Fig. 1.

PIGL synthetically induces HCC cell death with the combination of lenvatinib plus PD-1 blockade therapy. A, B The strategy using a sublibrary of metabolic enzyme sgRNA to screen the suppressive metabolic enzymes that improve the treatment efficiency of lenvatinib and PD-1 immunotherapy in liver cancer (A). The enrichment of positive and negative candidates in the sgRNA screen (B). C, D Subcutaneous transplantation of Hepa1-6 cells with the indicated genetic manipulation into C57BL/6 mice (n = 6) or C57BL/6 TCRα^−/− mice (n = 6). P, PD-1 antibody. E, F Cell proliferation (E) and cell death (F) were measured in vivo by Ki67 and TUNEL staining, respectively. Scale bar, 200 μm. G The tumors were subjected to single-cell sequencing. The subtypes of immune cells, macrophages (M), CD8⁺ T cells, CD4⁺ T cells, fibroblasts (Fb) and dendritic cells (DC) are shown. H, I The proportions of the T-cell subsets helper T-cell (CD3⁺CD4⁺), cytotoxic T lymphocytes (CD3⁺CD8⁺), regulatory T cells (CD4⁺CD25⁺) (H) and F4/80⁺ macrophages (I) were analyzed using flow cytometry (FCM). Len, lenvatinib; PD-1 Ab, antibody against PD-1; ⁺, positive. C, one-way ANOVA; E-F and H-I, two-tailed Student’s t test. (N.S., not significant; *p < 0.05; **p < 0.01; ***p < 0.001)