pld2 inhibitor cay10594  (Cayman Chemical)

Journal: Cancer Research

Article Title: DLGAP1-AS2–Mediated Phosphatidic Acid Synthesis Activates YAP Signaling and Confers Chemoresistance in Squamous Cell Carcinoma

doi: 10.1158/0008-5472.can-22-0717

Figure Lengend Snippet: Figure 6. D-AS2 promotes PLD/PA-mediated YAP activation. A, Western blot analyses of YAP and LATS1 phosphorylation. KYSE450 cells expressing EV or D-AS2 were pretreated with FIPI (30 mmol/L) or CAY10594 (20 mmol/L) for 1 hour. Representative results of at least three biological replicates are shown. B, qRT-PCR detection of the YAP target genes CTGF and CYR61. KYSE450 cells expressing EV or D-AS2 were pretreated with FIPI (30 mmol/L) or CAY10594 (20 mmol/L) for 1 hour. The data are presented as the mean SD values; two-tailed t test; , P < 0.001; n ¼ 3 technical replicates. C and D, IF detection of YAP localization in KYSE450 cells expressing EV or D-AS2. Cells were pre- treated with FIPI (30 mmol/L) or CAY10594 (20 mmol/L). YAP localization is shown in C. Scale bar, 30 mm. Cells from three different fields in three independent experiments were randomly selected for quantification of YAP localization (D). The data are expressed as the mean SEM values. E, Western blot analyses of YAP and LATS1 phosphorylation. Serum-starved KYSE30 cells expressing control or D-AS2–target- ing sgRNAs were pretreated with PA (100 mmol/L) for 1 hour. Representative results of at least three biological replicates are shown. F, qRT-PCR detection of the YAP target genes CTGF and CYR61. Serum- starved KYSE30 cells expressing control or D-AS2–targeting sgRNAs were pretreated with PA (100 mmol/L) for 1 hour. The data are presented as the mean SD values; two-tailed t test; , P < 0.001; ns, not significant; n ¼ 3 technical replicates. G and H, IF detection of YAP localization in KYSE30 cells expressing control and D-AS2–targeting sgRNAs and pretreated with PA (100 mmol/L) for 1 hour. YAP local- ization is shown in G. Scale bar, 30 mm. Cells from three different fields in three independent experiments were randomly selected for quantification of YAP localiza- tion (H). The data are expressed as the mean SEM values. I, qRT-PCR detection of the expression of the indicated genes in the samples with chemoresistance or chemosensitivity of patients with esoph- ageal SCC. The data are presented as the mean SEM values; two-tailed t test; n ¼ 17 for chemoresistant patients; n ¼ 17 for chemosensitive patients.

Article Snippet: FIPI (939055-18-2) and CAY10594 (1130067-34-3) were purchased from Topscience.

Techniques: Activation Assay, Western Blot, Phospho-proteomics, Expressing, Quantitative RT-PCR, Two Tailed Test, Control

Journal: bioRxiv

Article Title: DLGAP1-AS2-Mediated Phosphatidic Acid Synthesis Confers Chemoresistance via Activation of YAP Signaling

doi: 10.1101/2022.02.24.481869

Figure Lengend Snippet: ( A ) IB detection of YAP and LATS1 phosphorylation. KYSE450 cells expressing empty vector (EV) and D-AS2 were pretreated with FIPI (30 µM) or CAY10594 (20 µM) for 1 h. ( B ) RT-qPCR detection of the YAP target genes CTGF and CYR61. KYSE450 cells expressing EV and D-AS2 were pretreated with FIPI (30 µM) or CAY10594 (20 µM) for 1 h. The data are presented as the mean ± s.d. values; two-tailed t test, *** P < 0.001; n = 3. ( C, D ) IF detection of YAP localization in KYSE450 cells expressing EV and D-AS2. Cells were pretreated with FIPI (30 µM) or CAY10594 (20 µM). YAP localization is shown in (C); scale bar, 30 µm. Cells from three different fields in three independent experiments were randomly selected for quantification of YAP localization (D). The data are expressed as the mean ± s.e.m. values. ( E ) IB detection of YAP and LATS1 phosphorylation. Serum-starved KYSE30 cells expressing control or D-AS2-targeting sgRNAs were pretreated with PA (100 µM) for 1 h. ( F ) RT-qPCR detection of the YAP target genes CTGF and CYR61. Serum-starved KYSE30 cells expressing control or D-AS2-targeting sgRNAs were pretreated with PA (100 µM) for 1 h. The data are presented as the mean ± s.d. values; two-tailed t test, *** P < 0.001, ns: not significant; n = 3. ( G, H ) IF detection of YAP localization in KYSE30 cells expressing control and D-AS2-targeting sgRNAs and pretreated with PA (100 µM) for 1 h. YAP localization is shown in (G); scale bar, 30 µm. Cells from three different fields in three independent experiments were randomly selected for quantification of YAP localization (H). The data are expressed as the mean ± s.e.m. values. ( I ) RT-qPCR detection of the indicated genes expression in the samples with chemoresistance or chemosensitivity of esophageal SCC patients. The data are presented as the mean ± s.e.m. values; two-tailed t test; n = 17 for chemoresistant patients; n = 17 for chemosensitive patients.

Article Snippet: FIPI (939055-18-2) and CAY10594 (1130067-34-3) were purchased from Topscience (Topscience; Shanghai, China).

Techniques: Expressing, Plasmid Preparation, Quantitative RT-PCR, Two Tailed Test