Increased Immunogenicity and Induction of Class Switching by Conjugation of Complement C3d to Pneumococcal Serotype 14 Capsular Polysaccharide

Abstract

Previous studies have demonstrated an adjuvant effect for the C3d fragment of complement C3 when coupled to T-dependent protein antigens. In this study, we examined the antibody response to covalent conjugates of C3d and a T-independent antigen, the capsular polysaccharide of serotype 14 Streptococcus pneumoniae (PPS14). We prepared a conjugate of mouse C3d and PPS14 and compared its immunogenicity with that of a conjugate of PPS14 and ovalbumin (OVA). When BALB/c mice were immunized with PPS14-C3d, there was a significant increase in serum anti-PPS14 concentrations compared with either native PPS14 or control PPS14-glycine conjugates. This was accompanied by a switch in anti-PPS14 from predominantly immunoglobulin M (IgM) to IgG1 by day 25 following primary immunization. Following secondary immunization with PPS14-C3d, there was a marked booster response and a further increase in the ratio of IgG1 to IgM anti-PPS14. Although the primary antibody response to the PPS14-OVA conjugate exceeded that induced by immunization with PPS14-C3d, serum anti-PPS14 concentrations after a second injection of PPS14-C3d were nearly identical to those induced by secondary immunization with PPS14-OVA. Experiments with athymic nude mice suggested that T cells were not required for the adjuvant effect of C3d on the primary immune response to PPS14 but were necessary for enhancement of the memory response after a second injection of PPS14-C3d. These studies show that the adjuvant effects of C3d extend to T-independent antigens as well as T-dependent antigens. As a means of harnessing the adjuvant potential of the innate immune system, C3d conjugates may prove useful as a component of vaccines against encapsulated bacteria.

C3d Enhances Immune Responses Using Low Doses of DNA Expressing the HIV-1 Envelope from Codon-Optimized Gene Sequences

“…The molecular adjuvant, C3d, has been shown to enhance the immunogenicity of viral, bacterial, and self antigens (T-dependent and T-independent) when inoculated as either DNA or protein in both inbred and outbred mice strains [4,5,12,16,17,19,21,24,31,32,35,36,38,39]. Overall, the inoculation of C3d fused antigens as protein immunogens seems to produce more consistent results than DNA expressing the same protein in-vivo.…”

“…Independently, each approach enhances antibody titer and cellular responses against Env compared to DNA plasmidsexpressing wild-type (wt) env gene inserts only. The fusion of C3d to an antigen results in enhanced immunogenicity to the fused antigen [5,16,17,19,21,24,31,32,35,36]. Rodents inoculated with DNA plasmids expressing HIV-1 Env fused to multiple copies of human or murine C3d (mC3d) had enhanced anti-Env specific IgG titers and accelerated affinity maturation of antibody [16].…”

In Vivo Humoral Immune Responses to Isolated Pneumococcal Polysaccharides Are Dependent on the Presence of Associated TLR Ligands

“…PS Ags may also activate complement in vivo (27), resulting in cocross-linking of mIg and the type 2 complement receptor (CD21). Although coligation of mIg and CD21 is synergistic for B cell activation (28), and linkage of C3d to isolated PS can enhance anti-PS responses in vivo (29), it is unclear whether B cell differentiation and Ig class switching will occur solely on the basis of mIg/CD21 cocross-linking. However, complement can activate numerous immune cell types, potentially leading to the release of mediators that promote B cell differentiation.…”

Expression of complement receptors 1 (CR1/CD35) and 2 (CR2/CD21), and co-signaling molecule CD19 in cattle

“…In experiments using athymic mice, C3d bound antigens also induced class switching from IgM to IgG for T independent antigens (Test et al, 2001).…”