Two ApoE Mutations Decrease Risk for Alzheimer's Disease

ApoE just got a little more interesting—if that’s possible. While scientists may debate how this apolipoprotein contributes to Alzheimer’s disease pathology, nobody disputes that one of its isoforms, ApoE4, increases a person’s risk for developing the disease. Now, a twist. In the May 31 JAMA Neurology, researchers led by Yann Le Guen, Stanford University, California, report that two rare variants decrease risk for the disease. Both lie in the C-terminal, lipid-binding domain of the protein—and wouldn’t you know, one co-inherits with, and seems to neutralize, the E4 isoform. ApoE4 carriers who also inherit this R251G variant are at no more risk for AD than are ApoE2/3 carriers. The finding might prompt scientists to look for new ways to temper ApoE4 and protect against AD.

  • Large case-control study identified two rare ApoE variants.
  • V236E, which was known, co-inherits with ApoE3.
  • A novel variant, R251G, co-inherits with ApoE4, neutralizing it.

“While the allele frequencies of these variants are less than 0.1 percent, … it definitely appears that these coding changes are playing an important role in decreasing AD risk,” David Holtzman, Washington University, St. Louis, wrote to Alzforum. Guojun Bu, Mayo Clinic, Jacksonville, Florida, agreed. “… [T]he genetic data are in general convincing, despite the typical difficulties in genetic association studies on rare variants,” he wrote.

Protective Variants. The V236E and R251G variants in APOE reduced a person’s chances of getting AD to about that of an APOE2/3 carrier. Risks shown are relative to ApoE3/3 carriers whose odds ratios are 1. [Courtesy of Le Guen et al., JAMA Neurol 2022]

Beyond ApoE2, ApoE3, and ApoE4, the three major isoforms, gnomAD lists about 300 others, but most of them are rare and their relevance to AD unknown. The R136S Christchurch mutation in the N-terminal appears to be protective, but is based on only one person who also carried a pathogenic AD mutation in her presenilin 1 gene and whose disease onset was delayed by decades (see Nov 2019 news on Arboleda-Velasquez et al., 2019). Bu and colleagues at the Mayo Clinic, Jacksonville, discovered the V236E “Jac” mutation, which also seemed protective, based on data from a small cohort of 9,000 people (Oct 2021 news). Le Guen and colleagues, including senior author Michael Greicius at Stanford, wanted to look for variants in a much larger dataset.

The authors first analyzed whole-genome and whole-exome data from the Alzheimer’s Disease Sequencing Project, looking for associations between APOE variants and AD risk. Among 11,868 cases and 11, 934 controls of European ancestry, people who carried the V236E or R251G variants were four- and fivefold less likely to have AD. Next, the authors expanded the analysis in two more stages to almost 3.5 million volunteers across eight other cohorts. They were the UK Biobank, the European Alzheimer’s Disease DNA Biobank; the European Alzheimer’s Disease Initiative, the Genetic and Environmental Risk in Alzheimer’s Disease Consortium, the Norwegian Dementia Genetics Network, Genome Research at Fundació Alzheimer Center Barcelona/Dementia Genetics Spanish Consortium, the Copenhagen City Heart Study, and the Copenhagen General Population Study. All told, 544,384 people passed inclusion criteria, and their risk association was assessed.

This larger analysis confirmed that the two variants are protective. The odds ratio of having AD were 0.44 and O.37, for R251G and V236E, respectively, indicating two- and threefold lower risk. When stratified by ApoE2/3/4 genotype, the results were similar. In short, compared to ApoE3/3 carriers, risk for AD among ApoE3/3 V236E carriers was similar to that for ApoE2/3 carriers. Remarkably, the same held for R251G. That variant reduced risk for AD among ApoE3/4 carriers to that of ApoE2/3 carriers (see image above). This is the first variant ever found to mitigate the effect of ApoE4. One copy of APOE4 triples risk for AD and two copies increases it about 15-fold.

How these variants protect is unclear. Previously, Bu had reported that V236E reduces the tendency of the protein to aggregate. Whether the same goes for R251G remains to be seen, but evidence suggests the amino acid forms a salt bridge with glutamine 98, which would be abolished by swapping arginine with glycine (Chen et al., 2011). There are other salt bridges between the C-terminal and N-terminal ends of the protein that might also be disrupted. “Depending on the actual structure of ApoE in its native state, how the N- and C-terminals interact to influence ApoE structure will be critical to resolve, because it is likely that such interactions are key in determining how ApoE alters AD risk,” wrote Holtzman.

Alternatively, R251G might enhance ApoE binding with lipids, making it more like ApoE2/3. Scientists have found it challenging to study the structure of native proteins in their lipidated state, including ApoE. “New technologies such as cryoEM as well as small molecule FRET may be key in unlocking the high-resolution structure of lipidated APOE to get at some of these key issues,” Holtzman wrote.

Protective missense variations in APOE are very rare, but the magnitude of their observed effects is large and thus will further our understanding of the biological pathways through which APOE profoundly modifies the risk of AD,” wrote Gil Rabinovici and Dena Dubal, University of California, San Francisco, in a JAMA Neurology editorial. “A better understanding of these pathways will likely identify novel therapeutic targets that can delay or possibly prevent disease even in individuals at high genetic risk owing to APOE ε4 or autosomal dominant pathogenic gene variants, or even individuals with late-onset AD,” they wrote.

One caveat is that the study was limited to people of European ancestry. “It would also be interesting to conduct similar analyses in non-European populations given that the local ancestry at the APOE locus, and its surrounding regions, can also influence the AD risk effect of APOE-ε4,” noted Nancy Ip, Honk Kong University of Science and Technology. She has reported two genes, PVRL2 and APOC1, that associate with AD and are inherited as haplotypes with APOE and modulate its risk (Aug 2019 news on Zhou et al., 2019).  

Le Guen found that the results held when limiting the analysis to people with at most 55 per cent non-European ancestry. The authors note that the variants have been found in African American’s and Latinos based on gnomAD data, but they had insufficient number in their datasets to analyze the effects of these rare variants in non-Europeans.—Tom Fagan

Comments

  1. In this manuscript, Le Guen and colleagues report that two missense variants in APOE were associated with a twofold to threefold decreased AD risk: APOE ε4 (R251G) (odds ratio, 0.44; 95 percent CI, 0.33-0.59; P = 4.7 × 10−8) and APOE ε3 (V236E) (odds ratio, 0.37; 95 percent CI, 0.25-0.56; P = 1.9 × 10−6). While the allele frequencies of these variants are less than 0.1 percent, given the number of cohorts and number of individuals examined, it definitely appears that these coding changes are playing an important role in decreasing AD risk. The R251G mutation appears to be the first variant that protects against the increased risk caused by the APOE4 allele. The V236E mutation has been reported previously by the lab of Guojun Bu (the so-called Jacksonville variant).

    A key question is why do these variants, or, for that matter, other variants in APOE, lead to altered AD risk? While APOE has pleiotropic effects biologically, several of which may be relevant to influencing AD risk or clinical progression, it seems most likely that the variants in APOE sequence that affect AD risk are due to APOE’s ability to alter Aβ aggregation/clearance to influence to age of onset of Aβ accumulation in the brain. If one effect of APOE is to influence Aβ accumulation, then it would be critical to understand the molecular structure of APOE in its physiological state.

    In the brain, APOE is present in HDL-like lipoprotein particles. In amyloid plaques, APOE is found bound to plaques and, in this state, there is evidence that its conformation is different than in its physiological state (Liao et al., 2018). Factors that allow APOE to change its structure from its physiological form in lipidated particles to a non-lipidated form may be important in amyloid deposition.

    In assessing the structure of the V236E form of APOE, switching a nonpolar valine for an acidic glutamic acid might be predicted to reduce the hydrophobicity of this region, and reduce its tendency to oligomerize. This was tested by the Bu lab, where it was found that there were fewer Aβ aggregates and fewer ApoE aggregates in the brains of V236E carriers versus noncarriers. This APOE variant also resulted in less amyloid deposition in a mouse model (Liu et al., 2021). What structure in the C-terminal domain of ApoE in its physiological form explains these properties remains unclear.

    In regard to the fact that R251G appears to be directly in the lipid-binding C-terminal region of APOE (Dong et al., 1994), it is also possible that R251G confers a protective effect by influencing APOE’s ability to aggregate/form oligomers. Weisgraber, Agard, Mahley and colleagues originally proposed that an interaction between the N-terminal α-helical bundle and C-terminal α-helix of E4 with a salt bridge across Arg at position 61 and position Glu 255 is important in ApoE structure and DMPC binding (Hatters et al., 2005).

    Given that residue 251 sits directly above the E255 on the α-helix (3.6 A.A. per turn) that may interact with D65/E66 above the R61, the loss of positive charge in the R251G variant may impact a chain of salt bridges led by R61-E255. If so, the R251G may be altering this N- and C-terminal interaction in APOE to alter its stability in its lipidated state, making it more likely to aggregate as well. The ApoE4_R251G could also potentially become a key mutant to study the molecular characteristics in the N- and C-terminal interactions with biochemical assays.

    However, there are several structural models of APOE that have been more recently proposed by different groups (reviewed in Chen et al., 2021) when APOE is in its native, lipidated state. Depending on the actual structure of APOE in its native state, how the N- and C-terminals interact to influence ApoE structure will be critical to resolve, because it is likely such interactions are key in determining how ApoE alters AD risk.

    It has been difficult to determine the structure of apolipoproteins in their native lipidated state. New technologies such as cryoEM as well as small molecule FRET may be key in unlocking the high-resolution structure of lipidated APOE to get at some of these key issues.

    References:

    Liao F, Li A, Xiong M, Bien-Ly N, Jiang H, Zhang Y, Finn MB, Hoyle R, Keyser J, Lefton KB, Robinson GO, Serrano JR, Silverman AP, Guo JL, Getz J, Henne K, Leyns CE, Gallardo G, Ulrich JD, Sullivan PM, Lerner EP, Hudry E, Sweeney ZK, Dennis MS, Hyman BT, Watts RJ, Holtzman DM. Targeting of nonlipidated, aggregated apoE with antibodies inhibits amyloid accumulation. J Clin Invest. 2018 May 1;128(5):2144-2155. Epub 2018 Mar 30 PubMed.

    Liu CC, Murray ME, Li X, Zhao N, Wang N, Heckman MG, Shue F, Martens Y, Li Y, Raulin AC, Rosenberg CL, Doss SV, Zhao J, Wren MC, Jia L, Ren Y, Ikezu TC, Lu W, Fu Y, Caulfield T, Trottier ZA, Knight J, Chen Y, Linares C, Wang X, Kurti A, Asmann YW, Wszolek ZK, Smith GE, Vemuri P, Kantarci K, Knopman DS, Lowe VJ, Jack CR Jr, Parisi JE, Ferman TJ, Boeve BF, Graff-Radford NR, Petersen RC, Younkin SG, Fryer JD, Wang H, Han X, Frieden C, Dickson DW, Ross OA, Bu G. APOE3-Jacksonville (V236E) variant reduces self-aggregation and risk of dementia. Sci Transl Med. 2021 Sep 29;13(613):eabc9375. PubMed.

    Dong LM, Wilson C, Wardell MR, Simmons T, Mahley RW, Weisgraber KH, Agard DA. Human apolipoprotein E. Role of arginine 61 in mediating the lipoprotein preferences of the E3 and E4 isoforms. J Biol Chem. 1994 Sep 2;269(35):22358-65. PubMed.

    Hatters DM, Budamagunta MS, Voss JC, Weisgraber KH. Modulation of apolipoprotein E structure by domain interaction: differences in lipid-bound and lipid-free forms. J Biol Chem. 2005 Oct 7;280(40):34288-95. PubMed.

    Chen Y, Strickland MR, Soranno A, Holtzman DM. Apolipoprotein E: Structural Insights and Links to Alzheimer Disease Pathogenesis. Neuron. 2021 Jan 20;109(2):205-221. Epub 2020 Nov 10 PubMed.

    View all comments by David Holtzman
    • User Profile Image Nancy Ip
      Hong Kong University of Science & Technology
    • Posted:

    APOE is one of the most well-accepted genetic risk factors for Alzheimer’s disease, represented by the common coding variant APOE-ε4, which is known to modify Aβ metabolism. Along with APOE-ε4, recent studies have also identified coding and noncoding variants in this locus that modify disease risk. Specifically, indications are that some rare coding mutations, including APOE3-Jac (V236E) (Oct 2021 news) and Christchurch (R136S) (Nov 2019 news), alleviate disease risk by altering the function of the ApoE protein.

    In this study, the authors assembled a large group of participants (n = 544,384; who are predominantly European and admixed European individuals) to investigate APOE coding variants associated with the disease. The authors showed an AD protective effect of the V236E (APOE3-Jac, or rs199768005) mutation, which was co-inherited with the APOE-ε3 allele and exhibited a fourfold decrease in AD risk in the general population. Notably, the authors also identified another AD protective missense mutation, R251G (rs267606661), which was co-inherited with the APOE-ε4 allele and associated with a decreased chance of developing AD.

    It is interesting to note that all the identified AD protective missense mutations (Christchurch, APOE3-Jac, and R251G) are in the carboxyl-terminal of ApoE, which functions as the lipid-binding region. In particular, previous studies on V236E (APOE3-JAC) indicated that V236E can modify ApoE structure, reduce ApoE aggregation, and enhance its lipid-binding properties (Nov 2019 news; Liu et al., 2021). Further biochemical study is warranted to determine whether R251G would alter the structure and function of ApoE in a similar, or different, way, thereby helping us better understand the roles of APOE in health and disease.

    In addition, it would also be interesting to conduct similar analysis in populations of non-European descent, given that the local ancestry at the APOE locus, and its surrounding regions, can also influence the AD risk effect of APOE-ε4 (Zhou et al., 2021).

    References:

    Liu CC, Murray ME, Li X, Zhao N, Wang N, Heckman MG, Shue F, Martens Y, Li Y, Raulin AC, Rosenberg CL, Doss SV, Zhao J, Wren MC, Jia L, Ren Y, Ikezu TC, Lu W, Fu Y, Caulfield T, Trottier ZA, Knight J, Chen Y, Linares C, Wang X, Kurti A, Asmann YW, Wszolek ZK, Smith GE, Vemuri P, Kantarci K, Knopman DS, Lowe VJ, Jack CR Jr, Parisi JE, Ferman TJ, Boeve BF, Graff-Radford NR, Petersen RC, Younkin SG, Fryer JD, Wang H, Han X, Frieden C, Dickson DW, Ross OA, Bu G. APOE3-Jacksonville (V236E) variant reduces self-aggregation and risk of dementia. Sci Transl Med. 2021 Sep 29;13(613):eabc9375. PubMed.

    Zhou X, Fu AK, Ip NY. APOE signaling in neurodegenerative diseases: an integrative approach targeting APOE coding and noncoding variants for disease intervention. Curr Opin Neurobiol. 2021 Aug;69:58-67. Epub 2021 Feb 26 PubMed.

    View all comments by Nancy Ip

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References

News Citations

  1. Can an ApoE Mutation Halt Alzheimer’s Disease?
  2. Protective APOE3 Variant Binds More Lipids, Self-Aggregates Less
  3. Geneticists Seek Out Rare Contributors to Alzheimer’s

Paper Citations

  1. Arboleda-Velasquez JF, Lopera F, O'Hare M, Delgado-Tirado S, Marino C, Chmielewska N, Saez-Torres KL, Amarnani D, Schultz AP, Sperling RA, Leyton-Cifuentes D, Chen K, Baena A, Aguillon D, Rios-Romenets S, Giraldo M, Guzmán-Vélez E, Norton DJ, Pardilla-Delgado E, Artola A, Sanchez JS, Acosta-Uribe J, Lalli M, Kosik KS, Huentelman MJ, Zetterberg H, Blennow K, Reiman RA, Luo J, Chen Y, Thiyyagura P, Su Y, Jun GR, Naymik M, Gai X, Bootwalla M, Ji J, Shen L, Miller JB, Kim LA, Tariot PN, Johnson KA, Reiman EM, Quiroz YT. Resistance to autosomal dominant Alzheimer's disease in an APOE3 Christchurch homozygote: a case report. Nat Med. 2019 Nov;25(11):1680-1683. Epub 2019 Nov 4 PubMed.
  2. Chen J, Li Q, Wang J. Topology of human apolipoprotein E3 uniquely regulates its diverse biological functions. Proc Natl Acad Sci U S A. 2011 Sep 6;108(36):14813-8. Epub 2011 Aug 22 PubMed.
  3. Zhou X, Chen Y, Mok KY, Kwok TC, Mok VC, Guo Q, Ip FC, Chen Y, Mullapudi N, Alzheimer’s Disease Neuroimaging Initiative, Giusti-Rodríguez P, Sullivan PF, Hardy J, Fu AK, Li Y, Ip NY. Non-coding variability at the APOE locus contributes to the Alzheimer's risk. Nat Commun. 2019 Jul 25;10(1):3310. PubMed.

External Citations

  1. gnomAD

Further Reading

No Available Further Reading

Primary Papers

  1. Le Guen Y, Belloy ME, Grenier-Boley B, de Rojas I, Castillo-Morales A, Jansen I, Nicolas A, Bellenguez C, Dalmasso C, Küçükali F, Eger SJ, Rasmussen KL, Thomassen JQ, Deleuze JF, He Z, Napolioni V, Amouyel P, Jessen F, Kehoe PG, van Duijn C, Tsolaki M, Sánchez-Juan P, Sleegers K, Ingelsson M, Rossi G, Hiltunen M, Sims R, van der Flier WM, Ramirez A, Andreassen OA, Frikke-Schmidt R, Williams J, Ruiz A, Lambert JC, Greicius MD, Members of the EADB, GR@ACE, DEGESCO, DemGene, GERAD, and EADI Groups, Arosio B, Benussi L, Boland A, Borroni B, Caffarra P, Daian D, Daniele A, Debette S, Dufouil C, Düzel E, Galimberti D, Giedraitis V, Grimmer T, Graff C, Grünblatt E, Hanon O, Hausner L, Heilmann-Heimbach S, Holstege H, Hort J, Jürgen D, Kuulasmaa T, van der Lugt A, Masullo C, Mecocci P, Mehrabian S, de Mendonça A, Moebus S, Nacmias B, Nicolas G, Olaso R, Papenberg G, Parnetti L, Pasquier F, Peters O, Pijnenburg YA, Popp J, Rainero I, Ramakers I, Riedel-Heller S, Scarmeas N, Scheltens P, Scherbaum N, Schneider A, Seripa D, Soininen H, Solfrizzi V, Spalletta G, Squassina A, van Swieten J, Tegos TJ, Tremolizzo L, Verhey F, Vyhnalek M, Wiltfang J, Boada M, García-González P, Puerta R, Real LM, Álvarez V, Bullido MJ, Clarimon J, García-Alberca JM, Mir P, Moreno F, Pastor P, Piñol-Ripoll G, Molina-Porcel L, Pérez-Tur J, Rodríguez-Rodríguez E, Royo JL, Sánchez-Valle R, Dichgans M, Rujescu D. Association of Rare APOE Missense Variants V236E and R251G With Risk of Alzheimer Disease. JAMA Neurol. 2022 Jul 1;79(7):652-663. PubMed.
  2. Rabinovici GD, Dubal DB. Rare APOE Missense Variants-Can We Overcome APOE ε4 and Alzheimer Disease Risk?. JAMA Neurol. 2022 Jul 1;79(7):649-651. PubMed.

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