Journal: Journal for Immunotherapy of Cancer
Article Title: Cancer cell genotype associated tumor immune microenvironment exhibits differential response to therapeutic STING pathway activation in high-grade serous ovarian cancer
doi: 10.1136/jitc-2022-006170
Figure Lengend Snippet: Pten -deficient ovarian cancer cells alter their tumor immune microenvironment towards a suppressive state via polarizing macrophages to M2-like phenotype. Total RNA from untreated tumors of different genotypes were subjected to NanoString gene expression profiling using the PanCancer immune gene panel displayed as a (A) volcano plot showing differential expression pattern of genes associated with macrophage function. (B) Proportion of cells derived from ascites of mice injected with ID8- Trp53 −/− ; Pten −/− and ID8- Trp53 −/− ; Brca1 −/− cells expressing CD45+CD11b+F4/80+cells. (C) MCP-1 chemokine levels derived from the ascites fluid of untreated mice injected with ID8- Trp53 −/− ; Pten −/− and ID8- Trp53 −/− ; Brca1 −/− cells. (D) Transwell migration assay of IC21 peritoneal macrophage cells stimulated (from left to right) with media control (Dulbecco’s Modified Eagle’s Medium), positive control (20 ng/mL MCP-1), or 72 hours conditioned media of ID8- Trp53 −/− ; Pten −/− and ID8- Trp53 −/− ; Brca1 −/− cells. Stimulated for 16 hours; images 20x magnification. Flow cytometric analysis of IC21 macrophage proportions expressing (E) CD45+F4/80+CD206+ post-stimulation with concentrated ascites derived from mice injected with different genotypes of HGSC cells. US, unstimulated IC21 cells; M1 positive control represented by IC21 cells stimulated with IFN-ɣ (50 ng/mL) and LPS (100 ng/mL) in complete media; M2 positive control represented by IC21 cells stimulated with IL-4 (10 ng/mL) and IL-10 (20 ng/mL) in complete media. Averages of triplicate wells/experiment for three repeated experiments displayed. Mann-Whitney non-parametric test was used for (B–C) *p<0.05 **p<0.005, ***p<0.001, ****p<0.0001. One-way analysis of variance applied for D and E. Mean±SD. HGSC, high-grade serous ovarian cancer; IFN, interferon; IL, interleukin; MCP-1, macrophage chemoattractant protein 1; ns, not significant; PTEN, phosphatase and tensin homolog.
Article Snippet: IC21 murine peritoneal macrophage cell line was obtained from American Type Culture Collection (Manassas, Virginia, USA) and cultured in RPMI 1640 (Sigma-Aldrich, Ontario, Canada) supplemented with 10% FBS and 1% penicillin/streptomycin.
Techniques: Gene Expression, Quantitative Proteomics, Derivative Assay, Injection, Expressing, Transwell Migration Assay, Control, Modification, Positive Control, MANN-WHITNEY
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