Development of a Highly Selective NanoBRET Probe to Assess MAGL Inhibition in Live Cells

Area of Science:

• Biochemistry
• Pharmacology
• Drug Discovery

Background:

• Cell-free assays are scalable but fail to predict cellular pharmacology due to factors like permeability and protein binding.
• These limitations can lead to poor lead compound selection in early drug development.
• A need exists for cell-based assays that accurately reflect intracellular target engagement.

Purpose of the Study:

• To develop and validate a novel intracellular NanoBRET assay for assessing MAGL (monoacylglycerol lipase) inhibition in live cells.
• To enable simultaneous evaluation of target engagement and critical physicochemical properties.
• To accelerate the early stages of drug discovery by improving compound profiling.

Main Methods:

• Development of a NanoBRET assay using a BODIPY590-conjugated inhibitor and a MAGL-nanoluciferase fusion protein.
• Reverse design approach for inhibitor conjugation to maintain drug properties.
• Validation using known MAGL inhibitors and screening of over 1,900 drug candidates.

Main Results:

• Successful development of a robust intracellular NanoBRET assay for MAGL.
• Demonstrated ability to assess both target engagement and physicochemical properties (permeability, protein binding) in live cells.
• Facilitated selection of drug candidates with improved profiles for further development.

Conclusions:

• Cell-based target engagement assays, like the developed NanoBRET assay, are crucial for accurate early-stage drug discovery.
• This approach mitigates risks associated with cell-free assays, leading to more effective lead compound selection.
• The assay accelerates compound profiling and progression in drug discovery programs.