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Mucins synthesized by malignant cells may contribute (via decreased cellular adhesion and immune recognition) to cancer invasion and metastases. Human mucins are derived from a heterogeneous family of genes, labeled MUC1-6. Our aim was to determine the pattern of mucin gene expression in normal, preneoplastic (intestinal metaplasia), and malignant gastric specimens. Probes and antibodies for specific mucin tandem repeat sequences were used for RNA and immunohistochemical analysis. Normal stomach mucosa was characterized by expression of MUC1, MUC5, and MUC6 mRNA and immunoreactive protein, without MUC2, MUC3, and MUC4 gene expression. In contrast, high levels of MUC2 and MUC3 mucin mRNA and immunoreactive protein were found in specimens with intestinal metaplasia. Gastric cancers exhibited markedly altered secretory mucin mRNA levels compared with adjacent normal mucosa, with decreased levels of MUC5 and MUC6 mRNA and increased levels of MUC3 and MUC4 mRNA. Overall, immunoreactive MUC1 mucin was detected in 72% of 33 gastric cancers, and secretory mucin core peptides were expressed in 34% (MUC2), 45% (MUC3), 19% (MUC5), and 57% (MUC6) of these specimens. Coexpression of multiple (three or more) mucin core proteins occurred in 15 of 25 (60%) advanced (stages III and IV) cancers compared with 1 of 8 (12.5%) early (stages I and II) cancers (P < 0.048). We conclude that human gastric epithelium has a unique mucin gene pattern, which becomes markedly altered in preneoplastic and neoplastic specimens. Increased mucin gene heterogeneity in gastric adenocarcinomas is associated with advanced cancer stage.

To determine the relative expression of distinct mucin genes in normal and neoplastic tissue, antibodies and cDNA probes that recognize the core tandem repeat sequences of membrane-bound (MUC1) and secreted (MUC2 and MUC3) mucins were used for immunohistochemical and RNA Northern and slot-blot analysis. MUC1 mRNA was detected in all epithelial tissues tested. MUC1 core peptide, recognized by monoclonal antibodies 139H2 and DF3, was highly expressed on apical membranes of bronchus, breast, salivary gland, pancreas, prostate, and uterus, and was sparsely expressed in gastric surface cells, gallbladder, small intestine, and colonic epithelium. In contrast, MUC2 and MUC3 gene expression was primarily restricted to the intestinal tract. MUC2 mRNA was highly expressed in normal jejunum, ileum, and colon, compared with very low levels in normal bronchus and gallbladder. MUC3 mRNA was highly expressed in normal jejunum, ileum, colon, and gallbladder. Immunohistochemical studies using antibodies against synthetic MUC2 (anti-MRP) and MUC3 (anti-M3P) peptides indicate that MUC2- and MUC3-producing cells in the gastrointestinal tract are distinct. Goblet cells of the small intestine and colon reacted strongly with anti-MRP, whereas M3P reactivity was restricted to columnar cells of small intestinal villi, surface colonic epithelium, and gallbladder. Mucin protein epitopes and mRNA levels were frequently altered in adenocarcinomas compared to corresponding normal tissues. Alterations included increased expression, aberrant expression, and, less frequently, loss of expression. Increased MUC1 immunoreactivity was observed in most adenocarcinomas of the breast, lung, stomach, pancreas, prostate, and ovary. In addition, with the exception of prostate cancer, focal aberrant expression of MUC2 and MUC3 epitopes was frequently observed. Increased MUC1, MUC2, and MUC3 epitopes were present in colon adenocarcinomas of all histological subtypes, with the greatest increase of MUC2 epitopes observed in colloid (mucinous) colon cancers. MUC2 or MUC3 mRNA levels were increased in colloid colon cancer compared with normal colon, however in well- and moderately well-differentiated colon cancers MUC1, 2 and 3 mRNA levels were decreased. Compared with corresponding normal tissue, MUC1 mRNA levels were increased in breast cancer and well-differentiated lung cancers, and MUC3 mRNA was increased in gastric adenocarcinomas. Normal stomach lacked both MUC2 and MUC3 immunoreactivity and mRNA, however, MUC2 and MUC3 proteins and mRNA were highly expressed in gastric intestinal metaplasia. In conclusion, mucin genes are independently regulated and their expression is organ- and cell type-specific. Furthermore, neoplastic transformation is associated with dys-regulated expression of both membrane-bound and secreted mucin core protein epitopes and may be due to altered mucin mRNA levels and/or altered mucin glycosylation.

Pancreatic cancer mucins have several carbohydrate antigens that are potentially useful in the detection of pancreatic cancers, but little is known about the core polypeptides of pancreatic cancer mucins. In this study, purified mucin from SW1990 pancreatic cancer xenografts was deglycosylated by treatment with hydrogen fluoride to give pancreatic cancer apomucin. Consistent with near-complete removal of carbohydrate, the apomucin had 10- to 70-fold decreased binding of lectins and, unlike the native mucin, served as an acceptor for polypeptidyl N-acetylgalactosaminyl transferase. Antibodies prepared against the apomucin did not bind to native mucin, and antibodies that bound to native mucin did not bind to apomucin. On the basis of cross-reaction with deglycosylated colon cancer mucin and intestinal mucin repeat peptide, apomucins from SW1990 pancreatic cancer xenografts contain the intestinal mucin repeat peptide. On the basis of binding of breast cancer-reactive monoclonal antibodies 139H2, DF3, and HMFG-2, apomucins from SW1990 pancreatic cancer xenografts also have the mammary mucin repeat peptide. Using complementary DNA probes specific for intestinal mucin and breast mucin sequences, both types of apomucin mRNA were detected in nude mouse xenografts of SW1990 cells. In immunohistochemical staining, antibody against deglycosylated SW1990 mucin stained normal breast and pancreas but not normal colon. Some pancreatic and mammary cancers and most colonic cancers, however, were stained by antibodies against both intestinal apomucin and mammary apomucin. We conclude that pancreatic cancers can produce mucins with the intestinal repeat peptide as well as those with mammary repeat peptide sequences.

Sodium butyrate induces morphological and biochemical changes consistent with a more differentiated phenotype in some colon cancer cell lines. These changes include increased expression of carcinoembryonic antigen (CEA) and other oncodevelopmental markers. We utilized domain-specific probes and polyclonal antibodies against CEA-related antigens to study sodium butyrate-induced expression of the CEA gene family in a villous adenoma-derived cell line, which is nontumorigenic in nude mice (VACO 235), and two colonic carcinoma cell lines known to respond to sodium butyrate exposure by phenotypic differentiation (HT-29 and LS 174T). The induction begins as quickly as 24 h after exposure and occurs primarily at a transcriptional level, although some translational control is also evident. No evidence was found for gene amplification, rearrangement, or methylation to account for the mechanism of this transcriptional control. [35S]Cysteine pulse-labeled cell lysate immunoblots and polyadenylated RNA blot hybridization suggest that increases in mRNA transcript and CEA-related glycoprotein levels are primarily due to increased synthesis rather than decreased degradation. A considerable amount of heterogeneity is seen in the biosynthesis of the CEA-related glycoproteins, with each cell line showing a distinct pattern of CEA-related antigen expression from a limited number of mRNA transcripts.