Pharmaceutical Technology and Biopharmaceutics, Department of Pharmacy, Ludwig-Maximilians-Universitaet Muenchen, Munich, 81377, Germany.
Helmholtz Centre for Infection Research (HZI), Biogenic Nanotherapeutics Group (BION), Helmholtz Institute for Pharmaceutical Research Saarland (HIPS), Campus E8.1, Saarbruecken, 66123, Germany.
Adv Healthc Mater. 2022 Mar;11(5):e2100538. doi: 10.1002/adhm.202100538. Epub 2021 Jul 26.
Extracellular vesicles (EV) are an emerging technology as immune therapeutics and drug delivery vehicles. However, EVs are usually stored at -80 °C which limits potential clinical applicability. Freeze-drying of EVs striving for long-term stable formulations is therefore studied. The most appropriate formulation parameters are identified in freeze-thawing studies with two different EV types. After a freeze-drying feasibility study, four lyophilized EV formulations are tested for storage stability for up to 6 months. Freeze-thawing studies revealed improved colloidal EV stability in presence of sucrose or potassium phosphate buffer instead of sodium phosphate buffer or phosphate-buffered saline. Less aggregation and/or vesicle fusion occurred at neutral pH compared to slightly acidic or alkaline pH. EVs colloidal stability can be most effectively preserved by addition of low amounts of poloxamer 188. Polyvinyl pyrrolidone failed to preserve EVs upon freeze-drying. Particle size and concentration of EVs are retained over 6 months at 40 °C in lyophilizates containing 10 mm K- or Na-phosphate buffer, 0.02% poloxamer 188, and 5% sucrose. The biological activity of associated beta-glucuronidase is maintained for 1 month, but decreased after 6 months. Here optimized parameters for lyophilization of EVs that contribute to generate long-term stable EV formulations are presented.
细胞外囊泡 (EV) 作为免疫治疗和药物递送载体是一种新兴技术。然而，EV 通常储存在-80°C 下，这限制了其潜在的临床应用。因此，人们正在研究 EV 的冻干技术，以获得长期稳定的配方。通过两种不同类型 EV 的冻融研究确定了最合适的配方参数。在进行冻干可行性研究后，对四种冻干 EV 配方进行了长达 6 个月的储存稳定性测试。冻融研究表明，与使用磷酸盐缓冲盐水相比，在含有蔗糖或磷酸钾缓冲液的情况下，EV 的胶体稳定性得到了改善。与略酸性或碱性 pH 值相比，在中性 pH 值下，聚集和/或囊泡融合的发生较少。通过添加少量泊洛沙姆 188 可以最有效地保存 EV 的胶体稳定性。聚乙烯吡咯烷酮在冻干时未能保存 EV。在含有 10mM K-或 Na-磷酸盐缓冲液、0.02%泊洛沙姆 188 和 5%蔗糖的冻干物中，EV 的粒径和浓度在 40°C 下可保持 6 个月。与 EV 相关的β-葡糖苷酸酶的生物活性可维持 1 个月，但 6 个月后会下降。本文提出了优化 EV 冻干的参数，有助于生成长期稳定的 EV 配方。
