PRIMALVAC

Recombinant VAR2CSA protein as vaccine candidate for placental malaria

Press Release August 2012

Background

VAR2CSA is a 350 kDa transmembrane protein with a 300 kDa extracellular region composed of six Duffy-binding-like (DBL) domains and a cysteine-rich interdomain region module, as well as short inter-domain regions1,2.  It has been shown that, unlike the individual DBL domains, the full-length VAR2CSA extracellular region binds with high affinity and specificity to CSA3,4, and that the main binding site lies within the DBL1X-3X segment of VAR2CSA3,5,6.  It has been found that DBL3X is the principal target of the inhibitory antibodies.  Immunisation of the full length VAR2CSA protein as well as multiple domain constructs induced antibodies that efficiently abrogate parasite adhesion to CSA5,6.  Interestingly, naturally acquired antibodies and those induced by vaccination against the domain between the N-terminal sequence and the DBL2X segment7,8 target overlapping strain-transcendent anti-adhesion epitopes.  Taken together, these results indicate that DBL1X-3X region is an important target for inhibitory antibodies, and that strategies aimed at blocking P. falciparum infected Erythrocytes adhesion to CSA should focus on the N-terminal region of VAR2CSA.

Participants to the PRIMALVAC
Kick-off Meeting

PRIMALVAC Group

Infected erythrocytes binding to
placental cell line

Infected erythrocytes

Objectives

This project aims at developing a vaccine to prevent placental malaria and improve pregnancy outcomes using translational research.

The main objective is to obtain proof of concept that a VAR2CSA based vaccine inducing long lasting or rapidly boosted cross reactive and inhibitory antibodies can be designed for human use.  Recombinant forms of VAR2CSA will thus be generated, and their activity as immunogens that elicit functional and cross-reactive antibodies against placental parasite forms will be assessed.  The candidate antigens that best meet strict immunogenicity criteria will be transitioned to preclinical and clinical development.

Major Milestones

- Evaluation of various expression systems (Q3 2012)
- Batch of recombinant protein for immunogenicity testing (Q4 2012)
- Vaccine candidate for further preclinical and clinical development selected (Q2 2013)
- Development of Quality Control (QC) assays (Q2 2014)
- Batch release of the Master Cell Bank expressing the VAR2CSA clone (Q3 2014)
- Clinical phase I batch available (Q1 2015)
- Toxicity studies draft report (Q2 2015)
- Approval from Independent Ethics Committee and Regulatory Authorities (Q3 2015)
- Expected start of phase I clinical trial: (Q3 2015)

Achievements

- In 2012, evaluation of different constructs in various expression systems was performed in Pseudomonas fluorescens (Pfenex, USA) and in E. coli, L. lactis, P. pastoris and Chinese Hamster Ovary (CHO) cells (GTP Technology, France).
- Functional characterisation of micro-purified proteins was performed. 3D7 VAR2CSA DBL1-2 and FCR VAR2CSA DBL3-4 proteins expressed in CHO cells and E. coli were produced at larger analytical scale.  VAR2CSA DBL1-2 protein expressed in E. coli for further development.
- After removal of the His tag, the parameters of fermentation for the 50l scale production of the 3D7 VAR2CSA DBL1-2 protein were established at GTP Technology in November 2013.

References:                                                                                              

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  1. Bockhorst J, Lu F, Janes JH, Keebler J, Gamain B, Awadalla P, et al. Structural polymorphism and diversifying selection on the pregnancy malaria vaccine candidate VAR2CSA. Mol Biochem Parasitol. 2007 Oct;155(2):103-12
  2. Rask TS, Hansen DA, Theander TG, Gorm Pedersen A, Lavstsen T. Plasmodium falciparum erythrocyte membrane protein 1 diversity in seven genomes--divide and conquer. PLoS Comput Biol. 2010;6(9).
  3. Srivastava A, Gangnard S, Dechavanne S, Amirat F, Lewit Bentley A, Bentley GA, et al. var2CSA minimal CSA binding region is located within the N-terminal region. PLoS One. 2011;6(5):e20270.
  4. Srivastava A, Gangnard S, Round A, Dechavanne S, Juillerat A, Raynal B, et al. Full-length extracellular region of the var2CSA variant of PfEMP1 is required for specific, high-affinity binding to CSA. Proc Natl Acad Sci U S A. 2010 Mar 16;107(11):4884-9.
  5. Dahlback M, Jorgensen LM, Nielsen MA, Clausen TM, Ditlev SB, Resende M, et al. The chondroitin sulfate A-binding site of the VAR2CSA protein involves multiple N-terminal domains. J Biol Chem. 2011 May 6;286(18):15908-17.
  6. Avril M, Hathaway MJ, Srivastava A, Dechavanne S, Hommel M, Beeson JG, et al. Antibodies to a full-length VAR2CSA immunogen are broadly strain-transcendent but do not cross-inhibit different placental-type parasite isolates. PLoS ONE. 2011;6(2):e16622.
  7. Bordbar B, Gnidehou S, Ndam NT, Doritchamou J, Moussiliou A, Quiviger M, et al. Electroporation-mediated genetic vaccination for antigen mapping: Application to Plasmodium falciparum VAR2CSA protein. Bioelectrochemistry. 2011 Dec 30
  8. Bigey P, Gnidehou S, Doritchamou J, Quiviger M, Viwami F, Couturier A, et al. The NTS-DBL2X region of VAR2CSA induces cross-reactive antibodies that inhibit adhesion of several Plasmodium falciparum isolates to chondroitin sulfate A. J Infect Dis. 2011 Oct 1;204(7):1125-33.