Sen-itiroh Hakomori

Aberrant Glycosylation In Tumors And Tumor-Associated Carbohydrate Antigens

Publisher Summary Aberrant glycosylation is the most common phenomenon associated with oncogenic transformation expressed in cell membranes of animal and human cancer cells. Such aberrant structures at the surface membranes may well be effective targets in prevention, diagnosis, and treatment of human cancer. Many of the aberrant glycosylation products can be recognized by specific MAbs as tumor-associated carbohydrate antigens. Many of these antigens have been identified as carbohydrates, thus there is increasing evidence that essentially all human cancers are characterized by aberrant glycosylation. Aberrant glycosylation as such may be the basis of inappropriate cell/cell and cell/matrix interactions that may be reflected in the abnormal cell social behavior of tumor cells, such as uncontrolled cell growth, invasiveness, and metastatic potential. Whatever the genetic or epigenetic basis of expression of aberrant glycosylation is, the phenomenon is of crucial importance in understanding the antisocial behavior of tumor cells as well as in practical applications in diagnosis and treatment of human cancer.

Glycosylation defining cancer malignancy: New wine in an old bottle

Aberrant glycosylation occurs in essentially all types of experimental and human cancers, as has been observed for over 35 years, and many glycosyl epitopes constitute tumor-associated antigens. A long-standing debate is whether aberrant glycosylation is a result or a cause of cancer. Many recent studies indicate that some, if not all, aberrant glycosylation is a result of initial oncogenic transformation, as well as a key event in induction of invasion and metastasis. Glycosylation promoting or inhibiting tumor cell invasion and metastasis is of crucial importance in current cancer research. Nevertheless, this area of study has received little attention from most cell biologists involved in cancer research, mainly because structural and functional concepts of glycosylation in cancer are more difficult to understand than the functional role of certain proteins and their genes in defining cancer cell phenotypes. Glycosylation appears to be considered “in the shade” of more popular topics such as oncogenes and antioncogenes, apoptosis, angiogenesis, growth factor receptors, integrin and caderin function, etc., despite the fact that aberrant glycosylation profoundly affects all of these processes. The apoptotic signaling triggered by glycosylation presents an immense challenge for future study. The concept of glycosylation-dependent promotion or inhibition of tumor progression has developed in conjunction with clinicopathological studies. High expression of some glycosyl epitopes promotes invasion and metastasis, leading to shorter 5–10 year survival rates of patients, whereas expression of some other glycosyl epitopes suppresses tumor progression, leading to higher postoperative survival rates (for review see refs. 1 and 38). The former category of epitopes includes β6GlcNAc branching in N -linked structure; sialyl-Tn in O -linked structure; sialyl-Lex, sialyl-Lea, and Ley in either N -linked, O -linked, or lipid-linked structure; GM2, GD3, and sialyl-Gb5 in lipid-linked structure. The latter category includes β4GlcNAc competitive with β6GlcNAc; histo-blood group A …

Determination of aminosugar linkages in glycolipids by methylation: Aminosugar linkages of ceramide pentasaccharides of rabbit erythrocytes and of Forssman antigen☆

Gas chromatography-mass spectrometric identification of partially methylated aminosugars has been developed: (a) various kinds of O-methylated 2-deoxy-2-(N-methyl)-acetamidohexitols were prepared from partially O-(1-methoxy)-ethylated 2-deoxy-2-acetamidohexoses, and their gas chromatography-mass spectrometric patterns were determined; (b) permethylated glycolipids gave a satisfactory yield of 2-deoxy-2-N-methyl-amidohexoses by acetolysis with 0.5 n sulfuric acid in 95% acetic acid, followed by aqueous hydrolysis; (c) the resulting partially methylated aminosugars and neutral sugars were analyzed after borohydride reduction and acetylation according to the procedure of Lindberg and associates (Bjorndal, Lindberg and Svennson, 1967; Bjorndal, Hellerqvist, Lindberg and Svensson, 1970). This method was successfully applied to analysis of aminosugar linkages in blood group B-active ceramide pentasaccharide from rabbit erythrocytes and in Forssman antigen of equine spleen. The structure of blood group B-active glycolipid of rabbit erythrocyte was found to be Galα1 → 3Galβ1 → 4G1cNAcβ1 → 3Ga11 → 4Glc → Cer, and that of Forssman antigen to be GaNAcα1 → 3GalNAcβ1 → 3Galα1 → 4Ga11 → 4Glc → Cer.

ABH and related histo-blood group antigens : immunochemical differences in carrier isotypes and their distribution

Abstract. This review summarizes present knowledge of the chemistry of histo‐blood group ABH and related antigens. Recent advances in analytical carbohydrate chemistry (particularly mass spectrometry and NMR spectroscopy) and the introduction of monoclonal antibodies (MoAbs) have made it possible to distinguish structural variants of histo‐blood group ABH antigens. Polymorphism of ABH antigens is induced by: (i) variations in peripheral core structure, of which four (type 1, 2, 3 and 4) are known in man; (ii) variation in inner core by branching process (blood group iI), leading to variation of unbranched vs. branched ABH determinants; (iii) biosynthetic interaction with other glycosyltransferases (Lewis, P. T/Tn blood systems) capable of acting on the same substrate as the ABH‐defined transferases, and finally (iv) the nature of the glycoconjugate (glycolipid, glycoprotein of N‐ or O‐linked type). ABH variants induced by item (i) above have been clearly distinguished qualitatively by MoAbs; e.g., at least six types of A determinants can be distinguished by qualitatively different classes of antibody. The variants induced by item (ii) create mono‐ vs. bivalent antigens which may be responsible for observed differences in antibody‐binding affinity.

INAUGURAL ARTICLE by a Recently Elected Academy Member:The glycosynapse

Physically distinguishable microdomains associated with various functional membrane proteins are one of the major current topics in cell biology. Glycosphingolipids present in such microdomains have been used as “markers;” however, the functional role of glycosyl epitopes in microdomains has received little attention. In this review, I have tried to summarize the evidence that glycosyl epitopes in microdomains mediate cell adhesion and signal transduction events that affect cellular phenotypes. Molecular assemblies that perform such functions are hereby termed “glycosynapse” in analogy to “immunological synapse,” the membrane assembly of immunocyte adhesion and signaling. Three types of glycosynapses are so far distinguishable: (i) Glycosphingolipids organized with cytoplasmic signal transducers and proteolipid tetraspanin with or without growth factor receptors; (ii) transmembrane mucin-type glycoproteins with clustered O-linked glycoepitopes for cell adhesion and associated signal transducers at lipid domain; and (iii) N-glycosylated transmembrane adhesion receptors complexed with tetraspanin and gangliosides, as typically seen with the integrin–tetraspanin–ganglioside complex. The possibility is discussed that glycosynapses give rise to a high degree of diversity and complexity of phenotypes.

GM3-enriched Microdomain Involved in Cell Adhesion and Signal Transduction through Carbohydrate-Carbohydrate Interaction in Mouse Melanoma B16 Cells

Mouse melanoma B16 cells are characterized by the predominant presence of ganglioside GM3 and adhere to lactosylceramide- or Gg3-coated plates through interaction of GM3 with lactosylceramide or Gg3, whereby not only adhesion but also spreading and enhancement of cell motility occur (Kojima, N., Hakomori, S. (1991) J. Biol. Chem. 266, 17552–17558). We now report that the adhesion process is based essentially on a glycosphingolipid-enriched microdomain (GEM) at the B16 cell surface, since >90% of GM3 present in the original cells is found in GEM, and GEM is also enriched in several signal transducer molecules, e.g. c-Src, Ras, Rho, and focal adhesion kinase (FAK). GEM was isolated as a low density membranous fraction by homogenization of B16 cells in lysis buffer under two different conditions (i.e. buffer containing 1% Triton X-100, or hypertonic sodium carbonate without detergent), followed by sucrose density gradient centrifugation. A close association of GM3 with c-Src, Rho, and FAK was indicated by co-immunoprecipitation of GM3 present in GEM by anti-GM3 monoclonal antibody DH2, followed by Western blotting with antibodies directed to these transducer molecules. The following data indicate that GEM is a structural and functional unit for initiation of GM3-dependent cell adhesion coupled with signal transduction. 1) Tyrosine phosphorylation in FAK was greatly enhanced in B16 cells adhered to Gg3-coated plates but was minimal in cells adhered to GM3-coated, GlcCer-coated, or noncoated plates. 2) GTP loading on Ras and Rho increased significantly when cells were adhered to Gg3-coated plates, compared with GM3-coated, GlcCer-coated, or noncoated plates. Since Ras and Rho are closely associated with GM3 in GEM, cell adhesion/stimulation through GM3 in GEM may induce activation of Ras and Rho through enhanced GTP binding.

Glycosphingolipid antigens and cancer therapy

Specific types of glycosphingolipid (GSL), which are chemically detectable in normal cells, are more highly expressed in tumors. The high level of expression on the surfaces of tumor cells causes an antibody response to these GSLs, which can therefore be described as tumor-associated antigens. Some of these GSLs have been shown to be adhesion molecules involved in tumor cell metastasis, and to be modulators of signal transduction controlling tumor cell growth and motility. Tumor-associated GSL antigens have been used in the development of antitumor vaccines. GSLs and sphingolipids involved in adhesion and signaling are therefore targets for cancer therapy.

Correlation of Expression of H/LeY/LeB Antigens with Survival in Patients with Carcinoma of the Lung

BACKGROUND The level of expression of H/Le(y)/Le(b) antigens is high in various histologic types of lung cancer, a feature that may be related to deletion of A and B blood-group antigens. We evaluated the possibility that expression of this antigen, which can be defined by the monoclonal antibody MIA-15-5, might be of prognostic value, as suggested by our previous observation that MIA-15-5 inhibits tumor-cell motility and metastasis. METHODS We used MIA-15-5 to stain tissue sections from 149 patients with primary lung cancer whose clinico-pathological histories were well documented. The survival curves for patients whose tumors stained positively were compared with the curves for those whose tumors stained negatively. Multivariate analyses were performed with a Cox proportional-hazards regression model. RESULTS Among the 149 patients studied, five-year survival in the 91 patients with MIA-positive tumors was significantly lower than survival in the 58 with MIA-negative tumors (20.9 percent vs. 58.6 percent, P less than 0.001). Among the 67 patients with squamous-cell carcinoma, the rates also differed significantly (10.5 percent vs. 62.1 percent, P less than 0.001). The difference in survival between patients with MIA-positive tumors and those with MIA-negative tumors was significant among patients with blood groups A and AB (P less than 0.001), but not among those with blood group B or O (P = 0.071 and 0.068, respectively). Multivariate analysis with the Cox regression model indicated that positivity best correlated with five-year mortality, followed by lymph-node status (N stage) and tumor size status (T stage), whereas sex, age, and blood group did not correlate with mortality. CONCLUSIONS Positivity for MIA (i.e., immunohistologic staining by MIA-15-5, which defines H/Le(y)/Le(b) antigens) is inversely correlated with survival among patients with primary lung cancer and may be of prognostic value.

Structure, organization, and function of glycosphingolipids in membrane.

A large variety of glycosylation patterns in combination with different ceramide structures in glycosphingolipids provide a basis for cell type-specific glycosphingolipid pattern in membrane, which essentially reflects the composition of glycosphingolipid-enriched microdomains. Functions of glycosphingolipids as antigens, mediators of cell adhesion, and modulators of signal transduction are all based on such organization. Of particular importance is the assembly of glycosphingolipids with signal transducers and other membrane proteins to form a functional unit termed a glycosynapse, through which glycosylation-dependent cell adhesion coupled with signal transduction takes place. The microenvironment formed by interfacing glycosynapses of interacting cells plays a central role in defining phenotypic changes after cell adhesion, as occur in ontogenic development and cancer progression. These basic functional features of glycosphingolipids in membrane can also be considered roles of glycosphingolipids and gangliosides characteristic of neutrophils, myelocytes, and other blood cells. A series of sialyl fucosyl poly-N-acetylgalactosamine gangliosides without the sialyl-Lex epitope, collectively termed myeloglycan, have been shown to mediate E-selectin—dependent rolling and tethering under dynamic flow with physiologic shear stress conditions. Functional roles of myeloglycan in neutrophils during inflammatory processes are discussed

Possible functions of tumor-associated carbohydrate antigens

Expression of some tumor-associated carbohydrate antigens may define the stage, rate and phenotype of tumor progression and may have prognostic value. Some of these antigens are now recognized as adhesion molecules that define the site of metastasis. Monoclonal antibodies to tumor-associated carbohydrate antigens, or the antigens themselves, may serve not only as classic immunological reagents but also as anti-adhesion reagents for the prevention of tumor progression.