A Thermo-responsive Biopharmaceutical to Enhance the Tear Production of Lacritin

一种增强泪液分泌的热响应生物药物

• 批准号: 8448449
• 负责人: Sandeep Samudre
• 金额: $ 14.64万
• 依托单位: EYERX RESEARCH, INC.
• 依托单位国家: 美国
• 财政年份: 2013
• 资助国家: 美国
• 起止时间: 2013-09-30 至 2015-09-29
• 项目状态: 已结题
• 来源: https://reporter.nih.gov/project-details/8448449
• 关键词: Adhesives; Adverse effects; Affect; American; Behavior; Biocompatible; Biological Products; Body Temperature; Characteristics; Data; Development; Disease; Dose; Drainage procedure; Drops; Drug Delivery Systems; Drug Formulations; Drug vehicle; Dry Eye Syndromes; Elastin; Elderly; Equilibrium; Evaluation; Eye; Eyedrops; Figs - dietary; Film; Frequencies; Gel; Genetic Engineering; Glycoproteins; Goals; Heating; Hour; Human; In Vitro; Label; Length; Liquid substance; Modeling; Molecular Weight; Nanostructures; Oryctolagus cuniculus; Peptides; Pharmaceutical Preparations; Pharmacologic Substance; Phase; Phase Transition; Polymers; Postmenopause; Process; Production; Property; Proteins; Recombinants; Residencies; Route; Schedule; Solutions; Temperature; Time; Toxic effect; Transition Temperature; Tropoelastin; Viscosity; Woman; aqueous; combat; design; environmental change; eye dryness; in vivo; innovation; light scattering; macromolecule; novel; polypeptide; prevent; protein purification; public health relevance; research study; residence; scaffold; self assembly; small molecule

Project Summary/Abstract Ocular pharmaceuticals predominantly use the topical route of administration, which involves a number of benefits and limitations when compared to systemic drug routes. A major limitation of the ocular topical route is the rapid loss of drug via tearing and nasolacrimal drainage, requiring the use of frequent dosing and high drug concentrations that then create formulation difficulties, together with the potential for local and systemic adverse effects. The proposed innovation is to use the temperature sensitivity of the genetically engineered smart polymer class of elastin-like polypeptides (ELP) to control ocular clearance. ELPs have unique properties that promote phase separation, recombinant expression, protein purification, and self-assembly of nanostructures. ELPs are repeated pentameric peptides, (VPGXG)n, that have characteristic inverse phase transition temperatures, Tt, above which they phase separate from aqueous solution. By selecting X and the length n, ELPs of different Tt can be efficiently and precisely biosynthesized. Genetically engineered ELPs are pharmacologically relevant, being monodisperse, biodegradable, and biocompatible. Lacritin, a new-found, short glycoprotein promotes tear secretion in dry eye models but must be administered several times a day. Lacritin will be genetically fused with ELPs of different transition temperatures and molecular weights to optimize the construct for drug retention. Free, active lacritin is expected to be in equilibrium with the ELP aggregates, thus extending ocular lacritin residency. ELPs with a transition temperature greater than 37¿C should clear quickly from the eye; however, ELPs with transition temperature between room temperature (25¿C) and body temperature (37¿C) are expected to drain slowly. The following specific aims are designed to serve as a proof of concept of this idea: SA1) Synthesis and in vitro demonstration of lacritin-ELP thermal sensitivity. Thermally sensitive and insensitive ELPs with and without fluorescent labels will be prepared with molecular weights ranging from 10 to 50 kD and fused to lacritin. Our milestone will be to produce a thermally sensitive ELP-lacritin that will have a transition temperature of between 25 and 37¿C, suitable for use in an eye drop. SA2) Efficacy and tolerability evaluation of lacritin-ELP in in vivo normal rabbit and in vitro Ussing chamber ocular models. Optimal fusion peptides will be evaluated for residency, ocular tolerability and lacritin-stimulated tear formation in in vivo normal rabbit models and in in vitro Ussing chamber residency experiments. Our milestone is to demonstrate that the thermally-sensitive lacritin-ELP is more effective than the thermally-insensitive product at increasing tear production without significant local or systemic toxicity.

项目概要/摘要 眼部药物主要使用局部给药途径，其中涉及许多 与全身药物途径相比的优点和局限性 眼部局部途径的主要局限性是。 药物通过撕裂和鼻泪管引流迅速流失，需要频繁给药和高剂量药物 浓度会造成配制困难，以及局部和全身潜在的潜在风险 所提出的创新是利用基因工程的温度敏感性。 智能聚合物类弹性蛋白样多肽 (ELP) 具有独特的控制眼部清除能力。 促进相分离、重组表达、蛋白质纯化和自组装的特性 ELP 是重复的五聚肽 (VPGXG)n，具有反相特征。 转变温度 Tt，高于该温度时，通过选择 X 和 ，它们会从水溶液中发生相分离。 长度n，不同Tt的ELP可以被高效、精确地生物合成。 具有药理学相关性，具有单分散性、可生物降解性和生物相容性，是一种新发现的、 短糖蛋白可促进干眼模型的泪液分泌，但必须每天给药数次。 Lacritin 将与不同转变温度和分子量的 ELP 进行基因融合， 优化平衡药物保留的结构，预计会与 ELP 结合在一起。 聚集体，从而延长了 ELP 的眼部驻留时间，其转变温度大于 37°。 C 然而，ELP 的过渡温度在室温之间，应该会很快从眼睛中消失； (25°C) 和体温 (37°C) 预计会缓慢排出以下具体目标。 作为这个想法的概念证明：SA1）lacritin-ELP热的合成和体外演示 具有和不具有荧光标记的热敏感和不敏感ELP将通过以下方法制备。 分子量范围为 10 至 50 kD，并与乳泌素融合，我们的里程碑将是生产热热的。 敏感的 ELP-乳泌素，其转变温度在 25 至 37 之间C、适合用在眼睛上 SA2) Lacritin-ELP在正常兔体内和体外的功效和耐受性评价。 使用腔眼模型评估最佳融合肽的驻留性、眼耐受性。 体内正常兔模型和体外Ussing室驻留实验中泪液素刺激泪液形成 我们的里程碑是证明热敏乳泌素-ELP 比它更有效。 热不敏感产品，可增加泪液产生，且无显着的局部或全身毒性。