g(m1)-ganglioside and Liver-Neoplasms

The hepatitis B virus (HBV)-induced chronic liver diseases are serious health threats worldwide. There is evidence to display the alterations of salivary N-linked glycans related to the development of HBV-infected liver diseases. Here, we further investigated the alterations of fucosylated N/O-glycans recognized by LTL in saliva from 120 subjects (30 healthy volunteers (HV), 30 patients with hepatitis B (HB), 30 patients with hepatic cirrhosis (HC), and 30 patients with hepatocellular carcinoma (HCC)) using salivary microarrys and MALDI-TOF/TOF-MS. The results showed that the expression level of fucosylated glycans recognized by LTL was significantly increased in HCC compared with other subjects (P < 0.0001). Besides, the fucosylated glycoproteins were isolated from pooled saliva of HV, HB, HC, and HCC by LTL-magnetic particle conjugates. Then, N/O- glycans were released from the isolated glycoproteins with PNGase F and NaClO, and were identified by MALDI-TOF-MS, respectively. Totally, there were 21/20, 25/18, 29/19, and 28/24 N/O-glycan peaks that were identified and annotated with proposed structures in saliva of HV, HB, HC, and HCC. Among the total, there were 8 N-glycan peaks (e.g., m/z 1905.634, 2158.777 and 2905.036) and 15 O-glycan peaks (e.g., 1177.407, 1308.444 and 1322.444) that only presented in patients with HBV-induced liver diseases. One N-glycan peak (m/z 2205.766) was unique in HC, and 9 O-glycan peaks (e.g., m/z 1157.420, 1163.417 and 1193.402) were unique in HCC. This study could facilitate the discovery of biomarkers for HC and HCC based on precise alterations of fucosylated N/O-glycans in saliva.

Topics: Biomarkers, Tumor; Carcinoma, Hepatocellular; Female; Fibrosis; G(M1) Ganglioside; Hepatitis B virus; Hepatitis, Chronic; Humans; Lectins; Liver Neoplasms; Male; Polysaccharides; Protein Array Analysis; Saliva; Spectrometry, Mass, Matrix-Assisted Laser Desorption-Ionization

Metastases account for 90% of all cancer-related deaths, becoming a therapeutic problem. Approximately 50% of all uveal melanoma (UM) patients will develop metastases, mainly in the liver. Post-mortem analyses of livers from metastatic UM patients showed two different metastatic growth patterns: infiltrative and nodular. The infiltrative pattern exhibits tumor infiltration directly to the hepatic lobule and minimal angiogenesis. The nodular pattern shows clusters of tumor cells around the portal venules that efface the liver parenchyma. We recently demonstrated Natural Killer (NK) cells play a pivotal role in the control of hepatic metastases and the pigment epithelial-derived factor (PEDF) controls angiogenesis in the liver using our established ocular melanoma animal model. In this study we investigated the role of NK cells and PEDF in the development of metastatic growth patterns, as this can contribute to the development of novel therapeutics specific towards each growth pattern.. Our in vivo work showed two distinct metastatic growth patterns, the infiltrative and nodular, recapitulating the post-mortem analyses on human liver tissue. We discovered NK cells control the infiltrative growth. In contrast, PEDF controlled anti-angiogenic responses, showing higher MVD values compared to NK-depleted and WT animals. The myeloid lineage, comprised of monocytes, macrophages, and myeloid-derived suppressor cells, was reduced in the absence of NK cells or PEDF.. Our animal model recapitulates the metastatic growth patterns observed in the human disease. We demonstrated a role for NK cells in the development of the infiltrative growth pattern, and a role for PEDF in the development of the nodular pattern. The understanding of the complexity associated with the metastatic progression has profound clinical implications in the diagnostic and disease-management as we can develop and direct more effective therapies.

Topics: Animals; Antibodies; Cell Line, Tumor; Eye Proteins; Female; G(M1) Ganglioside; Gene Knockout Techniques; Killer Cells, Natural; Liver Neoplasms; Macrophages; Melanoma; Mice; Mice, Inbred C57BL; Monocytes; Myeloid-Derived Suppressor Cells; Neoplasm Transplantation; Nerve Growth Factors; Serpins; Uveal Neoplasms

Hepatocellular carcinoma (HCC) is one of the most common human malignancies. Therefore, developing the early, high-sensitivity diagnostic biomarkers to prevent HCC is urgently needed. Serum a-fetoprotein (AFP), the clinical biomarker in current use, is elevated in only ~60% of patients with HCC; therefore, identification of additional biomarkers is expected to have a significant impact on public health. In this study, we used glycan microarray analysis to explore the potential diagnostic value of several cancer-associated carbohydrate antigens (CACAs) as biomarkers for HCC. We used glycan microarray analysis with 58 different glycan analogs for quantitative comparison of 593 human serum samples (293 HCC samples; 133 chronic hepatitis B virus (HBV) infection samples, 134 chronic hepatitis C virus (HCV) infection samples, and 33 healthy donor samples) to explore the diagnostic possibility of serum antibody changes as biomarkers for HCC. Serum concentrations of anti-disialosyl galactosyl globoside (DSGG), anti-fucosyl GM1 and anti-Gb2 were significantly higher in patients with HCC than in chronic HBV infection individuals not in chronic HCV infection patients. Overall, in our study population, the biomarker candidates DSGG, fucosyl GM1 and Gb2 of CACAs achieved better predictive sensitivity than AFP. We identified potential biomarkers suitable for early detection of HCC. Glycan microarray analysis provides a powerful tool for high-sensitivity and high-throughput detection of serum antibodies against CACAs, which may be valuable serum biomarkers for the early detection of persons at high risk for HCC.

Topics: alpha-Fetoproteins; Antibodies, Viral; Antigens, Neoplasm; Biomarkers, Tumor; Carcinoma, Hepatocellular; Early Diagnosis; Female; G(M1) Ganglioside; Gangliosides; Hepacivirus; Hepatitis B virus; Hepatitis B, Chronic; Hepatitis C, Chronic; Humans; Liver Neoplasms; Male; Polysaccharides; Sensitivity and Specificity; Sex Factors; Tissue Array Analysis

To explore the role of natural killer T (NKT) cells in the development of liver metastases in mice harboring intraocular melanomas.. Cells derived from the cutaneous B16 melanoma cell line (B16LS9) were transplanted either into the vitreous body or under the spleen capsules of wild-type C57BL/6 mice and NKT-cell-deficient Jα18(-/-) and CD1d(-/-) mice. The development of liver metastases was evaluated by histopathology. The effect of NK cells on liver metastases was determined by selective depletion with anti-asialo-GM1 antiserum in vivo and NK-cell-mediated cytolysis of B16LS9 melanoma cells in vitro. The role of IL-10 and transforming growth factor (TGF)-β in the inhibition of liver NK resistance to liver metastases was determined by in vivo and in vitro neutralization with monoclonal antibodies.. Liver NKT cells, especially type I NKT cells, enhanced liver metastases arising from intraocular melanomas. NKT-cell-deficient mice developed significantly fewer liver metastases that were NK-cell dependent. Tumor-induced liver NKT cells, especially type I NKT cells, inhibited liver NK-cell cytotoxicity by an IL-10-dependent process.. NKT cells exert protective effects in many murine tumor models. However, the present results reveal that NKT cells exacerbate liver metastases arising from intraocular melanomas. To the authors' knowledge, this is the first report that liver NKT cells, especially type I NKT cells, inhibit liver NK-cell antimetastatic activity by the production of IL-10. These results suggest that hepatic NKT cell activity can have an important effect in the immune surveillance of liver metastases.

Topics: Animals; Cytotoxicity, Immunologic; Enzyme-Linked Immunosorbent Assay; Eye Neoplasms; Female; Flow Cytometry; G(M1) Ganglioside; Interleukin-10; Liver; Liver Neoplasms; Male; Melanoma, Experimental; Mice; Mice, Inbred C57BL; Natural Killer T-Cells; Neoplasm Transplantation; Reverse Transcriptase Polymerase Chain Reaction; Splenectomy; Splenic Neoplasms; Transforming Growth Factor beta; Vitreous Body

Galectins, beta-galactoside binding proteins, expressed selectively in human breast carcinoma are attractive targets to employ lectin-aimed therapeutics. We examined beta-galactoside binding potency of neoplastic cells using fluorescein-labelled synthetic glycoconjugates as probes for flow cytometry. As a result, surface beta-galactoside binding proteins/galectins were discovered on mouse mammary carcinoma cells in vitro and in vivo unlike non-malignant cells from the several tissues; and asialo-GM1 ganglioside carbohydrate part--containing probe was the most specific one. However, in liver and lung metastatic cells galectins seem to be expressed within cytoplasm and/or nuclei. Galectin expression correlated directly with aggressive tumour potential in the A/Sn transplantable model similar to findings in several human breast carcinoma cell lines. However, galectin expression was reduced during tumour progression in more aggressive forms of spontaneous BLRB mammary carcinomas like it was shown for human breast carcinoma specimens. Analysis of the histopathological data led, however, to the conclusion that galectin expression hardly might be a suitable marker of aggressiveness of heterogeneous mammary carcinomas as the observed level of galectin expression is influenced by the amount of the stroma in a tumour sample and/or probably, galectin expression inversely correlates with tumour aggressiveness during the initial and advanced steps of mammary tumour progression. We conclude that surface beta-galactoside binding proteins/galectins that are selectively expressed during mouse mammary carcinoma progression, similarly to human breast carcinomas, seem to be proper targets for asialo-GM1-vectored cytotoxics and our mouse model system might be a relevant instrument to further test novel modes of anti-breast cancer therapy.

Topics: Animals; Biomarkers, Tumor; Cell Membrane; Cell Nucleus; Cytoplasm; Disease Progression; Female; G(M1) Ganglioside; Galactosides; Galectins; Glycoconjugates; In Vitro Techniques; Liver Neoplasms; Male; Mammary Neoplasms, Experimental; Mice; Mice, Inbred Strains; Stromal Cells; Tumor Cells, Cultured

Hypoxia-inducible factor 1 alpha (HIF-1alpha), a component of HIF-1, is expressed in human tumors and renders cells able to survive and grow under hypoxic (low-oxygen) conditions. YC-1, 3-(5'-hydroxymethyl-2'-furyl)-1-benzylindazole, an agent developed for circulatory disorders that inhibits platelet aggregation and vascular contraction, inhibits HIF-1 activity in vitro. We tested whether YC-1 inhibits HIF-1 and tumor growth in vivo.. Hep3B hepatoma, NCI-H87 stomach carcinoma, Caki-1 renal carcinoma, SiHa cervical carcinoma, and SK-N-MC neuroblastoma cells were grown as xenografts in immunodeficient mice (69 mice total). After the tumors were 100-150 mm(3), mice received daily intraperitoneal injections of vehicle or YC-1 (30 microg/g) for 2 weeks. HIF-1 alpha protein levels and vascularity in tumors were assessed by immunohistochemistry, and the expression of HIF-1-inducible genes (vascular endothelial growth factor, aldolase, and enolase) was assessed by reverse transcription-polymerase chain reaction. All statistical tests were two-sided.. Compared with tumors from vehicle-treated mice, tumors from YC-1-treated mice were statistically significantly smaller (P<.01 for all comparisons), expressed lower levels of HIF-1 alpha (P<.01 for all comparisons), were less vascularized (P<.01 for all comparisons), and expressed lower levels of HIF-1-inducible genes, regardless of tumor type.. The inhibition of HIF-1 alpha activity in tumors from YC-1-treated mice is associated with blocked angiogenesis and an inhibition of tumor growth. YC-1 has the potential to become the first antiangiogenic anticancer agent to target HIF-1 alpha.

Topics: Angiogenesis Inhibitors; Animals; Antineoplastic Agents; Carcinoma; Carcinoma, Hepatocellular; Cell Hypoxia; Culture Media, Conditioned; Endothelial Growth Factors; Enzyme-Linked Immunosorbent Assay; Female; G(M1) Ganglioside; Gene Expression Regulation, Neoplastic; Humans; Hypoxia-Inducible Factor 1, alpha Subunit; Immunoblotting; Indazoles; Intercellular Signaling Peptides and Proteins; Kidney Neoplasms; Killer Cells, Natural; Liver Neoplasms; Lymphokines; Male; Mice; Mice, SCID; Neoplasms; Neovascularization, Pathologic; Neuroblastoma; Platelet Endothelial Cell Adhesion Molecule-1; Precipitin Tests; Rats; Reverse Transcriptase Polymerase Chain Reaction; Stomach Neoplasms; Transcription Factors; Transplantation, Heterologous; Tumor Cells, Cultured; Uterine Cervical Neoplasms; Vascular Endothelial Growth Factor A; Vascular Endothelial Growth Factors

Surgical manipulation of a tumor may result in increased influx of tumor cells into the systemic and portal circulation and give rise to formation of metastases. In addition, major surgery has been reported to cause profound immunosuppression. In an attempt to increase the host-antitumor immune mechanisms following surgery we have studied the effect of preoperative administration of interferon-gamma, related to the antimetastatic effects of Kupffer cells (KC) and natural killer cells (NK-cells) in the early phase of liver metastasis formation. Colon carcinoma cells were injected into the superior mesenteric vein of syngeneic mice and after 17 days metastases were quantified by weight, number, and uptake of [125I]iododeoxyuridine. Unstimulated control mice developed 10.5 surface nodules per liver 17 days following injection of colon carcinoma cells into the superior mesenteric vein of syngeneic mice. This figure was only 2.6 in mice stimulated with a single dose of 1000 IU IFN-gamma 4 h prior to inoculation of tumor cells. Administration of GdCl3, which is reported to deplete and block the function of Kupffer cells, 24 h prior to tumor cell inoculation resulted in a 5-fold tumor mass increase relative to control. Injection of anti-asiolo-GM1 antiserum, which eliminates the hepatic NK-cells, induced a 10-fold increase in tumor mass. These results indicate an important early antimetastatic function of hepatic NK-cells and KC and that presurgical administration of IFN-gamma may be important for eliminating circulating tumor cells and inhibiting development of residual tumors.

Topics: Animals; Antibodies; Carcinoma; Colonic Neoplasms; G(M1) Ganglioside; Gadolinium; Immune System; Injections, Intravenous; Interferon-gamma; Killer Cells, Natural; Kupffer Cells; Liver Neoplasms; Mesenteric Veins; Mice; Mice, Inbred BALB C; Mice, SCID; Neoplasm Transplantation; Tumor Cells, Cultured

Although the liver is a potent tumor cell killing organ it is frequently the site of lethal metastases often signifying the endstage for patients with colorectal cancers. Enhancing hepatic-associated immunity remains elusive until the interactions among hepatic nonparenchymal cells (NPC) are deciphered. We sought to modulate the cellular components of the hepatic immune system of mice with anti-NK and anti-T-cell-neutralizing antibodies in order to determine the cell type most efficacious in preventing liver metastasis.. Liver-derived murine colon adenocarcinoma (LD-MCA-38) cells were injected into the ileocolic vein (ICV) of immunocompetent and immunodeficient C57BL/6 mice. Mice were pretreated 1 day prior to tumor cell injection with one of three antibodies: anti-AsGM1, Anti-NK1.1, or Anti-Thy1.2. On Day 21 laparotomy was performed to determine the extent of hepatic tumor foci. The number of hepatic tumor foci was recorded and compared by the Wilcoxon rank sum test.. Mice pretreated with anti-AsGM1 or Anti-NK1.1 developed a massive increase in the number of hepatic tumor foci and decreased survival compared to the control treated mice. Pretreatment with anti-Thy1.2 antibody resulted in a significant decrease in the number of hepatic tumor foci. LD-MCA-38 tumor cells were unable to colonize the liver of C57BL/6 athymic nude mice; however, anti-AsGM1 antibody abolished this antimetastatic effect. There was no difference in the extent of hepatic metastasis and survival between immunodeficient C57BL/6 bg/bg and their conventional littermates bg/+.. AsGM1+ NK cells exhibit a significant antitumor response in the absence of T-cells. The concept of stimulating NK cell activity and suppressing T-cell function may enhance liver-associated immunity and serve as a deterrent for blood-borne tumor cells metastasizing to the liver.

Topics: Adenocarcinoma; Animals; Antibodies; Antibodies, Monoclonal; Antigens; Antigens, Ly; Antigens, Surface; Colonic Neoplasms; G(M1) Ganglioside; Immunocompetence; Killer Cells, Natural; Lectins, C-Type; Liver Neoplasms; Lymphocyte Count; Mice; Mice, Inbred BALB C; Mice, Inbred C57BL; Mice, Nude; Neoplasm Invasiveness; Neoplasm Transplantation; NK Cell Lectin-Like Receptor Subfamily B; Proteins; T-Lymphocytes; Thy-1 Antigens; Tumor Cells, Cultured

A novel peptide technology to produce mimicking peptides of carbohydrate moiety (which we propose to name glyco-replica peptides) is a useful tool to elucidate the functions of glycoconjugate. Carbohydrate moiety of ganglioside GD1alpha functions as a molecule involved in the adhesion between murine highly metastatic lymphoma RAW117-H10 cells and hepatic sinusoidal endothelial (HSE) cells. To prepare peptides which mimic the carbohydrate structure of GD1alpha, phage clones expressing peptides which bound to a monoclonal antibody against GD1alpha (KA17) were isolated from a phage-displayed random peptide library. Four phage clones having affinity to the monoclonal antibody KA17 were isolated, and these clones showed inhibitory effect on the binding of KA17 to GD1alpha. The amino acid sequences of the displayed pentadecamers were determined, and one of the phages displaying sequence WHWRHRIPLQLAAGR bound to HSE cells directly and showed the highest inhibitory effect on the adhesion between RAW117-H10 cells and HSE cells. The synthesized peptides having the same sequences to the displayed 15mers in the four isolated phage clones also showed the inhibitory effect on the adhesion of RAW117-H10 cells to HSE cells, and, again, the WHWRHRIPLQLAAGR peptide showed the highest inhibitory effect. Furthermore, intravenous injection of the peptide brought almost complete inhibition of the metastasis of RAW117-H10 cells to lung and spleen, and about 50% inhibition of the liver metastasis. These results indicate that GD1alpha plays an important role for metastasis of RAW117-H10 cells, and the peptides obtained by the present procedure are able to mimic the functional role of the glycoconjugate.

Topics: Amino Acid Sequence; Animals; Bacteriophages; Cell Adhesion; Cell Line; Epitopes; Female; G(M1) Ganglioside; Liver Neoplasms; Lung Neoplasms; Lymphoma; Mice; Mice, Inbred BALB C; Molecular Sequence Data; Neoplasm Metastasis; Peptide Library; Peptides; Splenic Neoplasms; Tumor Cells, Cultured

To elucidate the early events of blood-borne metastasis under actual blood flow, real-time trafficking of RAW117 large cell lymphoma cells, namely parental RAW117-P and liver-metastatic RAW117-H10 cells, was investigated using positron emission tomography (PET). Both types of cells accumulated in the liver immediately after injection via the portal vein, and were eliminated from the liver time-dependently. The elimination rate of RAW117-H10 cells, however, was slower than that of RAW117-P cells, suggesting that RAW117-H10 cells interact more strongly with hepatic sinusoidal endothelium than the parental cells. This result correlated with the metastatic potential of these cells: RAW117-H10 cells metastasized in the liver to a greater extent than RAW117-P cells after injection via this route. To investigate the role of sialylglycoconjugates in the interaction of RAW117-H10 cells with the hepatic endothelium after injection via the portal vein, the trafficking of RAW117-H10 cells was examined after the cells had been treated with sialidase. The elimination rate of RAW117-H10 cells from liver was observed to be greatly accelerated by sialidase treatment. To elucidate what kind of sialylglycoconjugates is related to this phenomenon, we analyzed the distribution of sialyl Lewis A and sialyl Lewis X antigens of both sublines of RAW117 by using flow cytometry. RAW117-H10 cells were found to express a much higher level of sialyl Lewis A than RAW117-P cells, whereas the amount of sialyl Lewis X did not differ significantly. These findings suggest that some sialylglycoconjugates, perhaps sialyl Lewis A in particular, play an important role in the initial interaction of RAW117-H10 cells with the hepatic endothelium, leading to metastasis.

Topics: Animals; Antigens, Neoplasm; CA-19-9 Antigen; Cell Adhesion; Cell Movement; Female; Flow Cytometry; G(M1) Ganglioside; Gangliosides; Injections, Intravenous; Liver Neoplasms; Lung Neoplasms; Lymphoma, Large B-Cell, Diffuse; Mice; Mice, Inbred BALB C; Neoplasm Metastasis; Neoplasm Transplantation; Neoplastic Cells, Circulating; Neuraminidase; Portal Vein; Sialyl Lewis X Antigen; Tomography, Emission-Computed; Tumor Cells, Cultured

Depletion of both natural killer 1.1+ (NK1+) intermediate alpha beta T-cell receptor (int T) cells and NK cells by in vivo treatment with anti-NK1 antibody greatly increased hepatic metastases of intravenously injected EL4 cells as well as pulmonary metastases of 3LL cells in C57BL/6 mice. However, depletion of NK cells alone by anti-asialo GM1 (AGM1) antibody treatment did not increase the metastases in either organ. Interleukin-12 (IL-12) administration into mice induced strong cytotoxicities of NK cell-depleted liver and lung mononuclear cells (MNC) comparable to those without NK-cell depletion and inhibited metastases in either organ. In contrast, in both NK cell- and NK1+ int T-cell-depleted mice, IL-12 could not induce cytotoxic activity of liver and lung MNC and metastases in both organs increased with or without IL-12 treatment. These results confirmed the fact that NK+ int T cells are more potent antitumour effectors than NK cells against experimental haematogenous tumour metastases.

Topics: Animals; Cytotoxicity, Immunologic; Flow Cytometry; G(M1) Ganglioside; Interleukin-12; Killer Cells, Natural; Leukocytes, Mononuclear; Liver Neoplasms; Lung Neoplasms; Lymphocyte Activation; Male; Mice; Mice, Inbred C57BL; Neoplasm Transplantation; T-Lymphocyte Subsets

Metastasis is a critical problem in the treatment of human lung cancer. Thus, a suitable animal model of metastasis of human lung cancer is required for in vivo biological and preclinical studies. In this study, we tried to establish a suitable model for this, using SCID mice. Neither human SCLC H69/VP cells (5 x 10(6)) nor squamous-cell carcinoma RERF-LC-AI cells (1 x 10(6)), injected through a tail vein, formed metastases in untreated SCID mice. Pre-treatment of SCID mice with anti-asialo GM1 serum resulted in only a few metastases of H69/VP cells, but pre-treatment with anti-mouse IL-2 receptor beta chain Ab (TM-beta 1) resulted in numerous lymph-node metastases 56 days after tumor inoculation. H69/VP-M cells, an in vivo-selected variant line, formed significant numbers of lymph-node metastases even in SCID mice pre-treated with anti-asialo GM1 serum. SCID mice depleted of NK cells by treatment with TM-beta 1 showed different patterns of metastasis when inoculated intravenously with the 2 different human lung cancer cell lines (H69/VP and RERF-LC-AI cells): H69/VP cells formed metastases mainly in systemic lymph nodes and the liver, whereas RERF-LC-AI cells formed metastases mainly in the liver and kidneys, with only a few in lymph nodes. A histopathological study showed that the metastatic colonies consisted of cancer cells. The numbers of metastatic colonies formed by the 2 cell lines increased with the number of cells inoculated. TM-beta 1 treatment of SCID mice efficiently removed NK cells from peripheral blood for at least 6 weeks, whereas, after treatment of the mice with anti-asialo GM1 serum, NK cells were recovered within 9 days. These findings suggest that NK-cell-depleted SCID mice may be useful as a model in biological and pre-clinical studies on metastasis of human lung cancer.

Topics: Animals; Antibodies; Disease Models, Animal; G(M1) Ganglioside; Humans; Immune Sera; Killer Cells, Natural; Liver Neoplasms; Lung Neoplasms; Lymphatic Metastasis; Male; Mice; Mice, SCID; Neoplasm Metastasis; Receptors, Interleukin-2; Tumor Cells, Cultured

Colorectal cancer frequently disseminates through the portal vein into the liver. In this study, outbred Swiss nude mice were adapted to facilitate the induction of liver metastases by a pre-grafting treatment with 6 Gy total body irradiation and i.v. injection of anti-asialo GM1 antibody. One day later, cultured LS 174T human colon cancer cells were injected into the surgically exposed spleen, which was resected 3 min later. In 48 of 65 mice, a few to several hundred liver metastases were macroscopically observed at dissection 3 to 4 weeks after transplantation. Ten of 10 mice, followed-up for survival, died with multiple large confluent liver metastases. By reducing the radiation dose to 4 or 0 Gy, or omitting the anti-asialo GM1 antibody injection, only 60%, 37% or 50% of mice, respectively, had visible metastases 3 weeks after transplantation. Carcinoembryonic antigen (CEA) measured in tumour extracts was in the mean 25.6 micrograms/g in liver metastases compared with 9.2 micrograms/g in s.c. tumours. Uptake of radiolabelled anti-CEA monoclonal antibody (MAb) in the metastases 12, 24 and 48 hr after injection gave a mean value of 39% of the injected dose per gram of tissue (ID/g). In comparison, MAb uptake in s.c. and intrasplenic tumours or lung metastases gave a mean percentage ID/g of 20, 18 and 15, respectively. Laser-induced fluorescence after injection of indocyanin-MAb conjugate allowed direct visual detection of small liver metastases, including some that were not visible under normal light. Preliminary results showed that mice, pre-treated with 4 Gy irradiation and the anti-asialo GM1 injection, were tolerant to radioimmunotherapy with a total dose of 500 muCi 131I labeled anti-CEA intact MAbs given in 3 injections.

Topics: Animals; Antibodies; Antibodies, Monoclonal; Carbocyanines; Carcinoembryonic Antigen; Colonic Neoplasms; Fluorescence; Fluorescent Dyes; G(M1) Ganglioside; Humans; Immunohistochemistry; Lasers; Liver Neoplasms; Mice; Mice, Nude; Neoplasm Transplantation; Radioimmunotherapy; Tumor Cells, Cultured; Whole-Body Irradiation

We have studied the neutral glycolipid composition of spontaneous hepatomas in LEC female rats. Neutral lipid fractions were isolated and purified by column chromatographies on DEAE-Toyopearl 650(M) and Iatrobeads. The neutral glycolipid fraction contained 3.2 to 4.4 micrograms lipid-bound glucose (Glc) per mg protein, and consisted of isogloboside (iso-Gb4, 50.8% of total neutral glycolipids) and IV3Gal, IV2Fuc, GgOse4Cer (asialo-BGM1, 13.5%) as the major neutral glycolipids and Gb3 and iso-Gb3 (9.2%), GlcCer (7.2%), LacCer (6.1%) as the other species. The structure of iso-Gb4 was elucidated by gas-liquid chromatography (GLC), permethylation study, liquid secondary ion (LSI) mass spectrometry, and nuclear Overhauser enhancement spectroscopy (NOESY) and that for asialo-BGM1 by GLC, LSI mass spectrometry, and high-performance thin-layer chromatography (HPTLC)-overlay method using anti-asialo-BGM1 antibody. Isogloboside and asialo-BGM1 which are found in negligible amounts in normal liver tissues may represent excellent markers for studying tumor metastasis and cellular adhesion.

Topics: ABO Blood-Group System; Animals; Biomarkers, Tumor; Carbohydrate Sequence; Carcinoma, Hepatocellular; Epitopes; Female; G(M1) Ganglioside; Globosides; Glycolipids; Liver Neoplasms; Male; Molecular Sequence Data; Rats

We have previously demonstrated that administration of killed streptococcal preparation (OK432), a biological modifier, increased the number of asialo GM1-positive cells in the liver, enhanced NK activity of hepatic mononuclear cells, and reduced the number of hepatic metastases of colon 38 adenocarcinoma that were inoculated into the superior mesenteric vein of C57BL/6 strain mice. In the present study, to clarify the role of the spleen in immune surveillance of the liver, the effect of splenectomy on hepatic metastasis of colon carcinoma and on hepatic NK activity has been examined. The number of hepatic metastasis increased in the splenectomized mice, compared with that in sham-operated mice. Administration of OK432 increased the number of asialo GM1-positive cells in the liver and enhanced NK activity of hepatic mononuclear cells in both groups, but NK activity of hepatic mononuclear cells in the splenectomized mice was less than that of the sham-operated mice. An enhanced NK activity of these cells was abolished by treatment with anti-asialo-GM1 antibody plus complement in vitro. Interleukin-2 mRNA expression was increased in the spleen 2 hr after OK432 administration and persisted until 8 hr, but was scarcely noted in the liver. On the other hand, NK activity of hepatic mononuclear cells in the asialo GM1-positive cell-depleted (previous administration of antiserum against asialo GM1) mice was enhanced after OK432 administration in the sham operated and splenectomized mice, but an enhanced NK activity in these mice was only partially or not at all abolished by treatment with anti-asialo GM1 antibody plus complement in vitro, respectively.(ABSTRACT TRUNCATED AT 250 WORDS)

Topics: Animals; Colonic Neoplasms; G(M1) Ganglioside; Interleukin-2; Killer Cells, Natural; Liver; Liver Neoplasms; Male; Mice; Mice, Inbred C57BL; Picibanil; RNA, Messenger; Spleen; Splenectomy

An experimental model for hepatic metastasis with a transplantation route of free-pedicled subcutaneous-embedded spleen was established in BALB/c mice. Colon-26 tumor cells to produce hepatic metastasis were inoculated into the spleen and the influence of surgical stress by means of a 20-min exposure of the abdominal cavity on the incidence of hepatic metastasis was examined. Hepatic metastasis was more promoted by the surgical stress in order when it was given on the same day, the 7th day and the 3rd day of the inoculation. Administration, without surgical stress, of ASGM 1, a specific inhibitor of the natural killer activity, also facilitated the hepatic disease. Administration of OK-432 prior to the surgical stress or ASGM 1 was at least partly effective for prevention of the hepatic metastasis and prolonged the survival of the inoculated mice. Preoperative immunotherapy utilizing OK-432 might be a possible means to prevent hepatic metastasis triggered in colorectal surgery for cancer.

Topics: Animals; Antibodies; Antineoplastic Agents; Colonic Neoplasms; Cytotoxicity, Immunologic; Disease Models, Animal; G(M1) Ganglioside; Killer Cells, Natural; Liver Neoplasms; Male; Mice; Mice, Inbred BALB C; Neoplasm Transplantation; Neoplastic Cells, Circulating; Picibanil; Stress, Physiological; Surgical Procedures, Operative

The ganglioside fucosyl-GM1 (FucGM1) has been suggested as a marker for small-cell lung cancer (SCLC). Immunohistochemical analyses have shown the expression of the ganglioside in tumors in 75 to 90% of patients with SCLC. We have demonstrated that the ganglioside is shedded from SCLC cells both in vitro and in vivo, and that the antigen can be detected in sera from SCLC patients by an immunochemical analysis. The FucGM1 antigen has recently been shown to act as a target for antibody-dependent cellular cytotoxicity. This may provide a rationale for developing immunotherapy against SCLC. We used an immunoassay based on the scintillation proximity assay to analyze the concentrations of FucGM1 in sera from 112 SCLC patients, 21 patients with non-SCLC, 4 patients with other cancer forms, and 20 healthy controls. Sera were collected at the time of diagnosis before initiation of chemotherapy. The expression of FucGM1 was related to age, sex, blood group of the patient, and to the stage of disease and organ site involvement of metastases. The sera of 50% of the patients with SCLC were positive for FucGM1, and 12 of 21 sera from non-SCLC patients were markedly elevated. In SCLC sera, the concentration of FucGM1 in positive sera ranged from 7 to more than 3000 ng/ml FucGM1. None of 20 controls were positive. FucGM1 correlated to organ site involvement of metastases (p = 0.0016). The ganglioside was detected both at significantly higher concentrations (p = 0.0005) and in significantly more patients (p = 0.0026) with metastases to both the liver and bone marrow, compared to patients with metastases to the liver only.(ABSTRACT TRUNCATED AT 250 WORDS)

Topics: Biomarkers, Tumor; Bone Marrow; Carcinoma, Small Cell; Female; Follow-Up Studies; G(M1) Ganglioside; Humans; Liver Neoplasms; Lung Neoplasms; Male; Middle Aged; Survival Rate

A 76-yr-old male patient with carcinomatous neuropathy associated with hepatic cell carcinoma, whose initial symptom was deep sensory disturbance followed by muscle weakness is described. The onset was subacute, followed by slow progression. Sural nerve biopsy, as well as electrophysiological examinations, revealed severe axonal degeneration without any evidence of demyelination. The autopsy findings were similar to findings described in the literature on carcinomatous neuropathy. Although carcinomatous neuropathy is usually associated with lung cancer, this report describes an association with hepatic cell carcinoma. The patient also had motor nerve involvement with positive serum anti-GM1 ganglioside antibody which decreased after immunosuppressant therapy in parallel with recovery of muscle weakness. The anti-GM1 ganglioside antibody may be involved in the pathogenesis of motor disturbance in the present case.

Topics: Aged; Antibodies; Carcinoma, Hepatocellular; G(M1) Ganglioside; Humans; Immunosuppressive Agents; Liver Neoplasms; Male; Nerve Degeneration; Neural Conduction; Neurons, Afferent; Peripheral Nervous System Diseases

This investigation aimed to develop a biologically relevant murine model of colorectal liver metastases and determine if Kupffer cells (KC) and hepatic natural killer cells (hNKC) regulate tumor growth. The model involves the injection of murine colon adenocarcinoma 26 (MCA 26) tumor cells into the portal vein of female-specific pathogen-free BALB/c mice. Metastases developed in all animals, and the growth was limited entirely to the liver. To determine if KC and hNKC control the development of liver metastases, the in vivo function of these hepatic effector cells was modulated. Tumor growth was quantitated by the uptake of 125I into tumor DNA. Stimulation of the KC and hNKC produced a significant (P less than 0.01) dose-dependent decrease in 125I uptake in the liver in both treatment groups, which was associated with a significant improvement in survival (P less than 0.05). The in vivo cytotoxic function of the liver was inhibited with an intravenous injection of gadolinium chloride (for KC) or asialo GM1 antiserum (for hNKC). Inhibition of KC and hNKC cytotoxic function led to a significant (P less than 0.01) increase in 125I uptake in the liver and a significant decrease in survival (P less than 0.05).

Topics: Adenocarcinoma; Animals; Antineoplastic Agents; Cell Division; Cell Line; Cell Survival; Colonic Neoplasms; Cytosine; Cytotoxicity, Immunologic; Female; G(M1) Ganglioside; Gadolinium; Immune Sera; Killer Cells, Natural; Kupffer Cells; Liver Neoplasms; Mice; Mice, Inbred BALB C; Propionibacterium acnes; Rectal Neoplasms

We have investigated the effect of recombinant polypeptides with cell-binding domain (C-274) or with heparin-binding domain (H-271) and their fusion polypeptide (CH-271) on liver metastasis of murine lymphoid tumor. The polypeptides containing heparin-binding domain, H-271 and CH-271, were able to inhibit liver metastasis when co-injected i.v. with L5178Y-ML25 T-lymphoma cells, while C-274 with cell-binding domain showed much weaker antimetastatic activity. Treatment with H-271 or CH-271 substantially prolonged the survival time of mice injected i.v. with L5178Y-ML25 cells. CH-271, containing cell- and heparin-binding domains, was more antimetastatic than H-271. The reason why CH-271 was more effective in inhibiting liver metastasis than H-271 can not be explained in terms of a difference in the stability in the circulation or in the molecular size of the polypeptide. The polypeptides used in this study did not affect the tumor cell growth or viability in vitro. CH-271 was found to be still active in inhibiting liver metastasis even when natural killer cells or macrophages were removed from this system. Furthermore, multiple administrations of CH-271 after tumor implantation effectively inhibited liver metastasis and enhanced the survival rate as compared with H-271, C-274 and untreated control. Thus, the fusion of H-271 with C-274 (i.e. CH-271) augments the antimetastatic property of H-271, possibly through the interaction between tumor cells and the heparin-binding domain of fibronectin.

Topics: 2-Chloroadenosine; Animals; Binding Sites; Cell Adhesion; Fibronectins; G(M1) Ganglioside; Glycosphingolipids; Heparin; Leukemia L5178; Liver Neoplasms; Mice; Neoplasm Metastasis; Recombinant Fusion Proteins; Structure-Activity Relationship; Survival Analysis

The ability of the host-immune defense mechanism of nude mice and their immunocompetent littermates to prevent liver metastases from the murine colon carcinoma, colon-26, was assessed. Give thousand tumor cells suspended in 0.05 ml of Hank's balanced salt solution were inoculated into the spleens of BALB/c nu/+ and BALB/c nu/nu mice. On the 21st day after inoculation, all the mice were sacrificed, and the liver metastases counted and the livers weighed. All the BALB/c nu/+ mice were found to have developed hepatic metastases with a mean of 10 nodules, whereas no hepatic metastases were observed in any of the 10 BALB/c nude mice. On the other hand, 4 of 6 nude mice developed hepatic metastases after treatment with anti-asialo GM1 antibody. These results indicate that the BALB/c nude mouse has an excellent host-immune defense mechanism for preventing liver metastasis, with NK cells in the liver and/or blood circulation perhaps playing an important role.

Topics: Animals; Antibodies; Colonic Neoplasms; Evaluation Studies as Topic; G(M1) Ganglioside; Glycosphingolipids; Immunity, Innate; Killer Cells, Natural; Liver Neoplasms; Male; Mice; Mice, Inbred BALB C; Mice, Nude; Time Factors; Transplantation, Homologous

This investigation was undertaken in order to assess both the preventive and antiproliferative effects of tumor necrosis factor (TNF) in a hepatic metastasis model, by means of inoculation of mouse colon-26 tumor cells into the portal vein via the superior mesenteric vein in male CDF1 mice, aged 5 weeks. Continuous 10-day administration of natural human TNF-alpha (nHuTNF-alpha) following the tumor cell inoculation caused no reduction but rather an increase in the number of hepatic metastases. However, pretreatment with this preparation daily for 10 days before the inoculation caused a remarkable decrease in the number of hepatic metastases. This prophylactic effect was reversed by the intravenous administration of anti-asialo GM1 antibody 24 h before the inoculation. The result of immunoperoxidase staining of liver specimens suggested that organ-associated natural killer cells might play a role in the metastatic inhibition. An apparent antiproliferative effect on metastatic liver tumors was also recognized following injection of nHuTNF-alpha from the 10th day after the inoculation. Thus, TNF appears to have important effects upon the host immune system, acting against liver metastases.

Topics: Animals; Antigen-Antibody Reactions; Cell Division; Colonic Neoplasms; G(M1) Ganglioside; Glycosphingolipids; Immunoenzyme Techniques; Liver Neoplasms; Mice; Neoplasm Metastasis; Tumor Necrosis Factor-alpha

The administration of anti-B16 monoclonal antibody of the IgG2b isotype to mice bearing established B16 melanoma liver metastases caused a significant and consistent reduction of up to 90% in the number of these metastases. No reduction in the number of metastases was noted when antigenically unrelated tumor or nonspecific immunoglobulin were employed. The antibody-mediated antitumor effect was completely abrogated by total body irradiation of the host. Treatment of the tumor-bearing host with antiserum directed against asialo GM1 prior to anti B16 antibody administration, abrogated the therapeutic effect indicating the involvement of a radiosensitive, ASGM1-positive cell in the tumor regression. The antitumor effect of the antibody treatment could be augmented by the concomitant administration of recombinant interleukin-2. The effect seen may have possible application in the treatment of liver metastases in humans by combined immunotherapy using recombinant interleukin-2 and specific antitumor monoclonal antibodies.

Topics: Animals; Antibodies, Monoclonal; Antibody Specificity; Dose-Response Relationship, Immunologic; G(M1) Ganglioside; Glycosphingolipids; Immunity; Immunotherapy; Interleukin-2; Liver Neoplasms; Melanoma, Experimental; Mice; Recombinant Proteins

Mice treated with anti-asialo GM1 (asGM1) serum exhibited increased formation of experimental metastases in lung and liver after i.v. challenge with B16 melanoma or Lewis lung carcinoma. This increased metastasis formation coincided with decreased splenic NK activity and increased survival of i.v. injected radiolabeled tumor cells. In contrast, the injection of mice with the pyran copolymer maleic anhydride divinyl ether (MVE-2) augmented NK activity in the spleen and significantly depressed the formation of experimental metastases in the lungs and liver. However, a single or double administration of anti-asGM1 antiserum to MVE-2-pretreated mice failed to inhibit the immunoprophylaxis associated with MVE-2 administration, although it did decrease splenic NK activity and also increased the survival of i.v.-injected radiolabeled tumor cells. To address the mechanism for this dichotomy, we examined NK activity not only in the spleen but also in the blood, lungs, and livers of MVE-2-treated mice. Levels of NK activity in the lungs and liver were several-fold higher than those observed in spleen and blood. However, MVE-2-augmented NK activity in lung and liver was more resistant to depletion by the standard regimen of anti-asGM1 treatment than was NK activity in blood and spleen, and required two high-dose administrations of a higher titered antiserum for depletion of the augmented response. This high-dose regimen removed all detectable NK activity from the lung and liver, and concomitantly eliminated the metastasis-inhibiting effect of MVE-2. These data are consistent with a role for organ-associated NK cells in inhibiting metastasis formation during the extravasation and/or early postextravasation phases of the metastatic process. The results also suggest that biologic effects of NK activity in spleen and blood can be dissociated from those mediated by NK activity in other organs by use of different treatment regimens with anti-asGM1 serum. Finally, because NK activity in target organs can be augmented to an even greater extent than in the blood and spleen by at least some biologic response modifiers (BRMs), organ-associated NK activity should be considered as a possible mechanism for the therapeutic effects of BRM treatment.

Topics: Animals; Antineoplastic Agents; Cytotoxicity, Immunologic; Female; G(M1) Ganglioside; Glycosphingolipids; Growth Substances; Immune Sera; Killer Cells, Natural; Liver Neoplasms; Lung Neoplasms; Lymphoma; Male; Melanoma; Mice; Mice, Inbred C3H; Mice, Inbred C57BL; Organ Specificity; Pyran Copolymer

We have studied the role of natural killer activity during the growth and dissemination of a transplantable renal adenocarcinoma (Renca) of spontaneous origin in BALB/c mice. The pattern of growth of this tumor accurately mimics that of adult human renal cell carcinoma in terms of clinical stages I-IV, particularly with regard to spontaneous metastasis to lung and liver. Renca is moderately sensitive to lysis by natural killer cells from normal mice and is more efficiently lysed by natural killer cells from mice treated with the biological response modifier maleic anhydride divinyl ether, a pyran copolymer. Our studies demonstrate that selective depression of natural killer activity by administration of antiserum specific for the neutral glycosphingolipid asialo GM1 correlated with increased formation of spontaneous metastases in the lungs, liver, and lymph nodes. Conversely, augmentation of natural killer activity by the biological response modifier decreased the formation of spontaneous metastases in lungs, liver and lymph nodes. Further, the suppression of natural killer activity and subsequent increased formation of metastases were accompanied by a significantly reduced survival time, whereas the augmented natural killer activity and decreased incidence of metastases in biological response modifier-treated mice were accompanied by an increase in time of survival. These results demonstrate a significant role for natural killer cells in the control of spontaneous metastasis during growth of this murine renal cancer.

Topics: Animals; Carcinoma, Renal Cell; Cell Line; Female; G(M1) Ganglioside; Glycosphingolipids; Immune Sera; Kidney Neoplasms; Killer Cells, Natural; Liver Neoplasms; Lung Neoplasms; Male; Mice; Mice, Inbred BALB C; Neoplasm Metastasis; Neoplasm Transplantation; Pyran Copolymer

The mechanism of artificial and spontaneous metastases of tumor was analyzed in B16 melanoma cells and C57BL/6 mice by using anti-asialo GM1 antibody and anticancer agents. Single administrations of 500 micrograms anti-asialo GM1 antibody resulted in significantly decreased NK activity in spleen cells of C57BL/6 mice, lasting 10 days from the day following administration. Treatment with anti-asialo GM1 antibody never decreased the function of T lymphocytes measured by blastogenesis with phytohemagglutinin or T cell growth factor. The tumoricidal functions of activated macrophages but not of resident macrophages were decreased by in vivo treatment with anti-asialo GM1 antibody. The anti-asialo GM1 antibody was evaluated in terms of the enhancing effect on pulmonary metastases with regard to the timing of administration. Treatment with anti-asialo GM1 antibody 1 day before or on the day of tumor inoculation resulted in a substantial increase in the number of artificial pulmonary metastases. In the experimental system of spontaneous metastases, anti-asialo GM1 antibody most effectively increased the number of pulmonary metastases when administered 1-2 weeks before the removal of primary tumor, when the tumor cells are thought to be released into blood circulation from the primary site. In addition, accelerated growth of transplanted tumors at the primary site was observed in mice treated with anti-asialo GM1 antibody. These results strongly suggest that anti-asialo GM1 antibody enhances the incidence of in vivo tumor metastases and the growth of transplanted tumor mainly by suppressing the function of NK cells. The maximum effective dose (MED) of mitomycin C or its derivative (M-83) suppressed NK activity significantly, and pretreatment with these anticancer agents enhanced the growth of the artificial pulmonary and liver metastases. In contrast, the MED of cDDP showed no effect on the NK activity or the numbers of pulmonary and liver metastases. These results indicate that the depression of NK activity induced by chemotherapy results in the promotion of metastatic disease. From these studies it can be concluded that NK cells have a key role in the control of metastases of malignant disease, and that support of NK activity is very important for the prevention of metastases.

Topics: Animals; Antibodies; Antineoplastic Agents; Cytotoxicity, Immunologic; G(M1) Ganglioside; Gangliosides; Killer Cells, Natural; Liver Neoplasms; Lung Neoplasms; Lymphocyte Activation; Macrophages; Male; Mice; Mice, Inbred C57BL; Neoplasm Metastasis; Neoplasms, Experimental

Hyperplastic nodules and hepatocellular carcinomas were induced in livers of rats by a low-protein diet containing 0.05% of the carcinogen N-2-fluorenylacetamide. Ganglioside amounts and composition were determined for histologically different hepatocellular carcinomas and compared with those for control livers, hyperplastic nodules, and liver tissue surrounding hepatomas and nodules as well as those for livers of fetal, newborn, 1-week-old, weanling, and adult Sprague-Dawley rats. Ganglioside sialic acid levels were elevated above those of normal adult liver in all liver tissues following the carcinogen treatment regimen. Livers of fetal and newborn rats contained nearly twice the amount of ganglioside sialic acid on a protein or DNA basis as did livers of adult rats. Analyses of individual nodules and hepatomas revealed two populations of tumors in which the levels of ganglioside sialic acid were 2.3 and 3.8 times normal. Ganglioside sialic acid content was at hepatoma levels in small nodules. Individual gangliosides were evenly distributed between products of the monosialoganglioside and disialoganglioside pathways in normal liver with a ratio of [N-acetylneuraminic acid (sialic acid)] (NAN)-galactose (Gal)-N-acetylgalactosamine (GalNAc)-(NAN)-Gal-glucose (Glc)-ceramide (Cer) (GD1a) to Gal-GalNAc-(NAN)2-Gal-Glc-Cer (GD1b) of about one. In contrast, the monosialogangliosides predominated in liver tissues following administration of the carcinogen. Increased levels of specific monosialogangliosides were present in nodules, in liver of carcinogen-treated animals prior to the appearance of tumors, and in the liver tissues surrounding nodules and hepatomas. In single hepatomas, ganglioside patterns correlated with tumorigenicity. A well-differentiated hepatoma had a normal complement of most gangliosides but was deficient in trisialogangliosides. In a poorly diferentiated but well-circumscribed hepatoma, the relative levels of all higher gangliosides were reduced. The monosialoganglioside Gal-GalNAc-(NAN)-Gal-Glc-Cer (GM1) accounted for 80% of the total ganglioside in a poorly circumscribed and poorly differentiated hepatoma. The ganglioside pattern of fetal livers most closely resembled that of a poorly differentiated hepatoma. During the first week post natum, levels of all higher monosialogangliosides and disialogangliosides declined, but the decline was most pronounced for gangliosides GM1 and GD1a. The ratio of GM1 + GD1a to GD1b + NAN-Gal-GalNAc-(NAN)

Topics: 2-Acetylaminofluorene; Animals; Animals, Newborn; Carcinoma, Hepatocellular; Fluorenes; G(M1) Ganglioside; G(M2) Ganglioside; G(M3) Ganglioside; Gangliosides; Hyperplasia; Liver; Liver Neoplasms; Neoplasms, Experimental; Precancerous Conditions; Rats; Sialic Acids