Extended Data Fig. 2: Tumor PDL1 analysis and Construction of engineered OA (eOA).

a, Overall survival of patients with melanoma, pancreatic cancer, or glioma with a high or low level of PDL1 analyzed by GEPIA. The source data was isolated from the TCGA. b, Representative images of PDL1 by immunohistochemistry (brown) of tumors from patients with pancreatic cancer, which were used to establish PDX model. The PDX mouse model was treated with saline, intratumorally (i.t.) injected with OA (1 × 10¹⁰ VP), i.v. administered with 1 × 10⁷ CAR-T cells, and combination OA (i.t.) with CAR-T cell (i.v.). Scale bar 100 μm. n = 5 biological independent mice, per tumor was random analyzed from three IHC slices. Score 0 (-, negative), score 1 (+, weak), score 2 (++, moderate), score 3 (+++, strong). c, Gating strategy (upper) and representative flow cytometry plot of PDL1 expression (lower) in EpCAM⁺ tumor cells after treatment. d, PDL1 expression in B16OVA tumor-bearing mice after treatment. e, The quantitative analysis of GFP-positive cells infected for 48 hours with eOA at an increasing MOI at B16OVA and SW1990 cells. f, Western blot (upper) analysis of Cas9 protein level and indel mutation (lower) of PDL1 locus in B16OVA cells 48 hours after infection. g, Western blot analysis (upper) of Cas9 proteins and indel mutation (lower) of human PDL1 locus in SW1990 cells after 48 hours of eOA infection at increasing MOIs. h, Schematic illustration of eOA-PLK1 design and indel mutations of PLK1 locus in B16F10 cells after 48 hours of eOA infection at increasing MOIs. i, Quantitative analysis of viral copy numbers in different cell lines by qRT-PCR after 48 hours of infection with eOA. j, Cell viability of B16OVA, L929, SW1990, and HUVEC cells with the indicated treatments for 72 h. k, Western blot analysis of PDL1 level in B16OVA cells after treatment for 72 h. l. Cell viability of B16OVA cells after treatment for 72 h. m. Illustration of eOA treatment in C57BL/6 mice bearing B16OVA tumor (Upper). T7E1 (lower left) and western blot (lower right) assays for the analysis of PDL1 levels after treatment. n. Tumor volume (left) and survival curves after the indicated treatment (n = 5 biologically independent mice). Red arrows (f-h) denote cleaved DNA fragments. Data are presented as the mean ± s.d. (b-e,i,l) and s.e.m. (n). n = 3 (e,i), n = 5 (b, c, d, j), and n = 6 (l) biological independent samples. One-way ANOVA with Tukey’s post hoc analysis was used in (c, i, l), and two-way ANOVA with Bonferroni post hoc test was used in (n), and log-rank (Mantel-Cox) test used in survival curves (a,n), with P values indicated on the graphs.

Source data