Abstract A056: TNFR2 is a novel marker of exhaustion and TNFR2 blockade improves subcutaneous tumor control
Authors: Hoyt-Miggelbrink Alexandra , Polania Jessica Waibl , Mohan Aditya , Wachsmuth Luke , Lorrey Selena , Lerner Emily , Wilkinson Daniel , Hardigan Andrew , Hotchkiss Kelly , Hemmers Saskia , Quackenbush Sarah Cook , Ayasoufi Katayoun , Fecci Peter 
Published: October 2024

Abstract
Checkpoint inhibitors have been very successful in various tumors, including melanoma and lung cancer. However, these treatments have failed in other tumors, including pancreatic cancer and glioblastoma. Approaches harnessing the immune response are hindered by multiple factors, including T cell exhaustion. Exhausted T cells are less capable of limiting tumor progression. TOX is particularly important for the transcriptional and epigenetic reprogramming of exhausted T cells, and without TOX, T cells cannot differentiate down the exhaustion pathway. While it is known that NFAT is directly upstream of TOX, our understanding of the pathways that lead to the upregulation of TOX remains incomplete. We hypothesized that tumor necrosis factor (TNF) could be involved in the upregulation of TOX, as TNF has been shown to induce exhaustion and can lead to the translocation of NFAT into the nucleus. While we did not detect an upregulation of the pro-inflammatory TNF receptor type I (TNFR1), we observed significant upregulation of the anti-inflammatory TNF receptor type II (TNFR2) in tumor-infiltrating T cells, but not in lymphoid organs. This local upregulation mimics expression patterns of canonical exhaustion markers and suggests that TNFR2 signaling could be involved in the progression of exhaustion. In support of this hypothesis, TNFR2 expression is correlated other markers of exhaustion, including PD1, TIM3, and TOX. Furthermore, TNFR2 knock out (KO) CD8 T cells have significantly lower expression of TOX, without the concomitant decrease of TIM3. Whereas previous studies have intricately linked TIM3 and TOX expression, these data suggest that TIM3 and TOX are regulated independently. To further evaluate transcriptional regulation of WT and KO T cells, we utilized bulk RNA-sequencing. After performing pathway analyses, we detected a significant reduction of T cell exhaustion, senescence, and immune checkpoint pathways in TNFR2 KO T cells. We identified significant decreases in various exhaustion-related transcription factors and coinhibitory markers. In contrast, we discovered a significant increase in AP1 transcription factors, commonly associated with T cell effector functions. Given this reduced exhaustion profile, we subsequently investigated the influence of TNFR2 on tumor burden. TNFR2 KO mice had significantly less tumor burden following subcutaneous tumor challenges. To make this approach more translatable, we treated mice with a TNFR2 antagonist. While the antagonist alone was not sufficient to improve tumor control, combination with immune checkpoint blockade (anti-PD1) significantly reduced tumor volumes in traditionally resistant models. These data provide evidence for a novel marker of exhaustion that could result in a novel therapeutic strategy.
Citation Format: Alexandra Hoyt-Miggelbrink, Jessica Waibl Polania, Aditya Mohan, Luke Wachsmuth, Selena Lorrey, Emily Lerner, Daniel Wilkinson, Andrew Hardigan, Kelly Hotchkiss, Saskia Hemmers, Sarah Cook Quackenbush, Katayoun Ayasoufi, Peter Fecci. TNFR2 is a novel marker of exhaustion and TNFR2 blockade improves subcutaneous tumor control [abstract]. In: Proceedings of the AACR Special Conference in Cancer Research: Tumor Immunology and Immunotherapy; 2024 Oct 18-21; Boston, MA. Philadelphia (PA): AACR; Cancer Immunol Res 2024;12(10 Suppl):Abstract nr A056.