Abstract 4588: Characterization of anti-CTLA-4 effects on tumor growth and immune microenvironment in a syngeneic CT26.WT colon carcinoma mouse model
Authors: Mäki-Jouppila Jenni H. , Kähkönen Tiina E. , Suominen Mari I. , Halleen Jussi M. , Bernoulli Jenni 
Published: August 2019

Colorectal cancer is the third most common type of cancer and one of the leading causes of cancer related deaths. It is a group of various diseases at molecular level, which can make targeted drug development challenging. However, novel therapeutics including immunotherapies have shown their potential to be utilized in the future also when treating colorectal cancer. One promising example is ipilimumab, a monoclonal antibody targeting CTLA-4 (cytotoxic T lymphocyte antigen-4, also known as CD152). CTLA-4 is a cell surface receptor expressed on activated T and B lymphocytes. It regulates immunological tolerance and downregulates immune responses. The aim of the study was to characterize the effects of anti-CTLA-4 on tumor growth, survival and tumor immune microenvironment of mice subcutaneously inoculated with mouse colon carcinoma CT26.WT cells.

CT26.WT cells were subcutaneously inoculated into female Balb/c mice (n=27). Body weights and tumor dimensions were determined two times a week during the course of the study. The mice were randomized to three groups (n=9/group) with similar tumor volume and variation and dosing was started when an average tumor size of approximately 50 mm³ was reached (day 7). Anti-CTLA-4 antibody (9D9, 10 mg/kg), an isotype control (mouse IgG2b) and vehicle (PBS) were administered (i.p.) twice a week. Tumor growth and survival were followed for 26 days after inoculation of the cancer cells. The mice were sacrificed individually when the maximum tumor volume (1500 mm³) was reached, the tumors were ulcerated or at the latest on study day 26. Tumor samples were collected for histological and immunohistochemical analysis and stained for helper T cells (CD4), cytotoxic T cells (CD8), myeloid-derived suppressor cells (MDSC; CD11b), tumor-associated macrophages (F4/80), and inflammatory monocytes and neutrophils (FCGR1/CD64) for characterization of tumor immune microenvironment.

Maximum tumor volume and tumor ulceration were observed the earliest on study day 17. Anti-CTLA-4 suppressed tumor growth, and all mice in the group receiving anti-CTLA-4 were still in the study on day 26. Only one and four mice were left in the vehicle and isotype control groups, respectively. Immunohistochemical staining suggested an increase of CD4+ tumor-infiltrating lymphocytes (TILs) in anti-CTLA-4 treated mice. Furthermore, more TAMs were observed in tumors of anti-CTLA-4 treated mice, especially in the borderlines of necrotic areas. Large quantity of CD11b+ MDSCs and inflammatory cells were observed in CT26.WT tumors.

CT26.WT subcutaneous model can be utilized for studying the effects of immunotherapies, and anti-CTLA-4 treatment suppressed tumor growth in this model. CT26.WT tumors represent a highly immunogenic hot tumor type with high immune cell infiltration, associated with a good efficacy of anti-CTLA-4 treatment in the model.

Citation Format: Jenni H. Mäki-Jouppila, Tiina E. Kähkönen, Mari I. Suominen, Jussi M. Halleen, Jenni Bernoulli. Characterization of anti-CTLA-4 effects on tumor growth and immune microenvironment in a syngeneic CT26.WT colon carcinoma mouse model [abstract]. In: Proceedings of the American Association for Cancer Research Annual Meeting 2019; 2019 Mar 29-Apr 3; Atlanta, GA. Philadelphia (PA): AACR; Cancer Res 2019;79(13 Suppl):Abstract nr 4588.