Aberrant N-Glycan Standards for Mass Spectrometry and Microarray Analysis

描述（由申请人提供）：大多数细胞表面和分泌蛋白质被共价连接的聚糖修饰，这些聚糖是生物过程（如蛋白质折叠、细胞信号传导、受精、胚胎发生和细胞增殖）的基本介质。此外，细胞表面聚糖的异常糖基化模式已显示在癌症的发展和进展中起作用。有越来越多的文献表明，蛋白质的异常糖基化提供了开发更可靠的癌症生物标志物用于早期诊断的可能性。细胞产生信息丰富的聚糖的能力创造了一个新的研究领域，称为“糖组学”，旨在识别和理解细胞类型和发育阶段特异性寡糖模式形成过程。关键的糖组学技术包括聚糖结构的质谱或HPLC分析、用于测量糖酶和聚糖结合蛋白的表达水平的糖基因微阵列以及用于筛选聚糖-蛋白质相互作用的凝集素阵列。虽然这些技术的使用开始显示出细胞类型特异性糖基化的功能意义，但由于缺乏明确定义的复杂寡糖标准品，进展受到阻碍。在许多情况下，定义明确的寡糖只能通过化学或酶促方法获得。虽然已经取得了巨大的进展，但复杂的寡糖合成仍然非常耗时，并且制备单个明确定义的衍生物可能需要长达一年的时间并不罕见。对更有效的低聚糖合成方法的需求刺激了化学-酶法的发展，其中合成低聚糖前体被一系列糖基转移酶修饰以得到更复杂的化合物。然而，这种方法的一个严重限制是，它只提供了对称的支链寡糖。化学酶促合成的范围进一步受到缺乏容易获得的糖基转移酶的限制。该项目的目的是开发一种新的化学酶促方法，该方法可以轻松访问在肿瘤细胞上发现的定义明确的对称和不对称分支寡糖库。在与“癌症检测糖生物学家联盟”联盟的合作中，将选择将通过这种新方法制备的目标。新的复合寡糖将被用作质谱分析的标准品，以确定肿瘤相关聚糖的确切结构。此外，聚糖将用于聚糖微阵列开发，以探测碳水化合物蛋白质相互作用，并分析癌症患者血清样品中具有诊断价值的肿瘤相关聚糖抗体。

DESCRIPTION (provided by applicant): Most cell surface and secreted proteins are modified by covalently-linked glycans which are essential mediators of biological processes such as protein folding, cell signaling, fertilization, embryogenesis, and cellular proliferation. Additionlly, abnormal glycosylation patterns of cell surface glycans have been shown to play a role in the development and progression of cancer. There is an expanding amount of literature that demonstrates that aberrant glycosylation of proteins offers the possibility to develop more reliable cancer biomarkers for early diagnosis. The ability of cells to generate information rich glycans has created a new field of research termed "glycomics," which seeks to identify and understand the processes involved in the formation of cell type and developmental stage specific oligosaccharide patterns. Key glycomic technologies include mass spectrometric or HPLC profiling of glycan structures, glyco-gene microarrays for measuring expression levels of glycoenzymes and glycan binding proteins, and lectin arrays for screening glycan-protein interactions. Although the use of these technologies is beginning to show the functional significance of cell type specific glycosylation, progress is hampered by the lack of well-defined complex oligsaccharide standards. In many cases, well-defined oligosaccharides can only be obtained by chemical- or enzymatic approaches. Although tremendous progress has been made, complex oligosaccharide synthesis remains very time consuming and it is not uncommon that the preparation of a single well-defined derivative can take as much as a year. The need for more efficient approaches for oligosaccharide synthesis has stimulated the development of chemo-enzymatic methods in which a synthetic oligosaccharide precursor is modified by a range of glycosyl transferases to give a more complex compound. However, a serious limitation of this approach is that it provides only access to symmetrical branched oligosaccharides. The scope of chemoenzymatic synthesis is further limited by a lack of readily available glycosyltransferases. The aim of this project is to develop a novel chemoenzymatic methodology that will give easy access to libraries of well-defined symmetrical and asymmetrical branched oligosaccharides found on tumor cells. In collaboration with the alliance of "Alliance of Glycobiologists for Detection of Cancer" targets will be selected that will be prepared by this new methodology. The new complex oligosaccharides will be employed as standards for mass spectrometry to identify exact structures of tumor associated glycans. Furthermore, the glycans will be employed for glycan microarray development to probe carbohydrate protein interactions and profile serum samples of cancer patients for antibodies against tumor associated glycans that can have diagnostic value.