p = 0 0433) (fig 3a ) neutrophil opa ( p = 0 081) (fig 3c ) and adrb ( p = 0 081) (fig 3d and supplementary data 2 ).; p = 0 0049 and ab-nk:cd107a p = 0 0322) (fig 3a-f and supplementary data 2 ).; ( r = 0 55 95% ci 0 3982 to 0 7536 p < 0 0001) (fig 3g supplementary data 2 ).; we further investigated the correlation among the various assays and readout parameters and observed the highest level of cross-correlation between c1q fixation (abc') thp1 opa neutrophil opa and adrb ( r = 0 74-0 88 p < 0 0001) and moderate correlation with the two ab- nk activities ( r = 0 38-0 65 p < 0 0001) (fig 3h supplementary data 2 ).; furthermore vaccinees with multifunctional fc-mediated immune responses were also those having the highest igg titers after the third vaccination ( p = 0 0047) (fig 3j supplementary data 2 ).; in comparison reactivity to p30 was negligible (fig 4 supplementary fig 4 and supplementary data 3 ).; to further investigate the strain-transcending nature of fc- mediated effector functions triggered after immunization with sumayavac-1 we compared the adrb activities for 3 dosages previously obtained for 3d7 and fcb1 at group level and found the data to moderately correlate ( r = 0 70 p = 0 0067) (supplementary fig 3 and supplementary data 1 ).; furthermore the adrb activity using the fcb1 strain also correlated significantly with igg titers from the volunteers ( r = 0 57 p = 0 0366) (supplementary fig 3 and supplementary data 1 ) as did adrb activity and igg titers when using the homologous3d7 strain .; both opa activity and c1q fixation using msp1-f were significantly correlated with igg titers ( r = 0 7441 p = 0 0014 for opa and r = 0 5147 p = 0 0436 for c1q) as for adrb using fcb1 (supplementary fig 3 and supplementary data 1 ).; furthermore purified igm antibodies recognized numerous epitopes spread across the entire msp1 protein as shown using an msp1 peptide chip consisting of 1706 15mer oligopeptides with a neighbor-to-neighbor overlap of 14 amino acids (fig 5b and supplementary data 4 ).; production by nk cells in the ab-nk assays (fig 6a supplementary data 5 ).; we further noted igm-mediated activation of the classical complement pathway as shown by c1q fixation c3b deposition and the formation of the membrane attack complex (fig 6a supplementary data 5 ).; in general the induction and decay kinetics of igm-mediated effector functions resembled those of igg although at a lower scale (fig 6b supplementary fig 7 supplementary table 1 and supplementary data 5 ).; ( p < 0 0001) and r = 0 38 ( p = 0 01) for abc' (fig 6c supplementary data 5 ).; varying degrees of correlation were further found between the different igm-mediated effector functions evaluated (fig 6c supplementary data 5 ).; igg purified from a semi-immune population from kenya inhibited parasite growth by 12% (fig 6d and supplementary data 5 ).; t cell populations b cell subsets (supplementary fig 8 supplementary data 6 ) t follicular helper (tfh) cells including tfh1 tfh2 and tfh1-17 (supplementary fig 8 supplementary fig 9 and supplementary data 6 ) and different memory phenotypes (naive central memory effector memory and temra) of cd4 + and cd8 + t cells (supplementary fig 9 and supplementary data 6 ).; there was only a significant decrease in the tfh17 subset on day 85 compared with day 0 ( p = 0 0138) (supplementary fig 8 and supplementary data 6 ).; we found a significant increase in the expression level of cd38 + in cd4 + and cd8 + t cells ( p < 0 0001 and p = 0 0105 respectively) and a trend in tfh cells (fig 7a and supplementary data 6 ) cd38 + is a marker for cell differentiation activation and cytotoxic potential and is associated with a reduced parasite burden - .; other markers of activation such as icos did not follow the same trend (supplementary fig 8 and supplementary data 6 ).; interestingly the proportion of pd1+ cd4+ t cells cd8+ t cells and tfh cells was significantly reduced on day 85 compared with day 0 ( p = 0 0028 p = 0 0020 and p < 0 0001 respectively) (fig 7a and supplementary data 6 ) pd1 is a marker of exhaustion .; we further investigated the expression of 38 cytokines using a bead-based multiplex assay but observed no alterations between pbmcs collected from msp1 fl vaccinees on day 0 and 85 (supplementary fig 10 and supplementary data 6 ).; and il-10 (fig 7b supplementary fig 11 and supplementary data 6 ) as demonstrated by transcript-specific real-time rt-pcr.; and il-2 ( p = 0 0006 and p = 0 0001 respectively) (fig 7c and supplementary data 6 ).; and r = 0 71 for il-2; p = 00003 p = 000121 and p < 0 0001 respectively) (fig 7d supplementary fig 11 supplementary data 6 ).; as seen in fig 8a there was a significant rise in granzyme a and b levels at day 85 compared with day 0 ( p = 0 0296 and p = 0 0006 respectively) (fig 8a and supplementary data 7 ).; this finding prompted us to sort magnetically cd8+ t cells from the pbmc collected from the msp1 fl vaccinees pre-and post-vaccination (days0 and 85) yielding an enrichment of >95% (fig 8b supplementary fig 12 and supplementary data 7 ).; we found significant increases in the production of spot forming units (sfu) between day 0 and day 85 upon stimulation with msp1 fl ( p = 0 0312) (fig 8c d and supplementary data 7 )."

"Competing interests The authors declare the following conflicts of interest. EB and ML are shareholders of Sumaya GmbH & Co. KG and RTL and EB are employed by the above company."

"Funding For the publication fee we acknowledge financial support by Deutsche Forschungsgemeinschaft within the funding programme "Open Access Publikationskosten" as well as by Heidelberg University. Open Access funding enabled and organized by Projekt DEAL."

"The trial was performed following the principles of good clinical practice, in accordance with the ethical principles described in the Declaration of Helsinki (6th revision, 2008) and registered with EudraCT (No. 2016-002463-33; date of first approval 31 January 2018). The protocol and amendments were approved by the Ethics Committee of the Medical Faculty of Heidelberg (Ethical vote AFmo-538/2016) and the relevant regulatory authority (Paul Ehrlich Institute, Langen, Germany). All volunteers were fully informed about the trial and gave their written consent. A single-center, randomized, double-blind, placebo and adjuvant-controlled phase Ia first-in-human dose escalation trial was conducted at the ISO-certified clinical trial unit of the Clinical Pharmacology and Pharmacoepidemiology Department of the Heidelberg University Hospital. Sixteen participants were included in two consecutive cohorts, each randomizing 12 volunteers into a vaccine + adjuvant group, and two participants each receiving placebo or adjuvant only. The safety of the study and any dose increase was supervised by an independent Data and Safety Monitoring Board (DSMB). Further information regarding the phase 1a can be found in Blank & Fürle et al., 2020. The clinical trial protocol and the ethical votes are available at DRYAD 10.5061/dryad.kwh70rz0f."