Archives of Pharmacal Research

, Volume 41, Issue 5, pp 544–553 | Cite as

Comparison of the size distributions and immunogenicity of human papillomavirus type 16 L1 virus-like particles produced in insect and yeast cells

  • Hyoung Jin Kim
  • Seo Young Cho
  • Min-Hye Park
  • Hong-Jin Kim
Research Article


Insect and yeast cells are considered the expression systems of choice for producing virus-like particles (VLPs), and numerous types of VLPs have been produced in these systems. However, previous studies were restricted to identifying the characteristics of individual VLP preparations. No direct comparison of the structures and immunogenic properties of insect and yeast-derived VLPs has so far been made. In the present study, the size distribution and immunogenic properties of human papillomavirus type 16 (HPV16) L1 VLPs produced in Spodoptera frugipedra-9 insect cells and Saccharomyces cerevisiae were compared. The insect cell-derived VLPs were larger than the yeast ones (P < 0.0001), with median sizes of 34 and 26 nm, respectively. In addition, the insect-derived VLPs appeared to be more diverse in size than the yeast-derived VLPs. Immunization of mice with 30 ng per dose of VLPs elicited 2.7- and 2.4-fold higher anti-HPV16 L1 IgG and anti-HPV16 neutralizing antibody titers than immunization with the same amounts of the yeast-derived VLPs after the 4th immunizations, respectively. Our results suggest that the choice of expression system critically affects the particle size and immunogenic property of HPV16 L1 VLPs.


Human papillomavirus Virus-like particles Vaccine Immunogenicity Spodoptera frugipedra-9 Saccharomyces cerevisiae 



This research was supported by a fund ‘(Grant no.: HD16A1337)’ by Research of Korea Centers for Disease Control and Prevention. This research was supported by the Chung-Ang University Research Scholarship Grants in 2017. We thank professor Ju-Won Kwak (Department of Life Sciences, Pohang University of Science and Technology) for providing pFastBac-HTb-HPV16 L1 and Yingji Jin (College of Pharmacy, Chung-Ang University) for her help with pseudovirus-based neutralizing assay.

Compliance with ethical standards

Conflicts of interest

The authors declared no conflicts of interest.

Supplementary material

12272_2018_1024_MOESM1_ESM.docx (295 kb)
Supplementary material 1 (DOCX 295 kb)


  1. Abdoli A, Soleimanjahi H, Fotouhi F, Teimoori A, Beiranvand SP, Kianmehr Z (2013) Human papillomavirus Type16-L1 VLP production in insect cells. Iran J Basic Med Sci 16:891–895PubMedPubMedCentralGoogle Scholar
  2. Buck CB, Day PM, Trus BL (2013) The papillomavirus major capsid protein L1. Virology 445:169–174CrossRefPubMedPubMedCentralGoogle Scholar
  3. Cardone G, Moyer AL, Cheng NQ, Thompson CD, Dvoretzky I, Lowy DR, Schiller JT, Steven AC, Buck CB, Trus BL (2014) Maturation of the human papillomavirus 16 capsid. Mbio 5:e01104–e01114PubMedPubMedCentralGoogle Scholar
  4. Carter JJ, Wipf GC, Benki SF, Christensen ND, Galloway DA (2003) Identification of a human papillomavirus type 16-specific epitope on the C-terminal arm of the major capsid protein L1. J Virol 77:11625–11632CrossRefPubMedPubMedCentralGoogle Scholar
  5. Chames P, Van Regenmortel M, Weiss E, Baty D (2009) Therapeutic antibodies: successes, limitations and hopes for the future. Br J Pharmacol 157:220–233CrossRefPubMedPubMedCentralGoogle Scholar
  6. Chang DY, Kim HJ, Kim HJ (2012) Effects of downstream processing on structural integrity and immunogenicity in the manufacture of papillomavirus type 16 L1 virus-like particles. Biotechnol Bioprocess Eng 17:755–763CrossRefGoogle Scholar
  7. Crosbie EJ, Einstein MH, Franceschi S, Kitchener HC (2013) Human papillomavirus and cervical cancer. Lancet 382:889–899CrossRefPubMedGoogle Scholar
  8. Effio CL, Hubbuch J (2015) Next generation vaccines and vectors: designing downstream processes for recombinant protein-based virus-like particles. Biotechnol J 10:715–727CrossRefPubMedGoogle Scholar
  9. Einstein MH, Baron M, Levin MJ, Chatterjee A, Edwards RP, Zepp F, Carletti I, Dessy FJ, Trofa AF, Schuind A, Dubin G (2009) Comparison of the immunogenicity and safety of Cervarix and Gardasil human papillomavirus (HPV) cervical cancer vaccines in healthy women aged 18-45 years. Hum Vaccin 5:705–719CrossRefPubMedGoogle Scholar
  10. Fernandes F, Teixeira AP, Carinhas N, Carrondo MJT, Alves PM (2013) Insect cells as a production platform of complex virus-like particles. Expert Rev Vaccines 12:225–236CrossRefPubMedGoogle Scholar
  11. Freivalds J, Dislers A, Ose V, Pumpens P, Tars K, Kazaks A (2011) Highly efficient production of phosphorylated hepatitis B core particles in yeast Pichia pastoris. Protein Expr Purif 75:218–224CrossRefPubMedGoogle Scholar
  12. Kanesashi SN, Ishizu K, Kawano M, Han S, Tomita S, Watanabe H, Kataoka K, Handa K (2003) Simian virus 40 VP1 capsid protein forms polymorphic assemblies in vitro. J Gen Virol 84:1899–1905CrossRefPubMedGoogle Scholar
  13. Karageosov I, Dimova R, Makaveeva V (1985) Electron microscopic detection of viruses in cervix papilloma. Zentralbl Gynakol 107:187–191PubMedGoogle Scholar
  14. Kim HJ, Kim H-J (2017a) Yeast as an expression system for producing virus-like particles: what factors do we need to consider? Lett Appl Microbiol 64:111–123CrossRefPubMedGoogle Scholar
  15. Kim HJ, Kim H-J (2017b) Current status and future prospects for human papillomavirus vaccines. Arch Pharm Res 40:1050–1063CrossRefPubMedGoogle Scholar
  16. Kim HJ, Kim SY, Lim SJ, Kim JY, Lee SJ, Kim H-J (2010) One-step chromatographic purification of human papillomavirus type 16 L1 protein from Saccharomyces cerevisiae. Protein Expr Purif 70:68–74CrossRefPubMedGoogle Scholar
  17. Kim HJ, Kwag HL, Jin Y, Kim H-J (2011) The composition of the carbon source and the time of cell harvest are critical determinants of the final yield of human papillomavirus type 16 L1 protein produced in Saccharomyces cerevisiae. Protein Expr Purif 80:52–60CrossRefPubMedGoogle Scholar
  18. Kim HJ, Lim SJ, Kwag HL, Kim H-J (2012) The Choice of resin-bound ligand affects the structure and immunogenicity of column-purified human papillomavirus type 16 virus-like particles. PLoS ONE 7:e35893CrossRefPubMedPubMedCentralGoogle Scholar
  19. Kim HJ, Jin Y, Kim H-J (2014) The concentration of carbon source in the medium affects the quality of virus-like particles of human papillomavirus type 16 produced in Saccharomyces cerevisiae. PLoS ONE 9:e94467CrossRefPubMedPubMedCentralGoogle Scholar
  20. Kim HJ, Kwag HL, Kim DG, Kang BK, Han SY, Moon H, Hwang JY, Kwon MG, Kang HA, Kim H-J (2016) Assembly of the capsid protein of red-spotted grouper nervous necrosis virus during purification, and role of calcium ions in chromatography. Biotechnol Bioprocess Eng 21:373–380CrossRefGoogle Scholar
  21. Le Tallec D, Doucet D, Elouahabi A, Harvengt P, Deschuyteneer M, Deschamps M (2009) Cervarix (TM), the GSK HPV-16/HPV-18 AS04-adjuvanted cervical cancer vaccine, demonstrates stability upon long-term storage and under simulated cold chain break conditions. Hum Vaccin 5:467–474CrossRefPubMedGoogle Scholar
  22. Mach H, Volkin DB, Troutman RD, Wang B, Luo Z, Jansen KU, Shi L (2006) Disassembly and reassembly of yeast-derived recombinant human papillomavirus virus-like particles (HPV VLPs). J Pharm Sci 95:2195–2206CrossRefPubMedGoogle Scholar
  23. Patel MC, Patkar KK, Basu A, Mohandas KM, Mukhopadhyaya R (2009) Production of immunogenic human papillomavirus-16 major capsid protein derived virus like particles. Indian J Med Res 130:213–218PubMedGoogle Scholar
  24. Rizk RZ, Christensen ND, Michael KM, Muller M, Sehr P, Waterboer T, Pawlita M (2008) Reactivity pattern of 92 monoclonal antibodies with 15 human papillomavirus types. J Gen Virol 89:117–129CrossRefPubMedGoogle Scholar
  25. Ryding J, Dahlberg L, Wallen-Ohman M, Dillner J (2007) Deletion of a major neutralizing epitope of human papillomavirus type 16 virus-like particles. J Gen Virol 88:792–802CrossRefPubMedGoogle Scholar
  26. Schadlicha L, Senger T, Kirschning CJ, Muller M, Gissmann L (2009) Refining HPV 16 L1 purification from E. coli: reducing endotoxin contaminations and their impact on immunogenicity. Vaccine 27:1511–1522CrossRefGoogle Scholar
  27. Shank-Retzlaff ML, Zhao QJ, Anderson C, Hamm M, High K, Nguyen M, Wang F, Wang N, Wang B, Wang Y, Washabaugh M, Sitrin R, Shi L (2006) Evaluation of the thermal stability of Gardasil (R). Hum Vaccin 2:147–154CrossRefPubMedGoogle Scholar
  28. Shirbaghaee Z, Bolhassani A (2016) Different applications of virus-like particles in biology and medicine: vaccination and delivery systems. Biopolymers 105:113–132CrossRefPubMedGoogle Scholar
  29. Van Oers MM, Pijlman GP, Vlak JM (2015) Thirty years of baculovirus-insect cell protein expression: from dark horse to mainstream technology. J Gen Virol 96:6–23CrossRefPubMedGoogle Scholar
  30. White WI, Wilson SD, Palmer-Hill FJ, Woods RM, Ghim SJ, Hewitt LA, Goldman DM, Burke SJ, Jenson AB, Koenig S, Suzich JA (1999) Characterization of a major neutralizing epitope on human papillomavirus type 16 L1. J Virol 73:4882–4889PubMedPubMedCentralGoogle Scholar
  31. Zahid M, Lunsdorf H, Rinas U (2015) Assessing stability and assembly of the hepatitis B surface antigen into virus-like particles during down-stream processing. Vaccine 33:3739–3745CrossRefPubMedGoogle Scholar
  32. Zahin M, Joh J, Khanal S, Husk A, Mason H, Warzecha H, Ghim SJ, Miller DM, Matoba N, Jenson AB (2016) Scalable production of HPV16 L1 protein and VLPs from tobacco leaves. PLoS ONE 11:e0160995CrossRefPubMedPubMedCentralGoogle Scholar
  33. Zeltins A (2013) Construction and characterization of virus-like particles: a review. Mol Biotechnol 53:92–107CrossRefPubMedGoogle Scholar
  34. Zhao QJ, Potter CS, Carragher B, Lander G, Sworen J, Towne V, Abraham D, Duncan P, Washabaugh MW, Sitrin RD (2014) Characterization of virus-like particles in GARDASIL (R) by cryo transmission electron microscopy. Hum Vaccin Immunother 10:734–739CrossRefPubMedGoogle Scholar

Copyright information

© The Pharmaceutical Society of Korea 2018

Authors and Affiliations

  • Hyoung Jin Kim
    • 1
  • Seo Young Cho
    • 1
  • Min-Hye Park
    • 1
  • Hong-Jin Kim
    • 1
  1. 1.Laboratory of Virology, College of PharmacyChung-Ang UniversitySeoulSouth Korea

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