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The high affinity selectin glycan ligand C2-O-sLex and mRNA transcripts of the core 2 beta-1,6-N-acetylglucosaminyltransferase (C2GnT1) gene are highly expressed in human colorectal adenocarcinomas.

St Hill CA, Farooqui M, Mitcheltree G, Gulbahce HE, Jessurun J, Cao Q, Walcheck B - BMC Cancer (2009)

Bottom Line: The most intense staining with CHO-131 mAb was observed at the advancing edge of tumors with the deepest invasive components.Finally, we analyzed C2GnT1 mRNA levels in 37 colorectal adenocarcinomas and 5 normal colorectal tissues by RT-PCR.Significantly, we observed a greater than 15-fold increase in C2GnT1 mRNA levels in colorectal adenocarcinomas compared to normal colorectal tissues.C2-O-sLex, detected by the CHO-131 mAb, is a tumor associated antigen whose expression is highly upregulated in colorectal adenocarcinomas and metastatic liver tumors compared to normal tissues.

View Article: PubMed Central - HTML - PubMed

Affiliation: Department of Veterinary Clinical Sciences, University of Minnesota, Room C339, Veterinary Medical Center, 1352 Boyd Avenue, St, Paul, MN 55108, USA. sthil001@umn.edu

ABSTRACT

Background: The metastasis of cancer cells and leukocyte extravasation into inflamed tissues share common features. Specialized carbohydrates modified with sialyl Lewis x (sLex) antigens on leukocyte membranes are ligands for selectin adhesion molecules on activated vascular endothelial cells at inflammatory sites. The activity of the enzyme core 2 beta1,6 N-acetylglucosaminyltransferase (C2GnT1) in leukocytes greatly increases their ability to bind to endothelial selectins. C2GnT1 is essential for the synthesis of core 2-branched O-linked carbohydrates terminated with sLex (C2-O-sLex). Our goal was to determine the expression profiles of C2-O-sLex in the malignant progression and metastasis of colorectal adenocarcinomas. The well characterized CHO-131 monoclonal antibody (mAb) specifically recognizes C2-O-sLex present in human leukocytes and carcinoma cells. Using CHO-131 mAb, we investigated whether C2-O-sLex was present in 113 human primary colorectal adenocarcinomas, 10 colorectal adenomas, 46 metastatic liver tumors, 28 normal colorectal tissues, and 5 normal liver tissues by immunohistochemistry. We also examined mRNA levels of the enzyme core 2 beta1,6-N-acetylglucosaminyltransferase (C2GnT1) in 20 well, 15 moderately, and 2 poorly differentiated colorectal adenocarcinomas, and in 5 normal colorectal tissues by using quantitative real-time polymerase chain reactions (RT-PCR).

Results: We observed high reactivity with CHO-131 mAb in approximately 70% of colorectal carcinomas and 87% of metastatic liver tumors but a lack of reactivity in colorectal adenomas and normal colonic and liver tissues. Positive reactivity with CHO-131 mAb was very prominent in neoplastic colorectal glands of well to moderately differentiated adenocarcinomas. The most intense staining with CHO-131 mAb was observed at the advancing edge of tumors with the deepest invasive components.Finally, we analyzed C2GnT1 mRNA levels in 37 colorectal adenocarcinomas and 5 normal colorectal tissues by RT-PCR. Significantly, we observed a greater than 15-fold increase in C2GnT1 mRNA levels in colorectal adenocarcinomas compared to normal colorectal tissues.

Conclusion: C2-O-sLex, detected by the CHO-131 mAb, is a tumor associated antigen whose expression is highly upregulated in colorectal adenocarcinomas and metastatic liver tumors compared to normal tissues. C2-O-sLex is a potentially useful early predictor of metastasis.

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Human well differentiated colorectal adenocarcinomas expressed low levels of C2-O-sLex. (A) Photomicrograph of a tissue section of a well differentiated colorectal adenocarcinoma stained with hematoxylin and eosin, scale bar = 500 μm. (B) A serial section of the same tissue as shown in (A) stained with CHO-131 mAb (15 μg/ml). Red color indicates positive reactivity with the CHO-131 mAb, scale bar = 500 μm. In (A) and (B) the arrow indicates adjacent normal colorectal mucosa. The arrowhead indicates nests of neoplastic cells in the tunica submucosa and muscularis. (C) Another well differentiated colorectal adenocarcinoma stained with CHO-131 mAb. Red color indicates positive reactivity with the CHO-131 mAb, scale bar = 100 μm. (D) Increased magnification of the boxed area shown in (C) demonstrating red stained CHO-131+ cells. Arrows in (D) indicate cytoplasmic and luminal reactivity with CHO-131 mAb, scale bar = 50 μm. (E) Mucosa of normal colorectal epithelium stained with CHO-131 mAb. Note the absence of red color, indicating a lack of reactivity with the CHO-131 mAb, scale bar = 150 μm. (F) A serial section of the same tissue as in (C) and (D), stained with CEA mAb (1.6 μg/ml). Brown color indicates positive reactivity with CEA mAb, scale bar = 100 μm. All tissue sections were 4 μm thickness. Figures (A) and (B) 20× magnification, (C) 200× magnification, (D) 400× magnification, (E) 100× magnification, and (F) 200× magnification. Mayer's hematoxylin was used as a counterstain for tissues stained with CHO-131 and CEA mAbs. Representative sections from multiple stained tissues are shown.
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Figure 2: Human well differentiated colorectal adenocarcinomas expressed low levels of C2-O-sLex. (A) Photomicrograph of a tissue section of a well differentiated colorectal adenocarcinoma stained with hematoxylin and eosin, scale bar = 500 μm. (B) A serial section of the same tissue as shown in (A) stained with CHO-131 mAb (15 μg/ml). Red color indicates positive reactivity with the CHO-131 mAb, scale bar = 500 μm. In (A) and (B) the arrow indicates adjacent normal colorectal mucosa. The arrowhead indicates nests of neoplastic cells in the tunica submucosa and muscularis. (C) Another well differentiated colorectal adenocarcinoma stained with CHO-131 mAb. Red color indicates positive reactivity with the CHO-131 mAb, scale bar = 100 μm. (D) Increased magnification of the boxed area shown in (C) demonstrating red stained CHO-131+ cells. Arrows in (D) indicate cytoplasmic and luminal reactivity with CHO-131 mAb, scale bar = 50 μm. (E) Mucosa of normal colorectal epithelium stained with CHO-131 mAb. Note the absence of red color, indicating a lack of reactivity with the CHO-131 mAb, scale bar = 150 μm. (F) A serial section of the same tissue as in (C) and (D), stained with CEA mAb (1.6 μg/ml). Brown color indicates positive reactivity with CEA mAb, scale bar = 100 μm. All tissue sections were 4 μm thickness. Figures (A) and (B) 20× magnification, (C) 200× magnification, (D) 400× magnification, (E) 100× magnification, and (F) 200× magnification. Mayer's hematoxylin was used as a counterstain for tissues stained with CHO-131 and CEA mAbs. Representative sections from multiple stained tissues are shown.

Mentions: There was a unique pattern of reactivity with CHO-131 mAb in cancer tissues. CHO-131 mAb positively stained both the cytoplasm and luminal surfaces of glandular structures in all colorectal adenocarcinomas. The intramucosal aspects of each adenocarcinoma stained less intensely with the CHO-131 mAb and the staining pattern was irregular. The deeper submucosal and muscularis areas of intestine that were infiltrated by tumor cells demonstrated a more uniform, intense staining for CHO-131 mAb and this finding was especially prominent at the advancing edge of the tumor with the deepest invasive components (Figure 2A, B). The positive reactivity with CHO-131 mAb was most prominent in neoplastic colorectal glands of well to moderately differentiated colorectal adenocarcinomas (Figure 2C, D and Figure 3A).


The high affinity selectin glycan ligand C2-O-sLex and mRNA transcripts of the core 2 beta-1,6-N-acetylglucosaminyltransferase (C2GnT1) gene are highly expressed in human colorectal adenocarcinomas.

St Hill CA, Farooqui M, Mitcheltree G, Gulbahce HE, Jessurun J, Cao Q, Walcheck B - BMC Cancer (2009)

Human well differentiated colorectal adenocarcinomas expressed low levels of C2-O-sLex. (A) Photomicrograph of a tissue section of a well differentiated colorectal adenocarcinoma stained with hematoxylin and eosin, scale bar = 500 μm. (B) A serial section of the same tissue as shown in (A) stained with CHO-131 mAb (15 μg/ml). Red color indicates positive reactivity with the CHO-131 mAb, scale bar = 500 μm. In (A) and (B) the arrow indicates adjacent normal colorectal mucosa. The arrowhead indicates nests of neoplastic cells in the tunica submucosa and muscularis. (C) Another well differentiated colorectal adenocarcinoma stained with CHO-131 mAb. Red color indicates positive reactivity with the CHO-131 mAb, scale bar = 100 μm. (D) Increased magnification of the boxed area shown in (C) demonstrating red stained CHO-131+ cells. Arrows in (D) indicate cytoplasmic and luminal reactivity with CHO-131 mAb, scale bar = 50 μm. (E) Mucosa of normal colorectal epithelium stained with CHO-131 mAb. Note the absence of red color, indicating a lack of reactivity with the CHO-131 mAb, scale bar = 150 μm. (F) A serial section of the same tissue as in (C) and (D), stained with CEA mAb (1.6 μg/ml). Brown color indicates positive reactivity with CEA mAb, scale bar = 100 μm. All tissue sections were 4 μm thickness. Figures (A) and (B) 20× magnification, (C) 200× magnification, (D) 400× magnification, (E) 100× magnification, and (F) 200× magnification. Mayer's hematoxylin was used as a counterstain for tissues stained with CHO-131 and CEA mAbs. Representative sections from multiple stained tissues are shown.
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Figure 2: Human well differentiated colorectal adenocarcinomas expressed low levels of C2-O-sLex. (A) Photomicrograph of a tissue section of a well differentiated colorectal adenocarcinoma stained with hematoxylin and eosin, scale bar = 500 μm. (B) A serial section of the same tissue as shown in (A) stained with CHO-131 mAb (15 μg/ml). Red color indicates positive reactivity with the CHO-131 mAb, scale bar = 500 μm. In (A) and (B) the arrow indicates adjacent normal colorectal mucosa. The arrowhead indicates nests of neoplastic cells in the tunica submucosa and muscularis. (C) Another well differentiated colorectal adenocarcinoma stained with CHO-131 mAb. Red color indicates positive reactivity with the CHO-131 mAb, scale bar = 100 μm. (D) Increased magnification of the boxed area shown in (C) demonstrating red stained CHO-131+ cells. Arrows in (D) indicate cytoplasmic and luminal reactivity with CHO-131 mAb, scale bar = 50 μm. (E) Mucosa of normal colorectal epithelium stained with CHO-131 mAb. Note the absence of red color, indicating a lack of reactivity with the CHO-131 mAb, scale bar = 150 μm. (F) A serial section of the same tissue as in (C) and (D), stained with CEA mAb (1.6 μg/ml). Brown color indicates positive reactivity with CEA mAb, scale bar = 100 μm. All tissue sections were 4 μm thickness. Figures (A) and (B) 20× magnification, (C) 200× magnification, (D) 400× magnification, (E) 100× magnification, and (F) 200× magnification. Mayer's hematoxylin was used as a counterstain for tissues stained with CHO-131 and CEA mAbs. Representative sections from multiple stained tissues are shown.
Mentions: There was a unique pattern of reactivity with CHO-131 mAb in cancer tissues. CHO-131 mAb positively stained both the cytoplasm and luminal surfaces of glandular structures in all colorectal adenocarcinomas. The intramucosal aspects of each adenocarcinoma stained less intensely with the CHO-131 mAb and the staining pattern was irregular. The deeper submucosal and muscularis areas of intestine that were infiltrated by tumor cells demonstrated a more uniform, intense staining for CHO-131 mAb and this finding was especially prominent at the advancing edge of the tumor with the deepest invasive components (Figure 2A, B). The positive reactivity with CHO-131 mAb was most prominent in neoplastic colorectal glands of well to moderately differentiated colorectal adenocarcinomas (Figure 2C, D and Figure 3A).

Bottom Line: The most intense staining with CHO-131 mAb was observed at the advancing edge of tumors with the deepest invasive components.Finally, we analyzed C2GnT1 mRNA levels in 37 colorectal adenocarcinomas and 5 normal colorectal tissues by RT-PCR.Significantly, we observed a greater than 15-fold increase in C2GnT1 mRNA levels in colorectal adenocarcinomas compared to normal colorectal tissues.C2-O-sLex, detected by the CHO-131 mAb, is a tumor associated antigen whose expression is highly upregulated in colorectal adenocarcinomas and metastatic liver tumors compared to normal tissues.

View Article: PubMed Central - HTML - PubMed

Affiliation: Department of Veterinary Clinical Sciences, University of Minnesota, Room C339, Veterinary Medical Center, 1352 Boyd Avenue, St, Paul, MN 55108, USA. sthil001@umn.edu

ABSTRACT

Background: The metastasis of cancer cells and leukocyte extravasation into inflamed tissues share common features. Specialized carbohydrates modified with sialyl Lewis x (sLex) antigens on leukocyte membranes are ligands for selectin adhesion molecules on activated vascular endothelial cells at inflammatory sites. The activity of the enzyme core 2 beta1,6 N-acetylglucosaminyltransferase (C2GnT1) in leukocytes greatly increases their ability to bind to endothelial selectins. C2GnT1 is essential for the synthesis of core 2-branched O-linked carbohydrates terminated with sLex (C2-O-sLex). Our goal was to determine the expression profiles of C2-O-sLex in the malignant progression and metastasis of colorectal adenocarcinomas. The well characterized CHO-131 monoclonal antibody (mAb) specifically recognizes C2-O-sLex present in human leukocytes and carcinoma cells. Using CHO-131 mAb, we investigated whether C2-O-sLex was present in 113 human primary colorectal adenocarcinomas, 10 colorectal adenomas, 46 metastatic liver tumors, 28 normal colorectal tissues, and 5 normal liver tissues by immunohistochemistry. We also examined mRNA levels of the enzyme core 2 beta1,6-N-acetylglucosaminyltransferase (C2GnT1) in 20 well, 15 moderately, and 2 poorly differentiated colorectal adenocarcinomas, and in 5 normal colorectal tissues by using quantitative real-time polymerase chain reactions (RT-PCR).

Results: We observed high reactivity with CHO-131 mAb in approximately 70% of colorectal carcinomas and 87% of metastatic liver tumors but a lack of reactivity in colorectal adenomas and normal colonic and liver tissues. Positive reactivity with CHO-131 mAb was very prominent in neoplastic colorectal glands of well to moderately differentiated adenocarcinomas. The most intense staining with CHO-131 mAb was observed at the advancing edge of tumors with the deepest invasive components.Finally, we analyzed C2GnT1 mRNA levels in 37 colorectal adenocarcinomas and 5 normal colorectal tissues by RT-PCR. Significantly, we observed a greater than 15-fold increase in C2GnT1 mRNA levels in colorectal adenocarcinomas compared to normal colorectal tissues.

Conclusion: C2-O-sLex, detected by the CHO-131 mAb, is a tumor associated antigen whose expression is highly upregulated in colorectal adenocarcinomas and metastatic liver tumors compared to normal tissues. C2-O-sLex is a potentially useful early predictor of metastasis.

Show MeSH
Related in: MedlinePlus