【作者】 梁建伟；

【导师】 赵平； 王明荣； 周志祥；

【作者基本信息】 中国协和医科大学 ， 肿瘤学， 2009， 博士

【摘要】 【背景与目的】基因组DNA的非随机改变在胰腺导管腺癌(PDAC)的发生发展中起重要的作用,既往研究多局限于以细胞系为研究对象,利用BAC探针的array CGH鉴定基因组DNA非随机改变。本研究通过寡核苷酸array CGH分析胰腺导管腺癌新鲜组织标本,寻找胰腺导管腺癌特异的DNA改变,筛选出可能与胰腺导管腺癌发生发展密切相关的DNA拷贝数改变和相应的基因。【材料与方法】收集2007年6月至2008年12月PDAC患者手术切除肿瘤组织标本15例,利用显微切割技术,富集肿瘤细胞,提取基因组DNA,通过array CGH分析,鉴定PDAC的基因拷贝数改变。应用荧光原位杂交(FISH)和实时定量PCR方法验证array CGH的结果。收集1999年～2008年间手术切除的PDAC65例,慢性胰腺炎8例,构建组织芯片,应用免疫组织化学(IHC)方法检测在array CGH中鉴定出的扩增基因PSCA和HMGA2的表达情况,分析其与PDAC临床病理参数的关系。【结果】Array CGH结果中,有11个增益频率在80%以上的染色体区段,分别为:1q31.3、3p22.1、3p22.31、3p22.3、8q24.3、10q11.23、14q12、16p13.3、17q24.1、Xp11.21和Xq28,其中包括畸变频率＞20%的高拷贝扩增基因80个。发现5个缺失频率≥80%的染色体区段,分别为:5p13.3、12q13.13、17p13.1、17q21.31、Xq11.1,其中包含畸变频率＞20%的缺失基因144个。Array CGH、FISH和q-PCR实验均表明8q24.3的PSCA基因扩增。IHC结果提示:PSCA和HMGA2在PDAC中的阳性表达率分别为67.2%和76.7%。PSCA和HMGA2的表达与患者年龄、性别、T分期、M分期、分化程度及预后无相关性,但均与淋巴结转移相关(PSCA:x~2=4.370,P=0.037;HMGA2:x~2=13.062,P=0.001)。【结论】PDAC基因拷贝数改变频率高、位点多,PSCA和HMGA2扩增和过表达可能作为PDAC淋巴结转移的分子标志。更多还原

【Abstract】 【Background and Purpose】DNA copy number alterations play an important role in the initiation and progression of pancreatic ductal adenocarcinoma(PDAC).There have been some reports on DNA changes of PDAC detected by BAC-based array comparative genomic hybridization(array CGH),but most of them were performed on pancreatic cancer cell lines.In the present study,we used oligonucleotide array CGH to identify genetic alterations of PDAC genome.【Patients and Methods】The tissue samples were obtained from 15 PDAC.Tumor cells collected by microdissection and genomic DNA were isolated.Array CGH was performed with the Agilent Human Genome CGH 44B array.DNA copy number alterations of PDAC were validated by fluorescence in situ hybridization(FISH) and real-time PCR.Tissue microarray was constructed,containing 65 PDAC and 8 chronic pancreatitis tissues.The expression of PSCA and HMGA2 proteins were detected by using immunohistochemical method and the relationships between the proteins expression and clinicopathological parameters of PDAC were analyzed.【Results】Profiles of 15 PDAC identified multiple chromosomal regions with DNA copy number alterations.Loci with(＞80%cases) gains were at 1q31.3,3p22.1,3p22.31, 3p22.3,8q24.3,10q11.23,14q12,16p13.3,17q24.1,Xp11.21,and Xq28.Eighty genes were amplified in≥4 out of 15 cases(＞20%).The most frequent losses(＞80%) were at 5p13.3,12q13.13,17p13.1,17q21.31,and Xq11.1.One hundred and forty-four loci with deletions were detected(＞20.0%).Q-PCR and FISH results confirmed that the PSCA gene was of amplification.IHC analysis showed that PSCA and HMGA2 expressed in 67.2%and 76.7%of tumors,respectively.The positive expression of PSCA and HMGA2 was associated with lymph node metastases(PSCA:x~2=4.370,P=0.037;HMGA2:x~2= 13.062,P=0.001).【Conclusions】There existed lots of chromosomal loci with frequent alterations of DNA copy number in PDAC.PSCA and HMGA2 might be two useful biomarkers in more advanced stage of PDAC.更多还原