MECHANISMS OF ABERRANT IG EXPRESSION IN CHRONIC LEUKEMIA

慢性白血病中 IG 表达异常的机制

• 批准号: 2653227
• 负责人: ALEXIS A THOMPSON
• 金额: $ 10万
• 依托单位: UNIVERSITY OF CALIFORNIA LOS ANGELES
• 依托单位国家: 美国
• 财政年份: 1997
• 资助国家: 美国
• 起止时间: 1997-09-30 至 1999-09-29
• 项目状态: 已结题
• 来源: https://reporter.nih.gov/project-details/2653227
• 关键词: cellular oncology; chronic lymphocytic leukemia; clinical research; gene expression; gene mutation; human subject; immunoglobulin genes; neoplasm /cancer genetics; neoplasm /cancer immunology

Chronic Lymphocytic leukemia (CLL) is the most prevalent form of leukemia in Western countries, and is characterized by a monoclonal proliferation of primarily mature CD5+B lymphocytes. Surface immuno-globulin (Ig) expression, which is often decreased in CLL, normally requires the protein product of the B29 gene for translocation of the B cell antigen receptor complex (BCR) to the cell surface and for signal transduction. Because B29 is essential for intracellular assembly and transport of the B cell antigen receptor complex to the cell surface, we postulate that a perturbationin the B29 gene could result in diminished expression and function of surface ig in leukemic Cll cells. Our studies have now revealed aberrations in the expression of the B29 gene in CLL. Analysis of 18 unselected cases of CLL demonstrate that over 80% had low to absent B29 expression which correlated directly to their level of surface ig expression. Half of these surface B29low/-cases had either no or barely detectable levels of B29 mRNA by RNAse protection assay. To date, all of the CLL samples with normal B29 mRNA levels have had point mutations or truncations identified by RT-PCR and sequencing which would significantly effect the structure and/or function of B29 protein. The primary objectives of this research proposal ae to define the spectrum of specific aberrations of the B29 gene in CLL, to determine their consequences and then finally to relate these abnormalities to disease pathogenesis. PCR/SSCP analysis of genomic DNA from a larger panel of CLL cases is expected to identify B29 mutations which will be confirmed by direct sequencing. RNAse protection assays and RT-PCR will be used to identify alternative spliced and other B29 mRNA variants. Cotransfection of mutant B29 constructs with othe rBCR component expression vectors should reproduce the defects seen in CLL. Primary CLL cells maintained on a CD40L expressing geeder layer will be infected with a normal B29 vaccinia viral expression vector and assayed for BCR surface expression. BCR signal transduction, cell cycle progression and sensitivity to apoptosis of "corrected" CLL cells will also be determined. From the studies proposed, it will be clearly demonstrated that mutations of directed at correctign these B29 mutations are expected to induce increased ig surface expression in CLL and may improve the sensitivity of CLL to cytotoxic chemotherapy.

慢性淋巴细胞白血病(CLL)是最常见的形式 白血病在西方国家的发病率，其特点是 CD5+B淋巴细胞的单克隆性增殖。 表面免疫球蛋白(Ig)的表达，通常是降低的 在慢性淋巴细胞性白血病中，通常需要B29基因的蛋白质产物 B细胞抗原受体复合体(BCR)转位到 细胞表面和信号转导。因为B29是必不可少的 用于B细胞抗原的细胞内组装和运输 受体复合体结合到细胞表面，我们假设一个 B29基因的突变可能导致表达减弱 白血病CLL细胞表面免疫球蛋白的功能。我们的研究已经 现在发现在B29基因的表达中出现了异常 CLL.对18例未选定的慢性淋巴细胞性白血病病例的分析表明 超过80%的人低表达或不表达B29，这与 直接表达到其表面的ig水平。其中的一半 表面B29low/-病例没有或几乎检测不到 核糖核酸酶保护实验检测B29基因的表达。到目前为止，所有的CLL B29mRNA水平正常的样本发生了点突变 或通过RT-PCR和测序确定的截断 显著影响B29蛋白的结构和/或功能。 这项研究提案的主要目标是定义 慢性淋巴细胞性白血病B29基因特异性畸变谱的研究 确定它们的后果，然后最后将它们联系起来 疾病发病机制的异常。应用聚合酶链式反应/单链构象多态性分析 来自更多CLL病例的基因组DNA预计将 确定B29突变，将通过DIRECT确认 测序。核糖核酸酶保护试验和RT-PCR将用于 确定可供选择的剪接和其他B29mRNA变种。 突变型B29构建体与其他rBCR共转染的研究 组件表达载体应该复制在 CLL.维持CD40L表达的原代CLL细胞 蛋鸡将感染一种正常的B29痘苗病毒 表达载体，检测bcr的表面表达。BCR 信号转导、细胞周期进程和对 “矫正”的CLL细胞的凋亡率也将被测定。从… 这些研究建议，将清楚地证明突变 这些B29突变的目的是纠正 诱导CLL免疫球蛋白表面表达增加，并可能改善 慢性淋巴细胞性白血病对细胞毒化疗的敏感性