Colitis-associated Carcinomas Research Articles

Estrogen receptor beta expression in colitis-associated carcinoma in comparison with sporadic colonic tumor: An immunohistochemical study.

The rate of ulcerative colitis (UC)-related colorectal cancer (colitis-associated carcinoma) is increasing. Estrogen receptor (ER) beta expression has been studied separately in patients with sporadic colorectal cancer and those with colitis-associated carcinoma. However, no study has compared the expression in both of these cancer types. The present study aimed to evaluate the relationship between colitis-associated carcinoma and ERs and assess whether the expression of ER beta influences cell proliferation. This study included 45 surgically operated colitis-associated carcinomas, 43 high-grade dysplasias, 34 low-grade dysplasias, 36 sporadic colorectal cancers, 44 high-grade adenomas, and 34 low-grade adenomas. ER beta expression was evaluated with immunohistochemistry. Colitis-associated carcinoma showed significantly lower ER beta immunoexpression than sporadic colorectal lesions and high- and low-grade dysplasia. In seven colitis-associated carcinoma harboring both intensity score 3 (strong immunoexpression) and score 1 (weak immunoexpression) areas, the correlation among ER beta intensity, Ki-67, and p21 labeling index was assessed; an area with an ER beta intensity score of 3 showed a higher Ki-67 labeling index than that with score 1. In four out of the seven lesions, p21 labeling index was higher in the area of ER beta score 1 than in that of ER beta score 3. The data suggest that ER beta expression is an accelerating factor in colorectal tumors. This association may be lower in colitis-associated carcinoma than in sporadic colorectal cancer.

Immunoprofiles and DNA Methylation of Inflammatory Marker Genes in Ulcerative Colitis-Associated Colorectal Tumorigenesis.

Immunological and epigenetic changes are interconnected and contribute to tumorigenesis. We determined the immunoprofiles and promoter methylation of inflammation-related genes for colitis-associated colorectal carcinomas (CA-CRC). The results were compared with Lynch syndrome (LS)-associated colorectal tumors, which are characterized by an active immune environment through inherited mismatch repair defects. CA-CRCs (n = 31) were immunohistochemically evaluated for immune cell scores (ICSs) and PDCD1 and CD274 expression. Seven inflammation-associated genes (CD274, NTSR1, PPARG, PTGS2, PYCARD, SOCS1, and SOCS2), the repair gene MGMT, and eight standard marker genes for the CpG Island Methylator Phenotype (CIMP) were investigated for promoter methylation in CA-CRCs, LS tumors (n = 29), and paired normal mucosae by multiplex ligation-dependent probe amplification. All but one CA-CRCs were microsatellite-stable and all LS tumors were microsatellite-unstable. Most CA-CRCs had a high ICS (55%) and a positive CD274 expression in immune cells (52%). NTSR1 revealed frequent tumor-specific hypermethylation in CA-CRC and LS. When compared to LS mucosae, normal mucosae from patients with CA-CRC showed significantly higher methylation of NTSR1 and most CIMP markers. In conclusion, CA-CRCs share a frequent ICShigh/CD274pos expression pattern with LS tumors. Elevated methylation in normal mucosa may indicate field cancerization as a feature of CA-CRC-associated tumorigenesis.

Molecular characterization of ulcerative colitis-associated colorectal carcinomas.

Patients with ulcerative colitis (UC) are at increased risk for developing colorectal cancer (CRC). In contrast to sporadic colorectal tumorigenesis, TP53 mutations occur early in the progression from inflamed colonic epithelium to dysplasia to CRC, and are sometimes readily detectable in inflamed, (yet) non-dysplastic mucosa. Here, we analyzed formalin-fixed paraffin-embedded tissue samples from 19 patients with long-standing UC (median 18 years, range 3 to 34) who had developed CRC as a consequence of chronic inflammation of the large bowel. We performed microsatellite instability testing, copy number analysis by array-based comparative genomic hybridization, mutation analysis by targeted next generation sequencing (48-gene panel) and TP53 immunostaining. The results were compared to The Cancer Genome Atlas (TCGA) data on sporadic CRC. All UC-CRC lesions in our cohort were microsatellite stable. Overall, genomic imbalances of UC-CRCs showed patterns of chromosomal aneuploidies characteristic for sporadic CRC with the exception of gains of chromosome arm 5p (12 of 23 UC-CRC, 52%), which are rare in sporadic CRCs from TCGA (21 of 144, 15%; FDR adjusted P = 0.006). UC-CRCs showed a predilection for TP53 alterations, which was the most frequently mutated gene in our cohort (20 of 23, 87%). Interestingly, spatially separated tumor lesions from individual patients tended to harbor distinct TP53 mutations. Similar to CRCs arising in a background of Crohn’s colitis, the genetic landscape of UC-CRCs was characterized by TP53 mutations and chromosomal aneuploidies including gains of chromosome arm 5p. Both alterations harbor the potential for early detection in precursor lesions, thus complementing morphologic diagnosis.

DOP51 Investigation on the role of Par4-associated cell polarity and associated barrier defects in inflammatory bowel diseases

Abstract Background In inflammatory bowel diseases (IBD), epithelial barrier defects occur as a consequence of chronic inflammation. Recent research suggested that cell polarity alterations may be upstream of barrier defects and additionally play a role in IBD-associated carcinogenesis (colitis-associated and small intestinal carcinoma). Par4 is a gene encoding a protein crucial in the development of cell polarity. LKB1, its human homologue, is mutated in Peutz–Jeghers syndrome (PJS), a genetic condition characterised by a higher risk of epithelial cancers. While the pivotal role of Par4/LKB1 in the development of epithelial cell polarity is established, its involvement in IBD-associated barrier defects and carcinogenesis is yet to be defined. Methods Endoscopic bowel mucosa samples from patients with Crohn’s disease, ulcerative colitis, PJS and controls were analysed to assess expression and localisation of Par4/LKB1. Cryosections were immunostained for Par4/LKB-1 along with a diverse set of markers of cell polarity and intercellular adhesion. The analysis was performed by confocal laser scanning microscopy in order to compare the expression of mucosal as well as the subcellular localisation of Par4/LKB1 in the gut mucosa. A quantitative analysis of protein expression with western blotting was performed as well. The function of Par4/LKB1 in intestinal epithelial cells (IEC) was evaluated by means of a Par4-deficient IEC model as well as in an IEC Par4-overexpression model. Moreover, a preliminary assessment of the possible role of Par4/LKB1-related polarity processes in IBD-associated carcinomas was performed through scanning genetic data from colitis-associated carcinomas for potential Par4-related mutations and altered Par4/LKB1 expression. Results The immunofluorescent staining allowed visualisation of intracellular expression of Par4 in epithelia from PJS, IBD patients and controls. In PJS-polyps, despite the alteration of regular tissue architecture typical of these lesions, the polarity of epithelial cells was maintained - contrary to control tissue, a punctate pattern of the Par4 staining was shown. In IBD tissue, no relevant differences in Par4 expression at confocal microscopy, as well as a quantitative assessment, were observed as compared with controls. No differences were observed in PJS or IBD samples as regards the expression of cell polarity markers such as Par3, CD71, crb3 or adhesion molecules such as ZO-1, e-cadherin, occludin, JAM-A. As regards the IEC model, we observed that Caco-2/BBe cells overexpressing Par4/LKB1 showed enhanced Par4/LKB1 cytosolic staining at immunofluorescence. In this model, a defective epithelial polarity and organisation on permeable supports could be observed in Caco2/BBe cysts in parallel with reduced expression of native LKB1 as seen in western blots. Preliminary analysis of data from our colitis-associated carcinomas database suggested various pathways potentially involving Par4/LKB1, yet no direct analysis of tissue samples for Par4-related mutations was performed yet. Conclusions Altered expression of Par4 in epithelial cells may influence the development of a functional epithelial barrier as suggested by IEC models. Par4/LKB1 as a known oncosuppressor gene is involved in a number of pathways potentially relevant for carcinogenesis in IBD; however, a direct connection between Par4-related alteration of the epithelial barrier and carcinogenesis is yet to be established.

Colitis-associated colorectal adenocarcinomas are frequently associated with non-intestinal mucin profiles and loss of SATB2 expression

The special AT-rich sequence binding protein 2 (SATB2) is a sensitive and specific diagnostic marker for colorectal adenocarcinoma and reduced expression of SATB2 is associated with a poor prognosis. Colitis-associated colorectal adenocarcinoma often shows distinct morphologic and molecular phenotypes compared to sporadic cases. However, the SATB2 expression profile in colitis-associated carcinoma has not been defined. We performed immunohistochemistry for SATB2 as well as CDX2, MUC5AC, MUC6 and mismatch repair proteins on 60 consecutive colitis-associated carcinomas from 58 inflammatory bowel disease patients and compared the expression profile to a control group of 32 sporadic colorectal carcinomas. Only 26 (43%) colitis-associated carcinomas expressed SATB2, compared to 29 (91%) sporadic colorectal carcinomas (p < 0.0001). MUC5AC expression was more frequently observed in colitis-associated carcinomas than sporadic colorectal caracinomas (52% and 25% respectively; p = 0.013). Eight (13%) cases of colitis-associated carcinoma showed loss of CDX2 expression, which was retained in all of the sporadic controls (p = 0.047). In colitis-associated carcinoma, 50% of SATB2 negative cases had lymph node metastasis compared to only 15% of SATB2 positive cases (p = 0.007). Loss of SATB2 was particularly frequent in mucinous-type tumors, occurring in 83% of these cases. There was no significant association between SATB2 expression and mismatch repair protein status. These data show that the immunoprofile of colitis-associated carcinoma is different than that seen in sporadic cases. In particular, SATB2 is significantly less sensitive in colitis-associated carcinoma and it should be interpreted cautiously as a marker of colorectal origin in colitis patients. The association between loss of SATB2 and lymph node metastasis suggests that it may have similar prognostic value in the setting of inflammatory bowel disease as in sporadic cases.

Heat shock protein 47 regulated by miR-29a to enhance glioma tumor growth and invasion

Heat shock protein 47 (HSP47), also known as SERPINH1, is a product of CBP2 gene located at chromosome 11q13.5, a region frequently amplified in human cancers. HSP47 has been demonstrated to effect on limiting tumor invasion and motility. The previous studies showed that HSP47 is overexpressed in many human cancers, including stomach cancer, lung cancer, pancreatic ductal adenocarcinoma, and ulcerative colitis-associated carcinomas. However, the role of HSP47 in human glioma is still unknown. Here, we examined the expression of HSP47 in a group of glioma tumors and matched non-tumor brain tissues using qRT-PCR. We found that HSP47 is significantly overexpressed in glioma tissues and cell lines and associated with glioma tumor grade. Next, we knockdown the expression of HSP47 in the glioma cells using small interfering RNAs. The result showed that knockdown of HSP47 inhibits glioma cell growth, migration and invasion in vitro. We further investigated the posttranscriptional regulation of HSP47 by microRNAs using bioinformatics analysis and experimental validation. The results suggested that the expression of HSP47 is regulated by miR-29a. Finally, stable knockdown of HSP47 using shRNA inhibits glioma tumor growth and induces apoptosis in mice models in vivo. Therefore, our data suggested that HSP47 regulated by miR-29a to enhance glioma tumor growth and invasion. Taken together, HSP47 plays important role in tumor growth and invasion and thus could be a therapeutic target for treating glioma in the future.

Abstract 856: Association of high levels of folate with increased risk for colitis-associated colorectal neoplasia

Abstract Low folate levels are known to be a risk factor for multiple human diseases including colorectal cancer. However, the relationship between folate levels and the prevention or development of colorectal cancer remains controversial. Patients with ulcerative colitis often possess low levels of folate due to inadequate intake and malabsorption of nutrients. Thus, folic acid supplementation is highly recommended despite very limited information regarding the effect of folic acid on the progression of colitis-associated colorectal cancer. The major objectives of the present study are to evaluate folate levels during the progression of colitis-associated colon cancer and to assess the potential functional role of folic acid supplementation on the growth of colon cancer cells. To study the effect of inflammation on levels of folate, acute and chronic colitis were induced within the mouse colon by administering 2 cycles of 3% dextran sulfate sodium (DSS) following one injection of azoxymethane (AOM; 7.4 mg/kg); a regimen that ultimately leads to the formation of colitis-associated dysplasias and carcinomas. At 10+, 12 and 19 wks of age, corresponding to acute colitis, chronic colitis, and colitis-associated dysplasia, respectively, blood samples were collected retro-orbitally and plasma folate levels were measured using stable isotope dilution liquid chromatography-multiple reaction monitoring/mass spectrometry. At sacrifice, the histopathology of the colon was correlated with plasma folate levels. Levels of plasma folate were reduced 33% (78.4 + 7.2 nM vs. 116.2 + 15.6 nM, p&amp;lt;0.05) and 21% (110. 4 + 5.2 nM vs. 138.2 + 10.8 nM, p&amp;lt;0.05) in AOM/DSS-treated mice at 10+ or 12 wks of age, respectively, as compared to age-matched mice treated with AOM alone. Surprisingly, plasma folate levels were elevated 12% in AOM/DSS-treated mice exhibiting multiple colitis-associated dysplasias (19 wks of age), as compared to control mice without tumors treated with AOM only. Administration of 5-aminosalicylic acid (5-ASA) to colitic mice at 75 mg/kg, a dose that inhibits the multiplicity of dysplasias by 50%, decreased plasma folate to a level lower than that of AOM/DSS controls (77.9 +10.7 nM vs. 199.0 + 25.1 nM, p&amp;lt;0.001). The effect of folic acid supplementation on cellular signaling pathways was evaluated in SW480 colon carcinoma cells cultured in the presence of low- or high-dose folic acid. Western blot analyses using antibodies against phospho-ERK1/2 and phospho-GSK3β revealed that exposure to high-dose folic acid increased phosphorylated ERK1/2 and reduced phosphorylated GSK3β by 2.0- and 2.5-fold, respectively, as compared to cells exposed to low-dose folic acid. These in vitro and in vivo data, when combined, suggest that high levels of folic acid may increase risk for colitis-associated neoplasia via the ERK/MAPK and GSK3β pathways. Supported by NIH R01 CA124693. Citation Format: {Authors}. {Abstract title} [abstract]. In: Proceedings of the 102nd Annual Meeting of the American Association for Cancer Research; 2011 Apr 2-6; Orlando, FL. Philadelphia (PA): AACR; Cancer Res 2011;71(8 Suppl):Abstract nr 856. doi:10.1158/1538-7445.AM2011-856

High Frequency of Aneuploidy Defines Ulcerative Colitis-Associated Carcinomas

Aneuploidy is an independent risk factor for forthcoming carcinogenesis in ulcerative colitis (UC). An inferior prognosis of patients with ulcerative colitis-associated colorectal cancer (UCC) compared with those with sporadic colorectal cancer (SCC) has been reported, but remains controversial. This prompted us to investigate if aneuploidy can be observed in UCCs as frequently as in their sporadic counterpart and if aneuploidy per se might be a driving feature of poor prognosis in UCC. We obtained clinical follow-up for 257 SCC patients (average observation time 57 months) and 31 UCC patients (51 months). Touch preparation slides or tissue sections were prepared of all 288 carcinomas for ploidy analysis. Ploidy status was assessed for 260 SCCs and 31 UCCs by image cytometry and correlated to clinical features. Survival data were analyzed using Kaplan-Meier estimates. Aneuploidy was detected in 74.6% of SCCs and in all 31 UCCs. Logistic regression analysis yielded age (odds ratio [OR], 1.05; 95% CI, 1.02-1.09; P = 0.003) and aneuploidy (OR, 4.07; 95% CI, 1.46-11.36; P = 0.007) as independent prognostic factors for R0-resected patients devoid of metastases. Diploid SCCs had a more favorable 5-year survival (88.2%) than aneuploid SCCs (69.0%) and UCCs (73.1%) (P = 0.074). UC-associated carcinomas presented aneuploidy at significantly higher frequency than sporadic colorectal carcinomas (P < 0.0006). UCCs and aneuploid SCCs share a similar prognosis inferior to that of diploid SCCs. Aneuploidy proved to be the strongest independent prognostic marker for R0-resected colorectal cancer patients overall.

High expression of HSP47 in ulcerative colitis-associated carcinomas: proteomic approach

Background:Ulcerative colitis (UC) is a chronic relapsing inflammatory bowel disease, known to be associated with a markedly increased risk of colorectal carcinoma development.Methods:Using proteomic analysis with two-dimensional gel electrophoresis and mass spectrometry, differentially expressed proteins were assessed between UC-associated cancer and sporadic colon cancer cell lines. Western blot and immunostaining were performed for confirming the expression.Results:Heat-shock protein of 47 kDa (HSP47) was identified as one of the proteins expressed more highly in UC-associated cancer cell lines, and an immunohistochemical examination confirmed significantly higher levels of HSP47 in UC-associated colon cancers than in sporadic counterparts, the expression increasing with a progression of neoplastic lesions. Heat-shock protein of 47 kDa was further found to be coexpressed with type I collagen in the cytoplasm, and both HSP47 and type I collagen were released from cultured cells into the culture medium.Conclusion:These results suggest that overexpression of HSP47 is a unique characteristic of UC-associated carcinoma related to type I collagen synthesis, with possible clinical applications.

Synchronous multifocal colorectal carcinoma in a patient with delayed diagnosis of ulcerative pancolitis

Patients with ulcerative colitis face an increased lifetime risk of developing colorectal cancer. Relatively often, the patients present with multiple synchronous or metachronous tumors. Here, we report a case of ulcerative colitis-associated synchronous multifocal colorectal carcinomas. A 36-year-old male presented with symptoms of persisting abdominal pain that had lasted for several months. Histology of the colonic biopsies showed active ulcerative pancolitis with extensive multifocal low- and high-grade dysplasia. Regardless of the diagnosis and medical advice, the patient initially refused therapy, and proctocolectomy was delayed for 12 months. In the resection specimen, four clinically unsuspected, partly mucinous adenocarcinomas accompanied by several foci of low- and high-grade dysplasia were found in the left colon and rectum. At the time of colectomy, advanced tumor stage was diagnosed and classified as pT3c(4) pN1(2/120) M0 V1 R0, UICC stage IIIB, G2. Furthermore, a mucinous cystadenoma was found in the appendix in the setting of ulcerative colitis. We discuss the neoplastic transformation, current surveillance guidelines, and the therapeutic management in ulcerative colitis.

Chromoendoscopy and ulcerative colitis

Background and aimsTimely diagnosis of intraepithelial neoplasias (IN) and colitis-associated colon carcinomas (CRC) is crucially important in the treatment of ulcerative colitis (UC). We performed a randomized, controlled trial to test whether chromoendoscopy (CE) can facilitate early detection of IN and CRC in ulcerative colitisMethodsA total of 263 patients with long-standing UC (8 years or more) were screened for inclusion in the study, 165 of whom were randomized at a 1:1 ratio to undergo conventional colonoscopy or colonoscopy with CE using 0.1% methylene blue. Five mucosal biopsy specimens were taken every 10 cm between the rectum and caecum. Circumscript lesions in the colon were evaluated according to a modified pit pattern classification.ResultsIn the CE group, there was a significantly higher correlation between the endoscopic assessment of degree (P = 0.0002) and extent (89 vs 52%; P < 0.0001) of colonic inflammation and the histopathologic findings, compared with the conventional colonoscopy group. More targeted biopsies resulted in the detection of significantly more IN in the CE group (32 vs 10; P = 0.003). Using a modified pit pattern classification, both the sensitivity and specificity for differentiation between non-neoplastic and neoplastic lesions measured 93%.ConclusionsBased on our prospective, randomized trial, CE permits more accurate diagnosis of the extent and severity of inflammatory activity in UC compared with conventional colonoscopy. In addition, CE with methylene blue provides a novel tool for the early detection of IN and CRC in patients with UC. These findings have important implications for medical and surgical interventions.

Methylene blue-aided chromoendoscopy for the detection of intraepithelial neoplasia and colon cancer in ulcerative colitis

Background & Aims: Timely diagnosis of intraepithelial neoplasias (IN) and colitis-associated colon carcinomas (CRC) is crucially important for the treatment of ulcerative colitis (UC). We performed a randomized, controlled trial to test whether chromoendoscopy (CE) might facilitate early detection of IN and CRC in UC. Methods: A total of 263 patients with long-standing UC (≥8 years) were screened for potential inclusion in the study, 165 of whom were randomized at a 1:1 ratio to undergo conventional colonoscopy or colonoscopy with CE using 0.1% methylene blue. Five mucosal biopsy specimens were taken every 10 cm between the rectum and cecum. Circumscript lesions in the colon were evaluated according to a modified pit pattern classification. Results: In the CE group, there was a significantly better correlation between the endoscopic assessment of degree (P = 0.0002) and extent (89% vs. 52%; P < 0.0001) of colonic inflammation and the histopathologic findings compared with the conventional colonoscopy group. More targeted biopsies were possible, and significantly more IN were detected in the CE group (32 vs. 10; P = 0.003). Using the modified pit pattern classification, both the sensitivity and specificity for differentiation between non-neoplastic and neoplastic lesions were 93%. Conclusions: Based on our prospective randomized trial, CE permits more accurate diagnosis of the extent and severity of the inflammatory activity in UC compared with conventional colonoscopy. In addition, CE with methylene blue is a novel tool for the early detection of IN and CRC in patients with UC. These findings have important implications for medical and surgical interventions.

Ulcerative colitis and colorectal carcinoma: DNA-profile, laminin-5 gamma2 chain and cyclin A expression as early markers for risk assessment.

Ulcerative colitis patients are at increased risk for developing colorectal carcinomas. Despite expensive surveillance programmes, clinical practice reflects an uncertainty in individual risk assessment. The aim of the study was to evaluate independent cellular features with possible predictive value. Two patient groups were selected: group A comprised 8 patients with ulcerative colitis-associated colorectal carcinomas, group B comprised 16 ulcerative colitis patients with risk factors (duration of disease, extent of inflammation, epithelial dysplasias). A total of 683 paraffin-embedded mucosal biopsies were retrospectively evaluated for inflammatory activity, grade of dysplasia, ploidy status, laminin-5 gamma2 chain and cyclin A expression. Mild or moderate inflammatory activity was present in 78% of all biopsies, low- or high-grade dysplasia in 5.5%. There was no difference in inflammatory activity and dysplasia between patient groups. In group A, 75% of the biopsies exhibited aneuploid DNA distribution patterns. Group B showed mainly proliferative-diploid cell populations (85% / P = 0.006). Laminin-5 gamma2 chain was expressed in 13% of all biopsies, with a higher frequency in group A (P = 0.002). Cyclin A expression was found in 98% of all biopsies, with a higher number of immunopositive cells in group A biopsies (P = 0.014). Combined nuclear DNA assessment, laminin-5 gamma2 chain and cyclin A expression may help to identify ulcerative colitis patients with an increased risk for cancer development.