Expression of Chimeric HPV-HIV Protein L1P18 in Pichia pastoris; Purification and Characterization of the Virus-like Particles.

Yoshiki Eto,Pau Ferrer,Narcís Saubi,Joan Joseph-Munné

doi:10.3390/pharmaceutics13111967

Abstract

Currently, three human papillomavirus (HPV) vaccines are already licensed and all of them are based on virus-like particles (VLPs) of HPV L1 capsid protein but not worldwide accessible. While about 38.0 million people were living with HIV in 2019, only 68% of HIV-infected individuals were accessing antiretroviral therapy as of the end of June 2020 and there is no HIV vaccine yet. Therefore, safe, effective, and affordable vaccines against those two viruses are immediately needed. Both HPV and HIV are sexually transmitted infections and one of the main access routes is the mucosal genital tract. Thus, the development of a combined vaccine that would protect against HPV and HIV infections is a logical effort in the fight against these two major global pathogens. In this study, a recombinant Pichia pastoris producing chimeric HPV-HIV L1P18 protein intracellularly was constructed. After cell disruption, the supernatant was collected, and the VLPs were purified by a combination of ammonium sulfate precipitation, size exclusion chromatography, ultracentrifugation, and ultrafiltration. At the end of purification process, the chimeric VLPs were recovered with 96% purity and 9.23% overall yield, and the morphology of VLPs were confirmed by transmission electron microscopy. This work contributes towards the development of an alternative platform for production of a bivalent vaccine against HPV and HIV in P. pastoris.

Highlights

More than 100 types of human papilloma virus (HPV) are already known and the HPV genotypes 16 and 18 are considered to be responsible for about 70% of cervical cancer worldwide [1,2,3]
The human immunodeficiency virus (HIV)-1 P18I10 peptide was inserted into the loop D-E of the HPV-16 L1 protein because Sadeyen et al [39] showed that Hepatitis B virus capsid (HBc) protein was more immunogenic when it was inserted in the D-E loop of HPV-16 L1 compared to other loops (Figure 1A)
The D-E loop is exposed to the exterior when 360 L1 proteins form a virus-like particles (VLPs) so that the inserted HIV-1 P18I10 proteins are supposed to be situated on the surface of the chimeric HPV-HIV VLP (Figure 1B)

Summary

Introduction

More than 100 types of human papilloma virus (HPV) are already known and the HPV genotypes 16 and 18 are considered to be responsible for about 70% of cervical cancer worldwide [1,2,3]. Gardasil and Gardasil 9 are produced by Saccharomyces cerevisiae (one type of yeast expression system) while the other one, Cervarix, is produced by baculovirus expression system. These three are still prohibitively expensive for the majority of women in developing countries and the cervical cancer caused by HPV is a lasting menace in those areas [7,8,9,10]. As for human immunodeficiency virus (HIV), in 2019, about 38.0 million people were living with HIV, 1.7 million individuals became newly infected with HIV, including.

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Results

Conclusion