False HDAC Inhibition by Aurone Compound.

Area of Science:

• Biochemistry
• Drug Discovery
• Chemical Biology

Background:

• Fluorescence assays are widely used for enzyme activity estimation and inhibitor screening.
• Auto-fluorescence and fluorescence quenching by compounds can compromise assay accuracy, leading to false results.

Purpose of the Study:

• To investigate the discrepancy between in vitro and cell-based assays for aurone compound 7, a potential histone deacetylase (HDAC) inhibitor.
• To identify the mechanism behind the false-positive results observed with compound 7 in fluorescence-based HDAC assays.

Main Methods:

• In vitro fluorescence-based HDAC activity assay.
• Cell-based HDAC activity assay.
• Fluorescence quenching experiments to assess compound interference.

Main Results:

• Aurone compound 7 exhibited false-positive histone deacetylase (HDAC) inhibitory activity in an in vitro fluorescence assay.
• Compound 7 did not demonstrate HDAC inhibitory activity in a cell-based assay.
• Verification experiments confirmed that compound 7 quenches the fluorescence signal, interfering with the assay.

Conclusions:

• Fluorescence assays can yield inaccurate results due to compound interference, such as fluorescence quenching.
• Careful interpretation of fluorescence assay data is crucial, especially when dealing with novel compounds.
• The findings highlight the importance of orthogonal validation methods in drug discovery.