Journal: Acta Pharmacologica Sinica

Article Title: CUDC-907 displays potent antitumor activity against human pancreatic adenocarcinoma in vitro and in vivo through inhibition of HDAC6 to downregulate c-Myc expression

doi: 10.1038/s41401-018-0108-5

Figure Lengend Snippet: CUDC-907 displays potent antiproliferative activity against pancreatic cancer cell lines. a The growth inhibition ratio in Aspc-1 cells after 6 days of treatment with four serial concentrations of various inhibitors as indicated. Red to blue, cell growth inhibition ratio from 100% to 0%. b A panel of human pancreatic adenocarcinoma cells were treated with CUDC-907 at gradient concentrations for 3 days, and the half-maximal inhibitory concentration (IC50) was measured using the SRB assay. Error bars represent the mean ± SD from three independent experiments. c Dose-dependent proliferation curves of Aspc-1, PANC-1, and Capan-1 cells over time. Cells were treated with CUDC-907 at gradient concentrations for 0, 1, 2, 3, or 4 days, and the relative cell growth ratio (%) was calculated using SoftMax Pro. Error bars represent the mean ± SD from three independent experiments. d Colony-formation assays. Aspc-1, PANC-1, and Capan-1 cells were measured with crystal violet staining after treatment with CUDC-907 at 5 to 20 nM for 14 days. e–g CUDC-907 arrests pancreatic cancer cells in the G2/M phase and induces apoptosis. Aspc-1 and Capan-1 cells were incubated with various concentrations (as indicated) of CUDC-907 for 24 or 48 h, and the percentage of each cell cycle phase (e) and apoptosis (f) was assessed by flow cytometry; (g) Aspc-1 and Capan-1 cells were collected for Western blotting with the indicated antibodies after treatment with CUDC-907 for 48 h

Article Snippet: Compounds CUDC-907 was purchased from Med Chem Express (MCE) (NJ, USA), dissolved in dimethylsulfoxide at 10 mg/mL to form a stock solution and stored at −80 °C for in vitro studies.

Techniques: Activity Assay, Inhibition, Concentration Assay, Sulforhodamine B Assay, Staining, Incubation, Flow Cytometry, Western Blot

Journal: Acta Pharmacologica Sinica

Article Title: CUDC-907 displays potent antitumor activity against human pancreatic adenocarcinoma in vitro and in vivo through inhibition of HDAC6 to downregulate c-Myc expression

doi: 10.1038/s41401-018-0108-5

Figure Lengend Snippet: CUDC-907 downregulates c-Myc protein to inhibit cell proliferation. a c-Myc expression in pancreatic cancer cell lines compared with the normal cell line H6C7 evaluated by Western blotting. b c-Myc protein level alteration. Protein levels in Aspc-1, PANC-1, and Capan-1 cells treated with CUDC-907 (0, 25, 50, 100, and 500 nM) for 24 h were measured. The cell lysates were subjected to Western blotting analysis using the indicated antibodies. c c-Myc mRNA level alteration. Cells pretreated as described in b followed by qRT-PCR analysis of c-Myc level. Error bars represent the mean ± SD from three independent experiments. d Cell growth assay. Pancreatic cancer cells were transfected with c-Myc-specific siRNAs for 24 h and then transferred to a 96-well plate to be cultured for 3 or 6 days or to a 6-well plate for 14 days to form colonies. Relative cell proliferation rates were calculated by OD values, and colonies were scanned for crystal violet staining. Interference efficiency was verified by Western blotting. e Aspc-1-MYC and PANC-1-MYC cells, which were stably transfected with a mock or MYC-overexpression plasmid, were transfected with siRNAs against c-Myc or scramble for 24 h and then were transferred to 96-well plates to be cultured for 6 days, and the relative cell proliferation rate was calculated. Overexpression efficiency was verified by Western blotting. Error bars represent the mean ± SD from triplicates or three independent experiments. ***P < 0.001

Article Snippet: Compounds CUDC-907 was purchased from Med Chem Express (MCE) (NJ, USA), dissolved in dimethylsulfoxide at 10 mg/mL to form a stock solution and stored at −80 °C for in vitro studies.

Techniques: Expressing, Western Blot, Quantitative RT-PCR, Growth Assay, Transfection, Cell Culture, Staining, Stable Transfection, Over Expression, Plasmid Preparation

Journal: Acta Pharmacologica Sinica

Article Title: CUDC-907 displays potent antitumor activity against human pancreatic adenocarcinoma in vitro and in vivo through inhibition of HDAC6 to downregulate c-Myc expression

doi: 10.1038/s41401-018-0108-5

Figure Lengend Snippet: CUDC-907 inhibits both the PI3K pathway and HDACs to suppress c-Myc function. a Individual PI3K or HDACs inhibitors suppress c-Myc. Aspc-1, PANC-1, and Capan-1 cells were treated with the indicated concentrations of the PI3K inhibitor GDC-0941 or the HDAC inhibitor vorinostat for 24 h, and the expression of c-Myc, phosphorylation of AKT and the acetylation of histone proteins were detected by Western blotting. b Synergistic effect of PI3K and HDAC inhibitors. IC50 values of vorinostat and GDC-0941 alone or in combination against Aspc-1, PANC-1, and Capan-1 cells were tested, and the combination index (CI) was calculated using the CompuSyn software. CI < 1, synergic effect; CI = 1, additive effect; CI > 1, antagonistic effect. Three independent experiments were exhibited. Error bars represent the mean ± SD from three independent experiments

Article Snippet: Compounds CUDC-907 was purchased from Med Chem Express (MCE) (NJ, USA), dissolved in dimethylsulfoxide at 10 mg/mL to form a stock solution and stored at −80 °C for in vitro studies.

Techniques: Expressing, Phospho-proteomics, Western Blot, Software

Journal: Acta Pharmacologica Sinica

Article Title: CUDC-907 displays potent antitumor activity against human pancreatic adenocarcinoma in vitro and in vivo through inhibition of HDAC6 to downregulate c-Myc expression

doi: 10.1038/s41401-018-0108-5

Figure Lengend Snippet: CUDC-907 inhibits the PI3K pathway and leads to degradation of c-Myc. a PI3K pathway inhibition upregulates phosphor-c-Myc (Thr58) protein level. Aspc-1 and PANC-1 cells were treated with CUDC-907 or the PI3K inhibitor GDC-0941 for 2 h, and phosphor-c-Myc (Thr58) and total c-Myc protein levels were detected by Western blotting analysis using the indicated antibodies. Relative accumulation of phosphor-c-Myc (Thr58) to total c-Myc (normalized by the same Actin using the Image J software) was shown. b Aspc-1 and PANC-1 cells were treated with CUDC-907 (0, 25, 50, 100, and 500 nM) with (right) or without (left) proteasome inhibitor MG-132 for 2 h and then the cell lysates were subjected to Western blotting analysis to detect the c-Myc protein level

Article Snippet: Compounds CUDC-907 was purchased from Med Chem Express (MCE) (NJ, USA), dissolved in dimethylsulfoxide at 10 mg/mL to form a stock solution and stored at −80 °C for in vitro studies.

Techniques: Inhibition, Western Blot, Software

Journal: Acta Pharmacologica Sinica

Article Title: CUDC-907 displays potent antitumor activity against human pancreatic adenocarcinoma in vitro and in vivo through inhibition of HDAC6 to downregulate c-Myc expression

doi: 10.1038/s41401-018-0108-5

Figure Lengend Snippet: CUDC-907 regulates c-Myc expression and cell proliferation through HDAC6 inhibition. a HDAC6 interference inhibits c-Myc protein levels. Aspc-1 and Capan-1 cells were transfected with individual siRNA against HDAC1, 2, 3, 4, and 6. HDAC6 interference inhibits c-Myc protein levels, as detected by Western blotting analysis using the indicated antibodies. b HDAC6 interference suppresses cell proliferation. Interference was performed as shown in Fig. 4a, and cells were transferred to a 96-well plate to be cultured for 3 days. The relative cell proliferation rate was calculated by OD values. Error bars represent the mean ± SD from three independent experiments. c HDAC6 reverses antiproliferation effect induced by CUDC-907 partially. Aspc-1-MOCK and Aspc-1-HDAC6 cells, which were stably transfected with a MOCK or HDAC6-overexpressing plasmid, were treated with CUDC-907 (0, 2, 5 nM) for 3 days, and the relative cell proliferation rate was calculated using SoftMax Pro. Error bars represent the mean ± SD from three independent experiments. d, e HDAC6 regulates c-Myc through the HDAC6–FOXO1–c-Myc axis. Aspc-1 cells were transfected with individual siRNA against HDAC6, FOXO1, or HDAC6-overexpressing plasmid, and the protein levels were detected by Western blotting analysis using the indicated antibodies (d). Also, a series of concentration of CUDC-907 were treated in Aspc-1, PANC-1, and Capan-1 cells for 24 h, and the FOXO1 and c-Myc protein levels were evaluated by Western blotting (e)

Article Snippet: Compounds CUDC-907 was purchased from Med Chem Express (MCE) (NJ, USA), dissolved in dimethylsulfoxide at 10 mg/mL to form a stock solution and stored at −80 °C for in vitro studies.

Techniques: Expressing, Inhibition, Transfection, Western Blot, Cell Culture, Stable Transfection, Plasmid Preparation, Concentration Assay

Journal: Acta Pharmacologica Sinica

Article Title: CUDC-907 displays potent antitumor activity against human pancreatic adenocarcinoma in vitro and in vivo through inhibition of HDAC6 to downregulate c-Myc expression

doi: 10.1038/s41401-018-0108-5

Figure Lengend Snippet: CUDC-907 inhibits pancreatic cancer growth in vivo. a Tumor growth inhibition upon CUDC-907 treatment in Aspc-1 pancreatic xenografts. The curve of relative tumor volume after 20 days of treatment with CUDC-907 or vehicle and the corresponding curve of body weight are shown. Error bars represent the mean ± SD from seven individuals in each group. **P < 0.01. b Tumor morphology and weight. After 20 days, all nude mice were sacrificed, and the tumors were resected to be imaged and weighed. c Protein levels' alteration. Equal amounts of proteins from tumor tissues were evaluated for the phosphorylation of AKT and Erk and the acetylation of histone. d Immunohistochemistry staining. (i) Representative images of Ki67 and c-Myc staining of untreated xenografts and xenografts treated with CUDC-907. Scale bar, 100 μm. (ii) Quantification of immunohistochemistry staining was calculated by quick score (QS) method. Ten random areas from three individually stained tumors were captured. Error bars represent the mean ± SD from three independent experiments. **P < 0.01

Article Snippet: Compounds CUDC-907 was purchased from Med Chem Express (MCE) (NJ, USA), dissolved in dimethylsulfoxide at 10 mg/mL to form a stock solution and stored at −80 °C for in vitro studies.

Techniques: In Vivo, Inhibition, Phospho-proteomics, Immunohistochemistry, Staining