【作者】 解亦斌；

【导师】 赵平； 马洁； 王成锋；

【作者基本信息】 中国协和医科大学 ， 肿瘤学， 2009， 博士

【摘要】 胰腺癌在最初诊断时即有约50%的患者出现肝转移,另外行根治性切除术后的患者约70%最先出现转移的部位包括肝脏,而目前对肝转移瘤缺乏有效治疗方法,这是胰腺癌治疗中面临的最大困难之一。当今临床医疗中对胰腺癌肝转移瘤最常采用的治疗方法是介入治疗,其所用药物借鉴胰腺癌全身化疗或肝癌介入化疗,包括吉西他滨、顺铂、5-Fu以及阿霉素等多种药物。但是所有这些药物都面临毒副作用大,患者耐受性差,用药剂量很难达到最佳治疗剂量,而且最重要的是有效率较低。胰腺癌由于缺乏大型动物细胞系,无法进行筛选最佳用药方案的介入化疗实验研究。脂质体阿霉素由于脂质体载药的原因改变了阿霉素的药代动力学特征及组织分布规律,提高疗效的同时降低了心脏及全身各系统的毒副作用。本课题采用裸鼠胰腺癌肝转移模型,首次通过脾脏注射给药模拟介入化疗来比较脂质体阿霉素与阿霉素相比用于胰腺癌肝转移介入化疗的优势,以筛选胰腺癌肝转移介入化疗新的药物剂型。本课题采用薄层水化法和主动载药法制备PEG-DSPE(polyethylene glycol-distearoylphasphatidyle-thano-lamine)修饰的脂质体阿霉素,经检测所制备的脂质体阿霉素粒径在110±10nm范围内,阿霉素药物包封率为94.21%,长期放置后再次检测性质稳定,适合用于动物实验研究。采用脾脏注射给药模拟介入化疗为本实验首次采用,其可行性原因有以下三点:(1)解剖学上脾脏内血液几乎全部首先回流肝脏,这点与肝动脉或门静脉给药类似。(2)染料法观察显示脾脏下极注射美兰溶液的同时肝脏左叶部分区域出现蓝染,继续给药蓝染区域扩散至全肝,说明肝左叶首先出现蓝染的区域为脾脏下极注射给药时药物首先到达的区域。(3)在门静脉高压模型的实验性治疗中有采用脾脏注射给药的报道。在采用脾脏注射给药模拟介入化疗和尾静脉注射给药模拟全身化疗的治疗实验中脾脏注射给药组疗效优于尾静脉给药组,进一步证实此给药途径虽不能替代肝动脉给药但可以模拟之。采用上下增减剂量法(up and down procedure,UPD)测得脂质体阿霉素和阿霉素经脾脏给药对裸鼠的LD50分别为30.4mg/Kg和17.65mg/Kg,说明脂质体阿霉素可以降低药物的毒副作用,并提高药物的安全剂量。通过药物组织分布实验发现脂质体阿霉素经脾脏注射给药与尾静脉注射给药相比降低了动物心脏、肺脏及肾脏内药物浓度,提高了肝脏及脾脏内药物浓度,同时血液内药物峰浓度降低,曲线下面积减小,从药代动力学角度解释了脾脏注射给药模拟介入化疗疗效更佳的原因。由于本实验采用的模型每只动物需要进行两次手术,费时较长,一次实验所能进行的动物只数有限,药效学实验中以剂量为3mg/kg和6mg/kg阿霉素做对照组分别进行了实验,结果显示脂质体阿霉素1.5mg/kg,3mg/kg和6mg/kg三个剂量疗效均有显著差异,呈剂量依赖性。3mg/kg脂质体阿霉素与相同剂量阿霉素相比肿瘤缩小更明显(P＜0.05),与相同剂量吉西他滨加顺铂无统计学差异(P＞0.05),脂质体阿霉素6mg/kg和9mg/kg疗效均较好,无显著差异。6mg/kg脂质体阿霉素与相同剂量阿霉素相比疗效无显著性差异,与相同剂量吉西他滨加顺铂相比肿瘤显著缩小(P＜0.05)。脂质体阿霉素各剂量组均未出现明显毒性反应,阿霉素组(6mg/kg)和吉西他滨加顺铂组(6mg/kg)出现明显毒性反应,均有约半数动物死亡。药效学实验进一步说明脂质体阿霉素用于介入化疗与阿霉素相比疗效显著提高,毒性显著降低,且6mg/kg脂质体阿霉素为最佳实验治疗剂量。上述实验资料表明,本实验所制备的PEG-DSPE修饰的脂质体阿霉素理化性质均一、稳定,应用于介入化疗能够取得较阿霉素更好的疗效,同时降低了阿霉素的毒副作用,有望成为新的介入化疗药物剂型。更多还原

【Abstract】 Nearly 50%of pancreatic cancer patients suffer liver metastasis at the time of diagnosis,and the first site of recurrence for more than 70%of radically resected patients includes liver,what makes thing worse is till now there are no effective treatmet for liver metastasis of pancreatic cancer.This is one of the largest difficulties we have to faced with in the treatment of pancreatic cancer.Now the mostly used method for liver metastasis of pancreatic cancer is interventional therapy,and the adopted drugs include gemcitabine,cisplatin, 5-fluorouracil and doxorubicin according to the systemic chemotherapy of pancreatic cancer or interventional chemotheary of primary liver cancer.However all these drugs used now have great toxic side effect,bad tolerance so it’s hard to reach optimal therapeutic dosage,and what’s more the response rate is low.As we know for pancreatic cancer there is no cell line for big animal transplantation,so the experimental study for screening optimal interventional chemotherapy drug schedule is hard to perform.For the reason of liposome,liposomal doxorubicin has different pharmacokinetics characteristic and tissue distribution pattern with doxorubicin,then the therapeutic effect was improved and the toxic side effect of heart and other system was decreased.The nude mouse pancreatic cancer liver metastasis model was adopted in this study,and for the first time injection through spleen imitating interventional chemotherapy was introduced in animal experiments to compare the therapeutic effect and toxic side effect of liposomal doxorubicin and doxorubicin,eventually find a new drug for the interventional therapy of liver metastasis of pancreatic cancer.PEG-DSPE（polyethylene glycol-distearoylphasphatidyle-thano-lamine） modified liposomal doxorubicin was prepared by lamellar hydration and initiative loading methods in our laboratory in this study.The diameter of the prepared liposomal doxorubicin was tested in the range of 110±10nm,and the entrapment rate was 94.21%.Lay up it for 3months and test again,we found it’s stable,so it’s suitable for animal experiment. It’s the first time that injection through spleen was adopted to imitate interventional chemotherapy,the feasibility can be explained from the 3 points mentioned below:（1）As to anatomy,the blood in spleen refluxs almost totally to liver, this is similar to injection through hepatic artery or portal vein.（2） When dyes was injected into spleen,you can see the left part of liver turned blue at the same time,and as injection going on,whole liver turned blue,this phenomenon told us this channel of administration is nearly unhindered.（3） As reported in the portal hypertension models, injection through spleen is an administration route.In the therapeutic experiments the group of injection through spleen showed better outcome than the group of injection through caudal vein,so it’s further confirmed that this administration route could imitate interventional chemotherapy.The LD₅₀ of liposomal doxorubicin and doxorubicin were 30.4mg/Kg and 17.65mg/Kg respectively examined by up and down procedure,so liposomal doxorubicin had lower toxic side effect and higher safty dose.In the tissue distribution experiment we found that the drug concentrations in heart,lung and kidney were lower when administrated through spleen injection compared to caudal vein,while the drug concentrations in liver and spleen were higher.In blood the peak concentration was lower and area under the curve was smaller for spleen injection group compared to caudal vein injection group.These results explained why the therapeutic effect for interventional chemotherapy was better from the point view of pharmacokinetics.For the model adopted in this study was time-consuming,the animal we can perform in one time of experiment was limited.So we complished 2 experiments about therapeutic effect of the drug accoding to the different dose of doxorubicin （3mg/kg and 6mg/kg） in control group.The results showed in the doses of 1.5mg/kg, 3mg/kg and 6mg/kg liposomal doxorubicin produced significantly different effect, that mean the effect was in a manner of dose dependent.In the doses of 6mg/kg and 9mg/kg the effect of liposomal doxorubicin turned to be same perfect,and there was no difference between them（P＞0.05）.In the dose level of 3mg/kg liposomal doxorubicin had a better effect than doxorubicin（P＜0.05）,while it’s no better than gemcitabine combined with cisplatin（P＞0.05）.In the dose level of 6mg/kg liposomal doxorubicin and doxorubicin had no significantly different effect,but it’s much better than gemcitabine combined with cisplatin（P＜0.05）.There was no obvious toxic side effect in all the four doses of liposomal doxorubicin,while doxorubicin（6mg/kg） and gemcitabine combined with cisplatin（6mg/kg） led about half of the animal dead in each group.The results of this part further confirmed liposomal doxorubicin had a significantly better effect and lower toxic side effect than doxorubicin,and 6mg/kg was the optimal dose in animal experiments.The results of above experiments showed the PEG-DSPE modified liposomal doxorubicin prepared in this laboratory was uniform and stable,when applied in interventional chemotherapy it could get a better therapeutic effect and lower toxic side effect than doxorubicin,so it’s a potential new pharmaceutical dosage form in interventional chemotherapy.更多还原