表皮生长因子受体EGFR和HER2配体寡肽与力达霉素组成的双特异性强化融合蛋白的构建与抗肿瘤活性研究
|
摘要
背景与目的:表皮生长因子受体EGFR/HER1和HER2/neu都是跨膜受体酪氨酸激酶家族(ErbB)的成员,ErbB受体家族与人类癌症的关系非常密切,EGFR和HER2在多种肿瘤细胞表面异常表达,且ErbB受体发生改变的肿瘤通常更具有侵袭性,病人也往往预后不良,这使得EGFR和HER2成为抗肿瘤靶向治疗的理想靶点。力达霉素(Lidamycin,LDM)是迄今报道过的对肿瘤细胞杀伤作用最强的大分子肽类抗肿瘤抗生素,由辅基蛋白(LDP)和烯二炔结构的发色团(AE)两部分组成,发色团是力达霉素分子的活性部分。本研究的目的是构建一种靶向EGFR和HER2的双特异性强化融合蛋白(Ec-LDP-Hr-AE),并对其体内外抗肿瘤活性进行检测。
方法:双特异性融合蛋白Ec-LDP-Hr的构建主要采用PCR扩增和DNA克隆技术。pelB信号肽序列、寡肽配体Ec基因序列、寡肽配体Hr基因序列以及(GGGGS)_2连接肽基因序列均采用人工合成的方法得到,并作为PCR引物,经过5轮PCR扩增以及DNA克隆后,得到了融合蛋白基因Ec-ldp-Hr,将该基因经过双酶切后插入到pET30a(+)表达载体中,得到重组表达载体pET-Ec-ldp-Hr。为了进行比较,本研究同时构建了单靶点的重组表达载体pET-Ec-ldp和pET-ldp-Hr。将三种表达载体均转化到大肠杆菌BL21(DE3)star~(TM)中,加入IPTG进行诱导表达,SDS-PAGE分析目的蛋白的表达情况。融合蛋白的提取采用渗透压震惊法,使用Ni~(2+)亲和层析法分离纯化融合蛋白,HPLC检测融合蛋白的纯度,Western Blot对纯化出的融合蛋白进行鉴定。融合蛋白与肿瘤细胞表面EGFR和HER2的亲和活性主要采用了ELISA、细胞免疫荧光化学法、基于流式细胞术的免疫荧光试验、竞争性免疫荧光试验以及免疫共沉淀技术进行分析。将LDM活性发色团分子(AE)与融合蛋白在体外进行分子组装,得到强化融合蛋白(Ec-LDP-Hr-AE),采用MTT法测定强化融合蛋白对肿瘤细胞的体外杀伤活性。细胞免疫荧光染色和基于流式细胞术的荧光检测用于分析融合蛋白是否可内吞进入细胞以及检测内吞效率。双特异性强化融合蛋白Ec-LDP-Hr-AE的体内抗肿瘤活性采用人卵巢癌SK-OV-3裸鼠移植瘤模型进行评价。结果:经过5轮PCR扩增以及DNA克隆后,得到了融合蛋白基因Ec-ldp-Hr,将其插入到表达载体后成功实现了在大肠杆菌中的表达。Ec-LDP-Hr蛋白和Ec-LDP蛋白主要以可溶性的形式分泌到了大肠杆菌周质腔中,而LDP-Hr蛋白除存在于周质腔中外,还分泌到了培养液上清中。Ni~(2+)亲和层析对周质腔中的蛋白进行纯化,得到的目的蛋白经HPLC检测纯度均在95%以上。Ec-LDP-Hr、Ec-LDP和LDP-Hr三种融合蛋白的产量分别为每升发酵液得到9 mg、18 mg和60 mg活性蛋白。将纯化的蛋白用Western blot方法进行鉴定,证实了纯化的蛋白为带有His-tag标签的目的蛋白。ELISA和基于流式细胞术的免疫荧光试验结果显示Ec-LDP-Hr蛋白对EGFR和HER2高表达的肿瘤细胞系,如A431、SK-BR-3、SK-OV-3、MCF-7均有很强的结合活性,而对不表达EGFR和HER2的NIH 3T3细胞则无亲和活性。细胞免疫荧光化学试验证实了Ec-LDP-Hr蛋白可以与A431和SK-BR-3细胞表面的受体结合,免疫共沉淀试验则进一步证明了Ec-LDP-Hr蛋白是与A431和SK-BR-3细胞表面的EGFR和HER2结合的;竞争性免疫荧光试验结果显示,Ec-LDP-Hr蛋白可部分抑制抗EGFR和抗HER2抗体与各自抗原的结合能力,表明该蛋白可竞争性的与EGFR和HER2结合。将AE分子与融合蛋白在体外进行分子组装后,经HPLC检测在350nm处出现了特定的吸收峰,表明AE分子与融合蛋白进行了成功组装,得到了强化融合蛋白Ec-LDP-Hr-AE。MTT试验结果显示强化融合蛋白Ec-LDP-Hr-AE对不同的肿瘤细胞系均表现出了强烈的杀伤作用,对A431、SK-BR-3、SK-OV-3、MCF-7和NIH 3T3细胞的IC_(50)值分别为:4.25×10~(-13)mol/L、4.04×10~(-14)mol/L、1.55×10~(-13)mol/L、4.7×10~(-12)mol/L和2.33×10~(-11)mol/L。对于高表达EGFR和HER2的肿瘤细胞系,双特异性强化融合蛋白Ec-LDP-Hr-AE表现出了比LDM和单靶点的强化融合蛋白Ec-LDP-AE和LDP-Hr-AE更强的杀伤活性,而对于EGFR和HER2不表达的NIH 3T3细胞,Ec-LDP-Hr-AE蛋白的IC_(50)值则高于LDM和Ec-LDP-AE、LDP-Hr-AE蛋白,表明双特异性的Ec-LDP-Hr-AE蛋白提高了对肿瘤细胞作用的特异性。细胞内吞试验结果表明,Ec-LDP-Hr和Ec-LDP-Hr-AE均可通过EGFR和HER2的介导进入细胞质中发挥作用,并且与单靶点的Ec-LDP和LDP-Hr蛋白相比,双特异性的Ec-LDP-Hr蛋白表现出更高的内吞效率。强化融合蛋白对人卵巢癌SK-OV-3裸鼠移植瘤的治疗结果显示,Ec-LDP-Hr-AE蛋白可显著的抑制移植瘤的生长,在0.3、0.4和0.5 mg/kg的剂量下,其抑瘤率分别为64.7%、79.5%和91.3%,其中0.5 mg/kg剂量组与LDM处理组(抑瘤率为65.6%)有显著性差异。在相同剂量下,双特异性Ec-LDP-Hr-AE蛋白同样表现出了比单靶点的Ec-LDP-AE、LDP-Hr-AE蛋白更强的抑制肿瘤生长的作用,三种强化融合蛋白的抑瘤率分别为:90.2%、83.9%和80.3%。结论:本研究成功构建并表达了一种含有EGFR和HER2寡肽配体以及力达霉素的双特异性强化融合蛋白Ec-LDP-Hr-AE,该融合蛋白可特异地与肿瘤细胞表面的EGFR和HER2受体结合,并对体外培养的不同肿瘤细胞系具有强烈杀伤作用,在体内对人卵巢癌裸鼠移植瘤有很强的生长抑制作用。Ec-LDP-Hr-AE蛋白具有双靶点、小型化和高效化的特点,具有发展成肿瘤靶向性治疗药物的潜能。
Background and Objective:The epidermal growth factor receptor EGFR/HER1 and HER2/neu are members of receptor tyrosine kinase family.Overexpression of EGFR and HER2 has been observed in a variety of human tumors,making these receptors promising targets for directed tumor therapy.Lidamycin(LDM) is an enediyne antibiotic with highly potent antitumor activity.This study was to construct a bispecific fusion protein consisting of EGFR and HER2 peptide ligands and lidamycin,and investigate its antitumor efficacy.Methods:Polymerase chain reaction and DNA cloning techniques were used to construct the gene of fusion protein.The DNA sequences of pelB signal peptide,peptide ligand Ec,(GGGGS)_2-linker and peptide ligand Hr were all synthesized and used as PCR primers.After five rounds of PCR and DNA cloning,the gene coding for fusion protein Ec-LDP-Hr was successfully constructed and was inserted into the expression vector pET30a(+) under an isopropyl-β-thiogalactopyranoside inducible promoter.Genes coding for monospecific fusion protein Ec-LDP and LDP-Hr were created by the same way.Three recombinant plasmids were transformed into E.coli strain BL21(DE3) star~(TM) and the fusion proteins were expressed after the IPTG induction. Proteins from periplasmic space of E.coli were prepared with osmotic shock method and the fusion proteins were purified by Ni~(2+) affinity chromatography.SDS-PAGE and HPLC were used to analyze the purity of fusion proteins and Western blot was used to verify the fusion proteins.The binding affinity of fusion proteins to EGFR and HER2 on the membrane of different cancer cell lines was analyzed by ELISA,immunofluorescent cytochemical staining,FCM-based immunofluorescence detection,competitive immunofluorescence assay and co-immunoprecipitation assay.The energized fusion proteins Ec-LDP-Hr-AE,Ec-LDP-AE and LDP-Hr-AE were prepared by integrating the active enediyne chromophore(AE) of lidamycin into the Ec-LDP-Hr,Fc-LDP and LDP-Hr protein,respectively.Immunoftuorescent staining and FCM-based immunofluorescence detection were used to analyze the internalization efficiency of fusion proteins.The cytotoxicity of energized fusion proteins was measured by MTT assay,and the in vivo efficacy of growth inhibition by energized fusion proteins was evaluated with human ovarian carcinoma SK-OV-3 xenografls model.Results:The genes coding for fusion proteins Ec-LDP-Hr,Ec-LDP,and LDP-Hr were successfully constructed and the encoded proteins were expressed in the periplasmic space of E.coli. Fusion proteins were purified by Ni~(2+) affinity chromatography and the purity of fusion proteins was all over 95%as determined by HPLC.The production of Ec-LDP-Hr, Ec-LDP and LDP-Hr was 9 mg,18 mg and 60 mg per liter fermentation broth, respectively.ELISA and FCM-based immunofluorescence assay revealed that Ec-LDP-Hr protein has strong binding activity to cancer cell lines highly expressing EGFR and HER2,such as A431,SK-BR-3,SK-OV-3 and MCF-7 cells.However, Ec-LDP-Hr has no binding activity to EGFR and HER2 negative NIH 3T3 cell.The results of imrnunofluorescent cytochemical staining showed that Ec-LDP-Hr protein bound to the receptors of A431 and SK-BR-3 cells,and the co-immunoprecipitation assay proved that Ec-LDP-Hr protein specifically binds to the EGF receptor and HER2. The inhibition on the binding of anti-EGFR and anti-HER2 mAbs with their antigens by Ec-LDP-Hr protein obtained by competitive immunofluorescence assay provided new evidence that Ec-LDP-Hr binds to EGFR and HER2 with high affinity.Internalization assay indicated that Ec-LDP-Hr was taken up by SK-OV-3 cells.Furthermore, Ec-LDP-Hr protein was more effectively taken up by SK-OV-3 and SK-BR-3 cells than the monospecific fusion proteins.The bispecific energized fusion protein Ec-LDP-Hr-AE showed potent cytotoxicity to A431,SK-BR-3,SK-OV-3,MCF-7,and NIH 3T3 cells with IC_(50) value of 4.25×10~(-13)mol/L,4.04×10~(-14)mol/L,1.55×10~(-13)mol/L,4.7×10~(-12)mol/L, and 2.33×10~(-11)mol/L,respectively.Ec-LDP-Hr-AE shows more potent cytotoxicity to EGFR and HER2 overexpressing cells(e.g.A431,SK-BR-3,SK-OV-3,MCF-7 cells), however,it is less cytotoxic to EGFR and HER2 negative NIH 3T3 cells compared with LDM and monospecific energized fusion proteins.Results of in vivo efficacy study have shown that the growth of established tumors was significantly inhibited when treated with Ec-LDP-Hr-AE.On day 30,Ec-LDP-Hr-AE at the dose of 0.3mg/kg,0.4mg/kg and 0.5mg/kg inhibited the growth of ovarian carcinoma SK-OV-3 xenografts by 64.7%, 79.5%and 91.3%,respectively.The group treated with 0.5 mg/kg Ec-LDP-Hr-AE showed significant differences with group treated with 0.05 mg/kg LDM which inhibited the xenografts by 65.6%.The antitumor activity of bispecific Ec-LDP-Hr-AE is more remarkable than that of monospecific energized fusion proteins Ec-LDP-AE and LDP-Hr-AE at the same dose,and these three energized fusion proteins inhibited the xenografts growth by 90.2%,83.9%,and 80.3%,respectively.Conclusion:A bispecific energized fusion protein Ec-LDP-Hr-AE was successfully constructed and expressed in E.coli.Ec-LDP-Hr-AE protein binds to EGF receptor and HER2 specifically and shows more potent cytotoxicity to different kinds of tumor cell lines overexpressing EGFR and HER2,as compared with corresponding monospecific energized fusion proteins.In addition,Ec-LDP-Hr-AE has remarkable antitumor activity in human ovarian carcinoma xenografts nude mice model.These properties,together with its two-receptor targeting and much smaller size,suggested that Ec-LDP-Hr-AE would be a promising candidate for cancer-targeting therapy.
方法:双特异性融合蛋白Ec-LDP-Hr的构建主要采用PCR扩增和DNA克隆技术。pelB信号肽序列、寡肽配体Ec基因序列、寡肽配体Hr基因序列以及(GGGGS)_2连接肽基因序列均采用人工合成的方法得到,并作为PCR引物,经过5轮PCR扩增以及DNA克隆后,得到了融合蛋白基因Ec-ldp-Hr,将该基因经过双酶切后插入到pET30a(+)表达载体中,得到重组表达载体pET-Ec-ldp-Hr。为了进行比较,本研究同时构建了单靶点的重组表达载体pET-Ec-ldp和pET-ldp-Hr。将三种表达载体均转化到大肠杆菌BL21(DE3)star~(TM)中,加入IPTG进行诱导表达,SDS-PAGE分析目的蛋白的表达情况。融合蛋白的提取采用渗透压震惊法,使用Ni~(2+)亲和层析法分离纯化融合蛋白,HPLC检测融合蛋白的纯度,Western Blot对纯化出的融合蛋白进行鉴定。融合蛋白与肿瘤细胞表面EGFR和HER2的亲和活性主要采用了ELISA、细胞免疫荧光化学法、基于流式细胞术的免疫荧光试验、竞争性免疫荧光试验以及免疫共沉淀技术进行分析。将LDM活性发色团分子(AE)与融合蛋白在体外进行分子组装,得到强化融合蛋白(Ec-LDP-Hr-AE),采用MTT法测定强化融合蛋白对肿瘤细胞的体外杀伤活性。细胞免疫荧光染色和基于流式细胞术的荧光检测用于分析融合蛋白是否可内吞进入细胞以及检测内吞效率。双特异性强化融合蛋白Ec-LDP-Hr-AE的体内抗肿瘤活性采用人卵巢癌SK-OV-3裸鼠移植瘤模型进行评价。结果:经过5轮PCR扩增以及DNA克隆后,得到了融合蛋白基因Ec-ldp-Hr,将其插入到表达载体后成功实现了在大肠杆菌中的表达。Ec-LDP-Hr蛋白和Ec-LDP蛋白主要以可溶性的形式分泌到了大肠杆菌周质腔中,而LDP-Hr蛋白除存在于周质腔中外,还分泌到了培养液上清中。Ni~(2+)亲和层析对周质腔中的蛋白进行纯化,得到的目的蛋白经HPLC检测纯度均在95%以上。Ec-LDP-Hr、Ec-LDP和LDP-Hr三种融合蛋白的产量分别为每升发酵液得到9 mg、18 mg和60 mg活性蛋白。将纯化的蛋白用Western blot方法进行鉴定,证实了纯化的蛋白为带有His-tag标签的目的蛋白。ELISA和基于流式细胞术的免疫荧光试验结果显示Ec-LDP-Hr蛋白对EGFR和HER2高表达的肿瘤细胞系,如A431、SK-BR-3、SK-OV-3、MCF-7均有很强的结合活性,而对不表达EGFR和HER2的NIH 3T3细胞则无亲和活性。细胞免疫荧光化学试验证实了Ec-LDP-Hr蛋白可以与A431和SK-BR-3细胞表面的受体结合,免疫共沉淀试验则进一步证明了Ec-LDP-Hr蛋白是与A431和SK-BR-3细胞表面的EGFR和HER2结合的;竞争性免疫荧光试验结果显示,Ec-LDP-Hr蛋白可部分抑制抗EGFR和抗HER2抗体与各自抗原的结合能力,表明该蛋白可竞争性的与EGFR和HER2结合。将AE分子与融合蛋白在体外进行分子组装后,经HPLC检测在350nm处出现了特定的吸收峰,表明AE分子与融合蛋白进行了成功组装,得到了强化融合蛋白Ec-LDP-Hr-AE。MTT试验结果显示强化融合蛋白Ec-LDP-Hr-AE对不同的肿瘤细胞系均表现出了强烈的杀伤作用,对A431、SK-BR-3、SK-OV-3、MCF-7和NIH 3T3细胞的IC_(50)值分别为:4.25×10~(-13)mol/L、4.04×10~(-14)mol/L、1.55×10~(-13)mol/L、4.7×10~(-12)mol/L和2.33×10~(-11)mol/L。对于高表达EGFR和HER2的肿瘤细胞系,双特异性强化融合蛋白Ec-LDP-Hr-AE表现出了比LDM和单靶点的强化融合蛋白Ec-LDP-AE和LDP-Hr-AE更强的杀伤活性,而对于EGFR和HER2不表达的NIH 3T3细胞,Ec-LDP-Hr-AE蛋白的IC_(50)值则高于LDM和Ec-LDP-AE、LDP-Hr-AE蛋白,表明双特异性的Ec-LDP-Hr-AE蛋白提高了对肿瘤细胞作用的特异性。细胞内吞试验结果表明,Ec-LDP-Hr和Ec-LDP-Hr-AE均可通过EGFR和HER2的介导进入细胞质中发挥作用,并且与单靶点的Ec-LDP和LDP-Hr蛋白相比,双特异性的Ec-LDP-Hr蛋白表现出更高的内吞效率。强化融合蛋白对人卵巢癌SK-OV-3裸鼠移植瘤的治疗结果显示,Ec-LDP-Hr-AE蛋白可显著的抑制移植瘤的生长,在0.3、0.4和0.5 mg/kg的剂量下,其抑瘤率分别为64.7%、79.5%和91.3%,其中0.5 mg/kg剂量组与LDM处理组(抑瘤率为65.6%)有显著性差异。在相同剂量下,双特异性Ec-LDP-Hr-AE蛋白同样表现出了比单靶点的Ec-LDP-AE、LDP-Hr-AE蛋白更强的抑制肿瘤生长的作用,三种强化融合蛋白的抑瘤率分别为:90.2%、83.9%和80.3%。结论:本研究成功构建并表达了一种含有EGFR和HER2寡肽配体以及力达霉素的双特异性强化融合蛋白Ec-LDP-Hr-AE,该融合蛋白可特异地与肿瘤细胞表面的EGFR和HER2受体结合,并对体外培养的不同肿瘤细胞系具有强烈杀伤作用,在体内对人卵巢癌裸鼠移植瘤有很强的生长抑制作用。Ec-LDP-Hr-AE蛋白具有双靶点、小型化和高效化的特点,具有发展成肿瘤靶向性治疗药物的潜能。
Background and Objective:The epidermal growth factor receptor EGFR/HER1 and HER2/neu are members of receptor tyrosine kinase family.Overexpression of EGFR and HER2 has been observed in a variety of human tumors,making these receptors promising targets for directed tumor therapy.Lidamycin(LDM) is an enediyne antibiotic with highly potent antitumor activity.This study was to construct a bispecific fusion protein consisting of EGFR and HER2 peptide ligands and lidamycin,and investigate its antitumor efficacy.Methods:Polymerase chain reaction and DNA cloning techniques were used to construct the gene of fusion protein.The DNA sequences of pelB signal peptide,peptide ligand Ec,(GGGGS)_2-linker and peptide ligand Hr were all synthesized and used as PCR primers.After five rounds of PCR and DNA cloning,the gene coding for fusion protein Ec-LDP-Hr was successfully constructed and was inserted into the expression vector pET30a(+) under an isopropyl-β-thiogalactopyranoside inducible promoter.Genes coding for monospecific fusion protein Ec-LDP and LDP-Hr were created by the same way.Three recombinant plasmids were transformed into E.coli strain BL21(DE3) star~(TM) and the fusion proteins were expressed after the IPTG induction. Proteins from periplasmic space of E.coli were prepared with osmotic shock method and the fusion proteins were purified by Ni~(2+) affinity chromatography.SDS-PAGE and HPLC were used to analyze the purity of fusion proteins and Western blot was used to verify the fusion proteins.The binding affinity of fusion proteins to EGFR and HER2 on the membrane of different cancer cell lines was analyzed by ELISA,immunofluorescent cytochemical staining,FCM-based immunofluorescence detection,competitive immunofluorescence assay and co-immunoprecipitation assay.The energized fusion proteins Ec-LDP-Hr-AE,Ec-LDP-AE and LDP-Hr-AE were prepared by integrating the active enediyne chromophore(AE) of lidamycin into the Ec-LDP-Hr,Fc-LDP and LDP-Hr protein,respectively.Immunoftuorescent staining and FCM-based immunofluorescence detection were used to analyze the internalization efficiency of fusion proteins.The cytotoxicity of energized fusion proteins was measured by MTT assay,and the in vivo efficacy of growth inhibition by energized fusion proteins was evaluated with human ovarian carcinoma SK-OV-3 xenografls model.Results:The genes coding for fusion proteins Ec-LDP-Hr,Ec-LDP,and LDP-Hr were successfully constructed and the encoded proteins were expressed in the periplasmic space of E.coli. Fusion proteins were purified by Ni~(2+) affinity chromatography and the purity of fusion proteins was all over 95%as determined by HPLC.The production of Ec-LDP-Hr, Ec-LDP and LDP-Hr was 9 mg,18 mg and 60 mg per liter fermentation broth, respectively.ELISA and FCM-based immunofluorescence assay revealed that Ec-LDP-Hr protein has strong binding activity to cancer cell lines highly expressing EGFR and HER2,such as A431,SK-BR-3,SK-OV-3 and MCF-7 cells.However, Ec-LDP-Hr has no binding activity to EGFR and HER2 negative NIH 3T3 cell.The results of imrnunofluorescent cytochemical staining showed that Ec-LDP-Hr protein bound to the receptors of A431 and SK-BR-3 cells,and the co-immunoprecipitation assay proved that Ec-LDP-Hr protein specifically binds to the EGF receptor and HER2. The inhibition on the binding of anti-EGFR and anti-HER2 mAbs with their antigens by Ec-LDP-Hr protein obtained by competitive immunofluorescence assay provided new evidence that Ec-LDP-Hr binds to EGFR and HER2 with high affinity.Internalization assay indicated that Ec-LDP-Hr was taken up by SK-OV-3 cells.Furthermore, Ec-LDP-Hr protein was more effectively taken up by SK-OV-3 and SK-BR-3 cells than the monospecific fusion proteins.The bispecific energized fusion protein Ec-LDP-Hr-AE showed potent cytotoxicity to A431,SK-BR-3,SK-OV-3,MCF-7,and NIH 3T3 cells with IC_(50) value of 4.25×10~(-13)mol/L,4.04×10~(-14)mol/L,1.55×10~(-13)mol/L,4.7×10~(-12)mol/L, and 2.33×10~(-11)mol/L,respectively.Ec-LDP-Hr-AE shows more potent cytotoxicity to EGFR and HER2 overexpressing cells(e.g.A431,SK-BR-3,SK-OV-3,MCF-7 cells), however,it is less cytotoxic to EGFR and HER2 negative NIH 3T3 cells compared with LDM and monospecific energized fusion proteins.Results of in vivo efficacy study have shown that the growth of established tumors was significantly inhibited when treated with Ec-LDP-Hr-AE.On day 30,Ec-LDP-Hr-AE at the dose of 0.3mg/kg,0.4mg/kg and 0.5mg/kg inhibited the growth of ovarian carcinoma SK-OV-3 xenografts by 64.7%, 79.5%and 91.3%,respectively.The group treated with 0.5 mg/kg Ec-LDP-Hr-AE showed significant differences with group treated with 0.05 mg/kg LDM which inhibited the xenografts by 65.6%.The antitumor activity of bispecific Ec-LDP-Hr-AE is more remarkable than that of monospecific energized fusion proteins Ec-LDP-AE and LDP-Hr-AE at the same dose,and these three energized fusion proteins inhibited the xenografts growth by 90.2%,83.9%,and 80.3%,respectively.Conclusion:A bispecific energized fusion protein Ec-LDP-Hr-AE was successfully constructed and expressed in E.coli.Ec-LDP-Hr-AE protein binds to EGF receptor and HER2 specifically and shows more potent cytotoxicity to different kinds of tumor cell lines overexpressing EGFR and HER2,as compared with corresponding monospecific energized fusion proteins.In addition,Ec-LDP-Hr-AE has remarkable antitumor activity in human ovarian carcinoma xenografts nude mice model.These properties,together with its two-receptor targeting and much smaller size,suggested that Ec-LDP-Hr-AE would be a promising candidate for cancer-targeting therapy.
引文
[1]Olayioye MA,Neve BM,Lane HA,et al.The ErbB signaling network:receptor heterodimerization in development and cancer[J].EMBO J,2000,19(13):3159-67
[2]Baselga J,Arteaga CL.Critical update and emerging trends in epidermal growth factor receptor targeting in cancer[J].J Clin Oncol,2005,23(11):2445-59
[3]Yarden Y,Sliwowski MX.Untangling the ErbB signaling network[J].Nat Rev Mol Cell Biol,2001,2(2):127-37
[4]Reid A,Vidal L,Shaw H,et al.Dual inhibition of ErbB 1(EGFR/HER1) and ErbB2 (HER2/neu)[J].Euro J Cancer,2007,43:481-89
[5]Rowinsky EK.The erbB family:targets for therapeutic development agadnst cancer and therapeutic strategies using monoclonal antibody and tyrosine kinase inhibitors[J].Annu Rev Med,2004,55:433-57
[6]Hynes NE,Lane HA.ERBB receptors and cancer:the complexity of targeted inhibitors[J].Nat Rev Cancer,2005,5(5):341-54
[7]Slamon DJ,Clark GM,Wong SG,et al.Human breast cancer:correlation of relapse and survival with amplification of the HER-2/neu oncogene[J].Science,1987,235(4785):177-82
[8]Slamon DJ,Godolphin W,Jones LA,et al.Studies of the HER-2/neu proto-oncogene in human breast and ovarian cancer[J].Science,1989,244(4905):707-12
[9]Toikkanen S,Helin H,Isola J,et al.Prognostic significance of HER-2 oncoprotein expression in breast cancer:a 30-year follow-up[J].J Clin Oncol,1992,10(7):1044-8
[10]Paez JG,Janne PA,Lee JC,et al.EGFR mutations in lung cancer:correlation with clinical response to gefitinib therapy[J].Science,2004,304(5676):1497-500
[11]Lynch TJ,Bell DW,Sordella R,et al.Activation mutations in the epidermal growth factor receptor underlying responsiveness of non-small-cell lung cancer to gefitinib[J].New Engl J Med,2004,350(21):2129-39
[12]Normanno N,De Luca A,Bianco C,et al.Epidermal growth factor receptor(EGFR) signaling in cancer[J].Gene,2006,366(1):2-16
[13]Kalyankrishna S,Grandis JR.Epidermal growth factor receptor biology in head and neck cancer[J].J Clin Oncol,2006,24(17):2666-72
[14]Argyriou A,Kalofonos HP.Recent advances relating to the clinical application of naked monoclonal antibodies in solid tumor[J].Mol Med,2009[Epub ahead of print]
[15]Kalofonos HP,Grivas PD.Monoclonal antibodies in the management of solid tumors[J].Curr Top Med Chem,2006,6(16):1687-705
[16]Yan L,Hsu K,Beckman RA.Antibody-based therapy for solid tumors[J].Cancer J,2008,14(3):178-83
[17]Wakeling AE.Epidermal growth factor receptor tyrosine kinase inhibitors[J].Curr Opin Pharmacol,2002,2(4):382-7
[18]Noonberg SB,Benz CC.Tyrosine kinase inhibitors targeted to the epidermal growth factor receptor subfamily:role as anticancer agents[J].Drugs,2000,59(4):753-67
[19]Harari PM.Epidermal growth factor receptor inhibition strategies in oncology[J].Endocr Relat Cancer,2004,11(4):689-708
[20]Goldstein NI,Prewett M,Zuklys K,et al.Biological efficacy of a chimeric antibody to the epidermal growth factor receptor in a human tumor xenograft model[J].Clin Cancer Res,1995,1(11):1311-8
[21]Harding J,Burtness B.Cetuximab:an epidermal growth factor receptor chimeric human-murine monoclonal antibody[J].Drugs Today(Bare),2005,41(2):107-27
[22]Ciardiello F,De Vita F,Orditura M,et al.Cetuximab in the treatment of colorectal cancer[J].Future Oncol,2005,1(2):173-81
[23]Cerea G,Ricotta R,Schiavetto I,et al.Cetuximab for treatment of metastatic colorectal cancer[J].Ann Oncol,2006,17(Suppl.7):vii66-7
[24]Lilenbaum RC.The evolving role of cetuximab in non-small cell lung cancer[J].Clin Cancer Res,2006,12(14 Pt 2):4432s-5s
[25]Burtness B,Goldwasser MA,Flood W,et al.Phase Ⅲ randomized trial of cisplatin plus placebo compared with cisplatin plus cetuximab in metastatic/recurrent head and neck cancer:an Eastern Cooperative Oncology Group study[J].J Clin Oncol,2005,23(34):8646-54
[26]Nahta R,Esteva FJ.Herceptin:mechanisms of action and resistance[J].Cancer Lett,2006,232(2):123-38
[27]Mariani G,Fasolo A,De Benedictis E,et al.Trastuzumab as adjuvant systemic therapy for HER2-positive breast cancer[J].Nat Clin Pract Oncol,2009,6(2):93-104
[28]Morrison C,Zanagnolo V,Ramirez N,et al.HER-2 is an independent prognostic factor in endometrial cancer:association with outcome in a large cohort of surgically staged patients[J].J Clin Oncol,2006,24(15):2376-85
[29]Metro G,Finocchiaro G,Toschi L,et al.Epidermal growth factor receptor(EGFR) targeted therapies in non-small cell lung cancer(NSCLC)[J].Rev Recent Clin Trials,2006,1(1):1-13
[30]Helfrich BA,Raben D,Varella-Garcia M,et al.Antitumor activity of the epidermal growth factor receptor(EGFR) tyrosine kinase inhibitor gefitinib(ZD1839,Iressa) in non-small cell lung cancer cell lines correlates with gene copy number and EGFR mutations but not EGFR protein levels[J].Clin Cancer Res,2006,12(23):117-25
[31]Chang GC,Yu CT,Tsai CH,et al.An epidermal growth factor receptor inhibitor,gefitinib,induces apoptosis thorough a p53-dependent upregulation of pro-apoptotic molecules and downregulation of anti-apoptotic molecules in human lung adenocarcinoma A549 cells[J].Eur J Pharmacol,2008,600(1-3):37-44
[32]Mendelson J,Baselga J.Epidermal growth factor receptor targeting in cancer[J].Semin Oncol,2006,33(4):369-85
[33]Giaccone G.Targeting HER1/EGFR in cancer therapy:experience with erlotinib[J].Future Oncol,2005,1(4):449-60
[34]Kokai Y,Myers JN,Wada T,et al.Synergistic interaction of p185c-neu and the EGF receptor leads to transformation of rodent fibroblasts[J].Cell,1989,58(2):287-92
[35]Motoyama AB,Hynes NE,Lane HA.The efficacy of ErbB receptor-targeted anticancer therapeutics is influenced by the availability of epidermal growth factor-related peptides[J].Cancer Res,2002,62(11):3151-8
[36]Xia W,Lau YK,Zhang HZ,et al.Combination of EGFR,HER-2/neu,and HER-3 is a strong predictor for the outcome of oral squamous cell carcinoma than any individual family members[J].Clin Cancer Res,1999,5(12):4164-74
[37]Tateishi M,Ishida T,Kohdono S,et al.Prognostic influence of the co-expression of epidermal growth factor receptor and c-erbB-2 protein in human lung adenocarcinoma[J].Surg Oncol,1994,3(2):109-13
[38]Britten CD.Targeting ErbB receptor signaling:a pan-ErbB approach to cancer[J].Mol Cancer Ther,2004,3(10)1335-42
[39]Brabender J,Danenberg KD,Metzger R,et al.Epidermal growth factor receptor and HER2-neu mRNA expression in non-small cell lung cancer is correlated with survival[J],Clin Cancer Res,2001,7:1850-5
[40]Ye D,Mendelsohn J,Fan Z.Augmentation of a humanized anti-HER2 mAb 4D5 induced growth inhibition by a human-mouse chimeric anti-EGF receptor mAb C225[J].Oncogene,1999,18(3):731-8
[41]Moulder SL,Yakes FM,Muthuswamy SK,et al.Epidennal growth factor receptor (HER1) tyrosine kinase inhibitor ZD18239(Iressa) inhibits HER2/neu (erbB2)-overexpressing breast cancer cells in vitro and in vivo[J].Cancer Res,2001,61(24):8887-95
[42]Moasser MM,Basso A,Averbuch SD,et al.The tyrosine kinase inhibitor ZD18239 (“Iressa”) inhibits HER2-driven signaling and suppresses the growth of HER2-overexpressing tumor cells[J].Cancer Res,2001,61(19):7184-8
[43]Normanno N,Campiglio M,De LA,et al.Cooperative inhibitory effect of ZD18239 (Iressa) in combination with trastuzumab(Herceptin) on human breast cancer cell growth[J].Ann Oncol,2002,13(1):65-72
[44]Schmidt M,Hynes NE,Groner B,et al.A bivalent single-chain antibody-toxinspecific for ErbB-2 and the EGF receptor[J].Int J Cancer,1996,65:538-546
[45]Finn RS,Wilson CA,Chen J,et al.Biologic effects of CP-724,714,a selective HER-2/neu kinase inhibitor,on human breast cancer cells with variable expression of EGFR and HER-2[A].In AACR,Mar,2004,2004:1052
[46]Ritter CA,Bianco R.HER2-overexpressing human breast cancer cells selected for Herceptin(trastuzumab) resistance in vivo retain HER2 gene amplification and overexpress HER1/EGF receptor ligands[A].In:Proceeding of the AACR,vol.44,2003,R4896
[47]Ye D,Mendelsohn J,Fan Z.Androgen and epidermal growth factor down-regulate cyclin-dependent kinase inhibitor p27kipl and costimulate proliferation of MDA PCa 2a and MDA PCa 2b prostate cancer cells[J].Clin Cancer Res,1999,5(8):2171-7
[48]Agus DB,Akita RW,Fox WD,et al.Targeting ligand-activated ErbB2 signaling inhibits breast and prostate tumor growth[J].Cancer Cell,2002,2(2):127-37
[49]Schaefer G,Fitzpatrick VD,Sliwkowski MX.Gamma-heregulin:a novel heregulin isoform that is an autocrine growth factor for the human breast cancer cell line,MDA-MB-175[J].Oncogene,1997,15(12):1385-94
[50]Mendoza N,Phillips GL,Silva J,et al.Inhibition of ligand-mediated HER2 activation in androgen-independent prostate cancer[J].Cancer Res,2002,62(19):5485-8
[51]Takai N,Jain A,Kawamata N,et al.2C4,a monoclonal antibody against HER2,disrupt the HER kinase signaling pathway and inhibits ovarian carcinoma cell growth[J]Cancer,2005,104(12):2701-8
[52]Jackson JG,St Clair P,Sliwkowski MX,et al.Blockade of epidermal growth factoror heregulin-dependent ErbB2 activation with the anti-ErbB2 monoclonal antibody 2C4 has divergent downstream signaling and growth effects[J].Cancer Res,2004,64(7):2601-9
[53]Pohl M,Striker I,Schoeneck A,et al.Antitumor activity of the HER2 dimerization inhibitor pertuzumab on human colon cancer cells in vitro and in vivo[J].J Cancer Res Clin Oncol,2009,[Epub ahead of print]
[54]Rusnak DW,Lackey K,Affleck K,et al.The effects of the novel,reversible epidermal growth factor receptor/ErbB-2 tyrosine kinase inhibitor,GW2016,on the growth of human normal and tumor-derived cell lines in vitro and in vivo[J].Mol Cancer Ther,2001,1(2):85-94
[55]Rusnak DW,Affieck K,Cockerill SG,et al.The characterization of novel,dual ErbB-2/EGFR,tyrosine kinase inhibitors:potential therapy for cancer[J].Cancer Res,2001,61(19):7196-203
[56]Xia W,Mullin RJ,Keith BR,et al.Anti-tumor activity of GW572016:a dual tyrosine kianse inhibitor blocks EGF activation of EGFR/erbB2 and downstream Erk1/2 and AKT pathways[J].Oncogene,2002,21(41):6255-63
[57]Lewis N,Marshall J,Amelsberg A,et al.A phase Ⅰ dose escalation study of BIBW 2992,an irreversible dual EGFR/HER2 receptor tyrosine kinase inhibitor,in a 3week on 1 week off schedule in patients with advanced solid tumors[J].J Clin Oncol,2006,24(June 20 suppl):3091
[58]Mom CH,Eskens FA,Gietema JA,et al.Phase Ⅰ study with BIBW 2992,an irreversible dual tyrosine kinase inhibitor of epidermal growth factor receptor 1(EGFR) and 2(HER2) in a 2 week on 2 week off schedule,2006 AS,CO annual meeting proceedings Part Ⅰ[J].J Clin Oncol,2006,24(June 20 suppl):3025
[59]Tsou HR,Overbeek-Klumpers EG,Hallett WA,et al.Optimization of 6,7-disubstituted-4-(arylamino) quinoline-3-carbonitriles as orally active,irreversible inhibitors of human epidermal growth factor receptor-2 kinase activity [J].J Med Chem,2005,48(4):1107-31
[60]Kwak EL,Sordella R,Bell DW,et al.Irreversible inhibitors of the EGF receptor may circumvent acquired resistance to gefitinib[J].Proc Natl Acad Sci USA,2005,102(21):7665-70
[61]Rabindran SK.Antitumor activity of HER-2 inhibitors[J].Cancer Lett,2005,227(1):9-23
[62]Ogiso H,Ishitani R,Nureki O,et al.Crystal structure of the complex of human epidermal growth factor and receptor extracellular domains[J].Cell,2002,110(6):775-87
[63]方华圣,张琳,卢圣栋,等.表皮生长因子介导的基因转移载体蛋白的制备[J].中国医学科学院学报,2002,24(4):381-384
[64]van de Poll ML,van vugt MJ,Lenferink AE,et al.Indentification of the minimal requirement for binding to the human epidermal growth factor(EGF) receptor using chimeras of human EGF and EGF repeat of Drosophila Notch[J].J Biol Chem,1998,273(26):16075-81
[65]Liu TF,Cohen KA,Ramage JG,et al.A diphtheria toxin-epidermal growth factor fusion protein is cytotoxic to human glioblastoma multiforme cells[J].Cancer Res, 2003,63(8):1834-7
[66]Pai LH,Gallo MG,FitzGerald DJ,et al.Antitumor activity of a transforming growth factor α-pseudomonas exotoxin fusion protein(TGF-α-PE40)[J].Cancer Res,1991,51(11):2808-12
[67]Chandler LA,Sosnowski BA,McDonald JR,et al.Targeting tumor cells via EGF receptors:selective toxicity of an HBEGF-toxin fusion protein[J].Int J Cancer,1998,78(1):106-11
[68]Azemar M,Schmidt M,Arlt F,et al.Recombinant antibody toxins specific for ErbB2 and EGF receptor inhibit the in vitro growth of human head and neck cancer cells and cause rapid tumor regression in vivo[J].Int J Cancer,2000,86(2):269-75
[69]Li ZH,Zhao R J,Wu XH,et al.Identification and characterization of a novel peptide ligand of epidermal growth factor receptor for targeted delivery of therapeutics[J].FASEB J,2005,19(14):1978-85
[70]Schier R,Marks JD,Wolf EJ,et al.In vitro an in vivo characterization of picomolar affinity anti-c-erbB-2 single-chain Fv isolated from a filamentous phage antibody library[J].Immunotechnology,1995,1(1):73-81
[71]Schier R,McCall A,Adams GP,et al.Isolation of high-affinity monomeric human anti-c-erbB-2 single chain Fv by molecular evolution of the complementarity determining regions in the center of the antibody binding site[J].J Mol Biol,1996,263(4):551-67
[72]Shirai H,Kidera A,Nakamura H.H3-rules:identification of CDR-H3 structures in antibodies[J].FEBS Lett,1999,455(1-2):188-97
[73]Kuroda D,Shirai H,Kobori M,et al.Structural classification of CDR-H3 revisited:a lesson in antibody modeling[J].Proteins,2008,73(3):608-20
[74]Qiu XQ,Wang H,Cai B,et al.Small antibody mimetics comprising two complementarity-determining regions and a framework region for tumor targeting [J].Nat Biotechnol,2007,25(8):921-9
[75]王晓良.应用分子药理学[M].北京:中国协和医科大学出版社,2005:279
[76]Shao RG,Zhen YS.Enediyne anticancer antibiotic lidamycin:chemistry,biology and pharmacology[J].Anticancer Agents Med Chem,2008,8(2):123-31
[77]甄永苏,薛玉川,邵荣光.烯二炔类新抗生素C-1027的抗肿瘤作用研究[J].中国抗生素杂志,1994,19:164-8
[78]Xu YJ,Zhen YS,Goldberg I.C-1027 chromphore,a potent new enediyne antitumor antibiotic,induces sequence-specific double-strand DNA cleavage[J].Biochemistry,1994,33:5947-54
[79]宋旭,包明敏,崔大鹏,等.抗肿瘤抗生素C-1027对人结肠癌HCT-8细胞凋 亡相关基因表达的影响[J].药学学报,1999,34(10):734-8
[80]崔大鹏,王真,李电东.力达霉素对人结肠癌HCT-8细胞侵袭调节基因的影响[J].药学学报,2001,36(4):246-9
[81]甄红英,薛玉川,甄永苏.抗肿瘤抗生素C-1027抑制血管生成及其抗肿瘤转移作用[J].中华医学杂志,1997,77:657-60
[82]丁健.抗肿瘤药物的研究趋势和进展——分子靶向药物[A].见:第二届抗肿瘤靶向药物研究与应用研讨会论文集[C].深圳:2008:19-28
[83]沈倍奋.靶向EGF受体家族成员的抗体药物[A].见:第二届抗肿瘤靶向药物研究与应用研讨会论文集[C].深圳:2008:9
[84]甄永苏.抗肿瘤药物研究与开发[M].北京:化学工业出版社,2004:381
[85]甄永苏.抗肿瘤靶向药物研究:抗体药物与配体寡肽药物[A].见:第二届抗肿瘤靶向药物研究与应用研讨会论文集[C].深圳:2008:1-8
[86]Konecny GE,Pegram MD,Nenkatesan N,et al.Activity of the dual kinase inhibitor lapatinib(GW572016) against HER-2-overexpressing and trastuzumab-treated breast cancer cells[J].Cancer Res,2006,66(3):1630-39
[87]Todhunter DA,Hall WA,Rustamzadeh E,et al.A bispecific immunotoxin(DTAT13)targeting human IL-13 receptor(IL-13R) and urokinase-type plasminogen activator receptor(uPAR) in a mouse xenograft model[J].Protein Eng Des Sel,2004,17(2):157-64
[88]Stish BJ,Chen H,Shu YQ,et al.Increasing anticarcinoma activity of an anti-erbB2recombinant immunotoxin by the addition of an anti-EpCAM sFv[J].Clin Cancer Res,2007,13(10):3058-67
[89]Stish BJ,Chen H,Shu YQ,et al.A bispecific recombinant cytotoxin(DTEGF13)targeting human interleukin-13 and epidermal growth factor receptors in a mouse xenografts model of prostate cancer[J].Clin Cancer Res,2007,13(21):6486-93
[90]Vallera DA,Todhunter DA,Kuroki DW,et al.A bispecific recombinant immunotoxin,DT2219,targeting human CD19 and CD22 receptors in a mouse xenograft model of B cell leukemia/lymphoma[J].Clin Cancer Res,2005,11(10):3879-88
[91]Rusnak DW,Affieck K,Cockerill SG,et al.The characterization of novel,dual ErbB-2/EGFR,tyrosine kinase inhibitors:potential therapy for cancer[J].Cancer Res,2001,61(19):7196-203
[92]Rathore D,Nayak SK,Batra JK,et al.Expression of ribonucleolytic toxin restriction in Escherichia coli:purification and characterization[J].FEBS Letters,1996,392(3):259-62
v[93]Bera TK,Onda M,Brinkmann U,et al.A bivalent disulfide stabilized Fv with
improved antigen binding to erbB2[J].J Mol Biol,1998,281(3):475-83
[94]Schmidt M,Hynes NE,Groner B,et al.A bivalent single-chain antibody-toxin specific for ErbB-2 and EGF receptor[J].Int J Cancer,1996,65(4):538-46
[95]Stoscheck CM,Carpenter G."Down-regulation" of EGF receptors:direct demonstration of receptor degradation in human fibroblasts[J].J Cell Biol,1984,98(3):1048-53
[96]Waterman H,Yarden Y.Molecular mechanisms underlying endocytosis and sorting of ErbB receptor tyrosine kinases[J].FEBS Lett,2001,490(3):142-52
[97]Sorkin A,Goh LK.Endocytosis and intracellular trafficking of ErbBs[J].Exp Cell Res,2008,314(17):3093-106
[98]Mori T.Cancer-specific ligands identified from screening of peptide-display libraries[J].Curr Pharm Des,2004,10(19):2335-2343
[99]Friedman PN,Chace DE Trail PA,et al.Antitumor activity of the single-chain immunotoxin BR96 sFv-PE40 against established breast and lung tumor xenografls [J].J Immunol,1993,150(7):3054-61
[100]甄永苏,李顺强,江敏.抗癌抗生素力达霉素与单链抗体的组装融合蛋白[P].中国专利:CNl251840,2000-05-03
[101]Shaw JP,Akiyoshi DE,Arrigo DA,et al.Cytotoxic properties of DAB486EGF and DAB389EGF,epidermal growth factor(EGF) receptor-targeted fusion toxins[J].J Biol Chem,1991,266(31):21118-24
[102]Kihara A,Pastan I.Small chimeric toxins containing only transforming growth factor α and domain Ⅲ of Pseudomonas exotoxin with good antitumor activity in mice[J].Cancer Res,1994,54(19):5154-9
[103]Miao QF,Liu XY,Shang BY,et al.An enediyne-energized single-domain antibody-containing fusion protein shows potent antitumor activity[J].Anti-cancer drugs,2007,18(2):127-37
[104]苗庆芳,尚伯杨,欧阳志刚,等.单域抗体与力达霉素的基因工程强化融合蛋白VL-LDP-AE的制备及其抗肿瘤作用[J].中国科学C辑,2007,37(2):143-51
[105]Berezov A,Zhang HT,Greene MI,et al.Disabling erbB receptors with rationally designed exocyclic mimetics of antibodies:structure-function analysis[J].J Med Chem,2001,44(16):2565-74
[106]Afshar S,Asai T,Morrison SL.Humanized ADEPT comprised of an engineered human purine nucleoside phosphorylase and a tumor targeting peptide for treatment of cancer[J].Mol Cancer Ther,2009,8(1):185-93
[1]Lacey Jr JV,Devesa SS,Brinton LA.Recent trends in breast cancer incidence and mortality[J].Environ Mol Mutagen,2002,39:82-88.
[2]Stoff-Khalili MA,Dall P,Curiel DT.Gene therapy for carcinoma of the breast.Cancer Gene Therapy 2006;13:633-647.
[3]Osborne RJ,Hamshere MG.A genome-wide map showing common regions of loss of heterozygosity/allelic imbalance in breast cancer.Cancer Res,2000;60:3706-3712.
[4]Miller BJ,Wang D,Krahe R,et al.Pooled analysis of loss of heterozygosity in breast cancer:a genome scan provides comparative evidence for multiple tumor suppressors and identifies novel candidate regions.Am J Hum Genet.2003,73:748-767
[5]Knudson Jr AG.Mutation and cancer:statistical study of retinoblastoma.Proc Natl Acad Sci USA,1971,68:820-23
[6]Johannsdottir HK,Johannesdottir G,Agnarsson BA,et al.Deletions on chromosome 4 in sporadic and BRCA mutated tumors and association with pathological variables.Anticancer Res,2004;24:2681-2687.
[7]Cleton-Jansen AM,Buerger H,Haar N,et al.Different mechanisms of chromosome 16 loss of heterozygosity in well- versus poorly differentiated ductal breast cancer.Genes Chromosomes Cancer,2004;41:109-116.
[8]Osborne C,Wilson P,Tripathy D.Oncogenes and tumor suppressor genes in breast cancer:potential diagnostic and therapeutic applications.Oncologist 2004;9:361-377.
[9]Miki Y,Swensen J,Shattuck-Eidens D,et al.A strong candidate for the breast and ovarian cancer susceptibility gene BRCA1.Science,1994,266:66-71
[10]Hall JM,Lee MK,Newman B,et al.Linkage of early-onset familial breast cancer to chromosome 17q21.Science,1990,250:1684-1689
[11]Phelan CM,Lancaster JM,Tonin P,et al.Mutation analysis of the BRCA2 gene in 49 site-specific breast cancer families.Nat Genet,1996,13:120-122
[12]de Jong MM,Nolte IM,te Meerman GJ,van der Graaf WT,Oosterwijk JC,Kleibeuker JH et al.Genes other than BRCA1 and BRCA2 involved in breast cancer susceptibility.J Med Genet,2002;39:225-242.
[13]Lancaster JM,Wooster R,Mangion J,et al.BBCA2 mutations in primary breast and ovarian cancers.Nat Genet,1996,13:238-240
[14]Lacroix M,Leclercq G.The“portrait”of hereditary breast cancer.Breast Cancer Res Treat,2005,89:297-304
[15]Deng CX,Brodie SG.Roles of BRCA1 and its interacting proteins.BioEssays,2000,22:728-737
[16]Lee WH,Bookstein R,Hong F,et al.Human retinoblastoma susceptibility gene:cloning,identification,and sequence.Science,1987,235:1394-1399
[17]Andersen TI,Gaustad A,Ottestad L,et al.Genetic alterations of the tumour suppressor gene regions 3p,11p,13q,17p,and 17q in human breast carcinomas.Genes Chromosomes Cancer,1992,4:113-121.
[18]Takahashi T,Nau MM,Chiba I,et al.p53:a frequent target for genetic abnormalities in lung cancer.Science,1989,246:491-494.
[19]Malkin D.Germline p53 mutations and heritable cancer.Annu Rev Genet,1994,28:443-465
[20]Hollstein M,Sidransky D,Vogelstein B,et al.p53 mutations in human cancers.Science,1991,253:49-53
[21]Bland KI.The 1999 James Ewing lecture:in pursuit of molecules of oncogenesis and neoplastic therapy.Ann Surg Oncol,1999,6:528-541
[22]King CR,Kraus MH,Aaronson SA.Amplification of a novel v-erbB-related gene in a human mammary carcinoma.Science,1985,229:974-976
[23]Funkhouser WK,Kaiser-Rogers K.Review:significance of,and optimal screening for,HER-2 gene amplification and protein overexpression in breast carcinoma.Ann Clin Lab Sci,2001,31:349-358.
[24]Nass SJ,Dickson RB.Defining a role for c-Myc in breast tumorigenesis.Breast Cancer Res Treat,1997,44:1-22.
[25]Varley JM,Brammer WJ.Alterations to either c-erbB2(neu) or c-myc proto-oncogenes in breast carcinoma correlate with poor short-term prognosis.Oncogene,1987,1:423-430
[26]Boveri T.The origin of malignant tumors.Williams and Wilkins:Baltimore,MD,1929
[27]Jiang Z,Zacksenhaus E.Activation of retinoblastoma protein in mammary gland leads to ductal growth suppression,precocious differentiation,and adenocarcinoma.J Cell Biol,2002,156:185-198.
[28]Campbell I,Magliocco A,Moyana T,et al.Adenovirus-mediated p16INK4 gene transfer significantly suppresses human breast cancer growth.Cancer GeneTher,2000,7:1270-1278
[29]Craig C,Wersto R,Kim M,et al.A recombinant adenovirus expressing p27Kipl induces cell cycle arrest and loss of cyclin-Cdk activity in human breast cancer cells.Oncogene,1997,14:2283-2289
[30]Shibata MA,Yoshidome K,Shibata E,et al.Suppression of mammary carcinoma growth in vitro and in vivo by inducible expression of Cdk inhibitor p21.Cancer Gene Ther,2001,8:23-25
[31]Wang NP,To H,Lee WH,et al.Tumor suppressor activity of RB and p53 genes in human breast carcinoma cells.Oncogene,1993,8:279-288.
[32]Sauane M,Gopalkrishnan RV,Choo HT,et al.Mechanistic aspects of mda-7/IL-24 cancer cell selectivity analyzed via a bacterial fusion protein.Oncogene,2004,23:7679-7690
[33]Holt JT,Thompson ME,Szabo C,et al.Growth retardation and tumor inhibition by BBCA1.Nat Genet,1996,12:298-302
[34]Holt JT.Breast cancer genes:therapeutic strategies.Ann NY Acad Sci 1997,833:34-41
[35]Shi HY,Liang R,Templeton NS,et al.Inhibition of breast tumor progression by systemic delivery of the maspin gene in a syngeneic tumor model.Mol Ther,2002,5:755-761
[36]Sarti M,Sevignani C,Calin GA,et al.Adenoviral transduction of TESTIN gene into breast and uterine cancer cell lines promotes apoptosis and tumor reduction in vivo.Clin Cancer Res,2005,11:806-813
[37]Xu M,Kumar D,Srinivas S,et al.Parenteral gene therapy with p53 inhibits human breast tumors in vivo through a bystander mechanism without evidence of toxicity.Hum Gne Ther,1997,8:177-185
[38]Reed JC,Miyashita T,Krajewski S,et al.Bcl-2 family proteins and the regulation of programmed cell death in leukemia and lymphoma.Cancer Treat Res,1996,84:31-72
[39]Owen-Schaub LB,Zhang W,Cusack JC,et al.Wild-type human p53 and a temperature-sensitive mutant induce Fas/APO-1 expression.Mol Cell Biol,1995,15:3032-3040
[40]Molinier-Frenkel V,Le Boulaire C,Le Gal FA,et al.Longitudinal follow-up of cellular and humoral immunity induced by recombinant adenovirus-mediated gene therapy in cancer patients.Hum Gene Ther,2000,11:1911-1920
[41]McCormick F.Cancer gene therapy:fringe or cutting edge? Nat Rev Cancer,2001,1:130-141
[42]Nielsen LL,Dell J,Maxwell E,et al.Efficacy of p53 adenovirus-mediated gene therapy against human breast cancer xenografts.Cancer Gene Ther,1997,4:129-138
[43]Dias N,Stein CA.Antisense oligonucleotides:basic concepts and mechanisms.Mol Cancer Ther,2002,1:347-355.
[44]McManaway ME,Neckers LM,Loke SL,et al.Tumor-specific inhibition of lymphoma growth by an antisense oligodeoxynucleotide.Lancet,1990,335:808-811
[45]Townsend PA,Villanova I,Uhlmann E,et al.An antisense oligonucleotide targeting the alpha V integrin gene inhibits adhesion and induces apoptosis in breast cancer cells.Eur J Cancer,2000,36:397-409
[46]Gardner-Thorpe J,Ito H,Ashley SW,et al.Ribosomal protein P2:a potential molecular target for antisense therapy of human malignancies.Anticancer Res,2003,23:4549-4560
[47]Brysch W,Magal E,Louis JC,et al.Inhibition of p185~(c-erbB-2) proto-oncogene expression by antisense oligodeoxynucleotides down-regulates p185-associated tyrosine-kinase activity and strongly inhibits mammary tumor-cell proliferation.Cancer Gene Ther,1994,1:90-105
[48]Sekhon J,Pereira P,Sabbaghian N,et al.Antisense inhibition of methylenetetrahydrofolate reductase reduces survival of methionine-dependent tumor lines.Br J Cancer,2002,87:225-230
[49]Fan Y,Borowsky AD,Weiss RH,et al.An antisense oligodeoxynucleotide to p21 (Waf1/Cip1) causes apoptosis in human breast cancer cells.Mol Cancer Ther,2003,2:773-782
[50]Roychowdhury D,Lahn M.Antisense therapy directed to protein kianse C-alpha (Affinitak,LY900003/ISIS 3521):potential role in breast cancer.Semin Oncol,2003,30:30-33
[51]Nahta R,Esteva FJ.Bcl-2 antisense oligonucleotides:a potential novel strategy for the treatment of breast cancer.Semin Oncol,2003,30:143-149
[52]Natarajan S,Chen Z,Wancewicz EV,et al.Telomerase reverse transcriptase(hTERT) mRNA and telomerase RNA(hTR) as targets for downregulation of telomerase activity.Oligonucleotides,2004,14:263-273
[53]Lee WJ,Robinson JA,Holman NA,et al.Antisense mediated inhibiton of the plasma membrane calcium-ATPase suppresses proliferation of MCF-7 cells.J Biol Chem 2005,280:27076-27084
[54]Salatino M,Schillaci R,Proietti CJ,et al.Inhibition of in vivo breast cancer growth by antisense oligonucleotides to type I insulin-like growth factor receptor mRNA involves inactivation of ErbBs,PI-3K/Akt and p42/p44 MAPK signaling pathways but not modulation of progesterone receptor activity.Oncogene,2004,23: 5161-5174
[55]Arteaga CL,Holt JT.Tissue-targeted antisense c-fos retroviral vector inhibits established breast cancer xenografts in nude mice.Cancer Res,1996,56:1098-1103
[56]Scherr M,Morgan MA,Eder M.Gene silencing mediated by small interfering RNAs in mammalian cells.Curr Med Chem,2003,10:245-256.
[57]Wang YH,Liu S,Zhang G,et al.Knockdown of c-myc expression by RNAi inhibits MCF-7 breast tumor cells growth in vitro and in vivo.Breast Caner Res,2005,7:R220-R228
[58]Rossi JJ.Ribozymes.Curr Opin Biotechnol,1992,3:3-7
[59]Citti L,Rainaldi G.Synthetic hammerhead ribozymes as therapeutic tools to control disease genes.Curr Gene Ther,2005,5:11-24
[60]Suzuki T,Anderegg B,Ohkawa T,Irie A,Engebraaten O,Halks-Miller M et al.Adenovirus-mediated ribozyme targeting of HER-2/neu inhibits in vivo growth of breast cancer cells.Gene Therapy,2000,7:241-248.
[61]Choi KS,Lee TH,Jung MH.Ribozyme-mediated cleavage of the human survivin mRNA and inhibition of antiapoptotic function of surviving in MCF-7 cells.Cancer Gene Ther,2003,10:87-95
[62]Iyo M,Kawasaki H,Taira K.Construction of an allosteric trans-maxizyme targeting for two distinct oncogenes.Nucleic Acids Res,2002,suppl 2:115-116
[63]Wright M,Grim J,Deshane J,et al.An intracellular anti-erbB-2 single-chain antibody is specifically cytotoxic to human breast carcinoma cells overexpressing erbB-2.Gene Therapy,1997,4:317-322
[64]Ueno NT,Yu D,Hung MC.E1A:tumor suppressor or oncogene? Preclinical and clinical investigations of E1A gene therapy.Breast Cancer,2001,8:285-293.
[65]Niculescu-Duvaz I,Springer CJ.Introduction to the background,principles,and state of the art in suicide gene therapy.Mol Biotechnol,2005,30:71-88.
[66]Chen SH,Chen XH,Wang Y,et al.Combination gene therapy for liver metastasis of colon carcinoma in vivo.Proc Natl Acad Sci USA,1995,92:2577-2581
[67]Kammertoens T,Gelbmann W,Karle P,et al.Combined chemotherapy of murine mammary tumors by local activation of the prodrugs ifofamide and 5-fluorocytosine.Cancer Gene Ther,2000,7:629-636
[68]Jaattela M.Escaping cell death:survival proteins in cancer.Exp Cell Res,1999,248:30-43
[69]Lin J,Page C,Jin X,et al.Suppression activity of pro-apoptotic gene products in cancer cells,a potential application for cancer gene therapy.Anticancer Res,2001,21:831-839
[70]Tai YT,Strobel T,Kufe D,et al.In vivo cytotoxicity of ovarian cancer cells through tumor-selective expression of the BAX gene.Cancer Res,1999,59:2121-2126
[71]Lin T,Huang X,Gu J,et al.Long-term tumor-free survival from treatment with the GFP-TRAIL fusion gene expressed from the hTERT promoter in breast cancer cells.Oncogene,2002,21:8020-8028
[72]Sanlioglu AD,Dirice E,Aydin C,et al.Surface TRAIL decoy receptor-4 expression is correlated with TRAIL resistance in MCF-7 breast cancer cells.BMC Cancer,2005,5:54
[73]Nagata S,Golstein P.The Fas death factor.Science,1995,267:1449-1456.
[74]Nylandsted J,Rohde M,Brand K,Bastholm L,Elling F,Jaattela M.Selective depletion of heat shock protein 70(Hsp70) activates a tumor-specific death program that is independent of caspases and bypasses Bcl-2.Proc Natl Acad Sci USA,2000,97:7871-7876.
[75]Hung MC,Hortobagyi GN,Ueno NT.Development of clinical trial of E1A gene therapy targeting HER-2/neu-overexpressing breast and ovarian caner.Adv Exp Med Biol,2000,465:171-180
[76]Davidoff AM,Nathwani AC.Antiangiogenic gene therapy for cancer treatment.Curr Hematol Rep,2004,3:267-273
[77]Chen QR,Kumar D,Stass SA,et al.Liposomes complexed to plasmids encoding angiostatin and endostatininhibit breast cancer in nude mice.Cancer Res,1999,59:3308-3312.
[78]Strong TV.Gene therapy for carcinoma of the breast:genetic immunotherapy.Breast Cancer Res,2000,2:15-21.
[79]Siegel JP,Puri RK.Interleukin-2 toxicity.J Clin Oncol,1991,9:694-704
[80]Coveney E,Clary B,lacobucci M,et al.Active immunotherapy with transiently transfected cytokine-secreting tumor cells inhibits breast caner metastases in tumor-bearing animals.Surgery,1996,129:265-272
[81]Lichtor T,Glick RP,Lin H,et al.Intratumoral injection of IL-sereting syngeneic/allogeneic fibroblasts transfected with DNA from breast cancer cells prolongs the survival of mice with intracerebral breast cancer.Cancer Gene Ther,2005,12:708-714
[82]Knuth A,Wolfel T,Meyer zum Buschenfelde KH.Cellular and humoral immune responses against cancer:implications for cancer vaccines.Curr Opin Immunol,1991,3:659-664
[83]Yip YL,Ward RL.Anti-ErbB-2 monoclonal antibodies and ErbB-2-directed vaccines.Cancer Immunol Immunother,2002,50:569-587.
[84]Chang SY,Lee KC,Ko SY,et al.Enhanced efficacy of DNA vaccination against Her-2/neu tumor antigen by genetic adjuvants.Int J Cancer,2004,111:86-95
[85]von Mehren M,Arlen P,Gulley J,et al.The influence of granulocyte macrophage colony-stimulating factor and prior chemotherapy on the immunological response to a vaccine(ALVAC-CEA B7.1) in patients with metastatic carcinoma.Clin Cancer Res,2001,7:1181-1191
[86]Thery C,Amigorena S.The cell biology of antigen presentation in dendritic cells.Curr Opin Immunol,2001,13:45-51.
[87]Sakai Y,Morrison BJ,Burke JD,et al.Vaccination by genetically modified dendritic cells expressing a truncated neu oncogene prevents development of breast cancer in transgenic mice.Cancer Res,2004,64:8022-8028
[88]Viehl CT,Tanaka Y,Chen T,et al.Tat mammaglobin fusion protein transduced dendritic cells stimulate mammaglobin-specific CD4 and CD8 t cells.Breast Cancer Res Treat,2005,91:271-178
[89]Hudson PJ.Recombinant antibodies:a novel approach to cancer diagnosis and therapy.Expert Opin Investig Drugs,2000,9:1231-1242.
[90]Larin SS,Georgiev GP,Kiselev SL.Gene transfer approaches in cancer immunotherapy.Gene Therapy,2004,11(suppl 1):S18-S25
[91]万福强 乳腺癌基因治疗新进展及展望 中华临床医学实践杂志 2005;6:484-486
[92]付晶鑫 乳腺癌基因治疗研究进展 使用肿瘤杂志 2007 22(2):183-186
[93]王深明,林颖 乳腺癌基因治疗进展 中华外科杂志 2006 44(1):55-58
[94]王深明,殷恒讳 乳腺癌基因治疗的现状与存在的问题 中华医学杂志 2005 85(1)3-7
[2]Baselga J,Arteaga CL.Critical update and emerging trends in epidermal growth factor receptor targeting in cancer[J].J Clin Oncol,2005,23(11):2445-59
[3]Yarden Y,Sliwowski MX.Untangling the ErbB signaling network[J].Nat Rev Mol Cell Biol,2001,2(2):127-37
[4]Reid A,Vidal L,Shaw H,et al.Dual inhibition of ErbB 1(EGFR/HER1) and ErbB2 (HER2/neu)[J].Euro J Cancer,2007,43:481-89
[5]Rowinsky EK.The erbB family:targets for therapeutic development agadnst cancer and therapeutic strategies using monoclonal antibody and tyrosine kinase inhibitors[J].Annu Rev Med,2004,55:433-57
[6]Hynes NE,Lane HA.ERBB receptors and cancer:the complexity of targeted inhibitors[J].Nat Rev Cancer,2005,5(5):341-54
[7]Slamon DJ,Clark GM,Wong SG,et al.Human breast cancer:correlation of relapse and survival with amplification of the HER-2/neu oncogene[J].Science,1987,235(4785):177-82
[8]Slamon DJ,Godolphin W,Jones LA,et al.Studies of the HER-2/neu proto-oncogene in human breast and ovarian cancer[J].Science,1989,244(4905):707-12
[9]Toikkanen S,Helin H,Isola J,et al.Prognostic significance of HER-2 oncoprotein expression in breast cancer:a 30-year follow-up[J].J Clin Oncol,1992,10(7):1044-8
[10]Paez JG,Janne PA,Lee JC,et al.EGFR mutations in lung cancer:correlation with clinical response to gefitinib therapy[J].Science,2004,304(5676):1497-500
[11]Lynch TJ,Bell DW,Sordella R,et al.Activation mutations in the epidermal growth factor receptor underlying responsiveness of non-small-cell lung cancer to gefitinib[J].New Engl J Med,2004,350(21):2129-39
[12]Normanno N,De Luca A,Bianco C,et al.Epidermal growth factor receptor(EGFR) signaling in cancer[J].Gene,2006,366(1):2-16
[13]Kalyankrishna S,Grandis JR.Epidermal growth factor receptor biology in head and neck cancer[J].J Clin Oncol,2006,24(17):2666-72
[14]Argyriou A,Kalofonos HP.Recent advances relating to the clinical application of naked monoclonal antibodies in solid tumor[J].Mol Med,2009[Epub ahead of print]
[15]Kalofonos HP,Grivas PD.Monoclonal antibodies in the management of solid tumors[J].Curr Top Med Chem,2006,6(16):1687-705
[16]Yan L,Hsu K,Beckman RA.Antibody-based therapy for solid tumors[J].Cancer J,2008,14(3):178-83
[17]Wakeling AE.Epidermal growth factor receptor tyrosine kinase inhibitors[J].Curr Opin Pharmacol,2002,2(4):382-7
[18]Noonberg SB,Benz CC.Tyrosine kinase inhibitors targeted to the epidermal growth factor receptor subfamily:role as anticancer agents[J].Drugs,2000,59(4):753-67
[19]Harari PM.Epidermal growth factor receptor inhibition strategies in oncology[J].Endocr Relat Cancer,2004,11(4):689-708
[20]Goldstein NI,Prewett M,Zuklys K,et al.Biological efficacy of a chimeric antibody to the epidermal growth factor receptor in a human tumor xenograft model[J].Clin Cancer Res,1995,1(11):1311-8
[21]Harding J,Burtness B.Cetuximab:an epidermal growth factor receptor chimeric human-murine monoclonal antibody[J].Drugs Today(Bare),2005,41(2):107-27
[22]Ciardiello F,De Vita F,Orditura M,et al.Cetuximab in the treatment of colorectal cancer[J].Future Oncol,2005,1(2):173-81
[23]Cerea G,Ricotta R,Schiavetto I,et al.Cetuximab for treatment of metastatic colorectal cancer[J].Ann Oncol,2006,17(Suppl.7):vii66-7
[24]Lilenbaum RC.The evolving role of cetuximab in non-small cell lung cancer[J].Clin Cancer Res,2006,12(14 Pt 2):4432s-5s
[25]Burtness B,Goldwasser MA,Flood W,et al.Phase Ⅲ randomized trial of cisplatin plus placebo compared with cisplatin plus cetuximab in metastatic/recurrent head and neck cancer:an Eastern Cooperative Oncology Group study[J].J Clin Oncol,2005,23(34):8646-54
[26]Nahta R,Esteva FJ.Herceptin:mechanisms of action and resistance[J].Cancer Lett,2006,232(2):123-38
[27]Mariani G,Fasolo A,De Benedictis E,et al.Trastuzumab as adjuvant systemic therapy for HER2-positive breast cancer[J].Nat Clin Pract Oncol,2009,6(2):93-104
[28]Morrison C,Zanagnolo V,Ramirez N,et al.HER-2 is an independent prognostic factor in endometrial cancer:association with outcome in a large cohort of surgically staged patients[J].J Clin Oncol,2006,24(15):2376-85
[29]Metro G,Finocchiaro G,Toschi L,et al.Epidermal growth factor receptor(EGFR) targeted therapies in non-small cell lung cancer(NSCLC)[J].Rev Recent Clin Trials,2006,1(1):1-13
[30]Helfrich BA,Raben D,Varella-Garcia M,et al.Antitumor activity of the epidermal growth factor receptor(EGFR) tyrosine kinase inhibitor gefitinib(ZD1839,Iressa) in non-small cell lung cancer cell lines correlates with gene copy number and EGFR mutations but not EGFR protein levels[J].Clin Cancer Res,2006,12(23):117-25
[31]Chang GC,Yu CT,Tsai CH,et al.An epidermal growth factor receptor inhibitor,gefitinib,induces apoptosis thorough a p53-dependent upregulation of pro-apoptotic molecules and downregulation of anti-apoptotic molecules in human lung adenocarcinoma A549 cells[J].Eur J Pharmacol,2008,600(1-3):37-44
[32]Mendelson J,Baselga J.Epidermal growth factor receptor targeting in cancer[J].Semin Oncol,2006,33(4):369-85
[33]Giaccone G.Targeting HER1/EGFR in cancer therapy:experience with erlotinib[J].Future Oncol,2005,1(4):449-60
[34]Kokai Y,Myers JN,Wada T,et al.Synergistic interaction of p185c-neu and the EGF receptor leads to transformation of rodent fibroblasts[J].Cell,1989,58(2):287-92
[35]Motoyama AB,Hynes NE,Lane HA.The efficacy of ErbB receptor-targeted anticancer therapeutics is influenced by the availability of epidermal growth factor-related peptides[J].Cancer Res,2002,62(11):3151-8
[36]Xia W,Lau YK,Zhang HZ,et al.Combination of EGFR,HER-2/neu,and HER-3 is a strong predictor for the outcome of oral squamous cell carcinoma than any individual family members[J].Clin Cancer Res,1999,5(12):4164-74
[37]Tateishi M,Ishida T,Kohdono S,et al.Prognostic influence of the co-expression of epidermal growth factor receptor and c-erbB-2 protein in human lung adenocarcinoma[J].Surg Oncol,1994,3(2):109-13
[38]Britten CD.Targeting ErbB receptor signaling:a pan-ErbB approach to cancer[J].Mol Cancer Ther,2004,3(10)1335-42
[39]Brabender J,Danenberg KD,Metzger R,et al.Epidermal growth factor receptor and HER2-neu mRNA expression in non-small cell lung cancer is correlated with survival[J],Clin Cancer Res,2001,7:1850-5
[40]Ye D,Mendelsohn J,Fan Z.Augmentation of a humanized anti-HER2 mAb 4D5 induced growth inhibition by a human-mouse chimeric anti-EGF receptor mAb C225[J].Oncogene,1999,18(3):731-8
[41]Moulder SL,Yakes FM,Muthuswamy SK,et al.Epidennal growth factor receptor (HER1) tyrosine kinase inhibitor ZD18239(Iressa) inhibits HER2/neu (erbB2)-overexpressing breast cancer cells in vitro and in vivo[J].Cancer Res,2001,61(24):8887-95
[42]Moasser MM,Basso A,Averbuch SD,et al.The tyrosine kinase inhibitor ZD18239 (“Iressa”) inhibits HER2-driven signaling and suppresses the growth of HER2-overexpressing tumor cells[J].Cancer Res,2001,61(19):7184-8
[43]Normanno N,Campiglio M,De LA,et al.Cooperative inhibitory effect of ZD18239 (Iressa) in combination with trastuzumab(Herceptin) on human breast cancer cell growth[J].Ann Oncol,2002,13(1):65-72
[44]Schmidt M,Hynes NE,Groner B,et al.A bivalent single-chain antibody-toxinspecific for ErbB-2 and the EGF receptor[J].Int J Cancer,1996,65:538-546
[45]Finn RS,Wilson CA,Chen J,et al.Biologic effects of CP-724,714,a selective HER-2/neu kinase inhibitor,on human breast cancer cells with variable expression of EGFR and HER-2[A].In AACR,Mar,2004,2004:1052
[46]Ritter CA,Bianco R.HER2-overexpressing human breast cancer cells selected for Herceptin(trastuzumab) resistance in vivo retain HER2 gene amplification and overexpress HER1/EGF receptor ligands[A].In:Proceeding of the AACR,vol.44,2003,R4896
[47]Ye D,Mendelsohn J,Fan Z.Androgen and epidermal growth factor down-regulate cyclin-dependent kinase inhibitor p27kipl and costimulate proliferation of MDA PCa 2a and MDA PCa 2b prostate cancer cells[J].Clin Cancer Res,1999,5(8):2171-7
[48]Agus DB,Akita RW,Fox WD,et al.Targeting ligand-activated ErbB2 signaling inhibits breast and prostate tumor growth[J].Cancer Cell,2002,2(2):127-37
[49]Schaefer G,Fitzpatrick VD,Sliwkowski MX.Gamma-heregulin:a novel heregulin isoform that is an autocrine growth factor for the human breast cancer cell line,MDA-MB-175[J].Oncogene,1997,15(12):1385-94
[50]Mendoza N,Phillips GL,Silva J,et al.Inhibition of ligand-mediated HER2 activation in androgen-independent prostate cancer[J].Cancer Res,2002,62(19):5485-8
[51]Takai N,Jain A,Kawamata N,et al.2C4,a monoclonal antibody against HER2,disrupt the HER kinase signaling pathway and inhibits ovarian carcinoma cell growth[J]Cancer,2005,104(12):2701-8
[52]Jackson JG,St Clair P,Sliwkowski MX,et al.Blockade of epidermal growth factoror heregulin-dependent ErbB2 activation with the anti-ErbB2 monoclonal antibody 2C4 has divergent downstream signaling and growth effects[J].Cancer Res,2004,64(7):2601-9
[53]Pohl M,Striker I,Schoeneck A,et al.Antitumor activity of the HER2 dimerization inhibitor pertuzumab on human colon cancer cells in vitro and in vivo[J].J Cancer Res Clin Oncol,2009,[Epub ahead of print]
[54]Rusnak DW,Lackey K,Affleck K,et al.The effects of the novel,reversible epidermal growth factor receptor/ErbB-2 tyrosine kinase inhibitor,GW2016,on the growth of human normal and tumor-derived cell lines in vitro and in vivo[J].Mol Cancer Ther,2001,1(2):85-94
[55]Rusnak DW,Affieck K,Cockerill SG,et al.The characterization of novel,dual ErbB-2/EGFR,tyrosine kinase inhibitors:potential therapy for cancer[J].Cancer Res,2001,61(19):7196-203
[56]Xia W,Mullin RJ,Keith BR,et al.Anti-tumor activity of GW572016:a dual tyrosine kianse inhibitor blocks EGF activation of EGFR/erbB2 and downstream Erk1/2 and AKT pathways[J].Oncogene,2002,21(41):6255-63
[57]Lewis N,Marshall J,Amelsberg A,et al.A phase Ⅰ dose escalation study of BIBW 2992,an irreversible dual EGFR/HER2 receptor tyrosine kinase inhibitor,in a 3week on 1 week off schedule in patients with advanced solid tumors[J].J Clin Oncol,2006,24(June 20 suppl):3091
[58]Mom CH,Eskens FA,Gietema JA,et al.Phase Ⅰ study with BIBW 2992,an irreversible dual tyrosine kinase inhibitor of epidermal growth factor receptor 1(EGFR) and 2(HER2) in a 2 week on 2 week off schedule,2006 AS,CO annual meeting proceedings Part Ⅰ[J].J Clin Oncol,2006,24(June 20 suppl):3025
[59]Tsou HR,Overbeek-Klumpers EG,Hallett WA,et al.Optimization of 6,7-disubstituted-4-(arylamino) quinoline-3-carbonitriles as orally active,irreversible inhibitors of human epidermal growth factor receptor-2 kinase activity [J].J Med Chem,2005,48(4):1107-31
[60]Kwak EL,Sordella R,Bell DW,et al.Irreversible inhibitors of the EGF receptor may circumvent acquired resistance to gefitinib[J].Proc Natl Acad Sci USA,2005,102(21):7665-70
[61]Rabindran SK.Antitumor activity of HER-2 inhibitors[J].Cancer Lett,2005,227(1):9-23
[62]Ogiso H,Ishitani R,Nureki O,et al.Crystal structure of the complex of human epidermal growth factor and receptor extracellular domains[J].Cell,2002,110(6):775-87
[63]方华圣,张琳,卢圣栋,等.表皮生长因子介导的基因转移载体蛋白的制备[J].中国医学科学院学报,2002,24(4):381-384
[64]van de Poll ML,van vugt MJ,Lenferink AE,et al.Indentification of the minimal requirement for binding to the human epidermal growth factor(EGF) receptor using chimeras of human EGF and EGF repeat of Drosophila Notch[J].J Biol Chem,1998,273(26):16075-81
[65]Liu TF,Cohen KA,Ramage JG,et al.A diphtheria toxin-epidermal growth factor fusion protein is cytotoxic to human glioblastoma multiforme cells[J].Cancer Res, 2003,63(8):1834-7
[66]Pai LH,Gallo MG,FitzGerald DJ,et al.Antitumor activity of a transforming growth factor α-pseudomonas exotoxin fusion protein(TGF-α-PE40)[J].Cancer Res,1991,51(11):2808-12
[67]Chandler LA,Sosnowski BA,McDonald JR,et al.Targeting tumor cells via EGF receptors:selective toxicity of an HBEGF-toxin fusion protein[J].Int J Cancer,1998,78(1):106-11
[68]Azemar M,Schmidt M,Arlt F,et al.Recombinant antibody toxins specific for ErbB2 and EGF receptor inhibit the in vitro growth of human head and neck cancer cells and cause rapid tumor regression in vivo[J].Int J Cancer,2000,86(2):269-75
[69]Li ZH,Zhao R J,Wu XH,et al.Identification and characterization of a novel peptide ligand of epidermal growth factor receptor for targeted delivery of therapeutics[J].FASEB J,2005,19(14):1978-85
[70]Schier R,Marks JD,Wolf EJ,et al.In vitro an in vivo characterization of picomolar affinity anti-c-erbB-2 single-chain Fv isolated from a filamentous phage antibody library[J].Immunotechnology,1995,1(1):73-81
[71]Schier R,McCall A,Adams GP,et al.Isolation of high-affinity monomeric human anti-c-erbB-2 single chain Fv by molecular evolution of the complementarity determining regions in the center of the antibody binding site[J].J Mol Biol,1996,263(4):551-67
[72]Shirai H,Kidera A,Nakamura H.H3-rules:identification of CDR-H3 structures in antibodies[J].FEBS Lett,1999,455(1-2):188-97
[73]Kuroda D,Shirai H,Kobori M,et al.Structural classification of CDR-H3 revisited:a lesson in antibody modeling[J].Proteins,2008,73(3):608-20
[74]Qiu XQ,Wang H,Cai B,et al.Small antibody mimetics comprising two complementarity-determining regions and a framework region for tumor targeting [J].Nat Biotechnol,2007,25(8):921-9
[75]王晓良.应用分子药理学[M].北京:中国协和医科大学出版社,2005:279
[76]Shao RG,Zhen YS.Enediyne anticancer antibiotic lidamycin:chemistry,biology and pharmacology[J].Anticancer Agents Med Chem,2008,8(2):123-31
[77]甄永苏,薛玉川,邵荣光.烯二炔类新抗生素C-1027的抗肿瘤作用研究[J].中国抗生素杂志,1994,19:164-8
[78]Xu YJ,Zhen YS,Goldberg I.C-1027 chromphore,a potent new enediyne antitumor antibiotic,induces sequence-specific double-strand DNA cleavage[J].Biochemistry,1994,33:5947-54
[79]宋旭,包明敏,崔大鹏,等.抗肿瘤抗生素C-1027对人结肠癌HCT-8细胞凋 亡相关基因表达的影响[J].药学学报,1999,34(10):734-8
[80]崔大鹏,王真,李电东.力达霉素对人结肠癌HCT-8细胞侵袭调节基因的影响[J].药学学报,2001,36(4):246-9
[81]甄红英,薛玉川,甄永苏.抗肿瘤抗生素C-1027抑制血管生成及其抗肿瘤转移作用[J].中华医学杂志,1997,77:657-60
[82]丁健.抗肿瘤药物的研究趋势和进展——分子靶向药物[A].见:第二届抗肿瘤靶向药物研究与应用研讨会论文集[C].深圳:2008:19-28
[83]沈倍奋.靶向EGF受体家族成员的抗体药物[A].见:第二届抗肿瘤靶向药物研究与应用研讨会论文集[C].深圳:2008:9
[84]甄永苏.抗肿瘤药物研究与开发[M].北京:化学工业出版社,2004:381
[85]甄永苏.抗肿瘤靶向药物研究:抗体药物与配体寡肽药物[A].见:第二届抗肿瘤靶向药物研究与应用研讨会论文集[C].深圳:2008:1-8
[86]Konecny GE,Pegram MD,Nenkatesan N,et al.Activity of the dual kinase inhibitor lapatinib(GW572016) against HER-2-overexpressing and trastuzumab-treated breast cancer cells[J].Cancer Res,2006,66(3):1630-39
[87]Todhunter DA,Hall WA,Rustamzadeh E,et al.A bispecific immunotoxin(DTAT13)targeting human IL-13 receptor(IL-13R) and urokinase-type plasminogen activator receptor(uPAR) in a mouse xenograft model[J].Protein Eng Des Sel,2004,17(2):157-64
[88]Stish BJ,Chen H,Shu YQ,et al.Increasing anticarcinoma activity of an anti-erbB2recombinant immunotoxin by the addition of an anti-EpCAM sFv[J].Clin Cancer Res,2007,13(10):3058-67
[89]Stish BJ,Chen H,Shu YQ,et al.A bispecific recombinant cytotoxin(DTEGF13)targeting human interleukin-13 and epidermal growth factor receptors in a mouse xenografts model of prostate cancer[J].Clin Cancer Res,2007,13(21):6486-93
[90]Vallera DA,Todhunter DA,Kuroki DW,et al.A bispecific recombinant immunotoxin,DT2219,targeting human CD19 and CD22 receptors in a mouse xenograft model of B cell leukemia/lymphoma[J].Clin Cancer Res,2005,11(10):3879-88
[91]Rusnak DW,Affieck K,Cockerill SG,et al.The characterization of novel,dual ErbB-2/EGFR,tyrosine kinase inhibitors:potential therapy for cancer[J].Cancer Res,2001,61(19):7196-203
[92]Rathore D,Nayak SK,Batra JK,et al.Expression of ribonucleolytic toxin restriction in Escherichia coli:purification and characterization[J].FEBS Letters,1996,392(3):259-62
v[93]Bera TK,Onda M,Brinkmann U,et al.A bivalent disulfide stabilized Fv with
improved antigen binding to erbB2[J].J Mol Biol,1998,281(3):475-83
[94]Schmidt M,Hynes NE,Groner B,et al.A bivalent single-chain antibody-toxin specific for ErbB-2 and EGF receptor[J].Int J Cancer,1996,65(4):538-46
[95]Stoscheck CM,Carpenter G."Down-regulation" of EGF receptors:direct demonstration of receptor degradation in human fibroblasts[J].J Cell Biol,1984,98(3):1048-53
[96]Waterman H,Yarden Y.Molecular mechanisms underlying endocytosis and sorting of ErbB receptor tyrosine kinases[J].FEBS Lett,2001,490(3):142-52
[97]Sorkin A,Goh LK.Endocytosis and intracellular trafficking of ErbBs[J].Exp Cell Res,2008,314(17):3093-106
[98]Mori T.Cancer-specific ligands identified from screening of peptide-display libraries[J].Curr Pharm Des,2004,10(19):2335-2343
[99]Friedman PN,Chace DE Trail PA,et al.Antitumor activity of the single-chain immunotoxin BR96 sFv-PE40 against established breast and lung tumor xenografls [J].J Immunol,1993,150(7):3054-61
[100]甄永苏,李顺强,江敏.抗癌抗生素力达霉素与单链抗体的组装融合蛋白[P].中国专利:CNl251840,2000-05-03
[101]Shaw JP,Akiyoshi DE,Arrigo DA,et al.Cytotoxic properties of DAB486EGF and DAB389EGF,epidermal growth factor(EGF) receptor-targeted fusion toxins[J].J Biol Chem,1991,266(31):21118-24
[102]Kihara A,Pastan I.Small chimeric toxins containing only transforming growth factor α and domain Ⅲ of Pseudomonas exotoxin with good antitumor activity in mice[J].Cancer Res,1994,54(19):5154-9
[103]Miao QF,Liu XY,Shang BY,et al.An enediyne-energized single-domain antibody-containing fusion protein shows potent antitumor activity[J].Anti-cancer drugs,2007,18(2):127-37
[104]苗庆芳,尚伯杨,欧阳志刚,等.单域抗体与力达霉素的基因工程强化融合蛋白VL-LDP-AE的制备及其抗肿瘤作用[J].中国科学C辑,2007,37(2):143-51
[105]Berezov A,Zhang HT,Greene MI,et al.Disabling erbB receptors with rationally designed exocyclic mimetics of antibodies:structure-function analysis[J].J Med Chem,2001,44(16):2565-74
[106]Afshar S,Asai T,Morrison SL.Humanized ADEPT comprised of an engineered human purine nucleoside phosphorylase and a tumor targeting peptide for treatment of cancer[J].Mol Cancer Ther,2009,8(1):185-93
[1]Lacey Jr JV,Devesa SS,Brinton LA.Recent trends in breast cancer incidence and mortality[J].Environ Mol Mutagen,2002,39:82-88.
[2]Stoff-Khalili MA,Dall P,Curiel DT.Gene therapy for carcinoma of the breast.Cancer Gene Therapy 2006;13:633-647.
[3]Osborne RJ,Hamshere MG.A genome-wide map showing common regions of loss of heterozygosity/allelic imbalance in breast cancer.Cancer Res,2000;60:3706-3712.
[4]Miller BJ,Wang D,Krahe R,et al.Pooled analysis of loss of heterozygosity in breast cancer:a genome scan provides comparative evidence for multiple tumor suppressors and identifies novel candidate regions.Am J Hum Genet.2003,73:748-767
[5]Knudson Jr AG.Mutation and cancer:statistical study of retinoblastoma.Proc Natl Acad Sci USA,1971,68:820-23
[6]Johannsdottir HK,Johannesdottir G,Agnarsson BA,et al.Deletions on chromosome 4 in sporadic and BRCA mutated tumors and association with pathological variables.Anticancer Res,2004;24:2681-2687.
[7]Cleton-Jansen AM,Buerger H,Haar N,et al.Different mechanisms of chromosome 16 loss of heterozygosity in well- versus poorly differentiated ductal breast cancer.Genes Chromosomes Cancer,2004;41:109-116.
[8]Osborne C,Wilson P,Tripathy D.Oncogenes and tumor suppressor genes in breast cancer:potential diagnostic and therapeutic applications.Oncologist 2004;9:361-377.
[9]Miki Y,Swensen J,Shattuck-Eidens D,et al.A strong candidate for the breast and ovarian cancer susceptibility gene BRCA1.Science,1994,266:66-71
[10]Hall JM,Lee MK,Newman B,et al.Linkage of early-onset familial breast cancer to chromosome 17q21.Science,1990,250:1684-1689
[11]Phelan CM,Lancaster JM,Tonin P,et al.Mutation analysis of the BRCA2 gene in 49 site-specific breast cancer families.Nat Genet,1996,13:120-122
[12]de Jong MM,Nolte IM,te Meerman GJ,van der Graaf WT,Oosterwijk JC,Kleibeuker JH et al.Genes other than BRCA1 and BRCA2 involved in breast cancer susceptibility.J Med Genet,2002;39:225-242.
[13]Lancaster JM,Wooster R,Mangion J,et al.BBCA2 mutations in primary breast and ovarian cancers.Nat Genet,1996,13:238-240
[14]Lacroix M,Leclercq G.The“portrait”of hereditary breast cancer.Breast Cancer Res Treat,2005,89:297-304
[15]Deng CX,Brodie SG.Roles of BRCA1 and its interacting proteins.BioEssays,2000,22:728-737
[16]Lee WH,Bookstein R,Hong F,et al.Human retinoblastoma susceptibility gene:cloning,identification,and sequence.Science,1987,235:1394-1399
[17]Andersen TI,Gaustad A,Ottestad L,et al.Genetic alterations of the tumour suppressor gene regions 3p,11p,13q,17p,and 17q in human breast carcinomas.Genes Chromosomes Cancer,1992,4:113-121.
[18]Takahashi T,Nau MM,Chiba I,et al.p53:a frequent target for genetic abnormalities in lung cancer.Science,1989,246:491-494.
[19]Malkin D.Germline p53 mutations and heritable cancer.Annu Rev Genet,1994,28:443-465
[20]Hollstein M,Sidransky D,Vogelstein B,et al.p53 mutations in human cancers.Science,1991,253:49-53
[21]Bland KI.The 1999 James Ewing lecture:in pursuit of molecules of oncogenesis and neoplastic therapy.Ann Surg Oncol,1999,6:528-541
[22]King CR,Kraus MH,Aaronson SA.Amplification of a novel v-erbB-related gene in a human mammary carcinoma.Science,1985,229:974-976
[23]Funkhouser WK,Kaiser-Rogers K.Review:significance of,and optimal screening for,HER-2 gene amplification and protein overexpression in breast carcinoma.Ann Clin Lab Sci,2001,31:349-358.
[24]Nass SJ,Dickson RB.Defining a role for c-Myc in breast tumorigenesis.Breast Cancer Res Treat,1997,44:1-22.
[25]Varley JM,Brammer WJ.Alterations to either c-erbB2(neu) or c-myc proto-oncogenes in breast carcinoma correlate with poor short-term prognosis.Oncogene,1987,1:423-430
[26]Boveri T.The origin of malignant tumors.Williams and Wilkins:Baltimore,MD,1929
[27]Jiang Z,Zacksenhaus E.Activation of retinoblastoma protein in mammary gland leads to ductal growth suppression,precocious differentiation,and adenocarcinoma.J Cell Biol,2002,156:185-198.
[28]Campbell I,Magliocco A,Moyana T,et al.Adenovirus-mediated p16INK4 gene transfer significantly suppresses human breast cancer growth.Cancer GeneTher,2000,7:1270-1278
[29]Craig C,Wersto R,Kim M,et al.A recombinant adenovirus expressing p27Kipl induces cell cycle arrest and loss of cyclin-Cdk activity in human breast cancer cells.Oncogene,1997,14:2283-2289
[30]Shibata MA,Yoshidome K,Shibata E,et al.Suppression of mammary carcinoma growth in vitro and in vivo by inducible expression of Cdk inhibitor p21.Cancer Gene Ther,2001,8:23-25
[31]Wang NP,To H,Lee WH,et al.Tumor suppressor activity of RB and p53 genes in human breast carcinoma cells.Oncogene,1993,8:279-288.
[32]Sauane M,Gopalkrishnan RV,Choo HT,et al.Mechanistic aspects of mda-7/IL-24 cancer cell selectivity analyzed via a bacterial fusion protein.Oncogene,2004,23:7679-7690
[33]Holt JT,Thompson ME,Szabo C,et al.Growth retardation and tumor inhibition by BBCA1.Nat Genet,1996,12:298-302
[34]Holt JT.Breast cancer genes:therapeutic strategies.Ann NY Acad Sci 1997,833:34-41
[35]Shi HY,Liang R,Templeton NS,et al.Inhibition of breast tumor progression by systemic delivery of the maspin gene in a syngeneic tumor model.Mol Ther,2002,5:755-761
[36]Sarti M,Sevignani C,Calin GA,et al.Adenoviral transduction of TESTIN gene into breast and uterine cancer cell lines promotes apoptosis and tumor reduction in vivo.Clin Cancer Res,2005,11:806-813
[37]Xu M,Kumar D,Srinivas S,et al.Parenteral gene therapy with p53 inhibits human breast tumors in vivo through a bystander mechanism without evidence of toxicity.Hum Gne Ther,1997,8:177-185
[38]Reed JC,Miyashita T,Krajewski S,et al.Bcl-2 family proteins and the regulation of programmed cell death in leukemia and lymphoma.Cancer Treat Res,1996,84:31-72
[39]Owen-Schaub LB,Zhang W,Cusack JC,et al.Wild-type human p53 and a temperature-sensitive mutant induce Fas/APO-1 expression.Mol Cell Biol,1995,15:3032-3040
[40]Molinier-Frenkel V,Le Boulaire C,Le Gal FA,et al.Longitudinal follow-up of cellular and humoral immunity induced by recombinant adenovirus-mediated gene therapy in cancer patients.Hum Gene Ther,2000,11:1911-1920
[41]McCormick F.Cancer gene therapy:fringe or cutting edge? Nat Rev Cancer,2001,1:130-141
[42]Nielsen LL,Dell J,Maxwell E,et al.Efficacy of p53 adenovirus-mediated gene therapy against human breast cancer xenografts.Cancer Gene Ther,1997,4:129-138
[43]Dias N,Stein CA.Antisense oligonucleotides:basic concepts and mechanisms.Mol Cancer Ther,2002,1:347-355.
[44]McManaway ME,Neckers LM,Loke SL,et al.Tumor-specific inhibition of lymphoma growth by an antisense oligodeoxynucleotide.Lancet,1990,335:808-811
[45]Townsend PA,Villanova I,Uhlmann E,et al.An antisense oligonucleotide targeting the alpha V integrin gene inhibits adhesion and induces apoptosis in breast cancer cells.Eur J Cancer,2000,36:397-409
[46]Gardner-Thorpe J,Ito H,Ashley SW,et al.Ribosomal protein P2:a potential molecular target for antisense therapy of human malignancies.Anticancer Res,2003,23:4549-4560
[47]Brysch W,Magal E,Louis JC,et al.Inhibition of p185~(c-erbB-2) proto-oncogene expression by antisense oligodeoxynucleotides down-regulates p185-associated tyrosine-kinase activity and strongly inhibits mammary tumor-cell proliferation.Cancer Gene Ther,1994,1:90-105
[48]Sekhon J,Pereira P,Sabbaghian N,et al.Antisense inhibition of methylenetetrahydrofolate reductase reduces survival of methionine-dependent tumor lines.Br J Cancer,2002,87:225-230
[49]Fan Y,Borowsky AD,Weiss RH,et al.An antisense oligodeoxynucleotide to p21 (Waf1/Cip1) causes apoptosis in human breast cancer cells.Mol Cancer Ther,2003,2:773-782
[50]Roychowdhury D,Lahn M.Antisense therapy directed to protein kianse C-alpha (Affinitak,LY900003/ISIS 3521):potential role in breast cancer.Semin Oncol,2003,30:30-33
[51]Nahta R,Esteva FJ.Bcl-2 antisense oligonucleotides:a potential novel strategy for the treatment of breast cancer.Semin Oncol,2003,30:143-149
[52]Natarajan S,Chen Z,Wancewicz EV,et al.Telomerase reverse transcriptase(hTERT) mRNA and telomerase RNA(hTR) as targets for downregulation of telomerase activity.Oligonucleotides,2004,14:263-273
[53]Lee WJ,Robinson JA,Holman NA,et al.Antisense mediated inhibiton of the plasma membrane calcium-ATPase suppresses proliferation of MCF-7 cells.J Biol Chem 2005,280:27076-27084
[54]Salatino M,Schillaci R,Proietti CJ,et al.Inhibition of in vivo breast cancer growth by antisense oligonucleotides to type I insulin-like growth factor receptor mRNA involves inactivation of ErbBs,PI-3K/Akt and p42/p44 MAPK signaling pathways but not modulation of progesterone receptor activity.Oncogene,2004,23: 5161-5174
[55]Arteaga CL,Holt JT.Tissue-targeted antisense c-fos retroviral vector inhibits established breast cancer xenografts in nude mice.Cancer Res,1996,56:1098-1103
[56]Scherr M,Morgan MA,Eder M.Gene silencing mediated by small interfering RNAs in mammalian cells.Curr Med Chem,2003,10:245-256.
[57]Wang YH,Liu S,Zhang G,et al.Knockdown of c-myc expression by RNAi inhibits MCF-7 breast tumor cells growth in vitro and in vivo.Breast Caner Res,2005,7:R220-R228
[58]Rossi JJ.Ribozymes.Curr Opin Biotechnol,1992,3:3-7
[59]Citti L,Rainaldi G.Synthetic hammerhead ribozymes as therapeutic tools to control disease genes.Curr Gene Ther,2005,5:11-24
[60]Suzuki T,Anderegg B,Ohkawa T,Irie A,Engebraaten O,Halks-Miller M et al.Adenovirus-mediated ribozyme targeting of HER-2/neu inhibits in vivo growth of breast cancer cells.Gene Therapy,2000,7:241-248.
[61]Choi KS,Lee TH,Jung MH.Ribozyme-mediated cleavage of the human survivin mRNA and inhibition of antiapoptotic function of surviving in MCF-7 cells.Cancer Gene Ther,2003,10:87-95
[62]Iyo M,Kawasaki H,Taira K.Construction of an allosteric trans-maxizyme targeting for two distinct oncogenes.Nucleic Acids Res,2002,suppl 2:115-116
[63]Wright M,Grim J,Deshane J,et al.An intracellular anti-erbB-2 single-chain antibody is specifically cytotoxic to human breast carcinoma cells overexpressing erbB-2.Gene Therapy,1997,4:317-322
[64]Ueno NT,Yu D,Hung MC.E1A:tumor suppressor or oncogene? Preclinical and clinical investigations of E1A gene therapy.Breast Cancer,2001,8:285-293.
[65]Niculescu-Duvaz I,Springer CJ.Introduction to the background,principles,and state of the art in suicide gene therapy.Mol Biotechnol,2005,30:71-88.
[66]Chen SH,Chen XH,Wang Y,et al.Combination gene therapy for liver metastasis of colon carcinoma in vivo.Proc Natl Acad Sci USA,1995,92:2577-2581
[67]Kammertoens T,Gelbmann W,Karle P,et al.Combined chemotherapy of murine mammary tumors by local activation of the prodrugs ifofamide and 5-fluorocytosine.Cancer Gene Ther,2000,7:629-636
[68]Jaattela M.Escaping cell death:survival proteins in cancer.Exp Cell Res,1999,248:30-43
[69]Lin J,Page C,Jin X,et al.Suppression activity of pro-apoptotic gene products in cancer cells,a potential application for cancer gene therapy.Anticancer Res,2001,21:831-839
[70]Tai YT,Strobel T,Kufe D,et al.In vivo cytotoxicity of ovarian cancer cells through tumor-selective expression of the BAX gene.Cancer Res,1999,59:2121-2126
[71]Lin T,Huang X,Gu J,et al.Long-term tumor-free survival from treatment with the GFP-TRAIL fusion gene expressed from the hTERT promoter in breast cancer cells.Oncogene,2002,21:8020-8028
[72]Sanlioglu AD,Dirice E,Aydin C,et al.Surface TRAIL decoy receptor-4 expression is correlated with TRAIL resistance in MCF-7 breast cancer cells.BMC Cancer,2005,5:54
[73]Nagata S,Golstein P.The Fas death factor.Science,1995,267:1449-1456.
[74]Nylandsted J,Rohde M,Brand K,Bastholm L,Elling F,Jaattela M.Selective depletion of heat shock protein 70(Hsp70) activates a tumor-specific death program that is independent of caspases and bypasses Bcl-2.Proc Natl Acad Sci USA,2000,97:7871-7876.
[75]Hung MC,Hortobagyi GN,Ueno NT.Development of clinical trial of E1A gene therapy targeting HER-2/neu-overexpressing breast and ovarian caner.Adv Exp Med Biol,2000,465:171-180
[76]Davidoff AM,Nathwani AC.Antiangiogenic gene therapy for cancer treatment.Curr Hematol Rep,2004,3:267-273
[77]Chen QR,Kumar D,Stass SA,et al.Liposomes complexed to plasmids encoding angiostatin and endostatininhibit breast cancer in nude mice.Cancer Res,1999,59:3308-3312.
[78]Strong TV.Gene therapy for carcinoma of the breast:genetic immunotherapy.Breast Cancer Res,2000,2:15-21.
[79]Siegel JP,Puri RK.Interleukin-2 toxicity.J Clin Oncol,1991,9:694-704
[80]Coveney E,Clary B,lacobucci M,et al.Active immunotherapy with transiently transfected cytokine-secreting tumor cells inhibits breast caner metastases in tumor-bearing animals.Surgery,1996,129:265-272
[81]Lichtor T,Glick RP,Lin H,et al.Intratumoral injection of IL-sereting syngeneic/allogeneic fibroblasts transfected with DNA from breast cancer cells prolongs the survival of mice with intracerebral breast cancer.Cancer Gene Ther,2005,12:708-714
[82]Knuth A,Wolfel T,Meyer zum Buschenfelde KH.Cellular and humoral immune responses against cancer:implications for cancer vaccines.Curr Opin Immunol,1991,3:659-664
[83]Yip YL,Ward RL.Anti-ErbB-2 monoclonal antibodies and ErbB-2-directed vaccines.Cancer Immunol Immunother,2002,50:569-587.
[84]Chang SY,Lee KC,Ko SY,et al.Enhanced efficacy of DNA vaccination against Her-2/neu tumor antigen by genetic adjuvants.Int J Cancer,2004,111:86-95
[85]von Mehren M,Arlen P,Gulley J,et al.The influence of granulocyte macrophage colony-stimulating factor and prior chemotherapy on the immunological response to a vaccine(ALVAC-CEA B7.1) in patients with metastatic carcinoma.Clin Cancer Res,2001,7:1181-1191
[86]Thery C,Amigorena S.The cell biology of antigen presentation in dendritic cells.Curr Opin Immunol,2001,13:45-51.
[87]Sakai Y,Morrison BJ,Burke JD,et al.Vaccination by genetically modified dendritic cells expressing a truncated neu oncogene prevents development of breast cancer in transgenic mice.Cancer Res,2004,64:8022-8028
[88]Viehl CT,Tanaka Y,Chen T,et al.Tat mammaglobin fusion protein transduced dendritic cells stimulate mammaglobin-specific CD4 and CD8 t cells.Breast Cancer Res Treat,2005,91:271-178
[89]Hudson PJ.Recombinant antibodies:a novel approach to cancer diagnosis and therapy.Expert Opin Investig Drugs,2000,9:1231-1242.
[90]Larin SS,Georgiev GP,Kiselev SL.Gene transfer approaches in cancer immunotherapy.Gene Therapy,2004,11(suppl 1):S18-S25
[91]万福强 乳腺癌基因治疗新进展及展望 中华临床医学实践杂志 2005;6:484-486
[92]付晶鑫 乳腺癌基因治疗研究进展 使用肿瘤杂志 2007 22(2):183-186
[93]王深明,林颖 乳腺癌基因治疗进展 中华外科杂志 2006 44(1):55-58
[94]王深明,殷恒讳 乳腺癌基因治疗的现状与存在的问题 中华医学杂志 2005 85(1)3-7