Livin反义寡核苷酸对膀胱癌细胞抑制作用的实验研究
摘要
近年来,人们发现凋亡过程受到抑制在肿瘤的发生、发展中起到非常关键的作用。其中IAPs在细胞的凋亡中扮演非常重要的角色,某些IAPs成员异常高表达常引起组织细胞凋亡受阻,与肿瘤的发生密切相关。目前已发现的人类IAP家族有8个,包括NAIP、ILP-2、c-IAPl、c-IAP2、XIAP、Bruce、survivin及Livin。其中Livin是最近发现的IAPs家族新成员,主要在一些肿瘤组织细胞中高表达,在正常组织不表达或极少表达,可能成为肿瘤治疗的靶点。我们的前期研究在膀胱癌患者尿液中也发现有Livin mRNA的表达,而健康志愿者无表达。
目的:
在我们的前期工作基础上,我们拟利用反义核酸技术,在体、内外阻断膀胱癌细胞Livin mRNA及蛋白质的表达,观察癌细胞的生长以及凋亡是否受到影响,以明确Livin在膀胱癌细胞中的作用,并初步探讨其机制。为以Livin为靶基因的膀胱癌的治疗提供理论依据。
方法:
一.设计、合成全硫代修饰Livin反义寡核苷酸序列及其功能鉴定。
1.参考文献设计、筛选、合成全硫代修饰Livin反义寡核苷酸。同时设计一条错义序列寡核苷酸作为对照。
2.制作Livin反义寡核苷酸- Lipofectamine?2000复合物,将其转染入5637膀胱癌细胞中。
3.荧光显微镜观察转染效率。
4.用RT-PCR、Western blot、细胞免疫荧光检测转染后膀胱癌细胞Livin mRNA及蛋白表达的变化。
二.细胞分组
根据向培养细胞中加入的成分不同,分为: (1)反义组:加入反义寡核苷酸; (2)错义组:加入错义的寡核苷酸; (3)脂质体组:只加入等量稀释的脂质体; (4)PBS(磷酸缓冲液)组:只加入等量的PBS。
三.Livin反义寡核苷酸对人膀胱癌细胞株5637生物学活性的影响
1.以不同浓度的Livin反义寡核苷酸转染培养的膀胱癌细胞,用MTT法测定细胞增殖活性。从中筛选出适当的Livin反义寡核苷酸浓度,进行下一步实验。
2.Livin反义寡核苷酸对膀胱癌细胞的生物学效应
①激光共聚焦显微镜、透射电子显微镜观察膀胱癌细胞转染Livin反义寡核苷酸后的形态学变化。
②AO/EB双染色、Annexin V-FITC细胞凋亡检测试剂盒流式细胞仪检测细胞凋亡情况。
四.Livin反义寡核苷酸诱导荷瘤鼠膀胱癌组织凋亡的研究
1.裸鼠膀胱癌移植瘤模型的建立
采用皮下注射法建立裸鼠膀胱癌移植瘤模型。
2.给药方法及分组
采用瘤体内5点注射法,根据注射核酸的不同,将裸鼠随机分为对照组(瘤体内注射错义寡核苷酸)和反义组(瘤体内注射反义寡核苷酸)。
3.Livin反义寡核苷酸对荷瘤鼠肿瘤组织Livin表达的影响
用RT-PCR、Western blot检测检测转染后膀胱癌细胞Livin mRNA及蛋白表达的变化。
4.Livin反义寡核苷酸对荷瘤鼠肿瘤组织的生物学效应
①用游标卡尺测量、计算肿瘤体积,绘制癌细胞体内生长曲线。接种后第30天剥瘤称重。
②H.E染色光镜观察肿瘤组织转染Livin反义寡核苷酸后的形态学变化。
③TUNEL法检测荷瘤鼠肿瘤细胞凋亡情况。
④免疫组化检测荷瘤鼠肿瘤caspas-3的表达情况。
结果:
一.Livin反义寡核苷酸能有效转染入膀胱癌细胞内并抑制Livin mRNA及蛋白的表达。
将FITC标记的Livin ASODN转染膀胱癌细胞后,在荧光显微镜下观察,见约55%的细胞染上绿色荧光,说明Livin ASODN能有效地转染入5637细胞中。
RT-PCR扩增后显示5637细胞表达livinα和livinβ,但反义组的电泳带亮度明显低于错义组、脂质体组及PBS组,而后三组间亮度相似。反义组livin两个片段的mRNA相对表达量(0.46±0.05)较错义组(0.88±0.05)、脂质体组(0.92±0.04)、PBS组(0.91±0.06)明显减弱,P<0.01,而各对照组间比较无显著性差异,P>0.05。
采用Western blot检测各组细胞中livin蛋白的表达,结果反义组livinα和livinβ杂交带亮度明显低于错义组、脂质体组和PBS组。细胞免疫荧光染色后用激光共聚焦显微镜观察,发现反义组癌细胞标记livin的绿色荧光明显减弱,且见有的细胞体积明显缩小,完全无绿色荧光。其余各对照组无明显变化。
说明Livin反义寡核苷酸能有效抑制膀胱癌细胞Livin mRNA及蛋白的表达。
二.Livin反义寡核苷酸对人膀胱癌细胞增殖活性的影响
MTT法发现Livin反义寡核苷酸能抑制人膀胱癌细胞的增殖活性,且其抑制作用呈明显的浓度依赖性。我们据此选择160nmol/L的寡核苷酸浓度进行下一步实验。
三.Livin反义寡核苷酸对膀胱癌细胞的生物学效应
①细胞形态的变化:激光共聚焦显微镜、透射电子显微镜下见反义组可见较多细胞体积肿大、细胞器肿胀、变性坏死。有的细胞体积缩小,发生凋亡,甚至形成凋亡小体。
②细胞凋亡率的变化:AO/EB双染色、Annexin V-FITC标记凋亡细胞,流式细胞仪检测细胞凋亡率,结果发现反义组的细胞凋亡率(46.39±9.23)%显著高于PBS组(4.54±1.84)%、脂质体组(5.70±1.61)%及错义组(5.10±1.56)%,P﹤0.01。而后三组间凋亡率差异无显著意义,P﹥0.05。
三.Livin反义寡核苷酸对荷瘤鼠肿瘤组织的抑制作用
1.Livin反义寡核苷酸对荷瘤鼠肿瘤组织Livin表达的影响
RT-PCR扩增后显示,反义组荷瘤鼠肿瘤组织表达livinα和livinβ的电泳带亮度明显低于对照组。
Western blot检测荷瘤鼠肿瘤组织livin蛋白的表达,结果反义组livinα和livinβ杂交带亮度明显低于对照组。
免疫组化也显示反义组livin表达减弱(其面积积分光密度为:0.1661±0.1028,对照组为0.2490±0.1483,P﹤0.05)说明Livin反义寡核苷酸在体内也能有效抑制膀胱癌细胞Livin mRNA及蛋白的表达。
2.Livin反义寡核苷酸对对荷瘤鼠肿瘤组织的生物学效应
①肿瘤生长的变化:通过测量、计算肿瘤体积,绘制癌细胞体内生长曲线。发现从肿瘤接种后的第18天开始至接种后第30天,反义组肿瘤体积显著小于对照组,P<0.05。接种后第30天剥瘤称重,反义组瘤块重量为1.31±0.88g,明显轻于对照组(2.41±0.41g),P<0.05。说明Livin反义寡核苷酸能在体内抑制了肿瘤的生长。
②H.E染色光镜观察肿瘤组织转染Livin反义寡核苷酸后的形态学变化:可见反义组肿瘤组织有局灶变性、坏死现象及炎症细胞浸润。
③肿瘤细胞凋亡情况:TUNEL法检测荷瘤鼠肿瘤组织凋亡指数,结果发现反义组为19.60±5.91,明显高于对照组(3.48±2.35),P﹤0.05。
④免疫组化检测荷瘤鼠肿瘤caspas-3的表达情况:反义组肿瘤组织caspas-3表达显著强于对照组(面积积分光密度分别为:反义组0.4501±0.1618,对照组0.3601±0.1065,P﹤0.05)。说明Livin反义寡核苷酸抑制了Livin的表达后,解除了Livin对caspas-3的抑制作用,从而使caspas-3的表达增强。
结论:
一.人膀胱癌细胞株5637表达Livin两个异构体。
二.Livin在人膀胱癌细胞的增殖及凋亡过程中扮演重要角色。
三.Livin抑制膀胱癌细胞凋亡的可能机制之一是其抑制了Caspase-3的活性。
四.我们合成的Livin反义寡核苷酸在体内、外均能有效抑制膀胱癌细胞Livin mRNA及蛋白的表达、抑制癌细胞的生长、诱导癌细胞凋亡。
五.Livin反义寡核苷酸可能成为膀胱癌基因治疗的潜在方法之一。
Background:
The inhibitor of apoptosis proteins (IAPs) are a family of highly conserved cell apoptosis inhibitors that have been found in yeast, invertebrates and vertebrates. The basal level of apoptosis is tightly controlled by endogenous IAPs in mammalian cells. The dysregulation of IAPs expression results in Tumorigenesis. Up to now, eight IAP-family members have been identified in human’s cells: NAIP, c-IAP1 (MIHB, HIAP-2), c-IAP2 (HIAP-1, MIHC, API2) .XIAP (hILP, MIHA, ILP-1), Survivin, BRUCE (apollon), ILP-2 and Livin (ML-IAP, KIAP). Livin is one of the novel human IAPs family members, which encodes negative regulatory proteins that prevent cell apoptosis. It was expressed during the development of embryonal. Interestingly, it was not detected in human adult tissues but expressed in the transformed cell lines and a variety of human tumors, including melanoma, bladder cancer, renal cancer, lung cancer, neuroblastomas and leukemia. It could be involved in carcinogenesis and it was important for cell survival in carcinomas. Clinical experiments also demonstrated that livin may be involved in the progression of superficial bladder cancer and used as a marker of early recurrence. In 30 patients with bladder cancer, Livin expression was measured by reverse transcription-PCR. In this study, patients with increased Livin had a shorter time to relapse compared with patients without increased Livin expression (3.5 versus 25.8 months).Our former study was to detect of Livin expression in urine of bladder cancer patients by using RT-PCR. We found that 42 of 52(86.5%)patients with bladder cancer showed positive Livinαsignals in urine,while none showed positive Livinβsignals in urine;none showed positive Livinαand Livinβsignals in urine from non-cancerous patients or healthy volunteers.It shows that Livinαis applicable to detect cancer cells in urine of patients with bladder cancer and may be helpful in screening bladder cancer.
Objective:
To investigate the effect of antisense o1igonuc1eotides targeting livin on the inhibition of livin mRNA and protein expression and apoptosis in human bladder cancer cells in vitro and in vivo. To discuss the mechanism of inhibition effect to bladder cancer cells apoptosis by Livin gene. To investigate the potency of Livin gene as a new target for bladder cancer’s treatment.
Methods:
1. Specific phosphorothioate antisense oligodeoxynucleotide targeting Livin was synthesized, and phosphorothioate missense oligodeoxy- nucleotide was synthesized as control nucleotide. And were transfected into bladder cancer cells.
2. The inhibiton effect of Livin antisense oligodeoxynucleotide (ASODN) on the biological effect of bladder cancer cell line (5637).
(1)Transfected Livin ASODN into bladder cancer cells in different concentrations. The inhibition of the proliferating of 5637 cells was assayed by MTT method.
(2) Investigate the effect of antisense o1igonuc1eotides targeting livin on the inhibition of livin mRNA and protein expression by RT-PCR, Western blot and immunofluorescence in 5637 cells.
(3)Biological effect of Livin ASODN on 5637 cells
①Morphology of 5637 cells transfected with Livin ASODN was observed by electromicroscope and confocal laser scanning microscope (CLSM).And the expression and location of livin were observed by CLSM.
②Apoptosis rate of 5637 cells transfected with Livin ASODN was assayed by AO/EB stain and flow cytometry.
3. Investigate the inhibitory effects of Livin ASODN on development and growth ability of Human bladder cancer cells xenograft in nude mice and it’s mechanism.
(1) A human bladder cancer animal model was set up by hypodermic injection of 5637 cells into nude mice.
(2) The tumor-bearing mice were randomized into 2 groups: ASODN group , and MSODN group. The o1igonuc1eotides were separately injected into the lumor bodies of mice.
(3) Investigate the effect of livin ASODN on the inhibition of livin mRNA and protein expression by RT-PCR, Western blot and immuno- histochemistry of human bladder cancer cells xenograft in nude mice.
(4) Biological effect of Livin ASODN on xenograft in nude mice.
①The tumor weight and volume were measured.
②Morphology of xenograft in nude mice was observed by microscope.
③Apoptosis index of xenograft in nude mice was assayed by TUNEL stain.
④The expression of caspase-3 of xenograft in nude mice was detected by immunohistochemistry.
Results:
1. Livin ASODN was transfected into bladder cancer cells, the transfection rate was about 55%.
2. The effect of Livin ASODN on the inhibition of livin mRNA and protein expression.
The expression of livinαand livinβin 5637 cells was detected by RT-PCR and Western blot.
After the transfection of Livin ASODN, the expression of livin mRNA and protein was decreased (P<0.01), which was detected by RT-PCR and Western blot.
In RT-PCR procedure and gel electrophoresis, the luminosity ratio of livin strip and GAPDH strip were represented the intensity of the expression of livin. The luminosity ratio of Livin ASODN group was 0.46±0.05, and the control group was 0.88±0.05 (MSODN group), 0.92±0.04 (Lipo group), 0.91±0.06 (PBS group) respectively.
In Western blot procedure, the luminosity ratio of livin strip was smaller than the control group too.
The fluorescence signal of livin observed by CLSM was significantly decreased.
3. Biological effect of Livin ASODN on 5637 cells
①Livin ASODN could inhibit the proliferating activity of bladder cancer cells, and the inhibiting effection depended on the concentration of Livin ASODN.
②Under electromicroscope and confocal laser scanning microscope, the morphology changes of 5637 cells transfected with Livin ASODN was: intumesce, degeneration, cellular organ swelling, some of cells necrosis and to form apoptotic body.
③The apoptosis rate of 5637 cells transfected with Livin ASODN was increased assayed by AO/EB stain and flow cytometry. The apoptosis rate was (46.39±9.23)%( ASODN group), (5.10±1.56)% (MSODN group), (5.70±1.61)% (Lipo group), (4.54±1.84)% (PBS group), P﹤0.01.
4. Experimentation of xenograft in nude mice
(1) The effect of livin ASODN on the inhibition of livin mRNA and protein expression.
The expression of livin mRNA and protein detected by RT-PCR, Western blot and immunohistochemistry of human bladder cancer cells xenograft in nude mice was decreased.
In RT-PCR and Western blot procedure, the luminosity of livin strip was smaller than the control group.
The expression of livin detected by immunohistochemistry of human bladder cancer cells xenograft in nude mice was decreased compare to control group(AIOD: 0.1661±0.1028 VS 0.2490±0.1483), P﹤0.05.
(2) Biological effect of Livin ASODN on xenograft in nude mice
①Tumor sizes were significantly smaller in Livin ASODN group than MSODN group from 18 to 30 days after inoculation, P < 0.05. Tumors weight were significantly lighter in Livin ASODN group(1.31±0.88g) than those in MSODN group(2.41±0.41g) at 30 days after inoculation,P<0.05
②Necrosis foci and inflammatory cell infiltration were observed in xenograft in nude mice of Livin ASODN group by microscopy.
③Apoptotic index detected by TUNEL stain in xenograft in nude mice of Livin ASODN group(19.60±5.91) was significantly higher than MSODN group(3.48±2.35), P﹤0.05.
④The expression of Caspase-3 detected by immunohistochemistry in xenograft in nude mice of Livin ASODN group (AIOD: 0.4501±0.1618)was increased compare to MSODN group(AIOD: 0.3601±0.1065), P﹤0.05.
Conclusions:
1. Human cancer cell line 5637 could express Livinαand Livinβwhich was detected by RT-PCR and Western blot.
2. Livin plays an important role in the inhibition of apoptosis and proliferation with bladder cancer cells.
3. One of the possible mechanical of the inhibition of apoptosis by Livin was the inhibition of the Caspase-3.
4. Livin ASODN could remarkably down-regulate the expression of livin gene in 5637cells, induce apoptosis and inhibit tumor growth.
5. Livin ASODN might be a potential target and strategies for the treatment of bladder cancer.
目的:
在我们的前期工作基础上,我们拟利用反义核酸技术,在体、内外阻断膀胱癌细胞Livin mRNA及蛋白质的表达,观察癌细胞的生长以及凋亡是否受到影响,以明确Livin在膀胱癌细胞中的作用,并初步探讨其机制。为以Livin为靶基因的膀胱癌的治疗提供理论依据。
方法:
一.设计、合成全硫代修饰Livin反义寡核苷酸序列及其功能鉴定。
1.参考文献设计、筛选、合成全硫代修饰Livin反义寡核苷酸。同时设计一条错义序列寡核苷酸作为对照。
2.制作Livin反义寡核苷酸- Lipofectamine?2000复合物,将其转染入5637膀胱癌细胞中。
3.荧光显微镜观察转染效率。
4.用RT-PCR、Western blot、细胞免疫荧光检测转染后膀胱癌细胞Livin mRNA及蛋白表达的变化。
二.细胞分组
根据向培养细胞中加入的成分不同,分为: (1)反义组:加入反义寡核苷酸; (2)错义组:加入错义的寡核苷酸; (3)脂质体组:只加入等量稀释的脂质体; (4)PBS(磷酸缓冲液)组:只加入等量的PBS。
三.Livin反义寡核苷酸对人膀胱癌细胞株5637生物学活性的影响
1.以不同浓度的Livin反义寡核苷酸转染培养的膀胱癌细胞,用MTT法测定细胞增殖活性。从中筛选出适当的Livin反义寡核苷酸浓度,进行下一步实验。
2.Livin反义寡核苷酸对膀胱癌细胞的生物学效应
①激光共聚焦显微镜、透射电子显微镜观察膀胱癌细胞转染Livin反义寡核苷酸后的形态学变化。
②AO/EB双染色、Annexin V-FITC细胞凋亡检测试剂盒流式细胞仪检测细胞凋亡情况。
四.Livin反义寡核苷酸诱导荷瘤鼠膀胱癌组织凋亡的研究
1.裸鼠膀胱癌移植瘤模型的建立
采用皮下注射法建立裸鼠膀胱癌移植瘤模型。
2.给药方法及分组
采用瘤体内5点注射法,根据注射核酸的不同,将裸鼠随机分为对照组(瘤体内注射错义寡核苷酸)和反义组(瘤体内注射反义寡核苷酸)。
3.Livin反义寡核苷酸对荷瘤鼠肿瘤组织Livin表达的影响
用RT-PCR、Western blot检测检测转染后膀胱癌细胞Livin mRNA及蛋白表达的变化。
4.Livin反义寡核苷酸对荷瘤鼠肿瘤组织的生物学效应
①用游标卡尺测量、计算肿瘤体积,绘制癌细胞体内生长曲线。接种后第30天剥瘤称重。
②H.E染色光镜观察肿瘤组织转染Livin反义寡核苷酸后的形态学变化。
③TUNEL法检测荷瘤鼠肿瘤细胞凋亡情况。
④免疫组化检测荷瘤鼠肿瘤caspas-3的表达情况。
结果:
一.Livin反义寡核苷酸能有效转染入膀胱癌细胞内并抑制Livin mRNA及蛋白的表达。
将FITC标记的Livin ASODN转染膀胱癌细胞后,在荧光显微镜下观察,见约55%的细胞染上绿色荧光,说明Livin ASODN能有效地转染入5637细胞中。
RT-PCR扩增后显示5637细胞表达livinα和livinβ,但反义组的电泳带亮度明显低于错义组、脂质体组及PBS组,而后三组间亮度相似。反义组livin两个片段的mRNA相对表达量(0.46±0.05)较错义组(0.88±0.05)、脂质体组(0.92±0.04)、PBS组(0.91±0.06)明显减弱,P<0.01,而各对照组间比较无显著性差异,P>0.05。
采用Western blot检测各组细胞中livin蛋白的表达,结果反义组livinα和livinβ杂交带亮度明显低于错义组、脂质体组和PBS组。细胞免疫荧光染色后用激光共聚焦显微镜观察,发现反义组癌细胞标记livin的绿色荧光明显减弱,且见有的细胞体积明显缩小,完全无绿色荧光。其余各对照组无明显变化。
说明Livin反义寡核苷酸能有效抑制膀胱癌细胞Livin mRNA及蛋白的表达。
二.Livin反义寡核苷酸对人膀胱癌细胞增殖活性的影响
MTT法发现Livin反义寡核苷酸能抑制人膀胱癌细胞的增殖活性,且其抑制作用呈明显的浓度依赖性。我们据此选择160nmol/L的寡核苷酸浓度进行下一步实验。
三.Livin反义寡核苷酸对膀胱癌细胞的生物学效应
①细胞形态的变化:激光共聚焦显微镜、透射电子显微镜下见反义组可见较多细胞体积肿大、细胞器肿胀、变性坏死。有的细胞体积缩小,发生凋亡,甚至形成凋亡小体。
②细胞凋亡率的变化:AO/EB双染色、Annexin V-FITC标记凋亡细胞,流式细胞仪检测细胞凋亡率,结果发现反义组的细胞凋亡率(46.39±9.23)%显著高于PBS组(4.54±1.84)%、脂质体组(5.70±1.61)%及错义组(5.10±1.56)%,P﹤0.01。而后三组间凋亡率差异无显著意义,P﹥0.05。
三.Livin反义寡核苷酸对荷瘤鼠肿瘤组织的抑制作用
1.Livin反义寡核苷酸对荷瘤鼠肿瘤组织Livin表达的影响
RT-PCR扩增后显示,反义组荷瘤鼠肿瘤组织表达livinα和livinβ的电泳带亮度明显低于对照组。
Western blot检测荷瘤鼠肿瘤组织livin蛋白的表达,结果反义组livinα和livinβ杂交带亮度明显低于对照组。
免疫组化也显示反义组livin表达减弱(其面积积分光密度为:0.1661±0.1028,对照组为0.2490±0.1483,P﹤0.05)说明Livin反义寡核苷酸在体内也能有效抑制膀胱癌细胞Livin mRNA及蛋白的表达。
2.Livin反义寡核苷酸对对荷瘤鼠肿瘤组织的生物学效应
①肿瘤生长的变化:通过测量、计算肿瘤体积,绘制癌细胞体内生长曲线。发现从肿瘤接种后的第18天开始至接种后第30天,反义组肿瘤体积显著小于对照组,P<0.05。接种后第30天剥瘤称重,反义组瘤块重量为1.31±0.88g,明显轻于对照组(2.41±0.41g),P<0.05。说明Livin反义寡核苷酸能在体内抑制了肿瘤的生长。
②H.E染色光镜观察肿瘤组织转染Livin反义寡核苷酸后的形态学变化:可见反义组肿瘤组织有局灶变性、坏死现象及炎症细胞浸润。
③肿瘤细胞凋亡情况:TUNEL法检测荷瘤鼠肿瘤组织凋亡指数,结果发现反义组为19.60±5.91,明显高于对照组(3.48±2.35),P﹤0.05。
④免疫组化检测荷瘤鼠肿瘤caspas-3的表达情况:反义组肿瘤组织caspas-3表达显著强于对照组(面积积分光密度分别为:反义组0.4501±0.1618,对照组0.3601±0.1065,P﹤0.05)。说明Livin反义寡核苷酸抑制了Livin的表达后,解除了Livin对caspas-3的抑制作用,从而使caspas-3的表达增强。
结论:
一.人膀胱癌细胞株5637表达Livin两个异构体。
二.Livin在人膀胱癌细胞的增殖及凋亡过程中扮演重要角色。
三.Livin抑制膀胱癌细胞凋亡的可能机制之一是其抑制了Caspase-3的活性。
四.我们合成的Livin反义寡核苷酸在体内、外均能有效抑制膀胱癌细胞Livin mRNA及蛋白的表达、抑制癌细胞的生长、诱导癌细胞凋亡。
五.Livin反义寡核苷酸可能成为膀胱癌基因治疗的潜在方法之一。
Background:
The inhibitor of apoptosis proteins (IAPs) are a family of highly conserved cell apoptosis inhibitors that have been found in yeast, invertebrates and vertebrates. The basal level of apoptosis is tightly controlled by endogenous IAPs in mammalian cells. The dysregulation of IAPs expression results in Tumorigenesis. Up to now, eight IAP-family members have been identified in human’s cells: NAIP, c-IAP1 (MIHB, HIAP-2), c-IAP2 (HIAP-1, MIHC, API2) .XIAP (hILP, MIHA, ILP-1), Survivin, BRUCE (apollon), ILP-2 and Livin (ML-IAP, KIAP). Livin is one of the novel human IAPs family members, which encodes negative regulatory proteins that prevent cell apoptosis. It was expressed during the development of embryonal. Interestingly, it was not detected in human adult tissues but expressed in the transformed cell lines and a variety of human tumors, including melanoma, bladder cancer, renal cancer, lung cancer, neuroblastomas and leukemia. It could be involved in carcinogenesis and it was important for cell survival in carcinomas. Clinical experiments also demonstrated that livin may be involved in the progression of superficial bladder cancer and used as a marker of early recurrence. In 30 patients with bladder cancer, Livin expression was measured by reverse transcription-PCR. In this study, patients with increased Livin had a shorter time to relapse compared with patients without increased Livin expression (3.5 versus 25.8 months).Our former study was to detect of Livin expression in urine of bladder cancer patients by using RT-PCR. We found that 42 of 52(86.5%)patients with bladder cancer showed positive Livinαsignals in urine,while none showed positive Livinβsignals in urine;none showed positive Livinαand Livinβsignals in urine from non-cancerous patients or healthy volunteers.It shows that Livinαis applicable to detect cancer cells in urine of patients with bladder cancer and may be helpful in screening bladder cancer.
Objective:
To investigate the effect of antisense o1igonuc1eotides targeting livin on the inhibition of livin mRNA and protein expression and apoptosis in human bladder cancer cells in vitro and in vivo. To discuss the mechanism of inhibition effect to bladder cancer cells apoptosis by Livin gene. To investigate the potency of Livin gene as a new target for bladder cancer’s treatment.
Methods:
1. Specific phosphorothioate antisense oligodeoxynucleotide targeting Livin was synthesized, and phosphorothioate missense oligodeoxy- nucleotide was synthesized as control nucleotide. And were transfected into bladder cancer cells.
2. The inhibiton effect of Livin antisense oligodeoxynucleotide (ASODN) on the biological effect of bladder cancer cell line (5637).
(1)Transfected Livin ASODN into bladder cancer cells in different concentrations. The inhibition of the proliferating of 5637 cells was assayed by MTT method.
(2) Investigate the effect of antisense o1igonuc1eotides targeting livin on the inhibition of livin mRNA and protein expression by RT-PCR, Western blot and immunofluorescence in 5637 cells.
(3)Biological effect of Livin ASODN on 5637 cells
①Morphology of 5637 cells transfected with Livin ASODN was observed by electromicroscope and confocal laser scanning microscope (CLSM).And the expression and location of livin were observed by CLSM.
②Apoptosis rate of 5637 cells transfected with Livin ASODN was assayed by AO/EB stain and flow cytometry.
3. Investigate the inhibitory effects of Livin ASODN on development and growth ability of Human bladder cancer cells xenograft in nude mice and it’s mechanism.
(1) A human bladder cancer animal model was set up by hypodermic injection of 5637 cells into nude mice.
(2) The tumor-bearing mice were randomized into 2 groups: ASODN group , and MSODN group. The o1igonuc1eotides were separately injected into the lumor bodies of mice.
(3) Investigate the effect of livin ASODN on the inhibition of livin mRNA and protein expression by RT-PCR, Western blot and immuno- histochemistry of human bladder cancer cells xenograft in nude mice.
(4) Biological effect of Livin ASODN on xenograft in nude mice.
①The tumor weight and volume were measured.
②Morphology of xenograft in nude mice was observed by microscope.
③Apoptosis index of xenograft in nude mice was assayed by TUNEL stain.
④The expression of caspase-3 of xenograft in nude mice was detected by immunohistochemistry.
Results:
1. Livin ASODN was transfected into bladder cancer cells, the transfection rate was about 55%.
2. The effect of Livin ASODN on the inhibition of livin mRNA and protein expression.
The expression of livinαand livinβin 5637 cells was detected by RT-PCR and Western blot.
After the transfection of Livin ASODN, the expression of livin mRNA and protein was decreased (P<0.01), which was detected by RT-PCR and Western blot.
In RT-PCR procedure and gel electrophoresis, the luminosity ratio of livin strip and GAPDH strip were represented the intensity of the expression of livin. The luminosity ratio of Livin ASODN group was 0.46±0.05, and the control group was 0.88±0.05 (MSODN group), 0.92±0.04 (Lipo group), 0.91±0.06 (PBS group) respectively.
In Western blot procedure, the luminosity ratio of livin strip was smaller than the control group too.
The fluorescence signal of livin observed by CLSM was significantly decreased.
3. Biological effect of Livin ASODN on 5637 cells
①Livin ASODN could inhibit the proliferating activity of bladder cancer cells, and the inhibiting effection depended on the concentration of Livin ASODN.
②Under electromicroscope and confocal laser scanning microscope, the morphology changes of 5637 cells transfected with Livin ASODN was: intumesce, degeneration, cellular organ swelling, some of cells necrosis and to form apoptotic body.
③The apoptosis rate of 5637 cells transfected with Livin ASODN was increased assayed by AO/EB stain and flow cytometry. The apoptosis rate was (46.39±9.23)%( ASODN group), (5.10±1.56)% (MSODN group), (5.70±1.61)% (Lipo group), (4.54±1.84)% (PBS group), P﹤0.01.
4. Experimentation of xenograft in nude mice
(1) The effect of livin ASODN on the inhibition of livin mRNA and protein expression.
The expression of livin mRNA and protein detected by RT-PCR, Western blot and immunohistochemistry of human bladder cancer cells xenograft in nude mice was decreased.
In RT-PCR and Western blot procedure, the luminosity of livin strip was smaller than the control group.
The expression of livin detected by immunohistochemistry of human bladder cancer cells xenograft in nude mice was decreased compare to control group(AIOD: 0.1661±0.1028 VS 0.2490±0.1483), P﹤0.05.
(2) Biological effect of Livin ASODN on xenograft in nude mice
①Tumor sizes were significantly smaller in Livin ASODN group than MSODN group from 18 to 30 days after inoculation, P < 0.05. Tumors weight were significantly lighter in Livin ASODN group(1.31±0.88g) than those in MSODN group(2.41±0.41g) at 30 days after inoculation,P<0.05
②Necrosis foci and inflammatory cell infiltration were observed in xenograft in nude mice of Livin ASODN group by microscopy.
③Apoptotic index detected by TUNEL stain in xenograft in nude mice of Livin ASODN group(19.60±5.91) was significantly higher than MSODN group(3.48±2.35), P﹤0.05.
④The expression of Caspase-3 detected by immunohistochemistry in xenograft in nude mice of Livin ASODN group (AIOD: 0.4501±0.1618)was increased compare to MSODN group(AIOD: 0.3601±0.1065), P﹤0.05.
Conclusions:
1. Human cancer cell line 5637 could express Livinαand Livinβwhich was detected by RT-PCR and Western blot.
2. Livin plays an important role in the inhibition of apoptosis and proliferation with bladder cancer cells.
3. One of the possible mechanical of the inhibition of apoptosis by Livin was the inhibition of the Caspase-3.
4. Livin ASODN could remarkably down-regulate the expression of livin gene in 5637cells, induce apoptosis and inhibit tumor growth.
5. Livin ASODN might be a potential target and strategies for the treatment of bladder cancer.
引文
[1]吴阶平主编.吴阶平泌尿外科学[M].济南.山东科学技术出版社,2004,第一版:965.
[2] Lee E, Park I, Lee C. Prognostic markers of intravesical bacillus Calmette–Guerìn therapy for multiple, high grade, stage T1 bladder cancer[J]. Int J Urol. 1997, 4: 552–556.
[3] LaCasse EC, Baird S, Korneluk RG, et al. The inhibitors of apoptosis (IAPs) and their emerging role in cancer [J]. Oncogene. 1998, 17(25):3247-59.
[4] Roy N, Mahadevan MS, McLean M, et al. The gene for neuronal apoptosis inhibitory protein is partially deleted in individuals with spinal muscular atrophy [J]. Cell. 1995, 80(1):167-78.
[5] Richter BW, Mir SS, Eiben LJ,et al. Molecular cloning of ILP-2, a novel member of the inhibitor of apoptosis protein family[J]. Mol Cell Biol. 2001, 21(13):4292-301.
[6] Rothe M, Pan MG, Henzel WJ, et al. The TNFR2-TRAF signaling complex contains two novel proteins related to baculoviral inhibitor of apoptosis proteins [J]. Cell. 1995, 83(7):1243-52.
[7] Liston P, Roy N, Tamai K,et al. Suppression of apoptosis in mammalian cells by NAIP and a related family of IAP genes[J]. Nature. 1996, 379(6563):349-53.
[8] Chen Z, Naito M, Hori S, et al. A human IAP-family gene, apollon, expressed in human brain cancer cells [J]. Biochem Biophys Res Commun. 1999, 264(3):847-54.
[9] Ambrosini G, Adida C, Altieri DC. A novel anti-apoptosis gene, survivin, expressed in cancer and lymphoma [J]. Nat Med. 1997, 3(8):917-21.
[10] Salvesen GS, Duckett CS. IAP proteins: blocking the road to death's door [J]. Nat Rev Mol Cell Biol. 2002, 3(6):401-10.
[11] Liu B, Han M, Wen JK, et al. Livin/ML-IAP as a new target for cancer treatment[J]. Cancer Lett. 2007, 250(2):168-76.
[12] Lin JH, Deng G, Huang Q, Morser J. KIAP, a novel member of the inhibitor of apoptosis protein family[J].Biochem Biophys Res Commun. 2000 ,279(3):820-31.
[13] Vucic D,Stennicke HR, Pisabarro MT et al. ML-IAP, a novel inhibitor of apoptosis that is preferentially expressed in human melanomas[J]. Curr Biol, 2000, 10(21):1359-1366.
[14] Kasof GM, Gomes BC. Livin, a novel inhibitor of apoptosis protein family member [J]. J Biol Chem, 2001,276(5):3238-3246.
[15] Ashhab Y, Alian A, Polliack A, et al. Two splicing variants of a new inhibitor of apoptosis gene with different biological properties and tissue distribution pattern [J].FEBS Lett. 2001,495(1-2):56-60.
[16] Crnkovi?-Mertens I, Semzow J, Hoppe-Seyler F,et,al. Isoform-specific silencing of the Livin gene by RNA interference defines Livin beta as key mediator of apoptosis inhibition in HeLa cells[J]. J Mol Med.2006, 84(3):232-40.
[17] Gong J, Chen N, Zhou Q,et al. Melanoma inhibitor of apoptosis protein is expressed differentially in melanoma and melanocytic naevus, but similarly in primary and metastatic melanomas[J]. J Clin Pathol. 2005,58(10):1081-5.
[18] Gazzaniga P,Gradilone A,Giuliani L, et al. Expression and prognostic significance of LIVIN, SURVIVIN and other apoptosis-related genes in the Progression of superficial bladder cancer [J]. Ann Oncol, 2003,14(1):85-89.
[19] Yin W, Chen N, Zhang Y,et al. Survivin nuclear labeling index: a superior biomarker in superficial urothelial carcinoma of human urinary bladder[J]. Mod Pathol. 2006, 19(11):1487-97.
[20] Kempkensteffen C, Hinz S, Christoph F,et al. Expression of the apoptosis inhibitor Livin in renal cell carcinomas: correlations with pathology and outcome[J]. Tumour Biol. 2007, 28 (3):132-8.
[21] Wagener N, Crnkovi?-Mertens I, Vetter C,et al. Expression of inhibitor of apoptosis protein Livin in renal cell carcinoma and non-tumorous adult kidney[J]. Br J Cancer. 2007, 97(9):1271-6.
[22] Kitamura H, Honma I, Torigoe T,et al. Expression of Livin in renal cell carcinoma and detection of anti-Livin autoantibody in patients[J]. Urology. 2007, 70(1):38-42.
[23] Tanabe H, Yagihashi A, Tsuji N,et al. Expression of survivin mRNA and Livin mRNA in non-small-cell lung cancer[J]. Lung Cancer. 2004 ,46(3):299-304.
[24] Yagihashi A, Asanuma K, Kobayashi D,et al. Detection of autoantibodies to Livin and survivin in Sera from lung cancer patients[J]. Lung Cancer. 2005,48(2):217-21.
[25] Choi J, Hwang YK, Sung KW,et al. Expression of Livin, an antiapoptotic protein, is an independent favorable prognostic factor in childhood acute lymphoblastic leukemia[J]. Blood. 2007,109(2):471-7.
[26] Qiuping Z, Jei X, Youxin J,et al. CC chemokine ligand 25 enhances resistance to apoptosis in CD4+ T cells from patients with T-cell lineage acute and chronic lymphocytic leukemia by means of Livin activation[J]. Cancer Res. 2004,64 (20):7579-87.
[27]易正金,杨林,吴小候,等.膀胱癌患者尿液脱落细胞Livin检测的临床意义[J].现代泌尿外科杂志,2007,12(3):179-181
[28] Chang H, Schimmer AD. Livin/melanoma inhibitor of apoptosis protein as a potential therapeutic target for the treatment of malignancy [J]. Mol Cancer Ther. 2007,6(1):24-30.
[29]张礼和.以核酸作为靶的药物研究[M].北京.科学出版社,1996,第一版:221-250.
[30] Geary RS, Bradley JD, Watanabe T, et al. Lack of pharmacokinetic interaction for ISIS 113715, a 2'-0-methoxyethyl modified antisense oligonucleotide targeting protein tyrosine phosphatase 1B messenger RNA, with oral antidiabetic compounds metformin, glipizide or rosiglitazone[J]. Clin Pharmacokinet. 2006,45(8):789-801.
[31] Chi KN, Eisenhauer E, Fazli L, et al. A phase I pharmacokinetic and pharmacodynamic study of OGX-011, a 2'-methoxyethyl antisense oligonucleotide to clusterin, in patients with localized prostate cancer [J]. J Natl Cancer Inst. 2005, 97(17):1287-1296.
[1] LaCasse EC, Baird S, Korneluk RG, et al. The inhibitors of apoptosis (IAPs) and their emerging role in cancer [J]. Oncogene. 1998, 17(25):3247-59.
[2] Roy N, Mahadevan MS, McLean M, et al. The gene for neuronal apoptosis inhibitory protein is partially deleted in individuals with spinal muscular atrophy [J]. Cell. 1995, 80(1):167-78.
[3] Richter BW, Mir SS, Eiben LJ,et al. Molecular cloning of ILP-2, a novel member of the inhibitor of apoptosis protein family[J]. Mol Cell Biol. 2001, 21(13):4292-301.
[4] Rothe M, Pan MG, Henzel WJ, et al. The TNFR2-TRAF signaling complex contains two novel proteins related to baculoviral inhibitor of apoptosis proteins [J]. Cell. 1995, 83(7):1243-52.
[5] Liston P, Roy N, Tamai K,et al. Suppression of apoptosis in mammalian cells by NAIP and a related family of IAP genes[J]. Nature. 1996, 379(6563):349-53.
[6] Chen Z, Naito M, Hori S, et al. A human IAP-family gene, apollon, expressed in human brain cancer cells [J]. Biochem Biophys Res Commun. 1999, 264(3):847-54.
[7] Ambrosini G, Adida C, Altieri DC. A novel anti-apoptosis gene, survivin, expressed in cancer and lymphoma [J]. Nat Med. 1997, 3(8):917-21.
[8] Salvesen GS, Duckett CS. IAP proteins: blocking the road to death's door [J]. Nat Rev Mol Cell Biol. 2002, 3(6):401-10.
[9] Liu B, Han M, Wen JK, et al. Livin/ML-IAP as a new target for cancer treatment. Cancer Lett. 2007,250(2):168-76.
[10]易正金,杨林,吴小候,等.膀胱癌患者尿液脱落细胞Livin检测的临床意义[J].现代泌尿外科杂志,2 0 0 7,12(3):179-181
[11] Ashhab Y, Alian A, Polliack A, et, al. Two splicing variants of a new inhibitor of apoptosis gene with different biological properties and tissue distribution pattern [J].FEBS Lett. 2001, 20; 495(1-2):56-60.
[12]张华东,袁守军,陈惠鹏.计算机辅助设计抗肿瘤凋亡基因Livin的反义核酸[J].中国临床药理学与治疗学2005 ;10(7) :750– 754
[13] Lin JH, Deng G, Huang Q, Morser J. KIAP, a novel member of the inhibitor ofapoptosis protein family[J].Biochem Biophys Res Commun. 2000 ,279(3):820-31.
[14] Vucic D,Stennicke HR, Pisabarro MT et al. ML-IAP, a novel inhibitor of apoptosis that is preferentially expressed in human melanomas[J]. Curr Biol, 2000, 10(21):1359-1366.
[15] Gazzaniga P,Gradilone A, Giuliani L, et al. Expression and prognostic significance of LIVIN, SURVIVIN and other apoptosis-related genes in the Progression of superficial bladder cancer[J]. Ann Oncol, 2003, 14(1):85-89.
[16]张礼和.以核酸作为靶的药物研究[M].北京.科学出版社,1996,第一版:221-250.
[17] No authors listed.Oblimersen: Augmerosen, BCL-2 antisense oligonucleotide - Genta, G 3139, GC 3139, oblimersen sodium [J]. Drugs R D. 2007,8(5):321-34
[18] Geary RS, Bradley JD, Watanabe T, et al. Lack of pharmacokinetic interaction for ISIS 113715, a 2'-0-methoxyethyl modified antisense oligonucleotide targeting protein tyrosine phosphatase 1B messenger RNA, with oral antidiabetic compounds metformin, glipizide or rosiglitazone[J]. Clin Pharmacokinet. 2006, 45(8):789-801.
[19] Chi KN, Eisenhauer E, Fazli L, et al. A phase I pharmacokinetic and pharmacodynamic study of OGX-011, a 2'-methoxyethyl antisense oligonucleotide to clusterin, in patients with localized prostate cancer [J]. J Natl Cancer Inst.2005, 97(17):1287-1296.
[1] Hong Chang,Aaron D. Shimmer. Livin/melanoma inhibitor of apoptosis protein as a potential therapeutic target for the treatment of malignancy [J]. Mol Cancer Ther.2007, 6(1):24-30.
[2] Liu B, Han M, Wen JK, Wang L. Livin/ML-IAP as a new target for cancer treatment [J]. Cancer Lett.2007, 250(2):168-176.
[3] Gazzaniga P,Gradilone A, Giuliani L, et al. Expression and prognostic significance of LIVIN, SURVIVIN and other apoptosis-related genes in the Progression of superficial bladder cancer[J]. Ann Oncol.2003; 14(1):85-89.
[4] Kato J, Kuwabara J, Mitani M, et al. Expression of survivin in esophageal cancer: correlation with the prognosis and response to chemotherapy [J]. Int J Cancer.2001, 95(2):92-95
[5] Kasof GM, Gomes BC. Livin, a novel inhibitor of apoptosis protein family member [J]. J Biol Chem.2001, 276(5):3238-3246.
[1]张礼和.以核酸作为靶的药物研究[M].北京.科学出版社,1996,第一版:221-250.
[2] Kasof GM, Gomes BC. Livin, a novel inhibitor of apoptosis protein family member [J]. J Biol Chem.2001 ,276 (5) :3238-46
[3] Vucic D, Franklin MC, Wallweber HJ,et al. Engineering ML-IAP to produce an extraordinarily potent caspase 9 inhibitor: implications for Smac-dependent anti-apoptotic activity of ML-IAP[J]. Biochem J. 2005,385 (Pt 1):11-20.
[1] LaCasse EC, Baird S, Korneluk RG, et al. The inhibitors of apoptosis (IAPs) and their emerging role in cancer [J]. Oncogene. 1998, 17(25):3247-59.
[2] Crook NE, Clem RJ, Miller LK. An apoptosis-inhibiting baculovirus gene with a zinc finger-like motif [J]. J Virol. 1993 ,67(4):2168-74.
[3] Birnbaum MJ, Clem RJ, Miller LK. An apoptosis-inhibiting gene from a nuclear polyhedrosis virus encoding a polypeptide with Cys/His sequence motifs [J]. J Virol. 1994 ,68(4):2521-8.
[4] Roy N, Mahadevan MS, McLean M,et al. The gene for neuronal apoptosis inhibitory protein is partially deleted in individuals with spinal muscular atrophy [J]. Cell. 1995, 80(1):167-78.
[5] Richter BW, Mir SS, Eiben LJ,et al. Molecular cloning of ILP-2, a novel member of the inhibitor of apoptosis protein family [J]. Mol Cell Biol. 2001, 21(13):4292-301.
[6] Rothe M, Pan MG, Henzel WJ, et al. The TNFR2-TRAF signaling complex contains two novel proteins related to baculoviral inhibitor of apoptosis proteins [J]. Cell. 1995, 83(7):1243-52.
[7] Liston P, Roy N, Tamai K,et al. Suppression of apoptosis in mammalian cells by NAIP and a related family of IAP genes [J]. Nature. 1996, 379(6563):349-53.
[8] Chen Z, Naito M, Hori S, et al. A human IAP-family gene, apollon, expressed in human brain cancer cells [J]. Biochem Biophys Res Commun. 1999, 264(3):847-54.
[9] Ambrosini G, Adida C, Altieri DC. A novel anti-apoptosis gene, survivin, expressed in cancer and lymphoma [J]. Nat Med. 1997, 3(8):917-21.
[10] Salvesen GS, Duckett CS. IAP proteins: blocking the road to death's door [J]. Nat Rev Mol Cell Biol. 2002, 3(6):401-10.
[11] Liu B, Han M, Wen JK, et al. Livin/ML-IAP as a new target for cancer treatment [J]. Cancer Lett. 2007, 250(2):168-76.
[12] Lin JH, Deng G, Huang Q, Morser J. KIAP, a novel member of the inhibitor of apoptosis protein family[J].Biochem Biophys Res Commun. 2000 ,279(3):820-31.
[13] Vucic D,Stennicke HR, Pisabarro MT et al. ML-IAP, a novel inhibitor of apoptosis that is preferentially expressed in human melanomas [J]. Curr Biol, 2000, 10(21):1359-1366.
[14] Kasof GM, Gomes BC. Livin, a novel inhibitor of apoptosis protein family member [J]. J Biol Chem, 2001; 276(5):3238-3246.
[15] Franklin MC, Kadkhodayan S, Ackerly H,et al. Structure and function analysis of peptide antagonists of melanoma inhibitor of apoptosis (ML-IAP) [J]. Biochemistry. 2003, 42(27):8223-31.
[16] Ashhab Y, Alian A, Polliack A, et al. Two splicing variants of a new inhibitor of apoptosis gene with different biological properties and tissue distribution pattern [J].FEBS Lett. 2001, 495(1-2):56-60.
[17] Gazzaniga P,Gradilone A, Giuliani L, et al. Expression and prognostic significance of LIVIN, SURVIVIN and other apoptosis-related genes in the Progression of superficial bladder cancer [J]. Ann Oncol.2003; 14(1):85-89.
[18] Crnkovi?-Mertens I, Semzow J, Hoppe-Seyler F,et,al. Isoform specific silencing of the Livin gene by RNA interference defines Livin beta as key mediator of apoptosis inhibition in HeLa cells [J]. J Mol Med. 2006,84 (3):232-40.
[19] Hariu H, Hirohashi Y, Torigoe T,et al. Aberrant expression and potency as a cancer immunotherapy target of inhibitor of apoptosis protein family, Livin/ML-IAP in lung cancer [J]. Clin Cancer Res. 2005,11 (3):1000-1009.
[20] Vucic D, Deshayes K, Ackerly H,et al. SMAC negatively regulates the anti-apoptotic activity of melanoma inhibitor of apoptosis (ML-IAP) [J].J Biol Chem. 2002,277 (14):12275-9.
[21] Ka H, Hunt JS. Temporal and spatial patterns of expression of inhibitors of apoptosis in human placentas [J].Am J Pathol. 2003, 163(2):413-22.
[22] Wagener N, Crnkovi?-Mertens I, Vetter C,et al. Expression of inhibitor of apoptosis protein Livin in renal cell carcinoma and non-tumorous adult kidney [J]. Br J Cancer. 2007, 97(9):1271-6.
[23] Kasof GM, Gomes BC. Livin, a novel inhibitor of apoptosis protein family member [J]. J Biol Chem. 2001,276 (5) :3238-46
[24] Vucic D, Franklin MC, Wallweber HJ,et al. Engineering ML-IAP to produce an extraordinarily potent caspase 9 inhibitor: implications for Smac-dependent anti-apoptotic activity of ML-IAP [J]. Biochem J. 2005,385 (Pt 1):11-20.
[25] Sanna MG, da Silva Correia J, Ducrey O,et al. IAP suppression of apoptosis involves distinct mechanisms: the TAK1/JNK1 signaling cascade and caspase inhibition [J]. Mol Cell Biol. 2002 ,22 (6) :1754-66.
[26] Duckett CS. IAP proteins: sticking it to Smac [J]. Biochem J. 2005, 385(Pt 1):e1-2.
[27] Yuan D, Liu L, Gu D. Transcriptional regulation of Livin by beta-catenin/TCF signaling in human lung cancer cell lines [J]. Mol Cell Biochem. 2007, 306(1-2):171-8.
[28] Gong J, Chen N, Zhou Q,et al. Melanoma inhibitor of apoptosis protein is expressed differentially in melanoma and melanocytic naevus, but similarly in primary and metastatic melanomas [J]. J Clin Pathol. 2005,58(10):1081-5.
[29] Yin W, Chen N, Zhang Y,et al. Survivin nuclear labeling index: a superior biomarker in superficial urothelial carcinoma of human urinary bladder [J]. Mod Pathol. 2006, (11):1487-97.
[30]宋涛,洪宝发,高江平,等.凋亡抑制基因Livin在膀胱癌中的表达及临床意义[J].中华医学杂志.2007,87(12):806-807
[31]宋希双,张志伟,车翔宇,等.凋亡抑制基因Livin在膀胱癌中的表达及意义[J].中华泌尿外科杂志.2006年,27,增刊:37-39
[32]李显文,杨罗艳,王红珊,等.凋亡抑制蛋白Livin在膀胱移行细胞癌中的表达及临床意义[J].临床泌尿外科杂志.2006,21(3):216-219
[33]曾剑,温端改,侯建全,等.凋亡抑制蛋白Livin在膀胱移行细胞癌中的表达及其临床意义[J].苏州大学学报(医学版) .2006 ,26 (2):247-249
[34]何远桥,曾甫,汪良. Livin在膀胱移行细胞癌中的表达及其与ki-67的关系[J].临床外科杂志.2006,l4(6):369-371
[35]鹿占鹏,王卫国.Livinα在膀胱移行细胞癌组织中表达的研究[J].中华肿瘤防治杂志.2007,14(14):1091-1093
[36]易正金,杨林,吴小候,等.膀胱癌患者尿液脱落细胞Livin检测的临床意义[J].现代泌尿外科杂志.2 0 0 7,12(3):179-181
[37] Kempkensteffen C, Hinz S, Christoph F,et al. Expression of the apoptosis inhibitor Livin in renal cell carcinomas: correlations with pathology and outcome [J]. Tumour Biol. 2007, 28(3):132-8.
[38] Crnkovi?-Mertens I, Wagener N, Semzow J, et al. Targeted inhibition of Livin resensitizes renal cancer cells towards apoptosis [J]. Cell Mol Life Sci. 2007, 64(9):1137-44.
[39] Kitamura H, Honma I, Torigoe T,et al. Expression of Livin in renal cell carcinoma and detection of anti-Livin autoantibody in patients [J]. Urology. 2007, 70(1):38-42.
[40] Tanabe H, Yagihashi A, Tsuji N,et al. Expression of survivin mRNA and Livin mRNA in non-small-cell lung cancer [J]. Lung Cancer. 2004, 46(3):299-304.
[41] Crnkovi?-Mertens I, Muley T, Meister M, et al. The anti-apoptotic Livin gene is an important determinant for the apoptotic resistance of non-small cell lung cancer cells [J]. Lung Cancer. 2006,54(2):135-42.
[42] Yagihashi A, Asanuma K, Kobayashi D,et al. Detection of autoantibodies to Livin and survivin in Sera from lung cancer patients [J]. Lung Cancer. 2005,48(2):217-21.
[43] Choi J, Hwang YK, Sung KW,et al. Expression of Livin, an antiapoptotic protein, is an independent favorable prognostic factor in childhood acute lymphoblastic leukemia [J]. Blood. 2007,109(2):471-7.
[44]徐璇,刘珊珊,龚萍.儿童急性白血病Livin基因表达及意义[J].中国医师杂志.2007,9(12):1630-1632
[45]何群力,苗立群,赵国新. Livin蛋白在急性淋巴细胞白血病儿童中的表达[J].实用儿科临床杂志.2007,22(15):1141-1142
[46] Qiuping Z, Jei X, Youxin J,et al. CC chemokine ligand 25 enhances resistance to apoptosis in CD4+ T cells from patients with T-cell lineage acute and chronic lymphocytic leukemia by means of Livin activation[J]. Cancer Res. 2004,64 (20):7579-87.
[47] Qiuping Z, Jie X, Youxin J,et al. Selectively frequent expression of CXCR5 enhances resistance to apoptosis in CD8(+)CD34(+) T cells from patients with T-cell-lineage acute lymphocytic leukemia [J]. Oncogene. 2005, 24(4):573-84.
[48]李家兵,李锦秀,姚惠.凋亡抑制蛋白Livin在良性前列腺增生和前列腺癌中的表达及临床意义[J].重庆医科大学学报.2007,32(7):689-691
[49]鞠文,何远桥,杨军,等. BPH和前列腺腺癌中Livin的表达及其意义[J].临床泌尿外科杂志.2005,20(12):762-764
[50] Kim DK, Alvarado CS, Abramowsky CR,et al. Expression of inhibitor-of-apoptosis protein (IAP) Livin by neuroblastoma cells: correlationwith prognostic factors and outcome [J]. Pediatr Dev Pathol. 2005,8(6):621-9.
[51] Yagihashi A, Ohmura T, Asanuma K,et al. Detection of autoantibodies to survivin and Livin in sera from patients with breast cancer [J]. Clin Chim Acta. 2005,362(1-2):125-30.
[52] Xiang Y, Yao H, Wang S,et al. Prognostic value of Survivin and Livin in nasopharyngeal carcinoma [J]. Laryngoscope. 2006,116(1):126-30.
[53] Gordon GJ, Mani M, Mukhopadhyay L,et al. Expression patterns of inhibitor of apoptosis proteins in malignant pleural mesothelioma [J]. J Pathol. 2007,211(4):447-54.
[54] Yagihashi A, Asanuma K, Tsuji N,et al. Detection of anti-Livin antibody in gastrointestinal cancer patients [J]. Clin Chem. 2003,49 (7):1206-8.
[55] Saif MW, Zalonis A, Syrigos K. The clinical significance of auto antibodies in gastrointestinal malignancies: an overview [J]. Expert Opin Biol Ther. 2007,7(4):493-507.
[56] Lopes RB, Gangeswaran R, McNeish IA,et al. Expression of the IAP protein family is dysregulated in pancreatic cancer cells and is important for resistance to chemotherapy [J]. Int J Cancer. 2007,120(11):2344-52.
[57]黎军和,何文静,何友兼. Survivin和Livin在Dukes'B期结直肠癌中的表达及其临床意义[J].癌症.2007,26(5):547-551
[58] Chang H, Schimmer AD. Livin/melanoma inhibitor of apoptosis protein as a potential therapeutic target for the treatment of malignancy [J]. Mol Cancer Ther. 2007,6(1):24-30.
[59] Wang R, Lin F, Wang X,et al. Silencing Livin gene expression to inhibit proliferation and enhance chemosensitivity in tumor cells [J]. Cancer Gene Ther. 2008, Mar 14 [Epub ahead of print]
[60] Crnkovic-Mertens I, Hoppe-Seyler F, Butz K. Induction of apoptosis in tumor cells by siRNA-mediated silencing of the Livin/ML-IAP/KIAP gene [J]. Oncogene. 2003, 22(51):8330-6.
[61] Schmollinger JC, Vonderheide RH, Hoar KM,et al. Melanoma inhibitor of apoptosis protein (ML-IAP) is a target for immune-mediated tumor destruction [J]. Proc Natl Acad Sci U S A. 2003, 100(6):3398-403.
[2] Lee E, Park I, Lee C. Prognostic markers of intravesical bacillus Calmette–Guerìn therapy for multiple, high grade, stage T1 bladder cancer[J]. Int J Urol. 1997, 4: 552–556.
[3] LaCasse EC, Baird S, Korneluk RG, et al. The inhibitors of apoptosis (IAPs) and their emerging role in cancer [J]. Oncogene. 1998, 17(25):3247-59.
[4] Roy N, Mahadevan MS, McLean M, et al. The gene for neuronal apoptosis inhibitory protein is partially deleted in individuals with spinal muscular atrophy [J]. Cell. 1995, 80(1):167-78.
[5] Richter BW, Mir SS, Eiben LJ,et al. Molecular cloning of ILP-2, a novel member of the inhibitor of apoptosis protein family[J]. Mol Cell Biol. 2001, 21(13):4292-301.
[6] Rothe M, Pan MG, Henzel WJ, et al. The TNFR2-TRAF signaling complex contains two novel proteins related to baculoviral inhibitor of apoptosis proteins [J]. Cell. 1995, 83(7):1243-52.
[7] Liston P, Roy N, Tamai K,et al. Suppression of apoptosis in mammalian cells by NAIP and a related family of IAP genes[J]. Nature. 1996, 379(6563):349-53.
[8] Chen Z, Naito M, Hori S, et al. A human IAP-family gene, apollon, expressed in human brain cancer cells [J]. Biochem Biophys Res Commun. 1999, 264(3):847-54.
[9] Ambrosini G, Adida C, Altieri DC. A novel anti-apoptosis gene, survivin, expressed in cancer and lymphoma [J]. Nat Med. 1997, 3(8):917-21.
[10] Salvesen GS, Duckett CS. IAP proteins: blocking the road to death's door [J]. Nat Rev Mol Cell Biol. 2002, 3(6):401-10.
[11] Liu B, Han M, Wen JK, et al. Livin/ML-IAP as a new target for cancer treatment[J]. Cancer Lett. 2007, 250(2):168-76.
[12] Lin JH, Deng G, Huang Q, Morser J. KIAP, a novel member of the inhibitor of apoptosis protein family[J].Biochem Biophys Res Commun. 2000 ,279(3):820-31.
[13] Vucic D,Stennicke HR, Pisabarro MT et al. ML-IAP, a novel inhibitor of apoptosis that is preferentially expressed in human melanomas[J]. Curr Biol, 2000, 10(21):1359-1366.
[14] Kasof GM, Gomes BC. Livin, a novel inhibitor of apoptosis protein family member [J]. J Biol Chem, 2001,276(5):3238-3246.
[15] Ashhab Y, Alian A, Polliack A, et al. Two splicing variants of a new inhibitor of apoptosis gene with different biological properties and tissue distribution pattern [J].FEBS Lett. 2001,495(1-2):56-60.
[16] Crnkovi?-Mertens I, Semzow J, Hoppe-Seyler F,et,al. Isoform-specific silencing of the Livin gene by RNA interference defines Livin beta as key mediator of apoptosis inhibition in HeLa cells[J]. J Mol Med.2006, 84(3):232-40.
[17] Gong J, Chen N, Zhou Q,et al. Melanoma inhibitor of apoptosis protein is expressed differentially in melanoma and melanocytic naevus, but similarly in primary and metastatic melanomas[J]. J Clin Pathol. 2005,58(10):1081-5.
[18] Gazzaniga P,Gradilone A,Giuliani L, et al. Expression and prognostic significance of LIVIN, SURVIVIN and other apoptosis-related genes in the Progression of superficial bladder cancer [J]. Ann Oncol, 2003,14(1):85-89.
[19] Yin W, Chen N, Zhang Y,et al. Survivin nuclear labeling index: a superior biomarker in superficial urothelial carcinoma of human urinary bladder[J]. Mod Pathol. 2006, 19(11):1487-97.
[20] Kempkensteffen C, Hinz S, Christoph F,et al. Expression of the apoptosis inhibitor Livin in renal cell carcinomas: correlations with pathology and outcome[J]. Tumour Biol. 2007, 28 (3):132-8.
[21] Wagener N, Crnkovi?-Mertens I, Vetter C,et al. Expression of inhibitor of apoptosis protein Livin in renal cell carcinoma and non-tumorous adult kidney[J]. Br J Cancer. 2007, 97(9):1271-6.
[22] Kitamura H, Honma I, Torigoe T,et al. Expression of Livin in renal cell carcinoma and detection of anti-Livin autoantibody in patients[J]. Urology. 2007, 70(1):38-42.
[23] Tanabe H, Yagihashi A, Tsuji N,et al. Expression of survivin mRNA and Livin mRNA in non-small-cell lung cancer[J]. Lung Cancer. 2004 ,46(3):299-304.
[24] Yagihashi A, Asanuma K, Kobayashi D,et al. Detection of autoantibodies to Livin and survivin in Sera from lung cancer patients[J]. Lung Cancer. 2005,48(2):217-21.
[25] Choi J, Hwang YK, Sung KW,et al. Expression of Livin, an antiapoptotic protein, is an independent favorable prognostic factor in childhood acute lymphoblastic leukemia[J]. Blood. 2007,109(2):471-7.
[26] Qiuping Z, Jei X, Youxin J,et al. CC chemokine ligand 25 enhances resistance to apoptosis in CD4+ T cells from patients with T-cell lineage acute and chronic lymphocytic leukemia by means of Livin activation[J]. Cancer Res. 2004,64 (20):7579-87.
[27]易正金,杨林,吴小候,等.膀胱癌患者尿液脱落细胞Livin检测的临床意义[J].现代泌尿外科杂志,2007,12(3):179-181
[28] Chang H, Schimmer AD. Livin/melanoma inhibitor of apoptosis protein as a potential therapeutic target for the treatment of malignancy [J]. Mol Cancer Ther. 2007,6(1):24-30.
[29]张礼和.以核酸作为靶的药物研究[M].北京.科学出版社,1996,第一版:221-250.
[30] Geary RS, Bradley JD, Watanabe T, et al. Lack of pharmacokinetic interaction for ISIS 113715, a 2'-0-methoxyethyl modified antisense oligonucleotide targeting protein tyrosine phosphatase 1B messenger RNA, with oral antidiabetic compounds metformin, glipizide or rosiglitazone[J]. Clin Pharmacokinet. 2006,45(8):789-801.
[31] Chi KN, Eisenhauer E, Fazli L, et al. A phase I pharmacokinetic and pharmacodynamic study of OGX-011, a 2'-methoxyethyl antisense oligonucleotide to clusterin, in patients with localized prostate cancer [J]. J Natl Cancer Inst. 2005, 97(17):1287-1296.
[1] LaCasse EC, Baird S, Korneluk RG, et al. The inhibitors of apoptosis (IAPs) and their emerging role in cancer [J]. Oncogene. 1998, 17(25):3247-59.
[2] Roy N, Mahadevan MS, McLean M, et al. The gene for neuronal apoptosis inhibitory protein is partially deleted in individuals with spinal muscular atrophy [J]. Cell. 1995, 80(1):167-78.
[3] Richter BW, Mir SS, Eiben LJ,et al. Molecular cloning of ILP-2, a novel member of the inhibitor of apoptosis protein family[J]. Mol Cell Biol. 2001, 21(13):4292-301.
[4] Rothe M, Pan MG, Henzel WJ, et al. The TNFR2-TRAF signaling complex contains two novel proteins related to baculoviral inhibitor of apoptosis proteins [J]. Cell. 1995, 83(7):1243-52.
[5] Liston P, Roy N, Tamai K,et al. Suppression of apoptosis in mammalian cells by NAIP and a related family of IAP genes[J]. Nature. 1996, 379(6563):349-53.
[6] Chen Z, Naito M, Hori S, et al. A human IAP-family gene, apollon, expressed in human brain cancer cells [J]. Biochem Biophys Res Commun. 1999, 264(3):847-54.
[7] Ambrosini G, Adida C, Altieri DC. A novel anti-apoptosis gene, survivin, expressed in cancer and lymphoma [J]. Nat Med. 1997, 3(8):917-21.
[8] Salvesen GS, Duckett CS. IAP proteins: blocking the road to death's door [J]. Nat Rev Mol Cell Biol. 2002, 3(6):401-10.
[9] Liu B, Han M, Wen JK, et al. Livin/ML-IAP as a new target for cancer treatment. Cancer Lett. 2007,250(2):168-76.
[10]易正金,杨林,吴小候,等.膀胱癌患者尿液脱落细胞Livin检测的临床意义[J].现代泌尿外科杂志,2 0 0 7,12(3):179-181
[11] Ashhab Y, Alian A, Polliack A, et, al. Two splicing variants of a new inhibitor of apoptosis gene with different biological properties and tissue distribution pattern [J].FEBS Lett. 2001, 20; 495(1-2):56-60.
[12]张华东,袁守军,陈惠鹏.计算机辅助设计抗肿瘤凋亡基因Livin的反义核酸[J].中国临床药理学与治疗学2005 ;10(7) :750– 754
[13] Lin JH, Deng G, Huang Q, Morser J. KIAP, a novel member of the inhibitor ofapoptosis protein family[J].Biochem Biophys Res Commun. 2000 ,279(3):820-31.
[14] Vucic D,Stennicke HR, Pisabarro MT et al. ML-IAP, a novel inhibitor of apoptosis that is preferentially expressed in human melanomas[J]. Curr Biol, 2000, 10(21):1359-1366.
[15] Gazzaniga P,Gradilone A, Giuliani L, et al. Expression and prognostic significance of LIVIN, SURVIVIN and other apoptosis-related genes in the Progression of superficial bladder cancer[J]. Ann Oncol, 2003, 14(1):85-89.
[16]张礼和.以核酸作为靶的药物研究[M].北京.科学出版社,1996,第一版:221-250.
[17] No authors listed.Oblimersen: Augmerosen, BCL-2 antisense oligonucleotide - Genta, G 3139, GC 3139, oblimersen sodium [J]. Drugs R D. 2007,8(5):321-34
[18] Geary RS, Bradley JD, Watanabe T, et al. Lack of pharmacokinetic interaction for ISIS 113715, a 2'-0-methoxyethyl modified antisense oligonucleotide targeting protein tyrosine phosphatase 1B messenger RNA, with oral antidiabetic compounds metformin, glipizide or rosiglitazone[J]. Clin Pharmacokinet. 2006, 45(8):789-801.
[19] Chi KN, Eisenhauer E, Fazli L, et al. A phase I pharmacokinetic and pharmacodynamic study of OGX-011, a 2'-methoxyethyl antisense oligonucleotide to clusterin, in patients with localized prostate cancer [J]. J Natl Cancer Inst.2005, 97(17):1287-1296.
[1] Hong Chang,Aaron D. Shimmer. Livin/melanoma inhibitor of apoptosis protein as a potential therapeutic target for the treatment of malignancy [J]. Mol Cancer Ther.2007, 6(1):24-30.
[2] Liu B, Han M, Wen JK, Wang L. Livin/ML-IAP as a new target for cancer treatment [J]. Cancer Lett.2007, 250(2):168-176.
[3] Gazzaniga P,Gradilone A, Giuliani L, et al. Expression and prognostic significance of LIVIN, SURVIVIN and other apoptosis-related genes in the Progression of superficial bladder cancer[J]. Ann Oncol.2003; 14(1):85-89.
[4] Kato J, Kuwabara J, Mitani M, et al. Expression of survivin in esophageal cancer: correlation with the prognosis and response to chemotherapy [J]. Int J Cancer.2001, 95(2):92-95
[5] Kasof GM, Gomes BC. Livin, a novel inhibitor of apoptosis protein family member [J]. J Biol Chem.2001, 276(5):3238-3246.
[1]张礼和.以核酸作为靶的药物研究[M].北京.科学出版社,1996,第一版:221-250.
[2] Kasof GM, Gomes BC. Livin, a novel inhibitor of apoptosis protein family member [J]. J Biol Chem.2001 ,276 (5) :3238-46
[3] Vucic D, Franklin MC, Wallweber HJ,et al. Engineering ML-IAP to produce an extraordinarily potent caspase 9 inhibitor: implications for Smac-dependent anti-apoptotic activity of ML-IAP[J]. Biochem J. 2005,385 (Pt 1):11-20.
[1] LaCasse EC, Baird S, Korneluk RG, et al. The inhibitors of apoptosis (IAPs) and their emerging role in cancer [J]. Oncogene. 1998, 17(25):3247-59.
[2] Crook NE, Clem RJ, Miller LK. An apoptosis-inhibiting baculovirus gene with a zinc finger-like motif [J]. J Virol. 1993 ,67(4):2168-74.
[3] Birnbaum MJ, Clem RJ, Miller LK. An apoptosis-inhibiting gene from a nuclear polyhedrosis virus encoding a polypeptide with Cys/His sequence motifs [J]. J Virol. 1994 ,68(4):2521-8.
[4] Roy N, Mahadevan MS, McLean M,et al. The gene for neuronal apoptosis inhibitory protein is partially deleted in individuals with spinal muscular atrophy [J]. Cell. 1995, 80(1):167-78.
[5] Richter BW, Mir SS, Eiben LJ,et al. Molecular cloning of ILP-2, a novel member of the inhibitor of apoptosis protein family [J]. Mol Cell Biol. 2001, 21(13):4292-301.
[6] Rothe M, Pan MG, Henzel WJ, et al. The TNFR2-TRAF signaling complex contains two novel proteins related to baculoviral inhibitor of apoptosis proteins [J]. Cell. 1995, 83(7):1243-52.
[7] Liston P, Roy N, Tamai K,et al. Suppression of apoptosis in mammalian cells by NAIP and a related family of IAP genes [J]. Nature. 1996, 379(6563):349-53.
[8] Chen Z, Naito M, Hori S, et al. A human IAP-family gene, apollon, expressed in human brain cancer cells [J]. Biochem Biophys Res Commun. 1999, 264(3):847-54.
[9] Ambrosini G, Adida C, Altieri DC. A novel anti-apoptosis gene, survivin, expressed in cancer and lymphoma [J]. Nat Med. 1997, 3(8):917-21.
[10] Salvesen GS, Duckett CS. IAP proteins: blocking the road to death's door [J]. Nat Rev Mol Cell Biol. 2002, 3(6):401-10.
[11] Liu B, Han M, Wen JK, et al. Livin/ML-IAP as a new target for cancer treatment [J]. Cancer Lett. 2007, 250(2):168-76.
[12] Lin JH, Deng G, Huang Q, Morser J. KIAP, a novel member of the inhibitor of apoptosis protein family[J].Biochem Biophys Res Commun. 2000 ,279(3):820-31.
[13] Vucic D,Stennicke HR, Pisabarro MT et al. ML-IAP, a novel inhibitor of apoptosis that is preferentially expressed in human melanomas [J]. Curr Biol, 2000, 10(21):1359-1366.
[14] Kasof GM, Gomes BC. Livin, a novel inhibitor of apoptosis protein family member [J]. J Biol Chem, 2001; 276(5):3238-3246.
[15] Franklin MC, Kadkhodayan S, Ackerly H,et al. Structure and function analysis of peptide antagonists of melanoma inhibitor of apoptosis (ML-IAP) [J]. Biochemistry. 2003, 42(27):8223-31.
[16] Ashhab Y, Alian A, Polliack A, et al. Two splicing variants of a new inhibitor of apoptosis gene with different biological properties and tissue distribution pattern [J].FEBS Lett. 2001, 495(1-2):56-60.
[17] Gazzaniga P,Gradilone A, Giuliani L, et al. Expression and prognostic significance of LIVIN, SURVIVIN and other apoptosis-related genes in the Progression of superficial bladder cancer [J]. Ann Oncol.2003; 14(1):85-89.
[18] Crnkovi?-Mertens I, Semzow J, Hoppe-Seyler F,et,al. Isoform specific silencing of the Livin gene by RNA interference defines Livin beta as key mediator of apoptosis inhibition in HeLa cells [J]. J Mol Med. 2006,84 (3):232-40.
[19] Hariu H, Hirohashi Y, Torigoe T,et al. Aberrant expression and potency as a cancer immunotherapy target of inhibitor of apoptosis protein family, Livin/ML-IAP in lung cancer [J]. Clin Cancer Res. 2005,11 (3):1000-1009.
[20] Vucic D, Deshayes K, Ackerly H,et al. SMAC negatively regulates the anti-apoptotic activity of melanoma inhibitor of apoptosis (ML-IAP) [J].J Biol Chem. 2002,277 (14):12275-9.
[21] Ka H, Hunt JS. Temporal and spatial patterns of expression of inhibitors of apoptosis in human placentas [J].Am J Pathol. 2003, 163(2):413-22.
[22] Wagener N, Crnkovi?-Mertens I, Vetter C,et al. Expression of inhibitor of apoptosis protein Livin in renal cell carcinoma and non-tumorous adult kidney [J]. Br J Cancer. 2007, 97(9):1271-6.
[23] Kasof GM, Gomes BC. Livin, a novel inhibitor of apoptosis protein family member [J]. J Biol Chem. 2001,276 (5) :3238-46
[24] Vucic D, Franklin MC, Wallweber HJ,et al. Engineering ML-IAP to produce an extraordinarily potent caspase 9 inhibitor: implications for Smac-dependent anti-apoptotic activity of ML-IAP [J]. Biochem J. 2005,385 (Pt 1):11-20.
[25] Sanna MG, da Silva Correia J, Ducrey O,et al. IAP suppression of apoptosis involves distinct mechanisms: the TAK1/JNK1 signaling cascade and caspase inhibition [J]. Mol Cell Biol. 2002 ,22 (6) :1754-66.
[26] Duckett CS. IAP proteins: sticking it to Smac [J]. Biochem J. 2005, 385(Pt 1):e1-2.
[27] Yuan D, Liu L, Gu D. Transcriptional regulation of Livin by beta-catenin/TCF signaling in human lung cancer cell lines [J]. Mol Cell Biochem. 2007, 306(1-2):171-8.
[28] Gong J, Chen N, Zhou Q,et al. Melanoma inhibitor of apoptosis protein is expressed differentially in melanoma and melanocytic naevus, but similarly in primary and metastatic melanomas [J]. J Clin Pathol. 2005,58(10):1081-5.
[29] Yin W, Chen N, Zhang Y,et al. Survivin nuclear labeling index: a superior biomarker in superficial urothelial carcinoma of human urinary bladder [J]. Mod Pathol. 2006, (11):1487-97.
[30]宋涛,洪宝发,高江平,等.凋亡抑制基因Livin在膀胱癌中的表达及临床意义[J].中华医学杂志.2007,87(12):806-807
[31]宋希双,张志伟,车翔宇,等.凋亡抑制基因Livin在膀胱癌中的表达及意义[J].中华泌尿外科杂志.2006年,27,增刊:37-39
[32]李显文,杨罗艳,王红珊,等.凋亡抑制蛋白Livin在膀胱移行细胞癌中的表达及临床意义[J].临床泌尿外科杂志.2006,21(3):216-219
[33]曾剑,温端改,侯建全,等.凋亡抑制蛋白Livin在膀胱移行细胞癌中的表达及其临床意义[J].苏州大学学报(医学版) .2006 ,26 (2):247-249
[34]何远桥,曾甫,汪良. Livin在膀胱移行细胞癌中的表达及其与ki-67的关系[J].临床外科杂志.2006,l4(6):369-371
[35]鹿占鹏,王卫国.Livinα在膀胱移行细胞癌组织中表达的研究[J].中华肿瘤防治杂志.2007,14(14):1091-1093
[36]易正金,杨林,吴小候,等.膀胱癌患者尿液脱落细胞Livin检测的临床意义[J].现代泌尿外科杂志.2 0 0 7,12(3):179-181
[37] Kempkensteffen C, Hinz S, Christoph F,et al. Expression of the apoptosis inhibitor Livin in renal cell carcinomas: correlations with pathology and outcome [J]. Tumour Biol. 2007, 28(3):132-8.
[38] Crnkovi?-Mertens I, Wagener N, Semzow J, et al. Targeted inhibition of Livin resensitizes renal cancer cells towards apoptosis [J]. Cell Mol Life Sci. 2007, 64(9):1137-44.
[39] Kitamura H, Honma I, Torigoe T,et al. Expression of Livin in renal cell carcinoma and detection of anti-Livin autoantibody in patients [J]. Urology. 2007, 70(1):38-42.
[40] Tanabe H, Yagihashi A, Tsuji N,et al. Expression of survivin mRNA and Livin mRNA in non-small-cell lung cancer [J]. Lung Cancer. 2004, 46(3):299-304.
[41] Crnkovi?-Mertens I, Muley T, Meister M, et al. The anti-apoptotic Livin gene is an important determinant for the apoptotic resistance of non-small cell lung cancer cells [J]. Lung Cancer. 2006,54(2):135-42.
[42] Yagihashi A, Asanuma K, Kobayashi D,et al. Detection of autoantibodies to Livin and survivin in Sera from lung cancer patients [J]. Lung Cancer. 2005,48(2):217-21.
[43] Choi J, Hwang YK, Sung KW,et al. Expression of Livin, an antiapoptotic protein, is an independent favorable prognostic factor in childhood acute lymphoblastic leukemia [J]. Blood. 2007,109(2):471-7.
[44]徐璇,刘珊珊,龚萍.儿童急性白血病Livin基因表达及意义[J].中国医师杂志.2007,9(12):1630-1632
[45]何群力,苗立群,赵国新. Livin蛋白在急性淋巴细胞白血病儿童中的表达[J].实用儿科临床杂志.2007,22(15):1141-1142
[46] Qiuping Z, Jei X, Youxin J,et al. CC chemokine ligand 25 enhances resistance to apoptosis in CD4+ T cells from patients with T-cell lineage acute and chronic lymphocytic leukemia by means of Livin activation[J]. Cancer Res. 2004,64 (20):7579-87.
[47] Qiuping Z, Jie X, Youxin J,et al. Selectively frequent expression of CXCR5 enhances resistance to apoptosis in CD8(+)CD34(+) T cells from patients with T-cell-lineage acute lymphocytic leukemia [J]. Oncogene. 2005, 24(4):573-84.
[48]李家兵,李锦秀,姚惠.凋亡抑制蛋白Livin在良性前列腺增生和前列腺癌中的表达及临床意义[J].重庆医科大学学报.2007,32(7):689-691
[49]鞠文,何远桥,杨军,等. BPH和前列腺腺癌中Livin的表达及其意义[J].临床泌尿外科杂志.2005,20(12):762-764
[50] Kim DK, Alvarado CS, Abramowsky CR,et al. Expression of inhibitor-of-apoptosis protein (IAP) Livin by neuroblastoma cells: correlationwith prognostic factors and outcome [J]. Pediatr Dev Pathol. 2005,8(6):621-9.
[51] Yagihashi A, Ohmura T, Asanuma K,et al. Detection of autoantibodies to survivin and Livin in sera from patients with breast cancer [J]. Clin Chim Acta. 2005,362(1-2):125-30.
[52] Xiang Y, Yao H, Wang S,et al. Prognostic value of Survivin and Livin in nasopharyngeal carcinoma [J]. Laryngoscope. 2006,116(1):126-30.
[53] Gordon GJ, Mani M, Mukhopadhyay L,et al. Expression patterns of inhibitor of apoptosis proteins in malignant pleural mesothelioma [J]. J Pathol. 2007,211(4):447-54.
[54] Yagihashi A, Asanuma K, Tsuji N,et al. Detection of anti-Livin antibody in gastrointestinal cancer patients [J]. Clin Chem. 2003,49 (7):1206-8.
[55] Saif MW, Zalonis A, Syrigos K. The clinical significance of auto antibodies in gastrointestinal malignancies: an overview [J]. Expert Opin Biol Ther. 2007,7(4):493-507.
[56] Lopes RB, Gangeswaran R, McNeish IA,et al. Expression of the IAP protein family is dysregulated in pancreatic cancer cells and is important for resistance to chemotherapy [J]. Int J Cancer. 2007,120(11):2344-52.
[57]黎军和,何文静,何友兼. Survivin和Livin在Dukes'B期结直肠癌中的表达及其临床意义[J].癌症.2007,26(5):547-551
[58] Chang H, Schimmer AD. Livin/melanoma inhibitor of apoptosis protein as a potential therapeutic target for the treatment of malignancy [J]. Mol Cancer Ther. 2007,6(1):24-30.
[59] Wang R, Lin F, Wang X,et al. Silencing Livin gene expression to inhibit proliferation and enhance chemosensitivity in tumor cells [J]. Cancer Gene Ther. 2008, Mar 14 [Epub ahead of print]
[60] Crnkovic-Mertens I, Hoppe-Seyler F, Butz K. Induction of apoptosis in tumor cells by siRNA-mediated silencing of the Livin/ML-IAP/KIAP gene [J]. Oncogene. 2003, 22(51):8330-6.
[61] Schmollinger JC, Vonderheide RH, Hoar KM,et al. Melanoma inhibitor of apoptosis protein (ML-IAP) is a target for immune-mediated tumor destruction [J]. Proc Natl Acad Sci U S A. 2003, 100(6):3398-403.