The purpose of these studies was to determine the effect of Ipilimumab treatment on T cell expansion, activation, cytokine production, tumor infiltration, and expression level of co-stimulatory and co-inhibitory receptors in a tumor bearing humanized NSG mouse model. Antibody therapies for immune modulation are proving to be effective in the oncology setting, as demonstrated by anti-CTLA-4 Ipilimumab and anti-PD-1 Pembrolizumab clinical activity. In order to assess efficacy of new therapies in the context of immune activation and tumor response, there is a need for suitable preclinical in vivo models. One approach to study immune cell function is to inject human peripheral blood mononuclear cells (PBMCs) into adult immunodeficient NSG (NOD/SCID/IL-2Rγnull) mice. This model, known as Hu-PBMC NSG, induces a Graft-versus-Host Disease (GvHD) state and has been used to study effector and memory T cell activity. Here we utilized the Hu-PBMC NSG model implanted with human cancer cell lines to investigate the effect of Ipilimumab on T cells and tumor growth in vivo. All studies were conducted in accordance with the GSK Policy on the Care, Welfare and Treatment of Laboratory Animals and were reviewed the Institutional Animal Care and Use Committee at GSK. The human biological samples were sourced ethically and their research use was in accord with the terms of the informed consents. We confirmed human PBMC engraftment in NSG mice, validated tumor growth of human A2058 melanoma and 786-O renal adenocarcinoma cell lines, and dosed Ipilimumab in tumor bearing NSG mice with different human PBMC donors. Naïve NSG mice were intravenously injected with 20x106 human PBMCs and the kinetics of human cell engraftment was monitored weekly. For studies including Ipilimumab dosing, mice were inoculated with a subcutaneous injection of 2.5x106 A2058 or 1x106 786-O tumor cells. As tumor size reached 100 mm3, mice were randomized and injected with 20x106 human PBMCs. Two days later, mice were intraperitoneally treated with Ipilimumab or human IgG1 isotype control twice weekly for 6 doses total. Tumor growth and body weight was evaluated over time. Peripheral blood was collected weekly for analysis of T cell activation, receptor expression levels, and human cytokine production until mice developed GvHD or tumor volume reached 2,000 mm3. Select tumors were also harvested for tumor infiltrating lymphocyte (TIL) analysis by flow cytometry. Our studies showed NSG mice demonstrated similar human CD45+ cell engraftment in blood with various PBMC donors. The frequency of circulating human CD45+ lymphocytes in mouse peripheral blood increased to 50% or greater until week 4. 95% of the CD45+ cells were CD3+ T cells, containing both CD4+ and CD8+ subsets. Signs of GvHD were observed at week 4, and serum cytokine analysis showed high levels of GvHD markers e.g. IL5, IL10, and TNF-alpha. Ipilimumab treatment delayed tumor growth, increased the expansion of human CD45+ cells, and induced higher levels of TNF-alpha, IL-12p70, IL-13, and IL-5 cytokines compared to isotype control. Ipilimumab also increased the surface expression level of CD69, PD-1, OX40, ICOS, CD137, TIM3, and LAG3 on circulating T cells, and increased the number of A2058 tumor infiltrating lymphocytes. TIL analysis showed that compared to isotype control, Ipilimumab increased expression of CD69, PD-1, OX40, and ICOS on tumor infiltrating lymphocytes. In conclusion, Ipilimumab treatment increased T cell expansion, activation, expression of co-stimulatory and co-inhibitory receptors, and cytokine production in this tumor-bearing Hu-PBMC NSG model. It also increased the number of tumor infiltrating lymphocytes with a corresponding tumor growth delay. Based on Ipilimumab activity, this model can be utilized to assess pre-clinical efficacy of novel immunotherapies.
Citation Format: Meixia Bi, Chris Hopson, Tianqian Zhang, James Smothers, Axel Hoos. In vivo characterization of Ipilimumab T cell modulation and antitumor activity in a tumor bearing humanized NSG mouse model. [abstract]. In: Proceedings of the CRI-CIMT-EATI-AACR Inaugural International Cancer Immunotherapy Conference: Translating Science into Survival; September 16-19, 2015; New York, NY. Philadelphia (PA): AACR; Cancer Immunol Res 2016;4(1 Suppl):Abstract nr A054.
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