Abstract
<jats:p>Histone deacetylases (HDACs) are key epigenetic regulators of gene expression and often dysregulated in cancers. In particular class I HDACs are frequently overexpressed in solid cancers, including glioblastomas. Whereas HDAC inhibitors mostly show limited effectiveness in monotherapy, they often act synergistically in combination with anticancer agents, particularly DNA-damaging agents. One approach to make use of this synergy is to combine two pharmacophores in one hybrid molecule. This study aimed to combine a class I selective HDAC inhibitor with an alkylating agent in a dual-targeting hybrid molecule to improve the efficacy of alkylating agents in glioblastomas. Structural variations in the cap region of the class I selective HDAC inhibitor VK1 were introduced to accommodate the nitrogen mustard chlorambucil into the HDAC inhibitor. This resulted in a chlorambucil-type series and a truncated series lacking the propyl linker of chlorambucil. Biological evaluation revealed that the truncated series acted primarily as HDAC inhibitor, whereas the chlorambucil-HDAC inhibitor hybrid compounds induced strong DNA damage estimated as γH2AX immunostaining. Notably, the most potent hybrid compound induced stronger DNA damage, caspase activation, and cytotoxicity due to synergistic interaction than the combination of chlorambucil plus the HDAC inhibitor. In conclusion, the synergy of the two pharmacophores, HDAC inhibitor and nitrogen mustard group, combined in one hybrid compound, highlights the superior efficacy and potency of the dual-targeting approach over the combination of two single-warhead inhibitors, which may be of particular interest in glioblastomas.</jats:p>