Increasing the Antigenicity of Synthetic Tumor‐Associated Carbohydrate Antigens by Targeting Toll‐Like Receptors

Abstract

Epithelial cancer cells often over express mucins that are aberrantly glycosylated. Although it has been realized that these compounds offer exciting opportunities for the development of immunotherapy for cancer, their use is hampered by the low antigenicity of classical immunogens composed of a glycopeptide derived from a mucin conjugated to a foreign carrier protein. We have designed, chemically synthesized, and immunologically evaluated a number of fully synthetic vaccine candidates to establish a strategy to overcome the poor immunogenicity of tumor-associated carbohydrates and glycopeptides. The compounds were also designed to study in detail the importance of TLR engagement for these antigenic responses. We have found that covalent attachment of a TLR2 agonist, a promiscuous peptide T-helper epitope, and a tumor-associated glycopeptide, gives a compound (1) that elicit in mice exceptionally high titers of IgG antibodies which recognize MCF7 cancer cells expressing the tumor-associated carbohydrate. Immunizations with glycolipopeptide (2), which contains lipidated amino acids instead of a TLR2 ligand, gave significantly lower titers of IgG antibodies demonstrating that TLR engagement is critical for optimum antigenic responses. Although mixtures of compound 2 with Pam3CysSK4 (3) or monophosphoryl lipid A (4) elicited similar titers of IgG antibodies compared to 1, the resulting antisera had an impaired ability to recognize cancer cells. It was also found that it is essential to covalently link the helper T-epitope to B-epitope probably because internalization of the helper T-epitope by B-cells requires assistance of the B-epitope. The results presented here show that synthetic vaccine development is amenable to structure activity relationship studies for successful optimization of carbohydrate-based cancer vaccines.

Translating Tumor Antigens into Cancer Vaccines

“…In particular, keyhole limpet hemocyanin (KLH) has been shown to be the most effective carrier for TACAs (86), and KLH conjugates of GM2 for melanoma (27) as well as those of sTn for breast cancer (73) have both entered phase III clinical trials. However, the disappointing outcomes of these vaccine clinical trials, in terms of time-to-disease progression and overall survival, have driven the generation of several forms of fully synthetic carbohydrate vaccines, which have been shown to be immunogenic regardless of the use of a protein carrier or external adjuvant (6,13,22,76,176,194). Unfortunately, despite all efforts, TACA-based cancer vaccines have failed to induce sufficient T cell-mediated immune responses in cancer patients, and none has been approved for clinical use yet (64).…”

Antitumor Vaccines Based on Synthetic Mucin Glycopeptides

“…An example of a MUC1 glycopeptide vaccine conjugated to both an immunostimulant and a T-cell epitope will here be described (Ingale et al 2009;Ingale et al 2006Ingale et al , 2007 The used stimulant, S-[(R)-2,3-dipalmitoyloxy-propyl]-N-palmitoyl-(R)-cysteine (Pam 3 Cys) (Bessler et al 1998) is a ligand of the Toll-like receptor 2. Through interaction with the receptor, pro-inflammatory cytokines and chemokines are produced, which stimulates the APCs that can process and present the immunogenic Scheme 11.11 Synthesis of a unimolecular pentavalent vaccine epitopes contained in the vaccine to the T-cells.…”

Molecular Recognition Elements for Toxin and Pathogen Detection

“…Because antibodies are generated in animals, it can be very difficult to raise them against molecules that are not well tolerated by the animal. Molecules of interest must also be sufficiently immunogenic if the desired antibody is to be generated [43,44]. They can be quite labor intensive and expensive to produce and identify.…”