Induction of an antigen-specific CTL response by a conformationally biasedagonist of human C5a anaphylatoxin as a molecular adjuvant

A conformationally biased decapeptide agonist of human C5a anaphylatoxin (Y SPKPMPLaR) was used as a molecular adjuvant in stimulating an Ag-specific C TL response against murine P815S target cells expressing an Ld-restricted C TL 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 (IPQS LDSWWTSL and IPQSLDSTaVTSLRR); the C5a agonist (YSFKPMPLaR); the C5a agonis t and HBsAg CTL epitopes admired (IPQSLDSWWTSL and IPQSLDSWWTSLRR + YSFKPMP LaR); the C5a-active, HBsAg CTL epitope-C5a agonist constructs (IPQSLDSWWTS LYSFKPMPLaR, IPQSLDSWWTSLRRYSFKPMPLaR, and IPQSLDSWWTSLRVRRYSFPMPLaR); a C5 a-inactive, reverse-moiety construct (YSFKPMPLaRRRIPQSLDSWWTSL); and a C5a- attenuated, carboxyl-terminal-blocked construct (IPQSLDSWWTSLRRYSFKPMPLaRG) . 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, IPQSLDSWWTSLRRYSFKPMPLaR and IPQSLDSWWTSLRVRRYSFKPMPL aR. These two C5a-active immunogens contained potential subtilisin-sensitiv e 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 intr oduction of these potentially cleavable sequences may be a method of increa sing the likelihood of liberating the CTL epitope from the C5a agonist by i ntracellular proteases, thereby facilitating entry of the epitope into Ag-p rocessing pathways via an exogenous route.