Chemical Interrogation of the neuronal kinome using a primary cell-based screening assay

ACS Chem Biol., 2013 · DOI: 10.1021/cb300584e · Published: May 17, 2013

Simple Explanation

This study addresses the challenge of limited axon regeneration in the adult central nervous system following spinal cord injury (SCI) and traumatic brain injury. The researchers developed a cell-based assay using primary rat hippocampal neurons to identify small molecule kinase inhibitors that promote neurite growth. A library of protein kinase inhibitors was screened, identifying compounds that consistently promoted neurite growth, with one hit demonstrating axon growth promotion in a cortical slice culture assay.

Study Duration

Not specified

Participants

Primary rat hippocampal neurons

Evidence Level

Not specified

Key Findings

1
A robust cell-based assay with a Z′-factor of 0.73 was developed, indicating high reliability and suitability for medium throughput screening.
2
Several compound groups within the screened kinase inhibitor library were found to strongly and consistently promote neurite growth in primary rat hippocampal neurons.
3
The best performing compound, ML-7, promoted axon growth in a postnatal cortical slice culture assay, suggesting potential for CNS application.

Research Summary

The study developed a high-content analysis (HCA) screening bioassay using primary mammalian neurons to identify small-molecule PKIs that can promote neurite growth. Screening a library of 240 PKIs revealed several hits, defined as compounds increasing neurite total length (NTL) ≥ 25% relative to control, with a Z score > 1.5. The best-performing hit, ML-7, was found to promote axon growth in a postnatal cortical slice culture, demonstrating potential for CNS applications.

Practical Implications

Drug Discovery

The study provides a starting point for developing PKIs with better neurite growth-promotion activity and toxicity profiles for treating SCI and related neurological pathologies.

Target Identification

Identified kinases, such as ROCKs, PKCs, CDKs, and RSKs, are potential targets for growth promoting PKIs.

Assay Validation

The developed phenotypic screen of PKIs, combined with knowledge of compound activities, can be used to identify suitable drug targets.

Study Limitations

1
The number of structurally related compounds within the library is too small for extensive structure-activity relationship (SAR) analyses.
2
The exact mechanisms of action of the identified compounds and their corresponding targets remain to be fully elucidated.
3
The possibility that correlations between neurite parameters may reflect target bias in the EMD library cannot be discounted.

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