Tailor-made expression hosts depleted in protease activity for recombinant protein production; PRODuCE (PROtease Depleted CEll line)

小分子药物（例如抗生素）传统上一直是人类治疗和疗法的支柱。然而，在过去的10-20年中，已经开发了基于蛋白质的药物（例如通常用于治疗乳腺癌的赫赛汀、胰岛素），使得这些药物现在构成了医药市场增长最快的部分。有几类基于蛋白质的药物，其中许多是在工业规模上从培养的哺乳动物或酵母细胞中生产的。由于生产这样的生物治疗剂所需的高精度，这样的“重组”基于蛋白质的药物（生物药物）通常由在限定条件下保持在培养物中的细胞生产。其中一个问题是，科学家用来制造治疗用蛋白质的细胞也会制造自己的蛋白质，包括蛋白酶，这些蛋白酶的表达可能对治疗性蛋白质的生产有害。这些类型的蛋白质实际上可以降解目标重组蛋白，使其在临床意义上无用。因此，科学家可能无法生产足够的这些药物和/或生产它们的成本可能太高，从而使医疗保健提供者无法推荐使用它们。该提案旨在解决支持从培养的哺乳动物和植物细胞合成重组蛋白的关键领域。我们的目标是开发用于生产生物制药的不同细胞系（动物和植物）中有害的蛋白水解活性的全面知识。我们将使用最先进的方法和技术来定义那些对这些表达系统的重组蛋白生产有害的蛋白酶，然后利用这些知识来减少或消除宿主细胞中这些蛋白酶的量。这将最终导致较少或没有蛋白水解切割和所产生的治疗性蛋白质的损伤。因此，该计划提出解决我们对蛋白酶在确定产品质量和产量方面的重要性的理解上的差距。总体目标是产生新的细胞和系统，其利用细胞蛋白水解机制的操作来增强工业规模的重组治疗性蛋白质的生产。该信息与工业非常相关，因为这些有害反应可能阻碍商业上有价值的治疗性蛋白质的生产。如果没有改进的表达系统，生物技术/制药工业将缺乏生产足够大量的这些有价值和有效的药物以满足需求的能力，这些药物的价格将允许将它们开给所有从中受益的患者。

Small molecule drugs (e.g. antibiotics) have traditionally been the mainstay of treatments and therapies in man. However in the last 10-20 years protein based drugs (e.g. Herceptin, often used to treat breast cancer, insulin) have been developed such that these now constitute the fastest growing section of the pharmaceutical market. There are several categories of protein-based drugs, many of which are produced from cultured mammalian or yeast cells at an industrial scale. Due to the high precision required to produce such biotherapeutics, such 'recombinant' protein-based drugs (biopharmaceuticals) are usually produced by cells kept in culture under defined conditions. One problem with this is that the cells scientists use to make proteins for therapeutic uses also make their own proteins including proteases whose expression can be detrimental to the production of therapeutic proteins. These types of proteins can actually degrade the target recombinant protein making them useless in a clinical sense. As a consequence, scientists may not be able to produce enough of these drugs and/or the cost of producing them may be too high, thus precluding health care providers from recommending their use. This proposal sets out to address a key area that underpins recombinant protein synthesis from cultured mammalian and plant cells. We aim to develop a comprehensive knowledge of the detrimental proteolytic activities in different cell lines (animals and plants) used for the production of biopharmaceuticals. We will use state-of-the-art approaches and technologies to define those proteases that are detrimental to recombinant protein production from these expression systems and then utilise this knowledge to reduce or eliminate the amount of these in the host cells. This will ultimately result in less or no proteolytic cleavage and damage of the therapeutic protein being produced. This programme therefore proposes to address the gap in our understanding as to the importance of proteases in defining product quality and yield. The overall aim is to generate new cells and systems that exploit manipulations of the cells proteolytic machinery to enhance the production of recombinant therapeutic proteins at the industrial scale. This information is of very substantial relevance to industry since the production of commercially valuable therapeutic proteins is potentially hindered by these detrimental reactions. Without improved expression systems, the biotechnology/pharmaceutical industries will lack the capability to produce large enough amounts of these valuable and effective drugs to meet the demand at a price that will allow them to be prescribed for all patients who would benefit from them.