scholarly journals A high-throughput screening strategy identifies cardiotonic steroids as alternative splicing modulators

2008 ◽  
Vol 105 (32) ◽  
pp. 11218-11223 ◽  
Author(s):  
P. Stoilov ◽  
C.-H. Lin ◽  
R. Damoiseaux ◽  
J. Nikolic ◽  
D. L. Black
2021 ◽  
pp. 247255522110262
Author(s):  
Jonathan Choy ◽  
Yanqing Kan ◽  
Steve Cifelli ◽  
Josephine Johnson ◽  
Michelle Chen ◽  
...  

High-throughput phenotypic screening is a key driver for the identification of novel chemical matter in drug discovery for challenging targets, especially for those with an unclear mechanism of pathology. For toxic or gain-of-function proteins, small-molecule suppressors are a targeting/therapeutic strategy that has been successfully applied. As with other high-throughput screens, the screening strategy and proper assays are critical for successfully identifying selective suppressors of the target of interest. We executed a small-molecule suppressor screen to identify compounds that specifically reduce apolipoprotein L1 (APOL1) protein levels, a genetically validated target associated with increased risk of chronic kidney disease. To enable this study, we developed homogeneous time-resolved fluorescence (HTRF) assays to measure intracellular APOL1 and apolipoprotein L2 (APOL2) protein levels and miniaturized them to 1536-well format. The APOL1 HTRF assay served as the primary assay, and the APOL2 and a commercially available p53 HTRF assay were applied as counterscreens. Cell viability was also measured with CellTiter-Glo to assess the cytotoxicity of compounds. From a 310,000-compound screening library, we identified 1490 confirmed primary hits with 12 different profiles. One hundred fifty-three hits selectively reduced APOL1 in 786-O, a renal cell adenocarcinoma cell line. Thirty-one of these selective suppressors also reduced APOL1 levels in conditionally immortalized human podocytes. The activity and specificity of seven resynthesized compounds were validated in both 786-O and podocytes.


2019 ◽  
Author(s):  
Laurence Jadin ◽  
Hiba Shaban ◽  
Anirban Kundu ◽  
Gregory Schreiber ◽  
Scooter Willis ◽  
...  

2019 ◽  
Author(s):  
Laurence Jadin ◽  
Hiba Shaban ◽  
Anirban Kundu ◽  
Gregory Schreiber ◽  
Scooter Willis ◽  
...  

2001 ◽  
Vol 6 (6) ◽  
pp. 429-440 ◽  
Author(s):  
Michael W. Pantoliano ◽  
Eugene C. Petrella ◽  
Joseph D. Kwasnoski ◽  
Victor S. Lobanov ◽  
James Myslik ◽  
...  

More general and universally applicable drug discovery assay technologies are needed in order to keep pace with the recent advances in combinatorial chemistry and genomics-based target generation. Ligand-induced conformational stabilization of proteins is a well-understood phenomenon in which substrates, inhibitors, cofactors, and even other proteins provide enhanced stability to proteins on binding. This phenomenon is based on the energetic coupling of the ligand-binding and protein-melting reactions. In an attempt to harness these biophysical properties for drug discovery, fully automated instrumentation was designed and implemented to perform miniaturized fluorescence-based thermal shift assays in a microplate format for the high throughput screening of compound libraries. Validation of this process and instrumentation was achieved by investigating ligand binding to more than 100 protein targets. The general applicability of the thermal shift screening strategy was found to be an important advantage because it circumvents the need to design and retool new assays with each new therapeutic target. Moreover, the miniaturized thermal shift assay methodology does not require any prior knowledge of a therapeutic target's function, making it ideally suited for the quantitative high throughput drug screening and evaluation of targets derived from genomics.


2011 ◽  
Vol 85 (3) ◽  
pp. 214-220 ◽  
Author(s):  
Zhi-Jian Lin ◽  
Ren-Chao Zheng ◽  
Li-Hua Lei ◽  
Yu-Guo Zheng ◽  
Yin-Chu Shen

2012 ◽  
Vol 21 (6) ◽  
pp. 1603-1610 ◽  
Author(s):  
Jun Tan ◽  
Ju Chu ◽  
Wenjuan Shi ◽  
Cheng Lin ◽  
Yuanxin Guo ◽  
...  

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