Identification of small-molecule inhibitors of human MUS81-EME1/2 by FRET-based high-throughput screening

Area of Science:

• Molecular biology
• Drug discovery
• Cancer research

Background:

• MUS81-EME1/2 endonucleases are crucial for processing DNA replication and recombination intermediates.
• These enzymes are recognized as potential drug targets to enhance cancer therapies involving DNA-damaging agents.
• Currently, no small-molecule inhibitors for MUS81 are available.

Purpose of the Study:

• To screen for and identify small-molecule inhibitors of MUS81 complexes.
• To characterize the identified inhibitor's mechanism of action and cellular effects.

Main Methods:

• High-throughput screening using a FRET-based DNA cleavage assay.
• Biochemical assays including surface plasmon resonance (SPR) and electrophoretic mobility shift assay (EMSA).
• Cellular assays measuring γH2AX phosphorylation in HeLa cells.

Main Results:

• Dyngo-4a was identified as a potent inhibitor of MUS81-EME1 and MUS81-EME2 complexes (IC50 values of 0.57 μM and 2.90 μM, respectively).
• Dyngo-4a directly binds to MUS81 complexes (KD ≈ 0.61 μM) and inhibits DNA substrate binding.
• Dyngo-4a suppressed bleomycin-induced γH2AX phosphorylation in HeLa cells.

Conclusions:

• Dyngo-4a is a potent MUS81 inhibitor.
• This compound represents a valuable chemical tool for further research into MUS81's physiological roles.
• Dyngo-4a has potential for development to potentiate anti-cancer drug efficacy.