RTHLVFFARK-NH: A potent and selective modulator on Cu-mediated amyloid-β protein aggregation and cytotoxicity.

Dysfunctional accumulation of amyloid-β (Aβ) protein stimulated by Cu2+ is considered as a key process in the pathogenesis of Alzheimer's disease (AD). Thus, bifunctional substances capable of chelating Cu2+ and inhibiting Aβ aggregation are promising therapeutic agents against AD. Herein, a novel bifunctional decapeptide RTHLVFFARK-NH2 (RK10) was developed by integrating a metal chelating tripeptide (RTH) and an Aβ aggregation inhibitor Ac-LVFFARK-NH2 (LK7). The high selectivity of RK10 for Cu2+ over other biologically relevant metal ions was demonstrated by isothermal titration calorimetry. RK10 bound Cu2+ with a dissociation constant of 0.02 μM. This enabled RK10 to sequester Cu2+ from Aβ40-Cu2+ species and to arrest the production of reactive oxygen species (ROS) catalyzed by Cu2+ or Aβ40-Cu2+ species. Extensive physical, biophysical and biological studies indicate that RK10 targeted free and Cu2+-bound Aβ40 species, suppressed Aβ40 aggregation, and diminished the cytotoxicity induced by Aβ40 and Cu2+-mediated Aβ40 in cultured SH-SY5Y cells. Taken together, the results proved the excellent selective roles of RK10 in inhibiting Cu2+-mediated Aβ40 aggregation and eliminating ROS generation catalyzed by Cu2+/Aβ40-Cu2+ species. Thus, this work provided new insight into the design and development of potent bifunctional inhibitors against Aβ aggregation and cytotoxicity.