Production of a covalent conjugate vaccine using cell-free protein expression (XpressCF™) and click chemistry

Abstract

Chlamydia trachomatis is the most common bacterial sexually transmitted infection globally and a vaccine is urgently needed. To address this, we determined that Chlamydia protease-like activity factor (CPAF) is the most immunoprevalent and immunodominant antigen for CD4 T cells and B cells in Chlamydia exposed women and mice. CPAF is a 70-kDa serine protease that is secreted into the host cell cytoplasm and subsequently released into the extracellular milieu where it paralyzes neutrophils to evade phagocytosis. CPAF contains a fold similar to other known serine proteases and is processed into C- and N-terminal fragments that heterodimerize. To improve and facilitate vaccine scale-up and immunogenicity, we developed CPAF variants incorporating non-native amino acids at specific sites using the proprietary cell free expression platform XpressCF™. These CPAF variants were covalently conjugated to DBCO-derivatized adjuvants using click chemistry to make novel antigen-adjuvant conjugate vaccines. Site-specific conjugation enables important T- and B-cell epitopes to be avoided for better immune recognition and provides a method to incorporate a specific number of conjugated adjuvants. These conjugated vaccines provide a means to simultaneously deliver antigen and adjuvant and potentially reduce the amount of adjuvant required for vaccine efficacy leading to fewer side effects and dose sparing.