Clobenpropit enhances anti-tumor effect of gemcitabine in pancreatic cancer

AIM: To evaluate the anti-tumor effect of clobenpropit, which is a specific H₃ antagonist and H₄ agonist, in combination with gemcitabine in a pancreatic cancer cell line.METHODS: Three kinds of human pancreatic cancer cell lines (Panc-1, MiaPaCa-2, and AsPC-1) were used in this study. Expression of H₃ and H₄ receptors in pancreatic cancer cells was identified with Western blotting. Effects of clobenpropit on cell proliferation, migration and apoptosis were evaluated. Alteration of epithelial and mesenchymal markers after administration of clobenpropit was analyzed. An in vivo study with a Panc-1 xenograft mouse model was also performed.RESULTS: H₄ receptors were present as 2 subunits in human pancreatic cancer cells, while there was no expression of H₃ receptor. Clobenpropit inhibited cell migration and increased apoptosis of pancreatic cancer cells in combination with gemcitabine. Clobenpropit up-regulated E-cadherin, but down-regulated vimentin and matrix metalloproteinase 9 in real-time polymerase chain reaction. Also, clobenpropit inhibited tumor growth (gemcitabine 294 ± 46 mg vs combination 154 ± 54 mg, P = 0.02) and enhanced apoptosis in combination with gemcitabine (control 2.5%, gemcitabine 25.8%, clobenpropit 9.7% and combination 40.9%, P = 0.001) by up-regulation of E-cadherin and down-regulation of Zeb1 in Panc-1 xenograft mouse.CONCLUSION: Clobenpropit enhanced the anti-tumor effect of gemcitabine in pancreatic cancer cells through inhibition of the epithelial-mesenchymal transition process.     

Experimental treatment of pancreatic cancer with two novel histone deacetylase inhibitors

AIM： To investigate in vitro and in vivo treatment with histone deacetylase inhibitors NVP-LAQ824 and NVP- LBH589 in pancreatic cancer.
METHODS： Cell-growth inhibition by NVP-LAQ824 and NVP-LBH589 was studied in vitro in 8 human pancreatic cancer cell lines using the 3-（4,5-dimethylthiazol-2- yl）-2,5-diphenyltetrazolium bromide （MTT） assay. In addition, the anti-tumoral effect of NVP-LBH589 was studied in a chimeric mouse model. Anti-tumoral activity of the drugs was assessed by immunoblotting for p21^WAF-1, acH4, cell cycle analysis, TUNEL assay, and immunohistochemistry for MIB-1.
RESULTS： In vitro treatment with both compounds significantly suppressed the growth of all cancer cell lines and was associated with hyperacetylation of nucleosomal histone H4, increased expression of p21^WAF-1, cell cycle arrest at G2/M-checkpoint, and increased apoptosis. In vivo, NVP-LBH589 alone significantly reduced tumor mass and potentiated the efficacy of gemcitabine. Further analysis of the tumor specimens revealed slightly increased apoptosis and no significant reduction of cell proliferation.
CONCLUSION： Our findings suggest that NVP-LBH589 and NVP-LAQ824 are active against human pancreatic cancer, although the precise mechanism of in vivo drug action is not yet completely understood. Therefore, further preclinical and clinical studies for the treatment of pancreatic cancer are recommended.     

Claudin-4: a new target for pancreatic cancer treatment using Clostridium perfringens enterotoxin

BACKGROUND & AIMS: Recently, several members of the claudin family have been identified as integral constituents of tight junctions. Using expression profiling, we previously found claudin-4 to be overexpressed in pancreatic cancer. Because claudin-4 has been described as a receptor for the cytotoxic Clostridium perfringens enterotoxin (CPE), we investigated the effect of CPE on pancreatic cancer cells. METHODS: Expression of claudin-4 was analyzed by Northern blots. In vitro toxicity of CPE was determined by trypan blue exclusion and the (86)Rb-release assay. The in vivo effect of CPE was studied in claudin-4-expressing nude mouse xenografts of the Panc-1 cell line. RESULTS: Expression analyses showed that claudin-4 was overexpressed in most pancreatic cancer tissues and cell lines and several other gastrointestinal tumors. CPE led to an acute dose-dependent cytotoxic effect, restricted to claudin-4-expressing cells and dependent on claudin-4 expression levels. Furthermore, transforming growth factor beta was identified as a negative modulator of both claudin-4 expression and susceptibility to CPE. In vivo, intratumoral injections of CPE in Panc-1 xenografts led to large areas of tumor cell necrosis and significant reduction of tumor growth. CONCLUSIONS: Our findings suggest that targeting claudin-4-expressing tumors with CPE represents a promising new treatment modality for pancreatic cancer and other solid tumors.     

塞来昔布对胰腺癌的放疗增敏作用及其机制研究

目的研究环氧合酶-2选择性抑制剂塞来昔布联合放射治疗对胰腺癌的作用，并探讨其作用机制。方法克隆形成实验和裸鼠移植瘤模型观察塞来昔布、放疗和两者联合对胰腺癌细胞SW1990体内外增殖的影响。Western印迹法检测细胞增殖相关蛋白表达。原位缺口末端标记法（TUNEL）研究胰腺癌细胞凋亡。RT-PCR检测凋亡相关基因表达的变化。体外血管生成和体外侵袭力测定方法检测塞来昔布、放疗及两者联合对胰腺癌细胞新生血管生成和细胞侵袭力的影响，RT-PCR、明胶酶谱和反式酶谱方法检测基质金属蛋白酶（MMP）-2、MMP-9及其组织抑制剂（TIMP）-1、TIMP-2的表达。结果塞来昔布在体内外均有放射增敏作用。胰腺癌细胞增殖细胞核抗原（PCNA）蛋白表达水平在塞来昔布组和联合放疗组均降低，联合组较塞来昔布组有进一步降低。塞来昔布诱导胰腺癌细胞凋亡，单独放疗并不能诱导SW1990细胞发生凋亡，两者联合凋亡细胞数明显增加。塞来昔布下调bel-2 mRNA表达，而放疗诱导bel-2 mRNA表达上调，联合组bel-2的表达最低。单剂量放疗时，裸鼠胰腺癌移植瘤的生长延迟时间为22d，联合塞来昔布为38d。单独放疗并不能抑制体外胰腺癌细胞的新生血管生成和侵袭，塞来昔布在体外抑制胰腺癌新生血管形成和侵袭，塞来昔布与放疗联合其抑制作用较单用塞来昔布无明显增强。塞来昔布抑制胰腺癌细胞合成、分泌和激活MMP-2、MMP-9，但对TIMP-1、TIMP-2的合成、分泌和激活无明显影响。结论COX-2选择性抑制剂塞来昔布对胰腺癌放疗有增敏作用，其机制涉及诱导细胞凋亡，并通过抑制胰腺癌细胞新生血管生成和细胞侵袭，间接对胰腺癌的放疗起增敏作用。     

Treatment of thyroid cancer with histone deacetylase inhibitors and peroxisome proliferator-activated receptor-gamma agonists.

Among the most promising new antineoplastic therapies for poorly differentiated or undifferentiated thyroid cancer are the histone deacetylase inhibitors and the peroxisome proliferator-activated receptor (PPAR)-gamma agonists. These two classes of drugs have been shown to inhibit growth and induce apoptosis and redifferentiation in a variety of hematologic and solid cancer cell lines and animal models. In this article we review the molecular mechanisms, in vitro and in vivo studies, and clinical applications of the histone deacetylase inhibitors and PPAR-gamma agonists in the treatment of thyroid cancer.     

姜黄素联合顺铂抗胰腺癌细胞增殖的效应

[目的]体外观察姜黄素（curcumin）联合顺铂（CDDP）抗胰腺癌细胞（Pane-1）增殖的效应。[方法]采用MTT法观察不同浓度姜黄素、CDDP单独应用对Panc-1细胞生长的抑制效应，并利用中效原理判定2药合用的效果，应用中位效应原理和联合作用指数法来定量姜黄素与CDDP的联合作用。[结果]姜黄素和CDDP对Panc-1有剂量依赖性的抑制增殖作用，作用48h姜黄素、CDDP的IC50值分别为（20．05±1．32）μmol／L、（0．89±0．31）μg／ml。中效原理分析显示，2药在联合应用时为协同效应，在相同的抑制效应下可降低姜黄素和明显降低CDDP的使用量。[结论]姜黄素可以促进Panc-1对CDDP的敏感性，降低其有效治疗浓度。     

Vimentin在胰腺癌细胞中的表达及临床意义

目的:研究vimentin在胰腺癌细胞中的表达及意义.方法:采用免疫组织化学的方法检测99例胰腺癌组织中vimentin的表达,并分析其与临床病理资料的关系.免疫荧光、Westem blot分析不同胰腺癌细胞中vimentin的表达,Transwell侵袭小室法检测细胞的运动能力.结果:在99例胰腺癌组织中vimentin阳性16例(16%),vimentin的阳性表达和淋巴结转移(P= 0.02)、远处转移(P<0.001)、肿瘤细胞的分化程度(P=0.002)存在明显的相关性.Vimentin在低分化的胰腺癌细胞Panc-1中呈高表达,而在中分化和高分化的胰腺癌细胞AsPC-1和BxPc-3中表达较低.vimentin的表达和细胞体外运动能力相关.结论:胰腺祖细胞标记物vimentin在胰腺癌细胞中存在表达,胰腺癌的发生和干细胞可能存在一定的联系,并可能预测胰腺癌的恶性程度.     

Apoptosis of human pancreatic cancer cells induced by Triptolide

AIM： To investigate apoptosis in human pancreatic cancer cells induced by Triptolide （TL）, and the relationship between this apoptosis and expression of caspase-3＇ bcl-2 and bax. METHODS： Human pancreatic cancer cell line SW1990 was cultured in DMEM media for this study. MTT assay was used to determine the cell growth inhibitory rate in vitro. Flow cytometry and TUNEL assay were used to detect the apoptosis of human pancreatic cancer cells before and after TL treatment. RT-PCR was used to detect the expression of apoptosis-associated gene caspase-3＇ bcl-2 and bax. RESULTS： TL inhibited the growth of human pancreatic cancer cells in a dose-and time-dependent manner. TL induced human pancreatic cancer cells to undergo apoptosis with typically apoptotic characteristics. TUNEL assay showed that after the treatment of human pancreatic cancer cells with 40 ng/mL TL for 12 h and 24 h, the apoptotic rates of human pancreatic cancer cells increased significantly. RT-PCR demonstrated that caspase-3 and bax were significantly up-regulated in SW1990 cells treated with TL while bcl-2 mRNA was not. CONCLUSION： TL is able to induce the apoptosis in human pancreatic cancer cells. This apoptosis may be mediated by up-regulating the expression of apoptosisassociated caspase-3 and bax gene.     

雷公藤内酯醇诱导胰腺癌细胞凋亡及对5-脂氧合酶和凋亡基因表达的影响

目的探讨雷公藤内酯醇(TL)在胰腺癌化学预防和治疗中的应用价值。方法采用人胰腺癌细胞株SW1990作为研究对象,MTT法和吖啶橙(AO)染色分别检测SW1990细胞的增殖和凋亡,采用半定量RT-PCR法检测5-脂氧合酶(5-LOX)mRNA、bcl-2 mRNA和bax mRNA的表达。结果10μg/ml的TL对SW1990细胞生长的抑制率达30%以上,0.1μg/mlTL处理24h后,SW1990细胞凋亡指数达38.5%,显著高于对照组的14.97%。5-LOX、bcl-2、bax mRNA表达从0.7160±0.0251,0.2446±0.0311和0.0260±0.0065分别变化为0.2400±0.0316,0.0700±0.0158和0.0700±0.0158。结论TL能抑制胰腺癌细胞增殖和诱导细胞凋亡,其凋亡机制可能与下调5-LOXmRNA和bcl-2mRNA、上调bax mRNA基因表达有关。     

DNA甲基转移酶1和组蛋白脱乙酰基酶1在胰腺组织中的表达及其临床意义

背景：近年针对DNA甲基化和组蛋白乙酰化修饰的表观遗传学研究是探索胰腺癌发生机制的新热点。目的：研究DNA甲基转移酶1（DNMT1）和组蛋白脱乙酰基酶1（HDAC1）在胰腺上皮内瘤变（PanIN）和胰腺癌组织中的表达．探讨其可能的临床意义。方法：以免疫组化SP法检测DNMT1、HDAC1在10例正常胰腺组织、39例证实含有PanIN的胰腺癌旁组织和54例胰腺导管腺癌（PDAC）组织中的表达。结果：DNMT1和HDAC1在胰腺组织中的阳性表达率均随组织异型程度的增加而逐渐增高，即正常胰腺导管（0％）〈PanIN—IA（7．2％±1．2％，10．7％±5．0％）〈PanIN-1B（24．5％±9．6％，40．1％±8．0％）〈PanIN-2（30．0％±11．9％，51．2％±13．4％）〈PanIN-3（46．7％±11．2％，73．1％±11.3％）〈PDAC（56．7％±27．5％，82．5％±19.4％），差异均有统计学意义（P〈0．05）。结论：DNMT1和HDAC1表达增强是胰腺癌发生的早期事件．两者共同促进了胰腺癌的进展，可能成为胰腺癌早期诊疗的新靶点。     

生长抑素2型受体基因转染对胰腺癌细胞蛋白表达的影响

目的 探讨生长抑素2型受体(hSSTR2)基因转染对胰腺癌细胞PANC1蛋白表达的影响,以寻找新的胰腺癌敏感治疗靶点.方法 利用前期构建的腺病毒载体Ad.CMV.hSSTR2.GFP将hSSTR2全长cDNA导入胰腺癌细胞PANC1.采用双向荧光差异凝胶电泳(2D-DIGE)技术分离并筛选转染hSSTR2的实验组、空载体对照组以及空白组胰腺癌细胞之间差异表达蛋白,用反射式基质辅助激光解吸附电离串联飞行时间质谱(MALDI-TOF/TOF)技术对差异蛋白进行鉴定.结果 hSSTB2成功地转染了胰腺癌细胞,获得了hSSTR2阴性和阳性表达的PANC1荧光差异蛋白表达图谱,经DeCyder v6.5软件分析,共有18个差异在1.3倍以上的蛋白质点,经质谱鉴定得到13个蛋白质.低表达的蛋白7个,为GMP合酶、磷酸化应激诱导蛋白、谷氨酸脱氢酶、Septin-11、波形蛋白、异柠檬酸脱氢酶α亚基、线粒体内膜易位酶;高表达蛋白6个,为真核延长因子1α1、丙酮酸激酶异构体M2型、烯酰-CoA水合酶、转录调节因子1-β、Mitofilin、HSP105.结论 hSSTR2基因转染胰腺癌细胞PANC1后引起蛋白表达发生变化,这些差异蛋白功能涉及到糖、脂肪、核酸代谢以及细胞生长调节和细胞凋亡等,从而为寻找新的胰腺癌敏感治疗靶点奠定基础.     

Celecoxib suppresses proliferation and metastasis of pancreatic cancer cells by down-regulating STAT3 / NF-kB and L1CAM activities

Objective

To explore the molecular mechanisms of celecoxib-induced pancreatic cancer suppression in vivo and in vitro.