Journal: Scientific Reports

Article Title: SHR1032, a novel STING agonist, stimulates anti-tumor immunity and directly induces AML apoptosis

doi: 10.1038/s41598-022-12449-1

Figure Lengend Snippet: SHR1032 specifically and potently activates STING signaling pathway. IRF luciferase reporter assays in ( a ) THP1-STING-R232, THP1-STING-H232, THP1-STING-HAQ, and THP1-STING-KO cells. Cells were stimulated for 24 h with the indicated STING compounds and assessed for luciferase activity. IFNβ production and activation of cGAS-STING signaling pathway in ( b ) human THP1-STING-R232, human PBMCs from a donor homozygous for the R232 variant, cynomolgus PBMC, and mouse RAW164.7 macrophage cells were treated with STING agonists for 5 h. IFNβ in media was quantified by human or mouse ELISA kits respectively. ( c ) THP1-STING-R232 cells and mouse RAW164.7 macrophage cells were stimulated with 40 µM of ADU-S100 or 10 µM of SHR1032 for the indicated time points. The amount of pTBK1, total TBK1, STING and α-tubulin was measured by western blotting.

Article Snippet: Human monocyte THP-1 Dual cells (InvivoGen), THP1-Dual™ KO-STING Cells (InvivoGen) and THP1-Dual KI-hSTING-R232 cells (InvivoGen) were cultured in RPMI 1640, 2 mM L-glutamine, 25 mM HEPES, 10% heat-inactivated fetal bovine serum, 100 μg/ml Normocin, Pen-Strep, and 10 µg/ml of blasticidin, 100 μg/ml of Zeocin.

Techniques: Luciferase, Activity Assay, Activation Assay, Variant Assay, Enzyme-linked Immunosorbent Assay, Western Blot

Journal: Scientific Reports

Article Title: SHR1032, a novel STING agonist, stimulates anti-tumor immunity and directly induces AML apoptosis

doi: 10.1038/s41598-022-12449-1

Figure Lengend Snippet: SHR1032 inhibits AML cell growth and displays specific activation of STING signaling pathway in AML cell lines. ( a – d ) AML cells were treated with STING agonists and standard of care cytarabine, and cell viability was measured by CellTiter Glo in ( a ) THP1-STING-R232, ( b ) THP1-STING-KO, ( c ) MV-4-11 and ( d ) MOLM-16. ( e ) MV-4-11, MOLM-16 and THP1-STING-KO cells were stimulated with 1 µM of SHR1032 for the indicated time points. The amount of pTBK1, total TBK1, pSTING, total STING and α-tubulin was measured by Western blot. pSTING phosphorylated STING.

Article Snippet: Human monocyte THP-1 Dual cells (InvivoGen), THP1-Dual™ KO-STING Cells (InvivoGen) and THP1-Dual KI-hSTING-R232 cells (InvivoGen) were cultured in RPMI 1640, 2 mM L-glutamine, 25 mM HEPES, 10% heat-inactivated fetal bovine serum, 100 μg/ml Normocin, Pen-Strep, and 10 µg/ml of blasticidin, 100 μg/ml of Zeocin.

Techniques: Activation Assay, Western Blot

Journal: Scientific Reports

Article Title: SHR1032, a novel STING agonist, stimulates anti-tumor immunity and directly induces AML apoptosis

doi: 10.1038/s41598-022-12449-1

Figure Lengend Snippet: Cell apoptosis induced by SHR1032 in AML cell lines is independent of type I interferon. ( a – d ) RNA-Seq analysis of THP1-STING-R232, THP1-STING-KO and MV4-11 AML cells treated with 1 µM of SHR1032 for 4 h. ( a ) Heatmaps showing gene expression patterns of upregulated (left) or downregulated (right) DEGs with SHR1032 treatment. ( b ) Venn diagram showing genes up- (above) or down-regulated (below) genes in MV4-11 and THP1-STING-R232 cells with SHR1032 treatment. ( c ) Gene Ontology analysis of those genes up-regulated in both MV4-11 and THP1-STING-R232 cells. ( d ) ANNEXIN V staining analysis with flow cytometry in MV4-11, MOLM-16, THP1-STING-WT and THP1-STING-KO cells. ( e ) Cell viability analysis by CellTiter Glo in THP1-IRFAR2-WT or -KO cells.

Article Snippet: Human monocyte THP-1 Dual cells (InvivoGen), THP1-Dual™ KO-STING Cells (InvivoGen) and THP1-Dual KI-hSTING-R232 cells (InvivoGen) were cultured in RPMI 1640, 2 mM L-glutamine, 25 mM HEPES, 10% heat-inactivated fetal bovine serum, 100 μg/ml Normocin, Pen-Strep, and 10 µg/ml of blasticidin, 100 μg/ml of Zeocin.

Techniques: RNA Sequencing Assay, Expressing, Staining, Flow Cytometry