氨基硅烷化Fe_3O_4磁性纳米载体介导野生型p53基因治疗肝癌的实验研究
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摘要
近年来,随着基因转移技术的日趋成熟,肝癌的基因治疗成为生物科学和临床医学的研究热点之一。
基因治疗的载体,以及载体相关的免疫反应、细胞毒性与安全性问题是限制基因治疗发展和临床应用的瓶颈。目前基因治疗的研究和临床应用中常用的病毒载体和非病毒载体都存在无法克服的局限性。纳米粒基因转运载体是近年发展起来的一种新型的非病毒基因转运载体。它是将DNA、RNA等基因治疗分子包裹在纳米颗粒之中或吸附在其表面,同时也在颗粒表面耦联特异性的靶向分子,如特异性配体、单克隆抗体等,通过靶向分子与细胞表面特异性受体结合,在细胞摄取作用下进入胞内,实现安全有效的靶向性基因治疗。
纳米粒材料的选择是成功进行纳米基因转运和治疗的关键。所选择的材料必须具有生物适应性和可降解型的,或者易于从体内排泄,而不产生有害的降解产物,且无免疫原性的物质,不会引起机体的免疫排斥反应。高分子生物降解材料制备的纳米颗粒具有稳定、无毒、无抗原性、生物相容性好、对所转运基因的表达有控释作用及对基因有保护作用等优点,是良好的纳米基因转运载体材料。
研究证实,纳米粒与DNA的连接多通过静电吸引作用来完成。DNA的磷酸骨架所带的负电荷只能与表面带正电性的载体结合。因此,需要利用生物活性分子对纳米颗粒的表面进行改性,使其表面携带阳离子物质,以起到防止纳米颗粒团聚,并有利于结合DNA分子的作用。
本研究中结果显示,以氨基硅烷化Fe_3O_4纳米颗粒作为基因转运载体所形成的氨基硅烷化Fe_3O_4纳米粒/DNA复合物,能保护所携带的DNA免受核酸酶的降解和超声剪切作用。同时,本研究发现,在较低浓度下Fe_3O_4纳米颗粒与氨基硅烷化Fe_3O_4纳米颗粒对肝癌(HepG2)细胞无明显的细胞毒性,只在高浓度下才会表现出一定的细胞毒性作用。在体外基因转运中,该纳米颗粒可有效转运pEGFP-C1报道基因表达质粒进入HepG2细胞,其转运效率为39%,略高于相同条件下脂质体的转染效率,但无统计学差异(*P＞0.05,N=10)。在外加磁场的条件下氨基硅烷化Fe_3O_4磁性纳米粒的转染效率明显增强。
在此基础之上,本研究以氨基硅烷化Fe_3O_4磁性纳米粒作为基因转运载体,以重组人真核细胞表达质粒pcDNA3.1(+)-p53作为治疗基因,对HepG2细胞的荷瘤小鼠模型进行以氨基硅烷化Fe_3O_4磁性纳米粒作为基因转运载体在体内转运重组人真核细胞表达质粒pcDNA3.1(+)-p53的转染,利用RT-PCR、Western blot等方法检测HepG2细胞中转染入的外源野生型p53的表达情况,结果发现,在HepG2细胞中成功导入了外源野生型p53基因,并能表达p53蛋白。
本实验结合局部热疗研究基因治疗与局部热疗的协同效应及对荷瘤小鼠的肿瘤抑制情况,设计了对照组、局部热疗组、氨基硅烷化Fe_3O_4纳米粒作转导pcDNA3.1(+)-p53基因治疗组和联合治疗组4个实验组,结果显示,热疗和野生型p53基因治疗都能抑制荷肝癌裸鼠转移瘤的生长。热疗和野生型p53基因联合应用可达到较好的协同作用,比单一的热疗或基因治疗能更好的抑制肝癌转移瘤的生长。联合治疗组、基因治疗组、热疗治疗组的抑瘤率分别为84.07%,59.43%,47.19%。
Recently,the methods of gene delivery are gradually developing and have made great progresses.So the gene therapy of HCC has been as a hot spot on the biology and clinical medicine field.
The gene delivery vectors and vectors associated immune response, cell toxicity and safety are the bottleneck of research and clinic application of gene therapy.The traditional gene delivery vectors including viral vectors and non-viral vectors have some disadvantages that are hard to overcome.Nanoparticle(NP) as a new non-viral gene delivery vector is recently developed.Gene therapeutical molecules including DNA,RNA,etc,which are encapsulated into or loaded on the surface of nanoparticles.While specific monoclonal antibodies of the target cell are linked on the surface of nanoparticle.The nanoparticles/DNA complexes can be taken into target cells by receptor-mediated endocytosis resulting in an effective target gene therapy.
Selecting proper materials is the key point for nanoparticle-mediated gene delivery and gene therapy.The nanoparticle materials must be very good to biodegradability or easy to be discharged,and to have non-immunogenic.The nanoparticles made by biodegradable polymer have some advantages,such as stability,non-toxicity,good biocompatibility and the loaded DNA delivery can be controlled and protected from degradation.
It is proved that the binding between nanoparticles and DNA was accomplished by electrostatic attraction.The negatively charge of phosphate backbone of DNA could be binded to the positively charged of vector.Therefore,modification of the nanoparticles surface properties with biomolecules makes the nanoparticles surface carring possitve charge which can protect the condensotion of NPs,it is avaliable to the carried gene delivery.
In this study,we used functional Fe_3O_4 nanoparticles modified by APTES as a gene delivery vector,and found that functional Fe_3O_4 nanoparticles -DNA complexes could protect the DNA from degradation of DNaseI and clip of ultrasound.While we found that Fe_3O_4 nanoparticles had no obvious cell toxicity to HepG2 at lower concentration,but showed cell toxicity at high concentration.The pEGEP-C1 expression plasmid could be transfected effectively into HepG2 cells by APTES-modified Fe_3O_4 nanoparticles in vitro.The transfection rate was 39%,which was slightly higher than that of liposome(P＞0.05),but the transfecation rate could be increased by addition of extra-magnetic field.
Using functional Fe_3O_4 NPs as gene delivery vector and eucryotic expression plasmid pEGEP-C1 as therapeutical gene were performed in this study,and the expression of wt-p53 transfected in HepG2 cells was measured by RT-PCR and Western blot.The results showed that transfection of exogenous wild-type p53 gene to HepG2 cells was successful and could be expression of p53 protein.
In this experiment,the nude rats were randomly diveded into 4 groups:control group,local thermotherapy group,amino-functional of Fe_3O_4 nanoparticles for transduction pcDNA3.1(+)-p53 gene(functional gene) therapy group,and combination of thermotherapy and group functional gene therapy.The results showed that thermotherapy and functional gene therapy can inhibit the growth of metastatic tumors in nude mice.However the effect of combined therapy was better than that of the thermotherapy or functional gene therapy alone.The inhibiting rate of the tumor growth in combined therapy group,functional gene therapy group and thermotherapy group was 84.07%,59.43%and 47.19%, respectively.
基因治疗的载体,以及载体相关的免疫反应、细胞毒性与安全性问题是限制基因治疗发展和临床应用的瓶颈。目前基因治疗的研究和临床应用中常用的病毒载体和非病毒载体都存在无法克服的局限性。纳米粒基因转运载体是近年发展起来的一种新型的非病毒基因转运载体。它是将DNA、RNA等基因治疗分子包裹在纳米颗粒之中或吸附在其表面,同时也在颗粒表面耦联特异性的靶向分子,如特异性配体、单克隆抗体等,通过靶向分子与细胞表面特异性受体结合,在细胞摄取作用下进入胞内,实现安全有效的靶向性基因治疗。
纳米粒材料的选择是成功进行纳米基因转运和治疗的关键。所选择的材料必须具有生物适应性和可降解型的,或者易于从体内排泄,而不产生有害的降解产物,且无免疫原性的物质,不会引起机体的免疫排斥反应。高分子生物降解材料制备的纳米颗粒具有稳定、无毒、无抗原性、生物相容性好、对所转运基因的表达有控释作用及对基因有保护作用等优点,是良好的纳米基因转运载体材料。
研究证实,纳米粒与DNA的连接多通过静电吸引作用来完成。DNA的磷酸骨架所带的负电荷只能与表面带正电性的载体结合。因此,需要利用生物活性分子对纳米颗粒的表面进行改性,使其表面携带阳离子物质,以起到防止纳米颗粒团聚,并有利于结合DNA分子的作用。
本研究中结果显示,以氨基硅烷化Fe_3O_4纳米颗粒作为基因转运载体所形成的氨基硅烷化Fe_3O_4纳米粒/DNA复合物,能保护所携带的DNA免受核酸酶的降解和超声剪切作用。同时,本研究发现,在较低浓度下Fe_3O_4纳米颗粒与氨基硅烷化Fe_3O_4纳米颗粒对肝癌(HepG2)细胞无明显的细胞毒性,只在高浓度下才会表现出一定的细胞毒性作用。在体外基因转运中,该纳米颗粒可有效转运pEGFP-C1报道基因表达质粒进入HepG2细胞,其转运效率为39%,略高于相同条件下脂质体的转染效率,但无统计学差异(*P＞0.05,N=10)。在外加磁场的条件下氨基硅烷化Fe_3O_4磁性纳米粒的转染效率明显增强。
在此基础之上,本研究以氨基硅烷化Fe_3O_4磁性纳米粒作为基因转运载体,以重组人真核细胞表达质粒pcDNA3.1(+)-p53作为治疗基因,对HepG2细胞的荷瘤小鼠模型进行以氨基硅烷化Fe_3O_4磁性纳米粒作为基因转运载体在体内转运重组人真核细胞表达质粒pcDNA3.1(+)-p53的转染,利用RT-PCR、Western blot等方法检测HepG2细胞中转染入的外源野生型p53的表达情况,结果发现,在HepG2细胞中成功导入了外源野生型p53基因,并能表达p53蛋白。
本实验结合局部热疗研究基因治疗与局部热疗的协同效应及对荷瘤小鼠的肿瘤抑制情况,设计了对照组、局部热疗组、氨基硅烷化Fe_3O_4纳米粒作转导pcDNA3.1(+)-p53基因治疗组和联合治疗组4个实验组,结果显示,热疗和野生型p53基因治疗都能抑制荷肝癌裸鼠转移瘤的生长。热疗和野生型p53基因联合应用可达到较好的协同作用,比单一的热疗或基因治疗能更好的抑制肝癌转移瘤的生长。联合治疗组、基因治疗组、热疗治疗组的抑瘤率分别为84.07%,59.43%,47.19%。
Recently,the methods of gene delivery are gradually developing and have made great progresses.So the gene therapy of HCC has been as a hot spot on the biology and clinical medicine field.
The gene delivery vectors and vectors associated immune response, cell toxicity and safety are the bottleneck of research and clinic application of gene therapy.The traditional gene delivery vectors including viral vectors and non-viral vectors have some disadvantages that are hard to overcome.Nanoparticle(NP) as a new non-viral gene delivery vector is recently developed.Gene therapeutical molecules including DNA,RNA,etc,which are encapsulated into or loaded on the surface of nanoparticles.While specific monoclonal antibodies of the target cell are linked on the surface of nanoparticle.The nanoparticles/DNA complexes can be taken into target cells by receptor-mediated endocytosis resulting in an effective target gene therapy.
Selecting proper materials is the key point for nanoparticle-mediated gene delivery and gene therapy.The nanoparticle materials must be very good to biodegradability or easy to be discharged,and to have non-immunogenic.The nanoparticles made by biodegradable polymer have some advantages,such as stability,non-toxicity,good biocompatibility and the loaded DNA delivery can be controlled and protected from degradation.
It is proved that the binding between nanoparticles and DNA was accomplished by electrostatic attraction.The negatively charge of phosphate backbone of DNA could be binded to the positively charged of vector.Therefore,modification of the nanoparticles surface properties with biomolecules makes the nanoparticles surface carring possitve charge which can protect the condensotion of NPs,it is avaliable to the carried gene delivery.
In this study,we used functional Fe_3O_4 nanoparticles modified by APTES as a gene delivery vector,and found that functional Fe_3O_4 nanoparticles -DNA complexes could protect the DNA from degradation of DNaseI and clip of ultrasound.While we found that Fe_3O_4 nanoparticles had no obvious cell toxicity to HepG2 at lower concentration,but showed cell toxicity at high concentration.The pEGEP-C1 expression plasmid could be transfected effectively into HepG2 cells by APTES-modified Fe_3O_4 nanoparticles in vitro.The transfection rate was 39%,which was slightly higher than that of liposome(P＞0.05),but the transfecation rate could be increased by addition of extra-magnetic field.
Using functional Fe_3O_4 NPs as gene delivery vector and eucryotic expression plasmid pEGEP-C1 as therapeutical gene were performed in this study,and the expression of wt-p53 transfected in HepG2 cells was measured by RT-PCR and Western blot.The results showed that transfection of exogenous wild-type p53 gene to HepG2 cells was successful and could be expression of p53 protein.
In this experiment,the nude rats were randomly diveded into 4 groups:control group,local thermotherapy group,amino-functional of Fe_3O_4 nanoparticles for transduction pcDNA3.1(+)-p53 gene(functional gene) therapy group,and combination of thermotherapy and group functional gene therapy.The results showed that thermotherapy and functional gene therapy can inhibit the growth of metastatic tumors in nude mice.However the effect of combined therapy was better than that of the thermotherapy or functional gene therapy alone.The inhibiting rate of the tumor growth in combined therapy group,functional gene therapy group and thermotherapy group was 84.07%,59.43%and 47.19%, respectively.
引文
[1] Aznavoorian S , Murphy AN , Stetler Stevenson WG, et al .Molecular aspects of tumor cell invasion and metastasis[J]. Cancer ,1993,71 (5): 1368.
[2] Parade,LF ,Land H ,Weinberg RA et al. Cooperation between genes encoding p53 tumor antigen and rad oncogene and transformation. Nature 1984,312:649-651.
[3] Finlay CA, Hinds PW, Levine AJ. The p53 proto-oncogene can act as a suppressor of transformation. Cell 1989,57:1083-93.
[4] J Frank Baker, A review of the important of the p53 gene in cancer research http://members.aye.net/-ifbaker/p53.htm
[5] Dameron KM, Volpert OV, TainskyMA, et al. Control of angiogenesis in fibroblasts by p53 regulation of thrombospondinl . Science, 1994, 265; 1582-1584.
[6] Barrett J. W, Brown wright A.J. and Primavera M.J. Studies of the nuclepolyhedrovirus infection process in insects by using the green fluorescence protein as a reporter. J Virol. 1998,7 2(4):3377-3382.
[7] Barthmaier, P .and E. Fyrberg. Monitoring development and pathology of Drosophila indirect flight muscles using green fuorescent protein. Dev. Biol. 1995.169:770-774.
[8] Brejc K .Sixma T.K.,and Kitts P.A. Structural basis for dual excitation and photoisomerization of the Aequorea victoria green fluorescent protein. Proc.Nad.Acad. Sci. USA. 1997, 94(6): 2306-2311.
[9] Chalfie, M .Green fluorescent protein. Photochem. Photobiol. 1995.62:651-656.
[10] Chalfie, M.Y. Tu ,G .Euskirchen, W.W.Ward, and D.C. Prasher. Green fluorescent protein as a marker for gene expression. Science 1994 :802- 805.
[11] Crameri, A, E.A. Whitehorn, E .Tate, and W.P.C.Stemmer, Improved green fluorescent protein by molecular evolution using DNA shuffling. Nat.Biotechnol. 1996.14:315-319.
[12] Ferreia GN,Monteiro GA,Prazeres DM,et al. Downstream processing of plasmid DNA for gene therapy and DNA vaccine applications. Trends Biotechnol. 2000, 18(9):380-388.
[13] Pankhurst Q.A..,Connolly J. Jones S.K., Dobson J. Applications of magnetic nanoparticles in biomedicine. J.Phys.D: Appl. Phys. 2003,36,R167.
[14] Tartaj P. Morales M.P. Veintemillas-Verdaguer S. et al. The preparation of magnetic nanoparticles for application in biomedicine. J.Phys. D:Appl. Phys. 2003 ,36 ,R182.
[15] Berry C.C. Curtis A.S.G. Functionalisation of magnetic nanoparticles for applications in biomedicine. J.Phys.D: Appl.Phys. 2003,36,r198.
[16] Ibrahim A,Couvreur P, Roland M. New magnetic drug carrier Journal of Pharmacy and Pharmacology (England), 1983,35:59-61.
[17] Hafeli U. Schutt W. Teller J. Scientific and Clinical Applications of Magnetic Carriers. Plenum Press: New York, 1997.
[18] Christopher H.D. Hui-Chen S. Wen-Jang C. et al. Normal T-cell response and in vivo magnetic resonance imaging of T cells loaded with HIV transactivator-peptide-derived supeparamagnetic nanoparticles. J. Immunological Mathods, 2001,256,89.
[19] Hergt R , Andra W. D Ambly .C.G. Higer I. et al. Physical limits of hyperthermia using magnetite fine particles. IEEE. Trans. Mag. 1998.34.3745.
[20] Jinquan C. Yongxian W. Junfeng Y. et al. Preparation and radiolabeling of surface-modified magnetic nanoparticles with rhenium-188 for magnetic targeted radiotherapy . J.Magn.Mater. 2004,277.165.
[21] Xin X,Xuz,Huan Z,Depu C. Weiyang F. Preparation and application of surface-coated superparamagnetic nanobeads in the isolation of genomic DNA. J. Magn. Magn. Mater. 2004.227.16.
[22] Wolfgang J.P. Daniele G. Teraes P. Daniela Z, et al. Biological applications of colloidal nanocrystals. Nanotechnology, 2003,14.R15.
[23] Patrick S.D.Jerome B,Aurelien B. Jean-Louis V. Self-assembled magetic matrices for DNA separation chips. Science. 2002,295,2237.
[24] Nam JM. Stoeva S.I. Mirkin C.A. Bio-bar-code based DNA detection with PCR-like sensitivit. J.Am.Chem.Soc. 2004.126.5932.
[25] Josephson L. Perez J.M.Weissleder R. Magnetic nanosensors for the detection of oligonucleotide sequences. Angew.Chem.Int.Ed. 2001,40.3204.
[26] Miller M.M. Sheehan P.E.Edelstein R.L,et al. A DNA array sensor utilizing magnetic microbeads and magnetoelectronic detection. J.Magn.Magn.Mater. 2001,225,138.
[27] Maite L . Nadia C. Ching-Hsuan T.et al. Tat peptied-derivatized magnetic nanoparticles allow in vivo tracking and recovery of progenitor cells. Nature Biotechnology,2000,18,410.
[28]O'Handley R C.Modern magnetic materials-principles and applications,2000,John Wiley & Sons press.
[29]Kim D K,Mikhaylova M,Zhang Y,et al.Protective coating of superparamagnetic iron oxide nanoparticles.Chem.Mater.,2003,15(8):1617-1627.
[30]Godovsky D Y,Varfolomeev A V,Efremova G D,Cherepanov V M,Kapustin G A,Volkov A V,Moskvina M A.Magnetic properties of polyvinyl alcohol-based composites containing iron oxide nanoparticles.Adv.Mater.Opt.Electron.1999,9,87-93.
[31]Feigner JH,Kumar R,Sridhar CN,et al.Enhanced gene delivery and mechanism studies with a novel series of cationic lipid formulations.J Biol Chem 1994;269:2550-2561.
[32]Van Craynest N,Santaella C,Boussif O,et al.Polycationic telomers and cotelomers for gene transfer:synthesis and evaluation of their in vitro transfection efficiency.Bioconjug Chem 2002;13:59-75.
[33]Lau C,Soriano HE,Ledley FD,et al.Retroviral gene transfer into the intestinal epithelium.Hum Gene Ther 1995;6:1145-1151.
[34]Scollay R.Gene therapy:a brief overview of the past,present,and future.Ann N YAcad Sci 2001;953:26-30.
[35]Cohen H,Levy RJ,Gao J,et al.Sustained delivery and expression of DNA encapsulated in polymeric nanoparticles.Gene Ther 2000;7:1896-1905.
[36]EngelhardtJF,YeX,DoranzB,et al.Ablation of E2A in recombinant adenoviruses improves transgene persistence and decreases inflammatory response in mouse liver.Proc Natl Acad Sci USA 1994;91:6196-6200.
[37]Niidome T,Ohmori N,Ichinose A,et al.Binding of cationic α-helical peptides to plasmid DNA and their gene transfer abilities into cells.J Biol Chem 1997;272:15 307-15 312.
[38]Chan CK,Senden T,Jans DA.Supramolecular structure and nuclear targeting efficiency determine the enhancement of transfection by modified polylysine.Gene Ther 2000;7:s1690-1697.
[39]杜仕国,施冬梅,韩其文,纳米颗粒的液相合成技术,粉末冶金技术,2000,18(1):46-50.
[40]Davis S S.Biomedical aplication of nanotechnology implications for drug targeting and gene therapy.Trends in Biotech,1997,15(2):217-224.
[41]Lambert G;Fatal E,Couvreur P.Nanoparticulate systems for the delivery of antisense oligonucleotides.Adv Drug Deliv Rev,2001,47(1):99-112.
[42]Schatzlein A G.Non-viral vectors in cancer gene therapy:principles and progress.Anti-Cancer Drugs,2001,12(5):275-304.
[43]Peppas A.Drug diffusion and binding in ionizable interpenetrating net-works fr m poly(vinyl alcohol) and poly(acrylic acid).Eru J Pharm Biopharm,1998,46(1):15-30.
[44]McDevitt MR,Ma D,Lai LT,et al.Tumor therapy with targeted atomic nanogenerators.Science,2001,29 4(12):1537-1540.
[45]Cosma-Cachita D,Bica D,Tarca A,et al.Magnetoliposomes obtained from lecithin and Fe3O4 nanoparticles.Rom J Physiol,1999,36(3-4):233-23.
[46]Kim D K,Mikhaylova M,Wang F H,et al.Starch-Coated Superparamagnetic Nanoparticles as MR Contrast Agents.Chem.Mater.,2003,15(23):4343-4351.
[47]Shen X,Fang X,Zhou Y,et al.Synthesis and characterization of 3-aminopropyltriethoxysilane-modified superparamagnetic magnetite nanoparticles.Chem.Lett.,2004,33(11):1468-1469.
[48]M Yamaura,R L Camilo,L C Sampaio,A Mac(?)do,M Nakamura,H E Toma.Preparation and characterization of(3-aminopropyl) triethoxysilane-coated magnetite nanoparticles.J.Magn.Magn.Mater.,2004,279(2-3):210-217.
[49]M Yamaura,R L Camilo,M Felinto.Synthesis and performance of organic-coated magnetite particles.J.Alloy.Compd.,2002,344(1-2):152-156.
[50]Finlay CA,Hinds PW,Levine AJ,et al.The p53 pro-oncogene can act as a suppressor of transformation.Cell,1989,57(7):1083-1093.
[51]Thapar K,Scheithauer BW,Kovacs K,et al.P53 expression in pituitary adenomas and carcinomas:correlation with invasiveness and tumor growth factions Neurosurgery,1996,38(4):763.
[52]Hollstein M,Sidransky D,and Harris CC.p53 mutations in human cancer.Science,1991,253:49-53.
[53]Y Lin,C.Y Shi,B Li et al.Tumor Suppressor P53 and Rb genes in human hepatocellular carcinoma.Ann Acad Med Singapore 1996,25:22-30.
[54]Alain Puisieux,Katherine Galvin,Frederic Troalen et al.Retinabiasloma and p53 tumor suppressor genes in human hepatoma cell lines.The FASEB Journal1993,7:1407-13.
[55] Brigite Bressac, Katherine M. Galvin, T Jake Liang et al .Abnormal structure and experssion of p53 gene in human hepatocellular carcinoma. PNAS 1990,87:1973-77.
[56] Kekule Asgari, Isabelle A .Sesterhenn, David G .McLeod et al . Inhibition of the growth of pre-established subcutaneous tumor nodules of human prostate cancer cells by single injection of the recombinant adenovirus p53 expression vector. Int J Cancer, 1997,71 :377-382.
[57] Dan M. Nguyen, Sandra A .Wiehle, Patricia E.Koch et al .Delivery of the p53 tumor suppressor gene into lung cancer cells by an adenovirus/DNA complex. Cancer Gene Therapy 1997,4(3): 191- 198.
[58] Kalpana Mujoo, Daniel C Maneal, Scot C Anderson et al. Adenoviral-mediated p53 tumor suppressor gene therapy of human ovariancarcinoma. Oncogene 1996,12:1617-1623.
[59] Jian-hua Li, Peixiang Li, Henry kiamut et al. Cytotoxic effects of Ad5CMV-p53 experssion in two human nasopharyngeal carcinoma cell lines. Clinical Cancer Research,1997,3:507-514.
[60] C.Dees,V.L.Godfrey,R.D.Schults et al. Wild type p53 reduces the size of tumors caused by bovine leukemia virus-infected cells. Cancer leters, 1996,101:115-122.
[61] Nielsen L L,et al. Adenovirus-mediated p53 gene therapy and paclitaxel have synergistic efficacy in models of human head and neck, ovarian, porstate, and berast cancer. Clin Cancer Res, 1998,4(4)835-846.
[62] W ILL IAMS D A. Gene therapy advances but struggles to interp ret safety data in small animal models [J]. Mol Ther, 2006, 13 ( 6): 1027-1028.
[63] TAND IA BM,LONEZ C,VANDENBRANDENM, et al. Lip id mix2 ing between lipop lexes and p lasma lipop roteins is a major barrier for intravenous transfection mediated by cationic lip ids[J]. JBiol Chem,2005,280(13) :12255-12261.
[64] LIEBERMAN DA, HOFFMAN B, STEINMAN RA. Molecular control of growth arrest and apoptosis: p532dependent and independent pathways[J]. Oncogene, 1995, 11 (1): 199-210.
[65] ANNAM, EDGARDOM, ALEXEIB, et al. Tumoral distribution of long circulating dextran-coated iron oxide nanoparticles in a rodent model[J]. Radiology, 2000, 214 (2): 568-576.
[66]向娟娟,聂新民,唐敬群,等.磁性氧化铁纳米颗粒用于体外基因的转染及其外加磁场对于转染效率的影响[J].中华肿瘤杂志,2004,26(2):71-74.
[67]DOBSON J.Gene therapy progress and prospects:magnetic nano particle based gene delivery[J].Gene Ther,2006,13(4):283-287.
[68]KIM J S,YOON T J,YU KN,et al.Cellular uptake of magnetic nanoparticle is mediated through energy dependent endocytosis in A549 cells[J].J Vet Sci,2006,7(4):321-326.
[69]Luo D,Saltzman WM.Enhancement of transfection by physical concerntration of DNA at the cell surface.Nat Biotechnol,2000;18:893-889.
[70]F Scheer,Manton,U Schiliinger,et al.Magnetofection:enhancing and targeting genedelivery by magneticforce in vitro and in vivo.Gene therapy,2002,9:102-109.
[71]Kro tz E Wit C,Sohn HY et al.Magnetofection A Highly efficient tool for antisense oligonuleotide delivery in vitro and in vivo[J].Mol Ther,2003,7(5):700-710.
[72]Plank C,Scherer F,Sohn HY et al.Magnetofection:enhancing and targeting gene delivery with superparamagnetic nanoparticles and magnetic fields.[J].J Liposome Res,2003,13(1):29-32.
[73]Lane DE p53,guardian of the genome[J].Nature,1992,358:15-16.
[74]Ressel A,Weiss C,Feyerabend T.Tumor oxygenation after radiotherapy,chemotherapy,and /or hyperthermia predicts tumor free survival[J].Int J Radiat Oncol Biol Phys,2001,49(4):111921125.
[75]Morita M,Kuwano H,Araki K,et al.Prognostic significance of lymphocyte infiltration following preoperative chemoradiotherapy and hyperthermia for esophageal cancer[J].Int J Radiat Oncol Biol Phys,2001,49(5):125921266.
[76]Masunaga S,Ono K,Takahashi A,et al.Usefulness of combined treat treatment of solid tumors:its independence of p53 status[J]Cancer Sci,2003,94(1):1252133.
[77]Roca C,Primo L,Valdembri D,et al.Hyperthermia inhibits angiogenesis by a plasminogen activator inhibitor I-dependent mechanism[J].Cancer Res,2003,63(7):150021507.
[78]Mohamed F,Marchettini P,Stuart OA,et al.Thermal enhancement of new chemotherapeutic agents at moderate hyperthermia[J].Ann Surg Oncol,2003,10(4):463 2468.
[79] Jin ZH ,Matsumoto H ,Hayashi S , et al. p53 independent thermo-sensitization by mitomycin C in human non-small-cell lung cancer cells[J] .Int J Radiat Oncol Biol Phys ,2004 ,59 (3) :8522860.
[80] Ohnishi K. Heat induced p53 dependents signal transduction and its role in hyperthermic cancertherapy [J]. Int J Hypeitheraiia ,2001 ,17 (5) :4152427.
[81] Qi V ,Weinrib L ,Ma N , et al . Adenoviral p53 gene therapy promotes heat-induced apoptosis in anasopharyngeal carcinoma cell line[J] . Int J Hyperthermia ,2001 ,17(1) :38247.
[1]Fox JL.Gene therapy safety issues come to fore.Nat Biotechnol.1999 Dec;17(12):1153.
[2]赵世巧,冯文莉.基因治疗的病毒载体研究进展.国外医学临床生物化学与检验学分册,2005,26(10):709-711.
[3]Wu X,Li Y,Crise B,Burgess SM.Transcription start regions in the human genome are favored targets for MLV integration.Science.2003 Jun 13;300(5626):1749-1751.
[4]Ragusa A,Garc(?)a I,Penad(?)s S.Nanoparticles as nonviral gene delivery vectors.IEEE Trans Nanobioscience.2007 Dec;6(4):319-30.Review.
[5]Wang JT,Peng DY,Chen M,Ye JS.Gene delivery for lung cancer using nonviral gene vectors.Pharmazie.2007 Oct;62(10):723-6.Review.
[6]Ilya Koltover,Kathrin Wagner,Cyrus R.Safinya.DNA condensation in two dimensions.Proc Natl Acad Sci U S A.2000,97(26):14046-14051.
[7]Jeong JH,Song SH,Lim DW.DNA transfection using linear poly(ethylenimine) prepared by controlled acid hydrolysis of poly (2-ethyl-2-oxazoline).J Control Release.2001,73(2-3):391-399.
[8]Kwoh DY,Coffin CC,Lollo CP,et al.Stabilization of poly-L-lysine/DNA polyplexes for in vivo gene delivery to the liver.Biochim Biophys Acta.1999,1444(2):171-190.
[9]Golan R,Pietrasanta LI,Hsieh W,et al.DNA toroids:stages in condensation.Biochemistry.1999,38(42):14069-14076.
[10]Tang M,Szoka FC Jr.The influence of polymer structure on the interactions of cationic polymer with DNA and morphology of resulting complexes.Gene Ther.1997,4:823-832.
[11]Shoichiro A,Toshihiro A,Atsushi M.Bi-phasic polycation for the DNA carrier responding to endosomal PH.Colloids and Surfaces B:Biointerfaces.2001,22:183-191.
[12]Zanta MA,SBelguise-Valladier P,Behr JP.Gene delivery:a single nuclear localization signal peptide is sufficient to carry DNA to the cell nucleus.Proc Natl Acad Sci USA.1999,96:91-96.
[13]向娟娟,朱诗国,吕文斌,等.用氧化铁磁性颗粒作为基因载体的研究[J].癌症,2001,20(10):1009-1014.
[14]Wei W,Xu C,Wu H.Magnetic iron oxide nanoparticles mediated gene therapy for breast cancer--an in vitro study.J Huazhong Univ Sci Technolog Med Sci.2006;26(6):728-30.
[15]Scherer F,Manton,Schillinger U,et al.Magnetofec2 tion:anhancing ang targeting genedelivery by magnet2 icforce in vitro and in vivo[J].Gene therapy,2002,9:102-109.
[16]常津,刘海峰,刘晓秋,等.一种新型纳米基因载体的制备及体外实验[J].中国生物医学工程学报,2002,21(6):515-519,529.
[17]Olbrich C,Bakowsky U,Lehr CM,et al.Cationic solid - lipid nanoparticles can efficiently bind and transfect plasmid DNA[J].J Control Release,2001,77(3):345-355.
[18]Janes KA,Calve P,Alonso MJ.Poly saccharide co12 loidal particles as delivery systems for macromolecule[J].Adv Drug Deliv Rev,2001,47(1):83-97.
[19]李拥军,管珩,郑日宏,等.纳米粒子介导特异基因转染的实验研究[J].中华医学杂志,2002,82(5):341-344.
[20]Danluo,Mark S.Enhancement of transfection by physial concentration of DNA at the cell surface[J].Nat Biot,2000,13:893-895.
[21]Carsten K,Mohammad S,Elenore G.Silica nanoparti2 cle modified with aminosilanes as carriers for plasmid DNA[J].Intern J of Pharmac,2000,196:257-261.
[22]Li Z,Zhu S,Gan K,Zhang Q,Zeng Z,Zhou Y,Liu H,Xiong W,Li X,Li G.Poly-L-lysine-modified silica nanoparticles:a potential oral gene delivery system.J Nanosci Nanotechnol.2005 Aug;5(8):1199-203.
[23]薛志刚,郑多,阮键明,等.硅纳米颗粒作为基因转染载体的研究[J].遗传学报,2003,30(7):606-610.
[24]Weissleder R,Elizondo G,Wittenberg J,et al.Ultra2 small superaramagnetic iron oxide;characterization of a new class of contrast agents for MR imaging [J].Rad,1990,175:489-493.
[25]Culver KW,Ram I,Wallbridge S,et al.In vivo trans2 fer with retroviral vector producer cells for treatment of experimental brain tumors[J].Sciences,1992,256:1550-1552.
[26]Kajiwara E,Kawano K,I-Hattori Y,Fukushima M,Hayashi K,Maitani Y.Long-circulating liposome-encapsulated ganciclovir enhances the efficacy of HSV-TK suicide gene therapy.J Control Release.2007 Jul 16;120(1-2):104-10.Epub 2007 Apr 21.
[27]何勤,张志荣,苏敏,等.载TK基因聚丙交酯乙交酯纳米粒的制备及有关性质研究[J].生物医学工程学杂志,2002,19(1):30-33.
[28]杨菁,宋存先,王彭延,等.载反义MCP-1纳米粒子的制备及应用[J].医学研究通讯,2002,31(4):11-14.
[29]黄伟,崔光华,贺俊峰,等.壳聚糖纳米粒用作基因递送载体的初步研究[J].药学学报,2002,37(12):981-985.
[30]Chorny M,Polyak B,Alferiev IS,Walsh K,Friedman G,Levy RJ.Magnetically driven plasmid DNA delivery with biodegradable polymeric nanoparticles.FASEB J.2007Aug;21(10):2510-9.Epub 2007 Apr 2.
[1]Tsuzuki T,Sugioka A,Ueda M.Hepatic resection for hepatoce Uular carcinoma [J].Surgery,1990,107:5111.
[2]Paquet K- J,Koussouris P,Mercado MA,et al.Limited hepatic resection for selected cirrhotic patients with hepatocellular or cholangiocellular carcinoma:a prospective study[J].Br J Surgrey,1991,78:4591.
[3]France D,Capussotti L,Smadja C,et al Resection of hepatocellular carcinoma:results in 72 European patients with cirrhosia[J].Gastroenterrology,1990,98:7331.
[4]Aznavoorian S,Murphy AN,Stetler Stevenson WG,et al.Molecular aspects of tumor cell invasion and metastasis[J].Cancer,1993,71(5):1368.
[5]Kiluchi A,Aoki Y,Sugaya S,et al.Development of novel cationic liposomes for efficient gene transfer into peritoneal disseminated tumor[J].Human Gene Therapy,1999,10(9):947.
[6]张腾飞,钱关祥,陈诗书.人类基因治疗研究的现状[J].生命的化学,1999,6(1):57.
[7]Hsu HC,Metcalf RA,Jeffery PD,et al.Crystal structure of a P53 gene in human hepatocellular carcinomas[J].Nature,1991,350:427.
[8]Bressac B,KewM,Wands J,et al.Selective G to Tmutation of P53 gene in hepatocellular carcinoma from southern Africa[J].Nature,1991,350:429.
[9]Kress S,John UR,Buchmann A,et al.Mutations in human hepatocellular carcinomas from Germany[J].Cancer Res,1992,52:220.
[10]Challen C,Lunac J,WarrenW,et al.Analysis of the P53 tumor - suppressor gene in hepatocellular carcinomas from Britain[J].Hepatology,1992,16(1):362.
[11]Murakami Y,Hayashi K,Hirohashi S,et al.Aberration of the tumor suppressor P53 and retain diastoma genes in human hep2 atocellular carcinomas[J].Cancer Res,1991,51(5):520.
[12]Aguilar F,Hassain SP,Ceruth P.Aflatoxin Bl inuces the trans - version of G to T in colon 249 of the P53 tumor - supp ressor gene in human hepatocyies[J].Proc NarlAcad SciUSA.1990,87(1):973.
[13]朱明华.肿瘤抑制基因P53的生物学功能研究进展和意义[J].中华病理学 杂志,2000,29(1):60.
[14]郭琳琅,曹长安,朱新生.Fas抗原和突变型P53蛋白在肝癌组织中的表达及意义[J].实用癌症杂志,1997,12(4):266.
[15]Itoh T,Shiro T,Seki T,et al.Relationship between P53 over - expression and the proliferation activity in hepatocellular carcinoma[J].Int J MolMed,2000,6(2):137.
[16]Brfssac B,Galvin KM,Hang TJ,et al.Abnormal structore and expression of P53 gene in human hepatocellular carcinoma[J].Proc Natl Acad Sci USA,1990,87(1):973.
[17]戴益民,许炳华,朱明华,等.人肝细胞癌P53异常的预后意义[J].华人消化杂志,1998,6(12):1043.
[18]Steele RJ,Thomp son AM,Hall PA,et al.The P53 tumor suppressor gene[J].Br J Surg,1998,85(11):1460.
[19]Guan YS,Liu Y,Sun L,et al.Successful management of postoperative recurrence of hepatocellular carcinoma with p53 gene therapy combining transcatheter arterial chemoembolization[J].World J Gastroenterol,2005,11(24):3803-3805.
[20]Gerolami R,Cardoso J,Lewin M,et al.Evaluation of HSV- tk gene therapy in a rat model of chemically induced hepatocellular carcinoma by intratumoral and intra hepatic artery routes[J].Cancer Res,2000,60(4):993-1001.
[21]Anderson SC,Johnson DE,Harris MP,et al.p53 gene therapy in a rat model of hepatocelluar carcinoma:intra- arterial delivery of a recombinant adenovirus[J].Clin Cancer Res,1998,4(7):1649-1659.
[22]Habib NA,Hodgson HJ,Lemoine N,et al.A phase Ⅰ/Ⅱ study of hepatic artery infusion with wtp53 - CMV - Ad in metastatic malignant liver tumours[J].Hum Gene Ther,1999,10(12):2019-2034.
[23]Atencio IA,Grace M,Bordens R,et al.Biological activities of a recombinant adenovirus p53(SCH58500) administered by hepatic arterial infusion in a phase 1 colorectal cancer trial[J].Cancer Gene Ther,2006,13(2):169-181.
[24]Shiba H,Okamoto T,Futagawa Y,et al.Efficient and cancer-selective gene transfer to hepatocellular carcinoma in a rat using adenovirus vector with iodized oil esters[J].Cancer Gene Ther,2001,8(10):713-718.
[25]Guan YS,Liu Y,Zhu XP,et al.p53 gene(Gendicine)and embolisation overcame recurrent hepatocellular carcinoma[J].World J Gastroenterol,2005,11(24): 3803- 3805.
[26] Zhu ZB, Liu JL, Sui J, et al. Clinical study of recombinant adenovirus p53 gene on advanced hepatocellular carcinoma[J]. Chin J Intern Med, 2004, 11(3): 11-14.
[2] Parade,LF ,Land H ,Weinberg RA et al. Cooperation between genes encoding p53 tumor antigen and rad oncogene and transformation. Nature 1984,312:649-651.
[3] Finlay CA, Hinds PW, Levine AJ. The p53 proto-oncogene can act as a suppressor of transformation. Cell 1989,57:1083-93.
[4] J Frank Baker, A review of the important of the p53 gene in cancer research http://members.aye.net/-ifbaker/p53.htm
[5] Dameron KM, Volpert OV, TainskyMA, et al. Control of angiogenesis in fibroblasts by p53 regulation of thrombospondinl . Science, 1994, 265; 1582-1584.
[6] Barrett J. W, Brown wright A.J. and Primavera M.J. Studies of the nuclepolyhedrovirus infection process in insects by using the green fluorescence protein as a reporter. J Virol. 1998,7 2(4):3377-3382.
[7] Barthmaier, P .and E. Fyrberg. Monitoring development and pathology of Drosophila indirect flight muscles using green fuorescent protein. Dev. Biol. 1995.169:770-774.
[8] Brejc K .Sixma T.K.,and Kitts P.A. Structural basis for dual excitation and photoisomerization of the Aequorea victoria green fluorescent protein. Proc.Nad.Acad. Sci. USA. 1997, 94(6): 2306-2311.
[9] Chalfie, M .Green fluorescent protein. Photochem. Photobiol. 1995.62:651-656.
[10] Chalfie, M.Y. Tu ,G .Euskirchen, W.W.Ward, and D.C. Prasher. Green fluorescent protein as a marker for gene expression. Science 1994 :802- 805.
[11] Crameri, A, E.A. Whitehorn, E .Tate, and W.P.C.Stemmer, Improved green fluorescent protein by molecular evolution using DNA shuffling. Nat.Biotechnol. 1996.14:315-319.
[12] Ferreia GN,Monteiro GA,Prazeres DM,et al. Downstream processing of plasmid DNA for gene therapy and DNA vaccine applications. Trends Biotechnol. 2000, 18(9):380-388.
[13] Pankhurst Q.A..,Connolly J. Jones S.K., Dobson J. Applications of magnetic nanoparticles in biomedicine. J.Phys.D: Appl. Phys. 2003,36,R167.
[14] Tartaj P. Morales M.P. Veintemillas-Verdaguer S. et al. The preparation of magnetic nanoparticles for application in biomedicine. J.Phys. D:Appl. Phys. 2003 ,36 ,R182.
[15] Berry C.C. Curtis A.S.G. Functionalisation of magnetic nanoparticles for applications in biomedicine. J.Phys.D: Appl.Phys. 2003,36,r198.
[16] Ibrahim A,Couvreur P, Roland M. New magnetic drug carrier Journal of Pharmacy and Pharmacology (England), 1983,35:59-61.
[17] Hafeli U. Schutt W. Teller J. Scientific and Clinical Applications of Magnetic Carriers. Plenum Press: New York, 1997.
[18] Christopher H.D. Hui-Chen S. Wen-Jang C. et al. Normal T-cell response and in vivo magnetic resonance imaging of T cells loaded with HIV transactivator-peptide-derived supeparamagnetic nanoparticles. J. Immunological Mathods, 2001,256,89.
[19] Hergt R , Andra W. D Ambly .C.G. Higer I. et al. Physical limits of hyperthermia using magnetite fine particles. IEEE. Trans. Mag. 1998.34.3745.
[20] Jinquan C. Yongxian W. Junfeng Y. et al. Preparation and radiolabeling of surface-modified magnetic nanoparticles with rhenium-188 for magnetic targeted radiotherapy . J.Magn.Mater. 2004,277.165.
[21] Xin X,Xuz,Huan Z,Depu C. Weiyang F. Preparation and application of surface-coated superparamagnetic nanobeads in the isolation of genomic DNA. J. Magn. Magn. Mater. 2004.227.16.
[22] Wolfgang J.P. Daniele G. Teraes P. Daniela Z, et al. Biological applications of colloidal nanocrystals. Nanotechnology, 2003,14.R15.
[23] Patrick S.D.Jerome B,Aurelien B. Jean-Louis V. Self-assembled magetic matrices for DNA separation chips. Science. 2002,295,2237.
[24] Nam JM. Stoeva S.I. Mirkin C.A. Bio-bar-code based DNA detection with PCR-like sensitivit. J.Am.Chem.Soc. 2004.126.5932.
[25] Josephson L. Perez J.M.Weissleder R. Magnetic nanosensors for the detection of oligonucleotide sequences. Angew.Chem.Int.Ed. 2001,40.3204.
[26] Miller M.M. Sheehan P.E.Edelstein R.L,et al. A DNA array sensor utilizing magnetic microbeads and magnetoelectronic detection. J.Magn.Magn.Mater. 2001,225,138.
[27] Maite L . Nadia C. Ching-Hsuan T.et al. Tat peptied-derivatized magnetic nanoparticles allow in vivo tracking and recovery of progenitor cells. Nature Biotechnology,2000,18,410.
[28]O'Handley R C.Modern magnetic materials-principles and applications,2000,John Wiley & Sons press.
[29]Kim D K,Mikhaylova M,Zhang Y,et al.Protective coating of superparamagnetic iron oxide nanoparticles.Chem.Mater.,2003,15(8):1617-1627.
[30]Godovsky D Y,Varfolomeev A V,Efremova G D,Cherepanov V M,Kapustin G A,Volkov A V,Moskvina M A.Magnetic properties of polyvinyl alcohol-based composites containing iron oxide nanoparticles.Adv.Mater.Opt.Electron.1999,9,87-93.
[31]Feigner JH,Kumar R,Sridhar CN,et al.Enhanced gene delivery and mechanism studies with a novel series of cationic lipid formulations.J Biol Chem 1994;269:2550-2561.
[32]Van Craynest N,Santaella C,Boussif O,et al.Polycationic telomers and cotelomers for gene transfer:synthesis and evaluation of their in vitro transfection efficiency.Bioconjug Chem 2002;13:59-75.
[33]Lau C,Soriano HE,Ledley FD,et al.Retroviral gene transfer into the intestinal epithelium.Hum Gene Ther 1995;6:1145-1151.
[34]Scollay R.Gene therapy:a brief overview of the past,present,and future.Ann N YAcad Sci 2001;953:26-30.
[35]Cohen H,Levy RJ,Gao J,et al.Sustained delivery and expression of DNA encapsulated in polymeric nanoparticles.Gene Ther 2000;7:1896-1905.
[36]EngelhardtJF,YeX,DoranzB,et al.Ablation of E2A in recombinant adenoviruses improves transgene persistence and decreases inflammatory response in mouse liver.Proc Natl Acad Sci USA 1994;91:6196-6200.
[37]Niidome T,Ohmori N,Ichinose A,et al.Binding of cationic α-helical peptides to plasmid DNA and their gene transfer abilities into cells.J Biol Chem 1997;272:15 307-15 312.
[38]Chan CK,Senden T,Jans DA.Supramolecular structure and nuclear targeting efficiency determine the enhancement of transfection by modified polylysine.Gene Ther 2000;7:s1690-1697.
[39]杜仕国,施冬梅,韩其文,纳米颗粒的液相合成技术,粉末冶金技术,2000,18(1):46-50.
[40]Davis S S.Biomedical aplication of nanotechnology implications for drug targeting and gene therapy.Trends in Biotech,1997,15(2):217-224.
[41]Lambert G;Fatal E,Couvreur P.Nanoparticulate systems for the delivery of antisense oligonucleotides.Adv Drug Deliv Rev,2001,47(1):99-112.
[42]Schatzlein A G.Non-viral vectors in cancer gene therapy:principles and progress.Anti-Cancer Drugs,2001,12(5):275-304.
[43]Peppas A.Drug diffusion and binding in ionizable interpenetrating net-works fr m poly(vinyl alcohol) and poly(acrylic acid).Eru J Pharm Biopharm,1998,46(1):15-30.
[44]McDevitt MR,Ma D,Lai LT,et al.Tumor therapy with targeted atomic nanogenerators.Science,2001,29 4(12):1537-1540.
[45]Cosma-Cachita D,Bica D,Tarca A,et al.Magnetoliposomes obtained from lecithin and Fe3O4 nanoparticles.Rom J Physiol,1999,36(3-4):233-23.
[46]Kim D K,Mikhaylova M,Wang F H,et al.Starch-Coated Superparamagnetic Nanoparticles as MR Contrast Agents.Chem.Mater.,2003,15(23):4343-4351.
[47]Shen X,Fang X,Zhou Y,et al.Synthesis and characterization of 3-aminopropyltriethoxysilane-modified superparamagnetic magnetite nanoparticles.Chem.Lett.,2004,33(11):1468-1469.
[48]M Yamaura,R L Camilo,L C Sampaio,A Mac(?)do,M Nakamura,H E Toma.Preparation and characterization of(3-aminopropyl) triethoxysilane-coated magnetite nanoparticles.J.Magn.Magn.Mater.,2004,279(2-3):210-217.
[49]M Yamaura,R L Camilo,M Felinto.Synthesis and performance of organic-coated magnetite particles.J.Alloy.Compd.,2002,344(1-2):152-156.
[50]Finlay CA,Hinds PW,Levine AJ,et al.The p53 pro-oncogene can act as a suppressor of transformation.Cell,1989,57(7):1083-1093.
[51]Thapar K,Scheithauer BW,Kovacs K,et al.P53 expression in pituitary adenomas and carcinomas:correlation with invasiveness and tumor growth factions Neurosurgery,1996,38(4):763.
[52]Hollstein M,Sidransky D,and Harris CC.p53 mutations in human cancer.Science,1991,253:49-53.
[53]Y Lin,C.Y Shi,B Li et al.Tumor Suppressor P53 and Rb genes in human hepatocellular carcinoma.Ann Acad Med Singapore 1996,25:22-30.
[54]Alain Puisieux,Katherine Galvin,Frederic Troalen et al.Retinabiasloma and p53 tumor suppressor genes in human hepatoma cell lines.The FASEB Journal1993,7:1407-13.
[55] Brigite Bressac, Katherine M. Galvin, T Jake Liang et al .Abnormal structure and experssion of p53 gene in human hepatocellular carcinoma. PNAS 1990,87:1973-77.
[56] Kekule Asgari, Isabelle A .Sesterhenn, David G .McLeod et al . Inhibition of the growth of pre-established subcutaneous tumor nodules of human prostate cancer cells by single injection of the recombinant adenovirus p53 expression vector. Int J Cancer, 1997,71 :377-382.
[57] Dan M. Nguyen, Sandra A .Wiehle, Patricia E.Koch et al .Delivery of the p53 tumor suppressor gene into lung cancer cells by an adenovirus/DNA complex. Cancer Gene Therapy 1997,4(3): 191- 198.
[58] Kalpana Mujoo, Daniel C Maneal, Scot C Anderson et al. Adenoviral-mediated p53 tumor suppressor gene therapy of human ovariancarcinoma. Oncogene 1996,12:1617-1623.
[59] Jian-hua Li, Peixiang Li, Henry kiamut et al. Cytotoxic effects of Ad5CMV-p53 experssion in two human nasopharyngeal carcinoma cell lines. Clinical Cancer Research,1997,3:507-514.
[60] C.Dees,V.L.Godfrey,R.D.Schults et al. Wild type p53 reduces the size of tumors caused by bovine leukemia virus-infected cells. Cancer leters, 1996,101:115-122.
[61] Nielsen L L,et al. Adenovirus-mediated p53 gene therapy and paclitaxel have synergistic efficacy in models of human head and neck, ovarian, porstate, and berast cancer. Clin Cancer Res, 1998,4(4)835-846.
[62] W ILL IAMS D A. Gene therapy advances but struggles to interp ret safety data in small animal models [J]. Mol Ther, 2006, 13 ( 6): 1027-1028.
[63] TAND IA BM,LONEZ C,VANDENBRANDENM, et al. Lip id mix2 ing between lipop lexes and p lasma lipop roteins is a major barrier for intravenous transfection mediated by cationic lip ids[J]. JBiol Chem,2005,280(13) :12255-12261.
[64] LIEBERMAN DA, HOFFMAN B, STEINMAN RA. Molecular control of growth arrest and apoptosis: p532dependent and independent pathways[J]. Oncogene, 1995, 11 (1): 199-210.
[65] ANNAM, EDGARDOM, ALEXEIB, et al. Tumoral distribution of long circulating dextran-coated iron oxide nanoparticles in a rodent model[J]. Radiology, 2000, 214 (2): 568-576.
[66]向娟娟,聂新民,唐敬群,等.磁性氧化铁纳米颗粒用于体外基因的转染及其外加磁场对于转染效率的影响[J].中华肿瘤杂志,2004,26(2):71-74.
[67]DOBSON J.Gene therapy progress and prospects:magnetic nano particle based gene delivery[J].Gene Ther,2006,13(4):283-287.
[68]KIM J S,YOON T J,YU KN,et al.Cellular uptake of magnetic nanoparticle is mediated through energy dependent endocytosis in A549 cells[J].J Vet Sci,2006,7(4):321-326.
[69]Luo D,Saltzman WM.Enhancement of transfection by physical concerntration of DNA at the cell surface.Nat Biotechnol,2000;18:893-889.
[70]F Scheer,Manton,U Schiliinger,et al.Magnetofection:enhancing and targeting genedelivery by magneticforce in vitro and in vivo.Gene therapy,2002,9:102-109.
[71]Kro tz E Wit C,Sohn HY et al.Magnetofection A Highly efficient tool for antisense oligonuleotide delivery in vitro and in vivo[J].Mol Ther,2003,7(5):700-710.
[72]Plank C,Scherer F,Sohn HY et al.Magnetofection:enhancing and targeting gene delivery with superparamagnetic nanoparticles and magnetic fields.[J].J Liposome Res,2003,13(1):29-32.
[73]Lane DE p53,guardian of the genome[J].Nature,1992,358:15-16.
[74]Ressel A,Weiss C,Feyerabend T.Tumor oxygenation after radiotherapy,chemotherapy,and /or hyperthermia predicts tumor free survival[J].Int J Radiat Oncol Biol Phys,2001,49(4):111921125.
[75]Morita M,Kuwano H,Araki K,et al.Prognostic significance of lymphocyte infiltration following preoperative chemoradiotherapy and hyperthermia for esophageal cancer[J].Int J Radiat Oncol Biol Phys,2001,49(5):125921266.
[76]Masunaga S,Ono K,Takahashi A,et al.Usefulness of combined treat treatment of solid tumors:its independence of p53 status[J]Cancer Sci,2003,94(1):1252133.
[77]Roca C,Primo L,Valdembri D,et al.Hyperthermia inhibits angiogenesis by a plasminogen activator inhibitor I-dependent mechanism[J].Cancer Res,2003,63(7):150021507.
[78]Mohamed F,Marchettini P,Stuart OA,et al.Thermal enhancement of new chemotherapeutic agents at moderate hyperthermia[J].Ann Surg Oncol,2003,10(4):463 2468.
[79] Jin ZH ,Matsumoto H ,Hayashi S , et al. p53 independent thermo-sensitization by mitomycin C in human non-small-cell lung cancer cells[J] .Int J Radiat Oncol Biol Phys ,2004 ,59 (3) :8522860.
[80] Ohnishi K. Heat induced p53 dependents signal transduction and its role in hyperthermic cancertherapy [J]. Int J Hypeitheraiia ,2001 ,17 (5) :4152427.
[81] Qi V ,Weinrib L ,Ma N , et al . Adenoviral p53 gene therapy promotes heat-induced apoptosis in anasopharyngeal carcinoma cell line[J] . Int J Hyperthermia ,2001 ,17(1) :38247.
[1]Fox JL.Gene therapy safety issues come to fore.Nat Biotechnol.1999 Dec;17(12):1153.
[2]赵世巧,冯文莉.基因治疗的病毒载体研究进展.国外医学临床生物化学与检验学分册,2005,26(10):709-711.
[3]Wu X,Li Y,Crise B,Burgess SM.Transcription start regions in the human genome are favored targets for MLV integration.Science.2003 Jun 13;300(5626):1749-1751.
[4]Ragusa A,Garc(?)a I,Penad(?)s S.Nanoparticles as nonviral gene delivery vectors.IEEE Trans Nanobioscience.2007 Dec;6(4):319-30.Review.
[5]Wang JT,Peng DY,Chen M,Ye JS.Gene delivery for lung cancer using nonviral gene vectors.Pharmazie.2007 Oct;62(10):723-6.Review.
[6]Ilya Koltover,Kathrin Wagner,Cyrus R.Safinya.DNA condensation in two dimensions.Proc Natl Acad Sci U S A.2000,97(26):14046-14051.
[7]Jeong JH,Song SH,Lim DW.DNA transfection using linear poly(ethylenimine) prepared by controlled acid hydrolysis of poly (2-ethyl-2-oxazoline).J Control Release.2001,73(2-3):391-399.
[8]Kwoh DY,Coffin CC,Lollo CP,et al.Stabilization of poly-L-lysine/DNA polyplexes for in vivo gene delivery to the liver.Biochim Biophys Acta.1999,1444(2):171-190.
[9]Golan R,Pietrasanta LI,Hsieh W,et al.DNA toroids:stages in condensation.Biochemistry.1999,38(42):14069-14076.
[10]Tang M,Szoka FC Jr.The influence of polymer structure on the interactions of cationic polymer with DNA and morphology of resulting complexes.Gene Ther.1997,4:823-832.
[11]Shoichiro A,Toshihiro A,Atsushi M.Bi-phasic polycation for the DNA carrier responding to endosomal PH.Colloids and Surfaces B:Biointerfaces.2001,22:183-191.
[12]Zanta MA,SBelguise-Valladier P,Behr JP.Gene delivery:a single nuclear localization signal peptide is sufficient to carry DNA to the cell nucleus.Proc Natl Acad Sci USA.1999,96:91-96.
[13]向娟娟,朱诗国,吕文斌,等.用氧化铁磁性颗粒作为基因载体的研究[J].癌症,2001,20(10):1009-1014.
[14]Wei W,Xu C,Wu H.Magnetic iron oxide nanoparticles mediated gene therapy for breast cancer--an in vitro study.J Huazhong Univ Sci Technolog Med Sci.2006;26(6):728-30.
[15]Scherer F,Manton,Schillinger U,et al.Magnetofec2 tion:anhancing ang targeting genedelivery by magnet2 icforce in vitro and in vivo[J].Gene therapy,2002,9:102-109.
[16]常津,刘海峰,刘晓秋,等.一种新型纳米基因载体的制备及体外实验[J].中国生物医学工程学报,2002,21(6):515-519,529.
[17]Olbrich C,Bakowsky U,Lehr CM,et al.Cationic solid - lipid nanoparticles can efficiently bind and transfect plasmid DNA[J].J Control Release,2001,77(3):345-355.
[18]Janes KA,Calve P,Alonso MJ.Poly saccharide co12 loidal particles as delivery systems for macromolecule[J].Adv Drug Deliv Rev,2001,47(1):83-97.
[19]李拥军,管珩,郑日宏,等.纳米粒子介导特异基因转染的实验研究[J].中华医学杂志,2002,82(5):341-344.
[20]Danluo,Mark S.Enhancement of transfection by physial concentration of DNA at the cell surface[J].Nat Biot,2000,13:893-895.
[21]Carsten K,Mohammad S,Elenore G.Silica nanoparti2 cle modified with aminosilanes as carriers for plasmid DNA[J].Intern J of Pharmac,2000,196:257-261.
[22]Li Z,Zhu S,Gan K,Zhang Q,Zeng Z,Zhou Y,Liu H,Xiong W,Li X,Li G.Poly-L-lysine-modified silica nanoparticles:a potential oral gene delivery system.J Nanosci Nanotechnol.2005 Aug;5(8):1199-203.
[23]薛志刚,郑多,阮键明,等.硅纳米颗粒作为基因转染载体的研究[J].遗传学报,2003,30(7):606-610.
[24]Weissleder R,Elizondo G,Wittenberg J,et al.Ultra2 small superaramagnetic iron oxide;characterization of a new class of contrast agents for MR imaging [J].Rad,1990,175:489-493.
[25]Culver KW,Ram I,Wallbridge S,et al.In vivo trans2 fer with retroviral vector producer cells for treatment of experimental brain tumors[J].Sciences,1992,256:1550-1552.
[26]Kajiwara E,Kawano K,I-Hattori Y,Fukushima M,Hayashi K,Maitani Y.Long-circulating liposome-encapsulated ganciclovir enhances the efficacy of HSV-TK suicide gene therapy.J Control Release.2007 Jul 16;120(1-2):104-10.Epub 2007 Apr 21.
[27]何勤,张志荣,苏敏,等.载TK基因聚丙交酯乙交酯纳米粒的制备及有关性质研究[J].生物医学工程学杂志,2002,19(1):30-33.
[28]杨菁,宋存先,王彭延,等.载反义MCP-1纳米粒子的制备及应用[J].医学研究通讯,2002,31(4):11-14.
[29]黄伟,崔光华,贺俊峰,等.壳聚糖纳米粒用作基因递送载体的初步研究[J].药学学报,2002,37(12):981-985.
[30]Chorny M,Polyak B,Alferiev IS,Walsh K,Friedman G,Levy RJ.Magnetically driven plasmid DNA delivery with biodegradable polymeric nanoparticles.FASEB J.2007Aug;21(10):2510-9.Epub 2007 Apr 2.
[1]Tsuzuki T,Sugioka A,Ueda M.Hepatic resection for hepatoce Uular carcinoma [J].Surgery,1990,107:5111.
[2]Paquet K- J,Koussouris P,Mercado MA,et al.Limited hepatic resection for selected cirrhotic patients with hepatocellular or cholangiocellular carcinoma:a prospective study[J].Br J Surgrey,1991,78:4591.
[3]France D,Capussotti L,Smadja C,et al Resection of hepatocellular carcinoma:results in 72 European patients with cirrhosia[J].Gastroenterrology,1990,98:7331.
[4]Aznavoorian S,Murphy AN,Stetler Stevenson WG,et al.Molecular aspects of tumor cell invasion and metastasis[J].Cancer,1993,71(5):1368.
[5]Kiluchi A,Aoki Y,Sugaya S,et al.Development of novel cationic liposomes for efficient gene transfer into peritoneal disseminated tumor[J].Human Gene Therapy,1999,10(9):947.
[6]张腾飞,钱关祥,陈诗书.人类基因治疗研究的现状[J].生命的化学,1999,6(1):57.
[7]Hsu HC,Metcalf RA,Jeffery PD,et al.Crystal structure of a P53 gene in human hepatocellular carcinomas[J].Nature,1991,350:427.
[8]Bressac B,KewM,Wands J,et al.Selective G to Tmutation of P53 gene in hepatocellular carcinoma from southern Africa[J].Nature,1991,350:429.
[9]Kress S,John UR,Buchmann A,et al.Mutations in human hepatocellular carcinomas from Germany[J].Cancer Res,1992,52:220.
[10]Challen C,Lunac J,WarrenW,et al.Analysis of the P53 tumor - suppressor gene in hepatocellular carcinomas from Britain[J].Hepatology,1992,16(1):362.
[11]Murakami Y,Hayashi K,Hirohashi S,et al.Aberration of the tumor suppressor P53 and retain diastoma genes in human hep2 atocellular carcinomas[J].Cancer Res,1991,51(5):520.
[12]Aguilar F,Hassain SP,Ceruth P.Aflatoxin Bl inuces the trans - version of G to T in colon 249 of the P53 tumor - supp ressor gene in human hepatocyies[J].Proc NarlAcad SciUSA.1990,87(1):973.
[13]朱明华.肿瘤抑制基因P53的生物学功能研究进展和意义[J].中华病理学 杂志,2000,29(1):60.
[14]郭琳琅,曹长安,朱新生.Fas抗原和突变型P53蛋白在肝癌组织中的表达及意义[J].实用癌症杂志,1997,12(4):266.
[15]Itoh T,Shiro T,Seki T,et al.Relationship between P53 over - expression and the proliferation activity in hepatocellular carcinoma[J].Int J MolMed,2000,6(2):137.
[16]Brfssac B,Galvin KM,Hang TJ,et al.Abnormal structore and expression of P53 gene in human hepatocellular carcinoma[J].Proc Natl Acad Sci USA,1990,87(1):973.
[17]戴益民,许炳华,朱明华,等.人肝细胞癌P53异常的预后意义[J].华人消化杂志,1998,6(12):1043.
[18]Steele RJ,Thomp son AM,Hall PA,et al.The P53 tumor suppressor gene[J].Br J Surg,1998,85(11):1460.
[19]Guan YS,Liu Y,Sun L,et al.Successful management of postoperative recurrence of hepatocellular carcinoma with p53 gene therapy combining transcatheter arterial chemoembolization[J].World J Gastroenterol,2005,11(24):3803-3805.
[20]Gerolami R,Cardoso J,Lewin M,et al.Evaluation of HSV- tk gene therapy in a rat model of chemically induced hepatocellular carcinoma by intratumoral and intra hepatic artery routes[J].Cancer Res,2000,60(4):993-1001.
[21]Anderson SC,Johnson DE,Harris MP,et al.p53 gene therapy in a rat model of hepatocelluar carcinoma:intra- arterial delivery of a recombinant adenovirus[J].Clin Cancer Res,1998,4(7):1649-1659.
[22]Habib NA,Hodgson HJ,Lemoine N,et al.A phase Ⅰ/Ⅱ study of hepatic artery infusion with wtp53 - CMV - Ad in metastatic malignant liver tumours[J].Hum Gene Ther,1999,10(12):2019-2034.
[23]Atencio IA,Grace M,Bordens R,et al.Biological activities of a recombinant adenovirus p53(SCH58500) administered by hepatic arterial infusion in a phase 1 colorectal cancer trial[J].Cancer Gene Ther,2006,13(2):169-181.
[24]Shiba H,Okamoto T,Futagawa Y,et al.Efficient and cancer-selective gene transfer to hepatocellular carcinoma in a rat using adenovirus vector with iodized oil esters[J].Cancer Gene Ther,2001,8(10):713-718.
[25]Guan YS,Liu Y,Zhu XP,et al.p53 gene(Gendicine)and embolisation overcame recurrent hepatocellular carcinoma[J].World J Gastroenterol,2005,11(24): 3803- 3805.
[26] Zhu ZB, Liu JL, Sui J, et al. Clinical study of recombinant adenovirus p53 gene on advanced hepatocellular carcinoma[J]. Chin J Intern Med, 2004, 11(3): 11-14.