ABSTRACT
High avidity for antigen and diversity of T cell receptor (TCR) repertoire are essential for effective immunity against cancer. We have previously created a transgenic mouse strain with increased TCR avidity in a diverse T cell population. In this report, we show that strong alloreactive responses of transgenic T cells against targets with low MHC class I expression can be used for effective adoptive transfer of tumor immunity in vivo. Alloreactive transgenic T cells could be an effective therapeutic approach counteracting tumor evasion of the immune system.
Subject(s)
ATP-Binding Cassette Transporters/physiology , CD8-Positive T-Lymphocytes/immunology , Lymphoma/immunology , Lymphoma/therapy , T-Lymphocytes, Cytotoxic/immunology , ATP Binding Cassette Transporter, Subfamily B, Member 2 , Animals , CD8-Positive T-Lymphocytes/metabolism , H-2 Antigens/immunology , H-2 Antigens/metabolism , Histocompatibility Antigen H-2D , Lymphoma/pathology , Major Histocompatibility Complex/immunology , Mice , Mice, Transgenic , Receptors, Antigen, T-Cell/physiology , Survival Rate , T-Lymphocytes, Cytotoxic/metabolism , beta 2-Microglobulin/physiologyABSTRACT
We have analyzed peptides associated with six human major histocompatibility complex (MHC) class I allomorphs expressed by the U937 cell line. Peptides were isolated by mild acid elution or by MHC class I immunoprecipitation by using W6/32 monoclonal antibody. Eighty-five peptides were sequenced by mass spectrometry, and their putative binding alleles were assigned using bioinformatic tools. Only three peptides isolated by the two approaches were identical, suggesting that the approaches may yield distinct partially overlapping peptide populations. Mild acid treatment-derived peptides manifested overall characteristics suggestive of relatively lower affinity of binding for MHC class I. Interestingly, a large proportion of putative HLA-B*5101-binding peptides was evident among the mild acid treatment-eluted peptides, and to a lesser degree in the affinity-purified peptide pool. These results suggest that HLA-B*5101 may bind a potentially large pool of peptides with relatively lower affinity. We suggest that lower affinity of peptide binding may be the basis for inefficient tolerance to HLA-B*5101-binding self-peptides, a predisposing factor for the development of Behçet disease.
Subject(s)
HLA-B Antigens/metabolism , Histocompatibility Antigens Class I/metabolism , Peptides/metabolism , Amino Acid Sequence , Behcet Syndrome/immunology , Binding Sites/genetics , Humans , Intracellular Space/metabolism , Mass Spectrometry , Molecular Sequence Data , Molecular Weight , Peptides/chemistry , Peptides/genetics , Peptides/isolation & purification , Protein Binding/drug effects , U937 CellsABSTRACT
We have previously shown that sigma-2 receptors are relatively difficult to solubilize (Eur. J. Pharmacol. 304 (1996) 201), suggesting possible localization in detergent-resistant lipid raft domains. Rat liver membranes were treated on ice with 1% Triton X-100 or 20 mM 3-[(3-cholamidopropyl)dimethylammonio]-1-propanesulfonate (CHAPS), and the extract subjected to centrifugation on a discontinuous gradient of 5%, 38%, and 40% sucrose. Gradient fractions were analyzed for sigma-1 receptors using [3H]+-pentazocine and for sigma-2 receptors using [3H]1,3-di-o-tolylguanidine ([3H]DTG), in the presence of dextrallorphan. Flotillin-2 was assessed by immunoblotting as a marker for lipid rafts. Sigma-2 receptors were found to discretely co-localize with flotillin-2 in lipid raft fractions. However, sigma-1 receptors were found throughout the gradient. Rafts prepared in CHAPS had sigma-2 receptors with normal pharmacological characteristics, whereas those in Triton X-100-prepared rafts had about seven-fold lower affinity for [3H]DTG and other ligands. Thus, sigma-2 receptors are resident in membrane lipid rafts, whereas sigma-1 receptors appear in both raft and non-raft membrane domains. Lipid rafts may play an important role in the mechanism of sigma-2 receptor-induced apoptosis.
