Glen Dranoff

To compare the ability of different cytokines and other molecules to enhance the immunogenicity of tumor cells, we generated 10 retroviruses encoding potential immunomodulators and studied the vaccination properties of murine tumor cells transduced by the viruses. Using a B16 melanoma model, in which irradiated tumor cells alone do not stimulate significant anti-tumor immunity, we found that irradiated tumor cells expressing murine granulocyte-macrophage colony-stimulating factor (GM-CSF) stimulated potent, long-lasting, and specific anti-tumor immunity, requiring both CD4+ and CD8+ cells. Irradiated cells expressing interleukins 4 and 6 also stimulated detectable, but weaker, activity. In contrast to the B16 system, we found that in a number of other tumor models, the levels of anti-tumor immunity reported previously in cytokine gene transfer studies involving live, transduced cells could be achieved through the use of irradiated cells alone. Nevertheless, manipulation of the vaccine or challenge doses made it possible to demonstrate the activity of murine GM-CSF in those systems as well. Overall, our results have important implications for the clinical use of genetically modified tumor cells as therapeutic cancer vaccines.

Granulocyte-macrophage colony-stimulating factor (GM-CSF) gene-transduced, irradiated tumor vaccines induce potent, T-cell-mediated antitumor immune responses in preclinical models. We report the initial results of a Phase I trial evaluating this strategy for safety and the induction of immune responses in patients with metastatic renal cell carcinoma (RCC). Patients were treated in a randomized, double-blind dose-escalation study with equivalent doses of autologous, irradiated RCC vaccine cells with or without ex vivo human GM-CSF gene transfer. The replication-defective retroviral vector MFG was used for GM-CSF gene transfer. No dose-limiting toxicities were encountered in 16 fully evaluable patients. GM-CSF gene-transduced vaccines were equivalent in toxicity to nontransduced vaccines up to the feasible limits of autologous tumor vaccine yield. No evidence of autoimmune disease was observed. Biopsies of intradermal sites of injection with GM-CSF gene-transduced vaccines contained distinctive macrophage, dendritic cell, eosinophil, neutrophil, and T-cell infiltrates similar to those observed in preclinical models of efficacy. Histological analysis of delayed-type hypersensitivity responses in patients vaccinated with GM-CSF-transduced vaccines demonstrated an intense eosinophil infiltrate that was not observed in patients who received nontransduced vaccines. An objective partial response was observed in a patient treated with GM-CSF gene-transduced vaccine who displayed the largest delayed-type hypersensitivity conversion. No replication-competent retrovirus was detected in vaccinated patients. This Phase I study demonstrated the feasibility, safety, and bioactivity of an autologous GM-CSF gene-transduced tumor vaccine for RCC patients.

Multiple clinical trials have shown the efficacy of adoptively transferred allogeneic antigen-specific T cells for the treatment of viral infections and relapsed hematologic malignancies. In contrast, the therapeutic potential of autologous antigen-specific T cells has yet to be established since it has been technically difficult to generate sufficient numbers of these T cells, ex vivo. A major obstacle to the success of this objective derives from our inability to simply and rapidly isolate and/or expand large numbers of highly efficient antigen presenting cells (APCs) for repetitive stimulations of antigen-specific T cells in vitro. We show that autologous CD40-activated B cells represent a readily available source of highly efficient APC that appear to have several important advantages over other APCs for ex vivo T cell expansion including: (a) methodological simplicity necessary to generate continuously large numbers of APCs from just 50 cm3 of peripheral blood without loss of APC function; (b) capacity to induce high peak T cell proliferation and interferon-gamma production without IL-10 production; (c) ease in cryopreservation; and (d) markedly reduced cost. We, therefore, contend that CD40-activated B cells are an alternative source of highly efficient APCs with which to generate antigen-specific T cells ex vivo for autologous adoptive immunotherapy.

Survival for patients with glioblastoma, the most common high-grade primary CNS tumor, remains poor despite multiple therapeutic interventions including intensifying cytotoxic therapy, targeting dysregulated cell signaling pathways, and blocking angiogenesis. Exciting, durable clinical benefits have recently been demonstrated for a number of other challenging cancers using a variety of immunotherapeutic approaches. Much modern research confirms that the CNS is immunoactive rather than immunoprivileged. Preliminary results of clinical studies demonstrate that varied vaccine strategies have achieved encouraging evidence of clinical benefit for glioblastoma patients, although multiple variables will likely require systematic investigation before optimal outcomes are realized. Initial preclinical studies have also revealed promising results with other immunotherapies including cell-based approaches and immune checkpoint blockade. Clinical studies to evaluate a wide array of immune therapies for malignant glioma patients are being rapidly developed. Important considerations going forward include optimizing response assessment and identifiying correlative biomarkers for predict therapeutic benefit. Finally, the potential of complementary combinatorial immunotherapeutic regimens is highly exciting and warrants expedited investigation.

Brain tumors have an immunoprivileged status which contributes to their refractoriness to treatment. In this study, immune rejection of GL261 glioma tumors in the mouse brain was achieved by subcutaneous vaccination with GM-CSF-transduced glioma cells. Cultured GL261 cells were transduced to secrete murine GM-CSF using a retrovirus vector, then irradiated, and injected subcutaneously into H-2 matched C57BL/6 mice. In prevaccination studies, the median survival time (MST) of animals vaccinated with 5 x 10(4) or 5 x 10(5) GM-CSF-transduced cells 7 days prior to intracranial injection of 10(6) nontransduced, nonirradiated GL261 cells was significantly prolonged by 45-50% compared with animals vaccinated in parallel with nontransduced, irradiated glioma cells. In treatment of established gliomas, the MST of animals, which were treated subcutaneously with 5 X 10(6) irradiated GM-CSF-transduced cells 3 days after intracranial injection of 2 x 10(4) nontransduced cells, was prolonged significantly by 36% compared with animals treated with the same number of nontransduced, irradiated cells or to sham-treated animals. In prevaccination studies, histology of brain tumors 4 days after intracranial tumor cell injection revealed infiltrates of CD8+ lymphocytes and eosinophils, the latter exclusively in animals vaccinated with GM-CSF-transduced cells, Thus, subcutaneous injection of irradiated GM-CSF-transduced glioma cells can induce a potent immune response to intracranial gliomas both as a vaccination against subsequent intracranial glioma cell implantation and for treatment of established intracranial glioma.