Event In Colon Carcinogenesis Research Articles

Loss of symbiotic and increase of virulent bacteria through microbial networks in Lynch syndrome colon carcinogenesis.

Through a pilot study, we performed whole gut metagenomic analysis in 17 Lynch syndrome (LS) families, including colorectal cancer (CRC) patients and their healthy first-degree relatives. In a second asymptomatic LS cohort (n=150) undergoing colonoscopy-screening program, individuals with early precancerous lesions were compared to those with a normal colonoscopy. Since bacteria are organized into different networks within the microbiota, we compared related network structures in patients and controls. Fecal prokaryote DNA was extracted prior to colonoscopy for whole metagenome (n=34, pilot study) or 16s rRNA sequencing (validation study). We characterized bacteria taxonomy using Diamond/MEGAN6 and DADA2 pipelines and performed differential abundances using Shaman website. We constructed networks using SparCC inference tools and validated the construction's accuracy by performing qPCR on selected bacteria. Significant differences in bacterial communities in LS-CRC patients were identified, with an enrichment of virulent bacteria and a depletion of symbionts compared to their first-degree relatives. Bacteria taxa in LS asymptomatic individuals with colonic precancerous lesions (n=79) were significantly different compared to healthy individuals (n=71). The main bacterial network structures, constructed based on bacteria-bacteria correlations in CRC (pilot study) and in asymptomatic precancerous patients (validation-study), showed a different pattern than in controls. It was characterized by virulent/symbiotic co-exclusion in both studies and illustrated (validation study) by a higher Escherichia/Bifidobacterium ratio, as assessed by qPCR. Enhanced fecal virulent/symbiotic bacteria ratios influence bacterial network structures. As an early event in colon carcinogenesis, these ratios can be used to identify asymptomatic LS individual with a higher risk of CRC.

Xanthine oxidoreductase promotes the progression of colitis-associated colorectal cancer by causing DNA damage and mediating macrophage M1 polarization

In addition to its pivotal role in purine metabolism, xanthine oxidoreductase (XOR) is one of the key enzymes involved in superoxide radical generation. Oxidative stress has been implicated in the etiology of colorectal cancer, but the contribution of XOR remains unclear. Here we investigated the role of XOR in colitis-associated colorectal cancer (CAC) and the underlying mechanisms. Using clinical samples, we demonstrated that XOR up-regulation was an early event in colonic carcinogenesis. Pharmacological inhibition of XOR effectively delayed the progression of CAC. Moreover, XOR activity positively correlated with tumor necrosis factor-alpha (TNFα) protein levels. Mechanistically, TNFα may activate XOR transcription via activator protein-1 and, thus, promote endogenous hydrogen peroxide generation, resulting in oxidative DNA damage in colon cancer cells. On the other hand, XOR may regulate the TNFα mRNA transcripts by mediating LPS-induced macrophage M1 polarization. Collectively, XOR promotes tumor development by programming the tumor microenvironment and stimulates CAC progression via DNA damage-induced genetic instability.

A Cell Model Suitable for a High-Throughput Screening of Inhibitors of the Wnt/β-Catenin Pathway.

A constitutive activation of the Wnt/β-catenin pathway is an initiating event in colon carcinogenesis. We developed colon cancer cells models that highlight the non-selectivity of previously described inhibitors of the Wnt pathway and we propose our model as a suitable screening system for inhibitors of the pathway.

Single Nucleotide Polymorphism Facilitated Down-Regulation of the Cohesin Stromal Antigen-1: Implications for Colorectal Cancer Racial Disparities

The biological underpinnings for racial disparities in colorectal cancer (CRC) incidence remain to be elucidated. We have previously reported that the cohesin SA-1 down-regulation is an early event in colon carcinogenesis which is dramatically accentuated in African-Americans. In order to investigate the mechanism, we evaluated single nucleotide polymorphisms (SNPs) for association with SA-1-related outcomes followed by gene editing of candidate SNP. We observed that rs34149860 SNP was significantly associated with a lower colonic mucosal SA-1 expression and evaluation of public databases showed striking racial discordance. Given that the predicted SNP would alter miR-29b binding site, we used CRISPR knock-in in CRC cells and demonstrated that the SNP but not wild-type had profound alterations in SA-1 expression with miR-29b inhibitor. This is the first demonstration of high-order chromatin regulators as a modulator of racial differences, risk alteration with SNPs and finally specific modulation by microRNAs.

Are Corrupted Non-dysplastic Colonic Crypts the First Histological Event in Experimental Colonic Carcinogenesis?

The colonic crypts in rats replicate by symmetric fission at the base of the crypts, and proceeds upwards, to generate two separate identical crypts. Recently, anomalous crypts (called corrupted colonic crypts, CCCs) were found in the colonic mucosa of Sprague-Dawley rats treated with the carcinogen dimethylhydrazine. Here it was investigated whether CCCs develop in the colonic mucosa of another rat strain, treated with a different carcinogen. Archived Swiss-roll colon sections from 25 male Fisher-344 rats treated with the mutagen 2-amino-6-methyldipyrido imidazole (GLU1) were reviewed. Non-dysplastic and dysplastic CCCs were regarded as those exhibiting asymmetric fission, asymmetric lateral sprouting/lateral fission, basal dilatations, or spatial aberrations of the normal (vertical) axis. Colonic adenomas were found in three out of the 25 specimens. In the entire colonic mucosa of the 25 GLU1-treated rats, 130 non-dysplastic CCCs were recorded amongst 357 non-dysplastic crypts with fission (36.4%). The mean number of non-dysplastic CCCs per animal was 5.2 (range=2-12). These numbers only mirror events taking place at a particular time (i.e. at sacrifice). Considering the high cell production rate of the colonic crypts, the actual number of CCCs/rat occurring during the usual mucosal turnover time of 72 hours might be substantial. In the three adenoma specimens, non-dysplastic CCCs were found underneath CCCs with dysplasia. For many years, the development of crypt dysplasia and adenoma have been considered the initial histological events in colonic carcinogenesis. This study demonstrates that non-dysplastic CCCs also develop in GLU 1-treated Fisher-344 rats. Non-dysplastic CCCs were found underneath CCCs with dysplasia. Non-dysplastic CCCs might act as scaffolds at the time of top-down cell replacement/transformation of the crypts by dysplastic cells. It is submitted that non-dysplastic CCCs might be the initial histological recordable event in experimental colonic carcinogenesis.

Su2014 Cohesin SA1 Regulates Early Metabolic Events in Colon Carcinogenesis: Implications for Racial Disparities

Su2014 Cohesin SA1 Regulates Early Metabolic Events in Colon Carcinogenesis: Implications for Racial Disparities

The Wnt signaling inhibitor ICAT is overexpressed in primary human colorectal carcinoma (CRC)

Hintergrund/Einleitung: Aberrant activation of Wnt/β-catenin signaling through mutations in APC or β-catenin is a key event in colon carcinogenesis. ICAT was recently identified as a β-catenin binding protein disrupting the interaction of β-catenin and the transcription factor TCF thereby negatively regulating Wnt signaling. The expression and localization of the endogenous ICAT protein in human colon cancer or normal mucosa has not yet been described.

Sa1911 Mitochondrial Alterations in Early Colon Carcinogenesis: Evidence for Warburg Effect in the Initiation of Colorectal Cancer (CRC) Carcinogenesis

Background: There is emerging realization of the central role of metabolism in carcinogenesis with the Warburg effect (preferential utilization of glycolysis in normoxic environment) (e.g. Weinberg, Cell 2012). We have previously noted an increase in microvascular blood supply in the predysplastic colonic mucosa (field carcinogenesis) (Gut 2005, Gastro 2008, Clin Cancer Res 2009, Cancer Prev Res 2010) which may reflect altered metabolism, however molecular determinants remain unexplored. The role of mitochondria in CRC is well established in frank tumors with dysfunction reflected by increased fission leading to less efficient oxidative phosphorylation, enabling shunting of precursors for cell growth. However, the presence of mitochondrial dysfunction and Warburg effect in early colon carcinogenesis has yet been explored. Therefore, we chose to investigate potential mitochondrial changes in the pre-malignant colon. Methods: To investigate mitochondrial changes we incorporated both human and animal studies: For human studies we obtained biopsies from endoscopically normal rectal muscosa from 80 patients undergoing screening colonoscopies. Samples were processed for gDNA and RNA isolation. Real Time PCRwas conducted to assess mitochondrial mass, OPA-1, UCP2, and PKM2 expression. For animal experiments we utilized the Polypopsis in Rat Colon (Pirc) rat, which possesses an APC truncation, consistent with the initiating factor in ~80% of sporadic CRCs. 7 Pirc rats with age-matched controls were euthanized after 24 wks of age. Colons were excised from these animals to obtain colonic gDNA and RNA. Real Time PCR was conducted to assess mitochondrial mass, OPA-1 and UCP2 expression. Results: In patient biopsies, mitochondrial mass was shown to be upregulated with adenomal presence (~72% increase, p=0.05). Pirc rats were also shown to possess a 63.6% induction of mitochondrial mass (p=0.03). OPA-1 expression was upregulated in both patients with adenomas as well as Pirc rats (49%, p=0.05 and 67%, p<0.03 respectively). Induction of UCP2 was detected in patients with adenomas (99%, p<0.04) and in Pircs (101%, p=0.05). Interestingly, patients with lesions had a 80% increase in PKM2 (p=0.002), suggestive of a Warburg effect in early colon carcinogenesis. Conclusions: We demonstrate for the first time, that in the predysplastic mucosa there is profound metabolic alterations consistent with a Warburg effect. Specifically, we noted increased mitochondria mass (fission) which suppresses oxidative phosphorylation. Aside from structural alterations, electron transport system is further inhibited by overexpression of UCP2. Biologically, this provides insight into early metabolic events in colon carcinogenesis. Clinically, these insights may enable novel biomarkers for early detection and also a myriad of potentially druggable targets for chemoprevention.

Pathological implications of Cx43 down-regulation in human colon cancer.

Connexin 43 is an important gap junction protein in vertebrates and is known for its tumor suppressive properties. Cx43 is abundantly expressed in the human intestinal epithelial cells and muscularis mucosae. To explore the role of Cx43 in the genesis of human colon cancer, we performed the expression analysis of Cx43 in 80 cases of histopathologically confirmed and clinically diagnosed human colon cancer samples and adjacent control tissue and assessed correlations with clinicopathological variables. Western blotting using anti-Cx43 antibody indicated that the expression of Cx43 was significantly down regulated (75%) in the cancer samples as compared to the adjacent control samples. Moreover, immunohistochemical analysis of the tissue samples confirmed the down regulation of the Cx43 in the intestinal epithelial cells. Cx43 down regulation showed significant association (p<0.05) with the histological type and tumor invasion properties of the cancer. Our data demonstrated that loss of Cx43 may be an important event in colon carcinogenesis and tumor progression, providing significant insights about the tumor suppressive properties of the Cx43 and its potential as a diagnostic marker for colon cancer.

End-binding protein 1 (EB1) up-regulation is an early event in colorectal carcinogenesis

End-binding protein (EB1) is a microtubule protein that binds to the tumor suppressor adenomatous polyposis coli (APC). While EB1 is implicated as a potential oncogene, its role in cancer progression is unknown. Therefore, we analyzed EB1/APC expression at the earliest stages of colorectal carcinogenesis and in the uninvolved mucosa (“field effect”) of human and animal tissue. We also performed siRNA-knockdown in colon cancer cell lines. EB1 is up-regulated in early and field carcinogenesis in the colon, and the cellular/nano-architectural effect of EB1 knockdown depended on the genetic context. Thus, dysregulation of EB1 is an important early event in colon carcinogenesis.

Novel multiplex method to assess insulin, leptin and adiponectin regulation of inflammatory cytokines associated with colon cancer

The role of altered levels of insulin, leptin and adiponectin in contributing to the observed increased risk of colon cancer associated with obesity remains to be determined. Elevated insulin and leptin associated with obesity are linked to inflammatory responses. Conversely, adiponectin levels are reduced in obese individuals and this hormone is generally associated with anti-inflammatory responses. Inflammatory cytokines are key components of processes linked with carcinogenesis. Insulin, leptin and adiponectin receptor expression profiles were assessed in human normal, adenomatous polyp and tumour tissue. Insulin, leptin and adiponectin regulation of inflammatory cytokines previously identified as being associated with early events in colon carcinogenesis were further investigated here using a surrogate colon epithelial cell line and a custom designed GeXP assay of the inflammatory cytokines (CCL20, CXCL1, CXCL2, CXCL3, CXCL11, IL1RN, CXCL4, IL8, CCL19, CCL21, CCL23, CCL5, IL10RB and TNFRSF1A). Mean insulin, leptin and adiponectin receptor expression levels were lower in adenomatous polyp samples in comparison with normal and tumour tissue. In contrast to leptin, insulin significantly reduced CCL20 and CXCL11 and increased CXCL3 expression. Full length adiponectin, but not globular adiponectin, induced CCL5, CXCL1, CXCL3 and CCL20 gene expression. GeXP assay permitted measurement of changes in gene expression of cytokines in response to insulin and adiponectin, indicating the potential for insulin and adiponectin regulation of mediators of inflammation associated with early events in colon carcinogenesis.

Paranuclear Dot-Like Immunostaining for CD99

Makhoul, Rami MD*; Schwartz, Arnold M. MD, PhD†; Williams, Russell PhD‡; Lee, Norman H. PhD‡ Author Information

Sustained proliferation and resistance to apoptosis after a cytotoxic insult are early alterations in rat colon carcinogenesis

To study the early alterations in carcinogenesis, we determined apoptosis and proliferation in rat mucin depleted foci (MDF), precancerous lesions in the colon under basal conditions and 24 h after treatment with 1,2-dimethylhydrazine (DMH), which induces apoptosis in the colon. Spontaneous apoptosis in MDF was higher than in normal mucosa (Apoptotic Index was 1.61 ± 0.30 and 0.21 ± 0.02 in MDF and normal mucosa, respectively, mean ± SE, p < 0.05). DMH (30 and 75 mg/kg) increased apoptosis in both normal mucosa and MDF (up to 20 times higher compared to basal levels in normal mucosa, but only two times in MDF). MDF had a higher and deregulated pattern of proliferation along the crypt compared to normal mucosa. After DMH, proliferation in normal mucosa was significantly depressed, but it did not vary in MDF. Survivin-Birc5 regulating apoptosis and proliferation was significantly over-expressed (RT-qPCR and immunohistochemistry experiments) in MDF vs. normal mucosa, but did not vary in response to DMH. The expression of the pro-apoptotic protein Bak did not vary in normal mucosa and MDF. Since inflammation is present in MDF, which may hamper apoptosis, we studied the effect of pre-treatment with aspirin (600 ppm in the diet for 10 days). No significant effects of aspirin were observed. In conclusion, MDF had a higher spontaneous apoptosis and proliferation coupled with a reduced response to apoptotic stimuli from cytotoxic compounds. Survivin over-expression in MDF indicates that this is an early event in colon carcinogenesis and suggests that down-regulation of Survivin may represent a strategy for cancer prevention.

Loss of Heterozygosity at the Glutathione Peroxidase 1 Locus Is Not an Early Event in Colon Carcinogenesis

It has been previously shown that loss of heterozygosity (LOH) at the cytosolic glutathione peroxidase (GPx-1) locus is a common event in the development of several cancer types, including colorectal cancer. GPx-1 is an antioxidant selenium-containing protein, and polymorphisms within this gene have been shown to be associated with the increased risk of cancer. In order to assess whether this genetic change was an early or late event in colon cancer development, we investigated whether LOH at this site was occurring in colorectal adenomas, a premalignant lesion. Twenty-four pairs of DNA samples, obtained from both whole-blood and adenoma tissue from the same individuals, were genotyped at 2 positions in the GPx-1 gene: a codon 198 variation resulting in either a leucine or proline at the corresponding position in the peptide, or a variable number of alanine repeat codons corresponding to the amino terminus of the GPx-1 protein. No evidence of GPx-1 LOH was observed in the examined sample sets. These data indicate that the genetic loss at the GPx-1 locus may be a late event in colon carcinogenesis.

Epigenetic Instability via Genomic-Wide DNA Methylation and Histone Modification Profile Predicts Colorectal Adenoma and Cancer

Background: Epigenetic silencing including chromatin modification likely precedes genetic alterations in early colorectal cancer (CRC) events. We aimed to investigate global histone acetylation, HDAC expression and genome wide DNA methylation (GWM) of all CpG island in normal, adenoma and CRC from African Americans (AA). Materials and methods: Tissue microarrays (TMA) with colon adenoma (134), cancer (58) and matched normal tissue from AA patients were stained for H3K18, H4K12 and HDAC2 by immunohistochemistry. Trends in intensity of staining were tested by analysis of variance. The correlation between expression of these proteins and various clinicopathological features were analyzed. In addition, DNA from 22 (two normal, eight adenoma, 12 cancer) fresh frozen samples was analyzed for GWM on an Illumina Infinium methylation array (HumanMethylation27). Data was normalized using the methylumi package for the R statistical analysis software. For each type of sample (normal, adenoma or cancer), the top 100-200 hyperand hypo-methylated genes were selected by comparing the confidence intervals for the methylation status to 0 (unmethylated) or 1 (methylated). We also compared the cancer and adenoma samples. From this comparison, 24 genes were found (by inspection) to have high methylation in CRC and adenoma compared to normal or vice versa which we did an Ingenuity Pathway Analysis (IPA). Methylation specific PCR (MSP) was used to validate few candidate genes. Comparative analysis was also done between our data and data from The Cancer Genome Atlas (TCGA) study. Results: HDAC2 nuclear expressionwas 81.9%, 62.1%, and 53.1% in cancer, adenoma, and normal tissue, respectively (P<0.002). H4K12 acetylation was 71.7%, 61%, and 43.6%, respectively (P<0.002). H4K12 acetylation and HDAC2 expression correlated with progression of adenoma to carcinoma (P<0.002). The GWM and IPA, revealed that Wnt/b-catenin signaling was one of the top canonical pathways implicated in the 24 genes differentially methylated between cancer and adenoma compared to normal. Hypermethylated genes were 307 in AA adenoma (including CBFA, ATP1A, ANKDD1, FLI, GDPD3. DRD5, TCF4 and SerpinB3) vs. normal, while there was 185 genes in Whites using TCGA. A near perfect [adenoma + normal] cluster and cancer cluster was observed indicating the significance and importance of DNA methylation in CRC progression. Methylation for TCF was 92%, in CRC and 70% of adenoma cases. Conclusion: These results suggest that global H4 acetylation and HDAC2 expression might be associated with colorectal tumor progression. Our data indicated a high methylation profile of SerpinB3 and TCF4 as potential markers in adenoma as part of the early events of colon carcinogenesis. DNA methylation profile and histone modification can predict colorectal adenoma and cancer.

Progesterone Receptor Upregulation is an Early Event in Colon Carcinogenesis: Potential Modulator of Neoplastic Risk in Women

Purpose: While colorectal cancer (CRC) impacts upon the genders equally, there are profound differences in biology and epidemiology (e.g., women tend to have proximal lesions, MSI high tumors etc). However, clinical practice largely treats CRC as gender-neutral. Our group has shown that CRC biomarkers are different in women thus having significant implications for screening (JAMA 2009, Can Prev Res 2010). The gender differences are generally attributed to sex steroids although precise hormones remain unclear. For instance, WHI data indicates estrogen/progesterone reduced CRC by 38% (Chlebowski et al., NEJM 2004) although estrogen alone had no effect (OR:1.08, Anderson et al., JAMA 2004) suggesting that the progesterone receptor (PR) was important. However, the role of PR in CRC remains unexplored. Methods: For these studies we employed the azoxymethane (AOM)-treated rat which is the best-accepted model of experimental colon carcinogenesis requiring 20 and 40 weeks for adenomas and CRCs to develop, respectively. PR expression from colonic mucosa was assessed by real time RT-PCR using high capacity cDNA synthesis kit and PR-specific taqman probe. In order to further assess PR's role, we employed a well-established chemopreventive agent, celecoxib in the human CRC cell line HCT 116. (Bertagnolli et al., NEJM 2006). We evaluated protein levels using quantitative immunoblot analysis. Results: In the AOM-treated rat, PR was markedly (319.6±12.2%) increased in the tumor when compared to colonic mucosal scrapings from age-matched saline-treated controls. Moreover, a comparable (392±4.2%, p<0.001) increase was noted in the microscopically-normal mucosa of these AOM-treated rats implicating field carcinogenesis. Typically, we examined the 10 week (premalignant) time-point since it best replicates human field carcinogenesis and noted a comparable induction of PR in the premalignant mucosa (313±4.2%, p<0.001). From a diagnostic perspective, the AUROC at the 10 week timepoint for discriminating AOM versus control rats was an impressive 0.97. Finally, we noticed that PR was markedly altered with celecoxib treatment (50μm) in HCT116 by 37.4% (p<0.01). Conclusion: We report, herein, for the first time that PR induction is an early event in colorectal carcinogenesis. Moreover, its reversal by chemoprevention underscores PR's biological significance. Importantly, the performance of PR expression was outstandingly supporting a possible role in CRC screening, potentially coupled with our recently developed ability to perform real time PCR on mucus layer fecal colonocytes (to detect field carcinogenesis). Furthermore, PR status may finally explain some of the gender-related differences in colon carcinogenesis.

In colon carcinogenesis, the cytoskeletal protein gelsolin is down-regulated during the transition from adenoma to carcinoma

The actin-binding protein gelsolin is involved in cell motility via the regulation of actin cytoskeleton, and its expression is modified in several human cancers. However, the potential implication of this protein in colorectal carcinogenesis is debated. By using immunohistochemistry, we studied gelsolin expression in 69 cases of colon adenocarcinomas and in 72 lesions representative of the different stages of colonic tumorigenesis. In addition, we performed Northern blot analysis of gelsolin messenger RNA in 12 paired samples of human colon cancer and normal corresponding mucosa. Gelsolin protein and messenger RNA expressions were severely down-regulated in all adenocarcinomas tested. Moreover, gelsolin protein was down-regulated in a large proportion of high-grade adenomas (14/16) before the acquisition of invasive properties but in only a small proportion of low grade adenomas and serrated adenomas (2/30) and in none of the 9 cases of nonneoplastic hyperplastic polyps tested. Our results therefore demonstrate that gelsolin down-regulation is an early and almost constant event in colon carcinogenesis and is associated with the transition from adenoma to carcinoma.

Identification of candidate cooperative genes of the Apc mutation in transformation of the colon epithelial cell by retroviral insertional mutagenesis

The mutation of Apc is an important early genetic event in colon carcinogenesis. However, it remains to be clarified what kinds of cooperative genes are required for complete carcinogenesis. To identify cooperative genes for the Apc(Min) mutation the authors carried out retroviral insertional mutagenesis (RIM) using Min mouse-derived IMCE colon epithelial cells. Anchorage-independent transformed colonies were induced by retroviral infection only in IMCE cells, while no transformation was found in young adult mouse colon (YAMC) cells that are normal for Apc. One hundred and fifty-seven retroviral integration sites (RIS) were identified in 101 independent transformants, and four common integration sites (CIS), Dnah3, Ahnak, Stk17b and Rbm9, were observed. Upregulation of Dnah3 and Ahnak, and truncation of Dnah3 due to the viral integration, was revealed. In addition, Dnah3-overexpressing IMCE cells showed impairment of microtubule function. These data suggest the importance of cytoskeletal function in Apc-related tumor development and the usefulness of RIM in non-hematopoietic tissues, providing new insight into the early stage of colon carcinogenesis.

Oncogenic K‐ras promotes early carcinogenesis in the mouse proximal colon

Oncogenic K-ras mutations are frequently observed in colon cancers and contribute to transformed growth. Oncogenic K-ras is detected in aberrant crypt foci (ACF), precancerous colonic lesions, demonstrating that acquisition of a K-ras mutation is an early event in colon carcinogenesis. Here, we investigate the role of oncogenic K-ras in neoplastic initiation and progression. Transgenic mice in which an oncogenic K-ras(G12D) allele is activated in the colonic epithelium by sporadic recombination (K-rasLA2 mice) develop spontaneous ACF that are morphologically indistinguishable from those induced by the colon carcinogen azoxymethane (AOM). Similar neoplastic changes involving the entire colon are induced in transgenic mice constitutively expressing K-ras(G12D) throughout the colon (LSL-K-ras(G12D)/Villin-Cre mice). However, the biochemistry and fate of K-ras-induced lesions differ depending upon their location within the colon in these mice. In the proximal colon, K-ras(G12D) induces increased expression of procarcinogenic protein kinase C beta II (PKC beta II), activation of the MEK/ERK signaling axis and increased epithelial cell proliferation. In contrast, in the distal colon, K-ras(G12D) inhibits expression of procarcinogenic PKC beta II and induces apoptosis. Treatment of K-rasLA2 mice with AOM leads to neoplastic progression of small ACF to large, dysplastic microadenomas in the proximal, but not the distal colon. Thus, oncogenic K-ras functions differently in the proximal and distal colon of mice, inducing ACF capable of neoplastic progression in the proximal colon, and ACF with little or no potential for progression in the distal colon. Our data indicate that acquisition of a K-ras mutation is an initiating neoplastic event in proximal colon cancer development in mice.

The Cables Gene on Chromosome 18q Is Silenced by Promoter Hypermethylation and Allelic Loss in Human Colorectal Cancer

Cables is a cyclin-dependent kinase-binding nuclear protein that maps to chromosome 18q11-12. Here, we assessed Cables expression in 160 colorectal cancers (CRCs), its role in colon cancer cell growth, and the potential mechanisms of Cables inactivation. Expression levels, promoter methylation, and mutational status of Cables were investigated in colon cancer cell lines and primary colon tumors. Chromosome 18q loss of heterozygosity (LOH) was evaluated with multiple polymorphic markers. Cables inhibited cellular proliferation and colony formation in colon cancer cell lines. Cables expression was reduced in 65% of primary CRCs. No mutations were detected in 10 exons of Cables in 20 primary colon tumors. Cables promoter was methylated in cell lines with decreased Cables expression and vice versa. 5-Aza-2'-deoxycytidine resulted in increased Cables expression in methylated cell lines. There was a significant correlation between promoter methylation and Cables gene expression in primary colon tumors. Sixty-five percent of primary colon tumors demonstrated chromosome 18q LOH. LOH involving the Cables region was observed in 35% of cases, including those in which more distal portions of chromosome 18q were retained, and Cables expression was decreased in all such cases. Loss of Cables expression in 65% of CRCs suggests that it is a common event in colonic carcinogenesis, with promoter methylation and LOH appearing to be important mechanisms of Cables gene inactivation.