Hemming ML, Elias JE, Gygi SP, Selkoe DJ.
Identification of beta-secretase (BACE1) substrates using quantitative proteomics.
PLoS One. 2009;4(12):e8477.
PubMed.
This is very interesting work. The identification of over 60 BACE1 substrates using an unbiased and quantitative proteomic method sheds light on the role of β-secretase in conditions where BACE1 levels are increased. Levels of BACE1 are normally very low in peripheral tissues as well as in the epithelial cell lines used in this study. Moreover, cerebral BACE1 levels decrease postnatally (Willem et al. 2006). Altogether these findings indicate that in normal conditions, β-secretase activity is low. Instead, several groups, including ours, have reported that β-secretase activity is increased in stress conditions (cerebral ischemia, traumatic brain injury, energy deprivation, hypoxia, apoptosis). Thus, this study is particularly interesting because it demonstrates that increased β-secretase activity results in alteration of normal cellular functions, e.g., the missorting of lysosomal hydrolases and increased cleavage of cell adhesion molecules, which may be detrimental for the cell. Given that BACE1 is elevated in AD brains, increased cleavage of additional substrates may contribute to AD pathology. Thus, β-secretase inhibition is expected to be beneficial in conditions associated with increased levels of BACE1.
References:
Willem M, Garratt AN, Novak B, Citron M, Kaufmann S, Rittger A, Destrooper B, Saftig P, Birchmeier C, Haass C.
Control of peripheral nerve myelination by the beta-secretase BACE1.
Science. 2006 Oct 27;314(5799):664-6.
PubMed.
The partial inhibition of both secretases is quite interesting, and the results could easily be viewed as implicating βCTF as well as Aβ in pathology. In addition to the mechanism Chow et al. propose of additive/synergistic effects on Aβ lowering, our data would suggest that an additional explanation for the combined inhibitor effects on memory deficits are a reversal of separate adverse effects of βCTF and Aβ on cognitive performance. A third copy of APP is known to be responsible for certain of the memory and learning deficits in the Ts65Dn mouse model of Down syndrome, which is associated with increased β-cleavage of APP but no measurable Aβ elevation, at least at the whole brain level (Choi et al. 2009; Salehi et al. 2006). In light of our recent evidence linking increased βCTF to AD-related endosome defects in Down syndrome and to FAD due to APP duplication, it is reasonable to suspect that βCTF may play a role in the rescue of cognitive deficits by BACE inhibition. This would be expected to add to effects of lowering Aβ levels in the APPswe/PS1DE9 model used in this study, which exhibits marked amyloid deposition/Aβ elevation in contrast to the Ts65Dn mouse.
Based on this framework, the caveats that our work suggested regarding the possible buildup of potentially toxic βCTF after using γ inhibitor alone would be offset by simultaneously blocking CTF buildup with a BACE inhibitor.
References:
Choi JH, Berger JD, Mazzella MJ, Morales-Corraliza J, Cataldo AM, Nixon RA, Ginsberg SD, Levy E, Mathews PM.
Age-dependent dysregulation of brain amyloid precursor protein in the Ts65Dn Down syndrome mouse model.
J Neurochem. 2009 Sep;110(6):1818-27. Epub 2009 Jul 10
PubMed.
Salehi A, Delcroix JD, Belichenko PV, Zhan K, Wu C, Valletta JS, Takimoto-Kimura R, Kleschevnikov AM, Sambamurti K, Chung PP, Xia W, Villar A, Campbell WA, Kulnane LS, Nixon RA, Lamb BT, Epstein CJ, Stokin GB, Goldstein LS, Mobley WC.
Increased App expression in a mouse model of Down's syndrome disrupts NGF transport and causes cholinergic neuron degeneration.
Neuron. 2006 Jul 6;51(1):29-42.
PubMed.
This is an interesting approach and could hold promise in the clinic. However, the paper describes a genetic knockdown approach. The progression of a small molecule combination therapy to the clinic would be more challenging.
Issues that would need to be addressed include assessing potential drug-drug interactions in humans. In the meantime, both tool γ-secretase and BACE inhibitors are available to test this theory in APP transgenic mice. These studies would need to be done to assess the potential of small molecule combinations and to assess any potential toxicity issues.
The β-secretase (BACE1) protease is one of the enzymes that cleaves the amyloid precursor protein (APP) and leads to the production of Aβ peptides. In Alzheimer disease, elevated expression of this enzyme is thought to underlie increased Aβ production and lead to pathological effects. As such, BACE1 is an attractive therapeutic target. However, as pointed out by the authors of this paper, the normal function of BACE1 is still not well understood. Although it was known that APP is not the only target of BACE1, few targets had been identified to date. In addition, in BACE1 knockout mice, various problems have been reported, suggesting that BACE1 has important roles in the nervous system. In this paper, the Selkoe laboratory performed a quantitative proteomic analysis to identify BACE1 substrates using cell culture systems in which they overexpress the protease. They identified 68 putative substrates and validated several of these. In addition, they identified various other proteins that are not membrane bound and are likely elevated by indirect effects. The putative substrates were identified in two cell lines that express very low levels of BACE1, and the results identified many differences between cells. This study illustrates the complexities associated with targeting the expression of BACE1 as a therapeutic strategy in Alzheimer disease and provides important fundamental information. It seems that the challenges moving forward will be to identify relevant substrates in neurons that express BACE1 and to determine the relative importance of various BACE1 substrates.
The new study by Chow and colleagues describes evidence that the combination of partial reductions in β-secretase and γ-secretase are effective in lowering Aβ production and deposition and preventing cognitive deficits in mice. This happens without the deleterious effects of complete knockout of either protease alone, something that many of us in the field have suspected for some time, but which had not yet been demonstrated.
The strategy for dual lowering of the two secretases was strictly genetic, not pharmacological: mice that are heterozygous knockouts of BACE1 and Aph1a were crossed into transgenic mice overexpressing AD mutant APP and Presenilin.
Nevertheless, the results provide an important proof of principle. From a practical perspective though, it will be difficult to test this combination approach in humans, as each secretase inhibitor must first be tested individually and shown to be efficacious before combination regimens can be tried.
Comments
Tufts University School of Medicine
This is very interesting work. The identification of over 60 BACE1 substrates using an unbiased and quantitative proteomic method sheds light on the role of β-secretase in conditions where BACE1 levels are increased. Levels of BACE1 are normally very low in peripheral tissues as well as in the epithelial cell lines used in this study. Moreover, cerebral BACE1 levels decrease postnatally (Willem et al. 2006). Altogether these findings indicate that in normal conditions, β-secretase activity is low. Instead, several groups, including ours, have reported that β-secretase activity is increased in stress conditions (cerebral ischemia, traumatic brain injury, energy deprivation, hypoxia, apoptosis). Thus, this study is particularly interesting because it demonstrates that increased β-secretase activity results in alteration of normal cellular functions, e.g., the missorting of lysosomal hydrolases and increased cleavage of cell adhesion molecules, which may be detrimental for the cell. Given that BACE1 is elevated in AD brains, increased cleavage of additional substrates may contribute to AD pathology. Thus, β-secretase inhibition is expected to be beneficial in conditions associated with increased levels of BACE1.
References:
Willem M, Garratt AN, Novak B, Citron M, Kaufmann S, Rittger A, Destrooper B, Saftig P, Birchmeier C, Haass C. Control of peripheral nerve myelination by the beta-secretase BACE1. Science. 2006 Oct 27;314(5799):664-6. PubMed.
New York University School of Medicine/Nathan Kline Institute
The partial inhibition of both secretases is quite interesting, and the results could easily be viewed as implicating βCTF as well as Aβ in pathology. In addition to the mechanism Chow et al. propose of additive/synergistic effects on Aβ lowering, our data would suggest that an additional explanation for the combined inhibitor effects on memory deficits are a reversal of separate adverse effects of βCTF and Aβ on cognitive performance. A third copy of APP is known to be responsible for certain of the memory and learning deficits in the Ts65Dn mouse model of Down syndrome, which is associated with increased β-cleavage of APP but no measurable Aβ elevation, at least at the whole brain level (Choi et al. 2009; Salehi et al. 2006). In light of our recent evidence linking increased βCTF to AD-related endosome defects in Down syndrome and to FAD due to APP duplication, it is reasonable to suspect that βCTF may play a role in the rescue of cognitive deficits by BACE inhibition. This would be expected to add to effects of lowering Aβ levels in the APPswe/PS1DE9 model used in this study, which exhibits marked amyloid deposition/Aβ elevation in contrast to the Ts65Dn mouse.
Based on this framework, the caveats that our work suggested regarding the possible buildup of potentially toxic βCTF after using γ inhibitor alone would be offset by simultaneously blocking CTF buildup with a BACE inhibitor.
References:
Choi JH, Berger JD, Mazzella MJ, Morales-Corraliza J, Cataldo AM, Nixon RA, Ginsberg SD, Levy E, Mathews PM. Age-dependent dysregulation of brain amyloid precursor protein in the Ts65Dn Down syndrome mouse model. J Neurochem. 2009 Sep;110(6):1818-27. Epub 2009 Jul 10 PubMed.
Salehi A, Delcroix JD, Belichenko PV, Zhan K, Wu C, Valletta JS, Takimoto-Kimura R, Kleschevnikov AM, Sambamurti K, Chung PP, Xia W, Villar A, Campbell WA, Kulnane LS, Nixon RA, Lamb BT, Epstein CJ, Stokin GB, Goldstein LS, Mobley WC. Increased App expression in a mouse model of Down's syndrome disrupts NGF transport and causes cholinergic neuron degeneration. Neuron. 2006 Jul 6;51(1):29-42. PubMed.
View all comments by Ralph NixonGlaxoSmithKline R&D
This is an interesting approach and could hold promise in the clinic. However, the paper describes a genetic knockdown approach. The progression of a small molecule combination therapy to the clinic would be more challenging.
Issues that would need to be addressed include assessing potential drug-drug interactions in humans. In the meantime, both tool γ-secretase and BACE inhibitors are available to test this theory in APP transgenic mice. These studies would need to be done to assess the potential of small molecule combinations and to assess any potential toxicity issues.
The Scripps Research Institute
The β-secretase (BACE1) protease is one of the enzymes that cleaves the amyloid precursor protein (APP) and leads to the production of Aβ peptides. In Alzheimer disease, elevated expression of this enzyme is thought to underlie increased Aβ production and lead to pathological effects. As such, BACE1 is an attractive therapeutic target. However, as pointed out by the authors of this paper, the normal function of BACE1 is still not well understood. Although it was known that APP is not the only target of BACE1, few targets had been identified to date. In addition, in BACE1 knockout mice, various problems have been reported, suggesting that BACE1 has important roles in the nervous system. In this paper, the Selkoe laboratory performed a quantitative proteomic analysis to identify BACE1 substrates using cell culture systems in which they overexpress the protease. They identified 68 putative substrates and validated several of these. In addition, they identified various other proteins that are not membrane bound and are likely elevated by indirect effects. The putative substrates were identified in two cell lines that express very low levels of BACE1, and the results identified many differences between cells. This study illustrates the complexities associated with targeting the expression of BACE1 as a therapeutic strategy in Alzheimer disease and provides important fundamental information. It seems that the challenges moving forward will be to identify relevant substrates in neurons that express BACE1 and to determine the relative importance of various BACE1 substrates.
University of Kansas
The new study by Chow and colleagues describes evidence that the combination of partial reductions in β-secretase and γ-secretase are effective in lowering Aβ production and deposition and preventing cognitive deficits in mice. This happens without the deleterious effects of complete knockout of either protease alone, something that many of us in the field have suspected for some time, but which had not yet been demonstrated.
The strategy for dual lowering of the two secretases was strictly genetic, not pharmacological: mice that are heterozygous knockouts of BACE1 and Aph1a were crossed into transgenic mice overexpressing AD mutant APP and Presenilin.
Nevertheless, the results provide an important proof of principle. From a practical perspective though, it will be difficult to test this combination approach in humans, as each secretase inhibitor must first be tested individually and shown to be efficacious before combination regimens can be tried.
View all comments by Michael WolfeMake a Comment
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