New promising transthyretin stabilizers for cardiac amyloidosis treatment

Transthyretin (TTR) is a plasma protein responsible for transport of thyroxine and retinol, the latter through binding with the retinol-binding protein. In contrast with its physiological role, TTR possesses an intrinsic amyloidogenic potential due to the high content of β-strands. Under pathological conditions, TTR undergoes a misfolding process forming protein aggregates and fibrils that cause organ damage and dysfunction, leading to the onset of TTR amyloidosis diseases (ATTRs). One therapeutic approach against ATTRs progression involves administering small molecules able to bind the inner channel of the protein enhancing tetramer stability. Here, we report the synthesis and characterization of 2-(((benzyloxy)imino)methyl)benzoic acids (1a-l), 3-(((benzyloxy)imino)methyl)benzoic acids (2a-l) and 4-(((benzyloxy)imino)methyl)benzoic acids (3a-l) as new potential inhibitors of TTR fibril formation. Our study shows that compounds 1a((E)-2-((((2-(trifluoromethyl)benzyl)oxy)imino)methyl)benzoic acid)), 1d((E)-2-((((2-chlorobenzyl)oxy)imino)methyl)benzoic acid), 2l((E)-3-((((2-chloro-6-fluorobenzyl)oxy)imino)methyl)benzoic acid) and 3g((E)-4-((((2-methoxybenzyl)oxy)imino)methyl)benzoic acid) effectively inhibit the aggregation of wild-type TTR (wt-TTR) and mutants (V30M, V122I). Among these derivatives 1a and 1d have stronger ability to contrast the fibril formations. The binding affinity of 1a, 1d, 2l with TTR (wt, V30M and V122I) was determined by fluorescence displacement assay and isothermal titration calorimetry. The binding modes of the selected ligands have been confirmed by X-ray crystallography. Moreover, these compounds show no toxicity in endothelial and cardiomyocyte cells, reinforcing their potential as TTR stabilizers.