Journal: FASEB BioAdvances

Article Title: Activation of AMPK sensitizes medulloblastoma to Vismodegib and overcomes Vismodegib‐resistance

doi: 10.1096/fba.2020-00032

Figure Lengend Snippet: HH/GLI1 activation and suppression of AMPK in Vismodegib‐resistant medulloblastoma cell lines. (A) To generate Vismodegib‐resistant MB cells, MB11 and DAOY cells were treated with Vismodegib daily with a gradual increase in concentration from 10 to 300 μM in 16 weeks. (B) From two pairs of MB parental and Vismodegib‐resistant cell lines (VisR), MB11 and DAOY, mRNA was collected, and the amount of HH/GLI1 downstream targets, GLI1 , PTCH1 , Cyclin D1 , C ‐ MYC , and BCL ‐ 2 mRNA was analyzed using qRT‐PCR with GAPDH mRNA as the internal control for normalization. The bars indicate mRNA level relative to that of parental cells. The experimental points were in triplicate and independently repeated three times (* p < 0.05, ** p < 0.01, *** p < 0.001). (C) Lysates from MB11 and DAOY (P: parental; VisR: Vismodegib‐resistant) cells were analyzed by immunoblot with the indicated antibodies

Article Snippet: AMPK activator Metformin was purchased from Sigma (St. Louis, MO, USA), A‐769662 and Vismodegib were purchased from LC Laboratories (Woburn, MA, USA).

Techniques: Activation Assay, Concentration Assay, Quantitative RT-PCR, Control, Western Blot

Journal: FASEB BioAdvances

Article Title: Activation of AMPK sensitizes medulloblastoma to Vismodegib and overcomes Vismodegib‐resistance

doi: 10.1096/fba.2020-00032

Figure Lengend Snippet: AMPK agonist synergizes with Vismodegib and suppresses medulloblastoma cell growth. Human MB cell lines, (A) MB11, (B) DAOY, (C) ONS‐76, and (D) MED8A were treated with DMSO, Vis (Vismodegib, 10 μM), Met (Metformin, 5 mM), Vis and Met (Vis+Met, 10 μM+5 mM) and subjected to cell growth assay for three consecutive days. (E) MB11 and (F) DAOY cells were treated with Vismodegib (from 20 to 140 μM), A769662 (from 20 to 140 μM), Vismodegib and A769662 for 48 hours and subjected for cell growth assay. (G) MB11 and (H) DAOY cells were treated with Vismodegib (from 40 to 100 μM), Metformin (10 mM), Vismodegib and Metformin for 48 hours and subjected to cell growth assay. All the experimental points were in triplicate and repeated three times independently (* p < 0.05, ** p < 0.01, *** p < 0.001)

Article Snippet: AMPK activator Metformin was purchased from Sigma (St. Louis, MO, USA), A‐769662 and Vismodegib were purchased from LC Laboratories (Woburn, MA, USA).

Techniques: Growth Assay

Journal: FASEB BioAdvances

Article Title: Activation of AMPK sensitizes medulloblastoma to Vismodegib and overcomes Vismodegib‐resistance

doi: 10.1096/fba.2020-00032

Figure Lengend Snippet: Combination of AMPK agonist and Vismodegib achieves better inhibitory effect on MB colony formation. (A) MB11 and (B) DAOY cells were seeded into six‐well plates and treated with DMSO, Vis, (Vismodegib, 10 μM), Met (Metformin, 5 mM), Vis and Met (Vis+Met, 10 μM+5 mM) for soft agar colony formation assay for 2 weeks. Colonies larger than 1.0 mm were counted (as indicated by arrows). The experiment was repeated three times (** p < 0.01,) Scale bar: 10 mm (C) MB11 and DAOY cells were seeded in 12‐wells plate and treated with DMSO, Vis, (Vismodegib, 10 μM), Met (Metformin, 5 mM), Vis and Met (10 μM+5 mM) for colony formation assay for 12 days. (D) MB11 and DAOY cells were treated with DMSO, Vis, (Vismodegib, 10 μM), Met (Metformin, 5 mM), Vis and Met (10 μM+5 mM) for 24 hours, and the cell lysates were analyzed by western blotting with the indicated antibodies

Article Snippet: AMPK activator Metformin was purchased from Sigma (St. Louis, MO, USA), A‐769662 and Vismodegib were purchased from LC Laboratories (Woburn, MA, USA).

Techniques: Soft Agar Assay, Colony Assay, Western Blot

Journal: FASEB BioAdvances

Article Title: Activation of AMPK sensitizes medulloblastoma to Vismodegib and overcomes Vismodegib‐resistance

doi: 10.1096/fba.2020-00032

Figure Lengend Snippet: Combining AMPK activator and SMO inhibitor sensitizes Vismodegib‐resistant MB. (A) MB11‐VisR and (C) DAOY‐VisR cells were treated with DMSO, Vismodegib (From 20 to 140 μM), A769662 (From 20 to 140 μM), Vismodegib (From 20 to 140 μM), and A769662 (From 20 to 140 μM) for 48 hours and subjected to cell growth assay. (B) MB11‐VisR and (D) DAOY‐VisR cells were treated with DMSO, Vismodegib (From 40 to 240 μM), Metformin (10 mM), Vismodegib (From 40 to 240 μM), and Metformin (10 mM) for 48 hours and subjected to cell growth assay. (E) MB11‐VisR and (F) DAOY‐VisR cells were treated with DMSO, Vismodegib (20 μM), Metformin (10 mM), Vismodegib (20 μM), and Metformin (10 mM) for 2 weeks and subjected to soft agar colony formation assay. (G) DAOY‐VisR and (H) MB11‐VisR cells were treated with DMSO, Vismodegib (10 μM), Metformin (5 mM), Vismodegib (10 μM), and Metformin (5 mM) for 24 hours, and lysates were analyzed by immunoblot with the indicated antibodies. (I) MB11‐ and DAOY‐SMO D473G (Smoothened mutation D473G) stable cell lines were treated with DMSO, Vismodegib (40 μM), Metformin (5 mM), Vismodegib (40 μM) and Metformin (5 mM) for 48 hours and subjected to cell growth assay. Similarly, (J) DAOY‐SMO W535L and (K) MB11‐SMO W535L stable cell lines were treated with DMSO, Vismodegib (40 μM), Metformin (5 mM), Vismodegib (40 μM), and Metformin (5 mM) for 48 hours and subjected to cell growth assay. (L) MB11‐ and DAOY‐SMO D473G (Smoothened mutation D473G) stable cell lines were treated with DMSO, Vismodegib (40 μM), Metformin (5 μM), Vismodegib (40 μM) and Metformin (5 μM) for 48 hours and subjected to cell growth assay. (M) MB11‐VisR cells were treated with DMSO, Vismodegib (10 μM), Metformin (5 μM), Vismodegib (10 μM) and Metformin (5 μM) for 24 hours and lysates were analyzed by immunoblot with the indicated antibodies. All the cell growth assay points were in triplicate and each experiment repeated three times (* p < 0.05, ** p < 0.01)

Article Snippet: AMPK activator Metformin was purchased from Sigma (St. Louis, MO, USA), A‐769662 and Vismodegib were purchased from LC Laboratories (Woburn, MA, USA).

Techniques: Growth Assay, Soft Agar Assay, Western Blot, Mutagenesis, Stable Transfection

Journal: FASEB BioAdvances

Article Title: Activation of AMPK sensitizes medulloblastoma to Vismodegib and overcomes Vismodegib‐resistance

doi: 10.1096/fba.2020-00032

Figure Lengend Snippet: Combination of AMPK activator and SMO inhibitor effectively suppresses MB growth in vivo. (A) Tumor volume of the MB11 xenografts treated with DMSO, vismodegib, Metformin, and vismodegib plus Metformin was measured for 28 days. (B) The tumor sections of two individual samples from four different treatment groups were subjected to immunohistochemistry with a Ki67 and cleaved caspase 3 antibody. Scale bar: 100 µm. Relative percentages of Ki67 (C) and cleaved caspase 3 (D) expression by individual xenograft tumors from B were analyzed and the mean values of Ki67 and cleaved caspase 3 expression in DMSO, Vismodegib, Metformin and Vismodegib plus Metformin treated group were indicated as bars (** p < 0.01, *** p < 0.001). (E) Intracranial MB11 xenografts treated with DMSO, Vismodegib, Metformin and Vismodegib plus Metformin were measured by IVIS image system for 5 weeks. Color indicates the quantification of tumor growth from small (blue) to big size (red)

Article Snippet: AMPK activator Metformin was purchased from Sigma (St. Louis, MO, USA), A‐769662 and Vismodegib were purchased from LC Laboratories (Woburn, MA, USA).

Techniques: In Vivo, Immunohistochemistry, Expressing

Journal: Clinical cancer research : an official journal of the American Association for Cancer Research

Article Title: CD44 expression denotes a subpopulation of gastric cancer cells in which Hedgehog signaling promotes chemotherapy resistance

doi: 10.1158/1078-0432.CCR-14-0011

Figure Lengend Snippet: Hedgehog pathway inhibition with Smo shRNA or vismodegib blocks spheroid formation. A. Western blot demonstrating knockdown of Smo in gastric cancer cell lines AGS, MKN-45, and N87 following transduction with Smo shRNA (Smo.shRNA) lentivirus compared to scrambled shRNA control (Scr.shRNA) lentivirus. B. Immunofluorescence photos for CD44 (green) and nuclei (blue) of gastric cancer cell lines treated with Smo.shRNA or Scr.shRNA and grown in spheroid formation conditions or treated with vismodegib (10 μM) or carrier (DMSO) and grown in spheroid formation conditions. C. Single cell assay of spheroid cells showing diameter of spheroids at selected time points following treatment with Smo.shRNA vs. Scr.shRNA or vismodegib (Vis, 10 μM) vs. carrier (DMSO). Bars represent standard deviation.

Article Snippet: Vismodegib was purchased from LC Laboratories (Woburn, MA).

Techniques: Inhibition, shRNA, Western Blot, Transduction, Immunofluorescence, Standard Deviation

Journal: Clinical cancer research : an official journal of the American Association for Cancer Research

Article Title: CD44 expression denotes a subpopulation of gastric cancer cells in which Hedgehog signaling promotes chemotherapy resistance

doi: 10.1158/1078-0432.CCR-14-0011

Figure Lengend Snippet: CD44(+) gastric cancer cells demonstrate chemotherapy resistance, which can be reversed with Hedgehog pathway inhibition. A. Immunofluorescence images of CD44 (green) and nuclei (blue) for CD44(+) and CD44(−) cells following magnetic bead sorting of AGS, MKN-45, and N87 gastric cancer cell lines. B. Western blot showing expression of Hedgehog pathway proteins Sonic hedgehog (Shh), Patch 1 (Ptch1), Smoothened (Smo), and Gli1 in CD44(+) and CD44(−) cells. Proliferation assay for CD44(+) and CD44(−) cells (C) and spheroid cells and monolayer cells (D) following treatment with 5-fluorouracil (5-FU) or cisplatin chemotherapy and vismodegib (Vis, 10 μM) compared to DMSO. Bars represent standard deviation.

Article Snippet: Vismodegib was purchased from LC Laboratories (Woburn, MA).

Techniques: Inhibition, Immunofluorescence, Western Blot, Expressing, Proliferation Assay, Standard Deviation

Journal: Clinical cancer research : an official journal of the American Association for Cancer Research

Article Title: CD44 expression denotes a subpopulation of gastric cancer cells in which Hedgehog signaling promotes chemotherapy resistance

doi: 10.1158/1078-0432.CCR-14-0011

Figure Lengend Snippet: Gastric cancer spheroid cells demonstrate increased migration, colony formation, and anchorage independent growth, which can be attenuated by Hedgehog pathway inhibition. A. Photos and graphs of migration assay of gastric cancer monolayer cells and spheroid cells treated with Smo shRNA (Smo.shRNA) compared to control (Scr.shRNA) or vismodegib (10 μM) compared to DMSO. B. Colony formation assay of gastric cancer monolayer cells and spheroid cells treated with Smo shRNA (Smo.shRNA) compared to control (Scr.shRNA) or vismodegib (10 μM) compared to DMSO. C. Photos and graph of soft agar assay of gastric cancer monolayer cells and spheroid cells treated with Smo shRNA (Smo.shRNA) compared to control (Scr.shRNA). Bars represent standard deviation.

Article Snippet: Vismodegib was purchased from LC Laboratories (Woburn, MA).

Techniques: Migration, Inhibition, shRNA, Colony Assay, Soft Agar Assay, Standard Deviation

Journal: Clinical cancer research : an official journal of the American Association for Cancer Research

Article Title: CD44 expression denotes a subpopulation of gastric cancer cells in which Hedgehog signaling promotes chemotherapy resistance

doi: 10.1158/1078-0432.CCR-14-0011

Figure Lengend Snippet: CD44 may be a response biomarker in advanced gastric and gastroesophageal cancer patients treated with chemotherapy with or without vismodegib. A. Schema of the clinical trial. B. CD44 immunohistochemistry of patient tumor samples showing low, intermediate, and high CD44 expression. C. Kaplan-Meier overall survival curves for patients receiving chemotherapy alone and patients receiving chemotherapy plus vismodegib stratified by low and high CD44 score.

Article Snippet: Vismodegib was purchased from LC Laboratories (Woburn, MA).

Techniques: Biomarker Assay, Immunohistochemistry, Expressing

Journal: Clinical cancer research : an official journal of the American Association for Cancer Research

Article Title: CD44 expression denotes a subpopulation of gastric cancer cells in which Hedgehog signaling promotes chemotherapy resistance

doi: 10.1158/1078-0432.CCR-14-0011

Figure Lengend Snippet: CD44 score, progression, and survival

Article Snippet: Vismodegib was purchased from LC Laboratories (Woburn, MA).

Techniques:

Journal: Cell Reports Methods

Article Title: RECOVER identifies synergistic drug combinations in vitro through sequential model optimization

doi: 10.1016/j.crmeth.2023.100599

Figure Lengend Snippet:

Article Snippet: Vismodegib (GDC-0449) , Selleck , S1082.

Techniques: Software, Recombinant

Journal: PLOS ONE

Article Title: Analysis of residual disease in periocular basal cell carcinoma following hedgehog pathway inhibition: Follow up to the VISORB trial

doi: 10.1371/journal.pone.0265212

Figure Lengend Snippet: A) Pathology results for excision samples after vismodegib treatment (error bars indicate 95% CI, n = 9 (left column), n = 18 (right column)), B) Pre- and post-vismodegib histology, C) Average total area and D) Average lesion size (pixels) of keratin 5 positive staining (normalized to section length, error bars indicate SEM, unpaired t-test *p<0.05, n = 7 (left column), n = 11 (right column)), E) Clinical photo, histology, and keratin staining pre and post-vismodegib treatment of patients scored as “no sign of disease” (black arrow–tumor location, yellow box—area of peripheral palisading).

Article Snippet: Vismodegib (Genentech Inc.) is a small-molecule SMO inhibitor approved by the United States Food and Drug Administration (FDA) to treat advanced and metastatic BCC [ ].

Techniques: Staining

Journal: PLOS ONE

Article Title: Analysis of residual disease in periocular basal cell carcinoma following hedgehog pathway inhibition: Follow up to the VISORB trial

doi: 10.1371/journal.pone.0265212

Figure Lengend Snippet: Keratin 5 expression in post-vismodegib excision samples.

Article Snippet: Vismodegib (Genentech Inc.) is a small-molecule SMO inhibitor approved by the United States Food and Drug Administration (FDA) to treat advanced and metastatic BCC [ ].

Techniques: Expressing

Journal: PLOS ONE

Article Title: Analysis of residual disease in periocular basal cell carcinoma following hedgehog pathway inhibition: Follow up to the VISORB trial

doi: 10.1371/journal.pone.0265212

Figure Lengend Snippet: A) Gli1 expression (dots/nuclei) pre- and post-vismodegib treatment in patient samples scored as disease present (red) or no sign of disease (green) (error bars indicate SEM, unpaired t-test *p<0.05, n = 7,6,9,11 columns left to right), B) Gli1 expression (brown dots) in excision samples from “disease present” (left panels) and “no sign of disease” (right panels) samples. C) Gli1 expression (left panels, brown) and proliferation (Ki67, green) in keratin 5 positive (red) persistent lesions from samples scored as disease present (top panels) and no sign of disease (bottom panels).

Article Snippet: Vismodegib (Genentech Inc.) is a small-molecule SMO inhibitor approved by the United States Food and Drug Administration (FDA) to treat advanced and metastatic BCC [ ].

Techniques: Expressing

Journal: PLOS ONE

Article Title: Analysis of residual disease in periocular basal cell carcinoma following hedgehog pathway inhibition: Follow up to the VISORB trial

doi: 10.1371/journal.pone.0265212

Figure Lengend Snippet: Gli1 expression in post-vismodegib excision samples.

Article Snippet: Vismodegib (Genentech Inc.) is a small-molecule SMO inhibitor approved by the United States Food and Drug Administration (FDA) to treat advanced and metastatic BCC [ ].

Techniques: Expressing

Journal: PLOS ONE

Article Title: Analysis of residual disease in periocular basal cell carcinoma following hedgehog pathway inhibition: Follow up to the VISORB trial

doi: 10.1371/journal.pone.0265212

Figure Lengend Snippet: A) Clinical photos and Gli1 expression (brown dots) in a patient pre- (left panels), post-vismodegib (center panels), and after recurrence (right panels), B) Allele frequency of PTCH1 and SMO variants pre-, post-vismodegib, and after recurrence (*variant not detected)(yellow star–predicted pathogenic variant, red star–known pathogenic variant).

Article Snippet: Vismodegib (Genentech Inc.) is a small-molecule SMO inhibitor approved by the United States Food and Drug Administration (FDA) to treat advanced and metastatic BCC [ ].

Techniques: Expressing, Variant Assay