Induction of microvariant-specific CTL lines reactive to a single amino acid mismatch in bulk cultures using a transfectant expressing a single HLA class I molecule.
Alloreactivity against micromismatches in MHC class I molecules is difficult to measure. Here, we describe an in vitro model with which it is possible to examine alloreactivity against a single HLA class I allotype. The HLA class I- and class II-negative myelocytic leukemia cell line K562 was transfected with a genomic DNA clone carrying B*4403 to express a single allotype. CTL lines were generated from normal individuals carrying B*4402, B*4403, or unrelated HLA-B alleles by stimulation with B*4403- transfected K562. The bulk CTL lines generated from B*4402+ T cells against B*4403 that carry a single amino acid disparity at position 156 were specific for B*4403+ targets and did not react with targets carrying any other HLA allotype. However, the CTL lines generated from B44-negative individuals exhibited killing of the targets bearing not only B44, but also B44 CREG and a few other B alleles. Reverse transcriptase-PCR analysis of TCRs, expressed in the CTL clones uniquely specific for B*4403, showed that TCR V beta usage of alloreactive T cells directed against B*4403 was diverse but nonrandom and was affected by the HLA background of the responder. Thus, the K562-HLA transfectant system provides a useful in vitro tool to analyze alloreactivity against a single class I allele and to aid in the prediction of alloreactivity in unrelated marrow transplantation.
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Department of Pediatrics, Memorial Sloan-Kettering Cancer Center, New York 10021, USA.
Yang
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SY
D000483:Alleles; D000939:Epitopes / Q000235:genetics; D005805:Genes, MHC Class I / Q000276:immunology*; D014644:Genetic Variation / Q000276:immunology*; D015235:HLA-B Antigens / Q000032:analysis / Q000235:genetics*; D006801:Humans; D007519:Isoantigens / Q000235:genetics; D007951:Leukemia, Myeloid; D007959:Lymphocyte Culture Test, Mixed; D005810:Multigene Family / Q000276:immunology; D013601:T-Lymphocytes / Q000637:transplantation; D013602:T-Lymphocytes, Cytotoxic / Q000276:immunology*; D014162:Transfection / Q000276:immunology*; D014407:Tumor Cells, Cultured
D000939:Epitopes; D015235:HLA-B Antigens; D007519:Isoantigens