Novel Approaches for Structural Determination of Cancer Stem Cell Glycans

描述（由申请人提供）：胰腺癌预计今年在美国造成> 32，000人死亡。这种高死亡率在很大程度上是由于缺乏可靠的早期肿瘤检测方法，以及缺乏产生治愈的治疗选择。传统癌症治疗的一个并发症是，它们很容易错过称为干细胞的小细胞亚群，这些细胞已被证明是肿瘤增殖能力的原因。异常蛋白质糖基化与癌症的发生和进展有关。特别地，具有唾液酸和硫酸酯基团的酸性聚糖通常被改变。有趣的是，最近鉴定的胰腺癌干细胞的细胞表面标志物是糖蛋白。然而，由于与不稳定、异质和分支聚糖的结构测定相关的巨大分析挑战，生成新诊断和治疗（包括癌症疫苗）所需的详细癌症相关聚糖结构是稀缺的。质谱法（MS）可以提供灵敏和准确的聚糖分析。然而，酸性分子MS的主要挑战是低电离效率。第二个挑战是糖分支和特异性连接的测定。此外，唾液酸和硫酸根基团非常不稳定，进一步损害电离并使硫酸根定位困难。本申请的重点是开发新的MS方法，用于识别和结构表征胰腺癌干细胞（潜在的生物标志物）独特表达的聚糖。具体而言，氧化锆和二氧化钛表面化学将用于从复杂混合物中富集唾液酸化和硫酸化聚糖，从而极大地提高了纳米级正相液相色谱傅立叶变换离子回旋共振（FTICR）MS的检测能力。对于这些聚糖的结构测定，我们将分别利用金属辅助电子捕获解离、电子分离解离和真空紫外光解离，以增加糖交叉环裂解，其提供连接信息并因此允许确定分支结构，以及确定硫酸根位置。 公共卫生相关性：胰腺癌是美国的主要死亡原因，五年生存率<5%。这项研究的重点是开发新的方法，用于改善胰腺癌干细胞表面存在的碳水化合物（糖分子）的检测和结构测定，即肿瘤内负责其生长和繁殖能力的细胞亚群。已知碳水化合物在癌症中发生改变，因此构成癌症疫苗开发的有希望的靶点。

DESCRIPTION (provided by applicant): Pancreatic cancer is expected to cause >32,000 deaths in he United States this year. This high mortality is largely due to lack of reliable methods for early tumor detection, and lack of treatment options that produce a cure. One complication with traditional cancer treatments is that they can easily miss the small subset of cells, termed stem cells, which have been shown to be responsible for a tumor's ability to proliferate. Aberrant protein glycosylation is linked to the onset and progression of cancer. Particularly, acidic glycans with sialic acid and sulfate groups are often altered. Interestingly, recently identified cell surface markers of pancreatic cancer stem cells are glycoproteins. However, due to tremendous analytical challenges associated with structural determination of labile, heterogeneous and branched glycans, detailed cancer-associated glycan structures, which are required for generation of novel diagnostics and therapeutics, including cancer vaccines, are scarce. Mass spectrometry (MS) can provide sensitive and accurate glycan analysis. However, a major challenge in acidic molecule MS is low ionization efficiency. A second challenge is the determination of saccharide branching and specific linkage. Also, sialic acids and sulfate groups are extremely labile, further compromising ionization and rendering sulfate localization difficult. This application focuses on developing novel MS approaches for identifying and structurally characterizing glycans uniquely expressed by pancreatic cancer stem cells (potential biomarkers). Specifically, zirconia and titania surface chemistry will be utilized to enrich sialylated and sulfated glycans from complex mixtures, thereby greatly improving their detection by nano-scale normal phase liquid chromatography Fourier transform ion cyclotron resonance (FTICR) MS. For structural determination of these glycans, we will utilize metal-assisted electron capture dissociation, electron detachment dissociation, and vacuum ultraviolet photodissociation, respectively, to increase sugar cross-ring cleavage, which provides linkage information and therefore allows determination of branched structures, and to determine sulfate location. PUBLIC HEALTH RELEVANCE: Pancreatic cancer is a major death cause in the United States with a five-year survival rate of <5%. This research focuses on developing novel approaches for improved detection and structural determination of carbohydrates (sugar molecules) present on the surface of pancreatic cancer stem cells, that is, a subset of cells within a tumor that is responsible for its ability to grow and propagate. Carbohydrates are known to be altered in cancer and therefore constitute promising targets for cancer vaccine development.