Almudena Fernández-Briera

Expression and enzyme activity of α(1,6)fucosyltransferase in human colorectal cancer

Changes in enzyme activity and the expression levels of α(1,6)fucosyltransferase [α(1,6)FT] have been reported in certain types of malignant transformations. To develop a better understanding of the role of α(1,6)FT in human colorectal carcinoma (CRC), we analysed the enzyme activity in healthy and tumour tissues. α(1,6)FT activity was considerably higher in tumour tissue than in healthy tissue and was related to gender, lymph node metastasis, type of growth and tumour stage. We also observed a significant increase in the α(1,6)FT expression in tumour tissues as compared to healthy and transitional tissues, inflammatory lesions and adenomas. The immunohistochemical expression in tumour tissues was correlated with the degree of infiltration through the intestinal wall. Finally, a statistical correlation was found between enzyme activity and expression obtained by Western blot in colorectal tumours when compared in the same patient. All these findings demonstrate an alteration of α(1,6)FT activity and expression in CRC.

Correlation Analysis between Tumorous Associated Antigen Sialyl-Tn Expression and ST6GalNAc I Activity in Human Colon Adenocarcinoma

Objectives: Sialyl-Tn (sTn) is a mucin carbohydrate-associated antigen that is strongly expressed in a large number of colorectal carcinomas. In this study, we combined immunohistochemical and enzymatic techniques in order to find the correlation between sTn tissue expression and the sialyltransferase activity (ST6GalNAc I) responsible for its synthesis in colorectal cancer (CRC) patients. Methods: We compared sTn expression in healthy (n = 46), tumorous (n = 60) and transitional tissue (n = 46) from CRC patients, and correlated sTn altered expression with clinicopathologic variables of the patient. Furthermore, we determined ST6GalNAc I tissue activity employing asialo-ovine submaxillary mucin (asialo-OSM) as glycoprotein acceptor (n = 27). Results: The rates of sTn positive expression obtained for healthy, tumorous and transitional tissues were 15, 67 and 63%, respectively. These rates led to statistically significant differences between healthy and tumorous or transitional tissue (p = 0.001); sTn expression was related to the first stages of the tumor invasion in transitional tissue. As regards ST6GalNAc I activity, we found an enhancement in transitional tissue. Statistical correlation analysis did not reveal association between sTn expression and ST6GalNAc I activity. Conclusions: Our findings indicated that sTn antigen tissue expression and ST6GalNAc I activity levels were not correlated in CRC, in spite of the overexpression of the antigen in tumorous and transitional tissue.

Clinical Interest of the Combined Use of Serum CD26 and Alpha-L-Fucosidase in the Early Diagnosis of Colorectal Cancer

The purpose of this study was to assess if the combination of CD26 and alpha-L-fucosidase has a role in the diagnosis of colorectal cancer, paying particular attention to the stages in which the tumour is not yet disseminated. CD26 concentration and alpha-L-fucosidase activity were determined in sera from 110 colorectal cancer patients and 46 donors. The combination of CD26 and alpha-L-fucosidase showed a specificity of 100% with a sensitivity of 64% in the diagnosis of colorectal cancer. Interestingly, the combination of both markers had a sensitivity of 75% in the stage I at the highest specificity (100%), providing also high sensitivity levels for the other non-disseminated stages (66% for stages II and III). In conclusion, the combined use of CD26 and alpha-L-fucosidase offers high sensitivity with high specificity in the diagnosis of colorectal cancer, especially at the earliest stage (TNM I).

α(1,6)Fucosyltransferase expression is an independent prognostic factor for disease-free survival in colorectal carcinoma ☆

We previously reported that α(1,6)fucosyltransferase (Enzyme class 2.4.1.68) activity and expression are increased in colorectal cancer, suggesting a role for this enzyme in tumor development and progression. However, the possible impact of α(1,6)fucosyltransferase activity or expression on clinical outcomes in colorectal cancer patients has never been studied. Thus, the present study was conducted to determine the value of α(1,6)fucosyltransferase as a prognostic factor for colorectal cancer. α(1,6)Fucosyltransferase expression was analyzed using immunohistochemistry in 141 colorectal tumors, and α(1,6)fucosyltransferase activity was determined in 39 tumors. A complete standardized follow-up of patients was documented until the end of the observation period of 5 years or patient death. Univariate analysis demonstrated the absence of a correlation between enzyme activity and disease evolution. However, in patients with moderate or strong α(1,6)fucosyltransferase expression, a significant decrease in the overall (P = .04) and disease-free (P = .03) survival rates was observed. In addition, when local and distant disease recurrence were considered separately, enzyme expression was found to correlate with local tumor recurrences (P = .01). Furthermore, multivariate analysis showed that α(1,6)fucosyltransferase expression has independent value for predicting tumor recurrences and, specifically, local recurrences. These findings suggest that α(1,6)fucosyltransferase expression may be a good indicator of poor prognosis in colorectal cancer and, therefore, a helpful tool to choose the most effective treatment.

Effect of Human Colorectal Carcinogenesis on the Neural Cell Adhesion Molecule Expression and Polysialylation

Objective: Although downregulation of neural cell adhesion molecule (NCAM) has been correlated with poor prognosis in colorectal cancer (CRC), it is also possible that colon cancer spreading comes from reducing tumor cell adhesion through NCAM polysialylation, as occurs in lung carcinoma or Wilms’ tumor. Methods: To prove this hypothesis, we have performed a prospective study on tumor and control specimens from 39 CRC patients, which were immunostained for NCAM and PSA (polysialic acid) expression. Results: Tumor versus control expression of NCAM and PSA epitopes in tissue specimens, as well as correlation between tumor expression and clinicopathological features, were statistically analyzed. Results showed a low constitutive expression of NCAM and PSA (PSA-NCAM) in control tissue, which reached a statistically significant increase in the tumor tissue. Likewise, the presence and number of lymph node metastases at surgery were correlated with NCAM expression and PSA/NCAM coexpression. Conclusions: These data highlight the importance of taking into account PSA-associated epitopes when dealing with NCAM cell expression studies in tumor development and progression. The analysis of PSA and NCAM expression in CRC suggests a new way, other than downregulation of NCAM, in order to escape contact inhibition and promote cell tumor spreading in colorectal cancer.

Superoxide dismutase and catalase: tissue activities and relation with age in the long-lived species Margaritifera margaritifera.

Free radicals are extremely reactive and produce damage and modify cell functions. Furthermore, superoxide dismutase and catalase are believed to play a key role in the enzymatic defence of the cells. Indeed, some authors have argued that reduced free-radical damage could explain increased longevity. Margaritifera margaritifera is one of the longest-lived animals in the world (up to 100-200 years). Furthermore, this organism may serve as a useful model for gerontologists interested in exploring the mechanisms that promote long life and the slowing of senescence. The present study estimated for the first time individual enzymatic activity for superoxide dismutase isozymes (Cu,Zn-SOD and Mn-SOD) and catalase in tissue preparations of gills, digestive glands and mantles of two natural populations of M. margaritifera. Superoxide dismutase activities showed significant differences in the tissues analysed of specimens from the same river and in specimens from different rivers for the same tissue. Catalase activity levels also showed significant variation, but differences among tissues, within tissues or between rivers were of relatively little interest. We failed to find any relationship between individual enzymatic activities and the age estimated for each mussel. Indeed, the wide variation found in activity levels can be principally interpreted as an adaptation to the unpredictable and changing nature of freshwater natural habitats.

Preoperative Serum Alpha-L-Fucosidase Activity as a Prognostic Marker in Colorectal Cancer

Objectives: The aim of this study was to examine the prognostic value of the preoperative serum α-L-fucosidase (AFU) activity in colorectal cancer and to assess whether it could add prognostic information that Dukes’ stages do not give. Methods: A postoperative follow-up of 137 colorectal cancer patients was performed, and survival analyses were carried out to evaluate the impact of AFU activity on disease-free survival. Dukes’ stage classification, preoperative serum carcinoembryonic antigen levels and six other clinicopathological features of the patients were also analysed. Results: In previous studies, we have stressed the diagnostic value of AFU activity in preoperatively obtained serum from colorectal cancer patients. In the present work, we have found that the enzymatic activity of serum AFU was not related to the Dukes’ stage of the primary tumour, but it was associated with the type of metastasis and recurrence of the disease. The mean value of preoperative serum AFU activity was higher in patients with distant metastases than in those with lymph node or peritoneal metastases, or without metastasis (p = 0.034). After a mean postoperative follow-up period of 22 months, three groups of patients with different recurrence rates could be distinguished (p = 0.0014). Similar results were found when only patients in Dukes’ stage B (p = 0.0439) or C (p = 0.0122) were considered. Conclusions: According to our findings, serum AFU activity appears to be a good prognostic factor of tumour recurrence in colorectal carcinoma. Furthermore, patients in Dukes’ stage B or C at high or very high risk of tumour recurrence could be spotted.

Synthesis and expression of CDw75 antigen in human colorectal cancer

BackgroundIncreased ST6Gal I activity has been associated with the α(2,6)sialylation enhancement of membrane glycoconjugates observed in metastatic colorectal carcinomas (CRC). Siaα(2,6)Galβ(1,4)GlcNAc sequence, known as CDw75, is a sialylated carbohydrate determinant generated by the ST6Gal I. This epitope has been reported to be associated with the progression of gastric and colorectal tumours, hence there are only a few conclusive studies to date.MethodsBy radioisotopic techniques we evaluated the ST6Gal I activity in healthy, transitional and tumour tissues from 43 patients with CRC. By immunohistochemistry we assessed the CDw75 expression in 25 colorectal adenomas, 43 tumours, 13 transitional and 28 healthy tissues of CRC patients.ResultsST6Gal I activity was likewise found to be statistically higher in tumour tissue respect to healthy tissue from CRC patients. CDw75 expression was positive in 20% of colorectal adenomas. Furthermore, 70% of tumour specimens and 8.3% of transitional specimens were positive for CDw75 expression, whereas none of the healthy ones showed the presence of the epitope.ConclusionThe major contribution of this study is the inclusion of data from transitional tissue and the analysis of CDw75 antigen expression in CRC and in colorectal adenomas, little known so far. ST6Gal I activity and CDw75 antigen expression were increased in CRC. Although their comparison did not reach the statistical significance, a great extent of patients showed both, an enhanced tumour ST6Gal I activity and an increased CDw75 expression in the tumour tissue. So, these two variables may play a role in malignant transformation. The expression of CDw75 in colorectal adenomas suggests that this antigen may be a tumour marker in CRC.

ALTERATIONS OF GLYCOSIDASES IN HUMAN COLONIC ADENOCARCINOMA

OBJECTIVES We have carried out a detailed study of some glycosidases in an attempt to explain the differential profile of enzyme activity between human colonic adenocarcinoma and normal mucosa. DESIGN AND METHODS Several glycosidase activities associated with human colonic adenocarcinoma and control tissues were submitted to a detailed structural and functional characterization. RESULTS Tumoral and control samples were assayed for beta-D-galactosidase, beta-D-glucuronidase, alpha-D-mannosidase, beta-NAc-D-glucosaminidase and beta-NAc-D-galactosaminidase activities. Tumoral tissue showed higher beta-D-galactosidase, beta-NAc-D-glucosaminidase, and beta-NAc-D-galactosaminidase activities than control tissue. Glycosidases from tumoral and control tissues demonstrated no differences in optimum pH, subcellular distribution, pH and thermal stability. However, the kinetic analysis showed a statistically significant increased Vmax in tumoral colon with respect to the control for beta-D-galactosidase, beta-NAc-D-glucosaminidase, and beta-NAc-D-galactosaminidase activities. The Km remained unaltered. CONCLUSIONS The increased Vmax detected for some glycosidase activities in human colonic adenocarcinoma could correspond with a greater presence of enzyme proteins in the tumoral cells, and not to changes in protein and/or active site structure.

Elevation of ST6Gal I Activity in Malignant and Transitional Tissue in Human Colorectal Cancer

Objectives: The aim of the present study was to investigate the activity of CMP-NeuAc:Galβ(1,4)GlcNAc sialyltransferase (ST6Gal I) in colorectal cancer (CRC). Methods: ST6Gal I activity was determined in healthy, transitional and tumor tissues from the same patient using asialotransferrin and N-acetyllactosamine as acceptors. Results: ST6Gal I activities with asialotransferrin (n = 85) and N-acetyllactosamine (n = 40) as acceptors were statistically significantly enhanced in CRC tissue compared with healthy mucosa from the same patient (p = 0.001). Using transitional tissue (n = 27), enhancement versus healthy tissue was observed (p < 0.05). A positive correlation was found between ST6Gal I activity with N-acetyllactosamine and asialotransferrin in healthy (n = 32), tumorous (n = 32) and transitional tissue (n = 27), supporting the fact that the same enzyme was detected using both acceptors. Furthermore, we studied the relationship between some patients’ clinicopathological features and ST6Gal I activity. Although the differences were not statistically significant, the levels of ST6Gal I activity in tumorous and transitional tissues varied with the histological grade of the tumor; however, we failed to find a correlation with the AJCC tumor classification. Conclusions: This work reports enhanced ST6Gal I activity in tumor and transitional tissues from CRC patients. However, our overall results suggest that ST6Gal I activity is not indicative of the patient’s outcome.