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Background: The role of ion channels in carcinogenesis and tumour progression remains unclear. CLCA1 and 2 are members of a novel [Ca.sup.2+]-dependent chloride channel family that are expressed mainly in [...]

The clinical significance of detecting supposed tumour cell-derived mRNA transcripts in blood using the polymerase chain reaction (PCR) remains unclear. We have used a fully quantitative 5′-nuclease RT-PCR assay to screen for the expression of cytokeratins (ck) 19 and 20 and guanylyl cyclase C (GCC) in the peripheral blood of 21 healthy controls and 27 colorectal cancer patients. Expression of cytokeratin 19 and 20 mRNA was detected in 30% and 100% of samples, respectively, taken from healthy volunteers. There was no apparent difference in ck19 and ck20 mRNA transcription levels between controls and patients, or between patients with different Dukes' stages. While GCC mRNA was detected in only 1/21 control samples, it was expressed in approximately 80% of patients, although again there was no correlation between GCC levels and disease stage. Transcription levels of all three markers varied considerably between samples, even between samples taken from the same person at different times. We conclude that neither ck19 nor ck20 are reliable markers for the detection of colon epithelial cells in peripheral blood and that an evaluation of the usefulness of GCC awaits further longitudinal studies. [ABSTRACT FROM AUTHOR]

The article offers information on the occurrence of enteritis cystica profunda, a condition similar to that of bowel adenocarcinoma. In a study observed to a man with the suspected case of the disease, it was noted that the mucin-filled cysts in the bowel wall outside of the mucosal layer are thought to arise after mucusal ulceration or inflammatory damage. Likewise, it was emphasized that the condition is a benign entity and similar changes can be found along the gastrointestinal tract.

Abstract Background Prevalence of colorectal cancer (CRC) in the British Bangladeshi population (BAN) is low compared to British Caucasians (CAU). Genetic background may influence mutations and disease features. Methods We characterized the clinicopathological features of BAN CRCs and interrogated their genomes using mutation profiling and high-density single nucleotide polymorphism (SNP) arrays and compared findings to CAU CRCs. Results Age of onset of BAN CRC was significantly lower than for CAU patients (p=3.0 x 10-5) and this difference was not due to Lynch syndrome or the polyposis syndromes. KRAS mutations in BAN microsatellite stable (MSS) CRCs were comparatively rare (5.4%) compared to CAU MSS CRCs (25%; p=0.04), which correlates with the high percentage of mucinous histotype observed (31%) in the BAN samples. No BRAF mutations was seen in our BAN MSS CRCs (CAU CRCs, 12%; p=0.08). Array data revealed similar patterns of gains (chromosome 7 and 8q), losses (8p, 17p and 18q) and LOH (4q, 17p and 18q) in BAN and CAU CRCs. A small deletion on chromosome 16p13.2 involving the alternative splicing factor RBFOX1 only was found in significantly more BAN (50%) than CAU CRCs (15%) cases (p=0.04). Focal deletions targeting the 5’ end of the gene were also identified. Novel RBFOX1 mutations were found in CRC cell lines and tumours; mRNA and protein expression was reduced in tumours. Conclusions KRAS mutations were rare in BAN MSS CRC and a mucinous histotype common. Loss of RBFOX1 may explain the anomalous splicing activity associated with CRC.

Abstract Background High-degree microsatellite instability (MSI-H) is a feature of approximately 15% of sporadic colorectal cancers. Patients with MSI-H cancers have been reported to have a better prognosis than those with non-MSI-H cancers. The MSI-H subset is also characterised by a dense infiltrate of intra-epithelial lymphocytes and the hypothesis that the latter represents an efficacious immune response contributing to improved outcome is very attractive. Methods Data for this review were identified by searches of MEDLINE, PubMed, and cross references from relevant articles using the search terms 'microsatellite instability', 'colorectal cancer' and 'immunology', 'immune response' or 'immunogenicity'. Results A total of 38 articles were identified by the search criteria and a further 95 articles by cross-referencing. The relevance of the articles to be interviewed was established by hand searching. Out of a total of 133 articles identified, 47 articles were rejected due to lack of relevance. A total of 86 articles were included in the review, pertaining to microsatellite instability in colorectal cancer, and immune mechanisms in colorectal cancer. Conclusion It is suggested that this distinct group of colorectal cancers may have inherent immunogenic properties and that further elucidation of these may be invaluable to the development of successful immunotherapy.

Abstract Background Colorectal cancers displaying high-degree microsatellite instability (MSI-H) have an improved prognosis compared to microsatellite stable (MSS) cancers. The observation of pronounced lymphocytic infiltrates suggests that MSI-H cancers are inherently more immunogenic. We aimed to compare the gene expression profiles of MSI-H and MSS cancers to provide evidence for an activated immune response in the former. Results We analysed tissue from 133 colorectal cancer patients with full consent and Local Ethics Committee approval. Genomic DNA was analysed for microsatellite instability in BAT-26. High-quality RNA was used for microarray analysis on the Affymetrix® HG-U133A chip. Data was analysed on GeneSpring software version 6.0. Confirmatory real-time RT-PCR was performed on 28 MSI-H and 26 MSS cancers. A comparison of 29 MSI-H and 104 MSS cancers identified 2070 genes that were differentially expressed between the two groups [P < 0.005]. Significantly, many key immunomodulatory genes were up-regulated in MSI-H cancers. These included antigen chaperone molecules (HSP-70, HSP-110, Calreticulin, gp96), pro-inflammatory cytokines (Interleukin (IL)-18, IL-15, IL-8, IL-24, IL-7) and cytotoxic mediators (Granulysin, Granzyme A). Quantitative RT-PCR confirmed up-regulation of HSP-70 [P = 0.016], HSP-110 [P = 0.002], IL-18 [P = 0.004], IL-8 [0.002] and Granulysin [P < 0.0001]. Conclusions The upregulation of a large number of genes implicated in immune response supports the theory that MSI-H cancers are immunogenic. The novel observation of Heat Shock Protein up-regulation in MSI-H cancer is highly significant in light of the recognised roles of these proteins in innate and antigen-specific immunogenicity. Increased mRNA levels of pro-inflammatory cytokines and cytotoxic mediators also indicate an activated anti-tumour immune response.

Aims: To determine whether Gene Expression Profiling can accurately predict staging in Colorectal Cancer. Methods: Ethical approval was obtained from East London and City Health Authority (P/01/234 and P/02/167). 112 colorectal cancer patient samples were collected at operation, snap frozen and stored at -70°. 88 patients underwent curative/R0 resections. In the remaining 24 patients, non-curative/R1 or R2 resections were performed. Total RNA was extracted using the RNeasy mini kit (Quiagen) and quantitated and quality assessed using RNA Chips on the Agilent Bioanalyser 2100 (Agilent Technologies). Microarray experiments were carried out on Affymetrix HU133 A and B chips using the standard Affymetrix protocols. Data analysis was carried out using Genespring 5·02 (SiliconGenetics). Results: 86 statistically (P < 0·05) differentially expressed genes between the two groups were filtered by Welch test. Multiple testing correction was by Bonferroni. The 30 most differentially expressed genes were then selected using the Class Predictor Algorithm and all 112 cancers were clustered using these 30 genes. 66 patients were predicted accurately as having undergone curative or noncurative resections. In 39 patients the status could not be predicted. 18 patients, in whom curative/R0 surgery was performed, were predicted by the class algorithm as having undergone non-curative/R1 or R2 resections. Conclusion: We have identified a new group of patients that have been restaged on tumour expression profiling. These patients may have a worse prognosis than their conventional staging would predict and remain under intensive follow up. [ABSTRACT FROM AUTHOR]

Aims: A significant proportion of colorectal cancer patients undergoing curative surgery develop treatment failure despite the absence of microscopic metastases in their lymph nodes (LN). Use of quantitative real-time RT-PCR allowed the development of a novel, more sensitive molecular metastasis index (MI). Methods: This study was approved by the local Research Ethics Committee with informed consent obtained from the patient prior to surgery. Total RNA was prepared from lymph nodes after surgery for non-malignant (n = 17 patients, 143 LN) or malignant disease (n = 48 patients, 448 LN, 30 of which were histologically positive). The 5′-nuclease RT-PCR assays were performed in triplicate, on an ABI Prism 7700 detecting cytokeratin 20 (ck20), carcinoembryonic antigen (CEA) and guanylyl cyclase C (GCC) mRNA. Absolute mRNA copy numbers were calculated using amplicon-specific standard curves. Results: Specificity: In patients with benign disease, GCC, ck20 and CEA were detected in 17, 76 and 124/143 LN, respectively. Sensitivity: In patients with histologically involved LN, GCC, ck20 and CEA were detected in 29, 28 and 30/30 LN, respectively. There was good correlation between CEA and GCC mRNA copy numbers above a threshold level. The mRNA copy numbers were used to develop a formula establishing MI with 99 per cent specificity and 97 per cent sensitivity which detected LN metastases in an additional 8/32 Dukes A/B patients. Conclusion: The MI can reliably identify LN likely to harbour metastases. Whether it can also predict metastatic potential to identify the patients most at risk of treatment failure requires longer follow-up. [ABSTRACT FROM AUTHOR]

Background: Microsatellite instability (MSI) occurs in 15-20% of colorectal cancers and confers a survival advantage independent from tumour stage. This might reflect an improved immunological response against such tumours. Previously we have used real-time RT-PCR to demonstrate that MSI+ tumours show increased expression of activated T-cell markers. By extending this work to protein expression we can now gain a clearer understanding of the immune environment within these tumours. Aims: To test the hypothesis that MSI+ colorectal cancers elicit an increased antigen-specific immune response compared with MSI-cancers. Methods: We identified 25 MSI+ and 75 MSI-colorectal cancers, matched for patient age and tumour location. Immunohistochemistry was performed on paraffin embedded sections for T-cell markers, CD3, CD4 and CD8, the specific activation marker, CD25 and the effector protease Granzyme B (GZMB). Intraepithelial lymphocyte (IEL) counts per unit area were scored by a single observer blinded to microsatellite status and independently verified by a consultant pathologist. MSI+ and MSI-groups were compared with a nonparametric test (Mann-Whitney, two-tailed) and significance taken at the 5% level. Results presented as (median, 5th-95th percentiles). Results: MSI+ tumours had both higher CD3+ and CD8+ than MSI-tumours. Additionally, MSI+ tumours expressed higher CD25+ IEL's and GZMB levels. Conclusions: These results support the hypothesis that patients with MSI+ colorectal cancers exhibit increased antigen-specific immunological activity. Clearly, these findings may underscore the improved patient survival in MSI+ colorectal cancer. [ABSTRACT FROM AUTHOR]