From Chromosome Aberrations to Cancer-Related Genes Identification: an Integrated Genomic Study for Squamous Cell Carcinoma of Esophagus

• Main Authors: Chueh-Chuan Yen, 顏厥全
• Other Authors: Po-Min Chen
• Format: Others
• Language: zh-TW
• Published: 2003
• Online Access: http://ndltd.ncl.edu.tw/handle/92129039036977341360

博士 === 國立陽明大學 === 臨床醫學研究所 === 91 === ENGLISH ABSTRACT By using state-of-the-art molecular cytogenetic techniques, we analyzed the chromosomal changes of one of the most important cancers in Taiwan-esophageal squamous cell carcinoma (EC-SCC). Then we used real-time quantitative polymerase chain reaction (Q-PCR) to assess the copy number changes of candidate genes in regions of chromosomal aberrations. We used comparative genomic hybridization (CGH) to analyze 46 samples of EC-SCC. Total 321 gains and 252 losses were found, and the average gains and losses per patient were 6.98 and 5.47, respectively. Frequent gains were found on chromosome 1q, 2q, 3q, 5p, 7p, 7q, 8q, 11q, 12p, 12q, 14q, 17q, 20q, and Xq. Frequent deletions were found on 1p, 3p, 4p, 5q, 8p, 9p, 9q, 11q, 13q, 16p, 17p, 18q, 19p, and 19q. Deletion of 4p and 13q12-q14 and gain of 5p were significantly correlated with pathologic staging. Loss of 8p22-pter and 9p were more easily found in advanced disease. Gain of 8q24-qter was more frequently seen in grade 3 tumor. A univariate analysis found that pathologic staging; gain of 5p and 7q; and deletion of 4p, 9p, and 11q were significant prognostic factors. However, pathologic staging became the only significant prognostic factor in a multi-variate analysis. We then combined CGH, spectral karyotyping (SKY) and fluorescence in situ hybridization (FISH) to perform a comprehensive analysis of 8 EC-SCC cell lines. The pooled CGH results were similar with clinical study. SKY detected 195 translocations, 13 deletions and 2 duplications. Among the 374 breakpoints found, 314 (84%) could be mapped by combining SKY and DAPI banding pattern. Most of them clustered at the centromeric regions, but some were found 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 aberrations than expected. We could choose appropriate bacterial artificial chromosome (BAC) clones from the hint of CGH to perform FISH for refinement of rearrangement. Gain of 3q was frequently found in EC-SCC, and the minimal common region identified by CGH is 3q26-qter. We used Q-PCR to measure copy number changes of candidate genes in this region. We first showed good correlation between FISH and Q-PCR in copy number assessment in cell lines. Next we showed different amplification pattern of PIK3CA and TP63 in 20 tumor samples known to have 3q amplification. Then we analyzed the copy number change of 13 candidate genes among 59 tumor samples. We found that there is a great variation of copy number changes of these genes, with highest copy number of hTERC, followed by PIK3CA and ECT2, then by TP63 and SSR3.CCNL1 and EIF4G1showed little changes in copy number. Cases with lymph node metastasis had lower copy number of SMC4L1, SKIL, EIF5A2 and TP63 than those without; cases with T3 tumor had higher SSR3 copy number; and copy number of CCNL1 and TP63 is lower in overall advanced stage tumor. There is no correlation of copy number changes of these 13 genes with tumor differentiation. In summary, novel chromosomal changes in EC-SCC were found by CGH, and some of them were associated with disease progression. CGH, SKY and FISH provide a comprehensive analysis of chromosomal changes in EC-SCC. For candidate genes on chromosome 3q, Q-PCR disclosed a great variation of amplification. Some genes were found with different copy number in different stages. Some amplified genes that have not been explored in cancer, such as ECT2 and SSR3, deserved further study.