Identification of a mechanism-based binding mode for a histone deacetylase 6 inhibitor
- Daniel A. Rodrigues
- Yu Wang
- Juana Goulart Stollmaier
- Graeme P. Sullivan
- Cian D’Arcy
- Aisling Y. Coughlan
- Andrew Roe
- Linda Bíró
- Paris R. Watson
- Jeremy D. Osko
- Brendan Twamley
- Kieran Wynne
- Gerard Cagney
- Péter Buglyó
- Yanli Liu
- Darren M. Griffith
- David W. Christianson
- Tríona Ní Chonghaile
2026-06-05
Histone deacetylase 6 (HDAC6) is a cytoplasmic enzyme that deacetylates non-histone substrates such as α-tubulin and cortactin. HDAC6 contains two catalytic domains, each containing a catalytic zinc ion, and a zinc-finger ubiquitin-binding domain. We have discovered BAS-2, a selective HDAC6 inhibitor with an isothiouronium core and no obvious zinc-binding group. To define its mechanism, we combine X-ray crystallography, structure-activity-relationships, molecular modeling and mutagenesis. BAS-2 potently inhibits human HDAC6 but it does not inhibit zebrafish HDAC6. Computational modeling highlighted Asp567 in human HDAC6 as critical for BAS-2 recognition and mutational analyses confirmed this. The corresponding zebrafish residue is Asn530 and the crystal structure of the N530D variant zHDAC6 revealed binding of a BAS-2–derived mercaptoacetamide that engages the catalytic zinc via strong thiolate–zinc coordination. Leveraging the orientation of BAS-2 binding, we designed a BAS-2–based proteolysis targeting chimera that induced proteasome-dependent HDAC6 degradation in cells, verified by global proteomics. Collectively, these insights clarify species selectivity and demonstrate that BAS-2 acts as a selective, mechanism-based inhibitor of human HDAC6. These discoveries will aid the development of the next generation of selective HDAC6 inhibitors and degraders.