Abstract
There is increasing demand for virus-like particles (VLPs) as a platform for prophylactic vaccine production. However, little attention has been paid to how downstream processing affects the structure and immunogenicity of the VLPs. In this study, we compared three methods of purifying human papillomavirus type 16 (HPV16) VLPs, each including the same cation-exchange chromatography (CEC) step. Method T-1 uses both ammonium sulfate precipitation (ASP) and a step to remove precipitated contaminating proteins (SRPC) prior to CEC, while T-2 uses only the SRPC step prior to CEC and T-3 includes neither step. We compared the structural integrity and immunogenicity of the HPV16 VLPs resulting from these three methods. All three preparations were highly pure. However, the final yields of the VLPs obtained with T-2 were 1.5 and 2 fold higher than with T-1 and T-3, respectively. With respect to structural integrity, T-1 and T-2 HPV16 VLPs had smaller hydrodynamic diameters and higher reactivity towards monoclonal anti-HPV16 neutralizing antibodies than T-3 VLPs, indicating higher potentials of T-1 and T-2 VLPs for eliciting anti-HPV16 neutralizing antibodies. Moreover, it was confirmed that the T-1 and T-2 HPV16 VLPs elicit anti-HPV16 neutralizing antibodies more efficiently than T-3 HPV16 VLPs do in mice immunizations: the abilities for eliciting neutralizing antibodies were in the order T-2 VLP > T-1 VLP > T-3 VLP. We conclude that the process design for purifying HPV VLPs is a critical determinant of the quality of the final product.
Article PDF
Similar content being viewed by others
Avoid common mistakes on your manuscript.
References
Morenweiser, R. (2005) Downstream processing of viral vectors and vaccines. Gene Ther. 12: 103–110.
Vicente, T., A. Roldao, C. Peixoto, M. J. Carrondo, and P. M. Alves (2011) Large-scale production and purification of VLP-based vaccines. J. Invertebr. Pathol. 107: 42–48.
Bachmann, A. S., G. Corpuz, W. P. Hareld, G. Wang, and B. A. Coller (2004) A simple method for the rapid purification of copia virus-like particles from Drosophila Schneider 2 cells. J. Virol. Methods 115: 159–165.
Pattenden, L. K., A. P. Middelberg, M. Niebert, and D. I. Lipin (2005) Towards the preparative and large-scale precision manufacture of virus-like particles. Trends Biotechnol. 23: 523–529.
Madrid-Marina, V., K. Torres-Poveda, G. Lopez-Toledo, and A. Garcia-Carranca (2009) Advantages and disadvantages of current prophylactic vaccines against HPV. Arch. Med. Res. 40: 471–477.
Conway, M. J. and C. Meyers (2009) Replication and assembly of human papillomaviruses. J. Dent. Res. 88: 307–317.
Seo, P. S., S. Y. Heo, E. J. Han, J. W. Seo, S. J. Ghim, and C. H. Kim (2009) Bacterial expression and purification of human papillomavirus type 18 L1. Biotechnol. Bioproc. Eng. 14: 168–174.
National Cancer Institute, Human Papillomavirus (HPV) Vaccines, http://www.cancer.gov/cancertopics/factsheet/prevention/HPV-vaccine.
Padmanabhan, S., T. Amin, B. Sampat, R. Cook-Deegan, and S. Chandrasekharan (2010) Intellectual property, technology transfer and manufacture of low-cost HPV vaccines in India. Nat. Biotechnol. 28: 671–678.
Walsh, G. (2010) Biopharmaceutical benchmarks 2010. Nat. Biotechnol. 28: 917–924.
National Cancer Institute (2007) Women’s Health Report, Fiscal Years 2005–2006. NCI Women’s Health Report FY2005-2006.
Kim, H. J., S. Y. Kim, S. J. Lim, J. Y. Kim, S. J. Lee, and H. -J. Kim (2010) One-step chromatographic purification of human papillomavirus type 16 L1 protein from Saccharomyces cerevisiae. Protein Expr. Purif. 70: 68–74.
Kim, H. J., S. J. Lee, and H. -J. Kim (2010) Optimizing the secondary structure of human papillomavirus type 16 L1 mRNA enhances L1 protein expression in Saccharomyces cerevisiae. J. Biotechnol. 150: 31–36.
Kim, H. J., H. L. Kwag, Y. Jin, and H. -J. Kim (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: 52–60.
Laemmli, U. K. (1970) Cleavage of structural proteins during the assembly of the head of bacteriophage T4. Nature 227: 680–685.
Kim, S. Y., H. J. Kim, and H. -J. Kim (2011) Simple and convenient chromatography-based methods for purifying the pseudovirus of human papillomavirus type 58. Protein Expr. Purif. 76: 103–108.
Mukherjee, S., M. V. Thorsteinsson, L. B. Johnston, P. A. DePhillips, and A. Zlotnick (2008) A quantitative description of in vitro assembly of human papillomavirus 16 virus-like particles. J. Mol. Biol. 381: 229–237.
Thones, N., A. Herreiner, L. Schadlich, K. Piuko, and M. Muller (2008) A direct comparison of human papillomavirus type 16 L1 particles reveals a lower immunogenicity of capsomeres than viruslike particles with respect to the induced antibody response. J. Virol. 82: 5472–5485.
Kim, S. N., H. S. Jeong, S. N. Park, and H. -J. Kim (2007) Purification and immunogenicity study of human papillomavirus type 16 L1 protein in Saccharomyces cerevisiae. J. Virol. Methods 139: 24–30.
Kim, H. J., J. K. Kim, S. B. Seo, H. J. Lee, and H. J. Kim (2007) Intranasal vaccination with peptides and cholera toxin subunit B as adjuvant to enhance mucosal and systemic immunity to respiratory syncytial virus. Arch. Pharm. Res. 30: 366–371.
Shi, L., G. Sanyal, A. Ni, Z. Luo, S. Doshna, B. Wang, T. L. Graham, N. Wang, and D. B. Volkin (2005) Stabilization of human papillomavirus virus-like particles by non-ionic surfactants. J. Pharm. Sci. 94: 1538–1551.
Culp, T. D. and N. D. Christensen (2004) Kinetics of in vitro adsorption and entry of papillomavirus virions. Virol. 319: 152–161.
Ryding, J., L. Dahlberg, M. Wallen-Ohman, and J. Dillner (2007) Deletion of a major neutralizing epitope of human papillomavirus type 16 virus-like particles. J. Gen. Virol. 88: 792–802.
Christensen, N. D., N. M. Cladel, C. A. Reed, L. R. Budgeon, M. E. Embers, D. M. Skulsky, W. L. McClements, S. W. Ludmerer, and K. U. Jansen (2001) Hybrid papillomavirus L1 molecules assemble into virus-like particles that reconstitute conformational epitopes and induce neutralizing antibodies to distinct HPV types. Virol. 291: 324–334.
Author information
Authors and Affiliations
Corresponding author
Rights and permissions
About this article
Cite this article
Chang, D.Y., Kim, H.J. & Kim, HJ. Effects of downstream processing on structural integrity and immunogenicity in the manufacture of papillomavirus type 16 L1 virus-like particles. Biotechnol Bioproc E 17, 755–763 (2012). https://doi.org/10.1007/s12257-012-0067-5
Received:
Revised:
Accepted:
Published:
Issue Date:
DOI: https://doi.org/10.1007/s12257-012-0067-5