FIGURE 2 from Exercise-induced β2-adrenergic Receptor Activation Enhances the Antileukemic Activity of Expanded γδ T-Cells via DNAM-1 Upregulation and PVR/Nectin-2 Recognition

• DOI: 10.1158/2767-9764.25810095.v1
• Publication Date: 2024-05-13T14:22:36Z
• Authors: Forrest L. Baker; Kyle A. Smith; Preetesh L. Mylab...; Tiffany M. Zúñiga; Douglass M. Diak; Helena Batatinha; Grace M. Niemiro; Michael D. Seckeler; Charles R. Pedlar; Daniel P. O'Connor; Jamie Colombo; Emmanuel Katsanis; Richard J. Simpson

<p>Acute exercise enhances the antileukemic activity of <i>ex vivo</i> expanded Vγ9Vδ2+ T-cells <i>in vitro</i> and <i>in vivo</i>. <b>A,</b> Schematic of the experimental design for <i>in vitro</i> and <i>in vivo</i> experiments. <b>B,</b> The total number (1 × 10⁶) of Vγ9Vδ2+ cells isolated before (REST) and during the final 5 minutes of exercise (EX) at day 0, and the total number of Vγ9Vδ2+ T-cells generated in the expanded cell products after stimulation with ZOL+IL2 for 14 days. The number of Vγ9Vδ2+ cells generated at day 14 relative to the number of γδ cells in the PBMC fractions at day 0. The cellular composition of the expanded Vγ9Vδ2+ T-cell products at day 14. The proportions of naïve (N), CM, EM, and CD45RA+ EMRA cells among Vγ9Vδ2+ T-cells after 14 days of expansion (<i>n</i> = 10). <b>C,</b> The <i>in vitro</i> anti-tumor activity of <i>ex vivo</i> expanded Vγ9Vδ2+ T-cells against K562, Daudi, and U266 with and without ZOL sensitization. Target cells were exposed to 5 µmol/L of ZOL for 20 hours and cytotoxicity of Vγ9Vδ2+ cells was assessed via flow cytometry–based assays. <b>D,</b> To determine the <i>in vivo</i> GvL effect of exercise expanded Vγ9Vδ2+, NSG-IL15 mice were injected with REST or EX expanded Vγ9Vδ2+ (10 × 10⁶) and challenged with 1 × 10⁶ luciferase tagged human chronic myeloid leukemia cells (K562-luc). A subset of mice in each group were also provided weekly injections of ZOL (REST+ZOL; EX+ZOL; VEHICLE +ZOL). The BLI (photons/second) scores, TFS, change in BW, and overall probability of survival (OS) after injection of expanded Vγ9Vδ2+ T-cells with or without ZOL sensitization (<i>n</i> = 8–11/group<b>)</b>. BLI significant difference from EX+ZOL and EX conditions indicated by * and ^#, respectively (all conditions were significantly different from vehicle controls). TFS, BW, and OS significant difference from vehicle controls were indicated by *. <b>E,</b> Representative bioluminescence images of leukemia-bearing mice that received (from left to right) vehicle, vehicle + ZOL, REST expanded Vγ9Vδ2+ T-cells, REST expanded Vγ9Vδ2+ T-cells + ZOL, EX expanded Vγ9Vδ2+ T-cells, and EX expanded Vγ9Vδ2+ T-cells + ZOL. BLI intensity on a scale from low (purple) to high (red). Data are represented as mean ± SEM; *, <i>P</i> < 0.05; **, <i>P</i> < 0.01; ***, <i>P</i> < 0.001; ****, <i>P</i> < 0.0001 by Student two-tailed paired <i>t</i> test (B); repeated measures two-way ANOVA or LMM with Bonferroni <i>post hoc</i> test (C–D); log-rank (Mantel–Cox) test (D).</p>