We carried out the most powerful genomewide study of Alzheimer disease (AD), involving over 16,000 people from eight countries, and identified two new genes that increase a person’s risk of developing the disease. These are CLU or clusterin and PICALM.

We compared over half a million differences in the DNA of each of 4,000 people with AD with 8,000 people without the disease. In addition to the APOE gene—a known risk factor—CLU and PICALM showed overwhelming evidence for a relationship with AD. These findings were replicated in a further sample of over 2,000 Alzheimer’s sufferers and 2,000 controls.

The findings are significant and conclusive.

We also found strong evidence that other genes play a role in disease risk. Putting our data together with the results of Philippe Amouyel’s study, a third risk gene was confirmed; CR1, complement receptor 1 gene.

This combination of discoveries forms an important breakthrough in the current impetus to discover the causes of AD.

Three of the risk genes, APOE, CLU, and CR1, have roles in protecting the brain from damage....  Read more

We carried out the most powerful genomewide study of Alzheimer disease (AD), involving over 16,000 people from eight countries, and identified two new genes that increase a person’s risk of developing the disease. These are CLU or clusterin and PICALM.

We compared over half a million differences in the DNA of each of 4,000 people with AD with 8,000 people without the disease. In addition to the APOE gene—a known risk factor—CLU and PICALM showed overwhelming evidence for a relationship with AD. These findings were replicated in a further sample of over 2,000 Alzheimer’s sufferers and 2,000 controls.

The findings are significant and conclusive.

We also found strong evidence that other genes play a role in disease risk. Putting our data together with the results of Philippe Amouyel’s study, a third risk gene was confirmed; CR1, complement receptor 1 gene.

This combination of discoveries forms an important breakthrough in the current impetus to discover the causes of AD.

Three of the risk genes, APOE, CLU, and CR1, have roles in protecting the brain from damage. Perhaps the changes we see in these genes remove this protection or may even turn them into killers.

Our results may highlight new targets for treatments. For example, clusterin has a role in dampening down inflammation in the brain. Up until now increased inflammation seen in the brains of Alzheimer’s sufferers had been viewed as a secondary effect of disease. Our results suggest the possibility that inflammation may be primary to disease development.

If we were able to remove the detrimental effects of these genes through treatments, we could reduce the proportion of people developing Alzheimer’s by 20 percent. In the UK alone this would prevent just under 100,000 people developing the disease. So the significance of these results is truly meaningful.

View all comments by Julie Williams

Two of the most important implications of this study are that it establishes that this approach works and suggests that larger studies are likely to identify other genes conferring similar risks to AD. For this reason we are planning an even larger study of 60,000 participants. This should allow us to identify other genes of relevance to AD.

We are also undertaking more complex analyses of our current data—using new analytic approaches we have developed in Cardiff. These allow many genes to be analyzed at once and identify patterns in the data that implicate specific biological processes.

The identification of multiple genetic risk factors should allow us to triangulate down onto specific biological processes. These might be related to β amyloid deposition, but they might also identify completely new disease mechanisms.

The key thing about genetics is that it allows us to distinguish events of primary importance from those that are occurring as a consequence of the disease—downstream if you like—and these primary events are likely to be the best targets for new...  Read more

Two of the most important implications of this study are that it establishes that this approach works and suggests that larger studies are likely to identify other genes conferring similar risks to AD. For this reason we are planning an even larger study of 60,000 participants. This should allow us to identify other genes of relevance to AD.

We are also undertaking more complex analyses of our current data—using new analytic approaches we have developed in Cardiff. These allow many genes to be analyzed at once and identify patterns in the data that implicate specific biological processes.

The identification of multiple genetic risk factors should allow us to triangulate down onto specific biological processes. These might be related to β amyloid deposition, but they might also identify completely new disease mechanisms.

The key thing about genetics is that it allows us to distinguish events of primary importance from those that are occurring as a consequence of the disease—downstream if you like—and these primary events are likely to be the best targets for new treatments.

It’s also possible that in the future we might be able to use the results of genetic tests as part of a battery of indicators to identify those who might benefit from early intervention with new therapies. I should stress that the current genes on their own are not strong predictors of risk and are not suitable for risk testing.

This study is one of the first major successes for the new MRC Centre in Cardiff which has been established this year to undertake genetic work in neurodegenerative diseases like AD and PD—as well as psychiatric disorders like schizophrenia and manic depression. This centre is allowing us to look not only at individual diseases, but also the overlap between them; we are currently looking to see whether the AD genes we have identified are involved in other forms of dementia such as Parkinson’s.

View all comments by Michael Owen

By Jean-Charles Lambert and Philippe Amouyel

We conducted a GWAS in 2,032 AD cases and 5,328 controls from the prospective population-based 3C study. We next replicated our most interesting hits in independent collections from Belgium, Finland, Italy and Spain totalling 3,978 AD cases and 3,297 controls. This replication work, the largest performed so far, strongly supports the association of CLU and CR1 with the risk of developing AD

It is with enthusiasm that we learned the validation of our observations in the Harold et al. GWA study. The combination of these genetic results and the available pathophysiological data seem to strongly support the involvement of these genes in AD.

However, it is important to keep in mind that in view of the large number of analyses performed, high-throughput approaches involve finding a balance between the risk of observing significant results by chance and the risk of rejecting biologically valid hypotheses on purely statistical grounds. Although no solution has been found to improve this dilemma, several approaches can be developed....  Read more

By Jean-Charles Lambert and Philippe Amouyel

We conducted a GWAS in 2,032 AD cases and 5,328 controls from the prospective population-based 3C study. We next replicated our most interesting hits in independent collections from Belgium, Finland, Italy and Spain totalling 3,978 AD cases and 3,297 controls. This replication work, the largest performed so far, strongly supports the association of CLU and CR1 with the risk of developing AD

It is with enthusiasm that we learned the validation of our observations in the Harold et al. GWA study. The combination of these genetic results and the available pathophysiological data seem to strongly support the involvement of these genes in AD.

However, it is important to keep in mind that in view of the large number of analyses performed, high-throughput approaches involve finding a balance between the risk of observing significant results by chance and the risk of rejecting biologically valid hypotheses on purely statistical grounds. Although no solution has been found to improve this dilemma, several approaches can be developed. It is possible to increase the statistical power associated with GWASs by performing meta-analyses. Other complementary approaches could also consist in better analyzing SNPs nominally associated with the risk of developing AD (p<0.05) using complex statistical and bioinformatics approaches. It is thus highly probable that in the next few years, we will be able to characterize new genetic determinants of AD and to approach in this way the complex pathophysiological processes involved in this disease.

View all comments by Jean-Charles Lambert

Marcello and colleagues demonstrate that synaptic activity regulates the levels of ADAM10 on the plasma membrane. Long-term potentiation (LTP) induces ADAM10 internalization by clathrin-mediated endocytosis (CME), whereas LTD induces its insertion into the plasma membrane. There is a long history of literature linking CME and Aβ; however, those studies generally revolve around APP internalization and Aβ generation. Work from our group and others shows that synaptic activity causes CME of APP, which increases Aβ production in endosomes. The data in the Marcello paper look at this from a different angle. Here, synaptic activity increases ADAM10 internalization, which decreases its ability to cleave APP; in theory, this would then increase Aβ generation. So taken together, this suggests that synaptic activity and CME may promote Aβ generation by two parallel pathways: 1) increasing amyloidogenic processing of APP within endosomes, and 2) decreasing non-amyloidogenic processing of APP at the plasma membrane.

As with any good study, lots of questions remain. Is ADAM10...  Read more

Marcello and colleagues demonstrate that synaptic activity regulates the levels of ADAM10 on the plasma membrane. Long-term potentiation (LTP) induces ADAM10 internalization by clathrin-mediated endocytosis (CME), whereas LTD induces its insertion into the plasma membrane. There is a long history of literature linking CME and Aβ; however, those studies generally revolve around APP internalization and Aβ generation. Work from our group and others shows that synaptic activity causes CME of APP, which increases Aβ production in endosomes. The data in the Marcello paper look at this from a different angle. Here, synaptic activity increases ADAM10 internalization, which decreases its ability to cleave APP; in theory, this would then increase Aβ generation. So taken together, this suggests that synaptic activity and CME may promote Aβ generation by two parallel pathways: 1) increasing amyloidogenic processing of APP within endosomes, and 2) decreasing non-amyloidogenic processing of APP at the plasma membrane.

As with any good study, lots of questions remain. Is ADAM10 internalized into separate endosomes than APP? Or can ADAM10 and APP still be within the same endosome but maybe the low pH prevents further cleavage? How is ADAM10 trafficked or segregated after CME? The authors demonstrate that activity modulates APP cleavage by ADAM10, which is consistent with a reduction in Aβ, but one thing lacking in this paper is a direct measurement that Aβ generation is changing as a consequence of the altered ADAM10 subcellular localization.

View all comments by John Cirrito

A disintegrin and metalloproteinase 10 (ADAM10) is apparently one of the most critical membrane-associated proteases in the central nervous system (CNS). Its prominent role in the embryonic and adult CNS has been revealed by a number of studies. Next to APP, an increasing number of transmembrane proteins, including Notch receptors and ligands, are subject to ADAM10-mediated shedding. These shedding events are of critical importance to modulate postsynaptic function and synaptic plasticity.

Based on their previous work, Monica Di Luca´s group convincingly addressed the post-transcriptional regulation of ADAM10 in neurons. Both its transport to the postsynaptic membrane and its removal are central events to regulate synaptic functions, morphology, and the processing of important substrates, including APP. In the current study, the authors focus on the endocytosis of ADAM10 from the postsynaptic membrane. Using mainly coimmunoprecipitation experiments, they showed that, like other surface molecules, ADAM10 endocytosis depends on binding to the clathrin adaptor AP2. This binding...  Read more

A disintegrin and metalloproteinase 10 (ADAM10) is apparently one of the most critical membrane-associated proteases in the central nervous system (CNS). Its prominent role in the embryonic and adult CNS has been revealed by a number of studies. Next to APP, an increasing number of transmembrane proteins, including Notch receptors and ligands, are subject to ADAM10-mediated shedding. These shedding events are of critical importance to modulate postsynaptic function and synaptic plasticity.

Based on their previous work, Monica Di Luca´s group convincingly addressed the post-transcriptional regulation of ADAM10 in neurons. Both its transport to the postsynaptic membrane and its removal are central events to regulate synaptic functions, morphology, and the processing of important substrates, including APP. In the current study, the authors focus on the endocytosis of ADAM10 from the postsynaptic membrane. Using mainly coimmunoprecipitation experiments, they showed that, like other surface molecules, ADAM10 endocytosis depends on binding to the clathrin adaptor AP2. This binding seems to be stronger in samples from AD patients. It was shown that abolishing this binding using mutants or pharmacological approaches also reduced ADAM10 endocytosis. Interestingly, the authors found that long-term potentiation induced this process, whereas long-term depression had an opposite effect by stimulating interaction of ADAM10 and SAP97, thereby promoting ADAM10 delivery to the plasma membrane. In a last set of experiments, they showed that this dynamic, and apparently ADAM10-dependent regulation, also led to a differential processing of APP at the α-secretase site. ADAM10 localization at the synaptic membrane more or less determines whether α-secretase processing occurs.

What are the consequences for AD? The findings certainly increase our basic understanding of how APP processing, synaptic remodeling, and cellular localization of ADAM10 are linked. I do not feel that immediate new therapeutic targets are apparent, since the factors involved affect a number of other proteins as well; for example, the fine-tuned localization of ADAM10 is necessary for the degree of shedding of most likely more than 10 other synaptic membrane proteins. Also, AP2 mediates endocytosis of a huge number of surface proteins. On the other hand, additional intracellular and extracellular factors are needed to control the activity of ADAM10. We and others showed recently, for example, that the integration of ADAM10 in the tetraspanin web is instrumental for forward trafficking of the protease. It is likely that additional factors directly contribute to the cellular localization of ADAM10.

As discussed by the authors, the Aβ levels in vivo are in part dependent on the activity of ADAM10. This study additionally provides evidence that the degree of neuronal and synaptic activity alters the function (and localization) of ADAM10 and the degree of Aβ production. Based on their initial observations that in AD brains, the endocytic route of ADAM10 is favored, this would directly explain both the increased Aβ production and the problems in synaptogenes as reported in AD.

View all comments by Paul Saftig

I read this paper by Marcello et al. with much interest, and I was impressed both by the number of data and the coherence of the hypothesis. It explains the role played by neuronal activity in Aβ secretion. It also sheds new light on the connection between endocytosis and Aβ.

In addition, it opens new research perspectives: The alteration of ADAM10/AP2 association in AD is currently not explained and could be related to changes in the cell membrane itself. We have, in this respect, shown that increases in membrane cholesterol favor endocytosis and production of Aβ (see Marquer et al., 2011; Cossec et al., 2010).

I'd add a word of caution on the neuropathology. Only six cases were examined at Braak stage IV—these were apparently the same cases the authors studied before (see Marcello et al., 2012). Braak stage IV pathology is common in asymptomatic aged persons. The diagnostic probability of Alzheimer's disease is only ranked as intermediate in the current diagnostic criteria (Hyman et al., 2012; Montine et al., 2012). Such cases are, by definition, free from tau pathology...  Read more

I read this paper by Marcello et al. with much interest, and I was impressed both by the number of data and the coherence of the hypothesis. It explains the role played by neuronal activity in Aβ secretion. It also sheds new light on the connection between endocytosis and Aβ.

In addition, it opens new research perspectives: The alteration of ADAM10/AP2 association in AD is currently not explained and could be related to changes in the cell membrane itself. We have, in this respect, shown that increases in membrane cholesterol favor endocytosis and production of Aβ (see Marquer et al., 2011; Cossec et al., 2010).

I'd add a word of caution on the neuropathology. Only six cases were examined at Braak stage IV—these were apparently the same cases the authors studied before (see Marcello et al., 2012). Braak stage IV pathology is common in asymptomatic aged persons. The diagnostic probability of Alzheimer's disease is only ranked as intermediate in the current diagnostic criteria (Hyman et al., 2012; Montine et al., 2012). Such cases are, by definition, free from tau pathology in the neocortex. Usually, Aβ deposits are, however, already present. This could be determined in the studied cases, for instance, by identifying the stage of amyloid pathology (see Thal et al., 2002). It would be interesting to compare the association of ADAM10/AP2 in the hippocampus (with tangle pathology) and in the frontal cortex (devoid of tangle pathology but possibly with Aβ deposition). Tau accumulation, which may affect synapses, could play a central role in the alteration of ADAM10/AP2 association, as it probably does for the clathrin adaptor PICALM, which we found to colocalize with tangles (Ando et al., 2013). New studies with more advanced cases would strengthen the human data.

References:
Marquer C, Devauges V, Cossec JC, Liot G, Lécart S, Saudou F, Duyckaerts C, Lévêque-Fort S, Potier MC. Local cholesterol increase triggers amyloid precursor protein-Bace1 clustering in lipid rafts and rapid endocytosis. FASEB J. 2011 Apr;25(4):1295-305. Abstract

Cossec JC, Simon A, Marquer C, Moldrich RX, Leterrier C, Rossier J, Duyckaerts C, Lenkei Z, Potier MC. Clathrin-dependent APP endocytosis and Aβ secretion are highly sensitive to the level of plasma membrane cholesterol. Biochim Biophys Acta. 2010 Aug;1801(8):846-52. Abstract

Marcello E, Epis R, Saraceno C, Gardoni F, Borroni B, Cattabeni F, Padovani A, Di Luca M. SAP97-mediated local trafficking is altered in Alzheimer disease patients' hippocampus. Neurobiol Aging. 2012 Feb;33(2):422.e1-10. Abstract

Hyman BT, Phelps CH, Beach TG, Bigio EH, Cairns NJ, Carrillo MC, Dickson DW, Duyckaerts C, Frosch MP, Masliah E, Mirra SS, Nelson PT, Schneider JA, Thal DR, Thies B, Trojanowski JQ, Vinters HV, Montine TJ. National Institute on Aging-Alzheimer's Association guidelines for the neuropathologic assessment of Alzheimer's disease. Alzheimers Dement. 2012 Jan;8(1):1-13. Abstract

Montine TJ, Phelps CH, Beach TG, Bigio EH, Cairns NJ, Dickson DW, Duyckaerts C, Frosch MP, Masliah E, Mirra SS, Nelson PT, Schneider JA, Thal DR, Trojanowski JQ, Vinters HV, Hyman BT, National Institute on Aging, Alzheimer’s Association. National Institute on Aging-Alzheimer's Association guidelines for the neuropathologic assessment of Alzheimer's disease: a practical approach. Acta Neuropathol. 2012 Jan;123(1):1-11. Abstract

Thal DR, Rüb U, Orantes M, Braak H. Phases of A β-deposition in the human brain and its relevance for the development of AD. Neurology. 2002 Jun 25;58(12):1791-800. Abstract

Ando K, Brion JP, Stygelbout V, Suain V, Authelet M, Dedecker R, Chanut A, Lacor P, Lavaur J, Sazdovitch V, Rogaeva E, Potier MC, Duyckaerts C. Clathrin adaptor CALM/PICALM is associated with neurofibrillary tangles and is cleaved in Alzheimer's brains. Acta Neuropathol. 2013 Apr 16. Abstract

View all comments by Charles Duyckaerts