Induction of Tumor Immunity Early after Syngeneic Bone Marrow Transplantation (BMT) by Transient Transfection of Mouse Neuroblastoma with Plasmid DNA Vectors Encoding a Panel of Immune Stimulatory Molecules.

Abstract

Autologous hematopoietic stem cell transplantation has resulted in improved event-free survival for patients with neuroblastoma, but the majority of these patients still relapse. We have shown that transient transfection (nucleofection) of mouse neuroblastoma (Neuro-2a) cells with plasmid DNA vectors encoding immune stimulatory molecules generates cell-based vaccines capable of inducing potent anti-tumor T cell immunity. We are exploring tumor vaccine administration early after hematopoietic stem cell transplantation because this may be an ideal setting to induce effective tumor immunity due to altered immune regulation and altered T cell homeostasis. The goal of this study was to determine the cellular components required to induce tumor-protective immunity in mice early after lethal total body irradiation (TBI) and syngeneic BMT. A/J mice were given 1100 cGy TBI, and one day later, transplanted with 107 syngeneic bone marrow cells with or without purified T cells or total splenocytes (as a source of T cells). An aggressive variant of Neuro-2a (AGN2a) was nucleofected with plasmid vectors encoding CD54 (ICAM-1), CD80, CD86 and CD137L (4-1BBL). The nucleofected cells were irradiated and then given to transplant recipients as a cell-based vaccine on days 7 and 14 post-transplant. To assess development of tumor immunity, groups of transplanted/vaccinated mice were challenged with different doses of tumor cells one week after the last vaccine. Vaccination of mice given BMT without added T cells failed to induce any protective anti-tumor immunity, and all of these mice died after challenge with 105 or 106 viable tumor cells. In contrast, 78.5% of vaccinated mice given 5x106 T cells at the time of BMT survived tumor challenge. Tumor protective immunity in the transplant recipients was dependent upon both CD4+ and CD8+ T cells, and tumor-specific CD8+ T cells in the tumor vaccine draining lymph nodes and spleens of vaccinated mice could be detected in IFN-g ELISPOT assays. From these results, we concluded that the adoptive transfer of T cells was absolutely required for induction of protective immunity by the tumor vaccine. We examined whether in vivo CD25 depletion of the transplant recipients, or depletion of CD25+ regulatory T cells from the adoptively-transferred T cells, would further increase vaccine-induced tumor immunity. This was based on previous results in non-transplanted mice, where in vivo-depletion of CD25+ cells increased effectiveness of cell-based neuroblastoma vaccines. Depletion of CD25+ cells in recipient mice or in the adoptively transferred T cells did not increase anti-tumor immunity as determined from tumor challenge experiments, suggesting that the lethal TBI itself had eliminated the majority of CD25+ regulatory T cells. Collectively, these results demonstrate that anti-neuroblastoma immunity can be induced early after BMT using a novel cell-based cancer vaccine; however, sufficient numbers of T cells must be included in the graft to achieve protective anti-tumor immunity.