Low Cost Production of Recombinant Proteins

描述(申请人提供)：有越来越多的蛋白质药物在流水线中，与合成的小分子药物相比，其生产成本非常高。此外，一些蛋白质药物将在未来几年内失去专利，这将为这些药物的仿制药创造一个市场，如果它们能够以低得多的成本生产并证明生物等效性的话。第一阶段的结果清楚地表明，DeltaPhone系统的一个强大和重要的商业应用是在重组蛋白质的生产中，其中一些蛋白质很难高水平表达为可溶性蛋白质。由于重组蛋白药物的重要性日益增加，以及对低成本表达和蛋白质易于纯化的明确需求，第二阶段研究的目标将是推进在未来三年内高表达和高纯度的重组蛋白的表达和纯化技术，这些重组蛋白将符合生物仿制药的条件。使阶段生物科学能够实现这一目标的技术是deltaPhaseTM系统，这是一种新的平台技术，使用一类称为弹性蛋白样多肽(ELPs)的多肽标签进行高水平表达和简单的非层析纯化。这项技术的基本基础是发现融合到ELP上的蛋白质和多肽保留了ELP在环境中触发的(温度、盐)可溶-不溶的相变行为，并且这种可逆的相变可以在细胞裂解液中进行，通过几个相变周期将ELP从其他细胞蛋白质的融合分离到均质。这项技术的影响将是为蛋白质行业提供生产蛋白质的能力，特别是生物遗传蛋白质，通过将层析从纯化过程中去掉，以显著降低成本。这将使这些药物与小分子药物更具竞争力，并降低患者的药物成本。该项目的具体目标包括：1.在大肠杆菌和CHO中表达6个与药物相关的、生物遗传的候选重组融合蛋白；2.用LC-MS法将这些蛋白纯化到至少90%的纯度；3.证明它们与商业化生产的蛋白相比具有生物活性。

DESCRIPTION (provided by applicant): There are an increasing number of protein drugs in the pipeline, whose cost of production is very high compared to synthetic, small molecule drugs. Further, a number of protein drugs are coming off patent in the next few years, which will create an opening for a market for generic version of these drugs, if they can be produced at significantly lower cost and demonstrate bioequivalence. The Phase I results clearly indicated that a powerful and commercially important application of the deltaPhase System is in the recombinant production of proteins, some of which are difficult to express at high levels as soluble proteins. Motivated by the increasing importance of recombinant protein drugs, and the clear need for low-cost expression and easy purification of proteins, the goal of the Phase II research will be to advance the technology for the expression and purification of recombinant proteins that will be eligible for biogenerics within the next three years, with high expression and at high purity levels. The enabling technology that will allow Phase Bioscience to achieve this objective is the deltaPhaseTM System, a novel platform technology for high-level expression and simple, non-chromatographic purification using a class of polypeptide tags, termed elastin-like polypeptides (ELPs). The fundamental basis of this technology is the discovery that proteins and peptides fused to an ELP retain the environmentally triggered (temperature, salt) soluble-insoluble phase transition behavior of ELPs, and that this reversible phase transition can be carried out in cell lysate to separate ELP fusions from other cellular proteins to homogeneity by a few phase transition cycles. The impact of the technology will be to provide the protein industry with the ability to produce proteins, especially biogeneric proteins, at significantly lower cost by removing chromatography from the purification process. This will make these drugs more competitive with small molecule drugs, and reduce the cost of drugs to patients. The Specific Aims of the project include: 1. Express six pharmaceutically-relevant, biogeneric candidate recombinant fusion proteins in E. coli and CHO; 2. Purify the proteins to at least 90% purity, as measured by LC-MS; and 3. Demonstrate their bioactivity compared to the commercially produced proteins.