Date of Award

2018

Degree Type

Dissertation

Degree Name

Doctor of Philosophy (PhD)

Graduate Group

Pharmacology

First Advisor

Saar Gill

Second Advisor

Carl H. June

Abstract

Despite recent landmark advances in chimeric antigen receptor (CAR) T cell immunotherapy for the treatment of human cancer, metastatic solid tumors remain an intractable challenge. Myeloid cells are actively recruited to the tumor microenvironment (TME), where tumor associated macrophages (TAMs) are often the most abundant infiltrating immune cell. Currently, macrophage orientated immunotherapeutic approaches under clinical development in oncology seek to reduce TAM infiltration or enhance TAM phagocytosis. We hypothesized that genetically engineering human macrophages with CARs against tumor-associated antigens could redirect their phagocytic activity and lead to therapeutic efficacy with the potential for the induction of an anti-tumor T cell response.

In this thesis, we demonstrate that CD3-zeta based CARs are capable of inducing phagocytosis by human macrophages. Notably, an active intracellular CAR signaling domain was required for activity. Targeted phagocytosis and clearance of CD19+, mesothelin+, and HER2+ cells by CARs targeted against each respective antigen was significantly superior to that by control untransduced (UTD) macrophages. Importantly, CAR macrophages were capable of polyphagocytosis and serial phagocytosis of tumor cells.

We demonstrate that primary human monocyte derived macrophages, which are resistant to most viral vectors, are efficiently transduced by the chimeric-fiber adenoviral vector Ad5f35. Ad5f35 transduced primary human CAR macrophages demonstrated targeted phagocytosis, with phagocytic activity dependent on both the CAR and antigen densities. CAR, but not UTD, macrophages led to potent dose-dependent killing of tumor cells in vitro and led to tumor regression and improved overall survival in murine xenograft models of human cancer.

Macrophage transduction with Ad5f35 leads to a broad gene expression change, an interferon signaling signature, and induction of a classically activated M1 phenotype. CAR macrophages upregulated co-stimulatory ligand and antigen processing/presentation genes and led to enhanced T cell stimulation in vitro and in vivo. Lastly, CAR, but not UTD, macrophages showed a broad resistance for M2 conversion in response to immunosuppressive cytokines.

In conclusion, human CAR macrophages display targeted tumor phagocytosis, lead to improved overall survival in xenograft models, and demonstrate enhanced T cell stimulation. Taken together, these data show that CAR macrophages are a novel cell therapy platform for the treatment of human cancer.

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Available to all on Tuesday, December 21, 2021

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