Induction of an Antigen-Specific CTL Response by a Conformationally Biased Agonist of Human C5a Anaphylatoxin as a Molecular Adjuvant

A conformationally biased decapeptide agonist of human C5a anaphylatoxin (YSFKPMPLaR) was used as a molecular adjuvant in stimulating an Ag-specific CTL response against murine P815S target cells expressing an L d -restricted CTL epitope of the hepatitis B surface Ag (HBsAg). Groups of BALB/c mice (H-2 d ) were immunized with aqueous solutions of the HBsAg CTL epitopes ( IPQSLDSWWTSL and IPQSLDSWWTSL RR); the C5a agonist (YSFKPMPLaR); the C5a agonist and HBsAg CTL epitopes admixed ( IPQSLDSWWTSL and IPQSLDSWWTSL RR + YSFKPMPLaR); the C5a-active, HBsAg CTL epitope-C5a agonist constructs ( IPQSLDSWWTSL YSFKPMPLaR, IPQSLDSWWTSL RRYSFKPMPLaR, and IPQSLDSWWTSL RVRRYSFPMPLaR); a C5a-inactive, reverse-moiety construct (YSFKPMPLaRRR IPQSLDSWWTSL ); and a C5a-attenuated, carboxyl-terminal-blocked construct ( IPQSLDSWWTSL RRYSFKPMPLaRG). Ag-specific CD8 + CTL responses were observed after the secondary boost in the absence of any added adjuvant only in mice that were immunized with C5a-active contructs, IPQSLDSWWTSL RRYSFKPMPLaR and IPQSLDSWWTSL RVRRYSFKPMPLaR. These two C5a-active immunogens contained potential subtilisin-sensitive linker sequences between the HBsAg CTL epitope and the C5a agonist; i.e., a double-Arg (RR) and a furin protease sensitive sequence (RVRR). The introduction of these potentially cleavable sequences may be a method of increasing the likelihood of liberating the CTL epitope from the C5a agonist by intracellular proteases, thereby facilitating entry of the epitope into Ag-processing pathways via an exogenous route.