crc-23-0259_fig3.png (1.3 MB)

FIGURE 3 from Rejuvenated iPSC-derived GD2-directed CART Cells Harbor Robust Cytotoxicity Against Small Cell Lung Cancer

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posted on 2024-03-11, 14:20 authored by Shintaro Kinoshita, Midori Ishii, Jun Ando, Takaharu Kimura, Tomoyuki Yamaguchi, Sakiko Harada, Fumiyuki Takahashi, Kazutaka Nakashima, Yozo Nakazawa, Satoshi Yamazaki, Koichi Ohshima, Kazuhisa Takahashi, Hiromitsu Nakauchi, Miki Ando

Robust tumor suppressive effect of GD2-CARrejTs in vivo.A, Graphical abstract of in vivo experiment. H446 cells labeled with FFluc/GFP were intravenously injected into NOG mice (1 × 10⁶ cells/mouse) 4 days before treatment. Mice treated with GD2-2840z-CARTs and GD2-CARrejTs were treated every 2 weeks (three doses). B, Bioluminescence imaging of mice engrafted with H446 cells in three groups: no treatment group (control group, n = 10), GD2-2840z-CARTs group (n = 7), and GD2-CARrejTs group (n = 7). Images of all mice from each group of three independent experiments are shown. C, Tumor burden of each mouse is shown, comparing the in vivo efficacy of GD2-2840z-CARTs and GD2-CARrejTs. Bioluminescence kinetics of all mice from each group of three independent experiments are shown: no treatment group (control group, n = 10), GD2-2840z-CARTs group (n = 7), and GD2-CARrejTs group (n = 7). D, Quantitation of tumor burden on day 35 is shown: no treatment group (n = 8), GD2-2840z-CARTs group (n = 7), and GD2-CARrejTs group (n = 7). ****, P < 0.0001 by one-way ANOVA. E, Kaplan–Meier curves representing percentage survival of groups: no treatment group (n = 10), GD2-2840z-CARTs group (n = 7), and GD2-CARrejTs group (n = 7). ***, P < 0.001; ****, P < 0.0001 by log-rank test. ns, not significant. F, Copy numbers of GD2-CAR transduced T cells in mouse treated with GD2-2840z-CARTs and GD2-CARrejTs on day 35. Error bars represent ± SD. The plots represent three independent experiments. G, Photomicrographs, representative hematoxylin and eosin–stained sections of liver of SCLC-bearing mouse in each group; no treatment, GD2-2840z-CART treatment, and GD2-CARrejT treatment (top row). Infiltrates are bracketed. Human CD3⁺ T-cell infiltrates in liver (bottom row). CD3⁺ T cells are stained in brown (black arrowhead). 4x, 40x: Objective lens. Scale bars, each 20 µm.

Funding

MEXT | Japan Society for the Promotion of Science (JSPS)

Ludwig Family Foundation (The Ludwig Family Foundation)

History

ARTICLE ABSTRACT

Small cell lung cancer (SCLC) is exceptionally aggressive, with limited treatment options. Disialoganglioside (GD2) is highly expressed on SCLC and is considered a good target for chimeric antigen receptor (CAR) T cells (CART). Although GD2-directed CARTs (GD2-CART) exhibit cytotoxicity against various GD2-expressing tumors, they lack significant cytotoxicity against SCLC. To enhance cytotoxicity of GD2-CARTs against SCLC, we introduced GD2-CAR into induced pluripotent stem cells (iPSC)-derived rejuvenated cytotoxic T lymphocytes (GD2-CARrejT). GD2-CARrejTs acted much more strongly against SCLC cells than did GD2-CARTs both in vitro and in vivo. Single-cell RNA sequencing elucidated that levels of expression of TIGIT were significantly lower and levels of expression of genes associated with cytotoxicity were significantly higher in GD2-CARrejTs than those in GD2-CARTs. Dual blockade of TIGIT and programmed death-1 (PD-1) increased the cytotoxicity of GD2-CARTs to some extent, suggesting that low TIGIT and PD-1 expression by GD2-CARrejTs is a major factor required for robust cytotoxicity against SCLC. Not only for robust cytotoxicity but also for availability as “off-the-shelf” T-cell therapy, iPSC-derived GD2-CARrejTs are a promising novel treatment for SCLC. This research introduces iPSC-derived rejuvenated GD2-CARTs (GD2-CARrejT) as a novel approach to combat SCLC. Compared with conventional GD2-CARTs, GD2-CARrejTs with reduced TIGIT and PD-1 expression demonstrate robust cytotoxicity against SCLC and would be a promising therapy for SCLC.