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Peer-reviewedCC BY-NC-NDACS chemical neuroscience

Development of NanoBRET Cellular Target Engagement Assays in Primary Neurons for Activating Mutants of p21-Activated Kinase 1.

  • Jacob L Capener1
  • Abdiel Badillo-Martinez2
  • Bassel Awada3
  • Zachary W Davis-Gilbert1
  • Thomas W Kramer1
  • Cameron S Blair1
  • Frances M Bashore1
  • Hassan Al-Ali3,4
  • Alison D Axtman1
4 affiliations
  1. 1Structural Genomics Consortium, UNC Eshelman School of Pharmacy, University of North Carolina at Chapel Hill, Chapel Hill, North Carolina27599, United States.
  2. 2Department of Biochemistry and Molecular Biology, University of Miami Miller School of Medicine, Miami, Florida33136, United States.
  3. 3The Miami Project to Cure Paralysis, University of Miami, Miami, Florida33136, United States.
  4. 4Department of Neurological Surgery, Peggy and Harold Katz Drug Discovery Center, Sylvester Cancer Comprehensive Center, and Frost Institute for Data Science and Computing, University of Miami, Miami, Florida33136, United States.
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Abstract

The p21-activated kinases (PAKs) are a group of serine-threonine kinases central to multiple signaling pathways that govern cell survival and proliferation. Aberrant activity of PAK1, the most well characterized member of the PAK family, drives progression of several malignancies and brain disorders, including Alzheimer's disease and neurodevelopmental disorders. Despite growing interest in PAK1 as a drug target for these diseases, there is no assay to evaluate the intracellular target engagement of PAK1 inhibitors. To address this need, we developed first-in-class NanoBRET assays for wild-type PAK1 and a neurodevelopmental disorder-causing gain-of-function PAK1 mutant. Furthermore, we executed our novel PAK1 NanoBRET assay to evaluate target engagement of PAK1 inhibitors in primary hippocampal neurons. To the best of our knowledge, this is the first demonstration of a NanoBRET cellular target engagement assay in primary neurons, thereby increasing the relevance of our work by confirming PAK1 inhibitor binding to the aberrant form of the protein in cerebral cells.

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Cite

BibTeXRIS
BibTeX
@article{capener2026development,
  title = {{Development of NanoBRET Cellular Target Engagement Assays in Primary Neurons for Activating Mutants of p21-Activated Kinase 1.}},
  author = {Jacob L Capener and Abdiel Badillo-Martinez and Bassel Awada and Zachary W Davis-Gilbert and Thomas W Kramer and Cameron S Blair and Frances M Bashore and Hassan Al-Ali and Alison D Axtman},
  journal = {ACS chemical neuroscience},
  year = {2026},
  month = oct,
  doi = {10.1021/acschemneuro.6c00326},
  url = {https://europepmc.org/article/MED/42904645},
  note = {CC BY-NC-ND},
}
RIS
TY  - JOUR
TI  - Development of NanoBRET Cellular Target Engagement Assays in Primary Neurons for Activating Mutants of p21-Activated Kinase 1.
AU  - Jacob L Capener
AU  - Abdiel Badillo-Martinez
AU  - Bassel Awada
AU  - Zachary W Davis-Gilbert
AU  - Thomas W Kramer
AU  - Cameron S Blair
AU  - Frances M Bashore
AU  - Hassan Al-Ali
AU  - Alison D Axtman
PY  - 2026
DA  - 2026/10/01
JO  - ACS chemical neuroscience
DO  - 10.1021/acschemneuro.6c00326
UR  - https://europepmc.org/article/MED/42904645
AB  - The p21-activated kinases (PAKs) are a group of serine-threonine kinases central to multiple signaling pathways that govern cell survival and proliferation. Aberrant activity of PAK1, the most well characterized member of the PAK family, drives progression of several malignancies and brain disorders, including Alzheimer's disease and neurodevelopmental disorders. Despite growing interest in PAK1 as a drug target for these diseases, there is no assay to evaluate the intracellular target engagement of PAK1 inhibitors. To address this need, we developed first-in-class NanoBRET assays for wild-type PAK1 and a neurodevelopmental disorder-causing gain-of-function PAK1 mutant. Furthermore, we executed our novel PAK1 NanoBRET assay to evaluate target engagement of PAK1 inhibitors in primary hippocampal neurons. To the best of our knowledge, this is the first demonstration of a NanoBRET cellular target engagement assay in primary neurons, thereby increasing the relevance of our work by confirming PAK1 inhibitor binding to the aberrant form of the protein in cerebral cells.
SN  - 1948-7193
ER  -