Comparison of Cancer-Associated Genetic Abnormalities in Columnar-Lined Esophagus with and without Goblet Cells. Homo sapiens

The requirement of goblet cells (intestinal metaplasia; IM) to define Barrett’s esophagus (BE) remains controversial as some studies have shown that the risk of progression to esophageal adenocarcinoma (EAC) is similar for non-goblet cell metaplasia (NGM) and IM. IM harbors genomic aberrations in known cancer-associated genes and these have been linked to increased EAC risk. No detailed studies have explored DNA alterations in NGM. We hypothesized that a comparison of DNA alterations in cancer-associated genes in IM and NGM samples would provide insight into their relative risks for progression to EAC. Patient biopsies were analyzed using Affymetrix SNP 6.0 arrays, FISH, and targeted resequencing of 20 frequently mutated EAC genes. Frequent CNA’s targeting cancer-associated genes were found in IM whereas no such changes were observed in NGM. In one subject, FISH confirmed loss of CDKN2A and amplification of chromosome 8 in IM but not in a nearby NGM biopsy. Targeted sequencing in a subset of metaplasia tissues revealed 11 non-synonymous mutations in 16 IM samples and 2 mutations in 19 NGM samples. Our results show that IM has a much higher frequency of cancer-associated mutations than NGM and is therefore likely to pose a higher risk for development of EAC. Overall design: 57 non-goblet metaplasia, intestinal metaplasias, and composite specimens from 45 subjects were studied. DNA copy number abnormalities (CNA’s) were identified using Affymetrix arrays and fluorescence in situ hybridization (FISH). Data was processed in Nexus 6.0 (BioDiscovery, CA). Targeted sequencing of all exons from twenty genes found to be frequently mutated in EAC was performed on metaplasia samples using Ion AmpliSeq™ DNA sequencing.

杯状细胞（goblet cells）相关的肠上皮化生（intestinal metaplasia, IM）作为巴雷特食管（Barrett’s esophagus, BE）的定义标准仍存在争议，因为部分研究显示，非杯状细胞化生（non-goblet cell metaplasia, NGM）与IM患者进展为食管腺癌（esophageal adenocarcinoma, EAC）的风险相似。IM在已知的癌症相关基因中存在基因组畸变，且这些畸变与EAC风险升高相关。目前尚无针对NGM中DNA改变的详细研究。我们假设，对比IM与NGM样本中癌症相关基因的DNA改变情况，可为二者进展为EAC的相对风险提供研究依据。患者活检样本采用Affymetrix SNP 6.0芯片、荧光原位杂交（fluorescence in situ hybridization, FISH）以及20种常见突变EAC基因的靶向重测序进行分析。研究发现，IM中存在针对癌症相关基因的高频染色体拷贝数异常（copy number aberrations, CNA），而NGM中未观察到此类改变。在1例受试者中，FISH证实IM样本存在CDKN2A基因缺失及8号染色体扩增，而邻近的NGM活检样本未出现此类改变。对部分化生组织的靶向测序结果显示，16例IM样本中检出11个非同义突变，19例NGM样本中仅检出2个突变。本研究结果表明，IM的癌症相关突变频率远高于NGM，因此其进展为EAC的风险可能更高。总体实验设计：本研究纳入45名受试者的57份非杯状化生、肠上皮化生及复合活检标本。通过Affymetrix芯片与荧光原位杂交（FISH）鉴定DNA拷贝数异常（CNA），数据采用Nexus 6.0（美国加利福尼亚州BioDiscovery公司）进行处理。采用Ion AmpliSeq™ DNA测序技术，对EAC中常见突变的20种基因的全部外显子进行靶向测序，以分析化生组织样本。