Final Progress Report

Proposal No. IBD-0099R
Principal Investigator:   Christoph Hanski, Ph.D.
Applicant Organization:  Charité Universitätsmedizin Berlin, Campus Benjamin Franklin (Germany)
Project Title:  Effects of deoxycholic acid and ursodeoxycholic acid on the development of carcinomas during chronic ulcerative colitis:  proof of principle in an animal model
Period of Award:  March 1, 2004 – June 30, 2006

Summary of the project aims

The following questions should be answered by the project

   1.    Does UDCA inhibit the development of colon carcinomas in the mouse model of ulcerative colitis?
Colon carcinogenesis should be monitored under conditions of feeding with cholic acid (CA) or ursodeoxycholic acid (which increases or decreases, respectively, the proportion of fecal deoxycholate). The concentration of DCA and UDCA should be monitored in the fecal water, which contains the biologically active portion of each bile acid.
The central hypothesis was that the inclusion of UDCA in the diet decreases the prevalence of colon carcinoma in mice with experimental ulcerative colitis.

   2.    Does the prevalence/multiplicity of colon carcinomas in the mouse model of ulcerative colitis increase when DCA/UDCA ratio in fecal water increases?
The second hypothesis of the project, based on the literature statements, was that the increase of DCA/UDCA ratio in fecal water enhances UC-associated colon carcinogenesis.

   3.    What is the influence of UDCA treatment on cell proliferation, apoptosis and the expression of PKC isozymes and COX-2?
The effect of UDCA on the proliferation as measured by the BrdU incorporation, on apoptosis and the expression of COX-2 and PKC should be determined.

How the questions were answered

   1.    Three groups of 25 mice were given 0.1 % dextran sulphate in drinking water for 7 days and pure water for 10 days and were fed a standard diet containing double iron concentration. In two groups, the diet was supplemented with 0.2% cholic acid (CA), the precursor of DCA, or with 0.4% UDCA. After 15 cycles the histology, the expression of MUC2, -catenin, p27 and p16 and the fecal water concentration of DCA and UDCA were investigated.
All animals showed colitis with similar severity and histologic as well as immunophenotypic alterations resembling those of human colitis. Among the animals fed the nonsupplemented diet 46% developed colorectal adenocarcinomas and 54% anal-rectal squamous cell carcinomas. In the animals given CA the prevalence of carcinomas was insignificantly lower. Among the mice fed with UDCA none developed adenocarcinomas and 20% squamous carcinomas. Dysplastic lesions were found in 88%, 67% and 40% of each group, respectively. The decrease of dysplasia and carcinoma prevalence correlated with the increase of UDCA concentration in the fecal water.

   2.    There was no correlation between the the concentration ration DCA/UDCA in the fecal water and the prevalence of carcinomas.

   3.    Inflammation was shown to suppress the cell cycle inhibitor protein p27. The treatment with UDCA did not affect the extent nor grade of inflammation but it prevented the inflammation-associated suppression of p27. The extent of epithelial proliferation and apoptosis is presently under investigation.

   4.    The cell proliferation was measured through BrdU incorporation, followed by immunohistiochemistry. The mice treated with UDCA exhibited a significantly lower incorporation of BrdU than the nontreated control mice.

List of significant results

   1.    UDCA treatment inhibits colitis-associated development of colonic adenocarcinomas in the animal model.

   2.    This inhibition is not the result of suppression of inflammation.

   3.    DCA/UDCA ratio does not affect colitis-associated colon carcinogenesis in the animal model.

   4.    The inflammation-induced suppression of the cyclin-dependent kinase inhibitor p27 is prevented by UDCA.

   5.    UDCA inhibits epithelial cell proliferation in the gut.

A lay summary of the final report

Chronic ulcerative colitis, depending on its extent and duration, increases 10-20 times the risk of developing colon cancer. Patients with diagnosed precancerous lesions usually undergo removal of the colon (colectomy) to prevent the development of the malignant tumor. Preventive measures which would inhibit or slow down the development of cancer are needed to avoid colectomy.

Patients with ulcerative colitis and primary sclerosing cholangitis (PSC), a condition associated with the obstruction of the bile ducts leading to the reduction of bile flow, are known to have a 3-5 fold higher risk of developing colon cancer than the ulcerative colitis patients without PSC. The treatment of PSC with ursodeoxycholic acid (UDCA) was recently shown to reduce the risk of colon cancer in these patients. Whether UDCA would also reduce the colon cancer risk of patients without PSC is not known at present and the analysis of this question in a clinical setting requires a long-term study.

In the present project this question was addressed by using mice with ulcerative colitis, which develop within several months adenocarcinomas, typical for ulcerative colitis, as well as squamous carcinomas, which do not occur in patients with colitis. The treatment of these mice with UDCA completely prevented the development of adenocarcinomas and significantly suppressed the development of squamous carcinomas. This model suggests, that UDCA may inhibit the development of colonic adenocarcinomas in patients with ulcerative colitis. It also allows to study the mechanism of the chemopreventive effect of UDCA in vivo.

Oulook

The present work, generously supported by the Broad Foundation, allowed us to show that oral administration of relatively high doses of UDCA completely prevent colitis associated carcinogenesis in mice. The dosage of the administered UDCA was, however, much higher than that achievable in patients. To analyse the applicability of the present finding to prevention of colitis-associated carcinogenesis in humans, we intend to analyse the dependence of the chemopreventive effect on the concentration of UDCA in fecal water. In addition, we will investigate the mechanism of UDCA action, in order to identify new potential targets for chemopreventive drugs.