Diverse Heparan Sulfate Oligosaccharide Libraries For Understanding and Therapeutically Manipulating Glycosaminoglycans

用于理解和治疗性操纵糖胺聚糖的多样化硫酸乙酰肝素寡糖文库

• 批准号: 10414669
• 负责人: Vijayakanth Pagadala
• 金额: $ 27.28万
• 依托单位: GLYCAN THERAPEUTICS CORPORATION
• 依托单位国家: 美国
• 财政年份: 2021
• 资助国家: 美国
• 起止时间: 2021-06-15 至 2023-02-28
• 项目状态: 已结题
• 来源: https://reporter.nih.gov/project-details/10414669
• 关键词: Award; Biology; Biomedical Research; California; Cell surface; Chemicals; Chemistry; Collection; Communities; Development; Disaccharides; Enoxaparin; Event; Exhibits; Extracellular Matrix; Funding; Glycosaminoglycans; Goals; Grant; Heparin; Heparin Lyase; Heparitin Sulfate; Institutes; Label; Libraries; Methodology; Methods; Michigan; Modification; Monosaccharides; National Institute of General Medical Sciences; Oligosaccharides; Parents; Pathologic; Pattern; Pharmaceutical Preparations; Phase; Physiological; Polysaccharides; Process; Production; Research; Research Personnel; Role; S Phase; Series; Small Business Innovation Research Grant; Solid; Source; Speed; Stable Isotope Labeling; Structure; Structure-Activity Relationship; Sulfate; System; Technology; Therapeutic; Time; Universities; base; commercialization; design; glycosylation; innovation; instrument; member; phase 1 study; phase 2 study; programs; scale up

Summary The purpose of this supplementary application is to request funding to purchase a Chemspeed Technologies Flex Liquidose System. This instrument will be utilized to automate the synthesis of heparan sulfate (HS)/heparin oligosaccharides and enable the production of large comprehensive libraries of defined, structurally diverse HS oligosaccharides for biomedical research. The project for the synthesis of HS libraries is currently under the support of the NIGMS SBIR grant (Dr. Vijayakanth Pagadala, 4R44GM134738). The Chemspeed Flex Liquidose workstation will significantly increase our speed and throughput, allowing us to automate the very labor-intensive, late-stage modification and purification steps of HS synthesis. This will drastically reduce the overall time required to synthesize HS oligosaccharides and enable the production of large collections of compounds. Notably, once developed, this system will be the first general, automated platform for glycosaminoglycan (GAG) synthesis. Developing this new capability is essential for completing the proposed oligosaccharide library synthesis and for sustaining the proposed commercialization plan and long-term goals of Glycan Therapeutics. Research Abstract/Summary of Funded Parent Award (Dr. Vijayakanth Pagadala, 4R44GM134738, funding period for phase I: 8/1/2019 to 1/31/2020; funding period for phase II 3/1/2020 to 2/28/2022) The overall goal of this fast-track SBIR program is to synthesize and commercialize the first large, comprehensive libraries of structurally-defined heparan sulfate (HS) oligosaccharides. Glycan Therapeutics is a company that specializes in HS-related research products. In this project, Glycan Therapeutics (Dr. Vijay Pagadala) has teamed up with leading experts in HS synthesis, including Prof. Linda Hsieh-Wilson (California Institute of Technology) and Prof. Xuefei Huang (Michigan State University), to produce large collections of structurally- diverse HS oligosaccharides for biomedical research. HS/heparin glycosaminoglycans (GAGs) are highly sulfated polysaccharides with important roles in many physiological and pathological events. The structural diversity of naturally-occurring HS, which exhibits diverse sulfation patterns, has hindered an understanding of its structure-activity relationships. To date, synthetic methodologies toward HS are mostly target oriented, leading to a small set of oligosaccharides. To enable a deeper understanding of HS biology and investigate its broader therapeutic potential (beyond drugs such as heparin, Lovenox, and Arixtra), the availability of large collections of HS oligosaccharides is critical. In Aim 1, we proposed to expedite the synthesis of key disaccharide building blocks. Rather than starting from monosaccharides, new methods were developed to hydrolyze natural heparin and heparosan polysaccharides to obtain disaccharides.1 These disaccharides were then transformed into suitably-protected building blocks ready for glycosylation and glycan chain extension. Notably, this process reduced the total number of synthetic steps needed to produce the disaccharide building blocks by ~50%. In Aim 2, we proposed to synthesize a comprehensive library of HS tetrasaccharides representing all possible 2-O, 6-O and N-sulfation sequences. A new protecting group strategy was designed to enable access to 256 different HS tetrasaccharides from only four universal building blocks by automated solid-phase synthesis. All tetrasaccharides were designed to be functionalized for bioconjugation and microarray studies. In Aim 3, we proposed to synthesize a library of 13C-labeled oligosaccharide standards carrying a DUA4,5- unsaturated residue at the nonreducing end. A common method for researchers to determine HS composition is to use heparin lyases to degrade HS into disaccharides, followed by disaccharide composition analysis using LC-MS. Currently, there are no commercial sources for stable isotopically-labeled 3-O-sulfated standards. Glycan Therapeutics is synthesizing a series of these standards to enable quantitative analysis of HS composition. In Phase I studies, four key disaccharides (200 mg each), four strategically-protected tetrasaccharide precursors (200 mg each), ten differentially-sulfated HS tetrasaccharides (2 mg each), as well as two 13C-labeled and 3-O- sulfated HS oligosaccharides (1 mg each), were prepared. In the current Phase II studies, the synthesis is being scaled up to produce the universal tetrasaccharide building blocks (10 g each), the 256-member library of HS tetrasaccharides representing all possible 2-O, 6-O and N-sulfation sequences (2 mg each), and a 13C-labeled and 3-O HS oligosaccharide library (28 compounds, 3 mg each). The new HS structures that will become available from this project will cover the entire chemical space of HS tetrasaccharides and represent the largest, most comprehensive HS libraries to date. Successful commercialization of these products will greatly accelerate research on the chemistry and biology of HS and facilitate the development of HS-based therapeutics.

摘要 此补充申请的目的是申请资金以购买一辆ChemSpeed. 科技公司Flex Liquidose系统。这个仪器将被用来自动合成 硫酸乙酰肝素(HS)/肝素低聚糖的合成和生产 已定义的、结构多样的HS寡糖文库，用于生物医学研究。该项目 目前正在NIGMS SBIR赠款(Dr。 Vijayakanth Pagadala，4R44GM134738)。ChemSpeedFlex Liquidose工作站将 显著提高我们的速度和吞吐量，使我们能够自动化非常劳动密集型、 HS合成的后期修饰和提纯步骤。这将大大减少 合成HS寡糖和能够生产大的 化合物的集合。值得注意的是，一旦开发出来，这个系统将是第一个通用的， 糖胺多聚糖(GAG)合成自动化平台。开发这一新功能是 对于完成拟议的寡糖库合成和维持 提出了葡聚糖治疗的商业化计划和长期目标。 研究摘要/资助父母奖摘要(Vijayakanth Pagadala博士， 4R44GM134738，第一阶段资金期限：2019年8月1日至2020年1月31日；阶段资金期限 II 2020年3月1日至2022年2月28日) 这个快速通道SBIR计划的总体目标是合成第一个大型的， 结构定义的硫酸乙酰肝素(HS)寡糖的全面文库。葡聚糖 Treateutics是一家专门从事HS相关研究产品的公司。在这个项目中， 葡聚糖治疗公司(Vijay Pagadala博士)与HS合成领域的领先专家合作， 包括Linda Hsieh-Wilson教授(加州理工学院)和黄学飞教授 (密歇根州立大学)，生产结构多样化的HS的大型集合 用于生物医学研究的低聚糖。 HS/肝素糖胺聚糖(GAG)是一种高度硫酸盐化的多糖，具有重要的作用 在许多生理和病理事件中。自然产生的HS的结构多样性， 表现出不同的硫酸盐化模式，阻碍了对其结构活性的理解 两性关系。到目前为止，HS的合成方法大多是面向目标的，导致 一小部分低聚糖。以加深对HS生物学的了解并调查 其更广泛的治疗潜力(超越肝素、Lovenox和Arixtra等药物)， 大量HS低聚糖的可获得性至关重要。 在目标1中，我们建议加快关键双糖构建块的合成。而不是 从单糖出发，开发了新的方法来水解天然肝素和 得到双糖，然后转化这些双糖。 为糖基化和糖链延伸做好准备的适当保护的构建块。值得注意的是， 这个过程减少了生产二糖所需的合成步骤的总数 构建块减少约50%。 在目标2中，我们建议合成一个全面的HS四糖文库，代表 所有可能的2-O、6-O和N-硫化序列。设计了一种新的保护群策略 通过以下方式仅从四个通用构建块访问256种不同的HS四糖 自动固相合成。所有四糖都被设计成官能化的 生物偶联和微阵列研究。 在目标3中，我们建议合成一个13C标记的寡糖标准品文库，携带 非还原末端的DUA4，5-不饱和残基。研究人员的一种常用方法 确定HS的组成是使用肝素裂解酶将HS降解为二糖，然后 用LC-MS分析双糖组成。目前，没有商业来源 用于稳定的同位素标记的3-O-硫酸盐标准。葡聚糖治疗公司正在合成一种 这些标准的一系列，以实现HS成分的定量分析。 在第一阶段的研究中，四种关键的双糖(每种200毫克)，四种战略保护 四糖前体(每个200毫克)，十个不同硫化的HS四糖(2毫克 每个)，以及两个13C标记的和3-O-硫化的HS寡糖(各1 mg)， 准备好了。在目前的第二阶段研究中，合成正在扩大规模，以生产通用的 四糖构建块(每个10克)，HS的256个成员文库 代表所有可能的2-O、6-O和N-硫化序列的四糖(各2毫克)，以及 13C标记的3-O HS寡糖库(28个化合物，每个3 mg)。新房协 该项目将提供的结构将覆盖HS的整个化学空间 四糖，是迄今为止最大、最全面的HS文库。成功 这些产品的商业化将极大地促进化学和生物学的研究 促进以HS为基础的治疗学的发展。