Centre for Pharmaceutical Nanotechnology, Department of Pharmaceutics, National Institute of Pharmaceutical Education and Research (NIPER), Sector 67, Phase X, SAS Nagar (Mohali), Punjab 160062, India.
Department of Pharmacoinformatics, National Institute of Pharmaceutical Education and Research (NIPER), Sector 67, Phase X, SAS Nagar (Mohali), Punjab 160062, India.
Int J Pharm. 2018 Jun 10;544(1):1-13. doi: 10.1016/j.ijpharm.2018.04.009. Epub 2018 Apr 7.
Amphotericin B is an antimicrobial membrane-acting drug used in the treatment of systemic fungal infections. However, the clinical utility of AmB is often low as a result of (i) dose-limiting toxicity which is closely associated with its aggregation wherein the selectivity for its target i.e. ergosterol in fungal membranes is diminished and (ii) limited oral bioavailablity. The latter is attributed to the unfavorable physicochemical properties of the AmB e.g., low solubility, gastrointestinal instability, and poor intestinal permeability. The hypothesis of present work was that by applying a lipid conjugation approach the aggregation induced toxicity of AmB vis-à-vis permeability can be overcome. From the array of fatty acids, the oleic acid (OA) was selected for conjugation due to its great impact on increasing the Caco-2 permeability of AmB. AmB-OA conjugate was synthesized using standard carbodiimide chemistry and characterized thoroughly. Due to the reported strong correlation between the self-aggregation of AmB and toxicity, the aggregation behavior of AmB and AmB-OA was studied by in silico modeling and confirmed experimentally. In vitro hemolytic studies and viability assays in kidney cells (HEK 293 cells) suggested that AmB in aggregated was state highly toxic but not AmB-OA. In silico modeling suggested possible aggregation conformation of AmB-OA dimers that retains the selectivity for cholesterol even in aggregated state when embedded in in silico generated lipid bilayers. The results were further confirmed by assessing the interactions of monomeric and aggregated state of AmB and AmB-OA with that of cholesterol and ergosterol containing liposomes employing circular dichroism spectroscopy. The findings were subsequently corroborated by in vivo nephrotoxicity studies. To conclude, the lipid conjugation approach may be a promising strategy for reducing the dose-limiting toxicity of AmB.
两性霉素 B 是一种抗微生物膜作用药物，用于治疗系统性真菌感染。然而，两性霉素 B 的临床应用往往受到限制，原因有二：（i）剂量限制毒性，这与它的聚集密切相关，其靶向真菌膜中的麦角固醇的选择性降低；（ii）有限的口服生物利用度。后者归因于两性霉素 B 的不利物理化学性质，例如低溶解度、胃肠道不稳定和差的肠道通透性。本工作的假设是，通过应用脂质缀合方法，可以克服两性霉素 B 的聚集诱导毒性和通透性。在一系列脂肪酸中，由于油酸（OA）对增加两性霉素 B 的 Caco-2 通透性具有很大的影响，因此选择 OA 进行缀合。两性霉素 B-OA 缀合物是使用标准的碳二亚胺化学合成的，并进行了全面的表征。由于两性霉素 B 的自聚集与其毒性之间存在很强的相关性，因此通过计算机模拟研究了两性霉素 B 和两性霉素 B-OA 的聚集行为，并通过实验进行了验证。体外溶血研究和肾脏细胞（HEK 293 细胞）活力测定表明，聚集状态的两性霉素 B 高度有毒，但两性霉素 B-OA 则不然。计算机模拟表明，两性霉素 B-OA 二聚体可能存在聚集构象，即使在嵌入计算机生成的脂质双层中处于聚集状态时，也能保持对胆固醇的选择性。通过评估单体和聚集状态的两性霉素 B 和两性霉素 B-OA 与含有胆固醇和麦角固醇的脂质体的相互作用，进一步证实了这一结果，方法是使用圆二色性光谱。随后，通过体内肾毒性研究对这些结果进行了验证。总之，脂质缀合方法可能是降低两性霉素 B 剂量限制毒性的一种有前途的策略。
