pretreatmentwith cmet kinase inhibitors su11274  (MedChemExpress)

Journal: Cancers

Article Title: Dual Targeting of CDK4/6 and cMET in Metastatic Uveal Melanoma

doi: 10.3390/cancers13051104

Figure Lengend Snippet: Merestinib overcomes HGF-mediated resistance to abemaciclib in metastatic uveal melanoma cells. ( A ) UM002B and UM004 cells were treated with merestinib for 4 h, followed by 10 ng/mL of HGF stimulation for 15 min. Phosphorylation of cMET was evaluated by Western immunoblotting of cell lysates with phospho-cMET antibody. Actin was used as the loading control. ( B ) UM002B and UM004 cells were treated with DMSO or 1 μM abemaciclib, in combination with 10 ng/mL of HGF and/or 0.2 μM merestinib for 4 days. Cell viability was determined by the MTS assay. Data are presented as mean ± SD. ** p < 0.01, based on the two sample t -test with unequal variance. ( C ) UM002B and UM004 cells were treated with DMSO or 1 μM abemaciclib, in combination with 10 ng/mL of HGF and/or 0.2 μM of merestinib for seven days. Cells were stained and representative images are shown. ( D ) UM002B and UM004 cells were pretreated with DMSO, 1 μM of abemaciclib, or 0.2 μM of merestinib for 48 h. Cells were then stimulated with 10 ng/mL HGF for 1 h, as indicated. Activation of cMET, RB, AKT, ERK1/2, and p70S6K was analyzed by Western immunoblotting.

Article Snippet: A CDK4/6 inhibitor, Abemaciclib (LY2835219), and a cMET inhibitor, merestinib (LY2801653), were provided by Eli Lilly and Company, Indianapolis, IN, USA.

Techniques: Phospho-proteomics, Western Blot, Control, MTS Assay, Staining, Activation Assay

Journal: Cancers

Article Title: Dual Targeting of CDK4/6 and cMET in Metastatic Uveal Melanoma

doi: 10.3390/cancers13051104

Figure Lengend Snippet: HGF prevents abemaciclib induced-cell senescence and merestinib reverses HGF effects. ( A ) UM002B and UM004 cells were treated with DMSO or 1 μM of abemaciclib, in combination with 10 ng/mL of HGF and/or 0.2 μM of merestinib for six days. Cells were fixed and stained overnight for senescence-associated β-galactosidase activity. Positive cells were shown with arrows. Data are presented as mean ± SD. *, p < 0.05; **, p < 0.01, based on Tukey analysis. ( B ) UM002B and UM004 cells were treated with 1 μM of abemaciclib, in combination with 10 ng/mL of HGF and/or 0.2 μM of merestinib for 72 h. Activation of cMET, RB, FOXM1, AKT, ERK1/2, and p70S6K was analyzed by Western immunoblotting. ( C ) UM004 cells were transfected with control or FOXM1 targeting siRNA (#1 and #2) and cultured with 1 μM of abemaciclib in the presence or absence of 10 ng/mL of HGF for six days. Following six-day treatment, cells were fixed and stained overnight for senescence-associated b-galactosidase activity. Positive cells were shown with arrows. Knockdown of FOXM1 reversed the evasion of cellular senescence mediated by HGF. ( D ) UM004 cells were transfected with control or FOXM1 targeting siRNA (#1 and #2) in the presence of 1 μM of abemaciclib and 10 ng/mL of HGF for 72 h. FOXM1 and β-actin were analyzed by Western immunoblotting.

Article Snippet: A CDK4/6 inhibitor, Abemaciclib (LY2835219), and a cMET inhibitor, merestinib (LY2801653), were provided by Eli Lilly and Company, Indianapolis, IN, USA.

Techniques: Staining, Activity Assay, Activation Assay, Western Blot, Transfection, Control, Cell Culture, Knockdown

Journal: Cancers

Article Title: Dual Targeting of CDK4/6 and cMET in Metastatic Uveal Melanoma

doi: 10.3390/cancers13051104

Figure Lengend Snippet: Effects of the abemaciclib and merestinib combination on a metastatic uveal melanoma xenograft tumor model in human HGF knockin mice. ( A ) Mice bearing subcutaneous UM004 tumors were dosed with vehicle ( n = 5), 50 mg/kg abemaciclib ( n = 5), 12 mg/kg merestinib ( n = 5), or abemaciclib plus merestinib combination ( n = 5) once daily for 28 days. Tumor size was measured by caliper twice weekly. ( B ) Tumor weight. Data represent mean volume ± SD. * p < 0.05; ** p < 0.01, based on Dunnett’s test. ( C ) Average percentage change in body weight after treatments. ( D ) UM004 tumors were excised from mice following seven-day treatment. Tumor lysates were prepared and analyzed by Western immunoblotting using phospho-cMET, cMET, phospho-RB, total RB, FOXM1, and β-actin. ( E ) Representative images of immunohistochemical staining for hematoxylin and eosin (H & E), IgG isotype control, phospho-cMET, phospho-RB, FOXM1, and Ki67 in UM004 tumor xenografts. Original magnification, ×400. Scale bar, 50 μm.

Article Snippet: A CDK4/6 inhibitor, Abemaciclib (LY2835219), and a cMET inhibitor, merestinib (LY2801653), were provided by Eli Lilly and Company, Indianapolis, IN, USA.

Techniques: Knock-In, Western Blot, Immunohistochemical staining, Staining, Control