Date of Graduation

5-2016

Document Type

Dissertation (PhD)

Program Affiliation

Experimental Therapeutics

Degree Name

Doctor of Philosophy (PhD)

Advisor/Committee Chair

Zhen Fan, M.D.

Committee Member

Bingliang Fang, M.D., Ph.D.

Committee Member

Gregory A. Lizee, Ph.D.

Committee Member

Dean A. Lee, M.D., Ph.D.

Committee Member

Gabriel Lopez Berestein, M.D.

Abstract

For decades, investigators have attempted to activate the immune system to prevent cancer metastasis or recurrence; however, owing to host immune tolerance to cancer antigens and the immunosuppressive environment at tumor sites, many such attempts have failed. The recent success of anti-CTLA4, PD-L1 and PD-1 antibodies targeting immune checkpoint pathways and HPV vaccines has renewed hope that patient survival can be increased through enhancing T-cell responses. We propose to test a novel approach that may bypass host immune tolerance to cancer cells. We hypothesize that host T-cell immunity acquired through vaccination against or natural infection with infectious diseases—e.g., influenza—can be re-routed to breast cancer cells if the cancer cells also express the vaccine antigens and present the antigens in complex with MHC on the cell surface. We chose HER2-breast cancer as a model for proof-of-principle.

In our study, we first examined MHC-I expression, which is required for mediating T cell-mediated response, in a panel of breast cancer cell lines with low or high levels of HER2. A previous study in literature reported an inverse correlation between the levels of HER2 and MHC-I expression in breast cancer cells. In contrast to that finding, we found no significant direct inverse correlation between the levels of HER2 and MHC-I expression. In the presence of peripheral blood mononuclear cells (PBMC), trastuzumab treatment resulted in a significant increase not only in MHC-I expression but also CD86 expression in the panel of breast cell lines. We demonstrated that this increase in MHC-I expression was correlated with an increase in IFN-γ in the co-culture of breast cancer cells and PMBC through trastuzumab-engaged PBMC. We further showed that trastuzumab treatment enhanced MHC-I expression in vivo in 4T1 mouse mammary tumors engineered to overexpress human HER2. To test our hypothesis of therapeutically redirecting preexisting non-cancer immunity developed through vaccination or contract with an infectious disease to cancer in vivo, we first immunized BALB/c mice with influenza PR8 virus to mimic flu vaccination, and then we challenged the mice with the highly aggressive 4T1 mouse mammary tumor cells or 4T1 cells lentivirally transduced to express hemagglutinin (HA) and nucleoprotein (NP) antigens of PR8 influenza virus, termed 4T1-HA+NP cells. We found a 70% rejection of 4T1-HA+NP tumors by day 12 and a significant reduction in tumor size and metastasis compared to mock (PBS) immunized mice. We also found that the anti-tumor responses in the influenza immunized group are associated with high percentage of memory CD8+ T cells, NK cells, mature DC’s, and low percentage of Treg cells and MDSC infiltration to the 4T1-HANP tumors. We next developed a trastuzumab-based immunoliposome to test our hypothesis of redirecting host influenza-induced immunity to cancer by therapeutic delivering influenza antigens to HER2-overexpressing breast cancer cells. The HER2-targeting immunoliposome was confirmed to retain its high affinity binding to HER2 in HER2-overexpressing breast cancer cells in vitro and in vivo. The immunoliposome effectively delivered labelled antigenic MHC-I influenza antigens in vivo and induce tumor regression in HER2-overexpressing tumors in influenza pre-immunized mice but not in naïve mice. Our data confirm that pre-existing non-cancer immunity can be rerouted to cancer cells through therapeutic delivery of relevant antigens using an immunoliposome approach.

Keywords

trastuzumab, immunoliposome, PD-L1, HLA-ABC, HER2, Breast cancer

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