Screen-detected colorectal cancers are associated with an improved outcome compared with stage-matched interval cancers.

BACKGROUND: Colorectal cancers (CRCs) detected through the NHS Bowel Cancer Screening Programme (BCSP) have been shown to have a more favourable outcome compared to non-screen-detected cancers. The aim was to identify whether this was solely due to the earlier stage shift of these cancers, or whether other factors were involved. METHODS: A combination of a regional CRC registry (Northern Colorectal Cancer Audit Group) and the BCSP database were used to identify screen-detected and interval cancers (diagnosed after a negative faecal occult blood test, before the next screening round), diagnosed between April 2007 and March 2010, within the North East of England. For each Dukes' stage, patient demographics, tumour characteristics, and survival rates were compared between these two groups. RESULTS: Overall, 322 screen-detected cancers were compared against 192 interval cancers. Screen-detected Dukes' C and D CRCs had a superior survival rate compared with interval cancers (P=0.014 and P=0.04, respectively). Cox proportional hazards regression showed that Dukes' stage, tumour location, and diagnostic group (HR 0.45, 95% CI 0.29-0.69, P<0.001 for screen-detected CRCs) were all found to have a significant impact on the survival of patients. CONCLUSIONS: The improved survival of screen-detected over interval cancers for stages C and D suggest that there may be a biological difference in the cancers in each group. Although lead-time bias may have a role, this may be related to a tumour's propensity to bleed and therefore may reflect detection through current screening tests.

Faecal calprotectin levels before and after weight loss in obese and overweight subjects.

Faecal calprotectin (FCP) is a non-invasive biomarker of intestinal inflammation, levels of which are reported to be elevated in individuals with increased body mass index (BMI). We investigated whether weight loss (WL), induced by dietary and behavioural change in a community weight loss programme (Slimming World), was associated with a reduction in FCP in a longitudinal cohort study. We obtained paired stool samples at the start and 11-15 weeks into the weight loss programme in 40 individuals with a BMI >25 kg m(-2) and no known colorectal or systemic inflammatory condition. Eight of 40 (20%) 'healthy' participants had a baseline FCP greater than 50 µg g(-1) (the accepted adult cutoff value for FCP), termed FCP(high). Although the degree (%) of WL was similar (5.1 FCP(high) versus 6.1 FCP(normal); P=0.63), only FCP(high) individuals, but not FCP(normal) participants, demonstrated a reduction in FCP level (median 0.3-fold versus 1.0-fold (that is, no change) during the study period (P=0.065). A large prospective cohort study of FCP(high) and FCP(normal) obese individuals is now required to determine the relationship between local mucosal inflammation and the systemic inflammatory state associated with obesity, as well as understand the relationship between FCP levels, WL and future risk of colorectal neoplasia.

Omega-3 polyunsaturated fatty acids for the treatment and prevention of colorectal cancer.

Omega (ω)-3 polyunsaturated fatty acids (PUFAs) are naturally occurring substances that are well tolerated and have been used extensively for the prevention of cardiovascular disease. More recently, ω-3 PUFAs have been recognised to have anticancer activity. There is also evidence suggesting improved efficacy and/or tolerability of conventional cancer chemotherapy when administered with ω-3 PUFAs. The purpose of this review is to (i) describe the mechanisms by which ω-3 PUFAs are thought to have antineoplastic activity, (ii) review published preclinical and clinical studies that support anti-colorectal cancer activity and (iii) summarise current clinical trials investigating the potential therapeutic role(s) of ω-3 PUFAs at different stages of colorectal carcinogenesis, from adenoma (polyp) prevention to treatment of established malignant disease and prevention of cancer recurrence.

Plasma and rectal mucosal oxylipin levels during aspirin and eicosapentaenoic acid treatment in the seAFOod polyp prevention trial.

BACKGROUND: Aspirin and eicosapentaenoic acid (EPA) have colorectal polyp prevention activity, alone and in combination. This study measured levels of plasma and rectal mucosal oxylipins in participants of the seAFOod 2 × 2 factorial, randomised, placebo-controlled trial, who received aspirin 300 mg daily and EPA 2000 mg free fatty acid, alone and in combination, for 12 months. METHODS: Resolvin (Rv) E1, 15-epi-lipoxin (LX) A4 and respective precursors 18-HEPE and 15-HETE (with chiral separation) were measured by ultra-high performance liquid chromatography-tandem mass spectrometry in plasma taken at baseline, 6 months and 12 months, as well as rectal mucosa obtained at trial exit colonoscopy at 12 months, in 401 trial participants. RESULTS: Despite detection of S- and R- enantiomers of 18-HEPE and 15-HETE in ng/ml concentrations, RvE1 or 15­epi-LXA4 were not detected above a limit of detection of 20 pg/ml in plasma or rectal mucosa, even in individuals randomised to both aspirin and EPA. We have confirmed in a large clinical trial cohort that prolonged (12 months) treatment with EPA is associated with increased plasma 18-HEPE concentrations (median [inter-quartile range] total 18-HEPE 0.51 [0.21-1.95] ng/ml at baseline versus 0.95 [0.46-4.06] ng/ml at 6 months [P<0.0001] in those randomised to EPA alone), which correlate strongly with respective rectal mucosal 18-HEPE levels (r = 0.82; P<0.001), but which do not predict polyp prevention efficacy by EPA or aspirin. CONCLUSION: Analysis of seAFOod trial plasma and rectal mucosal samples has not provided evidence of synthesis of the EPA-derived specialised pro-resolving mediator RvE1 or aspirin-trigged lipoxin 15­epi-LXA4. We cannot rule out degradation of individual oxylipins during sample collection and storage but readily measurable precursor oxylipins argues against widespread degradation.

Prostaglandin E2-EP4 receptor signalling promotes tumorigenic behaviour of HT-29 human colorectal cancer cells.

The predominant product of cyclooxygenase (COX) activity in the colon, prostaglandin (PG) E2 promotes intestinal tumorigenesis. Expression of the PGE2 receptor EP4 is upregulated during colorectal carcinogenesis. Therefore, we investigated the role of elevated PGE2-EP4 receptor signalling in the protumorigenic activity of PGE2 by increasing EP4 receptor expression in HT-29 human colorectal cancer (CRC) cells (HT-29-EP4) by stable transfection. Elevated PGE2-induced EP4 receptor activity in HT-29 cells increased resistance to spontaneous apoptosis and promoted anchorage-independent growth, but had no effect on proliferation of HT-29-EP4 cells. EP4 receptor activation by PGE2 in HT-29-EP4 cells also led to development of fluid-filled cysts, which was associated with increased tight junction protein (occludin and zonula occludens-1) expression. Overexpression of the EP4 receptor in HT-29 cells led to basal EP4 receptor signalling in the absence of exogenous PGE2, which was explained by autocrine activity of endogenous, COX-2-derived PGE2 and constitutive, ligand-independent EP4 receptor activity. The predominant signalling pathway mediating antiapoptotic activity downstream of PGE2-EP4 receptor activation in HT-29-EP4 cells was elevation of cyclic adenosine monophosphate (cAMP) levels, which was associated with phosphorylation of cAMP-response element binding protein. EP4 receptor activation led to a small increase in phosphorylated extracellular signal-regulated kinase (ERK) 2 protein levels but inhibition of ERK phosphorylation did not abrogate the antiapoptotic activity of PGE2. However, PGE2-EP4 receptor signalling did not lead to trans-activation of the epidermal growth factor receptor in HT-29 cells. Inhibition of protumorigenic PGE2-EP4 receptor signalling represents a potential strategy for anti-CRC therapy that may avoid the toxicity associated with systemic COX inhibition.

Abnormal T cell differentiation persists in patients with rheumatoid arthritis in clinical remission and predicts relapse.

OBJECTIVES: An abnormal CD4+ T cell subset related to inflammation exposure (inflammation-related cells, IRC) has been identified in rheumatoid arthritis (RA). Patients with inflammatory and non-inflammatory diseases were used to examine the relationship between inflammation and this T cell subset in vivo. METHODS: Blood was collected from healthy controls and patients with RA (active disease or in clinical remission), Crohn's disease and osteoarthritis. IRC and chemokine receptors were quantified by flow cytometry. Thymic activity and apoptotic factors were measured by real-time polymerase chain reaction. Circulating cytokines were measured by enzyme-linked immunosorbent assay. CXCR4 and SDF1 in synovial biopsies were measured using immunohistochemistry. RESULTS: IRC were identified in patients with RA (p<0.0001) and Crohn's disease (p = 0.005), but not in those with osteoarthritis. In RA in remission, IRC persisted (p<0.001). In remission, hyperproliferation of IRC was lost, chemokine receptor expression was significantly lowered (p<0.007), Bax expression dropped significantly (p<0.001) and was inversely correlated with IRC (rho = -0.755, p = 0.03). High IRC frequency in remission was associated with relapse within 18 months (OR = 6.4, p<0.001) and a regression model predicted 72% of relapse. CONCLUSIONS: These results suggest a model in which, despite the lack of systemic inflammation, IRC persist in remission, indicating that IRC are an acquired feature of RA. They have, however, lost their hyper-responsiveness, acquired a potential for survival, and no longer express chemokine receptors. IRC persistence in remission confirms their important role in chronic inflammation as circulating precursors of pathogenic cells. This was further demonstrated by much higher incidence of relapse in patients with high IRC frequency in remission.

Activity of the non-steroidal anti-inflammatory drug indomethacin against colorectal cancer.

A substantial body of evidence from rodent colon carcinogenesis models, in vitro experiments with human colorectal cancer cells and limited clinical observations in humans suggest that the non-steroidal anti-inflammatory drug indomethacin has anti-colorectal cancer activity. However, although many mechanisms of the anti-neoplastic activity of indomethacin have been suggested, e.g., cyclooxygenase inhibition and peroxisome proliferator-activated receptor gamma activation, the precise relevance of the majority of in vitro pharmacological observations to the in vivo anti-neoplastic activity of indomethacin remains unclear. Herein, we review the existing literature describing the chemopreventative and chemotherapeutic efficacy of indomethacin against colorectal cancer, and draw together the disparate literature describing potential mechanisms of action of indomethacin in human colorectal cancer cells in vitro. Although indomethacin itself has significant adverse effects, including serious upper gastrointestinal toxicity, the development of novel derivatives that may have an improved safety profile means that further investigation of the anti-colorectal cancer activity of indomethacin is warranted.

Cyclooxygenase-2 expression in colorectal cancer liver metastases.

Cyclooxygenase-2 (COX-2) is up-regulated in 85-90% of primary human colorectal cancers and is a putative target for the chemopreventative activity of non-steroidal anti-inflammatory drugs. However, COX-2 expression by human colorectal cancer liver metastases has been poorly characterized. We studied a consecutive series of 38 patients who underwent liver resection for metastatic disease, for whom long-term (up to 57 months), prospective follow-up data were available. Semi-quantitative immunohistochemistry for COX-2 was performed on 54 metastases from 35 patients, for whom adequate histological material was available. Diffuse cytoplasmic staining for COX-2 protein was detected in cancer cells in 100% of metastases (COX-2 score 1, n = 25; score 2, n = 29). There was no relationship between metastasis size or differentiation grade and the level of COX-2 protein expression. There was no difference in colorectal cancer-free or overall survival between patients with high (score 2) and low (score 1) COX-2 scores (Kaplan-Meier survival analysis and log rank test, both P = 0.97). Multivariate Cox regression analysis identified age, incomplete resection and presence of extra-hepatic disease as independent predictors of disease-free and overall survival following surgery. COX-2 protein was also localized to a subset of stromal fibroblasts and mononuclear cells within metastases as well as hepatocytes from resection specimens. COX-2 protein was expressed by cancer cells in all human colorectal cancer liver metastases which were studied. Investigation of the effect of selective COX-2 inhibition on metastasis growth and metastasis cancer cell proliferation/apoptosis in vivo are warranted.

The effect of non-steroidal anti-inflammatory drugs on human colorectal cancer cells: evidence of different mechanisms of action.

Non-steroidal anti-inflammatory drugs (NSAIDs) inhibit proliferation and induce apoptosis in human colorectal cancer cells in vitro. It remains unclear whether individual NSAIDs act by cyclooxygenase-2 (COX-2) inhibition and how NSAIDs exert their anti-proliferative effects. We investigated the effects of NS-398 (a selective COX-2 inhibitor), indomethacin (a non-selective COX inhibitor) and aspirin on four human colorectal cancer cell lines (HT29.Fu, HCA-7, SW480 and HCT116). NS-398 completely inhibited proliferation, induced G1 arrest and promoted apoptosis in COX-2-expressing cells (HT29.Fu and HCA-7). However, indomethacin had similar effects on all cells, regardless of COX-2 expression. NS-398 also had anti-proliferative activity on COX-2-negative cell lines (SW480 and HCT116). Aspirin inhibited proliferation of all cell lines but did not induce apoptosis. Indomethacin decreased beta-catenin protein expression in all cells (unlike NS-398 or aspirin). NSAIDs act on human colorectal cancer cells via different mechanisms. Decreased beta-catenin protein expression may mediate the anti-proliferative effects of indomethacin.

Non-steroidal anti-inflammatory drugs inhibit Helicobacter pylori-induced human neutrophil reactive oxygen metabolite production in vitro.

BACKGROUND: Helicobacter pylori infection is associated with increased production of gastric mucosal reactive oxygen metabolites which have been implicated in mucosal damage and carcinogenesis. In vitro, neutrophils produce reactive oxygen metabolites following activation by H. pylori. Non-steroidal anti-inflammatory drugs (NSAIDs) inhibit neutrophil activation by several factors, e.g. N-formyl-methionyl-leucyl-phenyalanine (f-MLP). AIM: To examine the effect of NSAIDs on H. pylori-induced reactive oxygen metabolite production by human peripheral blood neutrophils. METHODS: Neutrophils were stimulated by H. pylori (NCTC 11637) water extract or f-MLP in the presence or absence of NSAIDs. Reactive oxygen metabolite activity was measured by luminol-enhanced chemiluminescence. RESULTS: H. pylori water extract stimulated a sevenfold increase in chemiluminescence which was inhibited dose-dependently by diclofenac. All six NSAIDs studied (at 10-4 M) significantly inhibited H. pylori-and f-MLP-stimulated neutrophil reactive oxygen metabolite production. Meclofenamic acid and diclofenac had the greatest inhibitory effects on both H. pylori and f-MLP-stimulated neutrophil reactive oxygen metabolite production. The inhibitory effects of other NSAIDs varied with the activation stimulus. NSAIDs did not quench reactive oxygen metabolites generated in a cell-free xanthine:xanthine oxidase assay. CONCLUSION: Several NSAIDs attenuate H. pylori-induced neutrophil reactive oxygen metabolites production in vitro. This may be relevant to a potential chemopreventative role in gastric cancer and to a possible lack of synergy between H. pylori and NSAID use regarding peptic ulceration.

Percutaneous endoscopic gastrostomy (PEG): change in practice since 1988.

BACKGROUND AND AIMS: We previously reported a 30-day mortality following percutaneous endoscopic gastrostomy (PEG) of 8% (1988-92). Concerns over increasing mortality rates prompted us to survey current practice compared with 1988-92: assess case mix, outcome, risk factors for early death, and review practice guidelines. METHODS: 78 consecutive adults were referred for PEG over 7 months. Baseline characteristics, including age and functional status (Barthel Index), and outcome at 30 and 180 days were prospectively evaluated. RESULTS: 74 patients. Median age 69 years; male 55%. Major underlying diagnoses: cerebrovascular disease 42%, head and neck tumours 19%, motor neurone disease 4% (33%, 16% and 27% in 1988-92). Mortality rates at 30, 90 and 180 days were 19%, 35% and 42% respectively (8%, 20% and 37% in 1988-92). Univariate analysis showed that age >75 years, Barthel Index <1 and Glasgow Coma Scale < or =10 were significant risk factors for death at 30 days: odds ratios (95% confidence intervals) 3.9 (1.1-13), 5.9 (1.4-25) and 4.4 (1.2-15) respectively. CONCLUSIONS: 30-day mortality was increased from 8% to 19% between 1988-92 and 1998-99 reflecting a change in referral patterns: more elderly with cerebrovascular disease and fewer with motor neurone disease. Age and functional status should be considered when advising on PEG feeding.

The omega-3 polyunsaturated fatty acid eicosapentaenoic acid inhibits mouse MC-26 colorectal cancer cell liver metastasis via inhibition of PGE2-dependent cell motility.

BACKGROUND AND PURPOSE: The omega-3 polyunsaturated fatty acid (PUFA) eicosapentaenoic acid (EPA) has antineoplastic activity at early stages of colorectal carcinogenesis, relevant to chemoprevention of colorectal cancer (CRC). We tested the hypothesis that EPA also has anti-CRC activity at later stages of colorectal carcinogenesis, relevant to treatment of metastatic CRC, via modulation of E-type PG synthesis. EXPERIMENTAL APPROACH: A BALB/c mouse model, in which intrasplenic injection of syngeneic MC-26 mouse CRC cells leads to development of liver metastases, was used. Dietary EPA was administered in the free fatty acid (FFA) form for 2 weeks before and after ultrasound-guided intrasplenic injection of 1 × 10(6) MC-26 cells (n= 16 each group). KEY RESULTS: Treatment with 5% (w w(-1)) EPA-FFA was associated with a reduced MC-26 mouse CRC cell liver tumour burden compared with control animals (median liver weight 1.03 g vs. 1.62 g; P < 0.034). Administration of 5% EPA-FFA was also linked to a significant increase in tumour EPA incorporation and lower intratumoural PGE(2) levels (with concomitant increased production of PGE(3)). Liver tumours from 5% EPA-FFA- treated mice demonstrated decreased 5-bromo-2-deoxyuridine-positive CRC cell proliferation and reduced phosphorylated ERK 1/2 expression at the invasive edge of tumours. A concentration-dependent reduction in MC-26 CRC cell Transwell® migration following EPA-FFA treatment (50-200 µM) in vitro was rescued by exogenous PGE(2) (10 µM) and PGE(1)-alcohol (1 µM). CONCLUSIONS AND IMPLICATIONS: EPA-FFA inhibits MC-26 CRC cell liver metastasis. EPA incorporation is associated with a 'PGE(2) to PGE(3) switch' in liver tumours. Inhibition of PGE(2)-EP(4) receptor-dependent CRC cell motility probably contributes to the antineoplastic activity of EPA.

Differential CD74 (major histocompatibility complex Class II invariant chain) expression in mouse and human intestinal adenomas.

CD74 (major histocompatibility complex (MHC) Class II invariant chain) has recently been identified as the cell-surface receptor for the pro-tumorigenic cytokine macrophage migration inhibitory factor (MIF). Therefore, we investigated CD74 gene expression in intestinal adenomas in Apc(Min/+) mice and humans. CD74 mRNA (p31 and p41 splice variants) and immunoreactive CD74 protein levels were significantly lower in small intestinal and colonic Apc(Min/+) mouse adenomas compared with histologically normal mucosa. These findings were mirrored by a reduction in MHC Class II expression and Class II trans-activator type IV transcripts. Conversely, CD74 protein levels were actually increased in dysplastic epithelial cells in 47/55 (85%) human colorectal adenomas, with CD74 and MIF protein levels together predicting increasing dysplasia in individual adenomas (P=0.003). Down-regulation of CD74 during Apc(Min/+) mouse intestinal tumorigenesis does not model increased CD74 expression at the early, benign stages of human colorectal carcinogenesis. Epithelial cell CD74 represents a valid target for anti-CRC therapy.

Lack of interleukin-4 receptor alpha chain-dependent signalling promotes azoxymethane-induced colorectal aberrant crypt focus formation in Balb/c mice.

Interleukin (IL)-4 receptor (IL-4R) alpha chain-dependent signalling by IL-4 and IL-13 promotes tumour growth and metastasis in mouse models of colorectal cancer. However, the role of IL-4R alpha-dependent signalling during the early, pre-malignant stages of colorectal carcinogenesis has not been investigated. Therefore, we investigated the effect of deletion of the IL-4R alpha gene on azoxymethane-induced colorectal aberrant crypt focus (ACF) multiplicity and size in Balb/c mice. IL-4R alpha(-/-) mice developed significantly more ACFs [median 8, inter-quartile range (IQR) 4-11.5; n = 9] than wild-type (WT) animals (median 4, IQR 1-6; n = 9; p = 0.04, Mann-Whitney U-test). There were significantly higher levels of IL-4 in serum from azoxymethane- and sham-treated IL-4R alpha(-/-) mice than WT animals, but no difference in serum IL-13 levels. In the absence of functional IL-4Rs, IL-13 can also signal via the IL-13R alpha2 receptor, leading to induction of transforming growth factor (TGF) beta, which has pro-tumourigenic activity at early stages of intestinal tumourigenesis. We found that mucosal TGFbeta mRNA levels and intestinal epithelial cell TGFbeta immunoreactivity were significantly higher in IL-4R alpha(-/-) mice than in WT animals. In summary, IL-4R alpha-dependent signalling has a protective, anti-neoplastic role during the post-initiation phase of azoxymethane-induced colorectal carcinogenesis in Balb/c mice. Our data should prompt thorough investigation of the role of IL-4R alpha-dependent signalling during human colorectal carcinogenesis, particularly as antagonism of IL-4R signalling represents a therapeutic strategy for asthma and other allergic diseases.

Regulation of stromal cell cyclooxygenase-2 in the ApcMin/+ mouse model of intestinal tumorigenesis.

Cyclooxygenase-2 (Cox-2) is expressed predominantly by stromal cells in intestinal adenomas from the Apc(Min/+) mouse model of familial adenomatous polyposis. We investigated the mechanistic basis of stromal cell Cox-2 expression in Apc(Min/+) mouse adenomas, as well as Cox-2 expression and activity in histologically normal (HN) Apc(Min/+) mouse intestine, in order to gain further insights into regulation of Cox-2 as a potential chemoprevention target. Upregulation of Cox-2 in intestinal tumours is not an intrinsic feature of Apc(Min/+) macrophages as bone marrow-derived Apc(Min/+) macrophages did not exhibit an abnormality in Cox-2 expression or activity. Intestinal permeability to lactulose or mannitol was similar in Apc(Min/+) mice and wild-type littermates, implying that macrophage activation by luminal antigen is unlikely to explain stromal cell Cox-2 induction. Moreover, stromal cells exhibited differential expression of Cox-2 and inducible nitric oxide synthase, suggesting 'alternative' (M2) rather than 'classical' (M1) macrophage activation. Flow cytometric sorting of isolated stromal mononuclear cells (SMNCs), on the basis of M-lysozyme and specific macrophage marker expression, demonstrated that macrophages, neutrophils and non-myelomonocytic cells all contributed to lamina propria prostaglandin (PG) E(2) synthesis. However, the majority of PGE(2) synthesis by macrophages was via a Cox-2-dependent pathway compared with predominant Cox-1-derived PGE(2) production by non-myelomonocytic cells. SMNCs from HN Apc(Min/+) intestinal mucosa exhibited similar levels of Cox-2 mRNA and protein, but produced more Cox-2-derived PGE(2) than wild-type cells at 70 days of age. There was an age-dependent decline in PGE(2) synthesis by Apc(Min/+) SMNCs, despite tumour progression. These data suggest that other Cox-2-independent factors also control PGE(2) levels during Apc(Min/+) mouse intestinal tumorigenesis. Regulation of macrophage Cox-2 expression and other steps in PGE(2) synthesis (e.g. PGE synthase) are valid targets for novel chemoprevention strategies that could minimize or avoid systemic COX-2 inhibition.

Activation of peroxisome proliferator-activated receptor gamma does not explain the antiproliferative activity of the nonsteroidal anti-inflammatory drug indomethacin on human colorectal cancer cells.

The mechanism of the anticolorectal cancer activity of the nonsteroidal anti-inflammatory drug indomethacin is poorly understood. Indomethacin inhibits both cyclooxygenase (COX) isoforms, but it may also act via COX-independent targets. Indomethacin can bind and activate the transcription factor peroxisome proliferator-activated receptor (PPAR) gamma. Moreover, natural and synthetic PPARgamma ligands can induce growth arrest and apoptosis of human colorectal cancer cells in vitro. Therefore, we tested the hypothesis that the antiproliferative activity of indomethacin on human colorectal cancer cells in vitro is explained by a PPARgamma-dependent mechanism of action. Human colorectal cancer cell lines SW480 and HCT116 both expressed functional PPARgamma. Indomethacin directly activated PPARgamma in both cell lines (HCT116 > SW480). A dominant-negative PPARgamma strategy was used to demonstrate that endogenous PPARgamma represses proliferation of HCT116 cells (compatible with tumor suppressor activity) but that the presence of functional PPARgamma is not necessary for the antiproliferative activity (or reduction in cyclin D1 protein) associated with indomethacin in vitro. In summary, indomethacin (>100 microM) directly activates PPARgamma in human colorectal cancer cells. However, PPARgamma activation does not underlie the antineoplastic activity of indomethacin on human colorectal cancer cells in vitro.

Effect of nonsteroidal anti-inflammatory drugs on beta-catenin protein levels and catenin-related transcription in human colorectal cancer cells.

Elevated beta-catenin levels in human colorectal cancer (CRC) cells lead to increased trans-activation of 'protumorigenic' beta-catenin/T-cell factor (TCF) target genes such as cyclin D1. Therefore, possible targets for the anti-CRC activity of nonsteroidal anti-inflammatory drugs (NSAIDs) are beta-catenin and catenin-related transcription (CRT). We tested the antiproliferative activity and the effects on levels of beta-catenin and cyclin D1 protein, as well as CRT (measured using a synthetic beta-catenin/TCF-reporter gene [TOPflash]), of a panel of NSAIDs (indomethacin, diclofenac, sulindac sulphide and sulphone, rofecoxib; range 10-600 microM) on SW480 human CRC cells in vitro. Following NSAID treatment, there was no consistent relationship between reduced cell proliferation, induction of apoptosis and changes in beta-catenin protein levels or CRT. All the NSAIDs, except rofecoxib, decreased nuclear beta-catenin content and cyclin D1 protein levels in parallel with their antiproliferative activity. However, cyclin D1 downregulation occurred prior to a decrease in total beta-catenin protein levels and there was no correlation with changes in CRT, suggesting the existence of CRT-independent effects of NSAIDs on cyclin D1 expression. In summary, NSAIDs have differential effects on beta-catenin protein and CRT, which are unlikely to fully explain their effects on cyclin D1 and their antiproliferative activity on human CRC cells in vitro. British Journal of Cancer (2004) 91, 153-163. doi:10.1038/sj.bjc.6601901 www.bjcancer.com Published online 8 June 2004