Inhibiting and Remodeling Toxic Amyloid-Beta Oligomer Formation Using a Computationally Designed Drug Molecule That Targets Alzheimer's Disease

Abstract

Alzheimer's disease (AD) is rapidly reaching epidemic status among a burgeoning aging population. Much evidence suggests the toxicity of this amyloid disease is most influenced by the formation of soluble oligomeric forms of amyloid β-protein, particularly the 42-residue alloform (Aβ42). Developing potential therapeutics in a directed, streamlined approach to treating this disease is necessary. Here we utilize the joint pharmacophore space (JPS) model to design a new molecule [AC0107] incorporating structural characteristics of known Aβ inhibitors, blood-brain barrier permeability, and limited toxicity. To test the molecule's efficacy experimentally, we employed ion mobility mass spectrometry (IM-MS) to discover [AC0107] inhibits the formation of the toxic Aβ42 dodecamer at both high (1:10) and equimolar concentrations of inhibitor. Atomic force microscopy (AFM) experiments reveal that [AC0107] prevents further aggregation of Aβ42, destabilizes preformed fibrils, and reverses Aβ42 aggregation. This trend continues for long-term interaction times of 2 days until only small aggregates remain with virtually no fibrils or higher order oligomers surviving. Pairing JPS with IM-MS and AFM presents a powerful and effective first step for AD drug development. Graphical Abstract.

Keywords: Aggregation; Alzheimer’s disease; Amyloid-β protein; Atomic force microscopy; Aβ42; Inhibition; Ion mobility mass spectrometry; Joint pharmacophore space.

Figure 1.

a-c) Representative mass spectra for A03B242 alone, Aβ42 + 1:10 [AC0107], Aβ42 + 1:1 [AC0107], respectively. d) z/n= −5/2 ATD for Aβ42 alone. e-f) z/n= −5/2 ATDs at 1 hour co-incubation with 1:10 and 1:1 [AC0107], respectively. g-h) z/n= −5/2 ATD at 24 hours co-incubation with 1:10 and 1:1 [AC0107], respectively. The injection energy for each ATD is 40V. Each ATD has fitted structures with labels corresponding to Aβ42 dodecamer (n=12, purple fit), decamer (n=10, orange fit), hexamer (n=6, green fit), tetramer (n=4, red fit), and dimer (n=2, blue fit). The peak fitting procedure is outlined in the Supporting Information.

Figure 2.

AFM height images for representative 10 μM Aβ42 incubated alone at room temperature in solution for a) 5 min, b) 30 min, and c) 60 min. At 60 minutes, [AC0107] was added to the same solution of Aβ42 to a final concentration of 1:10 [AC0107] with aliquots taken and imaged at d) 5 min, e) 30 min, and f) 60 min co-incubation. g), h), i) are 5 min, 30 min, and 60 min co-incubation times, respectively, for 1:1 Aβ42:[AC0107] concentration after 60 minutes of Aβ42 incubated alone. Each image is 2 × 2 μm in dimension. Lighter colors are taller structures with the darkest representing the mica background surface. Particle height distributions are included in the Supporting Information.

Figure 3.

AFM phase images of 10 μM Aβ42 alone at a) 5 min, b) 30 min, and c) 60 min incubation in solution. After 60 minutes, an equimolar 1:1 Aβ42:[AC0107] solution was achieved in the same solution and aliquots were removed and imaged at d) 5min, e) 30 min, and f) 60 min co-incubation. Relative to background media of the image, darker colors are physically harder and more structured than the surrounding material. Lighter colors are softer and less structured. Each image is 500nm × 500nm.

Figure 4.

AFM images at a) 24 hours co-incubation of 1:10 Aβ42:[AC0107] and at c) 48 hours. AFM images at d) 24 hours co-incubation of 1:1 Aβ42:[AC0107] and at f) 48 hours. b) and e) are zoomed in (500nm × 500nm) images of a) 1:10 and d) 1:1, respectively.

Scheme 1.

Peptide Sequence of Aβ42.

Scheme 2.

[AC0107]. 4-({[3-(1-Pyrrolidinylmethyl)benzyl]amino}methyl)benzonitrile