Additional file 1 of Targeting FGFRs by pemigatinib induces G1 phase cell cycle arrest, cellular stress and upregulation of tumor suppressor microRNAs
Creators: name: Pace, Angelica
name: Scirocchi, Fabio
name: Napoletano, Chiara
name: Zizzari, Ilaria Grazia
name: Po, Agnese
name: Megiorni, Francesca
name: Asquino, Angela
name: Pontecorvi, Paola
name: Rahimi, Hassan
name: Marchese, Cinzia
name: Ferretti, Elisabetta
name: Nuti, Marianna
name: Rughetti, Aurelia
Publisher: name: figshare
Year: 2024
Subjects: subject: Biochemistry
subject: Medicine
subject: Cell Biology
subject: Genetics
subject: FOS: Biological sciences
subject: Molecular Biology
subject: Pharmacology
subject: Chemical Sciences not elsewhere classified
subject: Biological Sciences not elsewhere classified
subject: Cancer

description: Additional file 1: Table S1. FGFR expression in cancer cell lines. FGFR expression was evaluated by Western blot and q-RT‒PCR experiments in H1581, KATO III, RT-112, RCC4plusVHL and DU-145 cancer cell lines employed as cell models of lung, gastric, bladder, renal and prostate cancer, respectively. Figure S1. Serial dilution of Pem and proliferation analysis. A For selection of the optimal drug concentration, H1581, KATO III and RT-112 cells were exposed to serial dilutions of Pem (3–1000 nM) for 24 h and 48 h, and MTT proliferation assay was then performed. Data are reported as the mean of three independent experiments ± SEM as variability B H1581, KATO III and RT-112 cells were exposed to serial dilutions of Pem (25–400 nM) for 24 h and 48 h, and intracellular Ki67 levels were then evaluated by flow cytometry. The histograms represent the geo-mean of Ki67-BV421 MFI found in untreated (NT) and Pem-treated H1581, KATO III and RT-112 cells and are represented as mean of three independent experiments ± SEM. *p &lt; 0.05; **p &lt; 0.01; ***p ≤ 0.001 Student’s t test. Figure S2. Effect of Pem on the S and G2 phases of cell cycle. The histograms represent the count of PI-positive cells found in S- and G2-phase in untreated (NT, white) and Pem-treated (100 nM, grey) H1581, KATO III and RT-112 cells and are represented as mean of three independent experiments ± SEM. *p &lt; 0.05; **, p &lt; 0.01; ***p ≤ 0.001. Student’s t test. Figure S3. Effect of pemigatinib on ATP and HMGB1 release. A Extracellular HMGB1 release upon Pem treatment. The extracellular HMGB1 release was measured by means of an enzyme-linked immunosorbent assay (ELISA) kit (TECAN, Zürich, Switzerland), according to the manufacturer’s protocol. Histograms represent the mean values of three independent experiments ± SEM of HMGB1 released by untreated cells (NT, white) and Pem-treated cells (grey) at 24 and 48 h. ns, not significative. B Extracellular ATP release upon Pem treatment. The ATP release was measured by ENLITEN-Promega KIT as luminescence signals. The histograms represent the mean value of ATP moles of three independent experiments ± SEM by untreated cells (NT, white) and Pem-treated cells (Pem, grey) at 24 and 48 h. ns, not significative. Figure S4. Effect of pemigatinib on miRNA target transcripts. mRNA expression of miRNAs targets in untreated (NT) and Pem-treated (100 nM) cancer cell lines. Histograms represent the mean values of three independent experiments ± SEM.