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Silencing of CDX2 expression in colon cancer via a dominant repression pathway
Takao Hinoi1, Massimo Loda, Eric R Fearon
1Department of Internal Medicine, University of Michigan Medical School, Ann Arbor, Michigan 48109-0638, USA.
Abstract:
CDX2 is a caudal-related homeobox transcription factor whose expression in the adult is normally restricted to intestinal epithelium. Mice heterozygous for germ line Cdx2 inactivation develop intestinal polyps, and the lesions lack Cdx2 expression. Prior studies indicate some human colon carcinomas also lack CDX2 expression. To address the role of CDX2 defects in colon cancer development, we analyzed CDX2 expression in 45 primary colorectal carcinomas. Four carcinomas lacked CDX2 expression, and three others showed aberrant cytoplasmic localization of CDX2, although no significant CDX2 gene defects were seen in the seven tumors. Marked reductions in CDX2 transcript and protein levels were seen in five of 13 colorectal cell lines, and nuclear run-off data indicated reduced transcription was a major factor in CDX2 silencing. Treatment with the DNA demethylating agent 5-aza-2'-deoxycytidine and/or the histone deacetylase inhibitor trichostatin A did not restore CDX2 expression in CDX2-negative lines. However, consistent with a role for dominant repression mechanisms in CDX2 silencing, all somatic cell hybrids resulting from pairwise fusions between colon cancer lines with intact CDX2 expression and lines lacking CDX2 had reduced CDX2 transcripts and protein. A roughly 9.5-kb 5'-flanking region from the human CDX2 gene contained key cis elements for regulating transcription in colon cancer cells. Restoration of CDX2 expression suppressed proliferation and soft agar growth in the CDX2-negative HT-29 colon cancer cell line. Our findings suggest CDX2 inactivation in colon cancer results from defects in trans-acting pathways regulating CDX2 transcription, and CDX2 silencing contributes to the altered phenotype of some colorectal cancers.
Insights
CDX2, a key transcription factor, is often lost in colorectal cancers due to silencing, not gene defects. Restoring CDX2 suppresses tumor growth, indicating its importance in colon cancer development.
Area of Science:
- Molecular Biology
- Oncology
- Genetics
Background:
- CDX2 is a transcription factor crucial for intestinal epithelium development.
- Loss of CDX2 expression is observed in some human colorectal cancers and mouse models.
- CDX2 defects are implicated in the development of intestinal polyps and colon cancer.
Purpose of the Study:
- To investigate the role of CDX2 defects in the development of human colorectal carcinomas.
- To analyze CDX2 expression patterns in primary colorectal tumors and cell lines.
- To elucidate the mechanisms underlying CDX2 silencing in colon cancer.
Main Methods:
- Analysis of CDX2 expression (transcript and protein) in 45 primary colorectal carcinomas and 13 colorectal cancer cell lines.
- Investigation of CDX2 gene defects and localization.
- Nuclear run-off assays to assess transcription rates.
- Treatment with DNA demethylating and histone deacetylase inhibitor agents.
- Somatic cell hybrid fusions to study dominant repression.
- Analysis of the CDX2 5'-flanking region.
- Restoration of CDX2 expression in colon cancer cells.
Main Results:
- Four of 45 primary colorectal carcinomas lacked CDX2 expression; three showed aberrant localization.
- No significant CDX2 gene defects were found in tumors with altered expression.
- Marked reductions in CDX2 transcript and protein levels were observed in 5/13 cell lines, primarily due to reduced transcription.
- Epigenetic modifiers did not restore CDX2 expression.
- Somatic cell hybrids indicated dominant repression mechanisms.
- Restoring CDX2 expression suppressed proliferation and growth in HT-29 cells.
Conclusions:
- CDX2 inactivation in colon cancer arises from defects in trans-acting pathways regulating transcription.
- CDX2 silencing contributes to the altered cellular phenotype observed in some colorectal cancers.
- CDX2 plays a significant role in suppressing colon cancer cell proliferation and growth.