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. 2000 Aug;47(2):272–276. doi: 10.1136/gut.47.2.272

5'-CpG island methylation of the LKB1/STK11 promoter and allelic loss at chromosome 19p13.3 in sporadic colorectal cancer

J Trojan 1, A Brieger 1, J Raedle 1, M Esteller 1, S Zeuzem 1
PMCID: PMC1727990  PMID: 10896921

Abstract

BACKGROUND—In patients with Peutz-Jeghers syndrome (PJS), causative germline mutations in the LKB1/STK11 gene on chromosome 19p13.3 have been identified. Because of the loss of heterozygosity (LOH) at 19p13.3 in hamartomas and the cancer susceptibility of patients with PJS, LKB1/STK11 is suggested to act as a tumour suppressor. However, the frequency of genetic and epigenetic inactivation of LKB1/STK11 in sporadic tumours is unclear.
AIMS—To investigate the LKB1/STK11 gene for promoter hypermethylation and allelic loss in tumour specimens of patients with sporadic colorectal cancer.
METHODS—DNA from 50 consecutive paraffin embedded sporadic colorectal adenocarcinomas and corresponding normal epithelium was extracted. After bisulphite treatment, specimens were analysed for methylation of the LKB1/STK11 promoter 5'-CpG island by methylation specific polymerase chain reaction (MSP). In addition, tumours were analysed for LOH of chromosome 19p13.3. In tumours exhibiting LOH, LKB1/STK11 was sequenced.
RESULTS—MSP was successful in 48 of 50 tumour specimens. Of those, four (8%) demonstrated hypermethylation of the LKB1/STK11 promoter 5'-CpG island. Moreover, LOH at either D19S886 or D19S878 was observed in five of 38 (13%) informative tumours. All five tumours showing LOH at 19p13.3 were advanced and four of five were located in the left sided colon. There was no correlation between LOH and LKB1/STK11 promoter hypermethylation or somatic mutation.
CONCLUSIONS—In sporadic colorectal cancer, hypermethylation of the LKB1/STK11 promoter and allelic loss at the STK 11 gene locus are rare events. LOH at 19p13.3 was associated with advanced tumour stage and left sided location but not with LKB1/STK11 promoter hypermethylation or somatic mutation.


Keywords: sporadic colorectal adenocarcinoma; tumour suppressor genes; protein-serine-threonine kinase gene LKB1/STK11; promoter hypermethylation; loss of heterozygosity

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Figure 1  .

Figure 1  

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Methylation specific polymerase chain reaction (PCR) of the LKB1/STK11 promoter 5'-CpG island in sporadic colorectal cancer. In the left lane the 140 bp and 100 bp bands of a molecular weight standard are shown. The presence of a visible PCR product in those lanes marked U indicates unmethylated LKB1/STK11 promoter 5'-CpG islands; a visible PCR product in those lanes marked M indicates the presence of methylated LKB1/STK11 promoter 5'-CpG islands. Corresponding lanes are: positive controls for PCR reactions (for the unmethylated reaction DNA extracted from the colorectal cancer cell line HT29 and for the methylated reaction from the colorectal cancer cell line H6), primary colorectal cancers (T100 and T189), and negative controls for PCR reactions.

Figure 2  .

Figure 2  

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Fluorescent analysis of the microsatellite marker D19S878 (centromeric to the LKB1/STK11 gene locus) on chromosome 19p13.3. The 123 bp peak of the size standard is plotted in light grey in all electrophoretic profiles. The polymerase chain reaction (PCR) products of normal colon tissue (upper panels) and corresponding tumour tissue (lower panels) in patient No 31 (A) and No 3 (B) with sporadic colorectal adenocarcinoma were electrophoretically analysed on an automated ABI 310 DNA sequencer (Perkin Elmer). (A) Tumour without allelic loss at D19S878. (B) Tumour exhibiting loss of the 127 bp allele of D19S878 whereas the 134 bp allele is conserved.

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