Abstract 3979: PD-1 signals regulate NKG7 that is required for CD8+ T cells to sustain cytotoxicity and durable antitumor memory responses
Authors: Mao Zhiming , Gicobi Joanina , Hirdler Jacob , Liu Xin , Hsu Michelle , Dellacecca Emilia , Zhang Wenjing , Lucien-Matteoni Fabrice , dong Hai 
Published: March 2024

Abstract
Cytotoxic CD8+ T cells play a key role in response to anti-PD-1/PD-L1 immune checkpoint inhibitors (ICI) therapy. However, the exact effector molecules that are regulated by PD-1 signals in cytotoxic CD8+ T cells have not been completely defined. Lack of this knowledge is a crucial problem in addressing how to overcome the low clinical response rate to ICI therapy. When exploring why certain patients not responding to ICI therapy, we found non-responders did not have an elevated NKG7 expression in CD8+ T cells as responders to ICI therapy. The difference in NKG7 expression in CD8+ T cells between non-responders and responders to anti-PD-1 therapy prompted us to investigate the potential role of PD-1 signaling in regulating NKG7 expression. To that end, we generated a CD8+ T cell-specific Pdcd1 conditional knockout mice (CD8 E8ICre-Pdcd1fl/fl, also known as CD8-PD-1 cKO) where Pdcd1 is deleted after thymic selection of single positive CD8+ T cells. We observed a delayed tumor growth and prolonged survival in CD8-PD-1 cKO mice compared to control mice after B16-OVA melanoma tumor challenge. On Day 12, when the CD8-PD-1 cKO mice demonstrated their ability to control tumor growth, we conducted single cell RNA sequencing on isolated tumor infiltrating CD8+ T cells, revealing a significant increase of Nkg7 expression in effector CD8+ T cells in the Pdcd1 deficient CD8+ T cells compared to the control wild type T cells. Pdcd1-deficient effector CD8+ T cells also had increased expression of genes linked to NFAT/Calcineurin signaling pathway including Nfatc1, S100a4, Tbet and Tox, suggesting Nkg7 could be regulated by Pdcd1 via the NFAT pathway. To determine the impact of increased NKG7 expression in CD8+ T cells, we used OVA protein as a surrogate tumor antigen in vaccination. We found a complete B16-OVA tumor rejection in CD8-PD-1 cKO mice compared to control mice when tumor cells were injected on day 7 after vaccination with OVA protein and poly (I:C) (as an adjuvant), but only a partial rejection on day 21. The CD8-PD-1 cKO mice that were tumor-free for 30-40 days rejected 2nd tumor challenge with B16-OVA tumors. This long-term protection is tumor antigen (OVA) specific because these mice did not reject B16F10 tumor cells that do not express OVA antigen. We also found effector memory CD8+ T cells with high cytotoxicity increased in the spleen of CD8-PD-1 cKO mice after OVA antigen immunization compared to control. In conclusion, we found NKG7 is one of the effector molecules in cytotoxic CD8+ T cells, which can be regulated by PD-1 signals through NFAT signaling. Targeting the PD-1-NFAT pathway may be a new option to overcome low responses of T cells to ICI therapy in treatment of refractory cancer.
Citation Format: Zhiming Mao, Joanina Gicobi, Jacob Hirdler, Xin Liu, Michelle Hsu, Emilia Dellacecca, Wenjing Zhang, Fabrice Lucien-Matteoni, Hai dong. PD-1 signals regulate NKG7 that is required for CD8+ T cells to sustain cytotoxicity and durable antitumor memory responses [abstract]. In: Proceedings of the American Association for Cancer Research Annual Meeting 2024; Part 1 (Regular Abstracts); 2024 Apr 5-10; San Diego, CA. Philadelphia (PA): AACR; Cancer Res 2024;84(6_Suppl):Abstract nr 3979.