Kölble K, Barthel B, Ullrich O, Pidde H, Döhring C, Rüschoff J, Schlag P M, Dietel M
Institut für Pathologie, Universitätsklinikum Charité, Humboldt-Universität, Berlin.
Verh Dtsch Ges Pathol. 2000;84:182-6.
Various inherited and acquired alterations affecting the genes and gene products of the WNT pathway appear to be involved in the different molecular routes leading to colorectal cancer (CRC). This study was initiated to investigate the prevalence of somatic mutations in the beta-catenin gene (CTNNB1) and the associated pathology in CRC with defective DNA mismatch repair.
Paraffin and/or frozen sections of 33 primary CRC (including any liver and lymph node metastases present) with high-grade microsatellite instability (MSI-H; i.e. with > or = 5 unstable microsatellite markers of 10 tested) were polytopically fractionated by microdissection. Genomic and c-DNA samples were sequenced across exons 2-4 of CTNNB1 and the expression patterns of beta-catenin (beta-C) analyzed by immunohistology and Western blotting.
Seven somatic mutations affecting phosphorylation sites of exon 3 (2 deletions also encompassing parts of either intron 2 or exon 4 [delta X2/3 bzw. delta X3/4] and 5 missense mutations [2 x T41A, 2 x S45F, S45P]) were identified. Two mutations (delta X3/4 and S45F) were concordantly present in CRC primaries and their respective metastases whereas the S45P mutation was restricted to a hepatic metastasis. In the delta X2/3 CRC primary only a shortened 66 kD CTNNB1 gene product was present while its associated liver metastasis showed a total loss of beta-C expression.
Both exon 3 and the entire locus coding for beta-C are somatically altered in approximately 20% of CRC with MSI-H at different stages of tumor progression. Thus CTNNB1 appears to be a genomic target for complex oncogenic mutations and deletional processes in a substantial fraction of this molecular subset of CRC.
各种影响WNT信号通路基因及基因产物的遗传性和获得性改变，似乎参与了导致结直肠癌（CRC）的不同分子途径。开展本研究以调查β-连环蛋白基因（CTNNB1）的体细胞突变发生率以及DNA错配修复缺陷的CRC中的相关病理情况。
对33例伴有高度微卫星不稳定（MSI-H，即10个检测的微卫星标记中有≥5个不稳定微卫星标记）的原发性CRC（包括存在的任何肝和淋巴结转移灶）的石蜡切片和/或冰冻切片进行显微切割多区域分离。对CTNNB1基因外显子2-4进行基因组和c-DNA样本测序，并通过免疫组织化学和蛋白质免疫印迹分析β-连环蛋白（β-C）的表达模式。
鉴定出7个影响外显子3磷酸化位点的体细胞突变（2个缺失，也包括内含子2或外显子4的部分区域[ΔX2/3或ΔX3/4]以及5个错义突变[2×T41A、2×S45F、S45P]）。2个突变（ΔX3/4和S45F）在原发性CRC及其各自的转移灶中一致存在，而S45P突变仅限于肝转移灶。在ΔX2/3原发性CRC中仅存在一种缩短的66 kD CTNNB1基因产物，而其相关的肝转移灶显示β-C表达完全缺失。
在肿瘤进展的不同阶段，约20%的MSI-H CRC中，外显子3和编码β-C的整个基因座均发生体细胞改变。因此，在相当一部分这种分子亚型的CRC中，CTNNB1似乎是复杂致癌突变和缺失过程的基因组靶点。
