Combination metronomic chemotherapy and immunotherapy in preclinical mouse glioma models
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Cyclophosphamide (CPA) administered on a low dose, intermittent (metronomic) schedule has numerous defined mechanisms of immunomodulation. Certain immunotherapies also elicit anti-cancer responses through activation of host immune responses; however, combination with chemotherapy is often required for such responses. Therefore, additional studies are necessary to determine the best combination strategies that retain the benefits of chemotherapy without precluding the effectiveness of immunotherapy. This research characterized the anti-tumor efficacy of CPA chemotherapy combined with CpG oligodeoxynucleotide immunotherapy (CpG-1826) in a mouse model of glioma and used a high throughput sequencing approach to identify potential mechanisms of synergy. CpG-1826 treatment delayed growth and increased tumor-infiltrating immune cells in GL261 gliomas implanted in immunocompetent mice. Although anti-tumor responses varied between individual animals, both high and low CpG-1826-responsive mice showed increased anti-tumor responses when CpG-1826 was combined with CPA treatment; further, improved responses were observed with combination treatment in non-immunogenic B16F10 melanoma tumors. When CpG-1826 was combined with CPA, tumor-associated macrophages, B cells, dendritic cells, and cytotoxic T cells were increased and long-term GL261 glioma regression with immune memory was achieved. CpG oligodeoxynucleotide immunotherapy can thus be effectively combined with CPA treatment to enhance immune-based anti-tumor responses, even in poorly immunogenic cancer models. While gene expression studies have demonstrated an important link between immune infiltration and disease outcome in glioma, many different cell types are present in the tumor microenvironment, and the contributions of each cell type to composite tumor gene expression are largely uncharacterized. We used RNA-sequencing to analyze gene expression from total tumor tissue and isolated populations of CD45+ (total immune cells), F4/80+ (macrophages), and CD45- (tumor cells) cells from combination chemoimmunotherapy treated mice. Pathway analysis revealed that monotherapy or combination therapy with CPA and CpG-1826 enhanced immune processes in each cell population. Further, a number of pathways that may contribute to the efficacy of combination treatment were identified, including cell cycle progression, cell death, extracellular matrix remodeling and immune cell chemotaxis. Overall, the research presented in this thesis characterizes an effective combination of metronomic CPA and CpG-1826 immunotherapy and identifies key mechanisms of efficacy, providing novel insights into effective combination chemoimmunotherapy treatments.