Structure-Guided Discovery of Selective Indolizine-Based HDAC6 Inhibitors

Histone deacetylase 6 (HDAC6) is a cytoplasmic zinc-dependent epigenetic enzyme that mainly deacetylates non-histone proteins, which emerged as a novel molecular target to treat neuroinflammation. HDAC inhibitors have a general pharmacophoric architecture comprising a cap group (CAP), a linker region, and a zinc-binding group (ZBG), which corresponds to a hydroxamic acid for most ligands. However, all the approved inhibitors lack isoform selectivity and are therefore linked to off-target effects.3, , Selective HDAC6 inhibition represents a promising strategy to attenuate neuroinflammation while minimizing off-target effects associated with pan-HDAC inhibitors. Starting from structure–activity relationships (SAR) data of selective HDAC6 hydroxamic acid inhibitors featuring an indolizine-based recognition scaffold, we searched for a likely binding mode within HDAC6 that could account for the influence of CAP decoration on inhibitory potency and selectivity, to prospectively support the design of optimized analogues. The molecular modeling studies conducted in the present work showed that the indolizine CAP moiety occupied a cavity delimited by loop 1 (L1) and loop 2 (L2), two structural determinants for the design of selective inhibitors. Moreover, all compounds were involved in a hydrogen bond with the selectivity-determining residue Ser5684, . Experimental data from enzymatic assays was in accordance with the computational predictions, outlining the morpholine derivative (5f) as the optimal analogue in terms of potency and selectivity on HDAC6. Moreover, anti-inflammatory activity of compound 5f was validated in an in vitro hippocampal neuronal tissue model stressed with poly(I:C), which demonstrated an HDAC6-dependent reduction of IL-6 levels comparable to the one induced by approved pan-HDAC inhibitor Vorinostat (SAHA). SAR investigation through molecular docking guided the rational optimization of selective indolizine-based HDAC6 inhibitors. The findings highlight the indolizine scaffold as a valuable platform for the development of selective HDAC6 inhibitors, and the identification of a potent and selective morpholine-based derivative validates the proposed structure-based design strategy and supports the development of HDAC6-targeted modulators for neuroinflammation.