摘要
吡柔比星(pirarubicin)是一种蒽环类抗生素,对白血病、恶性淋巴瘤、乳腺癌、头颈部癌、消化系统癌、肝癌和肺癌等多种实体肿瘤有较好的疗效。但在临床应用中,仍具有以下不良反应：1、骨髓抑制和胃肠道反应；2、累积心脏毒性：同阿霉素相比较低,但多次给药后具有蓄积毒性。脂质体,作为一种新型药物载体,可以改变药物在体内的分布形式和分布规律,因此在抗癌药物的载体研究中备受关注。
利用热分析(DSC)、红外光谱(FTIR)以及基于分子模拟的量子化学计算深入研究了吡柔比星与处方中不同磷脂之间的作用。DSC和FTIR的结果表明,在低浓度时药物很容易嵌入二硬脂酰磷脂酰胆碱(DSPC)的双分子层膜内,从而改变DSPC的相变行为,影响磷脂双层膜的流动性和磷脂的碳链构象组成。而基于分子模拟的量子化学计算结果则从分子动力学角度表明：由于磷脂头部的特殊结构,药物更易与二硬脂酰磷脂酰甘油(DSPG)的头部基团作用,从而抑制了药物进一步嵌入DSPG双分子层膜中。在上述处方的研究基础上,依据药物自身性质,结合工业生产的可操作性,设计出了适合工业化生产的脂质体新型制备方法,并论述了新型制备工艺的可能机制。
建立了吡柔比星的体外、体内血浆和各组织中药物的分析方法和处理方法,并进行了方法学考察。动物实验结果表明,吡柔比星制备成脂质体后：1)可以大大降低其在心脏中的聚集；2)容易在脾、肺中聚集；3)体内清除快；4)虽然在血浆中生物利用度降低；但其总生物利用度仍然是显著升高的。
采用组织病理切片、血清酶活性结合尿液代谢组学来研究吡柔比星脂质体的减毒机制。体重、行为、组织病理切片、血清酶活性结果表明市售吡柔比星注射液2次给药后对机体造成严重的损伤,而吡柔比星脂质体组则相对较低。代谢组学则从分子机制上阐明毗柔比星引发心脏毒性的本质原因：1、药物和糖苷类代谢物的蒽环类自由基引发活性氧(ROS)的浓度增高,从而引发急性心肌损伤；2、代谢物通过单电子还原产生的二级醇代谢物,干扰心肌细胞的能量代谢和细胞平衡,从而引发慢性心肌损伤。从组织分布实验中得到,吡柔比星制备成脂质体后,不仅将大大降低其在心脏的蓄积；同时加速了其在体内的清除。因此本部分从分子机制上表明吡柔比星制备成脂质体不仅可以降低心脏毒性,同时还很难产生蓄积毒性。
Pirarubicin (THP), a commonly used anthracycline, has gained widespread applications against acute leukemia, malignant lymphoma, breast cancer, gastrointestinal cancer, liver cancer, lung cancer and some solid tumors. However, there are several adverse effects in the clinics:1, bone marrow suppression and severe gastrointestinal reactions; 2, cumulative cardiotoxicity:THP has a progressive dose-dependent cardiotoxicity that irreversible evolves toward congestive heart failure. Liposome drug delivery system, which can alter the in vivo behavior of the encapsulated drug, was expoited to provide a strategy for limiting or preventing the cardiotoxicity of THP.
Interaction between pirarubicin and distearylphosphatidylcholine (DSPC) or distearylphophatidylglycerol (DSPG) was investigated by means of differential scanning calorimetry (DSC), fourier transform infrared (FTIR) spectroscopy and quantum calculation based on molecular modeling. DSC and FTIR experiments indicated that THP could be more easily incorporated into the DSPC bilayers, whereas the-NH3+ of THP might have much stronger interactions with the PO2- in the polar head of DSPG. The quantum calculation part is to investigate the interaction between-NH3+ of THP and PO2- of phospholipids from the molecular dynamics perspective. The Eint value of the interaction between-NH3+ of THP and PO2- of phospholipids can not only reflect the thermodynamic stability, but also probe that the implying mechanism is closely related to the unique structure of the polar heads. The strong interaction between THP and the polar head of DSPG could prevent or inhibit the further penetration of THP into its bilayers; therefore, the bilayer mobility could not be easily changed.
A novel pirarubicin liposome powder (L-THP), comprising DSPC, DSPG, cholesterol and lactose, was appropriately prepared based on the unique physico-chemical properties of THP. The new preparation method has huge market prospective for its simple operation and good repeatability in the production. Meanwhile, the mechanism was well elucidated. Furthermore, the preparing process and formulations of L-THP were optimized.
In vitro and in vivo assay of pirarubicin were established and the corresponding analytical method were validated. Pharmacokinetic studies showed that the accumulation of THP in the heart could be greatly reduced in our L-THP. Meanwhile, it tended to accumulate in the spleen and lung. Meanwhile, there is a quick clearation of THP in the L-THP. Although the bioavability of THP in the plasma was relatively a bit lower than that of marketed pirarubicin injection, the overall bioavability could be significantly improved.
Safety mechanism of L-THP was investigated through the studies of enzyme activity, pathological and metabonomics. Results of weight, daily behavior, enzyme activity and pathological studies showed that the myocardial damage degree of THP could be greatly reduced when encapsulated into the liposomes. Metabonomics study of urine samples suggested that there were potential biomarkers closely related with the cardiotoxicity of pirarubicin:pirarubicin can generate ROS as well as the aglycones, thus inducing the acute cardiotoxicity; C-13 alcohol metabolites through the two-electron reduction may gain chronic cardiotoxicity. It further suggested that L-THP would have less cumulative cardiotoxicity since the content of biomarkers was low. This phenomenon should be attributed to the consequence of low accumution and high clearation rate of THP.
引文
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