The associations between methylation frequencies at CIMP-related markers and MSI or MSI-/LOH- sporadic CRCs suggest that the majority of these tumors evolve through CIMP. These findings suggest that CIN and CIMP represent 2 independent and inversely related mechanisms of genetic and epigenetic instability in sporadic CRCs and confirm that MSI cancers arise as a consequence of CIMP.

Loss of PTEN tumor suppressor function is observed in tumors of breast, prostate, thyroid, and endometrial origin. Allelic losses in the proximity of the PTEN locus (10q23) also occur in sporadic colorectal cancers (CRCs), but biallelic inactivation of this site has not been frequently demonstrated. We hypothesized that alternative mechanisms of PTEN allelic inactivation, such as promoter hypermethylation, might be operative in CRC and that PTEN inactivation may be related to recognized forms of genomic instability. We characterized a cohort of 273 sporadic CRCs by determining their microsatellite instability (MSI) status. Of these, 146 cancers were examined for PTEN promoter methylation by methylation-specific PCR. Mutations at the poly(A) 6 repeat sequences in PTEN exons 7 and 8 and deletions at the 10q23 locus were also identified using microsatellite analysis. The presence of PTEN protein was determined by immunostaining, and the results were correlated with the promoter methylation status. We observed that PTEN promoter hypermethylation was a frequent occurrence in MSI-high (MSI-H) tumors (19.1% of MSI-H versus 2.2% of MSI-low/microsatellite stable tumors; P ‫؍‬ 0.002). A PTEN mutation or a deletion event was present in 60% of the tumors with promoter region hypermethylation. Hypermethylation of the PTEN promoter correlated significantly with either decreased or complete loss of PTEN protein expression (P ‫؍‬ 0.004). This is the first demonstration of PTEN inactivation as a result of promoter hypermethylation in MSI-H sporadic CRCs. These data suggest that this silencing mechanism plays a major role in PTEN inactivation and, in colon cancer, may be more important than either allelic losses or inactivating mutations. The significant correlation of PTEN hypermethylation with MSI-H tumors further suggests that PTEN is an additional important "target" of methylation along with the hMLH1 gene in the evolution of MSI-H CRCs and also confers the "second hit" in the biallelic inactivation mechanism for some proportion of tumors.

Key words: RUNX3; methylation; colon cancerColorectal cancer (CRC) is one of the leading causes of cancerrelated deaths in the United States. In its early stages, colon carcinogenesis is associated with alterations in molecular signaling pathways and then progresses as a result of a sequential accumulation of events that either activate oncogenes or inactivate tumor suppressor genes. 1 Some alterations participating in multistep colon carcinogenesis affect APC, K-Ras, p53 and elements of the TGF-␤ signaling pathway, such as TGF␤RII and SMAD4. The TGF-␤ signaling pathway serves as a major tumor suppressor in a variety of human gastrointestinal tumors, including colon, gastric and pancreatic cancer. 2 The downstream transcriptional targets of the TGF-␤ signaling pathways are key mediators for regulating cellular proliferation, extracellular matrix production and immune surveillance, and also dictate gastrointestinal epithelial development.The runt-domain related (RUNX) transcription factors are mammalian homologues of the Drosophila genes runt and lozenge, and are heterodimeric proteins composed of 2 highly conserved DNA binding subunits, ␣ and ␤. 3 RUNX transcription factors are one of the important targets of TGF-␤ superfamily signaling and play critical functions in mammalian development. RUNX proteins have been shown to interact with downstream SMAD proteins in mediating the growth suppressive effects of TGF-␤ and play an important role in development and oncogenesis. 3 Three mammalian runt-related genes, RUNX1, RUNX2 and RUNX3 have been described. RUNX1 is an indispensable factor in hematopoiesis and angiogenesis, and anomalies in this gene are involved in about 30% of the cases of human acute leukemia. 4 RUNX2 collaborates with c-myc and Pim-1, and its over-expression in mice predisposes to the development of T-cell lymphoma. 5,6 RUNX3 is a putative tumor suppressor gene localized to chromosome 1p36, a region that exhibits frequent loss of heterozygosity (LOH) events in colon, gastric, breast and ovarian cancers. More recently, silencing of RUNX3 has been reported in gastrointestinal cancers in mice as well as humans. 7-10 Li and colleagues 8 demonstrated that the gastric mucosa of the RUNX3-null mouse exhibited hyperplasia due to stimulated proliferation and suppressed apoptosis of the epithelial cells. These cells were resistant to growth-inhibitory effects of TGF-␤, indicating that RUNX3 regulates the growth of gastric epithelial cells. 8 Physical interactions between the C-terminus of RUNX3 and SMAD that mediate transmission of TGF-␤ induced growth inhibitory signals to the nucleus have been already described. 11 Because the TGF-␤ mediated signaling pathway regulates cell growth in the human colon, it is possible that RUNX3 plays an important tumor suppressor role. In our study, we determined the prevalence of RUNX3 expression and its promoter methylation in human colon cancer cell lines and primary cancers, and defined the association of these events with MSI and TGF␤RII status. We further demonstrate t...

High-quality biospecimens with appropriate clinical annotation are critical in the era of personalized medicine. It is now widely recognized that biospecimen resources need to be developed and operated under established scientific, technical, business, and ethical/legal standards. To date, such standards have not been widely practiced, resulting in variable biospecimen quality that may compromise research efforts. The National Cancer Institute (NCI) Office of Biorepositories and Biospecimen Research (OBBR) was established in 2005 to coordinate NCI's biospecimen resource activities and address those issues that affect access to the high-quality specimens and data necessary for its research enterprises as well as the broader translational research field. OBBR and the NCI Biorepository Coordinating Committee developed NCI's "Best Practices for Biospecimen Resources" after consultation with a broad array of experts. A Biospecimen Research Network was established to fund research to develop additional evidence-based practices. Although these initiatives will improve the overall availability of high-quality specimens and data for cancer research, OBBR has been authorized to implement a national biobanking effort, cancer HUman Biobank (caHUB). caHUB will address systematically the gaps in knowledge needed to improve the state-of-the-science and strengthen the standards for human biobanking. This commentary outlines the progressive efforts by NCI in technical, governance, and economic considerations that will be important as the new caHUB enterprise is undertaken.