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930 Protein homeostasis promotes T cell stemness and enables exhausted TIL function in cancer
  1. Nicole Scharping1,
  2. Allison Cafferata1,
  3. Maximilian Heeg1,
  4. Maria Matias2,
  5. Quynhanh Nguyen1,
  6. Samuel Myers2 and
  7. Ananda Goldrath1
  1. 1University of California San Diego, San Diego, CA, USA
  2. 2La Jolla Institute for Immunology, San Diego, CA, USA
  • Journal for ImmunoTherapy of Cancer (JITC) preprint. The copyright holder for this preprint are the authors/funders, who have granted JITC permission to display the preprint. All rights reserved. No reuse allowed without permission.


Background In malignancies, CD8+ tumor-infiltrating lymphocytes (TIL) can target tumor cells, but often fail to cure disease due to chronic TIL activation in the tissue’s immunosuppressive tumor microenvironment, resulting in differentiation into an exhausted T cell state. In healthy tissues, T cells differentiate into tissue-resident memory (TRM) in response to infection, and after clearance of antigen can remain to survey and protect from reinfection. When TRM-like TIL are found in cancerous tissue, improved responses to immunotherapy and better patient outcomes are observed; therefore, understanding the connection between exhausted TIL and TRM can inform efforts to manipulate T cell fates towards TRM-like TIL to benefit cancer immunotherapy.

Methods To better understand the relationship among TRM-like TIL, exhaustion states, and TRM, we directly compared TRM from acute viral infection and exhausted TIL from tumors to find proteomic and transcriptional differences between these distinct T cell states. Focusing on genes highly expressed by TRM that may mediate their enhanced functions in tissues, we looked at genes that became downregulated as T cells transitioned to terminal exhaustion. This approach identified numerous genes related to protein homeostasis (proteostasis), including multiple under-characterized E3 ubiquitin ligases.

Results Protein regulation by the ubiquitin-proteosome system is an essential biological process, crucial for cell differentiation, and by investigating proteostasis in TIL, we found that exhausted T cells have an excess of misfolded proteins accumulated in their cytosol. CRISPR knockout studies in acute infection showed that loss of our E3 ubiquitin ligases decreased T cell stemness in tissues, and accelerated differentiation to their terminal fate via decreased ubiquitin-mediated proteasomal protein recycling. When we enforced expression of these ligases in tumor-specific T cells in cancer, we found the transduced T cells showed a decrease in cytosolic misfolded proteins and showed enhanced anti-tumor function: increased accumulation in the TME, upregulation of TRM markers, and increased IFN-gamma production, which resulted in better tumor control and improved mouse survival.

Conclusions These data highlight an underappreciated relationship between proteostasis and T cell stemness, and provide novel avenues of immunotherapeutic approaches for cancer.

Ethics Approval The study was approved by UCSD’s Institutional Animal Care and Use Committee, protocol number S04105.

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