Journal: Cancer Medicine
Article Title: A highly sensitive screening system to evaluate the reversibility of neuroendocrine prostate cancer to prostate adenocarcinoma
doi: 10.1002/cam4.70047
Figure Lengend Snippet: Validation of hit compounds: (A) plate layout of the second screening. The negative and positive control wells, located in either end row of the plate, were prepared in the same manner as in the first screening. KUCaP13_AREluc cells, treated with a 1:3 serial dilution series of eight compounds starting at a final concentration of 10 μM, were plated, adhering to the designated positions shown in the figure. A 10 nM R1881 was added to the wells containing the compound, while it was not added to the wells of the negative control plates. Duplicate plates were tested for each plate; (B)–(D) results of three representative compounds in the second screening; (B) AR‐A014418; (C) BMS‐345541; (D) Cyt387. Data represents mean ± standard deviation (SD). The graph on the left is with R1881, and the graph on the right is without R1881; (E) the scatter plot depicts the results of the second screening. For each compound, a plot was created with the maximum relative luminescence value of the wells with R1881 on the y ‐axis and the relative luminescence value of the wells without R1881 at the concentrations that exhibited the maximum values with R1881 on the x ‐axis. The red line on the y‐axis represents the cutoff value of relative luminescence from wells with R1881 added to the compound, set at 1 + 4 SD (SD = 0.14), or 1.56. The relative luminescence from wells without R1881 added to the compound was expected to fall between the two red lines, 0.8 and 1.2. The region enclosed by the solid red line is the area where true positives are expected. The compounds shown in (B)–(D) were plotted in blue; (F, G) mRNA expression levels of AR (F) and KLK3 (G) normalized by the expression of GAPDH using quantitative qPCR. All cell types were cultured with or without 10 nM R1881. After 24 h, LNCaP cells, KUCaP13 cells, and KUCaP13_AREluc_AR cells were treated with 0.1% DMSO and cultured for a total of 72 h. In addition, KUCaP13_AREluc cells were treated with 0.1% DMSO, or 10 μM of AR‐A014418, BMS‐345541, and CYT387 after 24 h, and cultured for a total of 72 h. Data represent mean ± standard error of the mean.
Article Snippet: A GSK‐3β inhibitor AR‐A014418 (Cat no. S7435), an IKK inhibitor BMS‐345541 (Cat no. S8044), and a JAK1/JAK2 inhibitor Momelotinib (CYT387, Cat no. S2219) were purchased from Selleck Chemicals.
Techniques: Positive Control, Serial Dilution, Concentration Assay, Negative Control, Standard Deviation, Expressing, Cell Culture
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