Abstract 4624: Profiling antitumor activity, immune cell infiltration, pharmokinetics, and receptor occupancy in a murine colon cancer model following immune checkpoint blockade
Authors: Allison Patrick , Chang Hua-Chen , Trampont Paul , Standarski Lindsey , Urisic Lauren , Rusk Jennifer , Mehta Jaydeep , Fohey Joshua , Thomas Eric , Agrawal Karan , Wilkinson Brandy 
Published: August 2019

Clinical success of immune checkpoint blocking antibodies has accelerated the evaluation of immune-related targets for novel anti-cancer therapies. Preclinical testing of immune-targeted oncology agents requires preclinical models with functional immune systems. Utilization of murine syngeneic tumor models provides a robust system to evaluate anti-tumor activity and mechanism of action. This study evaluates the anti-tumor activity and immune response of different immune-oncology therapies in the CT26.WT murine colon cancer model. In addition, an anti-PD-1 pharmacokinetic profile was established and receptor occupancy evaluated.

For this study, C57BL/6JOlaHsd mice were subcutaneously inoculated with CT26.WT cells. Tumor bearing mice were administered anti-CTLA-4, anti-PD-1, anti-PD-L1, anti-OX40, or anti-LAG3 monotherapy twice weekly for 3 weeks. Tumor volume was used to assess anti-tumor activity. Immune response to checkpoint blockade was monitored in a subset of tumors, and lymphocyte and myeloid populations were analyzed following two weeks of dosing. In addition, an anti-PD-1 pharmacokinetic profile experiment was performed using LC-MS/MS on mouse plasma from multiple time points. PD-1 receptor occupancy was also determined by flow cytometry utilizing a saturation/detection method.

Following 3 weeks of monotherapy treatment all immune checkpoint blocking therapies, anti-CTLA4, anti-PD-1, anti-PD-L1, anti-OX40, and anti-LAG3, resulted in significant antitumor response in CT26.WT derived tumors. Phenotypes of tumor infiltrating lymphocytes (TILs) resident in tumors were profiled across check point inhibitor treatments: CD45+ lymphocytes were analyzed for populations of cytotoxic lymphocytes (CTLs, CD8+ cells), T-helper cells (CD4+ cells), regulatory T-cells (T-regs, CD4+/CD25+/FoxP3+ cells). The immune cell responses to checkpoint inhibitors were distinct and varied across immune checkpoint blocking antibodies. Furthermore, an empirical exposure-response analysis of anti-PD-1 in the CT26.WT colon model was demonstrated using a novel LC/MS-MS method of detection. Dose dependent occupancy of PD-1 receptors by anti-PD-1 in CT26.WT tumors was also observed.

Our results demonstrate a significant anti-tumor response across five immune checkpoint blocking antibodies in the CT26.WT colon cancer model; however, immune response profiles are notably different across these blocking antibodies. Importantly, a novel LC/MS-MS method was developed specifically in mouse plasma to determine exposure response. Utilizing these multiple data sets will allow for PK/PD modeling to generate expectations for future dose response. This type of comprehensive analysis constitutes a highly-relevant tool to evaluate efficacy and mechanism of action for novel immune-targeted therapies for oncology.

Citation Format: Patrick Allison, Hua-Chen Chang, Paul Trampont, Lindsey Standarski, Lauren Urisic, Jennifer Rusk, Jaydeep Mehta, Joshua Fohey, Eric Thomas, Karan Agrawal, Brandy Wilkinson. Profiling antitumor activity, immune cell infiltration, pharmokinetics, and receptor occupancy in a murine colon cancer model following immune checkpoint blockade [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 4624.