Antigen-Loaded Monocytes as a Novel Cancer Vaccine

Gunn, Michael D

Huang, Min-Nung

2018-03-20T17:53:11Z

2018-03-20T17:53:11Z

2017

Immunology

Dendritic cells (DC) have been the key elements in developing cancer vaccines to induce potent T cell responses to eradicate tumors. However, the common approach adopted in clinical trials using ex vivo generated DC loaded with tumor antigens (Ag) has been challenged by its limited clinical response, complexity, and quality of the manufacturing process. Alternative efforts focused on in vivo Ag loading on endogenous primary DC have not yet been well validated in their efficacy for cancer treatment, suggesting the efficiency of in vivo Ag transfer to endogenous DC from currently available Ag-delivering vehicles needs to be further improved. Here, I aim to develop an alternative cellular vaccine platform that can circumvent the aforementioned problems. I reason that classical Ly-6Chi monocytes (i.e. monocytes hereafter) can be a promising candidate to be loaded with tumor Ag and induce effective T cell responses. With advantages including easy-purification from human peripheral blood, monocytes evidently can present antigens directly via in vivo differentiation into bona fide DC or indirectly via antigen transfer to lymphoid resident DC to induce strong Th1 or cytotoxic T lymphocyte (CTL) responses. However, whether monocytes exploit favorably direct or indirect pathway to present the same Ag they are carrying to trigger effective immune responses remains unclear. Furthermore, how exactly monocytes or monocyte-derived cells transfer antigens to lymphoid resident DC has yet to be elucidated. I hypothesized that Ag-loaded monocytes can induce strong anti-tumor immunity and began the research by investigating the immune responses that can be induced by Ag-loaded monocytes. I then went on to determine the mechanisms that mediate monocyte-induced immune responses and evaluate anti-tumor efficacy of this monocyte vaccine.

In the first part of this study, I characterized the immune responses induced by Ag-loaded monocytes. By using negative selection via magnetic-activated cell sorting (MACS) columns, I was able to purify monocytes from bone marrow (BM) cells and determined that these monocytes could be successfully loaded with Ag in the forms of proteins, peptides and mRNA. I found that intravenously (IV) injected Ag-loaded monocytes induced robust Ag-specific CD4+ and CD8+ T cell responses in mice without triggering antibody responses. This vaccine activity of Ag-loaded monocytes appeared to be dose-dependent and required live monocytes with no need of ex vivo stimulation. I found that Ag-specific CD8+ T cells induced by Ag-loaded monocytes were functionally more robust than those induced by protein Ag emulsified in a traditional adjuvant CFA.

In the second part of this study, I investigated how IV injected Ag-loaded monocytes stimulate T cell responses. I identified that the spleen is the primary immune niche for Ag-loaded monocytes to induce T cell responses. I found that Ag-loaded monocytes mainly retain in the spleen where they begin to differentiate into phenotypic DC. Surprisingly, major histocompatibility complex (MHC)-deficient monocytes maintain full capacity to stimulate T cell responses, suggesting that Ag-loaded monocytes do not present Ag by themselves. I determined that endogenous splenic DC is absolutely required for monocyte-induced T cell responses. Therefore, Ag-loaded monocytes induce T cell responses indirectly via transferring Ag to splenic DC even they do differentiate into phenotypic DC in the spleen. I elucidated that this monocyte-to-DC Ag transfer occurs via gap junctions for CD8+ T cell responses and via macrophages for CD4+ T cell responses.

In the final part of this study, I demonstrated that IV injected Ag-loaded monocytes have robust anti-tumor efficacy targeting both model and validated tumor Ag in prophylactic, memory and therapeutic murine SQ melanoma models. The anti-tumor efficacy is superior to that seen with traditional adjuvants or RNA-pulsed DC vaccines, and can be combined with checkpoint blockade to increase their efficacy. Furthermore, I demonstrated that Ag-loaded monocytes have a clear anti-tumor efficacy in an intracranial glioblastoma (GBM) model targeting against mutant isocitrate dehydrogenase 1-R132H (mIDH1-R132H), a validated tumor Ag of GBM.

In conclusion, IV injection of unactivated Ag-loaded monocytes without adjuvants induces highly efficacious anti-tumor T cell responses via dual independent and efficient Ag transfer pathways to splenic DC. These findings revise the paradigm that monocytes have to be activated ex vivo to achieve optimal vaccine efficacy and reveal unappreciated cell-associated Ag acquiring pathways of splenic DCs that can be specifically manipulated for future vaccine design in the treatment of human cancers.

https://hdl.handle.net/10161/16231

Immunology

Cancer

Dendritic cell

Immunotherapy

monocyte

Vaccine

Antigen-Loaded Monocytes as a Novel Cancer Vaccine

Dissertation