Abstract 4075: Overcoming glioma immunoediting and MHC class I loss during adoptive cellular therapy
Authors: Wildes Tyler J. , Flores Catherine T. , Dyson Kyle , Francis Connor , Grippin Adam , Dean Bayli Divita , Mitchell Duane A. 
Published: August 2019

Abstract
Introduction: Adoptive T cell immunotherapy (ACT) leads to 30% long-term cures of intractable models of malignant glioma. While promising, glioma-bearing hosts succumb to disease for unclear reasons. Therefore, we studied glioma escape variants after immunoediting by ACT. Since the escaped tumors displayed marked gene loss making them distinct from original tumor, we anticipated that successful future strategies would need to be tailored to the new antigenic signature while using combinatorial approaches to overcome other mechanisms of escape. We therefore studied the hypothesis that a polyclonal population of ex vivo-expanded T cells specific for the total tumor RNA of glioma escape variants could successfully retarget glioma escape variants, despite immunoediting pressure.
Methods: RNAseq revealed that tumor escape variants from KR158B (TOGA1.1/1.2) displayed a loss of ~80% of the genes that were present in KR158B and GL261 glioma. We regenerated polyclonal T cells against TOGA using total tumor RNA from TOGA tumors. We used restimulation co-cultures of T cells with tumor and IFN-γ ELISAs for T cell activation. In vivo therapeutic models required re-implantation of TOGA tumors intracranially after gross dissection and passaging in vitro. Flow cytometry determined marker expression in tumors.
Results: TOGA1.1 or TOGA1.2-specific T cells recognized cognate TOGA1.1 or TOGA1.2 cells but not KR158B or the opposite glioma escape variant. When we utilized TOGA-specific T cells in vivo for TOGA-bearing mice, median survival was prolonged from 24 to 33 days (p=.0003). However, all animals succumbed to disease. We then evaluated additional mechanisms of escape. At endpoint, MHC class I on tumors was ~50% of its original expression after antigen-matched ACT (KR158B-bearing: 70% to 40% post-ACT; TOGA-bearing: 22% to 10%). Additionally, KR158B and TOGA-specific T cells express ~50% PD-1 both post-expansion ex vivo and in the tumor-infiltrating lymphocytes post-treatment while tumor-infiltrating NK cells express 30% PD-1. Given the role of NK cells in targeting MHC Ilo tumors, and the recent discoveries that PD-1 checkpoint blockade works on T cells and NK cells, we investigated the impact of PD-1 and antigen-matched ACT in KR158B or TOGA-bearing animals. This revealed that the combination generates 71% long-term cures in KR158B primary glioma while promoting an increase in activated NK and T cells. Treatment of TOGA-bearing animals is underway and will include the depletion of either CD8 T cells or NK cells to determine the relative contributions of each cell type.
Conclusions: Glioma escape variants retain expression of immunogenic antigens. Generation of ACT with specificity for escape variants provides exquisite antigen-specific recognition of the immunoedited tumors. Combinatorial use of antigen-specific ACT with PD-1 blockade provides a flexible and specific platform for treatment of primary and escaped tumors.
Citation Format: Tyler J. Wildes, Catherine T. Flores, Kyle Dyson, Connor Francis, Adam Grippin, Bayli Divita Dean, Duane A. Mitchell. Overcoming glioma immunoediting and MHC class I loss during adoptive cellular therapy [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 4075.