Phenotypic Screening of Small-Molecule Inhibitors: Implications for Therapeutic Discovery and Drug Target Development in Traumatic Brain Injury
The inability of central nervous system (CNS) neurons to regenerate damaged axons and dendrites following traumatic brain injury (TBI) creates a substantial obstacle for functional recovery. Apoptotic cell death, deposition of scar tissue, and growth-repressive molecules produced by glia further complicate the problem and make it challenging for re-growing axons to extend across injury sites. To date, there are no approved drugs for the treatment of TBI, accentuating the need for relevant leads. Cell-based and organotypic bioassays can better mimic outcomes within the native CNS microenvironment than target-based screening methods and thus should speed the discovery of therapeutic agents that induce axon or dendrite regeneration. Additionally, when used to screen focused chemical libraries such as small-molecule protein kinase inhibitors, these assays can help elucidate molecular mechanisms involved in neurite outgrowth and regeneration as well as identify novel drug targets. Here, we describe a phenotypic cellular (high content) screening assay that utilizes brain-derived primary neurons for screening small-molecule chemical libraries.
Key wordsHigh-content screening Primary neurons Cell-based assay Axon regeneration CNS injury Kinase inhibitor Drug discovery
- 1.Centers for Disease Control and Prevention (2013) Injury prevention and control: traumatic brain injury. Centers for Disease Control and Prevention, Washington, DCGoogle Scholar
- 5.Blackmore MG, Moore DL, Smith RP, Goldberg JL, Bixby L, Lemmon VP (2011) Regulators of axon growth. Mol Cell 44:43–54Google Scholar
- 18.Al-Ali H, Blackmore M, Bixby JL, Lemmon VP (2013) High content screening with primary neurons. Assay Guid ManGoogle Scholar
- 33.Welsbie DS, Yang Z, Ge Y, Mitchell KL, Zhou X, Martin SE, Berlinicke CA, Hackler L, Fuller J, Fu J, Cao L-H, Han B, Auld D, Xue T, Hirai S-I, Germain L, Simard-Bisson C, Blouin R, Nguyen JV, Davis C-HO, Enke RA, Boye SL, Merbs SL, Marsh-Armstrong N, Hauswirth WW, Diantonio A, Nickells RW, Inglese J, Hanes J, Yau K-W, Quigley HA, Zack DJ (2013) Functional genomic screening identifies dual leucine zipper kinase as a key mediator of retinal ganglion cell death. Proc Natl Acad Sci U S A 110:4045–4050CrossRefPubMedPubMedCentralGoogle Scholar
- 36.Meberg PJ, Miller MW (2003) Culturing hippocampal and cortical neurons. Methods Cell Biol 71:111–127Google Scholar