Identification of an apoptotic cleavage product of BARD1 as an autoantigen : A potential factor in the antitumoral response mediated by apoptotic bodies

• Published in: Cancer research (Chicago, Ill.) Vol. 60; no. 24; pp. 6895 - 6900
• Main Authors: GAUTIER, Fabien, IRMINGER-FINGER, Irmgard, GREGOIRE, Marc, MEFLAH, Khaled, HARB, Jean
• Format: Journal Article
• Language: English
• Published: Philadelphia, PA American Association for Cancer Research 15.12.2000
• Subjects: Amino Acid Sequence; Animals; Antibodies, Monoclonal - metabolism; Antineoplastic agents; Apoptosis; Autoantigens - chemistry; Autoantigens - metabolism; Biological and medical sciences; Blotting, Western; BRCA1 Protein - metabolism; Breast Neoplasms - metabolism; Calpain - metabolism; Carrier Proteins - chemistry; Carrier Proteins - genetics; Carrier Proteins - metabolism; Cell Cycle; Cell Fractionation; Cloning, Molecular; Colonic Neoplasms - metabolism; Cysteine Proteinase Inhibitors - pharmacology; DNA, Complementary - metabolism; Egtazic Acid - pharmacology; Enzyme Inhibitors - pharmacology; Gene Library; Humans; Immunotherapy; Leupeptins - pharmacology; Mammary Neoplasms, Animal - metabolism; Medical sciences; Mice; Molecular Sequence Data; Pharmacology. Drug treatments; Precipitin Tests; Protein Binding; Protein Structure, Tertiary; Rats; Sequence Homology, Amino Acid; Tumor Cells, Cultured; Tumor Suppressor Proteins; Ubiquitin-Protein Ligases; Antineoplastic agent; Human; Immune response; Peptides; Rat; Rodentia; Autoantibody; Molecular interaction; In vitro; Biological activity; Vertebrata; Mammalia; Autoantigen; Immunogenicity; Cell death; Proteolysis; Animal; Cell cycle; Established cell line; Mechanism of action; Tumor cell; Humoral immunity; Apoptosis; Tumor suppressor gene
• ISSN: 0008-5472; 1538-7445

We have shown previously that rats can be cured from induced peritoneal colon carcinomatosis by injections of apoptotic bodies derived from tumor cells and interleukin 2. This curative treatment generated a tumor-specific cytotoxic T-cell response associated with a humoral response. Autoantibodies from sera of cured rats strongly recognized a Mr 67,000 protein from apoptotic bodies and weakly reacted with a protein of Mr approximately 97,000 in PROb parental cells. We now show that these autoantibodies are directed against BARD1, originally identified as a protein interacting with the product of the breast cancer gene 1, BRCA1. We demonstrate that the Mr 67,000 antigen is a cleaved form of BARD1 present in apoptotic bodies derived from rat and human colon and mammary carcinoma cell lines. Moreover, we show that the cleavage site of BARD1 is located NH2 terminally but downstream of the RING domain essential for BARD1 and BRCA1 protein interaction. In vitro studies using [35S]methionine-labeled human BARD1 and apoptotic cellular extracts derived from SW48 carcinoma cells indicate that BARD1 proteolysis occurs at an early stage of apoptosis and in a cell cycle-dependent manner. This hydrolysis is inhibited by EGTA, and the calpain inhibitor I, N-acetyl-leu-leu-norleucinal, but not by several caspases inhibitors, suggesting that BARD1 is hydrolyzed by the calcium-dependent cysteine proteases, calpains. Thus, the highly immunogenic form of cleaved BARD1 could contribute to the antitumoral response mediated by apoptotic bodies.