Production of antibodies with peptide-CpG-DNA-liposome complex without carriers

Abstract

BackgroundThe screening of peptide-based epitopes has been studied extensively for the purpose of developing therapeutic antibodies and prophylactic vaccines that can be potentially useful for treating cancer and infectious diseases such as influenza virus, malaria, hepatitis B, and HIV. To improve the efficacy of antibody production by epitope-based immunization, researchers evaluated liposomes as a means of delivering vaccines; they also formulated adjuvants such as flagella and CpG-DNA to enhance the magnitude of immune responses. Here, we provide a potent method for peptide-based epitope screening and antibody production without conventional carriers.ResultsWe present that a particular form of natural phosphodiester bond CpG-DNA encapsulated in a specific liposome complex (Lipoplex(O)) induces potent immunomodulatory activity in humans as well as in mice. Additionally, Lipoplex(O) enhances the production of IgG2a specific to antigenic protein in mice. Most importantly, immunization of mice with several peptides co-encapsulated with Lipoplex(O) without carriers significantly induces each peptide-specific IgG2a production in a TLR9-dependent manner. A peptide-specific monoclonal antibody produced against hepatocellular carcinoma-associated antigen has functional effects on the cancer cells.ConclusionsOur overall results show that Lipoplex(O) is a potent adjuvant and that complexes of peptide and Lipoplex(O) are extremely useful for B cell epitope screening and antibody production without carriers. Therefore, our strategy may be promptly used for the development of therapeutic antibodies by rapid screening of potent B cell epitopes.

Activation of Toll-like receptor 9 and production of epitope specific antibody by liposome-encapsulated CpG-DNA

“…The PO-ODN, specifically MB-ODN 4531(O) containing immunomodulatory CpG motifs, regulates expression of Th1 cytokines and does not have severe side effects. We also found that the PO-ODN encapsulated in a phosphatidyl-β-oleoyl-γ-palmitoyl ethanolamine (DOPE): cholesterol hemisuccinate (CHEMS) complex (Lipoplex(O)) is able to enhance the stimulation of immune responses in the cells of humans and mice (15). Here, we investigated the effects of liposome-encapsulated PO-ODN or PS-ODN on TLR9-mediated signal transduction pathways and performed screening of B cell epitopes using several peptides and Lipoplex(O).…”

“…We refer to MB-ODN 4531(O) encapsulated in a DOPE : CHEMS) complex as Lipoplex(O). Lipoplex(O) induced the production of cytokines such as IL-6, IL-12, and IFN-γ in mouse splenocytes and hPBMCs, and also induced secretion of IL-12 and IFN-γ for much longer time periods than MB-ODN 4531(O) did by in vivo application, in a TLR9-dependent manner (15). In this study, we determined the effects of liposome-encapsulated MB-ODN 4531(S) (PS-ODN) on IL-8 promoter activity in a human B cell line (RPMI 8226) and the mouse macrophage cell line, RAW 264.7.…”

“…However, PO-ODN encapsulated in DOTAP can elicit HLA-DRA and hBD-2 promoter activation and gene expression in RPMI 8226 cells in a CG sequence-dependent manner (20). We also compared the effects of liposome-encapsulated PO-ODN on the IL-8 promoter activation and cytokine production in mouse cells and human cells (15). After stimulation with PO-ODN encapsulated in liposomes such as DOTAP, the DOPE : CHEMS complex, and the DSPC : CHEMS : PEG-PE complex, the IL-8 promoter was significantly activated in both human cells and mouse cells.…”

“…Previously, we have shown that the Lipoplex(O) can elicit en-http://bmbreports.org BMB reports hanced intracellular uptake, endosomal localization, and biological activity of CpG-DNA (15). As CpG-DNA-mediated immune activation was initiated by recognition of TLR9 (1), we decided to evaluate the effects of DOPE : CHEMS complex encapsulation on binding of CpG-DNA to TLR9 and established HEK 293 cells stably expressing TLR9.…”

“…However, the PO-ODN is susceptible to nuclease activity, and the immunostimulatory activity of PO-ODN is relatively lower than that of PS-ODN (4). Previously, we developed potent immunomodulatory PO-ODN from chromosomal DNA sequences of M. bovis and E. coli (15,16). The PO-ODN, specifically MB-ODN 4531(O) containing immunomodulatory CpG motifs, regulates expression of Th1 cytokines and does not have severe side effects.…”