Journal: Respiratory Research

Article Title: TMPRSS2 is a tumor suppressor and its downregulation promotes antitumor immunity and immunotherapy response in lung adenocarcinoma

doi: 10.1186/s12931-024-02870-7

Figure Lengend Snippet: In vivo and in vitro experimental validation of the bioinformatics findings. TMPRSS2-knockdown tumors display increased tumor-infiltrating lymphocytes, expression of immune checkpoint molecules, and sensitization to immune checkpoint inhibitors. a TMPRSS2 knockdown markedly promoted proliferative and invasive abilities of A549 cells. b TMPRSS2 knockdown increased tumor volume and progression in Lewis tumor mouse models. Lewis tumor cells transfected with ShCon or ShTMPRSS2 lentivirus were subcutaneously injected into mice. The tumor volumes were measured every three days from the fifth day to the fifteenth. Data represent mean ± SEM. SEM, standard error of mean. ShTMPRSS2 versus ShCon group, n = 6 for each group, two-tailed Student’s t test, * p < 0.05, ** p < 0.01, *** p < 0.001. c TMPRSS2 knockdown increased MSH6 expression in A549 cells, as evidenced by Western blotting. d TMPRSS2 knockdown enhanced the expression of MHC class I genes ( HLA-A , HLA-B , and HLA-C ) in A549 cells, as evidenced by real-time qPCR. e NK cells co-cultured with TMPRSS2 -knockdown A549 cells showing higher proliferation capacity than NK cells co-cultured with TMPRSS2 -wildtype A549 cells, as evidenced by the EDU proliferation assay. f CD8, CD49b, and PD-L1 immunofluorescence staining in Lewis orthotopic tumors and H-score analysis. ShTMPRSS2 versus shCon group, n = 6 for each group, two-tailed Student’s t test, *** p < 0.001. g - j Comparisons of TNF-α, IFN-γ, PD-1, and LAG3 expression on CD8 + T cells from tumor-infiltrating lymphocytes (TILs) in tumor-bearing mice between TMPRSS2 -knockdown and TMPRSS2 -wildtype group (ShTMPRSS2 versus ShCon group, n = 6 for each group, two-tailed Student’s t test, * p < 0.05, ** p < 0.01, *** p < 0.001). TILs were stained with CD3, CD8, TNF-α, and IFN-γ and were then analyzed by flow cytometry. Lymphocytes were gated according to forward scatter and side scatter. CD3 and CD8 staining was used to identify CD8 + T cells. k - m TMPRSS2 -knockdown tumors formed by subcutaneous injection of Lewis cells, as mentioned in ( b ). shCon and shTMPRSS2 tumor-bearing mice were divided into vehicle and BMS-1 groups. The vehicle and BMS-1 groups of mice were treated with solvent and BMS-1, respectively. k Representative images of tumor-bearing mice shown on the left. The right graph showing the change of tumor size in the tumor-bearing mice over time. Data represent mean ± SEM ( n = 6 for each group, two-tailed Student’s t test, * p < 0.05, ** p < 0.01, *** p < 0.001); Comparison of the volume ratios of mice tumors after and before treatment with BMS-1 between TMPRSS2 -knockdown and TMPRSS2 -wildtype groups (two-tailed Student’s t test, *** p < 0.001). Comparisons of TNF-α ( l ) and IFN-γ ( m ) expression on CD8 + T cells from TILs in tumor-bearing mice ( n = 6 for each group, two-tailed Student’s t test, * p < 0.05, ** p < 0.01, *** p < 0.001)

Article Snippet: After 5 days, when the tumor volume was approximately 4–5 mm3, the mice were randomly divided into six groups, with half of the ShCon and ShTMPRSS2 mice treated with 150 U/L PD1/PDL1 inhibitor BMS-1 (concentration 500 mg/mL; i.p.) (MCE Cat. No. HY-19991) every 3 days.

Techniques: In Vivo, In Vitro, Biomarker Discovery, Knockdown, Expressing, Transfection, Injection, Two Tailed Test, Western Blot, Cell Culture, Proliferation Assay, Immunofluorescence, Staining, Flow Cytometry, Solvent, Comparison