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Assaying Protein Kinase Activity with Radiolabeled ATP
Published on: May 26, 2017
Identification and Profiling of a Selective and Brain Penetrant Radioligand for in Vivo Target Occupancy Measurement
Travis T Wager1, Paul Galatsis1, Ramalakshmi Y Chandrasekaran1,2
1Worldwide Medicinal Chemistry, Pfizer Worldwide Research and Development , 1 Portland, Cambridge, Massachusetts 02139, United States.
Abstract:
To enable the clinical development of our CNS casein kinase 1 delta/epsilon (CK1δ/ε) inhibitor project, we investigated the possibility of developing a CNS positron emission tomography (PET) radioligand. For this effort, we focused our design and synthesis efforts on the initial CK1δ/ε inhibitor HTS hits with the goal of identifying a compound that would fulfill a set of recommended PET ligand criteria. We identified [3H]PF-5236216 (9) as a tool ligand that meets most of the key CNS PET attributes including high CNS MPO PET desirability score and kinase selectivity, CNS penetration, and low nonspecific binding. We further used [3H]-9 to determine the binding affinity for PF-670462, a literature CK1δ/ε inhibitor tool compound. Lastly, [3H]-9 was used to measure in vivo target occupancy (TO) of PF-670462 in mouse and correlated TO with CK1δ/ε in vivo pharmacology (circadian rhythm modulation).
Insights
Researchers developed a novel positron emission tomography (PET) radioligand for casein kinase 1 delta/epsilon (CK1δ/ε) inhibitors. This tool enables assessment of drug efficacy and target engagement for CNS drug development.
Area of Science:
- Neuroscience
- Radiochemistry
- Pharmacology
Background:
- Casein kinase 1 delta/epsilon (CK1δ/ε) inhibitors are under investigation for central nervous system (CNS) applications.
- Developing CNS-penetrant positron emission tomography (PET) radioligands is crucial for clinical translation of CNS-targeted therapeutics.
Purpose of the Study:
- To identify and characterize a novel CNS-penetrant PET radioligand for CK1δ/ε inhibitors.
- To evaluate the suitability of the developed radioligand for assessing target engagement and pharmacology of CK1δ/ε inhibitors in vivo.
Main Methods:
- Design and synthesis of novel CK1δ/ε inhibitor-based PET ligands.
- In vitro characterization including kinase selectivity, CNS penetration, and non-specific binding assays.
- In vivo studies in mice to measure target occupancy and correlate with pharmacological effects.
Main Results:
- Identified [3H]PF-5236216 (9) as a promising PET ligand candidate with high CNS MPO PET desirability score, excellent kinase selectivity, CNS penetration, and low non-specific binding.
- Determined the binding affinity of a known CK1δ/ε inhibitor, PF-670462, using the developed radioligand.
- Demonstrated in vivo target occupancy of PF-670462 in mice and established a correlation between target occupancy and circadian rhythm modulation.
Conclusions:
- [3H]PF-5236216 (9) is a validated tool compound suitable for CNS PET imaging of CK1δ/ε.
- The developed PET radioligand facilitates the clinical development of CK1δ/ε inhibitors by enabling in vivo assessment of target engagement and pharmacology.
