载药高分子聚合物微泡超声造影剂的实验研究
摘要
癌症是威胁人类健康的重大疾病之一,恶性肿瘤的治疗在很大程度上仍以化疗为主。肿瘤化疗药物大多数为细胞毒性药物,具有较大的毒副作用。靶向给药系统能将药物定向输送到治疗部位,提高治疗部位的药物浓度,减少药物用量,降低不良反应,提高药物的安全性、有效性和患者的顺应性,是目前抗肿瘤治疗领域研究的重点。最新研究表明,超声空化过程中产生的“空化效应”、“声孔效应”能够实现药物的体内定位释放和提高治疗效果。微泡超声造影剂(UCA)作为一种新型的体内靶向给药载体系统受到广泛关注。
本研究采用具有良好生物相容性和生物降解性的乳酸/羟基乙酸共聚物(PLGA)作为成膜材料,通过双乳化法和冷冻干燥技术制备包裹丝裂霉素(MMC)和全氟丙烷(C_3F_8)的高分子聚合物微泡超声造影剂(MMC-PLGA微泡超声造影剂)。运用光学显微镜和扫描电子显微镜(SEM)进行形态学观察,通过体外显影和辐照实验对造影效果和药物定向释放进行研究。研究内容如下:
1、微泡超声造影剂的制备和表征
通过优化实验制备出了粒径在0.5～5μm大小均匀,外观规整圆滑的MMC-PLGA微泡超声造影剂;造影剂冻干粉复溶后分散度好,激光粒度分析仪检测其粒径分布窄,平均粒径2.86μm,且粒径大小与制备过程中匀质机的设置有关;载药微泡最大载药率0.93%,包封率达47.4%。
2、体外显影和辐照实验方面
制备的MMC-PLGA微泡超声造影剂具有良好体外显影效果;在1.0W/cm~2的超声辐照下微泡累积释药量达90.95%,远高于对照组未经任何超声辐照的微泡体外初始释药量31.38%,超声辐照控制药物释放效果明显。
研究结果证明,本研究制备的MMC-PLGA微泡超声造影剂具有良好的显影效果,同时初步实现了药物的控制释放,研究结果为应用超声波和微泡造影剂进行体内药物定位释放研究奠定了基础。
Cancer is a main disease to threat the health of human being. Chemical therapy is an important method to treat it comparing with operation and radial therapy. Most anticancer drugs are cytotoxinic with greater toxicity. Targeted drug delivery system can direct the drug to the treatment site, improve the drug concentration, reduce the drug consumption and adverse reactions, and improve drug safety, efficacy and patient compliance. So currently, it is the focus of anti-tumor therapy research. A recent study showed that "cavitation effects" and "sound-effect" that are generated in the ultrasonic cavitation process can achieve in vivo drug release positioning and improve the therapeutic effect. Ultrasound contrast agent (UCA) as a novel in vivo targeted drug delivery system receives widespread attention.
PLGA, a novel artificial and biodegradable high molecular polymer-polylactic-co-glycolic acid, was employed to fabricate the ultrasound contrast agent with Mytomycin(MMC) and C_3F_8 inside by a double emulsion method and lyophilization method. We used light microscope and scan electron microscope to study the shape and surface morphology of the MMC-PLGA ultrasound contrast agent. The experiment in vitro was employed to study the effects of MMC-PLGA ultrasound contrast agent on the enhancement of ultrasound imaging. The experiment of ultrasound irradiation was employed to study the drug controlled-release. The following are the research results:
1. The preparation and characterization of MMC-PLGA ultrasoundcontrast agent
MMC-PLGA ultrasound contrast agent was produced with a uniform size of 0.5～5μm through optimized experiment. Contrast agent lyophilized powder has a good dispersion when dissolved. Laser particle size analyzer was used to detect the size distribution. The results showed that the average particle size is 2.86μm, and the size of the microspheres has a relationship with the installation of heterogeneous machines in the preparation. The most loading capacity of the drug microbubbles were 3.39%, and the encapsulation rate was 32.2%.
2. In-vitro autoradiography and irradiation experiments
The experiment showed that MMC-PLGA ultrasound contrast agent had a good in-vitro auto -radiography effect. The amount of microbubble drag release could reach 90.95 % with 1.0W/cm~2 ultrasonic irradiation, which was much higher than 31.38%—the amount of microbubbles initial in vitro release—that without any ultrasonic irradiation. Ultrasonic irradiation had an evident effect in controlled drag release.
MMC-PLGA ultrasound contrast agent had a good autoradiography effect, and meanwhile realized the drag controlled release. The study laid a foundation for the use of ultrasonic and ultrasound contrast agent for in vivo drag delivery.
本研究采用具有良好生物相容性和生物降解性的乳酸/羟基乙酸共聚物(PLGA)作为成膜材料,通过双乳化法和冷冻干燥技术制备包裹丝裂霉素(MMC)和全氟丙烷(C_3F_8)的高分子聚合物微泡超声造影剂(MMC-PLGA微泡超声造影剂)。运用光学显微镜和扫描电子显微镜(SEM)进行形态学观察,通过体外显影和辐照实验对造影效果和药物定向释放进行研究。研究内容如下:
1、微泡超声造影剂的制备和表征
通过优化实验制备出了粒径在0.5～5μm大小均匀,外观规整圆滑的MMC-PLGA微泡超声造影剂;造影剂冻干粉复溶后分散度好,激光粒度分析仪检测其粒径分布窄,平均粒径2.86μm,且粒径大小与制备过程中匀质机的设置有关;载药微泡最大载药率0.93%,包封率达47.4%。
2、体外显影和辐照实验方面
制备的MMC-PLGA微泡超声造影剂具有良好体外显影效果;在1.0W/cm~2的超声辐照下微泡累积释药量达90.95%,远高于对照组未经任何超声辐照的微泡体外初始释药量31.38%,超声辐照控制药物释放效果明显。
研究结果证明,本研究制备的MMC-PLGA微泡超声造影剂具有良好的显影效果,同时初步实现了药物的控制释放,研究结果为应用超声波和微泡造影剂进行体内药物定位释放研究奠定了基础。
Cancer is a main disease to threat the health of human being. Chemical therapy is an important method to treat it comparing with operation and radial therapy. Most anticancer drugs are cytotoxinic with greater toxicity. Targeted drug delivery system can direct the drug to the treatment site, improve the drug concentration, reduce the drug consumption and adverse reactions, and improve drug safety, efficacy and patient compliance. So currently, it is the focus of anti-tumor therapy research. A recent study showed that "cavitation effects" and "sound-effect" that are generated in the ultrasonic cavitation process can achieve in vivo drug release positioning and improve the therapeutic effect. Ultrasound contrast agent (UCA) as a novel in vivo targeted drug delivery system receives widespread attention.
PLGA, a novel artificial and biodegradable high molecular polymer-polylactic-co-glycolic acid, was employed to fabricate the ultrasound contrast agent with Mytomycin(MMC) and C_3F_8 inside by a double emulsion method and lyophilization method. We used light microscope and scan electron microscope to study the shape and surface morphology of the MMC-PLGA ultrasound contrast agent. The experiment in vitro was employed to study the effects of MMC-PLGA ultrasound contrast agent on the enhancement of ultrasound imaging. The experiment of ultrasound irradiation was employed to study the drug controlled-release. The following are the research results:
1. The preparation and characterization of MMC-PLGA ultrasoundcontrast agent
MMC-PLGA ultrasound contrast agent was produced with a uniform size of 0.5～5μm through optimized experiment. Contrast agent lyophilized powder has a good dispersion when dissolved. Laser particle size analyzer was used to detect the size distribution. The results showed that the average particle size is 2.86μm, and the size of the microspheres has a relationship with the installation of heterogeneous machines in the preparation. The most loading capacity of the drug microbubbles were 3.39%, and the encapsulation rate was 32.2%.
2. In-vitro autoradiography and irradiation experiments
The experiment showed that MMC-PLGA ultrasound contrast agent had a good in-vitro auto -radiography effect. The amount of microbubble drag release could reach 90.95 % with 1.0W/cm~2 ultrasonic irradiation, which was much higher than 31.38%—the amount of microbubbles initial in vitro release—that without any ultrasonic irradiation. Ultrasonic irradiation had an evident effect in controlled drag release.
MMC-PLGA ultrasound contrast agent had a good autoradiography effect, and meanwhile realized the drag controlled release. The study laid a foundation for the use of ultrasonic and ultrasound contrast agent for in vivo drag delivery.
引文
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[1]冉海涛.新型高分子材料超声造影剂微泡声学造影剂制备及应用研究[D].重庆医科大学硕士学位论文,20050701.
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[2]Andrassy P,Zielinska M,Busch R,et al.Myocardial blood volume and the amount of viable myocardium early after mechanical reperftlsion of acute myocardial infarction:prospective study using venous contrast echocardiography[M].Heart,2002,87(4):350-355.
[3]钱梦騄.超声造影剂及其应用[J].声学技术,2004,23(3):01-02.
[4]Gramiak.Ultrasound cardiography:contrast studies in anatomy and function [J].Radiology,1969,92:929-935.
[5]Ziskin MC,Bonakdapour A,Wienstein DP.et al.Contrast agents for diagnostic ultrasound[J].Invest Radial,1972,7:500-503.
[6]王新房.肺小动脉嵌顿注射双氧水进行心脏声学造影的研究[J].中华医学杂志,1983,63:593-596.
[7]Feinstein SB,Ten Cate FJ,Zwehl W,et al.Two-dimensional contrast echo -cardiography:Ⅰ.In vitro development and quantitative of echo contrast agents[J].JACC,1984,3:14-18.
[8]Kelley MW,Feinstein SB,Watson DD.Successful left ventricular opaci -fication following peripheral venous injection of sonicated contrast agent:An experimental evaluation[J].Am Heart J,1987,114:570-579.
[9]Feinstein SB,Cheirif J,Ten Cate FJ.Safety and efficacy of a new tranpulmonary ultrasound contrast agent:Initial multicenter clinical results [J].JACC,1990,16:316-320.
[10]Barnhart J,Levene H,Villapando E,et al.Characteristics of Albunex:Air-filled albumin microspheres for echocardiography contrast enhancement[J].Invest.Radiol,1990,25(1):S162.
[11]Von bibra H,Becher H,Firschke,et al.Enhancement of mitral regurgitation and normal left atrial color Doppler flow signals with peripheral venousinje ction of a sccharide - based contrast agent[J].JACC,1993,22(2):21-26.
[12]Von bibra H,Sutherl G,Becher H,et al.Clinical evaluation of left heart Doppler contrast enhancement by a saccharide - based transpulmonary contrast agent[J].JACC,1995,25(2):500-503.
[13]Grayburn PA.Peripheral intravenous myocardial contrast echocardiography using a 2%emulsion:identification of myocardial risk are a and infarct size in the canine model ofischemia[J].JACC,1995,26(5):1340-1345.
[14]Skyba DM,Camarano G,Hemodynamic characteristics,myocardial kinetics and microvascular rheology of FS-069,a second -generation from a venous injection[J].Am Coll Cardiol,1996,28(5):1292-1299.
[15]Porter TR.Noninvasive identification of acute myocardial ischemia and reperfusion with contrast ultrasound using intravenous perfluoro propane exposed sonicated dextrose albumin[J].JACC,1995,26:33-36.
[16]Bokor D.Diagnostic efficacy of SonoVue[J].Am J Cardiol,2000,86(4):19-24.
[17]Chen W S,Matula T J,Brayman A A,Crum L A.A comparison of the frag -mentation thresholds and inertial cavitation doses of different ultrasound contrast agents[J].Acoust.Soc.Am,2003,113(1):643-651.
[18]Liu Jie hui,Gong Xiu fen,Zhang Dong et al.Acoustic properties of an ultrasound contrast[J].Journal of NaiJing University,2000,36(1):129-130.
[19]Burns PN.US contrast agents:Basic principle[J].Ultrasound in Med & Biol,2006,32(5):115-119.
[20]崔立刚,张武.超声造影剂基本原理[J].中华医学超声杂志,2006,13(6):372-375.
[21]Richard CW,Kwan OL,Michael H,et al.Initial safety and myocardial erfusion imaging results with AI2700:a newultrasound contrast agent[J].J Am Soc Echocardiogr,2001,14(5):457-459.
[22]赵应征,张彦.微泡超声造影剂的研究进展[J].Foreign Medical Sciences Section on Pharmacy,2003,30(5):298-302.
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