TACA is a promising epitope for anticancer vaccines targeting carbohydrates. Similar to HIV, carbohydrates are considered antigens capable of inducing a specific and effective IgG response. However, TACA typically demonstrates weak immunogenicity as a T cell-independent type II antigen, and it also exhibits low stability and availability in vivo. To address these challenges, glycoconjugate vaccines utilize immunogenic carriers with diverse structures. One such carrier with significant potential is KLH.
KLH is an immunogenic protein antigen that is recognized as foreign by the mammalian immune system. It possesses potent immunostimulatory properties in both animals and humans. KLH offers numerous primary amine groups that facilitate protein conjugation, allowing it to be used for immunizing animals and generating antibodies against small molecules through covalent conjugation. With 300-600 available lysine amine groups, KLH serves as an excellent carrier protein.
Fig.1 Unimolecular trivalent vaccine construct. (Ouerfelli, et al., 2005)
Globo-H antigen is first isolated from breast cancer cell line MCF-7. It is overexpressed on the surface of many common cancers, including breast, lung, gastric, and pancreatic cancers. This new class of tumor-associated antigens has potent immunological activity.
CD BioGlyco provides the preparation service of Globo H-KLH conjugates. Globo H hexasaccharide is synthesized using a glycosyl assembly method. The structure is equipped with a terminal allyl group (instead of a ceramide). The allyl group at the end of the fully synthesized structure is converted to an aldehyde group by ozonolysis and combined with the -NH2 group of the bifunctional crosslinker 4-(4-N-maleimethylene) cyclohexane-1-carbonyl hydrazine connected. Thiolation of KLH by treatment with 2-iminothiolane converts the -NH2 group in KLH to a sulfhydryl group. Activated Globo H is then mixed with thiolated KLH and incubated for 2 hours at room temperature. Wash Globo H-KLH conjugate and filter to confirm sterility. The conjugated article is then aseptically filled into individual vials containing a certain mass of saline.
Globo H and KLH molar ratios are determined by high pH anion-exchange chromatography with pulsed amperometric detection (HPAEC-PAD) assay of carbohydrates after acid hydrolysis and by Bio-Rad assay of proteins. The vaccine conjugate will be co-administered with QS-21.
Fig.2 Preparation of Globo H-KLH conjugates. (CD BioGlyco)
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