Development of a new vaccine formulation that enhances the immunogenicity of tumor-associated antigen CaMBr1

Abstract

Aberrant glycosylation is one of the most constant traits of malignant cells. The CaMBr1 hexasaccharide antigen, originally defined on the human breast carcinoma cell line MCF7, is expressed on some normal tissues but overexpressed in a high percentage of human breast, ovary, prostate and lung carcinomas. CaMBr1 overexpression is associated with poor prognosis. The epitope consists of the tetrasaccharide Fuc(alpha1-2)Ga1(beta1-3)GalNAc(beta1-3)Ga1alpha-O-spacer, which has recently become available as a synthetic oligosaccharide. Here we report the CaMBr1 tetrasaccharide conjugation to two different carrier proteins (CRM197 and KLH) and the evaluation of conjugate immunogenicity in mice following their administration in various vaccine formulations with two adjuvants (MPL-SE and Detox-PC). Radioimmunoassay to determine the level and isotype of anti-tetrasaccharide antibodies in mouse sera, and cytofluorimetric analysis and 51Cr-release assay on human tumor cells, to evaluate specificity of binding and complement-dependent lysis respectively, identified CaMBr1-CRM197, in association with the MPL-SE adjuvant, as the best vaccine formulation. This combination induced (1) production of tetrasaccharide-specific antibodies, with negligible side-effects; (2) antibodies with complement-mediated cytotoxic activity on human CaMBr1-positive cells and (3) a high titer of IgG1 detected in sera obtained 3 months after the first injection, indicating that the anti-tetrasaccharide antibody response was mediated by T cell activation. The availability of CaMBr1-glycoconjugate in the minimal and functional antigenic structure and the identification of an efficacious vaccine formulation opens the way to exploring the activity of this glycoconjugate in a clinical setting.

Immunization of metastatic breast cancer patients with a fully synthetic globo H conjugate: A phase I trial

“…Mouse studies have confirmed the generation of antibodies against globo H after vaccination with KLH or CRM197 (diptheria toxoid) conjugates, prepared by a direct conjugation protocol (18,19). There was evidence of a polyclonal antibody response, mediation of complement cytotoxicity, and binding of IgM antibodies with MCF-7 breast cancer cells (18).…”

Improvement of the Synthesis of Immunological Carbohydrate Vaccines Containing the Tumour Associate Antigen CaMBr1

“…[17] We decided to use quite a long spacer (adipic acid) between the saccharide moiety and the protein, in order to leave the sugar epitope exposed on the protein surface. The presence of the allyl group at the anomeric position permitted its manipulation in order to provide an anomeric linker for the conjugation.…”

“…The complete immunogenic evaluation is described in detail elsewhere. [17] In summary, the tetrasaccharide moiety was recognized by the Mab MBr1 in a dose-dependent manner with both conjugates. Such result indicates that the conjugation process neither compromises proper presentation of the epitope, nor modifies its conformation.…”

Chemical synthesis and immunological evaluation of entirely carbohydrate conjugate Globo H-PS A1

“…60 Globo H has therefore become one of the most attractive carbohydrate-based targets for cancer vaccine development. [61][62][63] Previous epitope mapping studies of the anti-Globo H mAb MBr1 indicated that the terminal tetrasaccharide (residue CDEF) is the minimal antigenic structure for Globo H recognition; 64,65 the structural rigidity is crucial for MBr1 binding. 66,67 The pursuit of higher Ag epitope specicity encouraged us to develop a chemically well-dened immunogenic glycoconjugate that carries Globo H hexasaccharide glycan with the specic aim of generating highly specic Abs that can bind to Globo H antigens on tumor cells.…”