Paál K, Müller J, Hegedûs L
Department of Pharmaceutical Research and Development, HUMAN Co. Ltd, Gödöllõ, Hungary.
Eur J Biochem. 2001 Apr;268(7):2187-91. doi: 10.1046/j.1432-1327.2001.02107.x.
Paclitaxel, a very potent antitumor agent is a hydrophobic molecule with low aqueous solubility. Its currently used formula (Taxol) contains the drug in a 1 : 1 (v/v) mixture of ethanol and Cremophor EL. To minimize vehicle-related toxicity, we developed a novel, water-soluble formulation in which paclitaxel is bound noncovalently to human serum albumin. For this purpose, studies of the paclitaxel-albumin binding equilibrium were performed. Paclitaxel dissolved in ethanol was added to the aqueous solution of human serum albumin. Precipitated paclitaxel was removed and unbound drug was separated by ultrafiltration. Paclitaxel concentration was measured by RP-HPLC. Binding data were evaluated based both on the Scatchard plot and the general binding equation describing binding equilibria with the stepwise stoichiometric binding constants. The Scatchard plot was found to be curvilinear with a slight positive slope of the final part. Parameters of high affinity specific binding were determined from the initial part of the curve (nsp = 1.3 and Ksp = 1.7 x 10(6) M(-1)). Stoichiometric binding constants were estimated by fitting the general binding equation to the experimental data (K1 = 2.4 x 10(6) M(-1) and K2 = 1.0 x 10(5) M(-1)). Saturation of the protein with paclitaxel, similarly to other ligands of albumin, could not be reached. The greatest observed value of r (number of paclitaxel molecules bound to one albumin molecule) was 6.6.
紫杉醇是一种非常有效的抗肿瘤药物，是一种疏水性分子，水溶性低。其目前使用的配方（泰素）中药物以乙醇和聚氧乙烯蓖麻油EL按1:1（v/v）的混合物形式存在。为了将与载体相关的毒性降至最低，我们开发了一种新型的水溶性制剂，其中紫杉醇与人类血清白蛋白非共价结合。为此，进行了紫杉醇-白蛋白结合平衡的研究。将溶解在乙醇中的紫杉醇加入到人类血清白蛋白的水溶液中。去除沉淀的紫杉醇，通过超滤分离未结合的药物。通过反相高效液相色谱法测定紫杉醇浓度。基于Scatchard图和描述具有逐步化学计量结合常数的结合平衡的一般结合方程对结合数据进行评估。发现Scatchard图呈曲线状，最后部分有轻微的正斜率。从曲线的初始部分确定高亲和力特异性结合的参数（nsp = 1.3，Ksp = 1.7×10⁶ M⁻¹）。通过将一般结合方程拟合到实验数据来估计化学计量结合常数（K1 = 2.4×10⁶ M⁻¹，K2 = 1.0×10⁵ M⁻¹）。与白蛋白的其他配体类似，无法使蛋白质被紫杉醇饱和。观察到的最大r值（与一个白蛋白分子结合的紫杉醇分子数）为6.6。
