Costimulation of γδ TCR and TLR7/8 promotes Vδ 2 T-cell antitumor activity by modulating mTOR pathway and APC function

Huaishan Wang, Hui Chen, Shujing Liu, Jie Zhang, Hezhe Lu, Rajasekharan Somasundaram, Robin Choi, Gao Zhang, Lingling Ou, John Scholler, Shifu Tian, Liyun Dong, Guo Yeye, Lili Huang, Thomas Connelly, Ling Li, Alexander Huang, Tara C. Mitchell, Yi Fan, Carl H. JuneGordon B. Mills, Wei Guo, Meenhard Herlyn, Xiaowei Xu

Research output: Contribution to journalArticlepeer-review

8 Scopus citations


Background Gamma delta (γδ) T cells are attractive effector cells for cancer immunotherapy. Vδ 2 T cells expanded by zoledronic acid (ZOL) are the most commonly used γδT cells for adoptive cell therapy. However, adoptive transfer of the expanded Vδ 2 T cells has limited clinical efficacy. Methods We developed a costimulation method for expansion of Vδ 2 T cells in PBMCs by activating γδT-cell receptor (γδ TCR) and Toll-like receptor (TLR) 7/8 using isopentenyl pyrophosphate (IPP) and resiquimod, respectively, and tested the functional markers and antitumoral effects in vitro two-dimensional two-dimensional and three-dimensional spheroid models and in vivo models. Single-cell sequencing dataset analysis and reverse-phase protein array were employed for mechanistic studies. Results We find that Vδ 2 T cells expanded by IPP plus resiquimod showed significantly increased cytotoxicity to tumor cells with lower programmed cell death protein 1 (PD-1) expression than Vδ 2 T cells expanded by IPP or ZOL. Mechanistically, the costimulation enhanced the activation of the phosphatidylinositol 3-kinase (PI3K)-protein kinase B (PKB/Akt)-the mammalian target of rapamycin (mTOR) pathway and the TLR7/8-MyD88 pathway. Resiquimod stimulated Vδ 2 T-cell expansion in both antigen presenting cell dependent and independent manners. In addition, resiquimod decreased the number of adherent inhibitory antigen-presenting cells (APCs) and suppressed the inhibitory function of APCs by decreasing PD-L1 and cytotoxic T-lymphocyte-associated protein 4 (CTLA-4) expression in these cells during in vitro Vδ 2 T-cell expansion. Finally, we showed that human Vδ 2 T cells can be expanded from PBMCs and spleen of humanized NSG mice using IPP plus resiquimod or ZOL, demonstrating that humanized mice are a promising preclinical model for studying human γδT-cell development and function. Conclusions Vδ 2 T cells expanded by IPP and resiquimod demonstrate improved anti-tumor function and have the potential to increase the efficacy of γδT cell-based therapies.

Original languageEnglish (US)
Article numbere003339
JournalJournal for immunotherapy of cancer
Issue number12
StatePublished - Dec 22 2021


  • adjuvants
  • adoptive
  • costimulatory and inhibitory T-cell receptors
  • immunological
  • immunotherapy
  • melanoma

ASJC Scopus subject areas

  • Immunology and Allergy
  • Immunology
  • Molecular Medicine
  • Oncology
  • Pharmacology
  • Cancer Research


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