Microfabricated cantilever methods as nanoscale screens for early indicators of protein aggregation; a feasibility study

目前，一种新的医药产品需要超过8亿美元和大约10-15年的时间才能进入市场。目前正在开发的产品中有三分之一以上是生物制药，即活性成分是大生物分子的药物，如蛋白质或核酸。含有单抗的制剂(例如，用于治疗免疫疾病、癌症和感染)是目前最大也是最重要的一类生物制药。因此，生物制药的开发既昂贵又耗时。用于设计/发现新的生物药剂的方法与传统的合成药物-分子药物非常不同，大生物分子在药物设计(配方)、制造和储存的开发过程中带来了新的挑战。目前，这往往反映在生物制药疗法的高市场价格上，而更快提供更便宜产品的压力导致了对新的分析方法/途径的需求，以解决传统药物方法难以满足的问题。其中一个直接影响以蛋白质为基础的药物，并可能对开发过程的任何阶段产生不利影响的问题是蛋白质分子粘连在一起形成称为聚集体的分子集合的不可预测的趋势。在药品生产过程中，凝聚会产生高度粘性的溶液，导致加工和产品包装过程中出现问题。它还可能导致产品稳定性降低，从而难以估计产品的保质期。在最终药物中，关键治疗活性成分的聚集最终会降低药物的疗效，在极端情况下可能会对患者产生严重的不良副作用。因此，通过在药物开发过程中尽早确定哪些生物分子有聚集的趋势(以及鼓励/阻止这种聚集的条件)，那么与治疗方法的开发相关的努力、成本和患者的风险可以显著减少。在这个项目中，我们的目标是探索使用微悬臂检测方法(使用超灵敏的弹簧来测量生物分子之间的相互作用)来解决这一困难挑战的可行性。最终，我们的目标是开发一种新的方法，通过使用能够检测聚集的非常早期阶段或与聚集行为一致的分子性质变化的设备，来筛选潜在的生物活性分子的聚集倾向。在这个为期一年的项目中，我们的目标是测试和探索实现这一目标的一系列潜在方法的可行性。重要的是，我们的实验计划将使用模型和治疗相关的蛋白质(包括单抗)。在与制药业的密切合作下，还将评估这些初步研究中确定的设备的可行性，并扩大其在工业背景下的使用。越来越多的学术界和工业界已经认识到蛋白质聚集的发生是许多领域的一个关键问题，并寻求新的方法来研究这一现象。这个项目有可能提供一种全新的方法，在最早的阶段检测和调查聚集的起源。从长远来看，这项工作如果建立在基础上，将对生物技术和医疗保健的重要领域产生影响。事实上，所开发的方法可以在以具有成本效益和及时的方式将新一代药物推向市场方面发挥重要作用，因此将对公众健康和生活质量产生非常重大的影响。

It currently takes over $800 million and around 10-15 years for a new medicinal product to reach the market. Over one third of the products currently under development are biopharmaceuticals; medicines in which the active ingredients are large biological molecules, such as proteins or nucleic acids. Preparations containing monoclonal antibodies (e.g. for the treatment of immune disorders, cancer and infection) are currently the largest, and most important class of biopharmaceutical. The development of biopharmaceuticals is therefore both costly and time intensive. The approaches employed to design/discover new biopharmaceutical agents are very different to traditional synthetic drug-molecule based medicines, with large biological molecules bringing new challenges in terms of development processes during medicine design (formulation), manufacture and storage. At present this is often reflected in the high market price of biopharmaceutical therapeutics, and the pressure to provide cheaper products more quickly has resulted in a demand for new analytical methods/approaches to address issues which conventional drug approaches are struggling to meet. One such issue which directly impacts on protein-based medicines, and which can adversely affect any stage of the development process, is the unpredictable tendency of protein molecules to stick together, to form assemblies of molecules termed aggregates. During medicine manufacture, aggregation can result in highly viscous solutions, causing problems during processing and product packaging. It can also lead to decreased product stability, and hence difficulties in estimating product shelf-life. Within the final medicine, aggregation of the key therapeutic active ingredient can ultimately reduce the efficacy of the medicine and can in extreme cases produce severe unwanted side effects in the patient. By identifying as early as possible within the drug development process, which biomolecules have a tendency to aggregate (and also the conditions which encourage/discourage this aggregation) then the effort, costs and risks to patients associated with development of the therapeutic could therefore be significantly reduced. In this project we aim to explore the feasibility of addressing this difficult challenge, using microcantilever detection approaches (which employ ultrasensitive springs to measure interactions between biological molecules). Ultimately we aim to develop a novel approach to screen potential biological 'active' molecules for their tendency to aggregate through the use of devices capable of detecting the very early stages of aggregation or changes in molecular properties consistent with aggregation behaviour. Within this one year project, our aim is to test and explore the feasibility of a range of potential approaches to achieve this goal. Importantly, our experimental programme will use both model and therapeutically relevant proteins (including monoclonal antibodies). In close collaboration with the pharmaceutical industry the devices identified in these preliminary studies will also be assessed for viability and scale up for use in an industrial context. A sizable and increasing academic and industrial community have identified the occurence of protein aggregation as a critical issue in a number of fields, and have sought new methods to study this phenomenon. This project has the potential to provide an entirely new approach to detecting and investigating the origins of aggregation at its earliest stage. In the longer-term, the work, if built upon will impact upon significant areas of biotechnology and healthcare. Indeed, the methods developed could play an important role in bringing new generation medicines to the market in a cost-efficient and timely manner, and would thus have a very significant impact on public health and quality of life.