Use of Zika virus recombinant proteins for development of diagnostic methods and vaccines.

Author(s)

Lennon Ramos Pereira

• Document type: Doctoral Thesis
• Press: São Paulo.
• Institution: Universidade de São Paulo (USP). Instituto de Ciências Biomédicas (ICB/SDI)
• Defense date: 2021-11-29

Examining board members

Luis Carlos de Souza Ferreira; Andrea Balan Fernandes; Wagner Quintilio; Carla Torres Braconi dos Santos

• Advisor: Luis Carlos de Souza Ferreira

Abstract

Zika virus (ZIKV) infection is acute and self-limited. However, some infected individuals have severe neurological manifestations, including Guillain-Barrée Syndrome and Congenital Zika Syndrome (CZS). Despite more than 70 years of studies, there are no specific therapeutic or prophylactic methods for ZIKV. In addition, the high antigenic similarity among ZIKV and other arboviruses makes it essential to establish serological tests that can effectively differentiating previous infection by these viruses. In this context, this project aimed to obtain and evaluate the serological specificity and vaccine potential of recombinant forms of the domain III envelope protein (ZIKV EDIII) and non-structural protein 1 of ZIKV (ZIKV NS1). Thus, the proposed recombinant proteins were expressed in a prokaryote system, purified and validated for preservation of antigenicity. Additionally, a fragment of the ZIKV NS1 protein, termed ZIKV &#916NS1, was designed by computational analysis based on sequence and structural homology compared to the corresponding region of NS1 of DENV1-4. The obtained proteins were used as solid phase antigens in the standardization of ELISA protocols aiming at the specific detection of IgG anti-ZIKV antibodies. The obtained results demonstrated that the implementation of the &#916NS1 ZIKV protein, but not the EDIII ZIKV, significantly reduced the detection of cross-reactive antibodies directed to other Flaviviruses, especially to DENV. Furthermore, the observed specificity was correlated with the presence of thermolabile conformational epitopes in the ZIKV &#916NS1. Given the promising results, the ELISA protocol based on the ZIKV &#916NS1 protein (ELISA ZIKA-v IgG) was adapted and transposed to conditions of a commercial kit in a public-private partnership. The validation of the ZIKA-v IgG ELISA against a human serum panel showed specificity and sensitivity of 94.12% and 90.7%, respectively, for which the marketing was approved by ANVISA. Furthermore, vaccine formulations based on ZIKV NS1 and EDIII proteins, in association with different adjuvants (Alum, LT-B or Poly [I:C]), were tested in a murine model. Although immunogenic, they were not able to provide protective immunity against ZIKV infection. Finally, we used a novel DNA vaccine strategy (pgDNS1-ZIKV) based on the expression of the ZIKV NS1 protein genetically fused to the glycoprotein D of the Herpes Simplex virus Type I (gD HSV-1). Interestingly, this approach was able to improve the induction of NS1-specific humoral and cellular immune responses, as well as increasing the protective efficacy against ZIKV infection. Thus, with the knowledge generated from this work, we hope to contribute to the improvement of serodiagnostic methods and vaccine strategies based on DNA vaccines for the ZIKV. (AU)

• FAPESP's process: 16/05570-8 - Use of Zika virus recombinant proteins for development of diagnostic methods and vaccines
• Grantee: Lennon Ramos Pereira
• Support Opportunities: Scholarships in Brazil - Doctorate