title: The use of TrkA-PathHunter assay in high-throughput screening to identify compounds that affect nerve growth factor signaling.
abstract: The TrkA-PathHunter cell-based assay was used in high-throughput screening (HTS) to identify compounds that inhibit nerve growth factor (NGF)/TrkA signaling. The assay was conducted in a 384-well format, and typical Z' values during HTS ranged from 0.3 to 0.8. The reproducibility of IC50 values was good, and the use of cryopreserved cells was well tolerated, as judged by assay parameters such as Z' and S/B and by comparison of IC50 values obtained with cells in culture. During hit deconvolution, TrkA-kinase inhibitors were identified with ATP-competitive as well as non-ATP-competitive mechanisms of action. Furthermore, other mechanisms of action such as NGF and TrkA antagonists were also identified. Because of the different molecular mechanisms identified, it is possible that subsequent optimization work to increase affinity and selectivity might lead to compounds that could have a better chance to evoke clinical efficacy without the adverse effects observed for nonselective TrkA inhibitors.
authors: AstraZeneca R&D, Neuroscience, iMed CNS&P, Södertälje, Sweden. pontus@alzacure.org
authors: Pontus
authors: P
authors: Forsell
authors: Helena
authors: H
authors: Almqvist
authors: Per
authors: P
authors: Hillertz
authors: Tomas
authors: T
authors: Akerud
authors: Magdalena
authors: M
authors: Otrocka
authors: Lina
authors: L
authors: Eisele
authors: Kai
authors: K
authors: Sun
authors: Henrik
authors: H
authors: Andersson
authors: Shephali
authors: S
authors: Trivedi
authors: Anna Ridderstad
authors: AR
authors: Wollberg
authors: Niek
authors: N
authors: Dekker
authors: Didier
authors: D
authors: Rottici
authors: Kristian
authors: K
authors: Sandberg
mesh_terms: D000255:Adenosine Triphosphate / Q000378:metabolism; D002460:Cell Line; D057166:High-Throughput Screening Assays / Q000379:methods*; D006801:Humans; D020128:Inhibitory Concentration 50; D020932:Nerve Growth Factor / Q000037:antagonists & inhibitors / Q000378:metabolism*; D016501:Neurites / Q000187:drug effects / Q000378:metabolism; D047428:Protein Kinase Inhibitors / Q000494:pharmacology*; D020917:Receptor, trkA / Q000037:antagonists & inhibitors / Q000378:metabolism*; D015398:Signal Transduction / Q000187:drug effects*
chemical_list: C502240:NGF protein, human; D047428:Protein Kinase Inhibitors; D000255:Adenosine Triphosphate; D020932:Nerve Growth Factor; D020917:Receptor, trkA
keywords: TrkA; assay development; high-throughput screening; nerve growth factor