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386 Sheltering telomeres from oxidative damage in therapeutic T cells protects against tumor-induced immune dysfunction
  1. Dayana Rivadeneira1,
  2. Sanjana Thosar1,
  3. William Gunn1,
  4. Victoria Dean1,
  5. Konstantinos Lontos2,
  6. Marcel Bruchez3,
  7. Patricia Opresko1 and
  8. Greg Delgoffe1
  1. 1University of Pittsburgh, Pittsburgh, PA, USA
  2. 2University of Pittsburgh Medical Center, Pittsburgh, PA, USA
  3. 3Carnegie Mellon University, Pittsburgh, PA, 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.

Abstract

Background Failure of adoptive cell therapies (ACTs) is associated with insufficient persistence within the patient, inability to infiltrate tumor sites, and cell-intrinsic loss of functionality. One of the key factors responsible for T cell dysfunction are metabolic barriers such as nutrient competition, low oxygen tension and damaging byproducts in the tumor microenvironment. This leads to a T cell metabolic deregulation affecting their ability to find and kill cancer cell targets effectively. Therefore, elucidating the metabolic pressures experienced in the TME could provide new therapeutic targets to improve ACTs. Reactive Oxidative Species (ROS) accumulation in the TME have detrimental effects on T cell function and anti-tumor response, although the precise targets of ROS are unclear. There are accumulating data showing that mitochondrial ROS can have profound effects on the telomere status of cells. However, there is little evidence describing the role or oxidative stress on telomere health, or the importance of telomere function in immune cells. Our current study demonstrates that tumor infiltrating lymphocytes (TIL) accumulate DNA damage at telomeres. Furthermore, inducing ROS accumulation at telomeres alone drives T cell dysfunction. Importantly we discovered that alleviating ROS specifically at telomeres improves the response to adoptive cell therapies in a mouse tumor model.

Methods We perform telomeric and centromeric FISH assays to analyze TIL for DNA damage accumulation. We used a chemo-optogenetic FAPS-TAPS to generate oxidative lesions specifically at telomeres.1 We tethered the antioxidant protein GPX1 to the telomere shelterin TRF1 to generate a telomere-guided ROS scavenger.

Results Telo-FISH analysis demonstrates an accumulation of telomeric DNA damage in TIL from B16 mouse tumors shown by the presence of 53BP1 and γH2AX at telomeres (figure 1A). Our data show that mitochondria and telomeric ROS cause the accumulation of DNA damage at telomeres, as well as the development of telomere fragility (figure 1B). These cells ultimately become dysfunctional showing a diminished capability for cytokine production. Importantly, localizing the ROS scavenger GPX1 directly to telomeres reduced telomere fragility and improved the function of therapeutic T cells in the B16 melanoma (figure 1C).

Conclusions Our data suggest that dysfunctional T cells in cancer are not classically senescent, bearing short telomeres, but rather harbor damaged telomeres due to exposure to oxidative stress. Telomeric damage is sufficient to drive a dysfunctional state in newly activated T cells. Protecting telomeres through expression of a telomere-targeted antioxidant protein may preserve T cell function in the tumor microenvironment and drive superior responses to adoptive cell therapies.

Reference

  1. Fouquerel E, Barnes RP, Uttam S, Watkins SC, Bruchez MP, Opresko PL. Targeted and persistent 8-oxoguanine base damage at telomeres promotes telomere loss and crisis. Mol Cell. 2019 Jul 11;75(1):117–130.e6. doi: 10.1016/j.molcel.2019.04.024. Epub 2019 May 14. PMID: 31101499; PMCID: PMC6625854.

Abstract 386 Figure 1

(A) Tumor infiltrating lymphocytes from B16 tumors and spleen sorted on PD1-Tim3 expression. FISH analysis of Telomere (Telo) and DNA damage response elements 53BP1 and γH2AX. (B) Activated T cells expression TRF1-FAP get oxidative damage induced at telomeres by chemoptogenetic method. T cells are arrested and perform metaphase spreads for telomere fragility analysis. (C) Therapeutic T cells co-expressing ROS scavenger GPX and TRF1 (telomere specific) transferred Into B16 tumor bearing mice. Control T cells express control plasmid PMIA.

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