lewis-y-antigen and Disease-Models--Animal

Tumor-associated macrophages (TAMs) are key components of tumor microenvironment (TME) during tumorigenesis and progression. However, the role of TAMs in lung adenocarcinoma is still unclear. In this study, we aimed to clarify the mechanism underlying the crosstalk between TAMs and epithelial-mesenchymal transition (EMT) of lung adenocarcinoma. Fucosyltransferase IV (FUT4) and its synthetic cancer sugar antigen Lewis Y (LeY) was aberrantly elevated in various solid tumors, it plays critical role in the invasion and metastasis. Here, we found that in lung adenocarcinoma samples, the density of TAMs correlates with E-cadherin level and LeY level. In vitro assays, M2 macrophages promoted FUT4/LeY expression through the transforming growth factor-β1(TGF-β1)/Smad2/3 signaling pathway. FUT4/LeY was indispensable in M2 macrophages-mediated cytoskeletal remodeling and EMT. Furthermore, fucosylation of Ezrin mediated by FUT4/LeY can promote the phosphorylation of Ezrin, which was the critical mechanism of M2 macrophages-induced EMT. In vivo assays confirmed that M2 macrophages promoted EMT through the up-regulation of LeY and phosphorylated Ezrin. Together, our results revealed that TAMs promote Ezrin phosphorylation-mediated EMT in lung adenocarcinoma through FUT4/LeY- mediated fucosylation. Targeting this newly identified signaling may offer new possibilities for immunotherapy in lung adenocarcinoma.

Topics: Adenocarcinoma; Adenocarcinoma of Lung; Animals; Biomarkers; Cadherins; Cell Count; Cell Line, Tumor; Cytoskeletal Proteins; Cytoskeleton; Disease Models, Animal; Epithelial-Mesenchymal Transition; Female; Fucosyltransferases; Heterografts; Humans; Lewis Blood Group Antigens; Lewis X Antigen; Lung Neoplasms; Macrophages; Mice; Models, Biological; Phosphorylation; Signal Transduction; Smad2 Protein; Smad3 Protein; Transforming Growth Factor beta1; Up-Regulation

The N-terminal lectin-like domain (domain 1 [D1]) of thrombomodulin (TM) is known to have an anti-inflammatory function. We previously showed that recombinant TM domain 1 (rTMD1) interacts with a carbohydrate molecule, Lewis Y (Le(y)), which is found to be expressed on adhesion molecules and involved in cell adhesion. Here, we tested the effect of rTMD1/Le(y) interaction on leukocyte recruitment in inflammation.. The expression of Le(y) on the surface of human umbilical vein endothelial cells was increased by tumor necrosis factor-α stimulation. Direct binding of rTMD1 to Le(y) on the cell surface was observed. rTMD1 inhibited Le(y)-mediated leukocyte adhesion on the Le(y)-immobilized flow chamber and activated endothelium under a shear flow. The following leukocyte transmigration to endothelium was also reduced by rTMD1 through binding Le(y). In vivo, treatment of rTMD1 reduced leukocyte recruitment to the inflammatory sites in carotid ligation injury and thioglycollate-induced peritonitis. rTMD1 administration in apolipoprotein E-deficient mice effectively suppressed atherosclerotic plaque formation and macrophage infiltration in atherosclerotic lesions. Increased Le(y) expression, as well as administered rTMD1, was observed in inflamed vessels.. rTMD1 suppresses vascular inflammation by inhibiting leukocyte recruitment to endothelium through attenuating Le(y)-mediated adhesion and further protects against atherosclerosis progression. The present study provides a mechanism showing that rTMD1 can inhibit inflammation by binding to its carbohydrate ligand Le(y).

Topics: Animals; Anti-Inflammatory Agents; Apolipoproteins E; Atherosclerosis; Carotid Artery Injuries; Cell Adhesion; Cell Line, Tumor; Chemotaxis, Leukocyte; Coculture Techniques; Disease Models, Animal; Dose-Response Relationship, Drug; Endothelial Cells; HEK293 Cells; Human Umbilical Vein Endothelial Cells; Humans; Leukocytes, Mononuclear; Lewis Blood Group Antigens; Macrophages; Mice; Mice, Inbred C57BL; Mice, Knockout; Neutrophils; Plaque, Atherosclerotic; Protein Structure, Tertiary; Recombinant Proteins; Thrombomodulin; Transendothelial and Transepithelial Migration; Tumor Necrosis Factor-alpha; Vasculitis

The SGN-15 monoclonal antibody-doxorubicin immunoconjugate is toxic to Lewis(Y) antigen-expressing cells and is effective against intracerebral tumors in nude rats when delivery is enhanced with osmotic disruption of the blood-brain barrier (BBB). We tested whether doxorubicin released locally in antigen-expressing cells would affect adjacent non-antigen-expressing cells in heterogeneously expressing intracerebral tumors.. Nude rats with intracerebral xenografts of human small cell lung carcinoma cells with high (n = 10) or low (n = 23) Lewis(Y) antigen expression were treated with SGN-15 at a low (10 mg/kg) or high (140 mg/kg) antibody dose, administered intra-arterially with BBB disruption, and tumor volumes and antigen expression were evaluated after 6 days.. BBB disruption-enhanced delivery of SGN-15 (10 mg/kg) reduced the high-expressor tumor volume from 26.1 +/- 3.7 mm(3) (n = 7) in untreated control animals to 6.7 +/- 4.6 mm(3) (n = 3, P < 0.05). Untreated high-expressor tumors exhibited uniform prominent Lewis(Y) antigen staining (97.6 +/- 0.9% positive, n = 4), whereas treated tumors demonstrated areas of nil, moderate, and prominent staining (71.0 +/- 8.5% positive, n = 3, P < 0.05). In intracerebral tumors with low initial antigen expression, BBB disruption-enhanced delivery of a low dose of immunoconjugate was significantly effective, but treated tumors demonstrated low levels of antigen expression. An increase in the immunoconjugate dose did not significantly alter either efficacy or antigen expression.. Immunoconjugate delivered across the BBB was effective against antigen-positive tumor cells, but there was not an effective chemical bystander effect against antigen-negative tumor cells.

Topics: Animals; Antibodies, Monoclonal; Antigens, Heterophile; Brain Neoplasms; Carcinoma, Small Cell; Cell Line, Tumor; Disease Models, Animal; Doxorubicin; Female; Humans; Lewis Blood Group Antigens; Rats; Rats, Nude; Xenograft Model Antitumor Assays

The population dynamics of Helicobacter pylori during colonization in an infected animal host provide a quantifiable experimental model of in vivo microbial phenotype evolution. Phenotype variability in H. pylori populations can be typed as polymorphic expression of Lewis antigens on their cell surfaces. The high mutational frequency of H. pylori for Lewis expression provides substrate for differential selection by the host. Experimental challenge and successful colonization of mice and gerbils allows tracking of H. pylori phenotype variability from the initial inoculation to the ultimate establishment of a quasispecies. Colonization data provide a quantitative experimental model of phenotype evolution in a relatively large population (>10(4) individuals) over a relatively long evolutionary time scale (>10(3) generations). A mathematical model is developed to interpret the data in terms of the dynamic processes occurring during colonization. The mathematical model distinguishes the roles of selection and mutation; quantifies the effects of initial phenotype diversity, mutational frequency, and selective advantage; and applies generally to phenotype evolution in biological populations.

Topics: Animals; Disease Models, Animal; Evolution, Molecular; Female; Gerbillinae; Helicobacter Infections; Helicobacter pylori; Lewis Blood Group Antigens; Lewis X Antigen; Lipopolysaccharides; Mathematical Computing; Mice; Mice, Inbred C3H; Models, Biological; Mutagenesis; Phenotype; Selection, Genetic

The humanized complementarity determining region-grafted anti-Lewis Y (Le(y)) monoclonal antibody [humanized 3S193 (hu3S193)] was developed for targeting Le(y)-expressing epithelial tumors such as breast, colon, lung, prostate, and ovarian carcinoma. We are exploring the potential use of smaller molecular size, bivalent analogues of hu3S193, because the faster blood clearance of M(r) approximately 54,000 diabody and M(r) approximately 110,000 F(ab')(2) molecules may be advantageous in achieving optimal and rapid tumor uptake for diagnostic and potential therapeutic applications. The single-chain variable fragment-5 residue linker construct (diabody) was expressed using the bacterial secretion vector pPOW3, and soluble product was purified without refolding processes. The F(ab')(2) fragment was obtained by pepsin digest of parental hu3S193. To facilitate evaluations, the radiometal (111)In was used to label C-functionalized trans-cyclohexyl diethylenetriaminepentaacetic acid chelated diabody and F(ab')(2). The immunoreactivity of the radiolabeled constructs was 41.3 and 58.6%, and the K(a) was 1.68 x 10(6) M(-1) and 5.33 x 10(6) M(-1) for the diabody and F(ab')(2), respectively. Radioconjugates were injected into mice bearing Le(y)-positive MCF-7 tumors, and biodistribution properties were determined at various time points after injection. The uptake of radiolabeled diabody in xenografts was maximal at 1 h after injection (4.7 +/- 0.6% injected dose/g), whereas the F(ab')(2) peaked at 8 h after injection (14.2 +/- 2.4% injected dose/g). The tumor:blood ratio at 4 h for the diabody and F(ab')(2) was 5:1 and 2:1, which increased to 20:1 and 5:1, respectively, at 8 h and increased further to 40:1 and 130:1, respectively, at 48 h. These results demonstrate that the diabody construct may have applications as a diagnostic imaging reagent, whereas F(ab')(2) displayed effective tumor targeting and may have potential as a therapeutic molecule in patients with Le(y)-expressing tumors.

Topics: Animals; Antibodies, Monoclonal; Antibody Affinity; Breast Neoplasms; Disease Models, Animal; Female; Gene Targeting; Humans; Immunoglobulin Fab Fragments; Indium Radioisotopes; Isothiocyanates; Lewis Blood Group Antigens; Mice; Mice, Inbred BALB C; Mice, Nude; Neoplasm Transplantation; Pentetic Acid; Transplantation, Heterologous; Tumor Cells, Cultured

Topics: Animals; Apoptosis; Disease Models, Animal; Escherichia coli Proteins; Helicobacter Infections; Helicobacter pylori; Humans; Hyperplasia; Lewis Blood Group Antigens; Lewis X Antigen; Lymphocytes; Transcription Factors

LMB-1 (B3-LysPE38) is an immunotoxin composed of the tumor-reactive monoclonal antibody B3 and a genetically engineered form of Pseudomonas exotoxin. Monoclonal antibody B3 reacts with a carbohydrate epitope that is found on a number of solid tumors (e.g., breast, ovarian, and lung carcinomas) that frequently invade the intrathecal space, causing neoplastic meningitis. The Pseudomonas exotoxin has been engineered to remove the binding domain to eliminate nonspecific binding. A model of human neoplastic meningitis using rats bearing the human epidermoid carcinoma A431 was used for therapeutic studies of immunotoxin LMB-1. Therapy was initiated 3 days after injection of the tumor cells, which was one third of the median survival time of untreated rats. A single intrathecal injection of 40 microgram increased median survival from 9 days with saline injection to 16 days (78%, P < 0.001), and a single dose of 200 microgram increased median survival to 25 days (188%, P < 0. 001). Three doses of 40 or 200 microgram given on days 3, 6, and 8 significantly increased the median survival of 9.5 days associated with saline injection to 40.5 days (326% increase) and 33.0 days (247% increase), respectively, with two long-term survivors (191-day survival) in each treatment group. LMB-1 had no therapeutic effect on the treatment of two B3 antigen-negative neoplastic meningitis models. Treatment of the antigen-positive A431 neoplastic meningitis with B3 alone or a nonspecific monoclonal, MOPC, coupled to the engineered Pseudomonas exotoxin produced no survival effects. Nontumor-bearing athymic rats showed no toxicity with a single dose of either 40 microgram or 200 microgram, or 3 doses of 40 microgram. However, when they were given three doses of 200 microgram, these rats showed weight loss and loss of neurological function, and two of eight animals died. These studies indicate that, in the range of the most therapeutically effective dosage, the immunotoxin LMB-1 is tolerated in the intrathecal space and should be considered for human intrathecal trials.

Topics: ADP Ribose Transferases; Animals; Antibodies, Monoclonal; Bacterial Toxins; Carcinoma; Disease Models, Animal; Drug Screening Assays, Antitumor; Exotoxins; Humans; Immunotoxins; Lewis Blood Group Antigens; Meningeal Neoplasms; Pseudomonas aeruginosa Exotoxin A; Rats; Rats, Nude; Tumor Cells, Cultured; Virulence Factors