Journal: Cancer Research

Article Title: Adipocyte Precursor-Derived NRG1 Promotes Resistance to FGFR Inhibition in Urothelial Carcinoma

doi: 10.1158/0008-5472.CAN-23-1398

Figure Lengend Snippet: Adipocyte precursors promote resistance against erdafitinib in bladder cancer cell lines. A, Western blot analysis of FGFR3 in four bladder cancer cell lines: RT4, RT112, TSCCUP, and T24. β-Actin served as a loading control. B, Proliferation analysis of five bladder cancer cell lines RT4, RT112, TSCCUP, T24, and MB49 treated with vehicle (Veh) or erdafitinib (1, 10, 100, 1,000 nmol/L). Crystal violet staining was done on day 7. Three biological replicates were performed. Data are represented as mean ± SD. C, Western blot analysis of pAKT and pERK1/2 in RT4 and RT112 cells treated with DMSO (Veh) or erdafitinib (10 and 100 nmol/L) for 16 hours. β-Actin served as a loading control. D, Schematic diagram of the proliferation assay performed to investigate the effect of CM of different stromal cells on erdafitinib response. E, Proliferation analysis of RT4 and RT112 cells treated with 10 nmol/L erdafitinib in media control (ctrl) or CM of 3T3-L1 cells or hADSC. Crystal violet staining was performed on day 7. Data are normalized to cells treated with vehicle. Four biological replicates were performed. Data are represented as mean ± SD. F, Western blot analysis of pAKT and pERK1/2 in RT4 and RT112 cells treated with 10 nmol/L erdafitinib in media control or CM of 3T3-L1 cells. β-Actin served as a loading control. G, Proliferation analysis of RT4 cells and RT112 cells treated with 10 nmol/L erdafitinib in CM of 3T3-L1 cells or heat-inactivated (HI) CM of 3T3-L1. Data were normalized to cells treated with vehicle. Crystal violet staining was performed on day 7. Three biological replicates were performed. Data are represented as mean ± SD. Two-way ANOVA was used for statistical analysis. *, P < 0.05; **, P < 0.01; ***, P < 0.001; ****, P < 0.0001.

Article Snippet: MB49 cell line was purchased from Addexbio, RT112 cell line was purchased from DSMZ (German Collection of Microorganisms and Cell Cultures), and human adipose-derived stem cells (hADSC) were purchased from Lonza.

Techniques: Western Blot, Staining, Proliferation Assay

Journal: International Journal of Molecular Sciences

Article Title: Combination of Cisplatin and Irradiation Induces Immunogenic Cell Death and Potentiates Postirradiation Anti–PD-1 Treatment Efficacy in Urothelial Carcinoma

doi: 10.3390/ijms22020535

Figure Lengend Snippet: Concurrent anti–PD-1 treatment facilitated abscopal effects induced by a single dose of irradiation in MB49 tumor-bearing mice. ( A ) Scheme for tumor inoculation and treatments. Six- to seven-week-old mice are inoculated subcutaneously with MB49 cells in the left hindlimb and in the right flank, and seven days later, receive irradiation with a single fraction of 10 Gy to the left hindlimb and anti–PD-1 treatment (or its isotype controls). ( B ) MB49 tumor growth curves of irradiated tumors in the left hindlimb (left) and nonirradiated tumors in the right flank (right). ( C ) Survival curves of mice. Log-rank test is used to compare survival curves. All data are shown as the mean ± standard error of the mean (SEM). Asterisks indicate p -values comparing two groups, as indicated in the figure. * p < 0.05; ** p < 0.01; *** p < 0.001.

Article Snippet: MB49, a mouse bladder UC cell line, was purchased from Merck KGaA (Darmstadt, Germany).

Techniques: Irradiation

Journal: International Journal of Molecular Sciences

Article Title: Combination of Cisplatin and Irradiation Induces Immunogenic Cell Death and Potentiates Postirradiation Anti–PD-1 Treatment Efficacy in Urothelial Carcinoma

doi: 10.3390/ijms22020535

Figure Lengend Snippet: The combination of cisplatin and irradiation potentiated the efficacy of postirradiation anti–PD-1 treatment in MB49 tumor-bearing mice. ( A ) Scheme for tumor inoculation, treatments, and T cell analysis. For the CRT model (top), six- to seven-week-old mice are injected with MB49 cells in the left hindlimb, and seven days later, receive cisplatin at 3 mg/kg and irradiation with a single fraction of 10 Gy to the left hindlimb. The mice are injected with MB49 cells in the right flank 14 days after irradiation, and seven days later, receive anti–PD-1 treatment (or its isotype controls). For the Non-CRT model (bottom), only a right flank tumor is established in the mice, and cisplatin and irradiation are not given. ( B ) MB49 tumor growth curves of irradiated tumors in the left hindlimb (left) and nonirradiated tumors in the right flank (right). ( C ) Survival curves of mice. The log-rank test is used to compare survival curves. ( D – G ) Proportions of CD45 + cells in the live cells ( D ), CD3 + cells in the CD45 + subpopulation ( E ), CD8 + CD4 − cells in the CD3 + subpopulation ( F ), and IFNγ + cells in the CD8 + CD4 − subpopulation ( G ) in single-cell suspensions of the irradiated and nonirradiated tumors on day seven in mice treated with CRT/postirradiation anti–PD-1 treatment compared to those of the nonirradiated tumors on day seven in mice treated with anti–PD-1 treatment alone ( n = 5/group). All data are shown as the mean ± standard error of the mean (SEM). Asterisks indicate p -values comparing two groups, as indicated in the figure. NS, not significant; * p < 0.05; ** p < 0.01; *** p < 0.001.

Article Snippet: MB49, a mouse bladder UC cell line, was purchased from Merck KGaA (Darmstadt, Germany).

Techniques: Irradiation, Cell Analysis, Injection

Journal: International Journal of Molecular Sciences

Article Title: Combination of Cisplatin and Irradiation Induces Immunogenic Cell Death and Potentiates Postirradiation Anti–PD-1 Treatment Efficacy in Urothelial Carcinoma

doi: 10.3390/ijms22020535

Figure Lengend Snippet: The combination of cisplatin and irradiation increases HMGB1 protein secretion and cell surface expression of calreticulin protein in MB49 cells. ( A ) MB49 cells are treated with cisplatin at 0.6 mg/L and/or irradiated with a single fraction of 10 Gy. Medium is collected two and five days after treatment, and HMGB1 protein expression levels in each medium are examined in duplicate by ELISA ( n = 3/group). ( B ) MB49 cells are treated with cisplatin at 0.6 mg/L and/or irradiated with a single fraction of 10 Gy. Cells are collected, and cell surface calreticulin protein expression levels are examined by flow cytometry ( n = 3/group). ( C , D ) Seven days after inoculation MB49 cells in the left hindlimb, mice are treated with cisplatin at 3 mg/kg and/or irradiation with a single fraction of 10 Gy. Tumor sections before and after CRT ( n = 3/group) are stained for HMGB1 ( C ) and calreticulin ( D ). Left, representative images; right, comparison of the mean (± standard error of the mean [SEM]) number of HMGB1- or calreticulin-positive cells counted in five fields of view (FOV). All data are shown as the mean ± SEM. Asterisks indicate p values comparing two groups, as indicated in the figure. NS, not significant; * p < 0.05; ** p < 0.01.

Article Snippet: MB49, a mouse bladder UC cell line, was purchased from Merck KGaA (Darmstadt, Germany).

Techniques: Irradiation, Expressing, Enzyme-linked Immunosorbent Assay, Flow Cytometry, Staining, Comparison