muramidase and Colorectal-Neoplasms

The tumor suppressor FBW7 targets oncoproteins such as c-MYC for ubiquitylation and is mutated in several human cancers. We noted that in a substantial percentage of colon cancers, FBW7 protein is undetectable despite the presence of FBW7 mRNA. To understand the molecular mechanism of FBW7 regulation in these cancers, we employed proteomics and identified the deubiquitinase (DUB) USP9X as an FBW7 interactor. USP9X antagonized FBW7 ubiquitylation, and Usp9x deletion caused Fbw7 destabilization. Mice lacking Usp9x in the gut showed reduced secretory cell differentiation and increased progenitor proliferation, phenocopying Fbw7 loss. In addition, Usp9x inactivation impaired intestinal regeneration and increased tumor burden in colitis-associated intestinal cancer. c-Myc heterozygosity abrogated increased progenitor proliferation and tumor burden in Usp9x-deficient mice, suggesting that Usp9x suppresses tumor formation by regulating Fbw7 protein stability and thereby reducing c-Myc. Thus, we identify a tumor suppressor mechanism in the mammalian intestine that arises from the posttranslational regulation of FBW7 by USP9X independent of somatic FBW7 mutations.

Topics: Animals; Colorectal Neoplasms; Endopeptidases; F-Box-WD Repeat-Containing Protein 7; Gene Expression Regulation, Enzymologic; Gene Expression Regulation, Neoplastic; HCT116 Cells; Humans; Mice; Mice, Mutant Strains; Muramidase; Mutation; Peptide Fragments; Proto-Oncogene Proteins c-myc; Tumor Suppressor Proteins; Ubiquitin Thiolesterase

Novel biomarker verification assays are urgently required to improve the efficiency of biomarker development. Benefitting from lower development costs, multiple reaction monitoring (MRM) has been used for biomarker verification as an alternative to immunoassay. However, in general MRM analysis, only one sample can be quantified in a single experiment, which restricts its application. Here, a Hyperplex-MRM quantification approach, which combined mTRAQ for absolute quantification and iTRAQ for relative quantification, was developed to increase the throughput of biomarker verification. In this strategy, equal amounts of internal standard peptides were labeled with mTRAQ reagents Δ0 and Δ8, respectively, as double references, while 4-plex iTRAQ reagents were used to label four different samples as an alternative to mTRAQ Δ4. From the MRM trace and MS/MS spectrum, total amounts and relative ratios of target proteins/peptides of four samples could be acquired simultaneously. Accordingly, absolute amounts of target proteins/peptides in four different samples could be achieved in a single run. In addition, double references were used to increase the reliability of the quantification results. Using this approach, three biomarker candidates, ademosylhomocysteinase (AHCY), cathepsin D (CTSD), and lysozyme C (LYZ), were successfully quantified in colorectal cancer (CRC) tissue specimens of different stages with high accuracy, sensitivity, and reproducibility. To summarize, we demonstrated a promising quantification method for high-throughput verification of biomarker candidates.

Topics: Adenosylhomocysteinase; Adult; Aged; Amino Acid Sequence; Biomarkers, Tumor; Calibration; Cathepsin D; Colorectal Neoplasms; Female; Humans; Male; Middle Aged; Muramidase; Peptide Fragments; Reference Standards; Reproducibility of Results; Sensitivity and Specificity; Staining and Labeling; Tandem Mass Spectrometry

Statistics from a 64 case study showed that mucinous adenocarcinoma was apt to invade the intestinal wall and to metastasize to lymph nodes (P < 0.05). The activity of arylsulfatase and lysozyme of mucinous adenocarcinoma was stronger than that of the papillary and tubular adenocarcinoma (P < 0.05). In RR staining for electron microscopic observation, a significant decrease of proteoglycan granules was found in the surrounding matrix of mucinous adenocarcinoma, which correlated with the amount of arylsulfatase and lysozyme secreted by mucinous adenocarcinoma. These enzymes reduced the degree of sulfation in heparan sulfate and degraded proteoglycans. The proteoglycan structural barrier having been destroyed, facilitates mucinous adenocarcinoma to infiltrate and metastasize.

Topics: Adenocarcinoma; Adenocarcinoma, Mucinous; Adenocarcinoma, Papillary; Arylsulfatases; Colorectal Neoplasms; Humans; Lymphatic Metastasis; Muramidase; Neoplasm Invasiveness; Proteoglycans

High levels of lysozyme have been reported to be present in the feces of patients with colorectal cancer. It has been suggested that fecal lysozyme could prove useful in the early detection of colorectal cancer, but that further work is needed. The present investigation reports on the measurement of fecal lysozyme in 23 colorectal cancer patients and 39 healthy controls. The mean fecal lysozyme level for the cases was 13.3 +/- 2.8 micrograms/g stool compared with 12.5 +/- 2.6 micrograms/g stool for the controls. The median level for the cases was 8.7 micrograms/g stool compared with 6.6 micrograms/g stool for the controls. Forty-three percent of the cases and 31% of the controls had fecal lysozyme levels above 10 micrograms/g stool. None of these differences were statistically significant. The results of this study indicate that fecal lysozyme would be of no use in the early detection of colorectal cancer.

Topics: Adenoma; Adult; Age Factors; Aged; Aged, 80 and over; Biomarkers, Tumor; Clinical Enzyme Tests; Colorectal Neoplasms; Feces; Female; Humans; Male; Middle Aged; Muramidase; Odds Ratio

In the last few years, an increase in the incidence of malignant tumours of the large bowel and the rectum has been observed in all advanced countries. Intensive search is therefore going on for biochemical and immunochemical markers which would help early diagnosis and monitoring of this disease. The most frequently used markers in colorectal cancer are CEA, CSi, CRP, CA 19-9 and sialic acid. The present study points out the possibility and advantage of using lysozyme determination in serum as another marker in patients with colorectal cancer.

Topics: Aged; Biomarkers, Tumor; Clinical Enzyme Tests; Colorectal Neoplasms; Female; Humans; Male; Middle Aged; Muramidase

Normal colonic epithelial cells consist of several cell types or lineages that are thought to arise from a common stem cell precursor. Neoplastic transformation may occur at different stages in the differentiation of a colonic stem cell to produce tumors that may retain characteristic cell lineage phenotypes. In this study, immunohistochemical techniques were used to identify cell lineage-related markers in fetal, normal, hyperplastic, adenomatous, and cancerous colonic tissue. These markers consisted of secretory component (columnar cells), a purified mucin antigen (mucous or goblet cells), chromogranin A (enteroendocrine cells), lysozyme (Paneth cells), and carcinoembryonic antigen (panepithelial cell marker). Colonic neoplasms, like normal mucosa, predominantly expressed the markers of columnar and goblet cell lineages. Chromogranin A was expressed in a small population of cells in most normal and fetal colonic crypts. Chromogranin A reactive cells were found in 55% of hyperplastic polyps, 31% of adenomatous polyps, and 33% of carcinomas. Lysozyme reactivity was rare in fetal, normal, and hyperplastic specimens, but was present in 86% of adenomas and 40% of carcinomas. Of 42 primary carcinomas, 9% were "pluripotent" and expressed markers of all four cell lineages. In addition to columnar and goblet cell markers, 7% expressed both enteroendocrine and Paneth cell markers, 17% expressed enteroendocrine cell markers, and 24% expressed Paneth cell markers. Two cases (5%) lacked expression of any of the cell lineage markers. The remainder expressed only columnar and goblet cell markers. The markers used in this study appear to identify the major cell lineages of fetal and normal colonic epithelium and can be used to delineate the altered cell lineage phenotypes in premalignant and malignant colonic mucosa.

Topics: Biomarkers, Tumor; Carcinoembryonic Antigen; Cell Transformation, Neoplastic; Chromogranin A; Chromogranins; Colon; Colonic Polyps; Colorectal Neoplasms; Humans; Immunoenzyme Techniques; Intestinal Mucosa; Mucins; Muramidase; Secretory Component; Stem Cells