Citation

  • Authors: Jneid B. et al.
  • Year: 2023
  • Journal: Sci Immunol 8 eabn6612
  • Applications: in vitro / DNA / PEIpro
  • Cell type: HEK-293T
    Description: Human embryonic kidney Fibroblast
    Known as: HEK293T, 293T

Method

cGAMP-VLP production for in vivo use A total of 7.5 million 293T cells were plated in a 150-cm2 cell culture flask and incubated overnight. One batch of cGAMP-VLP was made from four flasks. The following day, each flask was transfected with 13 μg of pVAX1-cGAS, 8.1 μg of HIV-1 psPAX2, 3.3 μg of pVAX1-VSVG-INDIANA2, and 50 μl of PEIpro according to the manufacturer’s instructions. The transfection mixes were prepared in Opti-MEM. The morning after transfection, the medium was changed with 52 ml of warm VLP production medium [293T culture medium with 10 mM Hepes and gentamicin (50 μg/ml)]. One day later, the cGAMP-VLP–containing supernatant was harvested from the cells, centrifuged for 10 min at 200g at 4°C, and filtered through 0.45-μm nylon mesh filters.

Abstract

T cells that recognize tumor antigens are crucial for mounting antitumor immune responses. Induction of antitumor T cells in immunogenic tumors depends on STING, the intracellular innate immune receptor for cyclic guanosine monophosphate-adenosine monophosphate (cGAMP) and related cyclic dinucleotides (CDNs). However, the optimal way to leverage STING activation in nonimmunogenic tumors is still unclear. Here, we show that cGAMP delivery by intratumoral injection of virus-like particles (cGAMP-VLP) led to differentiation of circulating tumor-specific T cells, decreased tumor regulatory T cells (Tregs), and antitumoral responses that synergized with PD1 blockade. By contrast, intratumoral injection of the synthetic CDN ADU-S100 led to tumor necrosis and systemic T cell activation but simultaneously depleted immune cells from injected tumors and induced minimal priming of circulating tumor-specific T cells. The antitumor effects of cGAMP-VLP required type 1 conventional dendritic cells (cDC1), whereas ADU-S100 eliminated cDC1 from injected tumors. cGAMP-VLP preferentially targeted STING in dendritic cells at a 1000-fold smaller dose than ADU-S100. Subcutaneous administration of cGAMP-VLP showed synergy when combined with PD1 blockade or a tumor Treg-depleting antibody to elicit systemic tumor-specific T cells and antitumor activity, leading to complete and durable tumor eradication in the case of tumor Treg depletion. These findings show that cell targeting of STING stimulation shapes the antitumor T cell response and identify a therapeutic strategy to enhance T cell-targeted immunotherapy.

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