Profiling immune functions to advance malaria vaccines

Multi-functional analysis of malaria vaccine-induced responses has the potential to better inform vaccine design and evaluation and accelerate malaria vaccine development goals. We propose a 3-tier approach to implementing assays to inform vaccine evaluation.

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Recent trends have shown a stagnation in the reduction of the global burden of malaria, with >200 million cases and >400,000 deaths reported in 2019. Vaccines are highly cost-effective for preventing infectious diseases and a vaccine to protect against malaria will greatly enhance control and elimination efforts. The RTS,S vaccine was recently recommended by WHO for infants and young children (5-22 months old) in regions with moderate-high malaria transmission. However, RTS,S displayed moderate efficacy in children that largely waned within 18 months. Vaccines with higher efficacy and that are suitable for use in other age groups remain a high priority.

Vaccines in development for malaria target a number of different antigens and parasite life-stages. They can be broadly grouped into pre-erythrocytic vaccines that target sporozoites or liver-stages, blood-stage vaccines targeting merozoites or infected erythrocytes, and transmission-blocking vaccines that aim to prevent transmission to mosquitoes.

Antibodies are a key mediator of immunity for malaria vaccines and function via multiple mechanisms. These include direct inhibition or neutralization, complement fixation and activation, and opsonic phagocytosis or cellular cytotoxicity by immune cells through interactions with Fc-receptors, and others.

To date, evaluation of vaccines has been largely restricted to quantifying antibody magnitude, or the application of a single assay of functional activity, depending on the vaccine type. This has limited understanding the protective potential of candidate vaccines.


Antibody responses can be categorized into 1) standard antibody parameters, 2) antibody Fc-dependent functions, and 3) antibody inhibitory functions. Note that some functional responses have been demonstrated specifically against pre-erythrocytic stage (PE), blood-stage (B) and transmission stage (T) parasites. Antibodies can also interact with monocytes to inhibit parasite growth (antibody-dependent cellular inhibition), disrupt or inhibit rosetting of pRBC and can and block parasite transmission to mosquitoes

Application of multi-functional antibody profiling in vaccine immunity

Although there are currently no well-established correlates of immunity, several assays are valuable for assessing candidate vaccines in pre-clinical development and clinical trials until clear immunological correlates are defined. Given the multiple ways in which antibodies can function to mediate protection, we argue that the application of multiple functional antibody assays for assessing vaccine responses should become routine, and vaccine evaluation should not rely on single reference assays. To achieve this, we propose a 3-tier approach of vaccine assays.


Three Tier approach to applying assays in vaccine development and clinical trial evalution

Tier 1 – Standard high-throughput immunoassays. This includes quantification of immunoglobulin isotypes (IgG, IgM, IgA) and IgG subclasses to the vaccine immunogen or target antigen. It also includes quantification of antibody avidity using antibody elution with chaotropic agents (e.g. thiocyanate). Assays can also quantify antibodies to specific regions or epitopes of target antigens or use competition approaches to assess epitope-specific or allele-specific antibodies. Typically, these assays are performed as plate-based ELISA, and throughput and reproducibility can be greatly enhanced by using automated liquid handling. Suspension bead arrays can also be used and are particularly suitable when antibodies to multiple antigens need to be quantified.

Tier 2 – Medium-high throughput functional assays. These include assays quantifying a functional activity of antibodies that can be performed without highly specialized resources and with sufficient throughput to enable testing of large sample numbers efficiently. Established assays in this category include GIA. Several other assays are also emerging, including complement fixation assays, Fcγ-receptor binding assays, and receptor-binding inhibition assays using recombinant proteins or parasites from culture. Additionally, assays of phagocytosis or ADCC can be performed using antigen-coated beads (to avoid the need for parasite culture) with reasonable throughput using standard flow cytometry approaches, and cell lines can be used to enhance standardization.

Tier 3 – Resource-intensive functional assays. These include assays that require higher technical capacity or specialized facilities/resources and are usually limited to specific laboratories with suitable expertise. Lower throughput and higher cost are significant considerations. In our model, these assays would only be performed after positive results in Tier 1 and 2 assays. Examples of assays in this category include i) inhibition of sporozoite motility and invasion for sporozoite vaccines, ii) inhibition of invasion using isolated merozoites with or without complement factors and ADCI for blood-stage vaccines, iii) and SMFA for transmission-blocking vaccines.


Future Directions

Quantification of multiple functional antibody parameters will provide a more complete assessment of responses generated by candidate vaccines and enable improved vaccine design and evaluation. Future research is needed to better understand the mechanisms mediating immunity and how these functional activities can be harnessed to maximize malaria vaccine efficacy. The wider application of multiple functional immunoassays in vaccine trials will be essential to accelerate the development of highly efficacious vaccines and will enable the identification of strong correlates of protection.

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This article was adapted from paper by D. Herbert Opi, Liriye Kurtovic, Jo-Anne Chan, Jessica L. Horton, Gaoqian Feng& James G. Beeson, Expert Review Vaccines, 2021

James Beeson

Deputy Director and Research Fellow, Burnet Institute

James is a public health physician and PhD graduate who has worked on the pathogenesis and immunology of malaria for many years through clinical and population studies and clinical trials. The major focus of his research is aimed at understanding the targets and mechanisms of protective immunity to malaria in humans, how protective immunity is acquired and maintained, and using this knowledge to advance malaria vaccine development and evaluation.