TLR-9 Activated Lymphoma B Cells Induce Anti-Tumor Immunity in a Murine Model

Abstract

Background: Tumor vaccines typically combine unique, tumor antigens with immune stimulants in an effort to elicit a tumor-specific immune response. In prior work we have described a vaccination maneuver that combines cytotoxic chemotherapy to release tumor antigens with intra-tumor injection of a Toll-like Receptor 9 (TLR9) ligand, CpG oligonucleotide (Li and Levy, J. Immunology, 2007). In this therapy, CpG can activate both host antigen presenting cells as well as the tumor itself. Stimulation of tumor B cells through TLR9 induces up-regulation of immune co-stimulatory molecules including CD80, CD86, and CD40 as well as increasing expression of MHC Class I and II. We have now developed an alternative vaccination approach in which B cell lymphoma tumor cells are stimulated with CpG ex vivo and administered as a whole-cell vaccine.Methods: A20 B cell lymphoma tumor cells were incubated with CpG for 72 hours, irradiated, and administered to naïve Balb/C mice. The complete vaccination regimen included six doses of 1×106 cells administered daily at a sub-cutaneous (s.c.) site. Vaccine-induced immune responses were assessed by measuring IFN-γ expression of peripheral blood lymphocytes (PBLs) in response to co-culture with A20 tumor cells. Tumor protection studies were conducted by challenging vaccinated mice with a lethal dose of 10×106 A20 tumor cells. Anti-tumor immunity generated by vaccination was also tested in adoptive cell transfer studies.Results: Sub-cutaneous vaccination with CpG-stimulated, whole-cell vaccine induces robust anti-tumor T cell immunity comparable to that induced by intra-tumor vaccination with CpG. This immunizing effect was dependent on tumor cell activation, since native tumor cells were less efficient at inducing anti-tumor immune responses. Both CD4+ and CD8+ T cells participated in this response. Mice vaccinated with this regimen were protected against tumor challenge. Splenocytes from vaccine-primed donors were adoptively transferred into irradiated, syngeneic recipients. These ‘immunotransplanted’ recipients had even greater immune protection against tumor challenge than the directly vaccinated donor mice.Conclusions: We have developed a vaccination approach that takes advantage of the antigen presentation capability of malignant B cells. Vaccination with CpG-activated lymphoma cells induced anti-tumor immune responses that were further enhanced by adoptive transfer of immune cells into lymphodepleted recipients. These vaccine maneuvers are directly translatable into therapeutic, human clinical trials.