Tamura G, Sakata K, Nishizuka S, Maesawa C, Suzuki Y, Iwaya T, Terashima M, Saito K, Satodate R
Department of Pathology, Iwate Medical University School of Medicine, Uchimaru, Morioka.
Jpn J Cancer Res. 1996 Nov;87(11):1153-9. doi: 10.1111/j.1349-7006.1996.tb03125.x.
We investigated the E (epithelial)-cadherin gene for mutations and loss of heterozygosity (LOH) in 24 primary gastric carcinomas (12 differentiated and 12 undifferentiated types, including 3 signet-ring cell carcinomas), as well as 4 gastric carcinoma cell lines of the undifferentiated type (MKN-45, GCIY, HGC-27 and GT3TKB). We utilized PCR-SSCP and RT-PCR followed by direct sequencing to detect gene mutations and skipped exons, and RT-PCR-SSCP to examine LOH. In primary carcinomas, gene mutations or skipped exons were detected in 4 of 9 (44%) undifferentiated carcinomas of the scattered type, including 2 signet-ring cell carcinomas, and in none of the 3 undifferentiated carcinomas of the adherent type and 12 differentiated carcinomas. Demonstrated mutations of the E-cadherin gene included an 18 bp deletion (codon 418-423) and a 3 bp deletion (codon 400, calcium-binding domain), both located in exon 9. Skipping of exon 9 with a 1 bp insertion at codon 337, and skipping of exon 8 with a 1 bp deletion at codon 336, also were detected. LOH was confirmed in all of the carcinomas in which gene mutations or skipped exons (3/3 informative cases) were demonstrated. The MKN-45 cell line exhibited an 18 bp deletion at the exon 6-intron 6 boundary with loss of the wild-type allele, and 2 of the remaining 3 cell lines (HGC-27 and GT3TKB) had lost expression without detectable structural alteration of the E-cadherin gene. These data provide support for classic two-hit inactivation of the E-cadherin gene in a high percentage of undifferentiated carcinomas of the scattered type.
我们研究了24例原发性胃癌（12例分化型和12例未分化型，包括3例印戒细胞癌）以及4例未分化型胃癌细胞系（MKN-45、GCIY、HGC-27和GT3TKB）中E（上皮）-钙黏蛋白基因的突变和杂合性缺失（LOH）情况。我们采用聚合酶链反应-单链构象多态性分析（PCR-SSCP）和逆转录聚合酶链反应（RT-PCR）后直接测序来检测基因突变和外显子跳跃，并采用RT-PCR-SSCP检测LOH。在原发性癌中，9例散在型未分化癌中的4例（44%）检测到基因突变或外显子跳跃，其中包括2例印戒细胞癌，而3例附着型未分化癌和12例分化型癌均未检测到。已证实的E-钙黏蛋白基因突变包括18 bp缺失（密码子418-423）和3 bp缺失（密码子400，钙结合结构域），均位于外显子9。还检测到外显子9跳跃且在密码子337处有1 bp插入，以及外显子8跳跃且在密码子336处有1 bp缺失。在所有证实有基因突变或外显子跳跃的癌中（3/3例信息充分的病例）均确认存在LOH。MKN-45细胞系在外显子6-内含子6边界处有18 bp缺失且野生型等位基因丢失，其余3例细胞系中的2例（HGC-27和GT3TKB）E-钙黏蛋白基因无结构改变但表达缺失。这些数据支持了在高比例的散在型未分化癌中E-钙黏蛋白基因经典的双打击失活机制。
