Exp Mol Pathol. 2015 Oct;99(2):330-4. doi: 10.1016/j.yexmp.2015.06.021.

Targeting synthetic Human Papillomavirus (HPV) L2 disulfide-induced N-terminus conformational epitopes for pan-HPV vaccine development.

 

Khanal S1, Ferraris ED2, Zahin M3, Joh J4, Ghim SJ5, Jenson AB6.
  • 1Department of Biochemistry and Molecular Genetics, University of Louisville School of Medicine, Louisville, KY, USA; James Graham Brown Cancer Center, University of Louisville School of Medicine, Louisville, KY, USA. Electronic address: sujita.khanal@louisville.edu.
  • 2James Graham Brown Cancer Center, University of Louisville School of Medicine, Louisville, KY, USA; Emergency Medicine Associate, Vancouver, WA, USA. Electronic address: ericdferraris@gmail.com.
  • 3James Graham Brown Cancer Center, University of Louisville School of Medicine, Louisville, KY, USA. Electronic address: m0zahi01@louisville.edu.
  • 4James Graham Brown Cancer Center, University of Louisville School of Medicine, Louisville, KY, USA; Department of Medicine, University of Louisville School of Medicine, Louisville, KY, USA. Electronic address: j0joh001@louisville.edu.
  • 5James Graham Brown Cancer Center, University of Louisville School of Medicine, Louisville, KY, USA; Department of Medicine, University of Louisville School of Medicine, Louisville, KY, USA. Electronic address: sjghim01@louisville.edu.
  • 6James Graham Brown Cancer Center, University of Louisville School of Medicine, Louisville, KY, USA. Electronic address: abjens01@louisville.edu.

 

Abstract

BACKGROUND: Current vaccines against Human Papillomavirus (HPV) are highly effective and based on recombinant virus-like particles (VLPs) of the major capsid protein L1. Since these vaccines are HPV type-specific and expensive for global implementation, an alternative, broader-spectrum immunogen would be the N-terminus of the minor capsid protein L2 that induces low titered broadly cross-neutralizing antibodies. Here we analyzed the reactivity of different synthetic L2 peptides containing N-terminus amino acids 17-36 in order to test their antigenicity.

METHODS: Different synthetic peptides were designed to target the 17-36 amino acid sequences, present in highly antigenic amino-terminus of L2 protein. Six different peptides including Cys22-Cys28 disulfide bonded cyclized L2 peptide were examined for their antigenicity against mouse monoclonal antibody RG-1 and rabbit polyclonal antisera to HPV L2 by enzyme-linked immunosorbent assay (ELISA).

RESULTS: Here we report that the cyclized form of synthetic L2 peptide, which is formed through Cys22-Cys28 disulfide bridges, has the highest reactivity to antibodies than other synthetic L2 peptides.

CONCLUSION: A cyclized L2 peptide has potential to be an excellent candidate to formulate a low-cost, broadly protective pan-oncogenic HPV vaccine.

KEYWORDS: Cyclized synthetic peptide; Human Papillomavirus (HPV); L2 protein; Neutralizing antibody

PMID: 26134615

 

Supplements:

The burden of human papillomavirus (HPV)-induced cancers (particularly cervical and oropharyngeal cancers) occurs worldwide. FDA approved HPV vaccines -Gardasil and Cervarix- contain recombinant virus-like-particles (VLPs) that self-assemble into the major HPV capsid protein, L1. These vaccines work well but are HPV type-specific and most importantly very expensive, which limits their global implementation. Therefore, there is a need for an inexpensive and efficient HPV vaccine to reduce global burden of HPV-associated cancers and our study focused on finding the possible candidate target.

HPV minor capsid protein, L2 could be an ideal target for the development of a cost-effective and broad-spectrum HPV vaccine. Amino acids (or epitopes) present in N-terminus of L2 are highly conserved among HPV types and are capable of inducing antibodies (1). However, the antibody titers induced are significantly lower compared to highly immunogenic L1 VLP (1,2), which is the main challenge in developing a L2-based vaccine. Thus, our study sought to improve the antigenicity of L2 by using synthetic disulfide bonded cyclized L2 peptide. We hypothesized that disulfide-induced cyclization of L2 peptide confers conformational stability and makes L2 highly antigenic.

We designed synthetic peptides covering highly conserved 17–36 amino acid residues of L2 protein and examined their antigenicity. We found that cyclized L2 peptide (which is formed through Cysteine22–Cysteine28 disulfide bridges) has the highest antigenic property as compared to other synthetic peptides. We further observed that the reactivity of non-cyclized peptide significantly increased in the presence of hydrogen peroxide, which induces disulfide bond formation. These results indicate the importance of oxidation or disulfide bond formation for the reactivity of L2 peptides. Our study suggests the requirement of disulfide-bonded L2 structure to enhance its reactivity and thus can be a potential target for L2-based HPV vaccine development. Further study on the cyclized L2 may open the door to develop the L2-based low-cost and broadly protective HPV vaccine.

 

References:

1.      Roden, R.B., Yutzy, W.H., Fallon, R., Inglis, S., Lowy, D.R., Schiller, J.T., 2000. Minor capsid protein of human genital papillomaviruses contains subdominant, cross-neutralizing epitopes. Virology 270 (2), 254–257.

2.      Pastrana, D.V., Gambhira, R., Buck, C.B., Pang, Y.Y., Thompson, C.D., Culp, T.D., et al., 2005. Cross-neutralization of cutaneous and mucosal papillomavirus types with anti-sera to the amino terminus of L2. Virology 337 (2), 365–372.

 

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