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Application of beta-lactamase enzyme complementation to the high-throughput screening of toll-like receptor signaling inhibitors.
| Content Provider | Semantic Scholar |
|---|---|
| Author | Lee, Hyun-Ku Brown, Steven J. Rosen, Hugh Tobias, Peter S. |
| Copyright Year | 2007 |
| Abstract | We describe a successful application of beta-lactamase fragment complementation to high-throughput screening (HTS) for Toll-like receptor 4 (TLR4) signaling inhibitors. We developed a stable cell line, HeLa/CL3-4, expressing MyD88/Bla(a) and TLR4/Bla(b), in which the two beta-lactamase fragments complement with each other by virtue of spontaneous MyD88-TLR4 binding via their Toll/IL-1R (TIR) domains. Inhibition of the MyD88-TLR4 binding leads to the disruption of the enzyme complementation and a loss of the lactamase activity. We used a 384-well plate format to screen 16,000 compounds using this assay and obtained 45 primary hits. After rescreening these 45 hits and eliminating compounds that directly inhibited beta-lactamase, we had five candidate inhibitors. We show that these five act as inhibitors of TLR4-MyD88 binding and are variously effective at inhibiting lipopolysaccharide-stimulated cytokine release from RAW264.7 cells. One compound is effective near 100 nM. None of the compounds showed any cytotoxicity at 20 microM. |
| File Format | PDF HTM / HTML |
| DOI | 10.1124/mol.107.038349 |
| PubMed reference number | 17615244 |
| Journal | Medline |
| Volume Number | 72 |
| Issue Number | 4 |
| Alternate Webpage(s) | http://molpharm.aspetjournals.org/content/molpharm/72/4/868.full.pdf?with-ds=yes |
| Alternate Webpage(s) | http://molpharm.aspetjournals.org/content/molpharm/72/4/868.full.pdf |
| Alternate Webpage(s) | https://doi.org/10.1124/mol.107.038349 |
| Journal | Molecular pharmacology |
| Language | English |
| Access Restriction | Open |
| Content Type | Text |
| Resource Type | Article |
