Bordetella pertussis and its components as delivery systems and adjuvants for the development of vaccines against Streptococcus pneumoniae

Author(s)

Júlia Tavares de Castro

• Document type: Doctoral Thesis
• Press: São Paulo.
• Institution: Universidade de São Paulo (USP). Instituto de Ciências Biomédicas (ICB/SDI)
• Defense date: 2019-11-28

Examining board members

Maria Leonor Sarno de Oliveira; Renato Mancini Astray; Ângela Silva Barbosa; Viviane Fongaro Botosso; Ivana Barros de Campos; Ana Paula de Mattos Arêas Dau

• Advisor: Maria Leonor Sarno de Oliveira; Alessandra Soares Schanoski

Abstract

Streptococcus pneumoniae (pneumococcus) is a major cause of pneumonia, meningitis and sepsis. About 400,000 children under the age of 5 die each year from pneumococcal infections worldwide. Pneumococcus Surface Protein A (PspA) is a well-characterized antigen that confers protection in animal models and represents a good alternative to current conjugate vaccines. However, few low-cost adjuvants have been proposed to date. In previous studies, our group has shown that the whole cell pertussis vaccine (wP) is a good adjuvant when tested in combination with PspA, inducing high antibody levels and protection against invasive challenges and pneumococcal nasal colonization in mice. These results support strategies for the development of combined vaccines for protection against pertussis and pneumococcus. In this work, two strategies for the composition of dual vaccines were tested. In the first, the PspA4Pro antigen (N-terminal region of PspA from clade 4) was expressed in the Bordetella pertussis vaccine strain for the production of an inactivated recombinant vaccine (wPPspA4Pro). PspA4Pro was expressed in fusion with the N-terminal portion of B. pertussis FHA (Filamentous hemagglutinin A) adhesin to target the surface of the bacterium. Expression was confirmed by western blot and different clones were used for the production of wPPspA4Pro vaccines by inactivation with formaldehyde. Immunization of BALB/c mice with different wPPspa4Pro vaccine preparations, at up to three doses, induced low levels of anti-PspA4Pro IgG in the sera. In addition, protection of vaccinated mice against a lethal pneumococcal challenge was not observed. Thus, the wPPspA4Pro recombinant vaccine was not effective against pneumococcal infections. The second strategy was to use an antigen delivery system based on B. pertussis adenylate cyclase toxin (CyaA). CyaA is capable of binding to receptors present on antigen presenting cells, modulating the immune response against associated antigens. Different PspA fragments from clades 2 and 4 (F1, F2, F4 and F5), as well as complete N-terminal regions (PspA2Pro and PspA4Pro) were cloned into a permissive region of the CyaA gene. Recombinant proteins were expressed in E. coli and purified by chromatographic methods. BALB/c mice were immunized in independent experiments with different CyaA-PspA proteins. CyaA-PspA4-F4 and CyaA-PspA4-F5 proteins were more immunogenic, inducing higher levels of anti-PspA4Pro IgG when compared to CyaA-PspA4-F1 and CyaA-PspA4-F2 proteins. In addition, 100 % of mice vaccinated with the CyaA-PspA4-F5 protein were protected against the lethal pneumococcal challenge, indicating that this antigen is a promising candidate for vaccine development. Thus, the CyaA-PspA2-F5 and CyaA-PspA4-F5 proteins were tested in additional experiments and the results showed the induction of high antibody levels and protection of mice against challenges with pneumococcal strains expressing PspAs from clade 2 and 4, respectively. The combination of the two proteins was protective against both challenges and was therefore a formulation with potential broader coverage of isolates. The protective potential against B. pertussis infections has yet to be evaluated. (AU)

• FAPESP's process: 16/17258-9 - Bordetella pertussis and its components as delivery systems and adjuvants for vaccine development against Streptococcus pneumoniae
• Grantee: Júlia Tavares de Castro
• Support Opportunities: Scholarships in Brazil - Doctorate (Direct)