MGMT protein stoichiometrically repairs O6 -alkylG-DNA adducts. Inactivation of MGMT by promoter-methylation can lead to G to A transition mutations in several genes, including KRAS. Thus, MGMT methylation could be associated with the metastatic process by increasing the rate of mutations. However, this has not yet been convincingly demonstrated in CRCs. Park et al. have reported that MGMT methylation in patients with gastric carcinoma is significantly associated with lymph-node metastasis, tumor stage and disease free survival. However, another study showed significant association between MGMT methylation and improved overall survival in diffuse large B-cell lymphoma. Thus, the relationship between MGMT methylation and metastasis or tumor prognosis might be tissue specific, or possibly coincidental. Our genome-wide analysis of hypermethylated genes at the liver metastatic tumor revealed that 7.4% of the genes showed hypermethylation in the metastatic tumors and 1.3% was commonly hypermethylated among three patients. These GDC-0199 numbers are quite large at face value, but when we validated the data by bisulfite-pyrosequencing, a change in methylation density was the explanation in most cases. One additional clue to explain this finding came from an analysis of resection time differences between the primary and metastatic lesions. Thus, the percentage of hypermethylated genes at liver metastasis was significantly higher in metachronous metastasis than in synchronous metastasis. In one patient, the time between surgery for the primary tumor and the liver metastasis was 46 months and 10.9% of genes analyzed using MCAM showed differential hypermethylation at the liver metastatic tumor. MCAM data in a patient with synchronous metastasis revealed 4.7% differential hypermethylated genes. Given that population doubling is a prime determinant of methylation in normal and neoplastic colon, our data could be explained by continued accumulation of methylation at the metastatic site. Overall, looking at methylation frequency, we find few differences between primary tumors and liver metastases, suggesting that aberrant DNA methylation is a very early event and that tumor cells acquire methylation changes before progression to liver metastasis. We cannot exclude the possibility that a few rare genes are highly selected for during the process of metastasis, but discovering these will require whole-genome methylation analysis technology that is more quantitative than what is currently available. In summary, our results indicate that methylation frequency between primary tumors and matched liver metastasis is similar, suggesting that tumor cells acquire methylation changes before progression to liver metastasis. While we cannot rule out rare consistent changes, it appears that DNA methylation frequency is very stable over time in CRC. In many areas of science, especially in biotechnology, the number of high-dimensional datasets recording multiple aspects of a single phenomenon is increasing.