Skip to main content
SpringerLink
Log in

A Plant-Based Transient Expression System for the Rapid Production of Malaria Vaccine Candidates

  • Protocol
  • First Online:
Book cover Vaccine Design

Part of the book series: Methods in Molecular Biology ((MIMB,volume 1404))

Abstract

There are currently no vaccines that provide sterile immunity against malaria. Various proteins from different stages of the Plasmodium falciparum life cycle have been evaluated as vaccine candidates, but none of them have fulfilled expectations. Therefore, combinations of key antigens from different stages of the parasites life cycle may be essential for the development of efficacious malaria vaccines. Following the identification of promising antigens using bioinformatics, proteomics, and/or immunological approaches, it is necessary to express, purify, and characterize these proteins and explore the potential of fusion constructs combining different antigens or antigen domains before committing to expensive and time-consuming clinical development. Here, using malaria vaccine candidates as an example, we describe how Agrobacterium tumefaciens-based transient expression in plants can be combined with a modular and flexible cloning strategy as a robust and versatile tool for the rapid production of candidate antigens during research and development.

This is a preview of subscription content, log in via an institution to check access.

Access this chapter

Log in via an institution
Protocol
USD 49.95
Price excludes VAT (USA)
  • Available as PDF
  • Read on any device
  • Instant download
  • Own it forever
eBook
USD 129.00
Price excludes VAT (USA)
  • Available as EPUB and PDF
  • Read on any device
  • Instant download
  • Own it forever
Softcover Book
USD 169.99
Price excludes VAT (USA)
  • Compact, lightweight edition
  • Dispatched in 3 to 5 business days
  • Free shipping worldwide - see info
Hardcover Book
USD 249.99
Price excludes VAT (USA)
  • Durable hardcover edition
  • Dispatched in 3 to 5 business days
  • Free shipping worldwide - see info

Tax calculation will be finalised at checkout

Purchases are for personal use only

Institutional subscriptions

References

  1. Crompton PD, Pierce SK, Miller LH (2010) Advances and challenges in malaria vaccine development. J Clin Invest 120:4168–4178

    CAS  PubMed  PubMed Central  Google Scholar 

  2. Vaughan AM, Kappe SHI (2012) Malaria vaccine development: persistent challenges. Curr Opin Immunol 24:324–331

    CAS  PubMed  Google Scholar 

  3. Hill AVS (2011) Vaccines against malaria. Philos Trans R Soc Lond B Biol Sci 366:2806–2814

    PubMed  PubMed Central  Google Scholar 

  4. Davey MR, Gartland KM, Mulligan BJ (1986) Transformation of the genomic expression of plant cells. Symp Soc Exp Biol 40:85–120

    CAS  PubMed  Google Scholar 

  5. Gleba Y, Klimyuk V, Marillonnet S (2007) Viral vectors for the expression of proteins in plants. Curr Opin Biotechnol 18:134–141

    CAS  PubMed  Google Scholar 

  6. Pogue GP, Vojdani F, Palmer KE et al (2010) Production of pharmaceutical-grade recombinant aprotinin and a monoclonal antibody product using plant-based transient expression systems. Plant Biotechnol J 8:638–654

    CAS  PubMed  Google Scholar 

  7. Gleba YY, Klimyuk VV, Marillonnet SS (2005) Magnifection—a new platform for expressing recombinant vaccines in plants. Vaccine 23:2042–2048

    CAS  PubMed  Google Scholar 

  8. Santi L, Giritch A, Roy CJ et al (2006) Protection conferred by recombinant Yersinia pestis antigens produced by a rapid and highly scalable plant expression system. Proc Natl Acad Sci U S A 103:861–866

    CAS  PubMed  PubMed Central  Google Scholar 

  9. Olinger GG, Pettitt J, KIM D et al (2012) Delayed treatment of Ebola virus infection with plant-derived monoclonal antibodies provides protection in rhesus macaques. Proc Natl Acad Sci U S A 109:18030–18035

    CAS  PubMed  PubMed Central  Google Scholar 

  10. D’Aoust M-A, Couture MMJ, Charland N et al (2010) The production of hemagglutinin-based virus-like particles in plants: a rapid, efficient and safe response to pandemic influenza. Plant Biotechnol J 8:607–619

    PubMed  Google Scholar 

  11. Jones RM, Chichester JA, Manceva S et al (2015) A novel plant-produced Pfs25 fusion subunit vaccine induces long-lasting transmission blocking antibody responses. Hum Vaccin Immunother 11:124–132

    PubMed  Google Scholar 

  12. Klimyuk V, Pogue G, Herz S et al (2014) Production of recombinant antigens and antibodies in Nicotiana benthamiana using “magnifection” technology: GMP-compliant facilities for small- and large-scale manufacturing. Curr Top Microbiol Immunol 375:127–154

    CAS  PubMed  Google Scholar 

  13. Chichester JA, Manceva SD, Rhee A et al (2013) A plant-produced protective antigen vaccine confers protection in rabbits against a lethal aerosolized challenge with Bacillus anthracis Ames spores. Hum Vaccin Immunother 9:544–552

    CAS  PubMed  PubMed Central  Google Scholar 

  14. Voepel N, Boes A, Edgue G et al (2014) Malaria vaccine candidate antigen targeting the pre erythrocytic stage of Plasmodium falciparum produced at high-level in plants. Biotechnol J 9:1435–1445

    CAS  PubMed  Google Scholar 

  15. Boes A, Spiegel H, Edgue G et al (2015) Detailed functional characterization of glycosylated and nonglycosylated variants of malaria vaccine candidate PfAMA1 produced in Nicotiana benthamiana and analysis of growth inhibitory responses in rabbits. Plant Biotechnol J 13:222–234

    CAS  PubMed  Google Scholar 

  16. Beiss V, Spiegel H, Boes A et al (2015) Heat-precipitation allows the efficient purification of a functional plant-derived malaria transmission-blocking vaccine candidate fusion protein. Biotechnol Bioeng 112:1297–1305

    CAS  PubMed  Google Scholar 

  17. Buyel JF, Gruchow HM, Boes A, Fischer R (2014) Rational design of a host cell protein heat precipitation step simplifies the subsequent purification of recombinant proteins from tobacco. Biochem Eng J 88:162–170

    CAS  Google Scholar 

  18. Koncz C, Schell J (1986) The promoter of TL-DNA gene 5 controls the tissue-specific expression of chimaeric genes carried by a novel type of Agrobacterium binary vector. Mol Gen Genet 204:383–396

    CAS  Google Scholar 

  19. Maclean J, Koekemoer M, Olivier AJ et al (2007) Optimization of human papillomavirus type 16 (HPV-16) L1 expression in plants: comparison of the suitability of different HPV-16 L1 gene variants and different cell-compartment localization. J Gen Virol 88:1460–1469

    CAS  PubMed  Google Scholar 

  20. von Itzstein M, Plebanski M, Cooke BM, Coppel RL (2008) Hot, sweet and sticky: the glycobiology of Plasmodium falciparum. Trends Parasitol 24:210–218

    Google Scholar 

  21. Wandelt CIC, Khan MRM, Craig SS et al (1992) Vicilin with carboxy-terminal KDEL is retained in the endoplasmic reticulum and accumulates to high levels in the leaves of transgenic plants. Plant J 2:181–192

    CAS  PubMed  Google Scholar 

  22. Remarque EJE, Faber BWB, Kocken CHMC, Thomas AWA (2008) A diversity-covering approach to immunization with Plasmodium falciparum apical membrane antigen 1 induces broader allelic recognition and growth inhibition responses in rabbits. Infect Immun 76:2660–2670

    CAS  PubMed  PubMed Central  Google Scholar 

  23. Faber BW, Younis S, Remarque EJ et al (2013) Diversity covering AMA1-MSP119 fusion proteins as malaria vaccines. Infect Immun 81:1479–1490

    CAS  PubMed  PubMed Central  Google Scholar 

  24. Chappel JA, Egan AF, Riley EM et al (1994) Naturally acquired human antibodies which recognize the first epidermal growth factor-like module in the Plasmodium falciparum merozoite surface protein 1 do not inhibit parasite growth in vitro. Infect Immun 62:4488–4494

    CAS  PubMed  PubMed Central  Google Scholar 

  25. Kumar S, Collins W, Egan A et al (2000) Immunogenicity and efficacy in aotus monkeys of four recombinant Plasmodium falciparum vaccines in multiple adjuvant formulations based on the 19-kilodalton C terminus of merozoite surface protein 1. Infect Immun 68:2215–2223

    CAS  PubMed  PubMed Central  Google Scholar 

  26. Chen L, Lopaticki S, Riglar DT et al (2011) An EGF-like protein forms a complex with PfRh5 and is required for invasion of human erythrocytes by Plasmodium falciparum. PLoS Pathog 7:e1002199

    CAS  PubMed  PubMed Central  Google Scholar 

  27. Reiling L, Richards JS, Fowkes FJI et al (2010) Evidence that the erythrocyte invasion ligand PfRh2 is a target of protective immunity against Plasmodium falciparum malaria. J Immunol 185:6157–6167

    CAS  PubMed  Google Scholar 

  28. Wang LL, Richie TLT, Stowers AA et al (2001) Naturally acquired antibody responses to Plasmodium falciparum merozoite surface protein 4 in a population living in an area of endemicity in Vietnam. Infect Immun 69:4390–4397

    CAS  PubMed  PubMed Central  Google Scholar 

  29. Gilson PR, Nebl T, Vukcevic D et al (2006) Identification and stoichiometry of glycosylphosphatidylinositol-anchored membrane proteins of the human malaria parasite Plasmodium falciparum. Mol Cell Proteomics 5:1286–1299

    CAS  PubMed  Google Scholar 

  30. Goschnick MW, Black CG, Kedzierski L (2004) Merozoite surface protein 4/5 provides protection against lethal challenge with a heterologous malaria parasite strain. Infect Immun 72:5840–5849

    CAS  PubMed  PubMed Central  Google Scholar 

  31. de Silva HD, Saleh S, Kovacevic S et al (2011) The antibody response to Plasmodium falciparum Merozoite Surface Protein 4: comparative assessment of specificity and growth inhibitory antibody activity to infection-acquired and immunization-induced epitopes. Malar J 10:266

    PubMed  PubMed Central  Google Scholar 

  32. Black CG, Wu T, Wang L et al (2001) Merozoite surface protein 8 of Plasmodium falciparum contains two epidermal growth factor-like domains. Mol Biochem Parasitol 114:217–226

    CAS  PubMed  Google Scholar 

  33. Black CG, Wang L, Wu T, Coppel RL (2003) Apical location of a novel EGF-like domain-containing protein of Plasmodium falciparum. Mol Biochem Parasitol 127:59–68

    CAS  PubMed  Google Scholar 

  34. Puentes A, Ocampo M, Rodríguez LE, Vera R (2005) Identifying Plasmodium falciparum merozoite surface protein-10 human erythrocyte specific binding regions. Biochimie 87:461–472

    CAS  PubMed  Google Scholar 

  35. Vermeulen AN (1985) Sequential expression of antigens on sexual stages of Plasmodium falciparum accessible to transmission-blocking antibodies in the mosquito. J Exp Med 162:1460–1476

    CAS  PubMed  Google Scholar 

  36. Duffy PE, Kaslow DC (1997) A novel malaria protein, Pfs28, and Pfs25 are genetically linked and synergistic as falciparum malaria transmission-blocking vaccines. Infect Immun 65:1109–1113

    CAS  PubMed  PubMed Central  Google Scholar 

  37. Yoshida N, Nussenzweig RS, Potocnjak P et al (1980) Hybridoma produces protective antibodies directed against the sporozoite stage of malaria parasite. Science 207:71–73

    CAS  PubMed  Google Scholar 

  38. Nussenzweig V, Nussenzweig RS (1985) Circumsporozoite proteins of malaria parasites. Cell 42:401–403

    CAS  PubMed  Google Scholar 

  39. Plassmeyer ML, Reiter K, Shimp RL et al (2009) Structure of the Plasmodium falciparum circumsporozoite protein, a leading malaria vaccine candidate. J Biol Chem 284:26951–26963

    CAS  PubMed  PubMed Central  Google Scholar 

  40. Campbell RE, Tour O, Palmer AE (2002) A monomeric red fluorescent protein. Proc Natl Acad Sci U S A 99:7877–7882

    CAS  PubMed  PubMed Central  Google Scholar 

  41. Wall MA, Socolich M, Ranganathan R (2000) The structural basis for red fluorescence in the tetrameric GFP homolog DsRed. Nat Struct Biol 7:1133–1138

    CAS  PubMed  Google Scholar 

  42. Shaner NC, Campbell RE, Steinbach PA et al (2004) Improved monomeric red, orange and yellow fluorescent proteins derived from Discosoma sp. red fluorescent protein. Nat Biotechnol 22:1567–1572

    CAS  PubMed  Google Scholar 

  43. Bevis BJ, Glick BS (2002) Rapidly maturing variants of the Discosoma red fluorescent protein (DsRed). Nat Biotechnol 20:83–87

    CAS  Google Scholar 

Download references

Acknowledgements

We thank Dr. Thomas Rademacher for cloning the pTRAkc vector series. The chimeric antibody 4G2 was kindly provided by Stefan Menzel. The work described in this chapter was partly supported by the Fraunhofer Future Foundation via the malaria vaccine project “Innovative technologies to manufacture ground-breaking biopharmaceutical products in microbes and plants”.

Author information

Authors and Affiliations

  1. Fraunhofer Institute for Molecular Biology and Applied Ecology IME, Forckenbeckstrasse 6, 52074, Aachen, Germany

    Alexander Boes, Andreas Reimann, Rainer Fischer, Stefan Schillberg & Holger Spiegel

  2. TRM Ltd, PO Box 93, York, YO43 3WE, UK

    Richard M. Twyman

  3. Institute for Molecular Biotechnology, RWTH Aachen University, Worringerweg 1, 52074, Aachen, Germany

    Rainer Fischer

Authors
  1. Alexander Boes
    View author publications

    You can also search for this author in PubMed Google Scholar

  2. Andreas Reimann
    View author publications

    You can also search for this author in PubMed Google Scholar

  3. Richard M. Twyman
    View author publications

    You can also search for this author in PubMed Google Scholar

  4. Rainer Fischer
    View author publications

    You can also search for this author in PubMed Google Scholar

  5. Stefan Schillberg
    View author publications

    You can also search for this author in PubMed Google Scholar

  6. Holger Spiegel
    View author publications

    You can also search for this author in PubMed Google Scholar

Corresponding author

Correspondence to Holger Spiegel .

Editor information

Editors and Affiliations

  1. Lankenau Institute for Medical Research, Wynnewood, PA, USA

    Sunil Thomas

Rights and permissions

Reprints and permissions

Copyright information

© 2016 Springer Science+Business Media New York

About this protocol

Cite this protocol

Boes, A., Reimann, A., Twyman, R.M., Fischer, R., Schillberg, S., Spiegel, H. (2016). A Plant-Based Transient Expression System for the Rapid Production of Malaria Vaccine Candidates. In: Thomas, S. (eds) Vaccine Design. Methods in Molecular Biology, vol 1404. Humana, New York, NY. https://doi.org/10.1007/978-1-4939-3389-1_39

Download citation

  • DOI: https://doi.org/10.1007/978-1-4939-3389-1_39

  • Published:

  • Publisher Name: Humana, New York, NY

  • Print ISBN: 978-1-4939-3388-4

  • Online ISBN: 978-1-4939-3389-1

  • eBook Packages: Springer Protocols

Publish with us

Policies and ethics

Search

Navigation