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.

项目摘要/摘要 眼药主要使用局部给药途径，其中涉及多个 与全身药物路线相比，益处和局限性。眼局部途径的主要限制是 通过撕裂和鼻泪排水迅速损失药物，需要使用频繁给药和高药物 然后造成公式困难的浓度，以及局部和系统性的潜力 不利影响。提出的创新是使用基因工程的温度灵敏度 弹性蛋白样多肽（ELP）的智能聚合物类，以控制眼清除。 ELP具有独特性 促进相分离，重组表达，蛋白质纯化和自组装的特性 纳米结构。 ELP是重复具有特征逆相的五聚肽（VPGXG）N 过渡温度，TT，上面它们与水溶液分离。通过选择X和 长度为n，不同TT的ELP可以有效，精确地化为生物合成。基因工程ELP是 在药理上相关，是单分散，可生物降解和生物相容性的。曲息蛋白，新发现， 短糖蛋白在干眼模型中促进撕裂分泌，但每天必须几次给药。 流质蛋白通常与不同的过渡温度和分子量的ELP融合到 优化用于保留药物的结构。预计游离活性乳葡萄蛋白将与ELP保持平衡 聚集体，因此扩展了眼绿蛋白的住所。过渡温度大于37 c的ELP 应该从眼睛迅速清楚；但是，在室温之间具有过渡温度的ELP （25¿C）和体温（37°C）预计会缓慢排水。以下特定目标旨在 作为这一想法的概念证明：SA1）合成和体外演示曲息蛋白-ELP热形成 灵敏度。带有和不具有荧光标签的热敏感和不敏感的ELP将使用 分子量范围从10到50 kD，融合到透眼素。我们的里程碑将是热产生 敏感的ELP曲息蛋白的过渡温度在25至37°C之间，适用于眼睛 降低。 SA2）在体内正常兔子和体外的基质素-ELP的功效和耐受性评估 使用室内眼模型。最佳的融合宠物将评估居住，眼部耐受性 在体内正常兔模型和体外用途的室内住宅中，在体内刺激的泪液形成和流质蛋白刺激 实验。我们的里程碑是证明热敏感的曲息蛋白-ELP比 没有明显的局部或全身毒性的泪液产生时增加的热敏感产物。