Dendritic Cell-specific Intercellular Adhesion Molecule 3-grabbing Non-integrin (DC-SIGN) Recognizes a Novel Ligand, Mac-2-binding Protein, Characteristically Expressed on Human Colorectal Carcinomas

Abstract

Dendritic cell (DC)-specific intercellular adhesion molecule-3-grabbing non-integrin (DC-SIGN) is a type II transmembrane C-type lectin expressed on DCs such as myeloid DCs and monocyte-derived DCs (MoDCs). Recently, we have reported that DC-SIGN interacts with carcinoembryonic antigen (CEA) expressed on colorectal carcinoma cells. CEA is one of the most widely used tumor markers for gastrointestinal cancers such as colorectal cancer. On the other hand, other groups have reported that the level of Mac-2-binding protein (Mac-2BP) increases in patients with pancreatic, breast, and lung cancers, virus infections such as human immunodeficiency virus and hepatitis C virus, and autoimmune diseases. Here, we first identified Mac-2BP expressed on several colorectal carcinoma cell lines as a novel DC-SIGN ligand through affinity chromatography and mass spectrometry. Interestingly, we found that DC-SIGN selectively recognizes Mac-2BP derived from some colorectal carcinomas but not from the other ones. Furthermore, we found that the α1-3,4-fucose moieties of Le glycans expressed on DC-SIGN-binding Mac-2BP were important for recognition. DC-SIGN-dependent cellular interactions between immature MoDCs and colorectal carcinoma cells significantly inhibited MoDC functional maturation, suggesting that Mac-2BP may provide a tolerogenic microenvironment for colorectal carcinoma cells through DC-SIGN-dependent recognition. Importantly, Mac-2BP was detected as a predominant DC-SIGN ligand expressed on some primary colorectal cancer tissues from certain parts of patients in comparison with CEA from other parts, suggesting that DC-SIGN-binding Mac-2BP bearing tumor-associated Le glycans may become a novel potential colorectal cancer biomarker for some patients instead of CEA. Dendritic cell (DC)-specific intercellular adhesion molecule-3-grabbing non-integrin (DC-SIGN) is a type II transmembrane C-type lectin expressed on DCs such as myeloid DCs and monocyte-derived DCs (MoDCs). Recently, we have reported that DC-SIGN interacts with carcinoembryonic antigen (CEA) expressed on colorectal carcinoma cells. CEA is one of the most widely used tumor markers for gastrointestinal cancers such as colorectal cancer. On the other hand, other groups have reported that the level of Mac-2-binding protein (Mac-2BP) increases in patients with pancreatic, breast, and lung cancers, virus infections such as human immunodeficiency virus and hepatitis C virus, and autoimmune diseases. Here, we first identified Mac-2BP expressed on several colorectal carcinoma cell lines as a novel DC-SIGN ligand through affinity chromatography and mass spectrometry. Interestingly, we found that DC-SIGN selectively recognizes Mac-2BP derived from some colorectal carcinomas but not from the other ones. Furthermore, we found that the α1-3,4-fucose moieties of Le glycans expressed on DC-SIGN-binding Mac-2BP were important for recognition. DC-SIGN-dependent cellular interactions between immature MoDCs and colorectal carcinoma cells significantly inhibited MoDC functional maturation, suggesting that Mac-2BP may provide a tolerogenic microenvironment for colorectal carcinoma cells through DC-SIGN-dependent recognition. Importantly, Mac-2BP was detected as a predominant DC-SIGN ligand expressed on some primary colorectal cancer tissues from certain parts of patients in comparison with CEA from other parts, suggesting that DC-SIGN-binding Mac-2BP bearing tumor-associated Le glycans may become a novel potential colorectal cancer biomarker for some patients instead of CEA. IntroductionDendritic cells (DCs) 4The abbreviations used are: DCdendritic cellDC-SIGNDC-specific intercellular adhesion molecule-3-grabbing non-integrinrhDC-SIGN-Fcrecombinant human DC-SIGN-human IgG-Fc fusion proteinpAbpolyclonal AbMac-2BPMac-2-binding proteinCEAcarcinoembryonic antigenMoDCmonocyte-derived DCLeLewisPHAphytohaemagglutininAALAleuria aurantia agglutininPNGasepeptide N-glycosidaseM-CSFmacrophage colony-stimulating factor. play a critical role in initiating adaptive immunity by uptaking foreign antigens at the periphery, undergoing maturation during migration to lymph nodes and presenting antigen-derived peptides to naïve T cells (1Banchereau J. Briere F. Caux C. Davoust J. Lebecque S. Liu Y.J. Pulendran B. Palucka K. Annu. Rev. Immunol. 2000; 18: 767-811Crossref PubMed Scopus (5568) Google Scholar, 2Lanzavecchia A. Sallusto F. Cell. 2001; 106: 263-266Abstract Full Text Full Text PDF PubMed Scopus (830) Google Scholar). For effective recognition of antigens, DCs exploit a wide range of pattern recognition receptors such as Toll-like receptors and C-type lectins (3Geijtenbeek T.B. van Vliet S.J. Engering A. ‘t Hart B.A. van Kooyk Y. Annu. Rev. Immunol. 2004; 22: 33-54Crossref PubMed Scopus (413) Google Scholar, 4van Vliet S.J. den Dunnen J. Gringhuis S.I. Geijtenbeek T.B. van Kooyk Y. Curr. Opin. Immunol. 2007; 19: 435-440Crossref PubMed Scopus (134) Google Scholar, 5van Kooyk Y. Rabinovich G.A. Nat. Immunol. 2008; 9: 593-601Crossref PubMed Scopus (544) Google Scholar). DC-specific intercellular adhesion molecule-3-grabbing non-integrin (DC-SIGN; CD209) is a type II transmembrane C-type lectin expressed on myeloid DCs in the dermis, mucosae, lymph nodes, and monocyte-derived DCs (MoDCs) (6Geijtenbeek T.B. Kwon D.S. Torensma R. van Vliet S.J. van Duijnhoven G.C. Middel J. Cornelissen I.L. Nottet H.S. KewalRamani V.N. Littman D.R. Figdor C.G. van Kooyk Y. Cell. 2000; 100: 587-597Abstract Full Text Full Text PDF PubMed Scopus (2026) Google Scholar, 7Geijtenbeek T.B. Torensma R. van Vliet S.J. van Duijnhoven G.C. Adema G.J. van Kooyk Y. Figdor C.G. Cell. 2000; 100: 575-585Abstract Full Text Full Text PDF PubMed Scopus (1443) Google Scholar). DC-SIGN binds to “self” glycan ligands found on human cells and to “foreign” glycans of bacterial or parasitic pathogens, and specifically recognizes glycoconjugates containing mannose (Man), fucose (Fuc), and nonsialylated Lewis (Le)a/Leb epitope structures in a Ca2+-dependent manner (8van Liempt E. Bank C.M. Mehta P. Garciá-Vallejo J.J. Kawar Z.S. Geyer R. Alvarez R.A. Cummings R.D. Kooyk Y. van Die I. FEBS Lett. 2006; 580: 6123-6131Crossref PubMed Scopus (202) Google Scholar). Through recognition of these glycans, DC-SIGN functions as an adhesion receptor and mediates the binding and internalization of foreign pathogens such as viruses (human immunodeficiency virus and hepatitis C virus), yeasts, bacteria (Mycobacterium tuberculosis), and parasites (3Geijtenbeek T.B. van Vliet S.J. Engering A. ‘t Hart B.A. van Kooyk Y. Annu. Rev. Immunol. 2004; 22: 33-54Crossref PubMed Scopus (413) Google Scholar).Recently, we and another group (9van Gisbergen K.P. Aarnoudse C.A. Meijer G.A. Geijtenbeek T.B. van Kooyk Y. Cancer Res. 2005; 65: 5935-5944Crossref PubMed Scopus (135) Google Scholar, 10Nonaka M. Ma B.Y. Murai R. Nakamura N. Baba M. Kawasaki N. Hodohara K. Asano S. Kawasaki T. J. Immunol. 2008; 180: 3347-3356Crossref PubMed Scopus (66) Google Scholar) found that DC-SIGN recognizes colorectal carcinoma cells through carcinoembryonic antigen (CEA). DC-SIGN-binding CEA contained tumor-associated Le glycans, which were important for cellular interactions between DCs and colorectal carcinoma cells in situ. It is evident that the expression of CEA is limited and can be detected only in cancer and embryonic tissues. This glycoprotein is one of the most widely used tumor markers for gastrointestinal cancers such as colorectal carcinomas. However, its sensitivity and specificity are not high, particularly for early stages of the disease such as Dukes A or B stages. Based on our studies, we hypothesized that DC-SIGN may also be involved in the recognition of other glycoproteins, as novel potential biomarkers, expressed on colorectal cancers.In this study, we first identified Mac-2-binding protein (Mac-2BP) expressed on colorectal carcinoma cells as a novel DC-SIGN ligand through affinity chromatography and mass spectrometry. Mac-2BP, also known as 90K or galectin-3BP, is a secretory glycoprotein expressed on various normal epithelial cells and in human bodily fluids. It has been reported that the level of Mac-2BP increases in patients with pancreatic, breast, and lung cancer, virus infections such as acquired immunodeficiency syndrome and hepatitis C, and autoimmune diseases (11Grassadonia A. Tinari N. Iurisci I. Piccolo E. Cumashi A. Innominato P. D'Egidio M. Natoli C. Piantelli M. Iacobelli S. Glycoconj. J. 2004; 19: 551-556Crossref PubMed Scopus (143) Google Scholar). However, most of the immunological significance of Mac-2BP on colorectal carcinomas remains unknown.In addition, we found that DC-SIGN-dependent cellular interactions between immature MoDCs and colorectal carcinoma cells mainly expressing Mac-2BP were able to significantly inhibit LPS-induced MoDC functional maturation. Furthermore, we also detected Mac-2BP as a predominant DC-SIGN ligand expressed on some primary colorectal cancer tissues from certain parts of patients in comparison with CEA from the other parts. Therefore, we suggest that Mac-2BP carrying fucosylated glycans may become a novel potential colorectal cancer biomarker for some CEA-low or -negative colorectal cancer patients, and the novel function of DC-SIGN may, at least in part, underlie for its potential use as a novel diagnostic sensor for some colorectal cancer patients through recognition of CEA and Mac-2BP.DISCUSSIONDuring the neoplastic process in colon tissues, genetic or epigenetic gene alteration often accompanies changes in cell-surface glycans. Glycosylation changes during malignant transformation lead to tumor-specific carbohydrate structures that interact with C-type lectins on dendritic cells. In general, these changes have been associated with a poor prognosis, as they are linked to the aggressiveness and metastatic capacity of tumors. Good examples of heavily glycosylated tumor antigens are CEA and CEACAM1 (13Midiri G. Amanti C. Benedetti M. Campisi C. Santeusanio G. Castagna G. Peronace L. Di Tondo U. Di Paola M. Pascal R.R. Cancer. 1985; 55: 2624-2629Crossref PubMed Scopus (34) Google Scholar, 14Hamada Y. Yamamura M. Hioki K. Yamamoto M. Nagura H. Watanabe K. Cancer. 1985; 55: 136-141Crossref PubMed Scopus (139) Google Scholar, 15Shi Z.R. Tsao D. Kim Y.S. Cancer Res. 1983; 43: 4045-4049PubMed Google Scholar, 16Ahnen D.J. Nakane P.K. Brown W.R. Cancer. 1982; 49: 2077-2090Crossref PubMed Scopus (155) Google Scholar, 17Nittka S. Böhm C. Zentgraf H. Neumaier M. Oncogene. 2008; 27: 3721-3728Crossref PubMed Scopus (27) Google Scholar, 18Shively J.E. Oncogene. 2004; 23: 9303-9305Crossref PubMed Scopus (9) Google Scholar, 19Nittka S. Günther J. Ebisch C. Erbersdobler A. Neumaier M. Oncogene. 2004; 23: 9306-9313Crossref PubMed Scopus (69) Google Scholar), which can be secreted or expressed by colon cancer cells. Because the function of dendritic cells may be dependent on their binding properties as to self-antigens and pathogens, it is essential to obtain a detailed insight into the carbohydrate-binding properties of DC-SIGN.Previous research indicated that tumor cells with their surface decorated with β1-6-branched N-linked oligosaccharides acquire invasiveness and metastatic potential through a change in the affinity level to the extracellular microenvironment (20Seelentag W.K. Li W.P. Schmitz S.F. Metzger U. Aeberhard P. Heitz P.U. Roth J. Cancer Res. 1998; 58: 5559-5564PubMed Google Scholar, 21Dennis J.W. Laferté S. Waghorne C. Breitman M.L. Kerbel R.S. Science. 1987; 236: 582-585Crossref PubMed Scopus (851) Google Scholar). Mac-2BP expressed on colorectal carcinoma cells was initially identified as a l-PHA-recognized major glycoprotein bearing polylactosamine glycans extending from β1-6 branched N-linked oligosaccharides (22Laferté S. Loh L.C. Biochem. J. 1992; 283: 193-201Crossref PubMed Scopus (31) Google Scholar, 23Fernandes B. Sagman U. Auger M. Demetrio M. Dennis J.W. Cancer Res. 1991; 51: 718-723PubMed Google Scholar). In this study, we have firstly identified Mac-2BP on colorectal carcinoma cells as a novel ligand for C-type lectin DC-SIGN and demonstrated the Ca2+-dependent interaction of rhDC-SIGN with Mac-2BP from colorectal carcinoma COLO205 cells. Generally, DC-SIGN exhibits dual specificities for high-mannose as well as Le-type glycans (8van Liempt E. Bank C.M. Mehta P. Garciá-Vallejo J.J. Kawar Z.S. Geyer R. Alvarez R.A. Cummings R.D. Kooyk Y. van Die I. FEBS Lett. 2006; 580: 6123-6131Crossref PubMed Scopus (202) Google Scholar, 24Appelmelk B.J. van Die I. van Vliet S.J. Vandenbroucke-Grauls C.M. Geijtenbeek T.B. van Kooyk Y. J. Immunol. 2003; 170: 1635-1639Crossref PubMed Scopus (378) Google Scholar, 25Guo Y. Feinberg H. Conroy E. Mitchell D.A. Alvarez R. Blixt O. Taylor M.E. Weis W.I. Drickamer K. Nat. Struct. Mol. Biol. 2004; 11: 591-598Crossref PubMed Scopus (479) Google Scholar). Foreign antigens recognized by DC-SIGN have both types of glycans. Whereas, so far, most endogenous ligands for DC-SIGN have been found to carry only Le-type glycans. Here we showed that specific removal of Le glycans on Mac-2BP by α1-3,4-fucosidase abrogated these interactions, indicating that the fucose moieties of Le glycans are required for the interactions. We also observed that DC-SIGN ligands are highly expressed with Le glycans on human colorectal carcinoma cells, suggesting that increased expression of DC-SIGN ligands on colorectal cancer cells in addition to altered glycosylation results in recognition by DC-SIGN and that further fucosylation of polylactosamine glycans and formation of Le epitopes on Mac-2BP during cancer progression may be important for induction of the interaction between DC-SIGN and Mac-2BP. In addition, we also found that the expression levels of Le epitopes were positively correlated with the staining levels with rhDC-SIGN in 30 colorectal carcinoma cell lines, including LoVo, LS513, LS174T, SNU1197, etc. (data not shown).CEA is one of the most widely used tumor markers for gastrointestinal cancer such as colorectal carcinomas. It has been pointed out that the change in distribution of CEA from apical into basolateral and stromal areas in malignant colorectal cancer tissues could be a cause of its entry into the blood and elevation of the serum CEA levels (14Hamada Y. Yamamura M. Hioki K. Yamamoto M. Nagura H. Watanabe K. Cancer. 1985; 55: 136-141Crossref PubMed Scopus (139) Google Scholar, 16Ahnen D.J. Nakane P.K. Brown W.R. Cancer. 1982; 49: 2077-2090Crossref PubMed Scopus (155) Google Scholar). However, the sensitivity and specificity of CEA are considered to be low, particularly for early stages of the disease such as Dukes A or B stages (26Duffy M.J. Clin. Chem. 2001; 47: 624-630Crossref PubMed Scopus (546) Google Scholar). We showed here that the distribution of Mac-2BP was more basolateral than that of CEA and that expression of Mac-2BP in primary colon adenocarcinoma tissues was increased compared with in proximal normal tissues and obtained various staining patterns for Mac-2BP such as luminal staining and punctate staining on the apical and basolateral faces (Fig. 7B). In addition, we observed here that DC-SIGN greatly associated with not only CEA but also Mac-2BP on the apical face of cancer epithelia and that DC-SIGN retained responsiveness to Mac-2BP in more basolateral areas. Because basolateral expression of Mac-2BP has been reported to be more prevalent in early stage Dukes A tumors than in advanced ones (12Ulmer T.A. Keeler V. Loh L. Chibbar R. Torlakovic E. André S. Gabius H.J. Laferté S. J. Cell Biochem. 2006; 98: 1351-1366Crossref PubMed Scopus (45) Google Scholar), DC-SIGN may preferably recognize early stage colorectal carcinoma cells through Mac-2BP. Therefore, similarly to that of CEA, aberrant expression of Mac-2BP and its polar breakdown in cancer tissues may allow its entry into blood vessels. Furthermore, it is also interesting that the type of Mac-2BP secreted by several colorectal carcinoma cells had DC-SIGN-detectable glycan structures, whereas we did not detect any binding of DC-SIGN to serum Mac-2BP from healthy donors (data not shown), suggesting that DC-SIGN can qualitatively distinguish colorectal carcinoma-derived Mac-2BP from serum Mac-2BP in normal and cancer patients.Recent evidence indicates that dendritic cell-associated DC-SIGN may have functions in both the induction of tolerance to self-antigens and the recognition of pathogens, respectively. A variety of cytokines, including IL-6, IL-10, and macrophage colony-stimulating factor (M-CSF), have been shown to affect the maturation of DCs from CD34+ precursors and from MoDCs in vitro (27Menetrier-Caux C. Montmain G. Dieu M.C. Bain C. Favrot M.C. Caux C. Blay J.Y. Blood. 1998; 92: 4778-4791Crossref PubMed Google Scholar, 28Kiertscher S.M. Luo J. Dubinett S.M. Roth M.D. J. Immunol. 2000; 164: 1269-1276Crossref PubMed Scopus (213) Google Scholar). Failure of the immune system to provide protection against tumor cells is an important immunological problem. It is now evident that inadequate functioning of the host immune system is one of the main mechanisms by which tumors escape from immune control, as well as an important factor that limits the success of cancer immunotherapy. In recent years, it has become increasingly clear that defects in DCs play a crucial role in non-responsiveness to tumors D. Nat. Rev. Immunol. 2004; PubMed Scopus Google Scholar). Here, we showed that DC-SIGN-dependent cellular interactions between immature MoDCs and colorectal carcinoma cells significantly inhibited MoDC functional maturation, suggesting to CEA, Mac-2BP bearing colorectal tumor-associated Le glycans may provide a tolerogenic microenvironment for colorectal carcinoma cells through interactions with also that the cellular interactions may in of through such as and IL-10, and that of these in tumor may be one of the mechanisms by which tumors the mechanisms of DC-SIGN were S.I. den Dunnen J. M. van B. van Kooyk Y. Geijtenbeek T.B. 2007; Full Text Full Text PDF PubMed Scopus Google Scholar, S.I. van M. van den den Dunnen J. M. Geijtenbeek T.B. Nat. Immunol. 11: PubMed Scopus Google Scholar, S.I. den Dunnen J. M. van M. Geijtenbeek T.B. Nat. Immunol. PubMed Scopus Google Scholar). DC-SIGN with a of protein and mediates with Toll-like receptor which results in of Toll-like interaction of foreign pathogens such as M. to of LPS-induced and MoDC maturation T.B. Vliet S.J. M. Vandenbroucke-Grauls C.M. B. Kooyk Y. J. 2003; PubMed Scopus Google Scholar). Therefore, the MoDC maturation as a of DC-SIGN with Le glycans of cancer cells may also be through a of with peptides are now as a for the of tumor the of patients immune which is by of human antigen of antigens is to obtain In this colorectal carcinoma-derived Mac-2BP, to CEA, become a cancer of such as MoDC by of that may be a critical for the effective in the on the expression patterns of Mac-2BP and CEA with the DC-SIGN recognition colorectal cancer patients could be into and of a of the interaction is a of the function and and a for the of cancer and novel For this of the that to and cancer both in vitro and in is a major for in the IntroductionDendritic cells (DCs) 4The abbreviations used are: DCdendritic cellDC-SIGNDC-specific intercellular adhesion molecule-3-grabbing non-integrinrhDC-SIGN-Fcrecombinant human DC-SIGN-human IgG-Fc fusion proteinpAbpolyclonal AbMac-2BPMac-2-binding proteinCEAcarcinoembryonic antigenMoDCmonocyte-derived DCLeLewisPHAphytohaemagglutininAALAleuria aurantia agglutininPNGasepeptide N-glycosidaseM-CSFmacrophage colony-stimulating factor. play a critical role in initiating adaptive immunity by uptaking foreign antigens at the periphery, undergoing maturation during migration to lymph nodes and presenting antigen-derived peptides to naïve T cells (1Banchereau J. Briere F. Caux C. Davoust J. Lebecque S. Liu Y.J. Pulendran B. Palucka K. Annu. Rev. Immunol. 2000; 18: 767-811Crossref PubMed Scopus (5568) Google Scholar, 2Lanzavecchia A. Sallusto F. Cell. 2001; 106: 263-266Abstract Full Text Full Text PDF PubMed Scopus (830) Google Scholar). For effective recognition of antigens, DCs exploit a wide range of pattern recognition receptors such as Toll-like receptors and C-type lectins (3Geijtenbeek T.B. van Vliet S.J. Engering A. ‘t Hart B.A. van Kooyk Y. Annu. Rev. Immunol. 2004; 22: 33-54Crossref PubMed Scopus (413) Google Scholar, 4van Vliet S.J. den Dunnen J. Gringhuis S.I. Geijtenbeek T.B. van Kooyk Y. Curr. Opin. Immunol. 2007; 19: 435-440Crossref PubMed Scopus (134) Google Scholar, 5van Kooyk Y. Rabinovich G.A. Nat. Immunol. 2008; 9: 593-601Crossref PubMed Scopus (544) Google Scholar). DC-specific intercellular adhesion molecule-3-grabbing non-integrin (DC-SIGN; CD209) is a type II transmembrane C-type lectin expressed on myeloid DCs in the dermis, mucosae, lymph nodes, and monocyte-derived DCs (MoDCs) (6Geijtenbeek T.B. Kwon D.S. Torensma R. van Vliet S.J. van Duijnhoven G.C. Middel J. Cornelissen I.L. Nottet H.S. KewalRamani V.N. Littman D.R. Figdor C.G. van Kooyk Y. Cell. 2000; 100: 587-597Abstract Full Text Full Text PDF PubMed Scopus (2026) Google Scholar, 7Geijtenbeek T.B. Torensma R. van Vliet S.J. van Duijnhoven G.C. Adema G.J. van Kooyk Y. Figdor C.G. Cell. 2000; 100: 575-585Abstract Full Text Full Text PDF PubMed Scopus (1443) Google Scholar). DC-SIGN binds to “self” glycan ligands found on human cells and to “foreign” glycans of bacterial or parasitic pathogens, and specifically recognizes glycoconjugates containing mannose (Man), fucose (Fuc), and nonsialylated Lewis (Le)a/Leb epitope structures in a Ca2+-dependent manner (8van Liempt E. Bank C.M. Mehta P. Garciá-Vallejo J.J. Kawar Z.S. Geyer R. Alvarez R.A. Cummings R.D. Kooyk Y. van Die I. FEBS Lett. 2006; 580: 6123-6131Crossref PubMed Scopus (202) Google Scholar). Through recognition of these glycans, DC-SIGN functions as an adhesion receptor and mediates the binding and internalization of foreign pathogens such as viruses (human immunodeficiency virus and hepatitis C virus), yeasts, bacteria (Mycobacterium tuberculosis), and parasites (3Geijtenbeek T.B. van Vliet S.J. Engering A. ‘t Hart B.A. van Kooyk Y. Annu. Rev. Immunol. 2004; 22: 33-54Crossref PubMed Scopus (413) Google Scholar).Recently, we and another group (9van Gisbergen K.P. Aarnoudse C.A. Meijer G.A. Geijtenbeek T.B. van Kooyk Y. Cancer Res. 2005; 65: 5935-5944Crossref PubMed Scopus (135) Google Scholar, 10Nonaka M. Ma B.Y. Murai R. Nakamura N. Baba M. Kawasaki N. Hodohara K. Asano S. Kawasaki T. J. Immunol. 2008; 180: 3347-3356Crossref PubMed Scopus (66) Google Scholar) found that DC-SIGN recognizes colorectal carcinoma cells through carcinoembryonic antigen (CEA). DC-SIGN-binding CEA contained tumor-associated Le glycans, which were important for cellular interactions between DCs and colorectal carcinoma cells in situ. It is evident that the expression of CEA is limited and can be detected only in cancer and embryonic tissues. This glycoprotein is one of the most widely used tumor markers for gastrointestinal cancers such as colorectal carcinomas. However, its sensitivity and specificity are not high, particularly for early stages of the disease such as Dukes A or B stages. Based on our studies, we hypothesized that DC-SIGN may also be involved in the recognition of other glycoproteins, as novel potential biomarkers, expressed on colorectal cancers.In this study, we first identified Mac-2-binding protein (Mac-2BP) expressed on colorectal carcinoma cells as a novel DC-SIGN ligand through affinity chromatography and mass spectrometry. Mac-2BP, also known as 90K or galectin-3BP, is a secretory glycoprotein expressed on various normal epithelial cells and in human bodily fluids. It has been reported that the level of Mac-2BP increases in patients with pancreatic, breast, and lung cancer, virus infections such as acquired immunodeficiency syndrome and hepatitis C, and autoimmune diseases (11Grassadonia A. Tinari N. Iurisci I. Piccolo E. Cumashi A. Innominato P. D'Egidio M. Natoli C. Piantelli M. Iacobelli S. Glycoconj. J. 2004; 19: 551-556Crossref PubMed Scopus (143) Google Scholar). However, most of the immunological significance of Mac-2BP on colorectal carcinomas remains unknown.In addition, we found that DC-SIGN-dependent cellular interactions between immature MoDCs and colorectal carcinoma cells mainly expressing Mac-2BP were able to significantly inhibit LPS-induced MoDC functional maturation. Furthermore, we also detected Mac-2BP as a predominant DC-SIGN ligand expressed on some primary colorectal cancer tissues from certain parts of patients in comparison with CEA from the other parts. Therefore, we suggest that Mac-2BP carrying fucosylated glycans may become a novel potential colorectal cancer biomarker for some CEA-low or -negative colorectal cancer patients, and the novel function of DC-SIGN may, at least in part, underlie for its potential use as a novel diagnostic sensor for some colorectal cancer patients through recognition of CEA and Mac-2BP.