DNA 3.0: Development of a novel, efficient and cost-effective enzymatic process for synthesis of DNA oligonucleotides

项目概要/摘要 自20世纪80年代生物技术诞生以来，有效的核酸合成一直是关键 生物发现和生物产品开发的驱动力，在最近出现的 “合成生物学”领域和核酸治疗学的快速发展。序列特异 RNA和DNA制造在医疗保健领域的影响尤其强烈，FDA 2018年批准的第一批基于阿替西汀的治疗药物很快导致了数百种 类似药物进入临床试验。尽管有这些积极的发展，新的DNA和RNA 迫切需要合成技术来满足对临床级药物的快速增长的需求。 寡核苷酸，因为目前的化学合成策略是昂贵的，难以规模化， 效率低且限于长度为200个核苷酸或更少的分子。在过去的十年里，酶 为此目的，已经探索和开发了寡核苷酸合成（EOS）策略。 在第一阶段的可行性项目中，原始遗传学公司。展示了新模板的使用- 独立的DNA聚合酶（TIDP），用于控制天然的、未修饰的核苷酸的添加， 在一个简单的过程中生长的DNA链，不需要化学解封闭步骤。这 创新有可能使一个强大的，廉价的，灵活的，环境友好的， 容易扩展的酶促途径来制造用于治疗剂，疫苗， 诊断和研发产品。 在第二阶段的小企业创新研究（SBIR）项目中，原始遗传学 建议继续第一阶段的工作，以优化已经发现的TIDP， 工业EOS所需的每个合成循环中的单核苷酸添加率（99%） 过程该公司的基因改良和筛选平台，特别是功能 Generator™技术将与人工智能和机器学习一起应用 我们的计算生物学合作者Koliber Biosciences的能力，以改善这些酶 达到所需的效率水平。一个原型EOS过程将开发使用优化的 作用于固定在固体支持物上的寡核苷酸的酶。 原型EOS过程有可能实现更有效的DNA和RNA合成 并消除目前阻碍核技术发展的制造瓶颈， 酸性药物随着时间的推移，我们将调整TIDP，以允许合成所有修饰的核苷酸 目前正被整合到基于DNA和RNA的治疗剂、疫苗以及试剂中 用于诊断、研发和DNA信息存储。

Project Summary/Abstract Since the birth of biotechnology in the 1980s, efficient nucleic acid synthesis has been a key driver of biological discovery and bio-product development, culminating in the recent emergence of the ‘synthetic biology’ field and rapid development of nucleic acid therapeutics. Sequence-specific RNA and DNA manufacturing is having an especially strong impact in healthcare, where FDA approval of the first oligonucleotide-based therapeutics in 2018 quickly led to many hundreds of similar drugs entering clinical trials. Despite these positive developments, new DNA and RNA synthesis technologies are urgently needed to meet the rapidly rising demand for clinical-grade oligonucleotides, because the current chemical synthesis strategies are costly, difficult to scale, inefficient and limited to molecules of 200 nucleotides or less in length. In the past decade, enzymatic oligonucleotide synthesis (EOS) strategies have been explored and developed for this purpose. In a Phase I feasibility project, Primordial Genetics Inc. demonstrated the use of novel template- independent DNA polymerases (TIDPs) for controlled addition of natural, unmodified nucleotides to a growing DNA strand in a simple process that does not require a chemical deblocking step. This innovation has the potential to enable a robust, inexpensive, flexible, environmentally friendly and easily scalable enzymatic route to manufacturing DNA and RNA used in therapeutics, vaccines, diagnostics and R&D products. In this Phase II Small Business Innovation Research (SBIR) project, Primordial Genetics proposes to continue the Phase I work to optimize the already discovered TIDPs to enable a high rate (99%) of single nucleotide addition in each synthesis cycle that is needed for an industrial EOS process. The company’s genetic improvement and screening platform, specifically the Function Generator™ technology, will be applied together with the artificial intelligence and machine learning capabilities of our computational biology collaborator Koliber Biosciences to improve these enzymes to the desired level of efficiency. A prototype EOS process will be developed using the optimized enzymes acting on oligonucleotides immobilized on a solid support. The prototype EOS process has the potential to enable more efficient DNA and RNA synthesis and remove the manufacturing bottlenecks that are currently holding back the development of nucleic acid medicines. Over time, we will adapt the TIDPs to allow synthesis of all modified nucleotides currently being incorporated into DNA- and RNA-based therapeutics, vaccines, as well as reagents for diagnostics, R&D and DNA-based information storage.