# TIGIT Blockade Potentiates the Anti-Leukemic Activity of Exercise-Mobilized Donor Lymphocytes and Expanded γδ T-Cells

## TL;DR

Exercise improves the effectiveness of immune cell therapies for leukemia when combined with TIGIT checkpoint inhibition.

## Contribution

Exercise primes donor lymphocytes and γδ T-cells to respond better to TIGIT blockade, enhancing their anti-leukemic activity.

## Key findings

- Exercise increases TIGIT and PD-1 expression on CD8+ and γδ T-cells, improving their cytotoxicity.
- TIGIT blockade amplifies the anti-leukemic activity of exercise-mobilized PBMCs and γδ T-cells.
- In mice, TIGIT-treated exercise-expanded γδ T-cells improved tumor suppression and survival.

## Abstract

Donor lymphocyte infusions and gamma delta T-cells are types of adoptive cell therapy that help treat leukemic relapse after stem cell transplantation, but their activity is inhibited by the tumor microenvironment. Immune checkpoint inhibitors can restore immune function, yet their effectiveness in blood cancers has been modest. Additionally, exercise is an emerging physiological strategy to enhance the composition and function of therapeutic immune cells. This study investigated whether exercise could improve the anti-leukemic activity of donor lymphocytes and expanded gamma delta T-cells when combined with the immune checkpoint inhibitor, TIGIT. Herein we demonstrate that exercise primes donor lymphocytes and gamma delta T-cells for enhanced responsiveness to checkpoint inhibition, resulting in improved leukemia cell killing. These results highlight the integration of exercise as a safe adjunct to immune checkpoint therapy to optimize cellular immunotherapy for leukemia.

Background: Donor lymphocyte infusion (DLI) is commonly used to prevent or treat leukemic relapse following allogeneic hematopoietic cell transplantation; however, efficacy is limited by immune exhaustion, checkpoint-mediated inhibition, and the risk of graft-versus-host disease (GvHD). Gamma delta (γδ) T-cells represent a promising “off-the-shelf” adoptive cell therapy (ACT) with favorable safety and MHC-independent cytotoxicity, yet their function is similarly constrained by the leukemic tumor microenvironment (TME). Acute exercise mobilizes cytotoxic lymphocyte subsets, and is an emerging strategy to enhance cellular immunotherapies, including DLI and expanded γδ T-cells. This study examined how exercise-mobilized lymphocytes and exercise-expanded γδ T-cells interact with TIGIT blockade to improve anti-leukemic activity. Methods: Healthy participants completed an acute cycling bout, after which peripheral blood mononuclear cells (PBMCs) and ex vivo expanded γδ T-cells were phenotyped and cytotoxicity was determined against leukemia cells with TIGIT checkpoint inhibition. The therapeutic relevance of combining TIGIT blockade with rest- or exercise-expanded γδ T-cells was further evaluated in NSG-IL15 mice challenged with K562-luc leukemia. Results: Acute exercise increased circulating CD8+ and γδ T-cells with higher TIGIT and PD-1 expression. Exercise-expanded γδ T-cells maintained increased PD-1 and TIGIT expression and exhibited increased co-expression of DNAM-1 and TIGIT. Exercise mobilized PBMCs and exercise-expanded γδ T-cells demonstrated enhanced cytotoxicity, further amplified by TIGIT blockade. In vivo, TIGIT-treated exercise-expanded γδ T-cells modestly improved tumor suppression and prolonged tumor-free survival compared to untreated controls. Conclusions: Exercise primes DLI and γδ T-cell products for enhanced responsiveness to TIGIT checkpoint inhibition. Targeting TIGIT likely augments DNAM-1 dependent cytotoxicity and improves anti-leukemic activity, supporting the integration of exercise-enhanced DLI and γδ T-cell therapies with immune checkpoint blockade as a safe strategy to improve relapse control in leukemia.