ganglioside--gd2 and Lymphoma

Four randomized phase III trials conducted recently in melanoma patients in the adjuvant setting have been based in part on the correlation between antibody responses in immunized patients and improved survival. Each of these randomized trials demonstrated no clinical benefit, although again there was a significant correlation between antibody response after vaccination and disease free and overall survival. To better understand this paradox, we established a surgical adjuvant model targeting GD2 ganglioside on EL4 lymphoma cells injected into the foot pad followed by amputation at variable intervals. Our findings are (1) comparable strong therapeutic benefit resulted from treatment of mice after amputation with a GD2-KLH conjugate vaccine or with anti-GD2 monoclonal antibody 3F8. (2) The strongest correlation was between antibody induction in response to vaccination and prolonged survival. (3) Antibody titers in response to vaccination in tumor challenged mice as compared to unchallenged mice were far lower despite the absence of detectable recurrences at the time. (4) The half life of administered 3F8 monoclonal antibody (but not control antibody) in challenged mice administered was significantly shorter than the half life of 3F8 antibody in unchallenged controls. The correlation between vaccine-induced antibody titers and prolonged survival may reflect, at least in part, increased tumor burden in antibody-negative mice. Absorption of vaccine-induced antibodies by increased, although not detected tumor burden may also explain the correlation between vaccine-induced antibody titers and survival in the adjuvant clinical trials described above.

Topics: Adjuvants, Immunologic; Animals; Antibodies, Monoclonal; Antibodies, Monoclonal, Murine-Derived; Antibodies, Neoplasm; Antibody-Dependent Cell Cytotoxicity; Cancer Vaccines; Cell Line, Tumor; Disease-Free Survival; Enzyme-Linked Immunosorbent Assay; Gangliosides; Hemocyanins; Immunoglobulin G; Lymphoma; Mice; Mice, Inbred C57BL; Tumor Burden; Vaccination; Vaccines, Conjugate

After 10 years of clinical trials in patients with advanced cancer, monoclonal antibodies (mAbs) against cell surface antigens have not lived up to their initial promise. One such cell surface antigen is the ganglioside GD2. GD2 is richly expressed at the cell surfaces of human neuroblastomas, sarcomas, and melanomas. We have described a murine lymphoma (EL4) that is syngeneic in C57BL/6 mice and expresses GD2, a mAb against GD2 (mAb 3F8), and we have prepared a conjugate vaccine (GD2-keyhole limpet hemocyanin plus immunological adjuvant QS-21) that consistently induces antibodies against GD2. We demonstrate here, for the first time in a syngeneic murine model, that passively administered and vaccine-induced antiganglioside antibodies prevent outgrowth of micrometastases, and we use this model to establish some of the parameters of this protection. The level of protection was proportional to antibody titer. Treatment regimens resulting in the highest titer antibodies induced the most protection, and protection was demonstrated even when immunization was initiated after tumor challenge. Treatment with 3F8 1, 2, or 4 days after i.v. tumor challenge was highly protective, but waiting until 7 or 10 days after challenge resulted in minimal protection. The results were similar whether number of liver metastases or survival was used as the end point. These results suggest that unmodified mAbs or antibody-inducing vaccines against GD2 (and possibly other cancer cell surface antigens) should be used exclusively in the adjuvant setting, where circulating tumor cells and micrometastases are the primary targets.

Topics: Adjuvants, Immunologic; Animals; Antibodies, Monoclonal; Cancer Vaccines; Gangliosides; Hemocyanins; Liver Neoplasms; Lymphoma; Mice; Mice, Inbred C57BL; Neoplasm Proteins; Neoplasm Transplantation

Carbohydrate antigens rarely provide target epitopes for cytotoxic T lymphocytes (CTL). Disialoganglioside GD2 is a glycolipid expressed at high levels in human tumors and a small group of murine lymphomas (EL4, RBL5, RMA, RMA-S, A13, and BALBRVE). Immunization of C57B1/6 mice with irradiated EL4 cells stimulated a specific CTL response and protected these animals from engraftment of EL4 lymphoma. The CTL activity resided in the CD4-CD8+ population, was dependent on T cell receptor alpha/beta, and was not removed by anti-natural killer cell immunoabsorption, but was restricted to GD2 and H-2b bearing targets. CTL activity could be completely inhibited by GD2-oligosaccharide-specific monoclonal antibodies and their F(ab')2 fragments, but not by immunoglobulin G3 myelomas or antibodies against GD3 or GM2. Soluble GD2 did not inhibit specific tumor lysis. RMA-S lymphoma cells (GD2+H-2b-TAP2 deficient) were resistant to GD2-specific CTL. Sialic acid-containing peptides eluted from EL4 lymphoma cells could (a) stabilize H-2 molecules on RMA-S cells and (b) sensitize them for GD2-specific CTL. Control peptides (derived from vesicular stomatitis virus nucleoprotein peptide and GD2-negative lymphomas) could also stabilize H-2 on RMA-S, but were resistant to GD2-specific CTL. These H-2-binding peptides could be purified by anti-GD2 affinity chromatography. We postulate a new class of naturally occurring epitopes for T cells where branched-chain oligosaccharides are linked to peptides with anchoring motifs for the major histocompatibility complex class I pocket. While analogous to the haptens trinitrophenyl and O-beta-linked acetyl-glucosamine, the potential implications of natural carbohydrates as antigenic epitopes for CTL in biology are considerable.

Topics: Animals; Antigens, Neoplasm; Epitopes; Gangliosides; H-2 Antigens; Immunization; Leukemia, Basophilic, Acute; Lymphoma; Mice; Mice, Inbred BALB C; Mice, Inbred C57BL; Mice, Inbred DBA; Mice, Nude; Neoplasm Transplantation; Neurons; Rats; Receptors, Antigen, T-Cell, alpha-beta; T-Lymphocytes, Cytotoxic; Tumor Cells, Cultured

The structure of the major gangliosides from murine lymphoma EL-4 was established by acid hydrolysis, methanolysis, methylation analysis, neuraminidase treatment and chromium trioxide oxidation. Two of these gangliosides were identified as the N-acetyl and N-glycoloyl forms of the GM2 ganglioside. The third ganglioside making up to 40% of the total ganglioside pool contained two sialic acid residues and was identified as a GD2 ganglioside which up to now had not been reported in extraneural tissues.

Topics: Animals; Ascitic Fluid; Chromatography, Thin Layer; Fatty Acids; G(M2) Ganglioside; Gangliosides; Lymphoma; Mice; Mice, Inbred C57BL; Sialic Acids