Skip to main content
Fig. 5 | Cell & Bioscience

Fig. 5

From: In vivo genome-wide CRISPR screening identifies ZNF24 as a negative NF-κB modulator in lung cancer

Fig. 5

Combinational inhibition of KRAS, NF-κB and PD-1 effectively shrinks KrasG12D/ZNF24−/− lung cancers. A Combination of NF-κB inhibitor (BAY11-7082) and/or KRAS inhibitor (BI3406) effectively inhibited proliferation of ZNF24 knockdown EKVX cells. EKVX-shZNF24 (1000 cells) were seeded in 96-well plates. Cells were treated with BAY (2 μM) and/or BI3406 (1 μM) for 5 days. Cell viability was checked with CCK8 after drug treatment. B NF-κB inhibitor (BAY11-7082) and/or KRAS inhibitor (BI3406) effectively inhibited colony forming ability of ZNF24 knockdown EKVX cells. EKVX-shZNF24 (1000 cells) seeded in 6-well plates. Cells were treated with BAY (2 μM) and/or BI3406 (1 μM) for 2 weeks before colony quantification. C Statistics of colony number of (B). D Induction of immunogenic cell death by NF-κB inhibitor (BAY11-7082) and/or KRAS inhibitor (BI3406) in EKVX-shZNF24 cells. EKVX-shZNF24 cells were treated with BAY (2 μM) and/or BI3406 (1 μM) for 24 h. Expression of CRT and HSP70 was determined by flow cytometry. E, F Statistics of (D). G Combinational treatment with BAY11-7082, BI3406 and PD-1 antibody (designated BAY + BI + PD-1) effectively shrinks lung tumor in K-sgZNF24 mice. Lsl-KrasG12D mice were treated with the recombinant lenti-virus co-expressed Cre and CRISPR/Cas9 by nasal drip. Mice were treated with BAY11-7082 (20 mg/kg/day, intraperitoneal), BI3406 (25 mg/kg/day, gavage) and PD-1 antibody (5 mg/kg, every other day, intraperitoneal). Tumor burdens were documented with CT. H Quantification of tumor burden of combinationally treated mice of (G). I Representative images of Hematoxylin and eosin (H&E) stained lung tissues obtained from different treatment groups. J Quantification of tumor numbers of mice of (G). K Relative tumor volume of mice of (G). L–O Combinational treatment with BAY11-7082, BI3406 and PD-1 antibody (designated BAY + BI + PD-1) activated T cell mediated immunity in tumor microenvironment. Expression of TIM-3 of tumor-infiltrating CD8 + T cells and CD4 + T cells were determined by flow cytometry. P, Q Combinational treatment with BAY11-7082, BI3406 and PD-1 antibody (designated BAY + BI + PD-1) activated expression of effector cytokine in tumor infiltrating CD8 + T cells. Tumor-infiltrating CD8 + T cells of (G) were intracellularly stained for FACS analysis of expression of IL-2. Data are means ± SEM of three independent experiments. *p < 0.05, **p < 0.01, ***p < 0.001 (student’s t-test)

Back to article page