摘要
胰腺有内分泌和外分泌两种功能，也就有内分泌和外分泌两种细胞。这两种细胞都会发生癌变，来源自内分泌细胞的癌，叫神经内分泌癌(neuroendocrine tumor, NET)，比较少见，多数情况下恶性程度比较低，病程比较长，治疗方式与常见的胰腺癌也有所不同。来自外分泌细胞的癌，就是我们常说的胰腺癌(pancreatic cancer, PC)，是一种恶性程度比较高的肿瘤。至今已是世界第8位肿瘤致死因，欧美癌症致死的第4位死因，是恶性程度最高的肿瘤之一，其死亡率接近发病率。胰腺癌是一个高度恶性的疾病，约90%的病人无法以手术根除治疗。因此，胰腺癌的药物治疗在胰腺癌治疗中占有非常重要的地位。
传统的化学治疗对于胰腺癌的控制并无太大的帮助。过去常用的药物5-氟尿嘧啶(Fluorouracil,5-FU)，但其反应率很少超过25%。目前，临床上治疗的主要手段仍然是在术后予以全身辅助的化学治疗，但目前用于PC的首选化疗药物吉西他滨(gemcitabine, GEM)也无突出新的显效率，且剂型单一，只有普通注射粉针剂（建议剂量是1000mg/m2），存在毒副作用大、靶向性差、用药顺应性差等缺点。
随着现代医学的发展，先进的医疗仪器和医疗诊断技术也在不断提高。超声内镜检查术(endoscopic ultrasonography, EUS)是一种在超声引导下的细针穿刺注射技术，能精准的找到癌组织，并利用内镜和超声相结合，将穿刺针有效的定位至胰腺癌新生血管等最易于药物发挥作用的部位，这为胰腺癌的瘤内注射治疗提供了有利的条件支持。
可注射性水凝胶因其独特的性质有望被广泛应用于药物释放等领域，用作多肽、蛋白质药物等生物活性物质的释放的载体。天然高分子可注射性水凝胶来源广泛，价廉易得；但由于材料来源不同，材料批次间差异明显，性能重复性差，结构与性能可调范围窄，且力学性能难以满足不同条件的医学需求。合成高分子可注射性水凝胶结构和性能可精细调控，重复性好，易于加工和工业化生产，尤其是降解性可控、具有良好生物相容性的聚酯/聚乙二醇聚合物水凝胶，颇具应用潜力。
本文采用聚乙二醇/聚酯嵌段共聚物PLGA-PEG-PLGA为材料，利用该材料的温敏成胶性质，即常温下成液体，遇生理温度快速变成凝胶的特性，于肿瘤局部注射，以期达到延长作用时间、提高药物肿瘤内浓度，同时使药物浓集在肿瘤部位并缓慢释药、达到安全有效的治疗目的。以吉西他滨(GEM)为模型药物，温度敏感型PLGA-PEG-PLGA聚合物为递药系统制备吉西他滨原位凝胶注射剂，并对其性质及体内外生物性能进行了考察。主要从以下几部分进行研究：
第一部分，以丙交脂(D, L-lactide, LA)，乙交酯(glycolide, GA)，PEG为原料，二乙基己酸亚锡为催化剂，在不同原料配比条件下，采用开环聚合的方式合成了三种不同比例的PLGA-PEG-PLGA聚合物，平均产率都在85%以上。对其结构、形态、分子量及多分散性、粒径、温敏特性、粘度-温度变化进行了考察。1H-NMR及傅立叶红外光谱表明其结构稳定，合成产物为目标产物。凝胶渗透色谱表明聚合物的分子量分布范围窄，无残留物质干扰，纯度较高。不同LA/GA比例的聚合物平均粒径分别为44.10±1.2nm、54.32±3.7nm、66.57±4.7nm，粒径分布相对均匀。采用试管倒置法测试相转变温度，结果表明所合成的嵌段聚合物在一定浓度时生理温度下均能产生凝胶，且溶液-凝胶转变温度Tsol-gel随LA/GA比例的增加而降低，同时也随聚合物浓度的增大而降低。温度粘度曲线也证实了聚合物从溶液向凝胶，从凝胶再向沉淀转变的粘度变化符合该状态下的粘度变化性质。
第二部分，对温度敏感性PLGA-PEG-PLGA聚合物拟作为医用材料的生物相容性和降解性进行了研究。在磷酸盐缓冲溶液（PBS）中考察了PLGA-PEG-PLGA聚合物的体外降解状态和性质变化，体外溶血的情况，体外细胞毒性；在裸鼠体内考察了PLGA-PEG-PLGA聚合物体内生物相容性。体外降解实验中，聚合物的体外降解时间接近一个月，通过GPC检测发现，聚合物的相对平均分子量逐渐降低，分子量分布逐渐变大。溶血实验表明，与对照组相比，不同量的聚合物的加入在观察的时间范围内无明显红细胞破裂或聚集的现象。采用CCK-8法测定细胞活性，聚合物对细胞增殖无明显影响，细胞活性在95%以上。裸鼠皮下注射的可立即形成凝胶，并保持其完整的形态超过3周，随着时间的延长凝胶体及逐渐变小直至消失，注射部位的肌肉组织没有明显的炎症反应。
第三部分，制备PLGA-PEG-PLGA吉西他滨原位凝胶，对制剂的表征、相转变温度及粘度性质进行了考察，建立了吉西他滨原位凝胶注射剂的含量测定方法。利用高效液相色谱法在20-500ug/mL范围内，线性良好，y=36387x+1.1E+5(R2=0.9996)，方法的精密度，准确度符合要求。考察了吉西他滨原位凝胶注射剂的体外释放情况，该制剂可缓慢释放近20天。
第四部分，通过胰腺癌裸鼠移植瘤模型的建立，考察了吉西他滨原位凝胶注射剂的抗胰腺癌效果及体内安全性。考察了凝胶制剂致胰腺细胞凋亡作用，参比各种治疗因子，考察并评价了吉西他滨原位凝胶注射剂在体内的非特异性毒性情况以及对胰腺癌的治疗情况及生存率。裸鼠的血常规检测结果表明，吉西他滨原位凝胶注射剂毒性较小，与GEM市售注射制剂相比，PLGA-PEG-PLGA聚合物对GEM的包裹可以有效地减少与正常组织细胞对GEM的非特异性摄取，减小了GEM在体内的非特异性毒性。在裸鼠的体内治疗试验中，吉西他滨原位凝胶注射剂对胰腺癌具有明确疗效，对肿瘤的生长产生了明显抑制作用。
第五部分，建立了吉西他滨LC-MS/MS的分析方法，吉西他滨在1.5~15000ng/mL的范围内，线性关系良好。该方法的准确度，日内及日间精密度良好，均符合药典标准。吉西他滨的冻融稳定性和24小时稳定性良好。经过预处理的血浆和肝脏组织采用LC-MS/MS对其中吉西他滨的含量进行了测定，结果表明，吉西他滨凝胶给药组比普通吉西他滨注射剂在体内的滞留时间更长，减少了给药次数，存在一定的缓释效果。
综上所述，本课题成功合成了PLGA-PEG-PLGA聚合物，优化了合成条件，对聚合物的理化性质进行了表征，该聚合物在特定LA/GA比例和浓度时具有温度敏感的性质；体内外实验均表明，该聚合物在生理环境下可降解，具备良好的生物相容性。以温度敏感性PLGA-PEG-PLGA聚合物为基础，吉西他滨为模型药物，制备了吉西他滨原位凝胶注射剂，对制剂进行了理化性质表征考察，结果表明该制剂同样具有温度敏感的性质，同时建立了吉西他滨含量测定的方法，考察了制剂的体外释放情况。吉西他滨原位凝胶注射剂体内外实验表明，该制剂具有良好的肿瘤抑制活性，毒副作用较低，是一种安全有效的抗胰腺癌肿瘤制剂。通过研究可为胰腺癌和其他肿瘤化疗和生物治疗提供研究方法和思路，也可为今后靶向药物控释系统设计提供试验资料和理论依据。
Pancreatic cancer (PC) is a malignant neoplasm originating from transformed cellsarising in tissues forming the pancreas. The most common type of pancreatic cancer,accounting for95%of these tumors, is adenocarcinoma (tumors exhibiting glandulararchitecture on light microscopy) arising within the exocrine component of the pancreas.Minorities arise from islet cells, and are classified as neuroendocrine tumors(NET). Usually, it is not easier to find the NETs rather than the PCs because of quitedifferent diagnosis and treatment. The PC is the fourth most common cause ofcancer-related deaths in the United States and the eighth worldwide.90%of the patientscannot be eradicated in the form of surgery. Nowadays, chemotherapy drugs still play animportant role in curing pancreatic cancer in this situation.
It was not quite helpful to control the pancreatic cancer with traditional chemotherapy.The5-fluorouracil (5-FU) was used with only less than25%effectiveness. Today, surgeryand adjuvant chemotherapy are the main treatment for pancreatic cancer in clinic.Gemcitabine (GEM) was the first choice for adjuvant chemotherapy after adequate surgeryat the moment, but there was also no interesting efficiency. The current clinical treatmentwith pancreatic cancer using GEM is mainly by systemic power injection (1000mg/m2recommended). The main advantages are significant toxicity, poor targeting and lack ofcompliance.
With the development of modern medicine, the diagnosis technique and medicalinstruments will be more and more advanced and impressive. The endoscopicultrasonography (EUS), for example, is a kind of fine needle puncture injection technologywith the guidance of ultrasound. The tumor tissue can be found easily and precisely withthis kind of technology. Also, the puncture needle can be localized effectively to the site ofpancreatic cancer neovascularization. It is the foundation for intra-tumor injection therapyof pancreatic cancer.
In situ injectable hydrogel formation makes it more feasible to apply hydrogels fordrug delivery systems. It can be the vehicle of bio-activators such as peptides and protein.There are numerous of sources of injectable hydrogels made of natural polymers. And theyare easy to get. It cannot be satisfied with the medical requirement because of differentkinds of material sources, lack of repeatability and other reasons. The structure andproperty of injectable hydrogels made of synthesis polymers can be controlled nicely andproduced easily. One of them is the polyester/poly (ethylene glycol) copolymers, which arebiodegradable and biocompatible.
The drug can be mixed with a polymer solution in vitro and the drug-loaded hydrogelscan form in situ after the in vivo administration immediately. Therefore, thermo-sensitiveblock copolymer hydrogels have many advantages, such as simple drug formulation andadministration procedures, site-specificity, a sustained drug release behavior, less systemictoxicity and ability to deliver both hydrophilic and hydrophobic drugs. In this study,gemcitabine is selected as the model drug for the intratumoral sustained release deliverysystem. There are four main sections included in the thesis as follow:
At the beginning, a series of copolymers PLGA-PEG-PLGA with different LA/GAratios were synthesized and purified through ting-opening polymerization of d, l-lactideand glycolide with the polyethylene glycol as the initiator in the presence of stannousoctoate. The average productivity was over85%. The chemical structure and compositionof the copolymers were characterized using1H-NMR and FI-IR spectrometry. Themolecular weight and molecular weight distribution of copolymers were determined by gelpermeation chromatography (GPC). The mean particle size were about44.10±1.2nm、54.32±3.7nm、66.57±4.7nm with different LA/GA ratios respectively. Typical phasediagram of PLGA-PEG-PLGA triblock copolymers at various concentrations from%(w/w)to%(w/w) showed that the sol-gel transition temperature was found to be a function ofboth the concentration and composition of the copolymers which was determined bytest-tube inverting method in study. The transition temperature was near the physiologicaltemperature. When the copolymer concentration increased, sol-gel transition temperature decreased and gel-sol transition temperature increased for the copolymers. There is anobvious change of viscosity during the increased temperature. The copolymer solution canbe administrated to tumors through the fine needles.
Secondly, the biodegradability and biocompatibility of thermosensitive triblockPLGA-PEG-PLGA copolymer were studied. This copolymer could be still found afternearly a month although the average molecular weight of triblock copolymer wasdecreased and the molecular weight distribution was increased step by step. It was going tobe slower with LA/GA ratio increasing. Compared with the control group, there was noobvious ruptured and gathered the red blood cell after adding triblock copolymers havingdifferent LA/GA ratios during the observation time. It was no significant effect for cellproliferation after adding different concentrations and LA/GA ratios of triblockcopolymers using CCK-8method to test cell viability. In situ gels could be formedimmediately after subcutaneous injection to nude mice. The whole hydrogels could befound after3weeks even though much smaller or disappeared without any inflammatoryreaction in muscle tissues.
Thirdly, the gemcitabine in situ gel injection was formulated with the solventevaporation method. The characterization of this formulation was investigated includingstructure, molecular weight, particle size, transition temperature and so on. The method todetect gemcitabine in hydrogels was established and it has better sensitivity and specificity.The results of in vitro study showed that the release of gemcitabine from hydrogel lastnearly20days.
Fourthly, the effect of antitumor and security in vivo of gemcitabine in situ gelinjection were studied through pancreas cancer xenograft model. Compared with varioustherapeutic agents, we investigated and evaluated the nonspecific toxicity in vivo andantitumor effect of gemcitabine in situ gel injection on pancreatic cancer. The nude routineblood test, liver and kidney function test results indicate that the toxicity of gemcitabine insitu gel injection is much lower than gemcitabine injection. Gemcitabine embedding inPLGA-PEG-PLGA can reduce the nonspecific intake of gemcitabine in normal tissue effectively. The results of in vivo treatment on nude demonstrate that gemcitabine in situgel injection has a much better therapeutic efficacy to inhibit tumor growth compared withgemcitabine injection and other controls.
Fifthly，the LC-MS/MS analysis method of gemcitabine was established and in therange of1.5~15000ng/mL, there was a good linear relationship. And the accurate andprecision was according with pharmacopoeia standards. The freeze-thaw stability and24-hour stability of gemcitabine were good. After pretreatment of plasma and liver tissue,the content of gemcitabine were determined by LC-MS/MS. The results showed that nanogel group was longer than normal gemcitabine injection in vivo. This kind ofadministration would decrease the injecting times and it may had controlled release ability.
In a word，the method of ring-opening polymerization for triblock copolymerPLGA-PEG-PLGA with different LA/GA ratio was described and the process wasoptimized．The characterization of triblock copolymer was investigated and this copolymersolution was thermosensitive at some LA/GA ratios and some concentrations. This triblockcopolymer could be biodegradable and biocompatible both in vivo and in vitro researches.It was simply to prepare gemcitabine in situ gel injection and the formulation was alsothermosensitive. The detection method of gemcitabine was established at the same time.The results of in vivo and in vitro research showed that gemcitabine in situ gel injection isa safe and effective novel antitumor formulation, which has high efficiency and significantantitumor activity as well as lower side-effect. This study could provide a new researchroute for the therapy of pancreatic cancer as well as the chemotherapy and targetingimmunotherapy for other cancers.
引文
[1] Parkin DM, Bray F, Ferlay J, et al. Global cancer statistics,2002[J]. CA Cancer J Clin.,2005,55(2):74.
[2] American Cancer Society. Facts&Figures2008. Atlanta: GA, American Cancer Society,2008.Ferlay J, Autier P, Boniol M, et al. Estimates of the cancer incidence and mortality in Europe in2006[J].Ann Oncol,2007,18:581.
[3] Cascium S, Craziano F, Catalana G. Chemotherapy for advanced pancreatic cancer: It may no longerbe ignored [J]. Ann Oncol,1999,10(1):105.
[4]孙燕.50年来我国抗肿瘤药物临床研究的进展[J].中国新药杂志,2009,18(18):1695-1700.
[5] Saif MW. New developments in the treatment of pancreatic cancer. Highlights from the “44thASCOAnnual Meeting”. Chicago, IL, USA. May30-June3,2008. JOP2008,9:391-397.
[6] Tanaka M, Javle M, Dong X, et al. Gemcitabine metabolic and transporter gene polymorphisms areassociated with drug toxicity and efficacy in patients locally pancreatic cancer [J]. Cancer,2010,116(22):5325-5335.
[7] Vervenne W, Bennouna J, Humblet Y, et al. Arandomized, doubleblind, placebo controlledmulticenter phase III trial to evaluate the efficacy and safety of adding bevacizumab to erlotinib andgemcitabine in patients with metastatic pancreatic cancer[J]. J Clin Oncol2008;26(supplement):4507.
[8] Howell SB. Clinical applications of a novel sustained-release injectable drug delivery system:DepoFoamTMtechnology [J]. Cancer J2001,7(3):219-227.
[9] Mccarron PA, et al. Sustained release of5-fluorouracil from polymeric nanoparticles[J]. J PhamPhamacol2000;52(12):1451-1459.
[10] Poorter RL, et al. Continuous infusion of chemotherapy: focus on5-fluorouracil andfluorodeoxyuridine[J]. Pharm World&Sci.,1998,20(2):45-49.
[11] Bibby DC, et al. Mechanisms by which cyclodextrins modify drug release from polymeric drugdelivery systems [J]. Int. J Pharm2000;197(1-2):1-11.
[12] Rosemurgy AS, et al. New directions in systemic therapy of pancreatic cancer[J]. Cancer Control2000;7(5):437-451.
[13] Gutowska A, Jeong B, Jasionowski M, et al. Injectable gels for tissue engineering [J]. Anat.Rec.2001,263,342-349.
[14]朱文,段世锋,丁建东.组织工程用水凝胶材料[J].功能高分子学报,2004,17,689-697.
[15] Hennink WE, Van Nostrum CF. Novel crosslinking methods to design hydrogels [J]. Adv. Drug.Deliv. Rev.2002,54,13-36.
[16] Hoffman AS. Hydrogels for biomedical applications [J]. Adv. Drug. Deliv. Rev.2002,54,3-12.
[17] sangeetha NM, Maitra U. Supermolecular gels: Functions and uses [J]. Chem. Soc. Rev.2005,34,821-836.
[18] Vinogradov SV, Bronich TK, Kabanov AV. Nanosized cationic hydrogels for drug delivery:preprafation, properties and interactions with cells[J].. Adv. Drug. Deliv. Rev,2002,54,135-147.
[19] Kim YJ, Kim SW. Controlled drug delivery from injectable biodegradable triblock copolymer[M].In Polymer Gels: Fundamentals And Applications, ed.; Amer. Chemical Soc.: Washington,2003; v01.833,300-311.
[20] Jeong B, Bae YH. Biodegradable block copolymers as injectable drug-delivery systems [J]. Nature1997,388,860-862.
[21] Zhang G, Wang DY, Gu ZZ, Hanmann J, et al. Two-dimensional non-close-packing arrays derivedfrom self-assembly of biomineralized hydrogel spheres and their patterning applications [J]. Chem.Mater.2005,17,5268-5274.
[22] Haraguchi k, Taniguchi S, Takehisa T. Reversible force generation in a temperature-responsivenanocomposite hydrogel consisting of poly(N-isopropylacrylamide) and clay [J]. Chem Phys Chem2005,6,238-241.
[23] Ji SC, Ding JD. A macroscopic helix formation induced by the shrinking of a cylindrical polymerichydrogel [J]. Polym. J.2001,33,701-703.
[24] Bae SJ, Suh JM, SohlI YS, Bac YH, et al. Thermogelling poly(caprolactone-b-ethyleneglycol-b-caprolactone) aqueous solutions [J]. Maromolecules2005,38,5260-5265.
[25] Hwang MJ, Suh JM, Bae YH, et al. Caprolactonic poloxamer analog: PEG-PCL-PEG[J].Biomacromolecules2005,6,885-890.
[26] Choi HS, Ymamoto K, Ooya T, Yui N. Synthesis of poly(epsilon-lysine)-grafted dextrans and theirpH and thermosensitive hydrogelation with cyclodextrins [J]. Chem Phys Chem2005,6,1081-1086.
[27] Jeong B, Bae YH, Lee DS, Kim SW. Biodegradable block copolymers as injectable drug-deliverysystems [J]. Nature1997,388,860-862.
[28] Jeong B, Bae YH, Kim SW. Thermoreversible gelation of PEG-PLGA-PEG triblock copolymeraqueous solution [J]. Macromolecules1999,32,7064-7069.
[29] Chenite A, Chaput C, Wang D. Novel injectable neutral solution of chitosan from biodegradablegels in situ [J]. Biomaterials2000,21,2155-2161.
[30] Wang J, Sun DDN. Stimuli-responsive hydrogel based on poly(propylene phasphate)[J].Macromolecules2004,37,670-672.
[31] Dagani R. Intelligent Gels [J]. Chem. Eng. News,1997,75(23):26-37.
[l]郑俊民．药用高分子材料学[M].北京:中国医药科技出版社,2000,1-2.
[2] Churchill JR, Hutchins FG. Biodegradable amphipathic copolymers[P]. US patent4,745,160,1988-05-17.
[3]王晶，周庆颂，袁悦，莫风奎.生物降解聚合物PLGA-PEG-PLGA的合成和表征[J].沈阳药科大学学报,2005,22(5):348-351
[4] Zentner GM, Rathi R, Shih C, et a1. Biodegradable block copolymers for drug delivery of proteinsand water-insoluble drugs[J]. J. Contro1Release,72(1-3)(2001)203-215.
[5] Chen SB, Pieper R, Webster DC, Singh J. Triblock copolymers: Synthesis, characterization, anddelivery of a model protein[J]. Int. J Pharm.,288(2005):207-218.
[6] Evertti DE, Avom J. Systemic effects of medications used to treat glaucoma[J]. Ann Intern Med，1990,112-120.
[7] Urtti A, Salminen L. Minimizing systemic absorption of topically administered ophthalmlic drugs[J].Surv Ophthalmol,1993,37:435-456.
[8] Qiao MX, Chen DW, Ma XC, Liu YJ, Injectable biodegradable temperature-responsivePLGA-PEG-PLGA copolymers: Synthesis and effect of copolymer composition on the drug releasefrom the copolymer-based hydrogel[J]. Int. J. Pharm.,294(2005)103-115.
[9] Beletsi A, Leontiadis L, Klepetsanis P, et a1.Effect of preparative variables on the properties of poly(d,1-lactide-co-glycolide)-methoxypoly(ethyleneglyco1)copolymers related to their application incontrolled drug delivery[J]. Int. J. Pharm.,82(2)(1999),187-197.
[1] Takahashi M, Onishi H, Machida Y. Development of implant tablet for a week-long sustainedrelease[J]. J. Control. Release,2004,100,63-74.
[2] Sun HF, Mei L, Song CX, Cui XM, Wang PY. The in vivo degradation, absorption and excretion ofPCL-based implant[J]. Biomaterials2006,27,1735-1740.
[3] Onishi H, Takahashi M, Machida Y. PLGA implant tablet of ketoprofen: comparison of in vitro andin vivo releases[J]. Biol. Pharm. Bull.,2005,28,2011-2015.
[4] Ma GL, Song CX, Sun HF, Yang J, Leng XG. A biodegradable levonorgestrel-releasing implantmade of PCL/F68compound as tested in rats and dogs[J]. Contraception2006,74,141-147.
[5] International conference on harmonization of technical requirements for the registration ofpharmaceuticals for human use. Residual Solvents, Draft No.11995.
[6] US Pharmacopia[M]. Organic volatile impurities,23rdrev. edition. US pharmacopeia convention,rockville, MD.1995,1746-1748.
[7] Anderson JM, Shive MS. Biodegradation and biocompatibility of PLA and PLGA microspheres[J].Adv. Drug Deliv. Rev.1997,28,5-24.
[8] Tamber H, Johansen P, Merkle HP, Gander B. Formulation aspects of biodegradable polymericmicrospheres for antigen delivery[J]. Adv. Drug Deliv. Rev.2005,57,357-376.
[9] Freiberg S, Zhu X. Polymer microspheres for controlled drug release[J]. Int. J. Pharm.2004,282,1-18.
[10] Freitas S, Merkle HP, Gander B. Microencapsulation by solvent extraction/evaporation: reviewingthe state of the art of microsphere preparation process technology[J]. J. Control. Release2005,102,313-332.
[11] Sawhney, A; Pathak, C.; Hubbell, J, Bioerodible hydrogels based on photopolymerizedpoly(ethylene glycol)-co-poly-(alpha hydroxy acid) diacrylate macromers. Macromolecules1993,26,581-587.
[1] Hidenobu Okino, Trans-tissue, sustained release of gemcitabine from photo cured gelatin gelinhibits the growth of heterotopic human pancreatic tumor in nude mice. Clinical Cancer Research.2003Nov;9:5786-5793.
[2]杨梅，李井泉，汤致强. PLGA吉西他滨缓释微球在荷胰腺癌裸鼠体内的药动学研究.中国药学杂志.2010Feb;45,295-299.
[1] Kyriazis A P, McCombs w B, Sandberg R, et aI. EstabIishment and characterization of humanpancreatic adenocarcinoma cell line Sw1990in tissue cuIture and the nude mice. Cancer Res,1983,4314393.
[2] Kowalczyk A, Gil M, Horwacik I. The GD2-specific14G2a monoclonal antibody induces apoptosisand enhances cytotoxicity of chemotherapeutic drugs in IMR-32human neuroblastoma cells. CancerLetters,2009,281:171–182.
[1]. Sun, Y., et al., Determination of gemcitabine and its metabolite in extracellular fluid of rat braintumor by ultra performance liquid chromatography-tandem mass spectrometry using microdialysissampling after intralesional chemotherapy. J Chromatogr B Analyt Technol Biomed Life Sci,2013.919-920:10-9.
[2]. Hertel, L.W., et al., Evaluation of the antitumor activity of gemcitabine(2',2'-difluoro-2'-deoxycytidine). Cancer Res,1990.50(14):4417-22.
[3]. Shord, S.S., et al., Gemcitabine pharmacokinetics and interaction with paclitaxel in patients withadvanced non-small-cell lung cancer. Cancer Chemother Pharmacol,2003.51(4):328-36.
[4]. Oguri, T., et al., The determinants of sensitivity and acquired resistance to gemcitabine differ innon-small cell lung cancer: a role of ABCC5in gemcitabine sensitivity. Mol Cancer Ther,2006.5(7):1800-6.
[5]. Shipley, L.A., et al., Metabolism and disposition of gemcitabine, and oncolytic deoxycytidineanalog, in mice, rats, and dogs. Drug Metab Dispos,1992.20(6):849-55.
[6].钟皎，赵文艳,反相高效液相色谱法测定人血浆中吉西他滨浓度.中国药师,2009.12(1):17-19.
[1] Moghimi SM, Hunter AC, Murray JC. Nano medicine: current statusand futureprospects[J]. FASEBJ.2005,19:311-330.
[2] Kempf RA, Mitchell MS, Effects of chemotherapeutic Agents on the immune response.I. CancerI nvest,1984,2:459-466．
[3]仇海镇,李娟.原位凝胶的研究进展及其在药剂学中的应用[J].医学信息2010,4(23):4
[4] E. Ruel-Gariepy, M. Shive, et al. A thermosensitive chitosan-based hydrogel for thelocal delivery of paclitaxel,[J] Eur. J. Pharm. Biopharm.,2004,57,53-63.
[5] Na Lei, ChangYang Gong, ZhiYong Qian, et al. Therapeutic application of injectablethermosensitive hydrogel in preventing local breast cancer recurrence and improvingincision wound healing in a mouse model.[J]Nanoscale.2012,4:5686-5693.
[6] Ding-Ding Guo, Seong-Ho Hong, Hu-Lin Jiang, et al, Synergistic effects of Akt1shRNA and paclitaxel-incorporated conjugated linoleic acid-coupled poloxamerthermosensitive hydrogel on breast cancer,[J]. Biomaterials,33(2012),2272-2281.
[7] Seki T，Tamai T，Ikeda K，da1．Rapid progression of hepatocel-hlar carcinoma aftertranseatheter arterial chemoembolization and Percutaneousr adiofrequencya blation in theprimary iunlour region[J]．Eur J C. aaroentWol IIepatol.2001,13(3):291-4．
[8] Qien F, Stowe N, Liu EH, et al. Quantification of in vivo doxombicin transport from Amillim in thennosblated xat livers[J]. Control Release,2003,91(1-2):157-66.
[9] Ahmed M, Ilu Z, hdCyanor AN, et al. Combination radioGequency ablation withintratumoral liposomel doxarubicin; effect on drug accumulation and coagulation inmultiple tissues and tumor types in animals[J]. Radiology,2005,235(2);469-77.
[10] Mok TS, Kanekal S, Lin XR, Pharmacokinetic study of intralesional cisplatin for thetreatment of hepatocellular carcinoma [J].Cancer,2001,91(12);23,69-77.
[11] Harbord M, Dawes RF, Bas H,et al. Palliation ad patients with dyapha a due toadvanced esophageal cancer by endoxopic inje lion‘ciaphUin/epinephrine injectable gel
[J]. Gashointert Endoac.2002.56:644-51.
[12]药晋鹏,戴立里,李娟,等.载阿霉素的液一固相变型原位凝胶注射治疗兔VX2肝癌HIFU消融后残癌[J].中国介入影像与治疗学,2012,9(11):34
[13] Parkin DM, Bray F, Ferlay J, et al. Global cancer statistics,2002[J], CA Cancer J Clin,2005,55(2):74.
[14] Saif MW. New developments in the treatment of pancreatic cancer. Highlights fromthe “44thASCOAnnual Meeting”. Chicago, IL, USA. May30-June3,2008. JOP2008,9:391-397
[15] Strocchi P, Dozza B, Pecorella I, Fresina M, Campos E,Stirpe F. Lesions caused byricin applied to rabbit eyes.[J]Invest Ophthalmol Vis Sci,2005;46:1113-1116.
[16] Ghahremankhani AA, Dorkoosh F, Dinarvand R. PLGA-PEG-PLGA tri-blockcopolymers as in situ gel-forming peptide delivery system: effect of formulation propertieson peptide release.[J] Pharm Dev Technol.2008;13:49-55.
[17] Jeong B, Bae YH, Lee DS, Kim SW. Biodegradable block copolymers as injectabledrug-delivery systems.[J]Nature.1997;388:860-862.
[18] Chen Zhi-Kui, Lin Li-Wu, Weng Xiu-Hua, Xue En-Sheng and Lin Yong-Hua,Interstitial chemotherapy with ricin-loaded thermosensitive hydrogel in pancreatic cancerxenograft,[J] Hepatobiliary Pancreat Dis Int，Vol8，No4. August15,2009
[19]Seruga B,Sterling L,Wang L,et.al.Reporting of serious adverse drug reactions oftargeted anticancer agents in pivotalphase Ⅲ clinical trials[J].J Clin Oncol,2011;9(2):174-185
[20] Yuan Gao, Fuzheng Ren, Baoyue Ding,et al. A thermo-sensitive PLGA-PEG-PLGAhydrogel for sustained release of docetaxel.[J] Journal of Drug Targeting,2010,1-12.
[21] Avudaiappan Maran,Mahrokh Dadsetan,Colleen M. Buenz,Kristen L.et al,Hydrogel-PLGA delivery system prolongs2-methoxyestradiol-mediated anti-tumor effectsin osteosarcoma cells,[J]Journal of biomedcal materials research month2013Vol00A,ISSUE00
[22]朱卫,徐元龙,李学明,智能纳米水凝胶及其药剂学研究新进展[J]，中国新药杂志,2011,20(6)