Idiotype TCR Vβ2 DNA Plasmid Constructe, Transfer and Express in K562 Cells.

Recently, combination of different therapeutic strategies have significantly increased survival in patients with hematologic malignancies. Specific immunotherapy is an important anticancer therapy to eradicate minimal residual disease in leukemia patients after chemotherapy or stem cells transplantation. DNA vaccines have been showed to induce strong and persistent cell-mediated and humoral immune responses and used in Hodgkin lymphoma patients, using the idotype Ig antigen. In order to develop the anti-lymphoblastic leukemia idiotypic TCR DNA vaccine, which was expected to induce the specific immune response anti T-cell lymphoblastic lymphoma /leukemia in vivo. The rearranged idiotypic CDR3 fragment coding TCR Vβ2, which was identified from a TCR Vβ2 clone-Molt4 cell line, was amplified using RT-PCR, and the PCR products were then cloned into pIRES vector. The recombinant plasmids contaning target gene (405 bp, 135 peptides) were analyzed by digestion with restriction enzyme (EcoRI and XbaI), PCR and sequencing. The correct fragment was transfected into K562 cells. The condition of idiotypic protein expression was tested by indirect immunophenotyping fluorescein dyeing with anti-TCR Vβ2 monoclonal antibody, SDS-PAGE and Western-Blot. The results showed that the recombinant DNA plasmids, pIRES-Molt4 Vβ2, containing idiotypic TCR Vβ2 frgments of the Molt4 cell line were developed successfully. A 15 KD protain which can bind with TCR Vβ2 antibody specially were identified from pIRES-Molt4 Vβ2 transduced K562 cells, indicating that can express special TCR Vβ2 protain in vitro. It should be further demonstrated whether the idiotype protein can elicit both humoral and cellular immune response for anit Vβ2+ leukemic cells in vivo.