Deletion of T and B Cells to Induce Tolerance

删除 T 和 B 细胞以诱导耐受

• 批准号: 7249386
• 负责人: Stanislaw M Stepkowski
• 金额: $ 18.17万
• 依托单位: UNIVERSITY OF TEXAS HLTH SCI CTR HOUSTON
• 依托单位国家: 美国
• 财政年份: 2004
• 资助国家: 美国
• 起止时间: 2004-07-01 至 2008-01-06
• 项目状态: 已结题
• 来源: https://reporter.nih.gov/project-details/7249386
• 关键词: Adoptive Cell Transfers; Adverse effects; Annamycin; Antibodies; Apoptosis; Apoptotic; B-Lymphocytes; BCL2 gene; Biological Assay; Cells; DNA; DNA Nucleotidylexotransferase; DNTT gene; Detection; Development; Dose; Doxorubicin; Dyes; Family; Fluorescence; Generations; Genes; Genetic Transcription; Goals; Graft Survival; Heart; Helix (Snails); Imagery; In Situ Nick-End Labeling; In Vitro; Interferon Type II; Interleukin-10; Interleukin-2; Interleukin-2/Interleukin-4; Interleukin-4; Interleukins; Knockout Mice; Label; Lymphocyte; Lymphoid Cell; Maintenance; Memory; Messenger RNA; Methods; Modeling; Monoclonal Antibodies; Morbidity - disease rate; Mus; NF-kappa B; New Agents; Nuclear; Numbers; Pathway interactions; Polymerase Chain Reaction; Production; Protein Tyrosine Kinase; Proteins; Protocols documentation; RNase protection assay; Rattus; Regulation; Resistance; Risk; Role; STAT protein; Sirolimus; Skin; T memory cell; T-Lymphocyte; TNFSF5 gene; Testing; Texas; Time; Transcription Coactivator; Transcriptional Activation; Transgenes; Transgenic Mice; Transgenic Organisms; Transplantation Tolerance; Tyrosine; Universities; Up-Regulation; Western Blotting; analog; annexin A5; cytokine; ear helix; heart allograft; improved; in vivo; inhibitor/antagonist; kidney allograft; mortality; research study; resistance mechanism; size; skin allograft; transcription factor; treatment duration

The ultimate goal--to radically improve graft survival without toxic side effects--must be achieved through induction of transplantation tolerance. However, tolerance induction requires initial deletion of donor-specific T and possibly B cell clones and subsequent generation of regulation to maintain tolerance. So far, no therapy induces controllable and selective deletion of donor-specific T and B cells without risking increased morbidity or mortality. Therefore, we will test two new families of apoptosis-indueing agents: 1) inducing single DNA breaks doxorubicin analogues a(nnamaycin; ANA, WP744, WP796, and WP853); and 2) selective Janus tyrosine kin ase (Jak)3 inhibitors (NC1153, WP938, WP988 and WP979). We already showed that WP744 induces apoptosis of only activated but not non-activated T cells; combination of WP744 with CD40Ligand (L) monoclonal antibody (mAb) induced tolerance to heart allografts. Similarly, NC 1153 alone induced T cell apoptosis resulting in tolerance to kidney allografts and in combination with CTL4-Ig to heart allografts. We will use unique models: T cells (but not B cells) from stimulators and activators of transcription (Stat)5a/b-deficient mice enter apoptosis after activation; whereas T cells from Stat4 and Stat6-deficient mice develop into interleukin (IL)-2-producing T helper (Th) 1 and IL-4-producing Th2, respectively; T cells from anti-apototic Bcl-2 gene over-expressing transgenic (Tg) mice display resistance to apoptosis. Using these mice we will examine the role of Stats in Bcl-2-dependent sensitivity or resistance to apoptosis in non-activated, activated and memory T cells following treatment with apoptosis-inducing agents. Apoptosis will be assessed by [a] visualization of karyolytic nuclear degeneration, [b] enzymatic detection of TdT-positive DNA degradation, [c] automated cytometric detection of annexin-V translocation, [d] expression of pro- versus antiapoptotic mRNAs in gene microarray/real-time PCR, and re] expression of Bcl-2 and Bcl-xL proteins by Western blot. In vivo, donor-specific T cell clones will be quantified by cytokine production by real-time PCR (IL-2, IL-4, IL10 and IFN-gamma) and RNase protection assay. Since T regulatory (Treg) cells are characterized as "memory" T helper 2- type CD4+/CD25 + (Th2reg) cells, we plan to explore the IL-4/Stat6-regulated apotosis-resistance mechanism. The new agents may provide potent and save method for deletion of donor-specific lymphocytes for tolerance.

最终目标——从根本上提高移植物存活率而无毒副作用——必须通过诱导移植耐受来实现。然而，耐受性诱导需要最初删除供体特异性T细胞和可能的B细胞克隆，并随后产生调节以维持耐受性。到目前为止，还没有一种治疗方法能在不增加发病率或死亡率的情况下诱导供体特异性T细胞和B细胞的可控和选择性删除。