Totally synthetic lipid-containing polyoxime peptide constructs are potent immunogens Weiguang Zeng a , David C. Jackson a, *, Julie Murray a , Keith Rose b , Lorena E. Brown a a Cooperative Research Centre for Vaccine Technology, Department of Microbiology and Immunology, The University of Melbourne, Parkville 3052, Vic., Australia b Biochimie Me´dicale, CMU, 1 rue Michel Servet, CH-1211 Geneva 4, Switzerland Received 31 May 1999; received in revised form 29 June 1999; accepted 15 July 1999 Abstract A synthetic peptide corresponding to a sequence from influenza hemagglutinin was used as a model antigen to study the immunogenicity of polyoxime constructs. In the absence of any adjuvant, tetrameric forms of dierent polyoxime constructs did not elicit an antibody response. High and long-lasting levels of antibody were induced, however, by polyoxime constructs to which Pam3Cys (tripalmitoyl-S-glyceryl cysteine) was attached. Comparable serum antibody levels were achieved with Tetraoxime-Pam3Cys administered by the intraperitoneal or intranasal routes to those obtained when the monomeric peptide was administered by the intraperitoneal route in complete Freund’s adjuvant (CFA). Mice receiving Tetraoxime-Pam3Cys and Pam3Cys-peptide intranasally developed peptide-specific antibody secreting cells (ASCs) in their lungs and mediastinal lymph nodes. At low dose, the Tetraoxime-Pam3Cys induced higher levels of antibody compared to those elicited by the monomeric Pam3Cys-peptide delivered by either route. These results show that lipo-tetraoxime constructs assembled by polyoxime chemistry can be potent inducers of systemic and mucosal immunity. # 2000 Elsevier Science Ltd. All rights reserved. Keywords: Peptide; Polyoxime; Lipid 1. Introduction In recent years there have been many reports describing the ability of peptide dendrimers to elicit immune responses and the potential of these constructs as vaccine candidates [1–4]. Compared to monomeric peptide, epitopes in the form of dendrimers, for example the multiple antigen peptide (MAP) system described by Tam et al. [1], often showed a significant increase in immunogenicity. Incorporation into these MAPs of a lipid with known adjuvant properties pro- duces a totally synthetic and self-adjuvanting vaccine candidate that not only induces antibody and cellular responses but does so in the absence of additional adjuvanting substances [5–7]. While these self-adju- vanting peptide dendrimers may circumvent the pro- blems associated with harmful and inadequate extraneous adjuvants, the ability to prepare these large and complex constructs by conventional synthetic methods is challenging [4,6]. Recently, chemical ligation [8–11] has been used in the synthesis of large complex molecules with defined structure. One such ligation strategy employs polyox- ime chemistry where unprotected peptides, functiona- lised to carry either aldehyde or aminooxy groups, react with an appropriate template through formation of oxime bonds to form homogeneous peptide dendri- mers [11,12]. This method has the advantages of using very mild conditions, producing a very pure product in good yield, the ability to incorporate native proteins [13] and unlike other methods, does not involve the use of sulfur which is prone to oxidation [12]. Using this approach polypeptide dendrimers have been syn- Vaccine 18 (2000) 1031–1039 0264-410X/00/$ - see front matter # 2000 Elsevier Science Ltd. All rights reserved. PII: S0264-410X(99)00346-1 www.elsevier.com/locate/vaccine * Corresponding author. Tel.: +61-3-9342-9819; fax: +61-3-9342- 9782. E-mail address: d.jackson@microbiology.unimelb.edu.au (D.C. Jackson).