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CpG island methylation is a common finding in colorectal cancer cell lines.

Suter CM, Norrie M, Ku SL, Cheong KF, Tomlinson I, Ward RL - Br. J. Cancer (2003)

Bottom Line: Overall only one cell line, SW1417, did not show methylation at any locus.Methylation was found with equal frequency in MSI and chromosomally unstable lines.We conclude that CpG island hypermethylation, whether acquired in vivo or in culture, is a ubiquitous phenomenon in CRC cell lines.

View Article: PubMed Central - PubMed

Affiliation: Department of Medical Oncology, St Vincent's Hospital, Darlinghurst, NSW, Australia.

ABSTRACT
Tumour cell lines are commonly used in colorectal cancer (CRC) research, including studies designed to assess methylation defects. Although many of the known genetic aberrations in CRC cell lines have been comprehensively described, no studies have been performed on their methylation status. In this study, 30 commonly used CRC cell lines as well as seven primary tumours from individuals with hereditary nonpolyposis colorectal cancer (HNPCC) were assessed for methylation at six CpG islands known to be hypermethylated in colorectal cancer: hMLH1, p16, methylated in tumour (MINT-)-1, -2, -12 and -31. The cell lines were also assessed for microsatellite instability (MSI), ploidy status, hMLH1 expression, and mutations in APC and Ki-ras. Methylation was frequently observed at all examined loci in most cell lines, and no differences were observed between germline-derived and sporadic cell lines. Methylation was found at MINT 1 in 63%, MINT 2 in 57%, MINT 12 in 71%, MINT 31 in 53%, p16 in 71%, and hMLH1 in 30% of cell lines. Overall only one cell line, SW1417, did not show methylation at any locus. Methylation was found with equal frequency in MSI and chromosomally unstable lines. MSI was over-represented in the cell lines relative to sporadic CRC, being detected in 47% of cell lines. The rate of codon 13 Ki-ras mutations was also over three times that expected from in vivo studies. We conclude that CpG island hypermethylation, whether acquired in vivo or in culture, is a ubiquitous phenomenon in CRC cell lines. We suggest that CRC cell lines may be only representative of a small subset of real tumours, and this should be taken into account in the use of CRC cell lines for epigenetic studies.

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Detection of mutations in codon 13 of Ki-ras in colorectal cell lines. RFLP analysis was performed on Ki-ras PCR amplicons with the restriction enzyme BslI. Mutations in the first or second base of codon 13 will abolish the BslI site; therefore Ki-ras codon 13 mutants are resistant to digestion. Wild-type (WT) amplicons, and mutants other than codon 13, are fully digested by BslI. Undigested amplicons are shown in the last lane. Both LIM1215 and RKO have WT Ki-ras; LS174T is a codon 12 mutant; HCT116, displaying undigested product, is a codon 13 mutant. MW is pUC19/MspI.
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fig3: Detection of mutations in codon 13 of Ki-ras in colorectal cell lines. RFLP analysis was performed on Ki-ras PCR amplicons with the restriction enzyme BslI. Mutations in the first or second base of codon 13 will abolish the BslI site; therefore Ki-ras codon 13 mutants are resistant to digestion. Wild-type (WT) amplicons, and mutants other than codon 13, are fully digested by BslI. Undigested amplicons are shown in the last lane. Both LIM1215 and RKO have WT Ki-ras; LS174T is a codon 12 mutant; HCT116, displaying undigested product, is a codon 13 mutant. MW is pUC19/MspI.

Mentions: A remarkably high rate of Ki-ras mutations was found in this data set. Of the 30 cell lines, 19 (63%) were found to have a point mutation in either codon 12 or codon 13 of Ki-ras (Figure 3Figure 3


CpG island methylation is a common finding in colorectal cancer cell lines.

Suter CM, Norrie M, Ku SL, Cheong KF, Tomlinson I, Ward RL - Br. J. Cancer (2003)

Detection of mutations in codon 13 of Ki-ras in colorectal cell lines. RFLP analysis was performed on Ki-ras PCR amplicons with the restriction enzyme BslI. Mutations in the first or second base of codon 13 will abolish the BslI site; therefore Ki-ras codon 13 mutants are resistant to digestion. Wild-type (WT) amplicons, and mutants other than codon 13, are fully digested by BslI. Undigested amplicons are shown in the last lane. Both LIM1215 and RKO have WT Ki-ras; LS174T is a codon 12 mutant; HCT116, displaying undigested product, is a codon 13 mutant. MW is pUC19/MspI.
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Related In: Results  -  Collection

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getmorefigures.php?uid=PMC2747532&req=5

fig3: Detection of mutations in codon 13 of Ki-ras in colorectal cell lines. RFLP analysis was performed on Ki-ras PCR amplicons with the restriction enzyme BslI. Mutations in the first or second base of codon 13 will abolish the BslI site; therefore Ki-ras codon 13 mutants are resistant to digestion. Wild-type (WT) amplicons, and mutants other than codon 13, are fully digested by BslI. Undigested amplicons are shown in the last lane. Both LIM1215 and RKO have WT Ki-ras; LS174T is a codon 12 mutant; HCT116, displaying undigested product, is a codon 13 mutant. MW is pUC19/MspI.
Mentions: A remarkably high rate of Ki-ras mutations was found in this data set. Of the 30 cell lines, 19 (63%) were found to have a point mutation in either codon 12 or codon 13 of Ki-ras (Figure 3Figure 3

Bottom Line: Overall only one cell line, SW1417, did not show methylation at any locus.Methylation was found with equal frequency in MSI and chromosomally unstable lines.We conclude that CpG island hypermethylation, whether acquired in vivo or in culture, is a ubiquitous phenomenon in CRC cell lines.

View Article: PubMed Central - PubMed

Affiliation: Department of Medical Oncology, St Vincent's Hospital, Darlinghurst, NSW, Australia.

ABSTRACT
Tumour cell lines are commonly used in colorectal cancer (CRC) research, including studies designed to assess methylation defects. Although many of the known genetic aberrations in CRC cell lines have been comprehensively described, no studies have been performed on their methylation status. In this study, 30 commonly used CRC cell lines as well as seven primary tumours from individuals with hereditary nonpolyposis colorectal cancer (HNPCC) were assessed for methylation at six CpG islands known to be hypermethylated in colorectal cancer: hMLH1, p16, methylated in tumour (MINT-)-1, -2, -12 and -31. The cell lines were also assessed for microsatellite instability (MSI), ploidy status, hMLH1 expression, and mutations in APC and Ki-ras. Methylation was frequently observed at all examined loci in most cell lines, and no differences were observed between germline-derived and sporadic cell lines. Methylation was found at MINT 1 in 63%, MINT 2 in 57%, MINT 12 in 71%, MINT 31 in 53%, p16 in 71%, and hMLH1 in 30% of cell lines. Overall only one cell line, SW1417, did not show methylation at any locus. Methylation was found with equal frequency in MSI and chromosomally unstable lines. MSI was over-represented in the cell lines relative to sporadic CRC, being detected in 47% of cell lines. The rate of codon 13 Ki-ras mutations was also over three times that expected from in vivo studies. We conclude that CpG island hypermethylation, whether acquired in vivo or in culture, is a ubiquitous phenomenon in CRC cell lines. We suggest that CRC cell lines may be only representative of a small subset of real tumours, and this should be taken into account in the use of CRC cell lines for epigenetic studies.

Show MeSH
Related in: MedlinePlus