Modification of insulin molecule for controlled delivery at basal level from triblock copolymers

修饰胰岛素分子以从三嵌段共聚物控制基础水平递送

基本信息

批准号：
8876907
负责人：
Jagdish Singh
金额：
$ 34.8万
依托单位：
NORTH DAKOTA STATE UNIVERSITY
依托单位国家：
美国
项目类别：
财政年份：
2015
资助国家：
美国
起止时间：
2015-05-01 至 2019-12-31
项目状态：
已结题

来源：
https://reporter.nih.gov/project-details/8876907
关键词：
American Animal Model Autoimmune Diseases Biocompatible Biological Assay Blood Body Temperature Body Weight Bolus Infusion Bromides Cerebrum Charge Child Chitosan Circular Dichroism Complex Development Diabetes Mellitus Diffuse Diffusion Drug Delivery Systems Drug Formulations Environment Enzyme-Linked Immunosorbent Assay Equilibrium Exhibits Foreign Bodies Forms Controls Gel Gel Chromatography Glucose Goals Health Hepatic Hydrogels Implant In Situ In Vitro Injection of therapeutic agent Insulin Insulin Antibodies Insulin-Dependent Diabetes Mellitus Length MALDI-TOF Mass Spectrometry Measures Modification Molecular Weight Non-Insulin-Dependent Diabetes Mellitus Patients Pharmaceutical Preparations Physiological Polyacrylamide Gel Electrophoresis Polymers Principal Investigator Production Property Proteins Quality of life Rattus Research Sampling Serum Site Sodium Dodecyl Sulfate Solutions Spectroscopy, Fourier Transform Infrared Stream Subcutaneous Injections System Techniques Temperature Testing Therapeutic Transition Temperature Water Weight Zinc absorption aqueous basal insulin base biodegradable polymer biomaterial compatibility controlled release copolymer diabetic diabetic rat dimer glucose output improved in vivo insulin dimers insulin secretion meetings monomer novel novel strategies poly(lactic acid)poly(lactic acid-ethylene glycol)programs public health relevance response subcutaneous therapeutic protein young adult

项目摘要

DESCRIPTION (provided by applicant): There are two types of insulin secretion in healthy people- basal and stimulated. The basal insulin is secreted continuously between meals and throughout the night at a rate of 0.5-1 unit/h. Although the basal insulin level is low, it modulats the rate of overnight hepatic glucose and glucose output during prolonged periods between meals. This allows for sufficient glucose level for cerebral energy production at bedtime. There is not a single controlled release delivery system available to deliver insulin continuously for longer duration after a single subcutaneous injection to meet the needs of the basal insulin. Burst release is a major issue in controlled delivery of therapeutics from polymeric formulations. In this application, we are proposing to modify the insulin molecule in order to eliminate the burst release. We propose a novel approach of modifying insulin molecule by utilizing the combination of distinctive properties of insulin (self-association in presence of zinc, and abilityto interact with cationic polymers such as chitosan) and subsequent addition of the complex into aqueous solution of thermo sensitive polymer for controlled delivery of insulin. Thermo sensitive polymer is solution in water at room temperature and turns into implant at body temperature at the site of injection. The proposed delivery system would circumvent the problems associated with insulin burst release by reducing the diffusion of zinc insulin-chitosan complex from the polymeric hydrogel matrix due to its larger size and help to stabilize the protein inside the delivery system, while providing controlled release of insulin at basal level. The long-term goal of this project is to develop novel controlled release delivery systems which can deliver insulin a basal level in a conformationally and chemically stable and biologically active form for longer duration (~3 months) after a single subcutaneous injection in patients with type 1 diabetes. We propose to study three specific aims: (1)To synthesize temperature sensitive poly (lactic acid)-poly (ethylene glycol)- poly(lactic acid) (PLAPEG-PLA) triblock copolymers with varying PLA and PEG chain lengths and characterize their critical gel concentration, gel transition temperature, weight average molecular weight by gel permeation chromatography (GPC), and number average molecular weight by 1H NMR. In situ gel-forming controlled delivery systems of insulin will be prepared, using the above copolymers and zinc-insulin-chitosan complex, and will be studied for the in vitro release of insulin. Further, the stability of insulin in the released samples and in the delivery systems will be investigated using Fourier transform infrared spectroscopy (FTIR), circular dichroism (CD), sodium dodecyl sulfate-polyacrylamide gel electrophoresis (SDS PAGE), native PAGE and Matrix-assisted laser desorption/ionization time-of-flight (MALDI-TOF) mass spectrometry techniques. (2). To evaluate the biocompatibility of polymeric delivery systems in vitro by 3-(4, 5-dimethylthiazol-2-yl)-2,5-diphenyltetrazolium bromide (MTT) assay and in vivo in rats by histological analysis. (3). To study in vivo absorption and bioactivity of insulin from the delivery systems in the streptozoticin (STZ) induced diabetic rats by measuring serum insulin and glucose levels, respectively. The anti-insulin antibody in serum will be assessed by ELISA in order to rule out the possibility of foreign body response. STZ induced diabetes leads to reduction in the body weight. Therefore the change in body weight in diabetic rats before and after induction of diabetes, as well as after insulin treatment will be evaluated. The proposed study will contribute significantly to the development of a novel delivery system to deliver insulin at a controlled rate for three months after a single subcutaneous injection in order to improve patients' quality of life, and decrease the long term complications associated with diabetes.

描述（由适用提供）：健康的人群中有两种类型的胰岛素分泌 - 基础和刺激。基础胰岛素在饭菜之间和整个夜晚之间连续分泌，以0.5-1单位/h。尽管基底胰岛素水平较低，但它在两餐之间的长时间内调节了过夜的肝葡萄糖和葡萄糖输出的速率。这允许在就寝时间产生足够的葡萄糖水平来产生脑能量。单个皮下注射后，没有一个可用的受控释放输送系统可以连续延长胰岛素，以满足基本胰岛素的需求。爆发是从聚合公式进行治疗的控制疗法方面的一个主要问题。在此应用中，我们提议修改胰岛素分子以消除爆发释放。我们提出了一种通过使用胰岛素的独特特性（存在锌的自我关联以及与阳离子聚合物（如壳聚糖）相互作用的能力）的新方法来修饰胰岛素分子，并随后将复合物添加到热敏聚合物的水溶液中，以控制胰岛素的递送。热敏感聚合物在室温下是水溶液，并在注射部位的体温下变成植入物。提出的输送系统将通过减少锌胰岛素 - 吉他络合物络合物从聚合物水凝胶基质中的扩散而避免与胰岛素爆发相关的问题，这是由于其尺寸较大，并有助于稳定递送系统内的蛋白质，同时在基本水平上提供控制的胰岛素释放。该项目的长期目标是开发新的受控释放递送系统，这些释放递送系统可以在1型糖尿病患者的单个皮下注射后，在构象和化学稳定和生物学活性的情况下以构象和化学稳定和生物活性形式提供基础水平。 We propose to study three specific aims: (1)To synthesize temperature sensitive poly (lactic acid)-poly (ethylene glycol)-poly(lactic acid) (PLAPEG-PLA) triblock copolymers with varying PLA and PEG chain lengths and characterize their critical gel concentration, gel transition temperature, weight average molecular weight by gel permeation chromatography (GPC), and number average分子量通过1H NMR。使用上述共聚物和锌 - 胰岛素 - 壳聚糖络合物将制备原位形成的胰岛素的受控递送系统，并将研究用于体外胰岛素的体外释放。此外，将使用傅立叶变换感染的光谱法（FTIR），圆形二色性（CD），十二烷基硫酸硫酸钠 - 丙烯酸丙烯酰胺凝胶电泳（SDS PAGE），本地页面和矩阵征服激光征服/离子化的时间范围（MASE IIN-IR-FLIMPLEST（MARDE），将研究释放样品中胰岛素和输送系统中胰岛素的稳定性。（2）。通过组织学分析，通过组织学分析评估3--（4，4-二甲基噻唑-2-基）的体外生物相容性 - 4（4，5-二甲基噻唑-2-基）-2,5-二苯基四唑烷（MTT）测定和体内通过组织学分析。（3）。通过分别测量血清胰岛素和葡萄糖水平，研究胰岛素（STZ）诱导的糖尿病大鼠的胰岛素的体内缓解和生物活性。 ELISA将评估血清中的抗胰岛素抗体，以排除异物反应的可能性。 STZ诱导的糖尿病会导致体重减轻。因此，将评估糖尿病前后糖尿病大鼠体重的变化以及胰岛素治疗后的体重变化。拟议的研究将对新型输送系统的发展产生重大贡献，以在单一皮下注射后三个月以受控速率传递胰岛素，以改善患者的生活质量，并减少与糖尿病相关的长期并发症。

项目成果

期刊论文数量（10）

专著数量（0）

科研奖励数量（0）

会议论文数量（0）

专利数量（0）

Synthesis and Characterization of Fatty Acid Grafted Chitosan Polymer and Their Nanomicelles for Nonviral Gene Delivery Applications.

DOI：
10.1021/acs.bioconjchem.7b00505
发表时间：
2017-11-15
期刊：
Bioconjugate chemistry
影响因子：
4.7
作者：
Sharma D;Singh J
通讯作者：
Singh J

Cell Penetrating Peptide Conjugated Chitosan for Enhanced Delivery of Nucleic Acid.

DOI：
10.3390/ijms161226142
发表时间：
2015-12-04
期刊：
International journal of molecular sciences
影响因子：
5.6
作者：
Layek B;Lipp L;Singh J
通讯作者：
Singh J

Chitosan-zinc-insulin complex incorporated thermosensitive polymer for controlled delivery of basal insulin in vivo.

DOI：
10.1016/j.jconrel.2012.07.035
发表时间：
2012-10-28
期刊：
Journal of controlled release : official journal of the Controlled Release Society
影响因子：
0
作者：
Oak M;Singh J
通讯作者：
Singh J

The Role of Cell-Penetrating Peptide and Transferrin on Enhanced Delivery of Drug to Brain.

细胞穿透肽和转铁蛋白在增强药物向大脑输送中的作用。