CN111450046A - 一种新冠病毒免疫凝胶制剂的合成方法 - Google Patents
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Abstract
本发明公布一种新冠病毒免疫凝胶制剂的合成方法。主要步骤包括:1)PLGA‑PEG‑PLGA凝胶溶液的制备；2)混合法制备SeV膜蛋白‑新冠S蛋白@ICG病毒免疫凝胶。SeV膜蛋白可全身性激活机体免疫功能，形成肌体对新冠病毒的免疫反应，并形成免疫记忆。吲哚菁绿(ICG)的荧光效应使疫苗接种的过程可视化，其产生的光热效应也可清除剩余的病毒。PLGA‑PEG‑PLGA无毒，生物相容性好，在常温下呈液态，35度左右相变为凝胶，可以很好地用于体内注射，延长药物在体内的滞留，从而提高效果。新冠病毒免疫凝胶可很好地预防新冠病毒的感染。
Description
技术领域
本发明涉及凝胶制剂的合成技术领域,具体涉及一种通过PLGA-PEG-PLGA包裹新冠病毒S蛋白、吲哚菁绿(ICG)和SeV膜蛋白的策略,合成病毒免疫凝胶制剂的方法。
背景技术
目前，冠状病毒是世界上对人类威胁最大的流行性疾病之一。免疫治疗通过激活人体免疫系统，依靠自身免疫机能杀灭病毒，从而达到预防病毒感染的作用。鉴于SeV膜蛋白可以作为异物在体内可以激发全身免疫系统产生细胞因子风暴(如干扰素和各种白细胞介素等)，并使在抗病毒过程中具有重要作用的树突状细胞(DC细胞，提呈病毒抗原)，巨噬细胞(RAW细胞，吞噬病毒)和自然杀伤细胞(NK细胞，杀伤病毒)等被大量激活，随后大幅提高T细胞的增值效率。这种免疫凝胶通过激活患者自身免疫系统，利用病毒的特征S蛋白诱导机体的特异性细胞免疫和体液免疫反应，增强机体的抗病毒能力，阻止病毒的感染，以达到清除或控制病毒扩散的目的。
ICG化学名为吲哚菁绿，是一种感光染料，是美国食品药品监督管理局(FDA)惟一批准的体内应用染料。它注入血液后会迅速与清蛋白及α1-脂蛋白结合(98％)，随血液经过肝脏时，90％以上被肝细胞摄取，再以原形由胆道排泄，不参与体内化学反应，无肠肝循环，无淋巴逆流，不从肾脏等肝外脏器排泄，无辐射，无毒副作用。
PLGA-PEG-PLGA是一种三嵌段聚合物。其中PLGA由两种单体——乳酸和羟基乙酸随机聚合而成，是一种可降解的功能高分子有机化合物，其降解产物是乳酸和羟基乙酸，同时也是人代谢途径的副产物，所当它应用在医药和生物材料中时不会有毒副作用，具有良好的生物相容性、无毒、良好的成囊和成膜的性能。PEG具有良好的水溶性，生物相容性好，并赋予了聚合物温敏相变的性质。
发明内容
本发明为克服现有技术的不足，提供一种通过PLGA-PEG-PLGA包裹新冠病毒S蛋白、吲哚菁绿(ICG)和SeV膜蛋白的策略,合成病毒免疫凝胶制剂的方法。利用病毒S蛋白诱导机体的特异性细胞免疫和体液免疫反应，增强机体的抗病毒能力，阻止病毒的感染，以达到清除或控制病毒扩散的目的。
本发明的技术方案是一种病毒免疫凝胶制剂的合成方法，通过PLGA-PEG-PLGA包裹新冠病毒S蛋白，吲哚菁绿(ICG)和SeV病毒膜蛋白的策略,具体步骤如下：
1)称取PLGA-PEG-PLGA材料，加入水中溶解，得到浓度为0.2-0.5mg/ml的PLGA溶液；
2)称取ICG材料，加入水中溶解，得到浓度为5-10mg/ml的ICG溶液；
3)各取适量混合成病毒免疫凝胶。
2、根据权利要求1所述的仿生模拟合成病毒免疫凝胶制剂的方法，其特征是，所述步骤3)具体如下：
(1)在室温下将盛有1ml的PLGA-PEG-PLGA溶液的单口瓶置于旋转搅拌器上，搅拌速度设定为500r/min；
(2)边搅拌边逐滴加入8uL的ICG水溶液和20uL病毒S蛋白溶液和20uL SeV膜蛋白溶液，继续搅拌直至完全混匀，最终得到SeV膜蛋白-新冠S蛋白@ICG病毒免疫凝胶。
本发明的优势在于：
1)SeV膜蛋白可以激活细胞免疫,增强机体的抗病毒能力，阻止病毒的感染，以达到清除或控制病毒扩散的目的。
2)吲哚菁绿(ICG)的荧光效应使疫苗接种的过程可视化，其产生的光热效应也可清除剩余的病毒蛋白。
3)PLGA-PEG-PLGA无毒，生物相容性好，在常温下呈液态，35度左右相变为凝胶，可以很好地用于体内注射，延长药物在体内的滞留，从而提高效果。
附图说明
图1：施用免疫凝胶后小鼠血液和腹腔液中干扰素数值柱状图，I组为空白组，II组为实验组。
具体实施方式
以下结合附图和具体实施例来对本发明作进一步的说明。
实施例1：
1)准确称取0.25mg的PLGA-PEG-PLGA材料，加入1mL水溶解，得到浓度为0.25mg/ml的PLGA-PEG-PLGA水溶液。
2)准确称取5mg的ICG材料，加入1mL水溶解，得到浓度为5mg/ml的ICG水溶液。
3)混合法合成SeV膜蛋白-新冠S蛋白@ICG病毒免疫凝胶的方法如下：
(1)在室温下将盛有1ml的PLGA-PEG-PLGA溶液的单口瓶置于旋转搅拌器上，搅拌速度设定为500r/min；
(2)边搅拌边逐滴加入8uL的ICG水溶液和20uL病毒S蛋白溶液和20uL SeV膜溶液，继续搅拌直至完全混匀，最终得到SeV膜蛋白-新冠S蛋白@ICG病毒免疫凝胶。
实施例2：
1)准确称取0.2mg的PLGA材料，加入1mL水溶解，得到浓度为0.2mg/ml的PLGA有机溶液。
2)准确称取8mg的ICG材料，加入1mL水溶解，得到浓度为8mg/ml的ICG水溶液。
3)混合法合成SeV膜蛋白-新冠S蛋白@ICG病毒免疫凝胶的方法如下：
(1)在室温下将盛有1ml的PLGA-PEG-PLGA溶液的单口瓶置于旋转搅拌器上，搅拌速度设定为500r/min；
(2)边搅拌边逐滴加入6uL的ICG水溶液和20uL病毒S蛋白溶液和20uL SeV膜蛋白溶液，继续搅拌直至完全混匀，最终得到SeV膜蛋白-新冠S蛋白@ICG病毒免疫凝胶。
实施例3：
1)准确称取0.4mg的PLGA材料，加入1mL水溶解，得到浓度为0.4mg/ml的PLGA有机溶液。
2)准确称取6mg的ICG材料，加入1mL水溶解，得到浓度为6mg/ml的ICG水溶液。
3)混合法合成SeV膜蛋白-新冠S蛋白@ICG病毒免疫凝胶的方法如下：
(1)在室温下将盛有1ml的PLGA-PEG-PLGA溶液的单口瓶置于旋转搅拌器上，搅拌速度设定为500r/min；
(2)边搅拌边逐滴加入10uL的ICG水溶液和20uL病毒S蛋白溶液和20uL SeV膜蛋白溶液，继续搅拌直至完全混匀，最终得到SeV膜蛋白-新冠S蛋白@ICG病毒免疫凝胶。
Claims (2)
1.一种新冠病毒免疫凝胶制剂的合成方法，其特征是，通过PLGA-PEG-PLGA包裹新冠病毒S蛋白、吲哚菁绿(ICG)和免疫激活剂SeV流感病毒膜蛋白的策略,具体步骤如下：
1)称取PLGA-PEG-PLGA材料，加入水中溶解，得到浓度为0.2-0.5mg/ml的PLGA溶液；
2)称取ICG材料，加入水中溶解，得到浓度为5-10mg/ml的ICG溶液；
3)各取6-10uL的ICG水溶液和20uL病毒S蛋白溶液和20uLCpG溶液加入1mlPLGA-PEG-PLGA水溶液中混合混合成病毒免疫凝胶。
2.根据权利要求1所述的新冠病毒免疫凝胶制剂的合成方法，其特征是，所述步骤3)具体如下：
(1)在室温下将盛有1ml的PLGA-PEG-PLGA溶液的单口瓶置于旋转搅拌器上，搅拌速度设定为500r/min；
(2)边搅拌边逐滴加入8uL的ICG水溶液和20uL新冠病毒S蛋白溶液和20uL SeV膜蛋白溶液，继续搅拌直至完全混匀，最终得到SeV膜蛋白-新冠S蛋白@ICG病毒免疫凝胶。
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| WEN-HSIANG CHEN ET AL.: ""The SARS-CoV-2 Vaccine Pipeline: an Overview"", 《CURRENT TROPICAL MEDICINE REPORTS》 * |
| 管晓燕等: ""温敏型生物降解水凝胶载体防龋基因疫苗pVAX1-spap/A经不同途径免疫新西兰大白兔的实验研究"", 《口腔医学研究》 * |
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