Abstract
There is an increasingly severe trend of antibiotic-resistant Neisseria gonorrhoeae strains worldwide and new therapeutic strategies are needed against this sexually-transmitted pathogen. Despite the urgency, progress towards a gonococcal vaccine has been slowed by a scarcity of suitable antigens, lack of correlates of protection in humans and limited animal models of infection. N. gonorrhoeae gene expression levels in the natural human host does not reflect expression in vitro, further complicating in vitro-basedvaccine analysis platforms. We designed a novel candidate antigen selection strategy (CASS), based on a reverse vaccinology-like approach coupled with bioinformatics. We utilized the CASS to mine gonococcal proteins expressed during human mucosal infection, reported in our previous studies, and focused on a large pool of hypothetical proteins as an untapped source of potential new antigens. Via two discovery and analysis phases (DAP), we identified 36 targets predicted to be immunogenic, membrane-associated proteins conserved in N. gonorrhoeae and suitable for recombinant expression. Six initial candidates were produced and used to immunize mice. Characterization of the immune responses indicated cross-reactive antibodies and serum bactericidal activity against different N. gonorrhoeae strains. These results support the CASS as a tool for the discovery of new vaccine candidates.
Highlights
Neisseria gonorrhoeae is the causative agent of the sexually transmitted infection (STI) gonorrhea, a multi-faceted disease with high morbidity worldwide and an estimated 87 million cases annually [1].N. gonorrhoeae infections in men are mostly symptomatic, while gonorrhea in women is often asymptomatic, leading to reproductive tract complications (pelvic inflammatory disease (PID), ectopic pregnancy, infertility) and disseminated gonococcal infections (DGI) [2]
Our studies highlighted three important observations: (1) the gonococcus responds to the male and female reproductive tract environments by expressing genes at different levels, (2) gonococcal genes are expressed and regulated differently in vivo and in vitro (genes detected as more expressed in vivo than in vitro were termed IVEFs) and (3) a large number of gonococcal genes expressed during human infection encode hypothetical proteins
A multi-pronged candidate antigen selection strategy (CASS) was designed for antigen mining by screening predicted immunogenicity, surface-exposure, sequence conservation/variability and structure features combined with expression levels
Summary
Neisseria gonorrhoeae is the causative agent of the sexually transmitted infection (STI) gonorrhea, a multi-faceted disease with high morbidity worldwide and an estimated 87 million cases annually [1]. Our studies highlighted three important observations: (1) the gonococcus responds to the male and female reproductive tract environments by expressing genes at different levels, (2) gonococcal genes are expressed and regulated differently in vivo and in vitro (genes detected as more expressed in vivo than in vitro were termed IVEFs (in vivo expressed factors)) and (3) a large number of gonococcal genes expressed during human infection encode hypothetical proteins Taking this information into account, we have designed a comprehensive, high-throughput in silico approach for vaccine antigen identification that combines gene expression levels with known antigen prerequisites such as immunogenicity, membrane association/surface exposure, conservation and structure features. Our results show induction of cross-reactive antibodies against diverse N. gonorrhoeae strains by the candidates and serum bactericidal activity These initial studies support the CASS for identification of gonococcal proteins not previously explored as vaccine antigens
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