g(m1)-ganglioside and Lymphoma--B-Cell

The variable regions of the immunoglobulin (Ig) expressed on the surface of a malignant B cell can be considered tumor-specific antigens and, as such, could be targets for immunotherapeutic approaches. However, because until now the immunization procedures have been complex and have given only partial protection, it was necessary to find new methods of immunotherapy. Here, we present a successful method of vaccination against B-cell tumors in a murine model. We produced recombinant vaccinia viruses (rVV) expressing the heavy and the light chain of surface Ig of a patient's malignant B cells and we tested the ability of these rVV to protect immunized mice against tumor growth of transfectomas producing the same human Ig. The protection of the mice was complete and specific to the variable region of the immunizing heavy chain although specific lymphoproliferative and cytotoxic responses were not detectable in vitro. The protection was strictly dependent on the presence of CD4 T cells and asialo GM1+ cells. Furthermore, tumor protection clearly required gamma-interferon and was partially inhibited by blocking the Fas-Fas ligand interaction. We also show, in a murine syngeneic model, that rVV expressing a poorly mutated Ig protects against the growth of Ig-producing tumor.

Topics: Amino Acid Sequence; Animals; Base Sequence; CD4-Positive T-Lymphocytes; Cricetinae; fas Receptor; Female; Flow Cytometry; G(M1) Ganglioside; Genetic Vectors; Humans; Immunoglobulin Idiotypes; Immunoglobulin Variable Region; Interferon-gamma; Lymphoma, B-Cell; Mice; Mice, Inbred BALB C; Molecular Sequence Data; Neoplasms, Experimental; Recombinant Proteins; T-Lymphocytes, Cytotoxic; Vaccination; Vaccinia virus

The B cell antigen receptor (BCR) serves both to initiate signal transduction cascades and to target antigen for processing and presentation by MHC class II molecules. How these two BCR functions are coordinated is not known. Recently, sphingolipid- and cholesterol-rich plasma membrane lipid microdomains, termed lipid rafts, have been identified and proposed to function as platforms for both receptor signaling and membrane trafficking. Here we show that upon cross-linking, the BCR rapidly translocates into ganglioside G(M1)-enriched lipid rafts that contain the Src family kinase Lyn and exclude the phosphatase CD45R. Both Igalpha and Lyn in the lipid rafts become phosphorylated, and subsequently the BCR and a portion of G(M1) are targeted to the class II peptide loading compartment. Entry into lipid rafts, however, is not sufficient for targeting to the antigen processing compartments, as a mutant surface Ig containing a deletion of the cytoplasmic domain is constitutively present in rafts but when cross-linked does not internalize to the antigen processing compartment. Taken together, these results provide evidence for a role for lipid rafts in the initial steps of BCR signaling and antigen targeting.

Topics: Animals; Cholesterol; Cross-Linking Reagents; G(M1) Ganglioside; Histocompatibility Antigens Class II; Horseradish Peroxidase; Leukocyte Common Antigens; Lymphoma, B-Cell; Membrane Lipids; Mice; Mutagenesis; Receptors, Antigen, B-Cell; Receptors, Antigen, T-Cell; Recombinant Proteins; Sequence Deletion; Signal Transduction; Sphingolipids; src Homology Domains; Tumor Cells, Cultured

Intraperitoneal injection of lymphoid cells from EBV+ donors into SCID mice might provide a useful tool for studying the pathways of B-cell lymphomagenesis in man. Since previous studies showed that donor T cells greatly favor B-cell proliferation and tumor generation in this model, we addressed the host and donor factors involved in limiting or promoting lymphoma development. The number of EBV-infected B-cell precursors was crucial, since purified B lymphocytes, which alone were unable to generate tumors, underwent expansion and established tumor masses when the animals were inoculated with an EBV-containing supernatant. Host factors were critical in limiting tumor development; in vivo NK-cell removal allowed purified B cells to expand and proceed to tumors in the absence of T lymphocytes, whereas potentiation of mouse NK-cell activity prevented tumor generation in PBMC- and LCL-injected animals. The T-cell-derived factors that favor lymphomagenesis could not be identified; IL-2, IL-4, IL-6, and soluble CD23 were not able to promote B-cell expansion, and treatment of PBMC-injected mice with the relevant anti-cytokine anti-sera did not counteract lymphoma development. These experiments also showed that IL-6 plays a minor role, if any, in B-cell lymphoproliferation in this model. Our data indicate that reconstitution of SCID mice with PBMC from EBV+ donors may constitute a useful model for determining the events involved in lymphomagenesis in humans, provided that strict control of all the experimental variables is guaranteed.

Topics: Animals; Antibodies; B-Lymphocytes; Base Sequence; Cell Separation; DNA, Viral; G(M1) Ganglioside; Gene Rearrangement; Herpesvirus 4, Human; Humans; Interleukin-6; Killer Cells, Natural; Leukocytes, Mononuclear; Lymphoma, B-Cell; Lymphoproliferative Disorders; Mice; Mice, SCID; Molecular Sequence Data; Polymerase Chain Reaction

Lethally irradiated C3H/HeN mice reconstituted with normal syngeneic bone marrow survived significantly longer than unmanipulated control mice following challenge with a lethal dose of 38C13 lymphoma cells 2 to 3 weeks post-bone marrow transplantation (BMT). Although the magnitude of this effect was modest, it was highly reproducible. This resistance-producing effect of BMT could be enhanced by interleukin 2 administration and could be abrogated by anti-asialo-GM1 antiserum treatment of recipients. These findings are consistent with the hypothesis that cells with a natural killer phenotype are activated by BMT and can mediate tumor resistance. These studies provide a model to explore the cellular basis, independent of donor alloreactivity, of the graft antitumor effect of BMT observed in humans.

Topics: Animals; Bone Marrow Transplantation; Female; G(M1) Ganglioside; Immunity, Innate; Immunization, Passive; Immunoglobulins; Interleukin-2; Killer Cells, Natural; Lymphocyte Activation; Lymphoma, B-Cell; Mice; Mice, Inbred C3H; Transplantation, Isogeneic