The role of galectins in colorectal cancer progression


  • Hannah Barrow,

    1. Department of Gastroenterology, Institute of Translational Medicine, University of Liverpool, Liverpool, United Kingdom
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  • Jonathan M. Rhodes,

    1. Department of Gastroenterology, Institute of Translational Medicine, University of Liverpool, Liverpool, United Kingdom
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  • Lu-Gang Yu

    Corresponding author
    1. Department of Gastroenterology, Institute of Translational Medicine, University of Liverpool, Liverpool, United Kingdom
    • Department of Gastroenterology, The Henry Wellcome Laboratory, Institute of Translational Medicine, University of Liverpool, Liverpool L69 3GE, United Kingdom
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    • Tel: +44-151-7946820, Fax: +44-151-7946825


Galectins constitute a family of 15 mammalian galactoside-binding proteins that share a consensus amino acid sequence in their carbohydrate binding sites. They are multi-functional molecules and are expressed widely in human tissues. Four galectins: galectin −1, −3, −4 and −8 are expressed in the human colon and rectum and their expressions show significant changes during colorectal cancer development and metastasis. These changes in galectin expression correlate with alterations in cancer cell growth, apoptosis, cell–cell and cell–matrix interactions and angiogenesis. This review summaries current knowledge of the expression and roles of these galectins in the progression of human colorectal cancer and discusses the relevance of galectins and their ligands as potential therapeutic targets for cancer treatment.

The galectins are a family of 15 mammalian galactoside-binding proteins that share a consensus amino acid sequence in their carbohydrate recognition domains (CRDs).1 On the basis of their structural differences, members of the galectin family are classified into three subgroups: the prototype, chimera-type and tandem-repeat type. The prototype galectins include galectin-1, −2, −5, −7, −10, −11, −13, −14 and −15. They contain a single CRD in their polypeptide sequences and are often identified as non-covalently linked homodimers. The chimera-type galectin, which has only one member (galectin-3), is composed of a non-lectin domain linked to a CRD and can precipitate as a pentamer. The tandem-repeat-type galectins include galectin-4, −6, −8, −9 and −12 and members of this subgroup of galectins contain two CRDs in a single polypeptide chain and these two CRDs are often different from each other. All galectins are bivalent or multivalent molecules in physiological or pathological conditions as a result either of the presence of two CRDs within a single polypeptide chain or by polymerization, and are able to form arrays and lattice structures on the cell surface following binding to their carbohydrate ligands.2

Expression of Galectins in Normal and Cancerous Human Colon and Rectum

In the normal human colon and rectum, four galectins, galectin-1, −3, −4 and −8, are expressed3–6 (Table 1).

Table 1. The expression of galectins in the human intestine and rectum
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Galectin-1 is expressed weakly in normal colonic epithelium but its expression is increased in inflammation and cancer. Over expression of galectin-1 correlates with increased metastasis of colorectal cancer.12 Immunohistochemistry reveals galectin-1 expression in 12% of normal colonic mucosa, 40% of adenomas and 84% of carcinomas.4 Galectin-1 expression is not confined to the epithelium and 91% of adenocarcinomas and 33% of mucinous adenocarcinomas also show strong stromal galectin-1 expression.4

Galectin-3, one of the most extensively studied galectins, is widely expressed in the human gastrointestinal tract including the colon and rectum. Normal colonic mucosa shows strong nuclear expression of galectin-3. The cells at the base of the crypt have a weak or negative galectin-3 expression, and the intensity of nuclear galectin-3 expression increases progressively from the base towards the surface of the gland.4 In the nucleus, galectin-3 is localized in the dense fibrillar component of the nucleolus, as well as the periphery of the fibrillar centres.13 Galectin-3 has also been demonstrated in the interchromatic spaces and along the borders of condensed chromatin of the nucleoplasm,13 where mRNA synthesis14 and the early events in pre-mRNA splicing15 take place. Although one study has shown a decreased galectin-3 expression in colorectal cancer,4 many other studies have demonstrated an increased galectin-3 expression.9, 16–18 Galectin-3 expression is greater in advanced cancer17 and metastases express higher levels of galectin-3 than the primary tumors from which they arise.19 There is a general change in galectin-3 subcellular localization from nucleus to the cytoplasm in colorectal cancer during progression from colorectal adenoma to carcinoma.4, 20 As cytoplasmic galectin-3 is known to be an apoptosis inhibitor (see below), it is very likely that this change in localization may contribute to cancer cell survival.

Galectin-4 is hydrophobic due to the high content of apolar amino acids in its linker sequence (residues 151–175). Galectin-4 is expressed in the human intestinal and colonic mucosa5 and its expression is generally lower in cancer than in normal mucosa.5, 11, 21 However, cancers that express high levels of galectin-4 tend to have a poor prognosis.11 Galectin-4 has a tendency to be associated with generally insoluble complex as a component of either adherent junction or lipid rafts in the microvillus membrane where it stabilizes these structures.5, 22 There is a progressive reduction in dense supra-nuclear galectin-4 aggregates and a subsequent increase in diffuse cytosolic galectin-4 throughout the progression of colorectal malignancy.5 Galectin-4 is strongly expressed in highly differentiated cell lines, such as the human colon adenocarcinoma T84 where it represents 38–60% of the total galectin content.22 In T84 cells, whereas galectin-3 is seen in subapical regions, galectin-4 is observed in a thick layer near the basal membrane. Following Ca2+ depletion, galectin-4 accumulates at break sites in T84 cell monolayers,22 indicating a possible role in initial cellular reattachment and cell spreading within a disrupted monolayer.

Galectin-8 is expressed widely in the gastrointestinal tract. Low basal levels of galectin-8 are observed in the human intestine.23, 24 In healthy colon tissue, galectin-8 can be detected both in the nuclei and cytoplasm whereas in colon cancer galectin-8 is found almost exclusively in the cytoplasm.6 Overall, there is reduced expression of galectin-8 in cancer that correlates with poor prognosis.6


CRD: galectin carbohydrate recognition domain; ECM: extracellular matrix; MCP: modified citrus pectin; TF antigen: Thomsen-Friedenreich (Galactoseβ1,3N-acetylgalactosamineα-) antigen

Contrasting Effects of Galectins on Apoptosis

Galectin-1 is an apoptosis promoter. Thus, over expression of galectin-1 in colon cancer Colo201 cells is associated with increased cell apoptosis.25 Interaction between galectin-1, expressed on the surface of tumor cells26 or extracellular matrix,27 and T cells induces T cell apoptosis.28 Galectin-1 binds to the N- or O-linked glycans on several molecules on the T cell surface including CD7, CD43, CD45 and CD95/Fas29, 30 (Table 2). Binding of galectin-1 to T cells can increase the expression of cell surface CD95/Fas,31 which initiates the activation of the extrinsic apoptosis pathway initiator caspase-8 and subsequently caspase-3, either directly or indirectly via the mitochondria-associated cytochrome c release (Fig. 1a). Galectin-1 cell surface binding can also induce CD43 cell surface clustering and initiation of apoptosis through cytochrome c release.32 Galectin-1-mediated activation of the transcription factor AP-1 has been shown to be involved in galectin-1-mediated T cell apoptosis.33 Binding of galectin-1 to CD43 on MOLT-4 or CEM T cell lines has been reported to induce T cell death in a caspase-independent mechanism.34 In this situation, galectin-1 induces rapid translocation of endonuclease G from mitochondria to nuclei and cell death without the involvement of cytochrome c or caspase activation. It is noted that the cells used in these studies have varied (e.g., some used freshly isolated T cells whereas others used T cell lines). These results indicate that galectin-1can induce T cell apoptosis by several signaling pathways dependent on its binding to specific cell surface ligands. As T cell-mediated immune reactivity is critical in immune surveillance, galectin-1-mediated T-cell apoptosis probably makes an important contribution to the successful escape of tumor cells from immune surveillance.

Figure 1.

Mechanisms underlying the effects of galectin-1 and -3 on apoptosis. (a) Extracellular galectin-1 promotes apoptosis. Binding of galectin-1 to the cell surface ligands CD45/CD43 induces increased expression of Fas/CD95 and initiation of the caspase-8-mediated extrinsic apoptosis signaling. Galectin-1 cell surface binding can also induce CD43 cell surface clustering and subsequent induction of cyochrome c release from the mitochondria and apoptosis. Galectin-1 also induces mitochondrial release of enduclease G and cell death through a caspase-independent mechanism. (b) Intracellular galectin-3 inhibits apoptosis but extracellular galectin-3 promotes apoptosis. Following apoptotic stimulation, galectin-3 binds to synxin and transports to the mitochondrial membrane where it interacts with Bcl-2 and prevents mitochondrial release of cytochrome c and apoptosis. Galectin-3 can also interact directly with Fas/CD95 and prevent the activation of caspase-8-mediated extrinsic apoptosis signaling. Extracellular galectin-3 interacts with the cell surface CD29/CD7 and promotes mitochondrial release of cytochrome c and apoptosis. Solid line arrows indicate direct interactions and dashed line arrows indicate indirect actions.

Table 2. Galectin binding ligands and the influences of galectin–ligand interactions in colorectal cancer
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In contrast, galectin-3 prevents apoptosis. This anti-apoptotic activity is conferred by a functional anti-death (NWGR) motif in its amino acid sequence, which is conserved in the BH1 domain of the Bcl2 gene family.35 Investigation of the molecular mechanism of galectin-3-mediated apoptosis inhibition has been largely conducted in T cells and breast cancer cells. Following apoptosis stimulation by stimuli such as cisplatin, genistein or nitric acid, intracellular galectin-3 interacts with Bcl-235 and synexin36 and translocates to the mitochondrial membrane at the peri-nuclear region where it prevents mitochondrial depolarization and cytochrome c release (Fig. 1b). Intracellular galectin-3 can also complex with CD95/Fas and inhibit CD95/Fas-mediated caspase-8 activation and apoptosis.37 The influence of galectin-3 on apoptosis inhibition is regulated by the phosphorylation/dephosphorylation status of its Ser-6 residue, which acts as an “on/off” switch in regulating the binding of galectin-3 to its ligands.38 Thus, substitution of the Ser-6 residue with Ala decreased the anti-apoptotic activity of galectin-3 in BT549 cells.39 Conversely, exogenous galectin-3 induces apoptosis.37 Galectin-3 binds to the cell surface CD29/CD7 complex and triggers mitochondrial release of cytochrome c and caspase-3 activation (Fig. 1b). Thus, although galectin-3 and -1 have many mutual cell surface ligands (Table 2) and both of them can also induce apoptosis when introduced exogenously; their actions on apoptosis are initiated through binding to different cell surface ligands.

A distinctive difference between the actions of intra- and extra-galectins on cell signaling and apoptosis is that the actions of extracellular galectins are often associated with galectin di/oligomerization and induction of their receptor clustering (lattice formation) whereas no evidence so far indicates this is the case for the actions of intracellular galectins. Extracellular galectins have to date all been shown to have their effects on cellular function as a consequence of lectin-carbohydrate interactions whereas the known actions of intracellular galectins are all mediated via protein-protein interactions. The galectin–monosaccharide interactions are usually weak (Kds ˜4 μM) but low affinity galectin binding events occur due to the multivalent nature of galectins, which often induce clustering. No information is currently available regarding the binding affinity of intracellular galectins with their protein ligands.

Galectins as Inducers of Tumor Angiogenesis

New blood vessel formation from pre-existing vasculature (angiogenesis) is an essential step in cancer progression. Galectin-1 and -3 have both been shown to promote tumor angiogenesis. Over-expression of galectin-1 is observed in the connective tissue surrounding cancer cells in high-grade colonic carcinomas43 as well as in tumor-associated vascular endothelial cells.42, 43, 59, 60 In galectin-1-null [gal-1(−/−)] mice, tumor angiogenesis is reduced in various tumor models.61 Two separate mechanisms have been proposed for galectin-1-mediated angiogenesis promotion. Galectin-1 can increase angiogenesis by interaction with neuropilin-1 on the endothelial cell surface.42 Binding of galectin-1 to europilin-1, which acts as a co-receptor of VEGF in endothelial cells, enhances VEGF receptor phosphorylation and subsequent activation of mitogen-activated protein kinase (MAPK). Galectin-1, secreted from tumor cells, can also be taken up by tumor-associated endothelial cells and stimulate endothelial cell proliferation, adhesion and migration as a result of activation of Ras-Raf-Erk endothelial signaling.61

Subcutaneous injection of galectin-3-expressing human breast 11-9-1-4 cells or recombinant galectin-3/Matrigel into nude mice is associated with increased formation of capillary density in tumors in comparison with those injected with non-galectin-3-expressing BT549 cells or control Matrigel. Exogenous addition of galectin-3 to HUVECs increases HUVECs tube formation in cell culture.62 This effect has been suggested to be dependent on galectin-3 interaction with aminopeptidase N/CD13 (APN), an endothelial cell surface enzyme.50 Recently, the galectin-3-mediated angiogenesis has also been shown to be attributed to the influence of galectin-3 on the function of vascular endothelial growth factor (VEGF) and basic fibroblast growth factor (bFGF).63 Galectin-3 binds to αvβ3 integrin on the endothelial cell surface and induces integrin clustering followed by activation of focal adhesion kinase and promotion of VEGF- and bFGF-mediated endothelial migration and vessel branch formation.63

These findings indicate that galectin-3, and maybe also galectin-1, can influence tumor angiogenesis by interactions with several endothelial cell surface receptors. It remains to be determined whether the other members of the galectin family, such as galectin-4 and galectin-8 which show altered expression in colorectal cancer, have similar effects on tumor angiogenesis. Recently, it has been reported that rat galectin-8 can induce bovine endothelial migration and morphogenesis in vitro and promotes angiogenesis in vivo in BALB/c mice as a result of galectin-8 interaction with CD166, a member of the immunoglobulin.64

Galectins in Cancer Cell Adhesion and Metastasis

Galectins are increasingly recognized as important players in metastasis by virtue of their effects on cancer cell–matrix, cancer cell–cell and cancer-endothelial adhesion. Transfection of human colon cancer Colo201 cells with galectin-1 is associated with increased adhesion of the cells to the extracellular matrix components fibronectin and laminin.25 Cell surface expression of galectin-3 enhances cancer cell–cell and cancer cell–matrix interactions and promotes cancer cell metastatic spread from primary to secondary tumor sites.65, 66 Laminin, carcinoembryonic antigen and lysosome-associated membrane glycoproteins (Lamps) are all potential galectin-3 cell surface ligands in galectin-3-mediated cell adhesion.48 When introduced exogenously, galectin-3 can bind to beta1,6-acetylglucosaminyltransferase V(Mgat5)-modified N-glycans on cancer cell surface and stimulates cellular FAK and PI3K activation and translocation of α5β1 integrin to fibrillar adhesions.49 Galectin-3 can also cross-link Mgat5-modified N-glycans expressed by epidermal growth factor receptor (EGFR) or transforming growth factor-β receptor (TGFβR) to form lattice-like structures on the cell surface and delay/prevent endocytosis of these receptors.54, 67 As EGFR and TGFβR are growth- and arrest-promoting receptors, respectively, delay of their endocytosis will influence cell growth and arrest. The formation of galectin lattices on the cell surface can also maintain the receptor densities at levels that promote the invasive phenotypes in transformed cells.

Reduction of galectin-3 expression using anti-sense technology before tumor cell inoculation in mice is associated with a marked reduction in liver colonization and spontaneous metastasis by the high-metastasing colonic adenocarcinoma cells LSLiM6 and HM7 whereas increase of galectin-3 expression is associated with an increase in liver metastasis by the low-metastasizing colonic LS174T cells.68 Higher galectin-3 expression in colon cancer patients is associated with increased risk of metastasis and has been suggested to be a useful prognostic marker.69, 70

Galectin-3 is also released into the circulation. Concentrations of circulating galectin-3 in the bloodstream of colorectal cancer patients can be increased up to 5-fold.10 Moreover, patients with metastasis have higher levels of circulating galectin-3 than those with localized tumors.10 The beta subunit of haptoglobin co-precipitates with galectin-3 from the serum of patients with colorectal cancer.53 Recent studies in our laboratory have suggested that the increased circulation of galectin-3 in the bloodstream of cancer patients can be an important promoter of cancer cell metastasis.45, 71 Galectin-3 interacts with the oncofetal Thomsen-Friedenreich (Galactoseβ1,3N-acetylgalactosamineα-, TF) antigen on the transmembrane mucin protein MUC1 expressed by many cancer cells46 and induces MUC1 cell surface polarization and exposure of the smaller cell adhesion molecules, which otherwise are concealed by the much larger and heavily glycosylated MUC1. This results in increased heterotypic adhesion of the cancer cells to vascular endothelium45 and increased homotypic aggregation of the cancer cells to form micro-tumor emboli that prolong tumor cell survival in the circulation46, 72 (Fig. 2).

Figure 2.

A model of the galectin-3-MUC1 interaction in promoting cancer cell–cell and cancer cell–endothelial adhesion and metastasis. Cancer-associated transmembrane mucin protein MUC1 is over expressed in most epithelial cancer cells. After invasion of epithelial (e.g., colorectal) cancer cells into the bloodstream from the primary tumor sites, the massive and heavily charged MUC1, which protrudes over ten times higher from the cell surface than the typical cell surface adhesion molecules, acts as a “protective shield” on the cell surface, covering the underlying adhesion molecules and blocking cancer cell interaction with adjacent cells. Binding to the TF antigen on cancer-associated MUC1 by circulating galectin-3 induces MUC1 cell surface polarization and exposes the smaller cell surface adhesion molecules. This results in adhesion of the cancer cells to the vascular endothelium and also increased cancer cell–cell homotypic aggregation to form micro-tumor emboli that prolong tumor cell survival in the circulation as a result of their increased resistance to anoikis.

There is evidence that the expression of galectin-4 and -8 can negatively regulate colorectal cancer cell behaviors essential for cancer progression. When transfected into galectin-4-negative colon cancer cells, galectin-4 reduces cell migration and motility. This effect of galectin-4 is related to its direct interaction with Wnt signaling protein.57 The SO3-→3-Galβ1→3GalNAc pyranoside,56 a carbohydrate structure present both in O-linked glycosides and glycosphingolipids,73 is recognized by galectin-4 on the surface of colon cancer cells. Galectin-8 transfection in human carcinoma cells reduces cell adhesion and colony formation.58 This effect is due to its selective interaction with the sugar moieties on integrin α3, α6, β1, and, to a lesser extent, α4 and β3, subunits.58

Galectins as Therapeutic Targets in Colorectal Cancer

There is strong in vitro and in vivo evidence that tumorigenesis and metastasis can be reduced by galectin inhibitors. Galectin-3-null mice are relatively healthy,74 indicating that inhibition of galectin-3-mediated actions may present a viable and relatively safe therapeutic approach for cancer treatment.

Natural small saccharides such as β-D-galactose, D-lactose, N-acetyllactosamine (LacNAc) galβ1,3Ara and galβ1,4Man are galectin ligands.75, 76 When administered intraperitoneally, D-galactose completely inhibited liver metastasis of L-1 sarcoma cells in mice.77 However, these natural galectin ligands are very sensitive to hydrolysis, which limits their effective use as therapeutic drugs. Modification of the sugar anomeric structure to increase their biological action has been investigated by several groups.78 Modifications of the carbohydrate structures by thio residues increase their resistance to acidic and enzymatic hydrolysis. Inoculation of 1-Methyl-β-D-lactoside in mice reduced lung metastases of the B16 murine melanoma by 35–45%.79 Daily intraperitoneal injections of glycoamines reduced metastasis of MDA-MB-435 human breast carcinoma in vivo in nude mice.80

Carbohydrate polymers have also been investigated for their potential to block galectin-mediated actions in cancer. Tree-shaped monodisperse glycodendrimers, obtained by repetitive assembly cycles with the carbohydrate ligands composing the outer sphere, inhibit galectin-1 and -3 binding to their carbohydrate ligands in a solid phase assay.78, 81 Small synthetic peptides, which bind specifically to the C-terminal CRD of galectin-3, can also inhibit galectin-3-mediated cancer cell adhesion in vitro and metastasis in vivo in mice.82

The potential for modified citrus pectin (MCP) to inhibit galectin-3-mediated cancer promotion has also been examined. Pectin is a carbohydrate polymer found in the peel and pulp of fruits and is composed of a complex multi-branched structure, rich in galactose. The dominant structure of pectin is the linear chain of poly-α-(1-4)-D-galacturonic acid with varying levels of carboxylic acid methylation, which makes up a “smooth region.”78 This region of pectin is occasionally broken up by side sugar chains rich in neutral sugars, such as arabinose, galactose and rhamnose. MCP produced by degradation of the galacturonic acid chain by α-elimination followed by partial acid degradation of the natural saccharides (MCP) results in a reduced molecular weight of pectin down from an average of 70-100 kDa to an average of 10 kDa. MCP is more soluble and better absorbed by the gut and inhibits galectin-3-mediated suppression of apoptosis, and galectin-3-mediated cell invasion, angiogenesis and cell resistance to chemotherapy.83 Intravenous injection of MCP decreased metastasis of B16-F1 melanoma cells to the lungs by more than 90%. When given orally to mice, MCP significantly reduced MDA-MB-435 breast carcinoma growth and metastasis to the lungs.84 It is unclear to what extent the pectin becomes absorbed into the bloodstream and further studies are needed to clarify the mechanisms involved.


Alterations of galectin expression are common in cancers of the human gastrointestinal tract. Accumulating evidence supports an active role of these multi-functional molecules in the regulation of colorectal cancer development, progression and metastasis. Future studies will provide greater insight into the molecular mechanisms that underlie the complex effects of galectin–ligand interactions. Selective targeting of the potentially harmful effects of galectins represents an attractive therapeutic approach for the development of new treatments for colorectal cancer.