First in class pyrrolo[2,3‐<i>d</i>]pyrimidine derivatives fused to fluorobenzylpiperidines as dual ligands of acetylcholinesterase and histamine H₃ receptor

Alzheimer's disease (AD) is a multifactorial neurodegenerative disease with manifold underlying pathophysiological mechanisms. Therefore, multitarget-directed ligands potentially offer beneficial therapeutic effects compared with classical therapies. Dual targeting of the histamine H₃ receptor (H₃R) and acetylcholinesterase (AChE) is a valid strategy for the treatment of AD. In this work, a new series of pyrrolo[2,3-d]pyrimidines fused to fluorobenzylpiperidine derivatives was designed, synthesized, and pharmacologically evaluated. Among the 16 derivatives reported here, compounds 4a (IC₅₀ = 2.19 µM for human acetylcholinesterase (hAChE) and K_i = 1.05 µM for H₃R) and 4f (IC₅₀ = 4.27 µM for hAChE and K_i = 1.31 µM for H₃R) show the most balanced dual targeting behavior coupled with moderate affinities at both targets. Selected compounds showed medium inhibition of butyrylcholinesterase (BuChE). Moreover, these compounds did not show any toxicity in the SH-SY5Y or HEK-293 cell lines at pharmacologically relevant concentrations. In silico studies allowed the proposition of binding modes and the prediction of favorable absorption, distribution, metabolism and excretion properties. The cumulative results suggest compounds 4a and 4f as lead structures for the further development of novel dual-targeted ligands for AD therapy.