关键词： 乳腺肿瘤   基因表达调控   钙黏着糖蛋白类   SASH1蛋白   含IQ模序的GTP酶活化蛋白1  

摘要： 目的:研究证实SASH1(SAM-and SH3-domain containing 1)作为一个新的抑癌基因,与许多肿瘤的发生、发展和转移密切相关。但SASH1基因调节乳腺癌转移的机制尚不明确。本文旨在探讨SASH1基因在调节乳腺癌转移中的分子作用机制。方法:收集80例重庆市肿瘤医院于2015年5月—2016年6月间收治的乳腺癌患者乳腺癌组织标本,均经病理检测(HE染色)确诊。免疫组织化学法检测人乳腺癌组织中SASH1、含IQ模序的GTP酶活化蛋白1(IQ motif-containing GTPase activating protein 1,IQGAP1)和E-钙黏蛋白(E-cadherin)的表达水平,并分析三者之间的相关性,以及SASH1和IQGAP1表达与乳腺癌患者临床指标的相关性。构建重组质粒HA-IQGAP1-pcDNA3.0和pEGFP-C3-SASH1,并将其转染人胚胎肾细胞HEK-293T,然后采用免疫沉淀-蛋白质印迹法分析SASH1与IQGAP1的相互作用。结果:人乳腺癌组织中,SASH1与IQGAP1的表达存在相关性(P<0.05),而且二者分别与E-cadherin的表达明显相关(P值均<0.001)。SASH1和IQGAP1蛋白表达水平分别与乳腺癌的肿瘤直径和肿瘤分级有相关性(P值均<0.05),但与淋巴结转移无相关性(P值均>0.05)。成功构建重组质粒HA-IQGAP1-pcDNA3.0和pEGFP-C3-SASH1;重组质粒转染HEK-293T细胞后,发现SASH1与IQGAP1之间存在蛋白-蛋白相互作用。结论:SASH1与IQGAP1结合后可发生蛋白-蛋白相互作用,并且与E-cadherin的表达密切相关。推测SASH1可能与IQGAP1和E-cadherin形成一条新的调节乳腺癌转移的信号轴。

关键词： IQGAP1   胃癌   化疗   顺铂   耐药性   上皮间质转化  

摘要： 研究目的本研究旨在探讨IQGAP1在胃癌细胞中调控上皮间质转化与化疗药物顺铂耐药性间的关系,为胃癌耐药方面提供新的治疗策略。研究方法1、CCK8法检测顺铂对胃癌细胞SGC7901和胃癌顺铂耐药细胞SGC7901/DDP的半数抑制浓度(IC50)。2、Western Blot检测SGC7901和SGC7901/DDP细胞的EMT相关蛋白E-cadherin、N-cadherin、Snail、Vimentin和Slug的表达,Transwell迁移和侵袭实验评价SGC7901和SGC7901/DDP细胞的迁移和侵袭能力。3、Western Blot检测SGC7901和SGC7901/DDP细胞的IQGAP1蛋白表达水平。4、在SGC7901细胞中转染质粒pFlag-IQGAP1,在SGC7901/DDP细胞中转染针对IQGAP1的小干扰RNA,并通过Western Blot验证转染的效率。5、通过Western Blot检测SGC7901和SGC7901/DDP细胞转染前后EMT相关蛋白E-cadherin、N-cadherin、Snail、Vimentin和Slug的表达变化,通过Transwell迁移和侵袭实验评价SGC7901和SGC7901/DDP细胞转染前后迁移和侵袭能力的变化。6、通过CCK8法检测SGC7901和SGC7901/DDP细胞转染前后顺铂IC50的变化。通过Western Blot检测SGC7901和SGC7901/DDP细胞转染前后耐药相关蛋白MRP1、MDR1及凋亡相关蛋白Bcl-2、Bax蛋白水平变化。通过流式细胞术(Annexin V/PI双染色法)检测顺铂处理下SGC7901和SGC7901/DDP细胞转染前后凋亡率的变化。研究结果1、顺铂对SGC7901/DDP细胞的IC50是SGC7901细胞的4.9倍。2、E-cadherin蛋白在SGC7901细胞相对高表达,在SGC7901/DDP细胞相对低表达;N-cadherin、Snail、Vimentin和Slug蛋白在SGC7901细胞相对低表达,在SGC7901/DDP细胞相对高表达。SGC7901/DDP细胞比SGC7901细胞具有更强的迁移和侵袭能力。3、与SGC7901细胞相比,IQGAP1蛋白在SGC7901/DDP细胞中表达增高。4、Western blot结果表明pFlag-IQGAP1明显上调SGC7901细胞中IQGAP1的表达,IQGAP1-siRNA明显抑制SGC7901/DDP细胞中IQGAP1的表达。5、SGC7901细胞转染质粒pFlag-IQGAP1上调了IQGAP1的表达后E-cadherin蛋白表达下降,N-cadherin、Snail、Vimentin和Slug蛋白表达升高,迁移及侵袭能力增强。SGC7901/DDP细胞转染IQGAP1-siRNA下调IQGAP1的表达后E-cadherin蛋白表达升高,N-cadherin、Snail、Vimentin和Slug蛋白表达降低,迁移及侵袭能力减弱。6、上调SGC7901的IQGAP1表达后顺铂的IC50升高,MRP1、MDR1、Bcl-2蛋白表达增高,Bax蛋白表达降低,顺铂处理下细胞凋亡率降低。下调SGC7901/DDP的IQGAP1表达后顺铂的IC50下降,MRP1、MDR1、Bcl-2蛋白表达降低,Bax蛋白表达升高,顺铂处理下细胞凋亡率升高。结论1、人胃癌顺铂耐药细胞株SGC7901/DDP中IQGAP1呈过表达,并且IQGAP1参与胃癌细胞在耐药形成过程中EMT的发生。2、IQGAP1可能通过影响EMT参与了胃癌细胞对顺铂的耐药性。

关键词： IQGAP1   ERK   基因沉默   舌鳞状细胞癌   增殖   侵袭   迁移  

摘要： IQ结构域三磷酸鸟苷合酶激活蛋白1(IQ-domain GTPase-activating protein 1,IQGAPI)是一种高度保守的细胞质支架蛋白,能与100余种其他蛋白质结合的多功能支架蛋白,是多种信号通路的关键调控因子。IQGAP1具有两个紧密相关的家族蛋白IQGAP2和IQGAP3,它们具有不同的功能,但是IQGAP1是更广泛表达的蛋白质,IQGAP1涉及骨骼重组和调节细胞粘附,极性和迁移。IQGAP1还通过IQ域和通过富含脯氨酸的WW域对RAF和MEK进行调节,以调节RAF/MEK/ERK信号。研究表明IQGAP1是MAPK和Wnt/β-catenin信号通路的重要调节因子。在肿瘤发生和癌症进展中起关键作用。IQGAP1在食管鳞状细胞癌中高表达,而IQGAP1敲低可以在体外和体内降低细胞增殖和转移能力。同时,过表达IQGAP1的癌细胞中AKT和ERK磷酸化水平上调。但IQGAP1在舌鳞癌中作用及机制尚不清楚。本课题进行了关于IQGAP1及ERK在舌鳞癌中的相关性研究。通过qPCR检测IQGAP1及ERK在舌鳞癌细胞株CAL27、SCC25、SCC9细胞中的表达量,结果显示,IQGAP1及ERK均在舌鳞癌细胞株中高表达。而当沉默IQGAP1后,通过qPCR及Westerm Blot实验检测IQGAP1及ERK表达,发现IQGAP1及ERK在舌鳞癌细胞中表达下调;舌鳞癌细胞的增殖、侵袭、迁移均受抑制。第一章IQGAP1基因沉默有效si-RNA的筛选目的:筛选沉默IQGAP1基因的有效si-RNA序列,选择最优序列进行后续实验。方法:1.常规培养舌鳞癌细胞株CAL27、SCC25、SCC9细胞;***检测IQGAP1及ERK在舌鳞癌细胞中的表达量;3.构建三个不同序列si-RNA,沉默IQGAPI基因,分别转染CAL27、SCC25及SCC9,qPCR实验检测沉默后各组细胞IQGAP1的表达量;***8增殖实验检测各组细胞增殖情况,根据各实验结果筛选最优序列进行后续实验。结果:***1在舌鳞癌细胞株中高表达,同时ERK在舌鳞癌细胞株中也表现为高表达;2.三个不同序列si-RNA沉默IQGAP1后,在CAL27、SCC25、SCC9细胞株中,IQGAP1均呈现低表达;3.在CCK8增殖实验中,三组细胞株中,三个不同序列si-RNA在不同时间段有不同程度差异,均有统计学意义,其中编码为si-RNA-3746序列在各组细胞株中均有差异,且其在SCC25中较为明显且稳定,选择si-RNA-3746及SCC25作为后续实验的最优序列及细胞株。第二章沉默IQGAP1基因对舌鳞癌细胞SCC25中ERK基因表达的影响目的:IQGAPI基因在舌鳞状细胞癌中呈现高表达,相应ERK基因也高表达。研究两者之间是否存在联系,当沉默IQGAP1基因,ERK基因表达是否降低。本实验通过沉默IQGAP1基因后检测ERK基因的表达,以验证两者的相关性。方法:1.沉默IQGAP1基因,以qPCR实验验证沉默IQGAP1基因后IQGAP1基因及ERK基因的表达水平;2.沉默IQGAP1基因,以Western Blot实验验证沉默IQGAP1基因后IQGAP1基因及ERK基因的表达水平。结果:1.沉默IQGAP1后,通过qPCR实验验证,结果显示SCC25细胞中IQGAP1基因表达量下降,同时ERK基因表达量也下降。2.沉默IQGAP1后,通过Wserern Blot验证,结果显示3CC25细胞中IQGAP1基因蛋白表达量下降,同时ERK基因蛋白表达量也下降。第三章沉默IQGAP1基因对舌鳞癌细胞SCC25的增殖、侵袭、迁移的影响目的:研究沉默IQGAP1基因后对舌鳞癌细胞SCC25的增殖、侵袭、迁移的影响。方法:1.沉默SCC25细胞IQGAP1基因;***8检测实验组及阴性对照组、空白对照组的细胞增殖情况;***检测实验组及阴性对照组、空白对照组的细胞侵袭性的改变;***检测实验组及阴性对照组、空白对照组的细胞迁移情况。结果:沉默SCC25细胞IQGAP1基因后,SCC25细胞的增殖性、侵袭性及迁移能力较对照组显著下降。全文总结***1与ERK基因在舌鳞癌细胞CAL27、SCC25、SCC9中高表达;2.当沉默SCC25细胞中IQGAP1基因时,ERK基因表达下调,癌细胞的增殖性、侵袭性、迁移能力下降;***1基因可能是舌鳞癌治疗的一个潜在新靶点。

关键词： 胰腺癌   IQGAP1   DVL2   上皮间质转化   预后  

摘要： 目的胰腺癌是目前致死率最高的恶性肿瘤之一,被称之为癌中之王,5年生存率极低。因此,研究胰腺癌快速增殖和高度侵袭转移的原因,寻找其重要的致病因子,对早期诊断胰腺癌、研发有效治疗药物、改善患者的预后有着极其重要的意义。IQGAP1(IQ-motif-containing GTPase-activating protein 1)是一种支架蛋白,研究发现其在甲状腺癌、结肠癌、肺癌中等多种实体瘤高表达,促进肿瘤进展。但IQGAP1在胰腺癌中研究甚少,作用机制仍不清楚。本研究从临床标本、体外细胞实验、动物模型阐述其在胰腺癌进展中的作用机制,为治疗胰腺癌提供新的靶点。方法1探讨IQGAP1影响胰腺癌增殖能力的体内外研究:首先,利用胰腺癌临床标本比较IQGAP1在癌组织及癌旁组织的差异表达及其与临床病理学特征的相关性,并分析其与胰腺癌患者预后的关系;然后,设计针对IQGAP1基因的2个干扰RNA(siRNA)和过表达质粒,选用干扰效率最好的siRNA转染IQGAP1高表达的2种胰腺癌细胞,利用过表达质粒转染IQGAP1低表达的细胞株;其次,分别应用细胞增殖、平板克隆实验证明其对胰腺癌细胞体外生长的作用;最后,利用稳定转染的shIQGAP1构建裸鼠胰腺癌皮下移植瘤模型,分析IQGAP1对肿瘤生长的影响。2探讨IQGAPI影响胰腺癌侵袭及转移能力的体内外研究:首先,将构建好的si1QGAP1转染高表达胰腺癌细胞株、过表达质粒转染低表达细胞株,利用划痕实验和Transwell迁移实验检测胰腺癌细胞迁移及侵袭能力的改变;然后,分别利用Western-blotting和PCR技术检测EMT相关蛋白及相关转录因子的变化;其次,分析IQGAP1与调控胰腺癌进展的关键信号通路的关系;最后,建立裸鼠胰腺原位癌肝转移模型,分析IQGAP1对肿瘤体内转移情况的影响。结果1探讨IQGAP1影响胰腺癌增殖能力的体内外研究:利用免疫组化染色,Western-blotting和PCR方法发现胰腺癌组织中IQGAP1mRNA和蛋白阳性表达率较癌旁组织明显升高,与肿瘤分化程度、淋巴结转移有关,而且IQGAP1高表达者提示预后较差,多因素分析提示IQGAP1为影响预后的独立因素。同时IQGAP1mRNA和蛋白在人胰腺癌细胞系中的表达都明显高于正常胰腺导管上皮细胞。通过转染siIQGAP1沉默SW1990和CFPAC-1细胞,过表达质粒转染PANC-1细胞证实siIQGAP1表达可抑制胰腺癌细胞增殖、克隆形成,而过表达IQGAP1则出现相反的结果。构建裸鼠胰腺癌移植瘤模型证实shIQGAP1组的瘤体积及重量均低于对照组。2探讨IQGAP1影响胰腺癌侵袭及转移能力的体内外研究:siIQGAP1可抑制胰腺癌细胞迁移及侵袭能力,siIQGAP1抑制DVL2、β-catenin及下游靶基因的表达,也抑制EMT相关蛋白及转录因子的表达;过表达IQGAP1则相反;免疫反应共沉淀提示IQGAP1能与DVL2、β-catenin形成复合物,在siIQGAP1干扰的细胞中发现DVL2与β-catenin结合明显减少;siDVL2则逆转了过表达IQGAP1引起的EMT过程。裸鼠胰腺原位癌肝转移模型提示shIQGAP1抑制胰腺癌肝转移。结论1 IQGAP1在胰腺癌组织及癌细胞系中均高表达,与病理分级、淋巴结转移及胰腺癌患者预后差明显相关,是影响胰腺癌患者预后独立危险因素。2 IQGAP1促进胰腺癌EMT及增殖、迁移,促进胰腺癌肝转移的发生。3 IQGAP1与DVL2、β-catenin形成复合物,干扰IQGAP1后DVL2与β-catenin结合明显减少,抑制DVL2可逆转IQGAP1过表达引起的EMT过程。

关键词： 食管癌   顺铂   紫杉醇   耐药   IQGAP1  

摘要： 目的:1.体外建立人食管癌顺铂耐药KYSE150/CDDP、人食管癌紫杉醇耐药KYSE150/PTX细胞系。2.观察顺铂和紫杉醇耐药对人食管癌细胞恶性表型的影响。3.探讨影响顺铂和紫杉醇耐药的分子机制。方法:1.采用体外低剂量间歇诱导+浓度递增的方式,分别使用顺铂、紫杉醇为诱导药物,建立人食管癌顺铂耐药KYSE150/CDDP细胞系、人食管癌紫杉醇耐药KYSE150/PTX细胞系,显微镜下拍照记录顺铂和紫杉醇诱导后KYSE150细胞的变化。MTT法测细胞对顺铂、紫杉醇和5-氟尿嘧啶的耐受程度。2.实验检测顺铂和紫杉醇耐药对食管癌细胞的影响,MTT法及细胞克隆形成实验检测增殖、DAPI染色法检测凋亡、粘附实验及划痕愈合实验检测粘附及迁移、体外小管形成实验检测小管形成。3.利用Western blot检测耐药细胞及亲代KYSE150细胞中Caspase-3、E-cadherin及MMP-2、VEGF、IQGAP1蛋白的表达水平。4.探讨IQGAP1基因干扰能否增强KYSE150/CDDP和KYSE150/PTX细胞的化疗敏感性。首先构建及鉴定IQGAP1基因短发卡RNA干扰质粒,然后稳定转染KYSE150/CDDP细胞、KYSE150/PTX细胞,通过Western blot鉴定IQGAP1蛋白的表达水平,最后通过上述实验方法检测IQGAP1对耐药细胞化疗敏感性、增殖、抗凋亡、粘附和迁移能力的影响,并采用Western blot法检测MMP-2蛋白及ERK通路的变化。结果:1.历时10个月,建立人食管癌顺铂耐药KYSE150/CDDP细胞系及人食管癌紫杉醇耐药KYSE150/PTX细胞系,显微镜下观察耐药细胞形态,发现KYSE150细胞对CDDP、PTX产生稳定耐药后并未发现细胞形态上的改变。MTT测得顺铂耐药KYSE150/CDDP细胞对顺铂耐药指数(RI)为6.175,紫杉醇耐药KYSE150/PTX细胞对紫杉醇RI为5.226,且两株耐药细胞有交叉耐药现象。2.实验结果显示,与亲代KYSE150细胞相比,KYSE150/CDDP细胞和KYSE150/PTX细胞生长速度减慢、增殖能力也降低、UV照射30 s后的凋亡细胞数减少、细胞间聚集能力减弱及分离能力增强(粘附力减弱)、划痕24 h后无细胞区域更小、形成的小管数更多。*** blot结果显示:与亲代KYSE150细胞相比,KYSE150/CDDP细胞和KYSE150/PTX细胞的Procaspase-3、MMP-2、VEGF、IQGAP1蛋白表达水平均增高,而E-cadherin蛋白表达水平降低。4.经测序证实IQGAP1基因短发卡RNA干扰质粒构建成功,荧光显微镜下观察绿色荧光蛋白(GFP)显示,实验组与对照组转染效率分别为(76±5.780)%和(83±3.851)%,RT-PCR和Western blot结果也表明构建的IQGAP1干扰载体能显著抑制KYSE150细胞中IQGAP1基因的表达;经G418筛选构建IQGAP1干扰的稳转细胞系及阴性对照细胞系,分别命名为K/D/sh RNA及K/D/Control、K/P/sh RNA及K/P/Control,Western blot结果显示IQGAP1干扰组细胞的IQGAP1蛋白表达水平较空载体转染组显著降低,提示IQGAP1基因干扰的稳转耐药细胞系构建成功;干扰IQGAP1基因的表达使耐药细胞的药物耐受性降低、抗凋亡能力减弱、细胞间粘附力增强而迁移能力减弱。Western blot检测发现,与K/D/Control细胞相比,K/D/sh RNA细胞中MMP-2、p-ERK水平降低。结论:1.成功建立人食管癌顺铂耐药KYSE150/CDDP细胞系及人食管癌紫杉醇耐药KYSE150/PTX细胞系,且获得顺铂及紫杉醇抗性后,细胞增殖能力减弱,抗凋亡、迁移及血管形成能力增强,细胞间粘附能力降低,提示这些改变可能是顺铂和紫杉醇耐药影响临床治疗效果的重要原因。*** blots结果提示IQGAP1、Caspase-3、E-cadherin、MMP-2、VEGF基因参与了食管癌顺铂和紫杉醇耐药机制的形成。3.成功构建IQGAP1基因干扰载体,并利用该重组质粒在KYSE150/CDDP、KYSE150/PTX细胞中构建了干扰IQGAP1基因表达的稳转细胞系,IQGAP1干扰后提高了耐药细胞的药物敏感性及细胞间粘附力,减慢了细胞的增殖,降低了耐药细胞的抗凋亡及迁移能力。而IQGAP1干扰可能通过ERK通路活化逆转食管癌顺铂耐药。

关键词： microRNA-124   IQ motif containing GTPase activating protein 1   colorectal cancer   extracellular signal regulated kinase 1/2   beta-catenin  

摘要： MicroRNA (miRNA/miR)-124 is a miRNA, which exerts tumor suppressive effects but is frequently absent in tumors. Although it has been validated to target oncogenic genes such as signal transducer and activator of transcription 3, forkhead box Q1, and Slug, the mechanistic link between miR-124 and potential target genes that contribute to tumor progression, is yet to be investigated. IQ motif containing GTPase activating protein 1 (IQGAP1) is a scaffold protein that participates in protein-protein interactions and integrating diverse signaling pathways. Previous studies suggest that overexpression of IQGAP1 enhances activity of mitogen activated protein kinase 1 and -catenin signaling cascades to facilitate tumor progression. The present study aimed to identify the regulative link between miR-124 and IQGAP1 in colorectal cancer (CRC). It was demonstrated that IQGAP1 was aberrantly overexpressed in CRC tissues and cell lines. Knockdown of IQGPA1 by introducing short hairpin-IQGAP1 lentivirus inhibited CRC cell growth and colony formation ability, and simultaneously suppressed phosphorylation of extracellular signal-regulated kinase (ERK)1/2 and -catenin expression. Furthermore, it was demonstrated that miR-124 was silenced in CRC. Restoration of miR-124 in CRC cells impeded cell growth and colony formation ability. The direct binding of miR-124 to the 3untranslated region of IQGAP1 mRNA was confirmed using a luciferase reporter gene assay. Importantly, downregulation of IQGAP1 expression was observed in miR-124-restoration cells with simultaneous reduction of phosphorylated-ERK1/2 and -catenin. In conclusion, the present study describes a potential mechanism underlying the miR-124/IQGAP1 link in CRC progression. Silencing of miR-124 may depress IQGAP1 expression, leading to increased activity of ERK1/2 and -catenin signaling.

关键词： molecular scaffolds   receptor tyrosine kinases   receptors   signaling  

摘要： Axl is a tyrosine kinase receptor that is important for hematopoiesis, the innate immune response, platelet aggregation, engulfment of apoptotic cells and cell survival. Binding of growth arrest-specific protein 6 (Gas6) activates Axl signaling, but the mechanism of inactivation of the Axl receptor is poorly understood. In the present study, we show that IQGAP1 modulates Axl signaling. IQGAP1 is a scaffold protein that integrates cell signaling pathways by binding several growth factor receptors and intracellular signaling molecules. Our in vitro analysis revealed a direct interaction between the IQ domain of IQGAP1 and Axl. Analysis by both immunoprecipitation and proximity ligation assays demonstrated an association between Axl and IQGAP1 in cells and this interaction was decreased by Gas6. Unexpectedly, reducing IQGAP1 levels in cells significantly enhanced the ability of Gas6 to stimulate both Axl phosphorylation and activation of Akt. Moreover, IQGAP1 regulates the interaction of Axl with the epidermal growth factor receptor. Our data identify IQGAP1 as a previously undescribed suppressor of Axl and provide insight into regulation of Axl function.

关键词： IQ-domain GTPase-activating protein 1   Head and neck squamous cell carcinoma   Prognosis   Transgenic mice   Tissue microarray  

摘要： IQ-domain GTPase-activating protein 1 (IQGAP1) is associated with the development and progression of many human cancers. We aimed to investigate the expression and clinicopathological significances of IQGAP1 in head and neck squamous cell carcinoma (HNSCC). In this study, immunohistochemical staining of IQGAP1, co-inhibitory immune checkpoint molecules and macrophage markers were performed in human HNSCC samples to analyze the expression and correlation with clinicopathological characteristics. Immunoreactivity of IQGAP1 was also detected in immunocompetent mouse HNSCC tissue. We found that IQGAP1 expression level was significantly increased in human HNSCC compared with dysplasia and normal mucosa, and the expression of IQGAP1 in HNSCC was positively associated with advanced lymph node status. Besides, our data indicated that patients with higher IQGAP1 expression exhibited poor overall survival compared with patients with lower IQGAP1 expression. Furthermore, this study demonstrated that IQGAP1 expression was positively associated with TIM3, Galectin-9 (TIM3 ligand), B7H4, macrophage markers CD68 and CD163. In conclusion, these findings suggest that over-expression of IQGAP1 in human HNSCC may indicate poor prognosis.

摘要： Despite major advances in targeted melanoma therapies, drug resistance limits their efficacy. Long noncoding RNAs (lncRNAs) are transcriptome elements that do not encode proteins but are important regulatory molecules. LncRNAs have been implicated in cancer development and response to different therapeutics and are thus potential treatment targets;however, the majority of their functions and molecular interactions remain unexplored. In this study, we identify a novel cytoplasmic intergenic lincRNA (MIRAT), which is upregulated following prolonged MAPK inhibition in NRAS mutant melanoma and modulates MAPK signaling by binding to the MEK scaffold protein IQGAP1. Collectively, our results present MIRAT's direct modulatory effect on the MAPK pathway and highlight the relevance of cytoplasmic lncRNAs as potential targets in drug resistant cancer.

关键词： interleukin 6   IL-6   CDC42   pancreatic cancer   metastasis   cell migration   small GTPase   Janus kinase 2   JAK2   STAT3   IQGAP1  

摘要： Inflammation is a major driver of tumor progression and metastasis, although the mechanisms by which proinflammatory cytokines drive metastatic invasion are unknown. Interleukin-6 (IL-6) is a potent proinflammatory cytokine that is elevated in individuals with pancreatic cancer (PDAC), is required for PDAC progression in mice, and increases tumor cell invasion in vitro. Here, we provide insights into the mechanisms by which IL-6 activates tumor cell invasion. We found that IL-6 stimulation rapidly and robustly activates the small GTPase cell division cycle 42 (CDC42) in human PDAC cells and promotes the formation of premigratory filopodia. The CDC42 activation was required for IL-6-induced invasion as blocking CDC42 activity rendered the cells insensitive to IL-6's proinvasive effects. Loss of Janus kinase 2 (JAK2) or signal transducer and activator of transcription 3 (STAT3) prevented IL-6-mediated CDC42 activation, indicating that IL-6 activates CDC42 through both JAK2 and STAT3. However, the rapid activation of CDC42 suggested that this activation may be distinct from canonical STAT3-mediated transcriptional activation. Importantly, we observed an interaction between STAT3 and IQ motif-containing GTPase-activating protein 1 (IQGAP1), a scaffolding platform that binds CDC42. STAT3 colocalized with CDC42 and IQGAP1 at the plasma membrane, suggesting cross-talk between IL-6-mediated STAT3 signaling and CDC42 activation. These results suggest that IL-6 promotes metastatic invasion, at least partially, through CDC42 and that, along with its pleiotropic effects on tumor growth and progression, IL-6 signaling also activates proinvasive GTPase signaling, priming tumor cells for metastatic invasion.