Polypeptide Multilayer Nanofilms for Synthetic Biomatrix Development

描述（由申请人提供）：SBIR一期项目将确定多肽多层纳米膜技术应用于合成生物基质开发和商业化的可行性。这些纳米膜是细胞和组织培养的一种表面涂层。合成生物基质材料的直接体外应用包括基础医学研究、诊断测试、高通量药物筛选、细胞表型选择、生物制剂生产和各种治疗细胞技术的开发等许多领域。据估计，全球合成生物基质市场的年价值接近10亿美元。组织中任何非细胞的部分称为细胞外基质（ECM）。在活体中，ECM包围细胞，影响细胞形态、增殖、活化和存活。科学研究已经揭示了许多分子信号，其中ECM影响细胞行为。研究还表明，细胞生长的表面在体外决定细胞行为和优化特定细胞类型的培养条件方面起着关键作用。多种方法用于修饰体外细胞培养的表面。在一种方法中，一种或多种来自哺乳动物外基质的蛋白质，例如胶原蛋白，通过非特异性吸附沉积。这种涂层提供了相对先进的功能，但ECM蛋白的纯化成本很高，它们必须冷藏，它们是从动物源制备的，并且它们自身控制体外细胞行为的能力有限。更复杂的产品，如MatrigelTM在某些用途上是有用的，但它们在分子组成方面定义不清，不适合许多用途。合成生物基质的功能是在不使用动物产品的情况下模仿ECM的某些特征，并且在分子组成方面具有高度定义。所制备的多肽多层纳米膜在体外细胞培养中具有明显的优势。这些纳米膜完全是人工合成的，它们可以模拟ECM蛋白的关键特征，并且可以针对特定的细胞类型和培养目标进行定制，这对于体外细胞行为的某些方面的控制很重要。此外，这些纳米膜可能在室温下长时间保持货架稳定性，并且与市场上现有产品相比具有成本竞争力。本研究主要包括两个部分：优化多肽纳米膜在多孔板中制备的可靠工艺，确定这些纳米膜的不同物理性质和生物功能对不同类型细胞的增殖、附着和形态的影响。如果多肽多层纳米膜相对于目前市场上的涂层产品能够可靠地控制细胞附着和增殖，那么将多肽多层纳米膜技术应用于合成生物基质开发和商业化的可行性将得到证明。哺乳动物细胞的体外培养已成为医学研究、诊断、药物筛选、治疗性蛋白的大规模生产以及其他与医疗保健有关的用途中不可或缺的一部分；对细胞培养方法发展的兴趣持续快速增长。多肽多层纳米膜是合成生物基质材料或人工细胞外基质的重要研究方向。这些材料在体外对细胞特性具有相当或更好的控制，在室温下的保质期延长，制造成本具有竞争力，它们可以促进对健康和疾病生物过程的了解，促进新的蛋白质和细胞疗法的发展。

DESCRIPTION (provided by applicant): This SBIR Phase I project will determine the feasibility of applying polypeptide multilayer nanofilm technology to synthetic biomatrix development and commercialization. These nanofilms are a type of surface coating for cell and tissue culture. Immediate in vitro uses of synthetic biomatrix materials include many areas of basic medical research, diagnostic testing, high-throughput drug screening, cell phenotype selection, biologics production, and development of various therapeutic cell technologies. The synthetic biomatrix market has an estimated annual value of close to $1 billion worldwide. Any part of a tissue that is not a cell is called the extracellular matrix (ECM). In the living body the ECM surrounds cells and influences cell shape, proliferation, activation, and survival. Scientific research has revealed numerous molecular signals whereby the ECM affects cell behavior. Research has also shown that the surface on which cells are grown plays a key role in determining cell behavior in vitro and in optimizing culture conditions for a particular cell type. A variety of methods are used to modify surfaces for in vitro cell culture. In one approach, one or more proteins from mammalian ECM, for example collagen, are deposited by non-specific adsorption. Such coatings offer relatively advanced functionality, but ECM proteins are expensive to purify, they must be refrigerated, they are prepared from an animal source, and they have a limited ability on their own to control cell behavior in vitro. More complex products such as MatrigelTM are useful for some purposes, but they are poorly defined with regard to molecular composition and unsuited for many purpose. The function of a synthetic biomatrix is to mimic some features of the ECM without the use of animal products and in a way that is highly defined with regard to molecular composition. The polypeptide multilayer nanofilms of the propose research offer significant advantages over existing coatings for in vitro cell culture. These nanofilms are entirely synthetic, they can mimic crucial features of ECM proteins, and they can be tailored to a particular cell type and culture objective, important for control of some aspects of cell behavior in vitro. Moreover, these nanofilms are likely to be shelf-stable at room temperature for extended periods and cost competitive with existing products in the marketplace. The proposed research has two main parts: optimization of a reliable process of polypeptide nanofilm fabrication in multi-well plates, and determination of the effect of different physical properties and biofunctionalization of these nanofilms on the proliferation, attachment, and morphology of various cell types. Feasibility of applying polypeptide multilayer nanofilm technology to synthetic biomatrix development and commercialization will have been demonstrated if polypeptide multilayer nanofilms are shown to enable reliable control over cell attachment and proliferation relative to coating products currently on the market.Project Narrative In vitro culture of mammalian cells has become indispensable for medical research, diagnostics, drug screening, large-scale production of therapeutic proteins, and other purposes touching on healthcare; interest in development of cell culture methods continues to grow rapidly. Polypeptide multilayer nanofilms are promising for the development of synthetic biomatrix materials, or artificial extracellular matrices. Such materials offer comparable or superior control over cell properties in vitro, extended shelf-life at room temperature, and cost-competitive manufacture, and they can advance knowledge of biological processes in health and disease and promote the development of new protein and cellular therapeutics.