GLYCOSPHINGOLIPIDS IN ONCOGENESIS AND DIFFERENTIATION

我们以前的工作主要集中在结构表征上 人类白细胞和白血病细胞抗原。我们已经确立了 人类白细胞表达谱系特异性的神经鞘糖脂 抗原，以及几种白细胞特异性的单抗 结合到由神经节苷脂携带的碳水化合物序列。这个 两种复杂髓系特异结构的完整结构 这些抗体识别的鞘糖脂抗原有 最近才确定的。最后，白血病的存在-- 相关的神经鞘糖脂抗原GD3也已经被 已经成立了。 在当前的提案中，我们将通过使用 包括薄层色谱在内的生化和化学技术 碳水化合物序列特异性单抗的免疫染色 抗体和快原子轰击质谱仪(FAB/MS)， 测定其他神经节苷脂抗原的结构 携带髓系特异性抗体识别的表位， Vim-8。使用这些技术和一个结合了 薄层层析法的分辨率与灵敏度 对于FAB/MS，我们建议获取高度的结构信息 复杂的髓系特异性(VIM-2)神经节苷脂。我们还提议 测定其活性和特异度的实验 合成白细胞的糖基转移酶，谱系- 特异性和白血病相关的鞘糖脂抗原 都是之前被确认的。一种新的基于抗体的酶联免疫吸附试验 程序将用于确定其表达水平 正常人白细胞和白血病中的这些糖基转移酶 细胞。这些拟议的实验将(1)提供以下信息 正常人白细胞表面标志物的结构 和白血病细胞，以及(2)为它们的 生物合成。需要这些基本信息来确定 导致特定细胞表达的机制 正常分化和白血病发生过程中的表面标志物。

Our previous work has focused on the structural characterization of human leukocyte and leukemia cell antigens. We have established that human leukocytes express lineage-specific glycosphingolipid antigens, and that several leukocyte-specific monoclonal antibodies bind to carbohydrate sequences carried by glycosphinoglipids. The complete structures of two complex myeloid-specific glycosphingolipid antigens recognized by these antibodies have recently been determined. Finally, the existence of a leukemia- associated glycosphingolipid antigen, GD3, has also been established. In the current proposal, we will extend this work by using biochemical and chemical techniques which include TLC immunostaining with carbohydrate sequence-specific monoclonal antibodies and fast atom bombardment mass spectrometry (FAB/MS), to determined the structure of other glycosphingolipid antigens that carry the epitope recognized by the myeloid-specific antibody, VIM-8. With these techniques and a procedure that combines the resolving power of thin-layer chromatography with the sensitivity of FAB/MS, we propose to obtain structural information on highly complex myeloid-specific (VIM-2) gangliosides. We also propose experiments to measure the activity and specificity of glycosyltransferases which synthesize the leukocyte, lineage- specific and leukemia-associated glycosphingolipid antigens that have previously been identified. A new antibody-based ELISA procedure will be used to determine the level of expression of these glycosyltransferases in normal human leukocytes and leukemia cells. These proposed experiments will (1) provide information on the structures of cell surface markers of normal human leukocytes and leukemia cells, and (2) establish the enzymatic basis for their biosynthesis. This fundamental information is needed to determine the mechanisms which lead to the expression of specific cell surface markers during normal differentiation and leukemogenesis.