CHARACTERIZATION OF THE ANTI-RH ALLOIMMUNE RESPONSE

抗 RH 同种免疫反应的特征

• 批准号: 6302367
• 负责人: DONALD Lawrence SIEGEL
• 金额: $ 40.59万
• 依托单位: UNIVERSITY OF PENNSYLVANIA
• 依托单位国家: 美国
• 财政年份: 2000
• 资助国家: 美国
• 起止时间: 2000-02-15 至 2001-12-31
• 项目状态: 已结题
• 来源: https://reporter.nih.gov/project-details/6302367
• 关键词: CD40 molecule; antibody specificity; antigen antibody reaction; antiidiotype antibody; blood group incompatibility; blood transfusion; cell transformation; clinical research; epitope mapping; erythroblastosis fetalis; histocompatibility; homologous transplantation; human subject; human tissue; humoral immunity; immune response genes; immunogenetics; immunoglobulin idiotypes; immunoglobulin structure; isoantibody; laboratory mouse; monoclonal antibody; protein sequence; tissue /cell culture; transplantation immunology

Alloimmunization to the Rhesus blood group system through transfusion or pregnancy can limit the effectiveness of transfusions, can lead to clinically-significant transfusion reactions, and can cause hemolytic disease of the newborn. In order to design specific and effective methods for the prevention and management of the problems associated with Rh alloimmunization, it is first necessary to characterize the anti-Rh immune response at the molecular level. To this end, the proposed research will utilize a number of novel cellular and recombinant approaches to examine the repertoire of variable regions genes used by anti-Rh alloantibodies as well as the structural heterogeneity of their antigenic binding sites. Specifically, we will express human anti-Rh alloantibodies in vitro from the B-cells of sensitized patients by first expanding the B-cells with crosslinked anti- CD40 and IL-4, and then screening for anti-Rh producing clones prior to cell transformation with EBV or somatic cell hybridization. In parallel, filamentous phage immunoglobulin display libraries will be created from the initial populations of B-cells, as well as from expanded and transformed clones, to rescue cells that are unstable or refractory to immortilization. This combined approach will provide both the expressed immunoglobulins and the cDNAs that encode them. This will enable the identification of immunodominant epitopes on Rh antigens and conserved DNA sequences on anti- Rh antibodies. Finally, we will isolate peptides that inhibit anti-Rh binding to RBCs in vitro through the analysis of hypervariable loops of blocking (anti-idiotypic) antibodies and the screening of random peptide libraries with anti-Rh antibodies. Collectively, these studies will provide an understanding of the nature of the human immune response to Rh antigens on a molecular genetic level. This will facilitate t he design of therapeutically useful inhibitors that modulate anti-Rh/Rh antigen interactions in vivo. It will also lead to the development of genetic vaccines that modulate anti-Rh/Rh antigen interactions in vivo. It will also lead to the development of genetic vaccines that induce T-cell responses that down-regulate athe production of anti-Rh alloantibodies. In addition, the in vitro production of monoclonal anti-Rh antibodies/recombinant Fab fragments and the structural analogs of their antigenic epitopes will provide useful biochemical and serological reagents to increase our understanding of the biochemistry and immunology of this important blood group system.

通过输血对恒河猴血型系统进行同种免疫，或 怀孕会限制输血的有效性， 具有临床意义的输血反应，并可导致溶血性 新生儿的疾病。 为了设计具体有效的方法 用于预防和管理与Rh相关的问题 同种异体免疫，首先需要表征抗Rh免疫 分子水平的反应。 为此，拟议的研究将利用一些新的细胞 和重组方法来检查可变区的库 抗Rh同种异体抗体所用的基因以及 其抗原结合位点的异质性。 具体来说，我们将 在体外从以下B细胞表达人抗Rh同种异体抗体： 致敏患者首先扩大B细胞与交联抗- CD 40和IL-4，然后筛选产生抗Rh的克隆， 用EBV或体细胞杂交进行细胞转化。 同时， 丝状噬菌体免疫球蛋白展示文库将从 B细胞的初始群体，以及来自扩增和转化的 克隆，以拯救不稳定或难以永生化的细胞。 这种组合的方法将提供表达的免疫球蛋白和 编码它们的cDNA 这将有助于识别 Rh抗原上的免疫显性表位和抗- Rh抗体。 最后，我们将分离抑制抗Rh的肽 通过分析红细胞高变环， 封闭（抗独特型）抗体和随机肽的筛选 抗Rh抗体的文库。 总的来说，这些研究将提供一个了解的性质， 在分子遗传水平上对Rh抗原的人类免疫应答。 这将有助于设计治疗上有用的抑制剂， 调节体内抗Rh/Rh抗原相互作用。 这也将导致 调节抗Rh/Rh抗原的基因疫苗的开发 体内的相互作用。 这也将导致遗传学的发展。 诱导T细胞应答的疫苗， 抗Rh同种抗体。 此外，在体外生产的单克隆 抗Rh抗体/重组Fab片段及其结构类似物 它们的抗原表位将提供有用的生物化学和血清学 增加我们对生物化学和免疫学的理解 这个重要的血型系统。