pd173952  (MedChemExpress)

Journal: iScience

Article Title: Decoding sunitinib resistance in ccRCC: Metabolic-reprogramming-induced ABAT and GABAergic system shifts

doi: 10.1016/j.isci.2024.110415

Figure Lengend Snippet: Inhibiting LYN or SYK enhances sunitinib efficacy against resistant ccRCC (A and B) Decreased SYK and LYN phosphorylation in ABAT-knockdown ccRCC cells (786-O and 769-P) (see Figures S7 A and S7B for statistical graph). (C) Dose-response curves and corresponding IC50 of sunitinib in ABATsh LYN Y397F and ABATsh SYK Y323/525F ccRCC (786-O and 768-P) cells and ABATsh SYK Y323/525F ccRCC cells treated with R406 and ABATsh LYN Y397F ccRCC cells treated with PD173952. x axis, log10 drug concentrations. y axis, Activity%, the percentage of cellular or biological activity in the drug-treated group relative to the control group. (D) Tube formation assays of HUVECs cultured in conditioned medium from ABATsh SYK Y323/525F , ABATsh SYK WT , ABATsh treated with R406, ABATsh LYN Y397F , ABATsh LYN WT , or ABATsh treated with PD173952 ccRCC cells (786-O and 769-P) treated with DMSO or sunitinib for 24 h (see Figures S7 C and S7D for statistical graph). (E) Chick embryo chorioallantoic membrane (CAM) assays with conditioned medium from ABATsh SYK Y323/525F , ABATsh SYK WT , ABATsh treated with R406, ABATsh LYN Y397F , ABATsh LYN WT , and ABATsh treated with PD173952 ccRCC cells (786-O and 769-P) treated with DMSO or sunitinib for 24 h (see Figures S7 E and S7F for statistical graph). (F) Compared to those in the monotherapy with sunitinib, there were fewer EdU-positive cells in the combination therapy of sunitinib with R406 or PD173952 in ABATsh SYK Y323/525F or ABATsh LYN Y397F ccRCC cells. (G and H) Mutual interaction between SYK phosphorylation and LYN phosphorylation. Representative blots ( n = 3 experiments) are shown (see Figures S7 G and S7H for statistical graph). (I) Treatments were combined with vehicle (vegetable oil), sunitinib (40 mg/kg in vegetable oil), R406 (5 mg/kg PD173952 in vegetable oil), a combination of sunitinib and R406 (40 mg/kg sunitinib and 5 mg/kg PD173952 in vegetable oil), a combination of sunitinib and PD173952 (40 mg/kg sunitinib and 20 mg/kg PD173952 in vegetable oil), or PD173952 (20 mg/kg PD173952 in vegetable oil) separately in heterograft tumors (769-P S, 769-P R cells), and tumor growth was recorded ( n = 4/group) (see Figures S7 I and S7J for statistical graph). The data are presented as the means ± SEMs. Groups were compared using one-way ANOVA with Tukey’s multiple comparisons test, two-way ANOVA with Tukey’s multiple comparisons test, or unpaired t test where appropriate. Each spot represents one subject. ns, not significant; ∗ p < 0.05, ∗∗ p < 0.01, ∗∗∗∗ p < 0.0001. See also Figure S7 .

Article Snippet: PD173952 , MedChemExpress , CAS: 305820-75-1.

Techniques: Phospho-proteomics, Knockdown, Activity Assay, Control, Cell Culture, Membrane

Journal: iScience

Article Title: Decoding sunitinib resistance in ccRCC: Metabolic-reprogramming-induced ABAT and GABAergic system shifts

doi: 10.1016/j.isci.2024.110415

Figure Lengend Snippet: Elevated phosphorylation levels of SYK and LYN activate the Akt and STAT pathways (A and B) Differential gene expression analysis of wild-type ccRCC cells and stable SYK- and LYN-phosphorylated ccRCC cell activation by sequencing. (C) KEGG enrichment analysis of the DEGs. (D) Western blot of key genes enriched in the pathway in LYN Y397E ccRCC cells (786-O and 769-P) treated with PD173952 (see Figure S8 A for statistical graph). Representative blots ( n = 3 experiments) are shown. (E) Western blot of key genes enriched in the pathway comprising LYN Y397E or LYN Y397F in ccRCC cells (786-O and 769-P) (see Figure S8 B for statistical graph). Representative blots ( n = 3 experiments) are shown. (F) Western blot of key genes enriched in the pathway in SYK Y323/525E or SYK Y323/525F ccRCC cells (786-O and 769-P) (see Figure S8 C for statistical graph). The data represent one of two independent experiments. (G) Western blot of key genes enriched in the pathway in SYK Y323/525E ccRCC cells (786-O and 769-P) treated with R406 (see Figure S8 D for statistical graph). Representative blots ( n = 3 experiments) are shown. The data are presented as the means ± SEMs. Groups were compared using two-way ANOVA with Tukey’s multiple comparisons test. Each spot represents one subject. POS, positive control spot; ns, not significant; ∗∗∗∗ p < 0.0001. See also Figure S8 .

Article Snippet: PD173952 , MedChemExpress , CAS: 305820-75-1.

Techniques: Phospho-proteomics, Gene Expression, Activation Assay, Sequencing, Western Blot, Positive Control

Journal: iScience

Article Title: Decoding sunitinib resistance in ccRCC: Metabolic-reprogramming-induced ABAT and GABAergic system shifts

doi: 10.1016/j.isci.2024.110415

Figure Lengend Snippet: Elevated phosphorylation levels of SYK and LYN upregulate PGF to promote angiogenesis (A–C) The intersection of angiogenesis genes and key genes enriched in the pathway (A). (B) A protein-protein interaction (PPI) network of the top 15 key genes from (A) was constructed. Partial vascular-related key genes (C). (D) Western blot of PDPK1, PIK3CA, SHH, EREG, and PGF in ccRCC cells (786-O and 769-P) treated with PD173952 (see Figure S9 A for statistical graph). The data represent one of two independent experiments. (E) Western blot of PDPK1, PIK3CA, SHH, EREG, and PGF in ccRCC cells (786-O and 769-P) treated with R406 (see Figure S9 B for statistical graph). The data represent one of two independent experiments. (F) Western blot of PDPK1, PIK3CA, SHH, EREG, and PGF in LYN Y397E or LYN Y397F ccRCC cells (786-O and 769-P) (see Figure S9 C for statistical graph). The data represent one of two independent experiments. (G) Western blot of PDPK1, PIK3CA, SHH, EREG, and PGF in SYK Y323/525E or SYK Y323/525F ccRCC cells (786-O and 769-P) (see Figure S9 D for statistical graph). The data represent one of two independent experiments. (H and I) Inhibited EREG or PGF in ABAT-knockdown ccRCC cells (786-O and 769-P) (see Figures S10 A and S10B for statistical graph). (J) Tube formation assays using HUVECs cultured in CM from ABATsh PGFsi, ABATsh EREGsi, and ABATsh NCsi ccRCC cells (786-O and 769-P) treated with DMSO or sunitinib for 24 h (see Figures S10 C and S10D for statistical graph). (K) Chick embryo chorioallantoic membrane (CAM) assays with conditioned medium from ABATsh PGFsi, ABATsh EREGsi, and ABATsh NCsi ccRCC cells (786-O and 769-P) treated with DMSO or sunitinib for 24 h (see Figures S10 E–S10G for statistical graph). The data are presented as the means ± SEMs. Groups were compared using one-way ANOVA with Tukey’s multiple comparisons test, two-way ANOVA with Tukey’s multiple comparisons test, or unpaired t test where appropriate. Each spot represents one subject. NC, negative control. ∗∗ p < 0.01, ∗∗∗ p < 0.0001, ∗∗∗∗ p < 0.0001. See also Figures S9 and .

Article Snippet: PD173952 , MedChemExpress , CAS: 305820-75-1.

Techniques: Phospho-proteomics, Construct, Western Blot, Knockdown, Cell Culture, Membrane, Negative Control

Journal: iScience

Article Title: Decoding sunitinib resistance in ccRCC: Metabolic-reprogramming-induced ABAT and GABAergic system shifts

doi: 10.1016/j.isci.2024.110415

Figure Lengend Snippet:

Article Snippet: PD173952 , MedChemExpress , CAS: 305820-75-1.

Techniques: Recombinant, Lysis, Electrophoresis, Cell Counting, Fluorescence, Staining, Phospho-proteomics, Ab Array, Plasmid Preparation, Software

Journal: Thrombosis Research

Article Title: Involvement of Src kinases and PLCγ2 in clot retraction

doi: 10.1016/j.thromres.2006.09.003

Figure Lengend Snippet: The Src kinase inhibitors and α IIb β 3 blockade by GRGDS peptide reduced time course of clot retraction and inhibited protein tyrosine phosphorylation during clot retraction. (A) Human washed platelets (5 × 10 8 /ml) were preincubated with DMSO, 50 μM PP2, 40 μM PD173952. Clot retraction assays were started by adding 250 μl of 2 U/ml thrombin to 250 μl of platelets in the presence of 2 mg/ml fibrinogen and 2 mM CaCl 2 (final concentrations: 2.5 × 10 8 /ml of platelets, 1 U/ml of thrombin, 1 mg/ml of fibrinogen, 1 mM CaCl 2 ). The volume of remaining fluid was measured to assay the degree of clot retraction. The volume was expressed as mean ± SE ( n = 5–18 from 2–5 experiments). Results were analyzed using unpaired Student's t -test. One asterisk denotes p < 0.05 and two denote p < 0.005 between control and PP2/PD173952. (B) Human washed platelets in the presence or absence of 1 mM GRGDS peptide were stimulated by 250 μl of 2 U/ml thrombin as described. Reactions were terminated by addition of 2× lysis buffer. Samples were sonicated 3 periods of 15 s and insoluble debris removed by centrifugation at 15,000 g for 10 min. The supernatant was solubilized by addition of 4× SDS sample buffer. The platelet proteins were separated by SDS-PAGE and blotted with anti-phosphotyrosine antibody (4G10). (C) Human washed platelets (5 × 10 8 /ml) were preincubated with 50 μM PP3 or PP2 before starting assays of clot retraction. Protein tyrosine phosphorylation was analyzed as described above.

Article Snippet: PD173952 was a gift from Pfizer (Ann Arbor, Michigan, USA) .

Techniques: Phospho-proteomics, Control, Lysis, Sonication, Centrifugation, SDS Page

Journal: Thrombosis Research

Article Title: Involvement of Src kinases and PLCγ2 in clot retraction

doi: 10.1016/j.thromres.2006.09.003

Figure Lengend Snippet: A role for PLCγ2 in clot retraction. (A) Human washed platelets (5 × 10 8 /ml) were preincubated with 50 μM PP3 or PP2 before starting assays of clot retraction. Clot retraction was stimulated as described in the legend of and reactions were terminated by addition of 2× lysis buffer. Samples were sonicated 3 periods of 15 s and insoluble debris removed by centrifugation at 15,000 g for 10 min. PLCγ2 was immunoprecipitated using the supernatant. The immunoprecipitates were separated by SDS-PAGE and blotted with anti-phosphotyrosine antibody (4G10) or anti-PLCγ2. (B, C) Murine diluted-PRP (400 μl, 3 × 10 8 /ml) from wild type mice or PLCγ2-deficient mice was added 10 U/ml of thrombin and 2 mM CaCl 2 . Where indicated, PLCγ2-deficient platelets were preincubated with 40 μM PD173952 before starting clot retraction assay. Clot retraction was recorded by digital camera (B) and the volume of remaining fluid was measured at 120 min to assay the degree of clot retraction (C). The results were shown as a percentage of fluid that could be removed. The percentage of fluid was expressed as mean ± SE ( n = 6–9 from 3 experiments). Data were analyzed with a one-tailed Mann–Whitney U -test. P value less than 0.05 was considered statistically significant.

Article Snippet: PD173952 was a gift from Pfizer (Ann Arbor, Michigan, USA) .

Techniques: Lysis, Sonication, Centrifugation, Immunoprecipitation, SDS Page, One-tailed Test, MANN-WHITNEY