Alzheimer's disease (AD) is a progressive disorder resulting in severe dementia which currently cannot be stopped. Abnormal accumulation of extracellular brain beta-amyloid is thought to cause AD and compounds that block beta-amyloid production are predicted to be effective for treating AD. Beta-amyloid is generated by cleavage from a larger protein, amyloid precursor protein (APR), by proteases called beta-secretases and gamma-secretases. Compounds that inhibit either of these secretases reduce beta-amyloid production and, therefore, are thought to be potentially effective AD drugs. Brain neurons secrete beta-amyloid by regulated and constitutive secretory pathways. We find that neurons secrete the vast majority of beta-amyloid via the regulated secretory pathway and that the cysteine proteases, cathepsin B and cathepsin L, are the beta- secretases of that pathway. Consequently, we find cathepsin B and cathepsin L are responsible for producing the majority of secreted beta-amyloid. Evidence for our finding is based on a wide range of experimental data. We purified beta-secretase activity from isolated regulated secretory vesicles and determined by microsequencing that the beta-secretase activity is due cathepsin B and cathepsin L. We found by immunoelectron microscopy that cathepsin B and cathepsin L are co-localized with APP and beta-amyloid in isolated secretory vesicles. We observed that cathepsin B and/or cathepsin L inhibitors block endogenous beta-secretase activity in isolated regulated secretory vesicles and stop the continued natural production of beta-amyloid that occurs after isolation of regulated secretory vesicles. Moreover, we found that cathepsin B and cathepsin L cleave with excellent efficiency the beta-secretase site found in wild-type APP, the APP form found in 99% of AD patients. Further, we found that a cathepsin B specific inhibitor, CA-074, dramatically reduces regulated secretion of Ab, but not its constitutive secretion, by neuronal chromaffin cells. Significantly, we show in this application that the cysteine protease inhibitor, e64d, reduces in vivo total brain and synaptosomal beta-amyloid in the guinea pig AD animal model. In this project, we will evaluate peptidomimetics that specifically inhibit cathepsin B or cathepsin L and which we find also inhibit beta-secretase activity in regulated secretory vesicles. Specifically, we will assess the affects of these peptidomimetics on beta-amyloid secretion in primary neuronal cultures and in vivo.
|Hook, Vivian; Funkelstein, Lydiane; Wegrzyn, Jill et al. (2012) Cysteine Cathepsins in the secretory vesicle produce active peptides: Cathepsin L generates peptide neurotransmitters and cathepsin B produces beta-amyloid of Alzheimer's disease. Biochim Biophys Acta 1824:89-104|
|Kindy, Mark S; Yu, Jin; Zhu, Hong et al. (2012) Deletion of the cathepsin B gene improves memory deficits in a transgenic ALZHeimer's disease mouse model expressing A?PP containing the wild-type ?-secretase site sequence. J Alzheimers Dis 29:827-40|
|Hook, Gregory; Hook, Vivian; Kindy, Mark (2011) The cysteine protease inhibitor, E64d, reduces brain amyloid-? and improves memory deficits in Alzheimer's disease animal models by inhibiting cathepsin B, but not BACE1, ?-secretase activity. J Alzheimers Dis 26:387-408|
|Hook, Vivian; Hook, Gregory; Kindy, Mark (2010) Pharmacogenetic features of cathepsin B inhibitors that improve memory deficit and reduce beta-amyloid related to Alzheimer's disease. Biol Chem 391:861-72|
|Hook, Vivian Y H; Kindy, Mark; Reinheckel, Thomas et al. (2009) Genetic cathepsin B deficiency reduces beta-amyloid in transgenic mice expressing human wild-type amyloid precursor protein. Biochem Biophys Res Commun 386:284-8|