Yen Chueh-Chuan, Chen Yann-Jang, Lu Kai-Hsi, Hsia Jiun-Yi, Chen Jung-Ta, Hu Cheng-Po, Chen Po-Min, Liu Jin-Hwang, Chiou Tzeon-Jye, Wang Wei-Shu, Yang Muh-Hwa, Chao Ta-Chung, Lin Chi-Hung
Division of Medical Oncology, Department of Medicine, Taipei Veterans General Hospital, Taipei, Taiwan.
Int J Oncol. 2003 Oct;23(4):871-81.
We performed an integrated cytogenetic study using a combination of comparative genomic hybridization (CGH), spectral karyotyping (SKY) and fluorescence in situ hybridization (FISH) to analyze chromosomal aberrations associated with 8 human esophageal squamous cell carcinoma (EC-SCC) cell lines, and used real-time quantitative PCR (Q-PCR) to study the copy number changes of two candidate genes of chromosome 3q, PIK3CA and TP63, in 20 primary tumors of EC-SCC. The pooled CGH results revealed frequent gain abnormalities on chromosome arms 1p, 1q, 3q, 5p, 6p, 7p, 7q, 8q, 9q, 11q, 12p, 14q, 15q, 16p, 16q, 17q, 18p, 19q, 20q, 22q, and Xq, while frequent losses were found on 3p, 4, 5q, 6q, 7q, 9p, and 18q. SKY detected 195 translocations, 13 deletions and 2 duplications. Among the 374 breakpoints, most clustered at the centromeric regions, such as 8q10, 13q10, 7q10, 9q10, 14q10, 15q10, 16q10, 21q10, and 22q10, but also at other regions, including 3q (3q21, 3q22, 3q25), 7p (7p22, 7p14, 7p12), 7q (7q21, 7q31, 7q32), 8q (8q21.1, 8q23), 11q (11q21, 11q24), 13q (13q14) and 18q (18q21). There was a good correlation between the number of aberrations identified by CGH and SKY (r=0.667; p=0.035). Combined CGH and SKY analyses indicated that chromosomes 3, 7, 9, 11, 14, 16, 18, 19, 20, and 22 harbored higher frequency of chromosomal aberrations than expected. FISH using BAC clones containing oncogene PIK3CA and TP63 found that both genes were amplified in 6 and 5 cell lines, respectively. Q-PCR analysis of primary tumors revealed amplification of PIK3CA and TP63 in 100% and 80% of the cases. Average copy number of PIK3CA per haploid genome was greater than that of TP63 (6.27 vs 2.73), and the difference showed statistical significance (p<0.001). Combination of CGH, SKY and FISH could reveal detailed chromosomal changes associated with esophageal cancer cells, and Q-PCR could assess the change of the candidate genes in clinical samples in a high throughput way.
我们采用比较基因组杂交（CGH）、光谱核型分析（SKY）和荧光原位杂交（FISH）相结合的方法，对8种人食管鳞状细胞癌（EC - SCC）细胞系进行了综合细胞遗传学研究，分析与之相关的染色体畸变情况，并运用实时定量PCR（Q - PCR）研究了20例原发性EC - SCC肿瘤中3号染色体上两个候选基因PIK3CA和TP63的拷贝数变化。汇总的CGH结果显示，1p、1q、3q、5p、6p、7p、7q、8q、9q、11q、12p、14q、15q、16p、16q、17q、18p、19q、20q、22q和Xq染色体臂上频繁出现增益异常，而3p、4、5q、6q、7q、9p和18q上则频繁出现缺失。SKY检测到195处易位、13处缺失和2处重复。在374个断点中，大多数聚集在着丝粒区域，如8q10、13q10、7q10、9q10、14q10、15q10、16q10、21q10和 22q10，但也出现在其他区域，包括3q（3q21、3q22、3q25）、7p（7p22、7p14、7p12）、7q（7q21、7q31、7q32）、8q（8q21.1、8q23）、11q（11q21、11q24）、13q（13q14）和18q（18q21）。CGH和SKY所识别的畸变数量之间存在良好的相关性（r = 0.667；p = 0.035）。CGH和SKY联合分析表明，3、7、9、11、14、16、18、19、20和22号染色体上染色体畸变的频率高于预期。使用包含癌基因PIK3CA和TP63的BAC克隆进行FISH发现，这两个基因分别在6个和5个细胞系中发生扩增。对原发性肿瘤的Q - PCR分析显示，PIK3CA和TP63在100%和80%的病例中发生扩增。单倍体基因组中PIK3CA的平均拷贝数大于TP63（6.27对2.73），差异具有统计学意义（p < 0.001）。CGH、SKY和FISH相结合能够揭示与食管癌细胞相关的详细染色体变化，而Q - PCR能够以高通量方式评估临床样本中候选基因的变化。
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