Ex vivo pretreatment with venetoclax boosts antileukemic efficacy of therapeutic γδ T cells in AML models
Xingchi Chen, Lin Zhang, Bingbing Yan, Yinqiang Sui, Weiwei Ma, Hui Zhao, Yining Wang, Kepeng Yang, Jiewen Ma, Baolin Tang, Yonghui Zhang, Xiaoyu Zhu
Xingchi Chen, Lin Zhang, Bingbing Yan, Yinqiang Sui, Weiwei Ma, Hui Zhao, Yining Wang, Kepeng Yang, Jiewen Ma, Baolin Tang, Yonghui Zhang, Xiaoyu Zhu
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Research Article Hematology Immunology

Ex vivo pretreatment with venetoclax boosts antileukemic efficacy of therapeutic γδ T cells in AML models

Abstract

Ex vivo engineering strategies for adoptive αβ T cell therapies increasingly use pharmacological modulation to improve survival, expansion, and antitumor activity. Short-term exposure to the B cell lymphoma-2 (BCL-2) inhibitor venetoclax during αβ T cell manufacturing enhances apoptotic priming and effector persistence, suggesting a route to strengthen other T cell lineages. γδ T cells share cytotoxic properties with αβ T cells but recognize targets independently of major histocompatibility complex and show low alloreactivity, supporting off-the-shelf use in acute myeloid leukemia (AML). Whether such conditioning benefits γδ T cells was unknown. Here, we show that ex vivo venetoclax pretreatment enhances the antileukemic efficacy of therapeutic γδ T cells and chimeric antigen receptor (CAR) γδ T cells. Venetoclax-pretreated γδ T cells displayed increased cytotoxicity and proliferation with reduced exhaustion, yielding superior control of AML blasts and xenografts. These functional gains coincided with elevated mitochondrial content and a fatty acid oxidation metabolic profile. In vivo, venetoclax-pretreated γδ T cells achieved durable disease suppression, and the same conditioning improved CAR γδ T cell efficacy. Together, these results show that short-term BCL-2 inhibition enhances γδ T cell cytotoxicity and persistence. Incorporating venetoclax pretreatment into γδ T cell manufacturing may improve therapeutic efficacy and inform next-generation γδ T cell therapies for AML.

Authors

Xingchi Chen, Lin Zhang, Bingbing Yan, Yinqiang Sui, Weiwei Ma, Hui Zhao, Yining Wang, Kepeng Yang, Jiewen Ma, Baolin Tang, Yonghui Zhang, Xiaoyu Zhu

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Figure 2

Venetoclax enhances the antileukemic activity of γδ T cells.

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Venetoclax enhances the antileukemic activity of γδ T cells. (A) Schemat...
(A) Schematic workflow for screening compounds that enhance γδ T cell function. (B) Killing efficiency of venetoclax-pretreated γδ T cells (ven-γδ T cells) against KG-1α cells. γδ T cells from 7 healthy donors were pretreated with dimethyl sulfoxide (DMSO) or 1 μM venetoclax for 18 hours and cocultured with KG-1α cells at a 1:1 effector-to-target (E:T) ratio for 14–18 hours. Cytotoxicity was determined by luminescence. Each paired symbol represents 1 donor (n = 7). (C) Dose-dependent killing efficiency of ven-γδ T cells against MV4-11 and OCI-AML3 cells at a 1:1 E:T ratio (n = 3 biological replicates). Data represent mean ± SD and were analyzed using repeated measures 1-way ANOVA with Dunnett’s multiple comparisons test. (D) Representative flow cytometry plots and quantification of ven-γδ T cell cytotoxicity against primary AML blasts from relapsed or newly diagnosed patients (n = 12 donors). Representative plots from 1 donor sample are shown. Cytotoxicity was assessed by annexin V/ 7-aminoactinomycin D (7-AAD) staining after 12 hours of coculture at a 1:1 E:T ratio. Data were analyzed using Wilcoxon’s matched pairs signed-rank test. (E) Schematic of cytotoxic assay using γδ T cells freshly derived from peripheral blood mononuclear cells (PBMCs) of relapsed or newly diagnosed patients with AML. (F) Cytotoxicity of patient-derived ven-γδ T cells against autologous AML blasts from relapsed AML patients (n = 5) and newly diagnosed AML patients (n = 4), assessed after 12 hours of coculture at a 4:1 E:T ratio. Data were analyzed using 2-tailed paired Student’s t test (B and F). *P < 0.05; **P < 0.01; ***P < 0.001; ****P < 0.0001.