ganglioside--gd2 and Lymphoma--T-Cell

Monoclonal antibodies ME361 specific to ganglioside GD2 were isolated from the conditioned medium of hybridoma HB9326 and mouse ascitic fluid by the method of affinity chromatography; their Fab-fragments were obtained by proteolytic cleavage with papain. Evaluation of Fab-fragment specificity by flow cytometry and dot-blot analysis showed that binding effectiveness of fragments with antigens was close to that for the full-length molecule of antigen. It was shown that Fab-fragments and whole antibodies ME361 dose-dependently inhibit the proliferation of cells of mice T-lymphoma EL-4, and induce apoptosis of these cells 24 h after incubation.

Topics: Animals; Antibodies, Monoclonal; Apoptosis; Ascitic Fluid; Cell Line, Tumor; Cell Proliferation; Gangliosides; Hybridomas; Immunoglobulin Fab Fragments; Lymphoma, T-Cell; Mice

Immunocytokines (ICKs) targeting cytokines to the tumor environment using antibodies directed against a tumor-associated antigen often have a higher therapeutic index than the corresponding unconjugated cytokines. Various ICKs displaying significant antitumoral effects in several murine tumor models have already been developed, and some of them, in particular interleukin (IL)-2-based ICKs, are in Phase II clinical trials. Although sharing common biological activities with IL-2 in vitro, IL-15 is now considered as having a better potential in antitumor immunotherapeutical strategies and has been shown to be less toxic than IL-2 in preclinical studies. We previously developed the fusion protein RLI, linking a soluble form of human IL-15Rα-sushi+ domain to human IL-15. RLI showed better biological activities than IL-15 in vitro as well as higher antitumoral effects in vivo in murine and human cancer models. Here, we investigated, in the context of an ICK, the effect of associating RLI with an antibody targeting the GD2 ganglioside, a validated tumoral target expressed on many neurectodermal tumors. Anti-GD2-RLI fully retained the cytokine potential of RLI and the antibody effector functions (antibody-dependent cellular cytotoxicity and complement-dependent cytotoxicity). It displayed strong antitumor activities in two syngeneic cancer models in immunocompetent mice (subcutaneous EL4 and metastatic NXS2). Its therapeutic potency was higher than those of RLI and anti-GD2 alone or in combination. We suggest that this is related to its bifunctional (cytokine and antibody) nature.

Topics: Animals; Antibodies, Monoclonal; Antibodies, Neoplasm; Antibody-Dependent Cell Cytotoxicity; Antigens, Neoplasm; Cell Line, Tumor; Cell Proliferation; Female; Gangliosides; Humans; Immunotherapy; Interleukin-15; Interleukin-15 Receptor alpha Subunit; Liver Neoplasms; Lymphoma, T-Cell; Male; Mice; Mice, Inbred A; Mice, Inbred C57BL; Neoplasm Metastasis; Neuroblastoma; Protein Binding; Recombinant Fusion Proteins

IgM antibodies to gangliosides, sialic acid-containing glycosphingolipids, have been shown to mediate anti-tumor effects in cancer patients with melanoma and neuroblastoma and to correlate with survival. Mechanisms by which the antibodies induce tumor suppression, however, have not been systematically studied. To investigate this point, we produced and characterized C57BL/6 mice transgenic for IgM antibody to ganglioside GD2. The transgenic (TG) mice showed high IgM, but not IgG antibody titers against GD2 in their sera. No significant clinical symptoms were observed. When EL4 cells, syngeneic T lymphoma that express ganglioside GD2, were injected into TG mice, prolonged survival was observed. Complement-dependent cytotoxicity (CDC) of EL4 cells was mediated with TG mice sera. Neither antibody-dependent cellular cytotoxicity with their sera nor cytotoxic T lymphocyte activity to EL4 cells was shown in TG mice. Spleen lymphocytes from TG mice had increased numbers of natural killer (NK) cells, but not T cells, B cells, or macrophages compared with wild-type mice. Depletion of NK cells with anti-asialo GM1 rabbit serum reduced or abrogated the observed anti-tumor effects, suggesting that NK cells play a major role in tumor eradication or suppression. NK cell activity in TG mice was much higher than wild-type mice. Moreover, TG mice showed prolonged survival after injection with syngeneic B16 melanoma cells, which express GM3, but not GD2 or GD3. Taking these results together, our studies demonstrate that the TG mice have significant anti-tumor characteristics, probably due to CDC and NK cell expansion and activation with anti-ganglioside GD2 antibody.

Topics: Animals; Antibodies, Monoclonal; Antibody-Dependent Cell Cytotoxicity; Complement System Proteins; Female; Gangliosides; Humans; Immunoglobulin M; Interferon-gamma; Killer Cells, Natural; Lymphocyte Depletion; Lymphoma, T-Cell; Macrophages; Melanoma, Experimental; Mice; Mice, Inbred C57BL; Mice, Transgenic; Spleen; Survival Rate; T-Lymphocytes, Cytotoxic; Tumor Cells, Cultured