The cysteine protease inhibitor, E64d, reduces brain amyloid-β and improves memory deficits in Alzheimer's disease animal models by inhibiting cathepsin B, but not BACE1, β-secretase activity

J Alzheimers Dis. 2011;26(2):387-408. doi: 10.3233/JAD-2011-110101.


The cysteine protease cathepsin B is a potential drug target for reducing brain amyloid-β (Aβ) and improving memory in Alzheimer's disease (AD), as reduction of cathepsin B in transgenic mice expressing human wild-type amyloid-β protein precursor (AβPP) results in significantly decreased brain Aβ. Cathepsin B cleaves the wild-type β-secretase site sequence in AβPP to produce Aβ, and cathepsin B inhibitors administered to animal models expressing AβPP containing the wild-type β-secretase site sequence reduce brain Aβ in a manner consistent with β-secretase inhibition. But such inhibitors could act either by direct inhibition of cathepsin B β-secretase activity or by off-target inhibition of the other β-secretase, the aspartyl protease BACE1. To evaluate that issue, we orally administered a cysteine protease inhibitor, E64d, to normal guinea pigs or transgenic mice expressing human AβPP, both of which express the human wild-type β-secretase site sequence. In guinea pigs, oral E64d administration caused a dose-dependent reduction of up to 92% in brain, CSF, and plasma of Aβ40 and Aβ42, a reduction of up to 50% in the C-terminal β-secretase fragment (CTFβ), and a 91% reduction in brain cathepsin B activity, but increased brain BACE1 activity by 20%. In transgenic AD mice, oral E64d administration improved memory deficits and reduced brain Aβ40 and Aβ42, amyloid plaque, brain CTFβ, and brain cathepsin B activity, but increased brain BACE1 activity. We conclude that E64d likely reduces brain Aβ by inhibiting cathepsin B and not BACE1 β-secretase activity and that E64d therefore may have potential for treating AD patients.

Publication types

  • Research Support, N.I.H., Extramural
  • Research Support, Non-U.S. Gov't

MeSH terms

  • Alzheimer Disease / drug therapy*
  • Alzheimer Disease / metabolism
  • Alzheimer Disease / pathology
  • Amyloid Precursor Protein Secretases / antagonists & inhibitors
  • Amyloid Precursor Protein Secretases / metabolism*
  • Amyloid beta-Peptides / metabolism*
  • Animals
  • Aspartic Acid Endopeptidases / antagonists & inhibitors
  • Aspartic Acid Endopeptidases / metabolism*
  • Brain / drug effects*
  • Brain / metabolism
  • Brain / pathology
  • Cathepsin B / antagonists & inhibitors*
  • Cathepsin B / metabolism
  • Cysteine Proteinase Inhibitors / pharmacology*
  • Cysteine Proteinase Inhibitors / therapeutic use
  • Disease Models, Animal
  • Guinea Pigs
  • Leucine / analogs & derivatives*
  • Leucine / pharmacology
  • Leucine / therapeutic use
  • Memory / drug effects*
  • Mice
  • Mice, Transgenic


  • Amyloid beta-Peptides
  • Cysteine Proteinase Inhibitors
  • Amyloid Precursor Protein Secretases
  • Cathepsin B
  • Aspartic Acid Endopeptidases
  • Bace1 protein, mouse
  • Leucine
  • aloxistatin