Expression of MUC1, MUC2, MUC3 and MUC4 mucin mrnas in human pancreatic and intestinal tumor cell lines

We examined the steady-state expression levels of mRNA for the MUC1, MUC2, MUC3 and MUC4 gene products in 12 pancreatic tumor cell lines, 6 colon tumor cell lines, and one ileocecal tumor cell line. The results showed that 10 of 12 pancreatic tumor cell lines expressed MUCI mRNA and that 7 of these 12 lines also expressed relatively high levels of MUC4 mRNA. In contrast, MUC2 mRNA was expressed at only low levels and MUC3 was not detected in the pancreatic tumor cell lines. All 7 intestinal tumor cell lines examined expressed MUC2, and 5 of 7 expressed MUC3; however only one expressed significant levels of MUCI and 2 expressed low levels of M0C4 mRNA. This report of high levels of MUC4 mRNA expression by pancreatic tumor cells raises the possibility that mucin carbohydrate epitopes defined by antibodies such as DuPan 2 may be expressed on a second mucin core protein produced by pancreatic tumor cells. © 1994 Wiley-Liss, Inc.     

Gefitinib inhibits MUC5AC synthesis in mucin-secreting non-small cell lung cancer cells

Gefitinib, an epidermal growth factor receptor (EGFR) tyrosine kinase inhibitor, is an active agent in non-small cell lung cancer, and rapidly relieves bronchorrhea in patients with bronchioloalveolar carcinoma before the improvement of radiological findings. In addition, epidermal growth factor regulates mucin secretion in normal airway goblet cells. The present study was designed to clarify whether gefitinib modifies mucin production in lung cancer cell lines apart from its anti-proliferative effects, using A549 adenocarcinoma and NCI-H292 mucoepidermoid carcinoma cells expressing EGFR and MUC5AC mRNA. Mucin synthesis was measured by RT-PCR and ELISA, and MAPK and Akt, the downstream targets of EGFR, were examined by Western blotting assay. The clinically-achievable concentration of 1muM gefitinib inhibited the growth of both cells by only 10%, but gefitinib suppressed MUC5AC mRNA levels subsequent to a decrease in intracellular and secreted MUC5AC protein. Gefitinib also inhibited the phosphorylation of MAPK and Akt, and the selective inhibitors PD98059 and LY294002 also suppressed MUC5AC protein synthesis. These findings suggest that gefitinib may inhibits MUC5AC synthesis, at least in part, through MAPK and Akt signaling pathways. Thus, gefitinib inhibits mucin production, which is encouraging for trials involving its use against bronchorrhea in patients with lung cancer.     

Mucin gene expression in colonic tissues and cell lines.

Complementary DNA clones encoding four different mucin core peptides have been isolated. However, the expression of these mucin genes in the colon has not been systematically studied. The present investigation used Northern blot analysis to study the expression of MUC1, MUC2, MUC3, and MUC4 mRNA in paired normal and cancerous colonic tissues, and nine colon cancer cell lines. Results were correlated with the clinicopathological features of the tumors and with the immunohistochemical expression of several carbohydrate tumor-associated antigens that may reside on mucins. MUC1 mRNA was expressed in all colonic tissues, and levels in paired normal and cancer tissues were similar in most cases. MUC2 and MUC3 mRNAs were expressed in both normal and cancer tissues, but levels were often decreased in the cancers. MUC4 mRNA was present in normal mucosa with comparable or sometimes greater expression in cancers. There was no apparent correlation between the expression of any particular mucin gene or pattern of mucin genes and the site, stage, or histological type of tumor. In addition, the expression of mucin-associated carbohydrate antigens did not correlate with any individual mucin gene or group of mucin genes. In colon cancer cell lines all four MUC genes were expressed rather weakly or not at all. These results indicate that the human colon expresses a broad repertoire of mucin genes which are differentially regulated in malignancy. Whether this differential regulation of mucin genes affect the behavior of the tumor and results in the altered glycosylation commonly seen in these requires further investigation.     

Heterogeneity of MUC1 expression by human breast carcinoma cell lines in vivo and in vitro

Increased expression of the epithelial mucin MUC1 has been linked to tumor aggressiveness in human breast carcinoma. Recent studies have demonstrated that overexpression of MUC1 interferes with cell-substrate and cell-cell adhesion by masking cell surface integrins and E-cadherin. Additionally, the cytoplasmic tail of MUC1 is involved in signal transduction and interactions with catenins. In the present study, we have examined the in vitro expression of MUC1 mRNA and protein in a panel of 14 human breast cancer cell lines using northern blotting, western blotting, immunocytochemistry, and flow cytometry. Considerable variability of expression was noted not only between cell lines but also within several individual lines. Many cell lines such as BT 20, KPL-1, and T47D expressed abundant MUC1 whilst others such as MDA-MB-231 and MCF-7 showed intermediate expression, and MDA-MB-435 and MDA-MB-453 expressed very low levels. Low levels of MUC1 expression were associated with decreased expression of cytokeratin and increased expression of vimentin. Additionally, 12 of the cell lines were established as xenografts in immunocompromised (SCID) mice, and MUC1 expression in both the primary tumors as well as metastases was assessed immunohistochemically. In general, in vivo expression mirrored in vitro expression, although there was reduced in vivo expression in T47D and ZR-75-1 xenografts. Although we showed no correlation between tumorigenicity or metastasis and MUC1 expression, this study will assist development of experimental models to assess the influence of MUC1 of on breast cancer progression.     

Real-time RT-PCR detection of CK19, CK7 and MUC1 mRNA for diagnosis of lymph node micrometastases in non small cell lung carcinoma

Metastatic lymph nodes (LNs) are the major prognostic factor in resected non small cell lung carcinoma (NSCLC). However, almost 50% of pN0 patients relapse, suggesting metastatic cells undetected by current staging procedures. A combination of markers [cytokeratins 19 and 7 (CK19, CK7) and mucin type 1 (MUC1) mRNAs] was therefore evaluated by real-time RT-PCR in order to detect occult cancer cells. Forty-three NSCLC tumor samples, 4 micrometastatic, 6 metastatic and 84 histologically negative mediastinal LNs from 19 patients with NSCLC were evaluated as well as blood mononuclear cells from 29 healthy volunteers and 17 benign LNs. When tested on cell lines, RT-PCR was particularly efficient for evaluation of CK19, CK7 and MUC1 mRNA expression. All tumor samples were positive for at least 1 marker and 74% of samples were positive for all 3 markers. CK7 and CK19 mRNA were not detected in benign LN and blood cells from healthy donors in contrast with MUC1 mRNA. Only CK7 and CK19 mRNA were therefore used for evaluation of mediastinal LNs: the 6 histologically metastatic and the 4 micrometastatic LNs were positive for at least one marker. Among the 84 histologically negative LNs, 6 (7%) were positive for at least one marker, potentially changing the stage of 2 out of 19 patients. In conclusion, in our feasibility study, parallel molecular detection of CK19 and CK7 mRNA can be considered a specific diagnostic tool for the assessment of microscopic lymphatic spread. Its prognostic impact remains to be evaluated in a prospective study.     

Expression and immunogenicity of a tumor-associated antigen, 90K/Mac-2 binding protein,in lung carcinoma

BACKGROUND: The authors attempted to obtain shared proteins among lung carcinoma cells by column chromatographies. A glycoprotein with approximately 500 kDa isolated from QG56 cells showed an identical amino acid sequence to 90K/Mac-2 binding protein (M2BP). This protein has been reported to be highly expressed and to modulate the expression of surface molecules involved in immune responses on cultured cancer cells. Therefore, it would be beneficial for M2BP to be targeted in cancer immunotherapy. METHODS: The authors analyzed the expression of M2BP in lung carcinoma cells and M2BP's immunogenicity as a tumor antigen. Eight cultured lung carcinoma cell lines and 28 tumor tissues from patients with lung carcinoma were examined for the expression of M2BP mRNA and protein. Sera from cancer patients (n = 23) and healthy donors (n = 19) were studied for their reactivity to M2BP peptides by enzyme-linked immunosorbent assay. RESULTS: Seven of the 8 (87.5%) lung carcinoma cell lines and 17 of the 28 (60.7%) tumor tissues expressed high levels of M2BP mRNA. Most of the M2BP mRNA-positive cancer cell lines and tumors also showed M2BP protein expression. The serum levels of antibodies to M2BP were elevated in 30.4% of the patients. In addition, M2BP-specific immunoglobulin G was observed in all patients with anti-M2BP antibodies. CONCLUSIONS: M2BP is highly expressed in lung carcinoma cells and is sufficiently immunogenic to elicit specific immunity to this molecule in patients with lung carcinoma. M2BP is expected to be useful as a tumor marker and a target antigen in cancer immunotherapy.     

Heterogeneity of MUC1 expression by human breast carcinoma cell lines in vivo and in vitro

Increased expression of the epithelial mucin MUC1 has been linked to tumor aggressiveness in human breast carcinoma. Recent studies have demonstrated that overexpression of MUC1 interferes with cell-substrate and cell–cell adhesion by masking cell surface integrins and E-cadherin. Additionally, the cytoplasmic tail of MUC1 is involved in signal transduction and interactions with catenins. In the present study, we have examined the in vitro expression of MUC1 mRNA and protein in a panel of 14 human breast cancer cell lines using northern blotting, western blotting, immunocytochemistry, and flow cytometry. Considerable variability of expression was noted not only between cell lines but also within several individual lines. Many cell lines such as BT 20, KPL-1, and T47D expressed abundant MUC1 whilst others such as MDA-MB-231 and MCF-7 showed intermediate expression, and MDA-MB-435 and MDA-MB-453 expressed very low levels. Low levels of MUC1 expression were associated with decreased expression of cytokeratin and increased expression of vimentin. Additionally, 12 of the cell lines were established as xenografts in immunocompromised (SCID) mice, and MUC1 expression in both the primary tumors as well as metastases was assessed immunohistochemically. In general, in vivo expression mirrored in vitro expression, although there was reduced in vivo expression in T47D and ZR-75-1 xenografts. Although we showed no correlation between tumorigenicity or metastasis and MUC1 expression, this study will assist development of experimental models to assess the influence of MUC1 of on breast cancer progression.     

结肠直肠癌组织中粘蛋白MUC1和MUC2的表达及临床意义

背景与目的：粘蛋白MUC1的表达上调或MUC2的表达下调可能参与了肿瘤的发生,而MUC1和MUC2的表达中国人结肠直肠癌发生发展的关系未见报道。本研究的目的是探讨结肠直肠癌组织中粘蛋白MUC1和MUC2的表达与不同的临床病理参数间的关系及其临床意义。方法：采用免疫组化方法检测126例结肠直肠癌和20例正常结肠直肠粘膜中粘蛋白MUC1和MUC2的表达。结果：20例正常结肠直肠粘膜中粘蛋白MUC1和MUC2的表达阳性率分别为0和100％,而126例结肠直肠癌中粘蛋白MUC1和MUC2的阳性表达率分别为42．1％和36．5％（P＜0．01）;结肠直肠癌中粘蛋白MUC1的表达与肿瘤的浸润深度、淋巴结转移和Dukes分期呈正相关（P＜0．05）,与肿瘤的分化程度呈负相关（P＜0．01）;粘蛋白MUC2在结肠直肠癌中的表达与肿瘤的浸润浓度、淋巴结转移和Dukes分期密切相关（P＜0．05）,而与肿瘤的分化程度无关（P＞0．05）。结论：粘蛋白MUC1表达上调或MUC2表达下调可能参与了结肠直肠癌的发生发展,检测结肠直肠癌中粘蛋白MUC1和MUC2的表达可间接反映肿瘤的预后。     

MUC4 expression correlates with poor prognosis in small-sized lung adenocarcinoma

The mortality of lung cancer remains high, despite improved diagnostic techniques that allow small lung tumors to be detected. In this study, we evaluated the prognostic significance of the tracheal mucin MUC4 by immunohistochemical investigation of the expression profiles of MUC4, ErbB2, p27 and MUC1 in lung adenocarcinoma specimens (non-bronchiolo-alveolar type, < or =3cm) from 185 patients. MUC4 is a membrane mucin, similarly to MUC1, and in addition MUC4 functions as an intra-membrane ligand for receptor tyrosine kinase ErbB2 and is associated with regulation of p27. However, MUC4 expression was found to be unrelated to expression of MUC1, ErbB2 and p27 in small-sized lung adenocarcinomas. The disease-free interval (DFI) and survival rate of 25 patients with high MUC4 expression (> or =25% of neoplastic cells stained) were significantly lower than those of 160 patients with low MUC4 expression (<25% of neoplastic cells stained) (P<0.05), whereas ErbB2 and p27 expression showed no significant correlation with DFI and survival. Univariate analysis showed that high MUC4 and p27 expression correlated with blood vessel invasion (P=0.0004), and MUC4 expression was frequently detected in regions of stromal invasion. In addition, the survival rate of stage IA patients with high MUC4 expression was significantly lower than that of stage IA patients with low MUC4 expression (P<0.05). In conclusion, high MUC4 expression in small-sized lung adenocarcinomas correlates with a short DFI and a poor survival rate. Therefore, MUC4 expression might be a new independent factor for prediction of outcome and indication of poor prognosis in lung adenocarcinoma.     

结直肠癌组织中黏蛋白MUC2和MUC3的表达及临床意义

目的:探讨散发性结直肠癌组织中MUC2和MUC3的表达与临床病理参数间的关系及其临床意义。方法:采用免疫组化SP法检测90例结直肠癌和30例正常结直肠黏膜组织中MUC2和MUC3的表达。结果:30例正常结直肠黏膜组织中黏蛋白MUC2和MUC3的表达阳性率均为100％,而90例结直肠癌组织中MUC2和MUC3的阳性表达率分别为35．5％和52．2％(P< 0．05);MUC2和MUC3的表达与肿瘤的浸润深度、淋巴结转移和Dukes分期密切相关(P<0．05),黏蛋白MUC3与肿瘤的分化程度呈负相关(P<0．05)。结论:黏蛋白MUC2和MUC3在人结直肠癌组织中表达下调。     

Mucin 21 is a key molecule involved in the incohesive growth pattern in lung adenocarcinoma

Decreased cell adhesion has been reported as a significant negative prognostic factor of lung cancer. However, the molecular mechanisms responsible for the cell incohesiveness in lung cancer have not yet been elucidated in detail. We herein describe a rare histological variant of lung adenocarcinoma consisting almost entirely of individual cancer cells spreading in alveolar spaces in an incohesive pattern. A whole exome analysis of this case showed no genomic abnormalities in CDH1 or other genes encoding cell adhesion molecules. However, whole mRNA sequencing revealed that this case had an extremely high expression level of mucin 21 (MUC21), a mucin molecule that was previously shown to inhibit cell‐cell and cell‐matrix adhesion. The strong membranous expression of MUC21 was found on cancer cells using mAbs recognizing different O‐glycosylated forms of MUC21. An immunohistochemical analysis of an unselected series of lung adenocarcinoma confirmed that the strong membranous expression of MUC21 correlated with incohesiveness. Thus, MUC21 could be a promising biomarker with potential diagnostic and therapeutic applications for lung adenocarcinoma showing cell incohesiveness.     

脂质体包裹MUC2反义核酸抑制胃癌细胞增殖的初步探讨

背景与目的：我们前期研究发现粘蛋白MUC2在胃癌组织中的表达与肿瘤的生物学行为密切相关。本研究拟观察MUC2反义脱氧寡核苷酸(antisense oligodeoxynucleotide,ASODN)对人胃癌细胞SGC7901生长抑制作用。方法：应用硫代磷酸修饰的MUC2 ASODN经阳离子脂质体包裹后转染入SGC7901细胞,采用MTT法、形态学观察测定MUC2 ASODN对SGC27901细胞的增殖抑制作用,免疫组化检测细胞中MUC2和p16蛋白的表达情况。结果：不同浓度ASODN均能抑制SGC7901细胞的增殖,在48h抑制作用最强,0．5μmol／L ASODN对细胞的抑制率为55％,随时间延长抑制作用逐渐减弱。SGC7901细胞转染MUC2 ASODN后,与对照组相比,光镜下观察到细胞数量减少、体积变小、核分裂明显减少、可见较多的坏死。透射电镜下见线粒体肿胀、细胞内脂滴增多、髓样结构、染色质边集等。免疫组化SP法染色显示转染ASODN后,SGC7901细胞MUC2蛋白表达水平明显降低,p16蛋白表达明显增强。结论：使用人工合成MUC2 ASODN能有效抑制人胃癌细胞SGC790l的增殖。     

MUC6 expression in breast tissues and cultured cells: Abnormal expression in tumors and regulation by steroid hormones

Neoplastic transformation of epithelial cells is commonly associated with alterations in the expression of mucin genes. The mechanisms involved in this process are largely unknown. MUC6, isolated from a stomach cDNA library, is mainly expressed in stomach antral glands, as detected by using in situ hybridization and immunohistochemistry. We examined MUC6 expression in normal and pathological breast tissues using immunohistochemistry with MUC6-specific antibodies and in cultured breast cancer cells using immunocytochemistry and Northern blotting. MUC6 was generally not detected in normal breast (1/11) but was detected in fibrocystic disease without atypia (7/17, 41%), in atypical fibrocystic disease (11/11, 100%) and in carcinoma (57/60, 95%). To study the mechanisms involved in mucin gene up-regulation in breast cancer, we examined baseline, growth-related and steroid-induced levels of MUC1, MUC3 and MUC6 in 4 breast cancer cell lines, 2 of which express estrogen receptors. MUC6 levels were up-regulated at post-confluence in 2/4 cell lines, whereas no changes were detected for the other mucin genes examined. MUC6 and MUC3 were constitutively expressed, and steroid-induced, in BT-474 and MCF-7 cells, respectively. As a control, pS2 was induced in both cell lines. Our results indicate that (1) MUC6 is overexpressed in breast cancer and in benign breast disease, (2) in vitro, MUC6 and MUC3 are up-regulated by steroids and (3) abnormal expression of MUC6 in breast cancers may, in part, be explained by hormonal changes associated with tumor development. Int. J. Cancer 77:193–199, 1998.© 1998 Wiley-Liss, Inc.     

Quantitative analysis of MUC1 and MUC5AC mRNA in pancreatic juice for preoperative diagnosis of pancreatic cancer

Pancreatic juice is a promising type of diagnostic sample for pancreatic cancer, and members of the mucin (MUC) family are diagnostic candidates. To evaluate the utility of MUC family members as diagnostic markers, we measured MUC mRNA expression in pancreatic tissues and pancreatic juice obtained from patients with different pancreatic diseases as well as in pancreatic cancer cell lines by real-time PCR. Furthermore, to support the possibility of early diagnosis by quantification of MUC1 and MUC5AC, immunohistochemistry and microdissection-based quantitative analysis of mRNA were carried out. There was no significant correlation between MUC1 and MUC5AC expression in cell lines. When beta-actin was used as a reference gene, median MUC1 and MUC5AC mRNA expression levels were remarkably greater in tumoral tissues than in non-tumoral tissues, but median MUC4 and MUC6 mRNA expression levels were not. Receiver operating characteristic curve analysis showed that quantitative analysis of MUC1 and MUC5AC mRNA in pancreatic juice is better diagnostic modality than that of MUC4 and MUC6 mRNA. Immunohistochemistry showed that MUC1 and MUC5AC were highly expressed in invasive ductal carcinomas (IDC) and moderately expressed in high-grade pancreatic intraepithelial neoplasia (PanIN); no staining was observed in normal ducts. Analysis of cells isolated by microdissection showed stepwise upregulation of MUC1 and MUC5AC in the development of high-grade PanIN to IDC. Our results suggest that MUC1 and MUC5AC are upregulated stepwise in pancreatic carcinogenesis and that quantitative assessment of MUC1 and MUC5AC mRNA in pancreatic juice has high potential for preoperative diagnosis of pancreatic cancer.