Engineering cells for concurrent protein drug biosynthesis and polysialylation

项目概要/摘要 治疗性蛋白质或生物制剂代表了一个1000亿美元的市场，其中包括药物 如抗体、激素等。生物制剂的临床疗效至关重要， 由它们的循环半衰期决定。因此，已经开发了各种方法来 通过降低清除率来增加其循环半衰期。这通常通过以下方式实现： 在体外将生物制剂与生物相容性聚合物化学缀合。然而，化学 缀合昂贵、复杂，并且常常导致比活性的实质性损失 以及非均相产物混合物。这些严重的弊端产生了一种需求， 该技术可以将生物相容性聚合物添加到生物制剂中，而无需体外化学反应。 为了满足这一需求，GlycoBac提出了一种新的创新方法， 在生物合成过程中转化为生物制剂。聚唾液酸（PSA）是人体内天然存在的物质， 并且是完全生物相容的、可生物降解的和非免疫原性的聚合物。体外化学 聚唾液酸化的生物制剂已经显示出改善的耐受性和药代动力学， to parent父母drugs药物.此外，唾液酸生物学通过半个多世纪的研究已被充分理解。 research.因此，PSA是添加到生物制剂中的极好选择，其目的是增加其生物活性。 半衰期我们的新方法使用糖蛋白生物制剂上现有的N-聚糖作为支架， PSA添加。用于生物生产的细胞已经将N-聚糖添加到明确定义的位置。 我们建议在生物降解过程中将PSA酶促添加到这些预先存在的N-聚糖中。 生物合成（体内）。与化学缀合相反，我们的方法是位点特异性的， 需要额外的处理步骤，并且不引入额外的成本和复杂性。 该SBIR项目旨在证明体内聚唾液酸化作为下一个- 生成平台技术。我们将通过专注于生产原型的目标来实现这一目标 具有多聚唾液酸化途径的细胞系。这些细胞将用于产生两个聚唾液酸化的， 商业上相关的糖蛋白生物制剂。在第一阶段，我们将使用糖工程昆虫 细胞，因为GlycoBac对这种细胞类型有丰富的经验。我们的聚唾液酸化技术 也与哺乳动物细胞系如CHO和PerC.6相容，这些细胞系通常用于 来生产生物制品。第一阶段的成功将为更大的第二阶段项目奠定基础， 证明体内聚唾液酸化生物制剂的药代动力学和活性。III期 我们与私营部门合作伙伴的体内聚唾液酸化技术的商业化， 预计将通过生产更有效的 糖蛋白生物制剂，其需要较低频率的给药和/或减少的剂量。

Project Summary / Abstract Therapeutic proteins, or biologics, represent a $100 billion market that includes drugs such as antibodies, hormones, and many others. The clinical efficacy of biologics is critically determined by their circulating half-lives. Hence, various methods have been developed to increase their circulating half-lives by reducing clearance rates. This is commonly achieved by chemically conjugating biologics with biocompatible polymers in vitro. However, chemical conjugation is expensive, complicated, and often results in substantial losses of specific activity as well as a heterogeneous product mixture. These serious drawbacks have created a demand for a technology that can add biocompatible polymers to biologics without in vitro chemistry. To meet this demand, GlycoBac proposes a new, innovative method to add polysialic acid to biologics during their biosynthesis. Polysialic acid is (PSA) naturally found in the human body, and is a fully biocompatible, biodegradable and non-immunogenic polymer. In vitro chemically polysialylated biologics have already shown improved tolerance and pharmacokinetics compared to parent drugs. Moreover, sialic acid biology is well-understood through over half a century of research. Thus, PSA is an excellent choice to add to biologics with the goal of increasing their half-lives. Our new method uses existing N-glycans on glycoprotein biologics as a scaffold for PSA addition. Cells used for biologic production already add N-glycans to well-defined positions. We propose to enzymatically add PSA to these pre-existing N-glycans during biologic biosynthesis (in vivo). In contrast to chemical conjugation, our method is site-specific, does not require additional processing steps, and does not introduce additional cost and complexity. This SBIR project is designed to prove the feasibility of in vivo polysialylation as a next- generation platform technology. We will achieve this by Aims focused on producing a prototype cell line with a polysialylation pathway. These cells will be used to produce two polysialylated, commercially relevant glycoprotein biologics. For Phase I, we will use glycoengineered insect cells, as GlycoBac has extensive experience with this cell type. Our polysialylation technology is also compatible with mammalian cell lines such as CHO and PerC.6, which are commonly used to produce biologics. Phase I success will set the stage for a larger Phase II project focused on demonstrating the pharmacokinetics and activity of in vivo polysialylated biologics. Phase III commercialization of our in vivo polysialylation technology with private-sector partners is expected to significantly impact human health by enabling production of more efficacious glycoprotein biologics that require less-frequent dosing and/or reduced dosages.