Solid-Phase Fragment Coupling Synthesis of Granulocyte Colony-Stimulating Factor

描述（由申请人提供）：粒细胞集落刺激因子（G-CSF）是一种重要的造血生长因子，属于一类更大的生物相关糖蛋白细胞因子。两种治疗形式的G-CSF用于临床治疗化疗引起的中性粒细胞减少症。这两种形式的主要区别在于苏氨酸-133上是否有o链碳水化合物。通过化学合成方法制备两种治疗形式的既定平台可以解决有关G-CSF糖基化重要性的问题。此外，该研究策略的进一步发展将允许模块化访问更有效的G-CSF变体。由于这些原因，糖蛋白G-CSF被认为是化学合成的靶标。

DESCRIPTION (provided by applicant): Granulocyte colony-stimulating factor (G-CSF) is an important hematopoietic growth factor belonging to a larger class of biologically relevant glycoprotein cytokines. Two therapeutic forms of G-CSF are used clinically for the treatment of neutropenia induced by chemotherapy. The principal difference between these forms is the presence or absence of an O-linked carbohydrate at threonine-133. An established platform to prepare both therapeutic forms by means of chemical synthesis could address concerns with regard to the importance of G-CSF glycosylation. Additionally, further development of this research strategy would allow for modular access to more efficacious variants of G-CSF. For these reasons, glycoprotein G-CSF is considered a target for chemical synthesis. The development of efficient synthetic methods for the preparation of such therapeutic glycoproteins is an ongoing goal in our laboratory. This proposal entails the investigation of reaction scope and limitations with regard to our recently described solid-phase fragment coupling (SPFC) method. This method will be useful for the convergent preparation of large polypeptides and glycopeptides by the coupling of C-terminal polypeptide thioacids and N-terminal polypeptides on solid-phase. The application of SPFC technology would expedite the synthesis of target glycoprotein, G-CSF. To advance this method, the following synthetic challenges will be addressed: (1) the efficient preparation of C-terminal thioacids (+15-mers) and (2) the isonitrile mediated fragment coupling of C-terminal thioacids with resin-bound N-terminal polypeptides to access large polypeptides (+30-mers). Following this advance, the utility of SPFC technology will be examined in the context of target polypeptide fragments relevant to G-CSF synthesis. These targeted polypeptides will be assembled by chemical ligation techniques to prepare homogenously glycosylated and non-glycosylated forms of G-CSF.