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Updated: Mar 10, 2026
Single-Step Enrichment of a TAP-Tagged Histone Deacetylase of the Filamentous Fungus Aspergillus nidulans for Enzymatic Activity Assay
Published on: May 1, 2019
UHPLC-MS-based HDAC Assay Applied to Bio-guided Microfractionation of Fungal Extracts
Vincent Zwick1, Pierre-Marie Allard1, Lucie Ory1
1School of Pharmaceutical Sciences, University of Geneva, University of Lausanne, CMU - Rue Michel Servet 1, 1211, Geneva, 11, Switzerland.
Introduction:
Histone deacetylases (HDAC) are considered as promising targets for cancer treatment. Today, four HDAC inhibitors, vorinostat, romidepsin, belinostat, and panobinostat, have been approved by the Food and Drug Administration (FDA) for cancer treatment, while others are in clinical trials. Among them, several are naturally occurring fungal metabolites.
Objective:
To develop and optimise an enzyme assay for bio-guided identification of HDAC inhibitors in fungal strains.
Methods:
Fluorescence and MS-based HDAC enzymatic assays were compared during the bio-guided fractionation of Penicillium griseofulvum. The MS-based approach was then optimised to evaluate HDAC selectivity using the human recombinant class I isoform HDAC1 and the class II isoform HDAC6.
Results:
Fluorescence-based assays have several drawbacks when used for bio-guided fractionation because of the native fluorescence and the trypsin inhibitory ability of compounds present in many extracts. The MS-based method led to the isolation of gliocladride C, which is selective for HDAC1 and salirepol, which showed an HDAC6 selectivity. Their activity and presence in P. griseofulvum is described here for the first time.
Conclusion:
The UHPLC-ESI-MS/MS-based method using specific HDAC isoforms is suitable to isolate selective HDAC inhibitors by bio-guided fractionation of fungal strains. Also, it decreases potential interferences with natural products compared to the fluorescence-based assay.
Insights
A new mass spectrometry (MS)-based assay effectively identifies selective histone deacetylase (HDAC) inhibitors from fungal extracts. This method overcomes limitations of fluorescence assays, enabling discovery of novel cancer drug candidates.
Area of Science:
- Biochemistry
- Natural Product Chemistry
- Pharmacology
Background:
- Histone deacetylases (HDAC) are crucial targets in cancer therapy, with several FDA-approved inhibitors.
- Naturally occurring fungal metabolites represent a promising source of novel HDAC inhibitors for drug development.
Purpose of the Study:
- To develop and optimize an enzyme assay for the bio-guided identification of HDAC inhibitors from fungal strains.
- To compare fluorescence-based and MS-based assays for efficiency and accuracy in identifying HDAC inhibitors.
Main Methods:
- Comparative analysis of fluorescence and MS-based HDAC enzymatic assays during bio-guided fractionation of Penicillium griseofulvum.
- Optimization of the MS-based approach for evaluating HDAC isoform selectivity (HDAC1 and HDAC6).
Main Results:
- Fluorescence assays exhibited drawbacks due to native fluorescence and trypsin inhibitory compounds in extracts.
- The MS-based method successfully isolated gliocladride C (HDAC1 selective) and salirepol (HDAC6 selective) from P. griseofulvum.
- The activity and presence of these selective inhibitors in P. griseofulvum were reported for the first time.
Conclusions:
- The UHPLC-ESI-MS/MS-based method with specific HDAC isoforms is effective for isolating selective HDAC inhibitors via bio-guided fractionation of fungal strains.
- This MS-based assay minimizes interferences from natural products compared to traditional fluorescence-based methods.
