SYNTHESIS OF MULTI-EPITOPIC GLYCOPEPTIDE-BASED CANCER VACCINES - Biologically-Responsive Hybrid Biomaterials - page 152

thus designed vaccine prototypes 38 combining a cluster of Tn antigen on the
upper domain of the template and helper T-cell peptide epitope from the type I-
poliovirus protein (PV) on the second addressable domain (Figure 15). A linear
orthogonally protected decapeptide was first prepared using an automated
peptide synthesizer on an acido-labile support such as the SASRIN resin. After
cleavage of the linear peptide, the head-to-tail cyclization was achieved by
treatment with PyBOP (benzotriazol-1-yl-oxytripyrrolidinophosphonium
hexafluorophosphate) under high dilution to prevent intermolecular side
reactions. The protecting groups were next removed regioselectively, then
protected serine residues as masked aldehydes [74] and aminooxy acetic acid
were incorporated to the lysine side chain pointing on the upper and lower
addressable domains respectively. The full deprotection of the template by
acidolysis provides 39 on which PV peptide functionalised with aldehyde at N -
terminus can be coupled by oxime ligation. This coupling reaction was
performed in sodium acetate buffer pH 4. Further oxidative cleavage of serines
by treatment with sodium periodate liberates four glyoxo-aldehyde functions that
were finally used for linking aminooxy Tn. The final glycocluster 38 displaying
PV and Tn was thus obtained in excellent yield and purity. Biological
investigations clearly showed that the cyclopeptide template revealed an efficient
non-immunological carrier molecule. Moreover, this multi-epitopic construction
elicited a specific immune response towards tumors expressing the human form
of Tn antigen, highlighting his potential for the design of more elaborated tumor
vaccine prototypes.
A new generation of TACA-based vaccine prototype was indeed optimised
recently [75-76]. For the first time, four essential components for immune
activation were assembled within the same molecular construction 40 (Figure
15). This molecule contains: a cluster of carbohydrate antigens to elicit a B-cell
response and trigger high affinity tumor-specific antibodies; T helper and T
cytotoxic epitopes to ensure both humoral and cellular eradication of tumors; a
palmitic acid rather than potentially toxic external immunoadjuvants to provide
self-adjuvanting properties. Therefore, a chimeric CTL-Th peptide with a
cysteine at C -terminus, composed of a T-cell epitope from ovalbumin
(OVA 257-264 ) in line with a universal T helper epitope (PADRE) and bearing a
palmitic acid moiety at the N -terminal end was prepared. This lipopeptide was
inserted to a cyclopeptide template 41 displaying oxime-linked clustered Tn
analogues using disulfide bond formation. The resulting glycolipopeptide 40
was finally purified by RP-HPLC and charaterized by mass spectroscopy.
Immunological investigations with OVA-expressing mouse B16 melanoma