多次玻璃体内注射,这是痛苦和昂贵的,在视网膜疾病的治疗中通常需要。因此,目前正在评估一种使用水凝胶的新型药物递送系统作为替代方案。本研究旨在评估四臂聚乙二醇(tetra-PEG)凝胶的体外缓释能力。贝伐单抗负载的四-PEG凝胶和5-羧基荧光素N-琥珀酰亚胺酯(FAM-NHS)标记的IgG负载的四-PEG凝胶通过混合四-PEG与硫醇末端(四-PEG-SH)溶液制备,马来酰亚胺末端(四-PEG-MA)溶液,和贝伐单抗或FAM-NHS标记的IgG。凝胶是用三种不同的聚合物浓度为1.5%,5%,10%,然后进行体外释放研究以评估负载药物的四PEG凝胶的持续释放能力。使用高效液相色谱(HPLC)来测试从四-PEG凝胶释放的贝伐单抗的结构稳定性。使用SDS-聚丙烯酰胺凝胶电泳(PAGE)评估贝伐单抗与四-PEG-SH或MA的结合。使用KDR/NFAT-REHEK293细胞测试释放的贝伐单抗的生物活性。此外,还进行了体外降解和肿胀研究。体外释放分析表明,贝伐单抗在5%和10%四-PEG凝胶中的释放比1.5%四-PEG凝胶中的释放更慢。同样,FAM-NHS标记的IgG的释放最慢,为1.5%,5%,和10%的四PEG凝胶,按这个顺序。5%和10%四-PEG凝胶在1-2周的时间内释放贝伐单抗和FAM-NHS标记的IgG。贝伐单抗和FAM-NHS标记的IgG均未在2周内完全释放。HPLC分析显示,从装载贝伐单抗的四PEG凝胶释放的样品的保留时间与贝伐单抗标准品的保留时间相似。SDS-PAGE分析显示贝伐单抗结合四-PEG-MA。生物活性测定测试显示释放的贝伐单抗的生物活性没有降低。体外降解和溶胀研究表明,1.5%,5%,和10%的四PEG凝胶膨胀约1.4-,2-,和3倍,分别。根据释放和溶胀试验的结果,5%四-PEG凝胶被认为是治疗性抗体如贝伐单抗的控释系统的良好候选物。PEG与治疗性抗体的结合可降低可释放的治疗性抗体的可用性。
Multiple intravitreal injections, which are painful and costly, are often required in the treatment of retinal disorders. Therefore, a novel drug delivery system using hydrogels is currently being evaluated as an alternative. This study aimed to evaluate the ability of tetra-armed polyethylene glycol (tetra-PEG) gel for sustained release in vitro. Bevacizumab-loaded tetra-PEG gel and 5-Carboxyfluorescein N-succinimidyl ester (FAM-NHS)-labeled IgG-loaded tetra-PEG gel were prepared by mixing tetra-PEG with thiol termini (tetra-PEG-SH) solution, maleimide termini (tetra-PEG-MA) solution, and bevacizumab or FAM-NHS labeled IgG. The gels were prepared with three different polymer concentrations of 1.5%, 5%, and 10%, then an in vitro release study performed to assess the sustained release ability of the drug-loaded tetra-PEG gels. High performance liquid chromatography (HPLC) was used to test the structural stability of the bevacizumab released from the tetra-PEG gel. The binding of bevacizumab to tetra-PEG-SH or MA was assessed using SDS-polyacrylamide gel electrophoresis (PAGE). The bioactivity of released bevacizumab was tested using KDR/NFAT-RE HEK293 cells. In addition, in vitro degradation and swelling studies were also performed. The in vitro release analysis showed that the release of bevacizumab was slower in the 5% and 10% tetra-PEG gels than that of 1.5% tetra-PEG gels. Similarly, the release of FAM-NHS-labeled IgG was slowest in the 1.5%, 5%, and 10% tetra-PEG gels, in that order. The 5% and 10% tetra-PEG gels released bevacizumab and FAM-NHS-labeled IgG over a period of 1-2 weeks. Both bevacizumab and FAM-NHS-labeled IgG were not fully released in 2 weeks. HPLC analysis showed that the retention time of the samples released from the bevacizumab-loaded tetra-PEG gel was similar to that of the bevacizumab standard. The SDS-PAGE analysis showed that bevacizumab binds to tetra-PEG-MA. The bioactivity assay test revealed no decrease in the bioactivity of the released bevacizumab. In vitro degradation and swelling studies revealed that 1.5%, 5%, and 10% tetra-PEG gels expanded by approximately 1.4-, 2-, and 3-fold, respectively. Based on the results of the release and swelling tests, 5% tetra-PEG gels are considered good candidates for controlled release systems for therapeutic antibodies such as bevacizumab. The binding of PEG to the therapeutic antibodies may reduce the availability of therapeutic antibodies that can be released.