Overview

Pathogenicity: Cerebral Amyloid Angiopathy : Pathogenic
ACMG/AMP Pathogenicity Criteria: PS3, PM1, PM2, PM5, PP2, PP3, PP4
Clinical Phenotype: Hereditary Cerebral Hemorrhage with Amyloidosis of the Dutch type
Reference Assembly: GRCh37/hg19
Position: Chr21:27264168 G>C
dbSNP ID: rs63750579
Coding/Non-Coding: Coding
DNA Change: Substitution
Expected RNA Consequence: Substitution
Expected Protein Consequence: Missense
Codon Change: GAA to CAA
Reference Isoform: APP Isoform APP770 (770 aa)
Genomic Region: Exon 17
Research Models: 4

Findings

Carriers of this mutation develop a severe hereditary form of cerebral amyloid angiopathy (CAA), known as hereditary cerebral hemorrhage with amyloidosis, Dutch type (HCHWA-D). This disease is associated with recurrent strokes during the fifth and sixth decades of life often causing focal neurological symptoms and signs, including focal seizures. In addition to severe hemorrhages, extensive amyloid develops in the vasculature. The mutation has been described in three Dutch families (Wattendorff et al., 1982; Luyendijk and Bots, 1986) and it is estimated that as many as 500 descendants of these families are at risk of the disease. An additional affected family of Dutch descent was described in Western Australia (Panegyres et al., 2005). The discovery of this mutation was an early demonstration that a variant in the APP gene could cause severe amyloid deposition (Levy et al., 1990; van Broeckhoven et al., 1990; Fernandez-Madrid et al., 1991).

This variant was absent from the gnomAD variant database (v2.1.1, Oct 2021).

Neuropathology

This mutation is associated with severe amyloid deposition in cerebral vessels, hemorrhages, secondary microvascular degeneration, and diffuse plaques in brain parenchyma (Timmers et al., 1990, Vinters et al., 1998). Extensive Aβ accumulates in the cerebral vessels, especially the meningeal arteries and the cerebro-cortical arterioles. Angiopathy in the cerebellar cortex has also been reported (Wattendorff et al., 1995). In this same study, hemorrhages and hemorrhagic infarcts were observed in the subcortical white matter, a region particularly vulnerable to impaired cortical circulation. In people with HCHWA-D, the amount of CAA is correlated with dementia, whereas parenchymal plaque density and intraneuronal neurofibrillary tangles are not (Natté et al., 2001). In contrast to patients with AD, neurofibrillary tangles are not a prominent neuropathological feature of HCHWA-D.

Interestingly, one study found abundant fibrin deposits and fibrinogen/Aβ co-deposits in the occipital cortices of mutation carriers (Cajamarca et al., 2020). The mutation appears to increase Aβ’s affinity for fibrinogen which may exacerbate CAA, perturb clot structure, and delay fibrinolysis.

Biological Effect

This mutation results in the accumulation of Aβ in cerebral vessel walls which is associated with cell death and loss of vessel wall integrity. Vessels become prone to obstruction and rupture, manifesting clinically as hemorrhages and infarcts. Toxicity has been reported in cerebral microvessels and aortic smooth muscle cells (Wang et al., 2000), as well as in human cerebrovascular smooth muscle cells (Van Nostrand et al., 2001), cerebral endothelial cells (Miravalle et al., 2000), and PC12 cells (Murakami et al., 2003). Moreover, a transcriptomic analysis of post-mortem frontal and occipital cortical brain tissue from nine affected carriers indicated downregulation of genes involved in aerobic respiration and upregulation of genes related to extracellular matrix interactions compared with healthy controls (Grand Moursel et al., 2018). The latter alteration was similar to that observed in presymptomatic transgenic mice carrying the E693 mutation.

In vitro, this mutation accelerates Aβ aggregation, leading to increased fibril formation (Wisniewski et al., 1991). Subsequent studies have shown that the mutant peptide, a.k.a. E22Q, forms long fibrillar bundles that may be twisted, as well as short fibrils, densely packed fibril clusters, and amorphous structures. Aggregation lag time is much shorter than that of wildtype Aβ, but longer than those of other Aβ mutant peptides (Fawzi et al., 2008, Hatami et al., 2017, Yang et al., 2018, Illes-Toth et al., 2021, Lee et al., 2021).

The E22Q substitution is a fairly conservative substitution regarding the volume of the amino acid side-chain and hydrogen-bonding, but it eliminates a negative charge (Yang et al., 2018). Although the structures of wildtype and mutant monomeric peptides in solution are similar (Esler et al., 2000), the mutant appears to adopt a more disordered, more flexible conformation than wildtype peptides (Esler et al., 2000, Massi et al., 2001) and a larger hydrophobic surface area is exposed to the solvent which facilitates aggregation (Massi et al., 2001, Massi and Straub, 2003, Bitan et al., 2003). Also, the elimination of the negative charge relative to the wildtype peptide has been proposed to lower the repulsion between Aβ monomers and the barrier for nucleation, as well as accelerate fibrillization (Lee et al., 2021). The peptide has high levels of β-sheet conformation (Miravalle et al., 2000), and, in particular, a β-turn at positions 21-23 has been associated with enhanced aggregation (Murakami et al., 2003). Adding mutations that increase α-helical structure decreased E22Q aggregation (He et al., 2020).

One study using molecular dynamics simulations predicted mutant peptide oligomers are more internally stable than wildtype (Kassler et al., 2010). Another simulation-based study predicted destabilization of hydrogen bonding required for α-helix formation and the adoption of a short β-sheet lacking some of the longer-range contacts present in the wildtype peptide (Davidson et al., 2022). The authors of this latter study hypothesized that increased solvent exposure at the C-terminus could fuel fibril formation.

A few studies have suggested E22Q may insert into cell membranes and cause cytotoxicity. One in vitro study, for example, indicated E22Q trimers and/or hexamers destabilize membranes and disrupt their integrity (McKnelly et al., 2022). Similarly, a structural modeling study predicted the mutant peptide partitions into membranes more than wild type Aβ and suggested this may disrupt the normal function of these peptides in the endosomal lumen where they have been proposed to play important roles in synaptic and neuronal function (Kim and Bezprozvanny, 2023). Also of note, E693 lies within a cholesterol-binding site as determined by NMR resonance spectroscopy and site-directed mutagenesis (Barrett et al., 2012).

Altered turnover of E22Q may also play a role in its toxicity. One study, for example, found that mutant Aβ40 is poorly cleared from the cerebrospinal fluid and its transport from the central nervous system into the bloodstream is reduced compared with wildtype Aβ40 (Monro et al., 2002). Moreover, E22Q aggregates appear to be resistant to proteolytic degradation by neprilysin, a peptidase that degrades Aβ in the brain (Tsubuki et al., 2003). On the other hand, production and activation of matrix metalloprotease 2 in human brain endothelial cells increased after E22Q exposure. This could help degrade the peptide and delay toxicity, but may also compromise the integrity of the blood-brain barrier (Hernandez-Guillamon et al., 2010). 

In addition to altered clearance, E22Q may suffer from disrupted production. One study reported cells transfected with E693Q generated elevated levels of extracellular Aβ oligomers compared with cells transfected with wildtype APP (Ohshima et al., 2018). Moreover, in at least one study, the processing of APP was found to be disrupted in human embryonic kidney cells at peptide positions D1, V18, and F19 (Watson et al., 1999). A study in H4 neuroglioma cells reported no significant changes in the amounts of full-length APP, secreted APPα, C83, or Aβ40 (Van Nostrand et al., 2001).

This variant's PHRED-scaled CADD score, which integrates diverse information in silico, was above 20, consistent with it having a deleterious effect (CADD v.1.6, Oct 2021).

Pathogenicity

Cerebral Amyloid Angiopathy : Pathogenic*

*Although not AD, the condition associated with this variant is inherited in an autosomal dominant manner, so its pathogenicity was classified using the ACMG-AMP guidelines.

This variant fulfilled the following criteria based on the ACMG/AMP guidelines. See a full list of the criteria in the Methods page.

Research Models

Transgenic APP with the Dutch mutation has been introduced into several mouse lines that model CAA and vascular amyloid in AD. The models, such as the well-characterized Tg-SwDI and APPDutch mice, develop prominent vascular amyloid.

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References

Research Models Citations

1. Tg-SwDI (APP-Swedish,Dutch,Iowa)
2. APPDutch

Paper Citations

1. Wattendorff AR, Bots GT, Went LN, Endtz LJ. Familial cerebral amyloid angiopathy presenting as recurrent cerebral haemorrhage. J Neurol Sci. 1982 Aug;55(2):121-35. PubMed.
2. Luyendijk W, Bots GT. Hereditary cerebral hemorrhage. Scand J Clin Lab Invest. 1986 Jun;46(4):391. PubMed.
3. Panegyres PK, Kwok JB, Schofield PR, Blumbergs PC. A Western Australian kindred with Dutch cerebral amyloid angiopathy. J Neurol Sci. 2005 Dec 15;239(1):75-80. Epub 2005 Oct 5 PubMed.
4. Levy E, Carman MD, Fernandez-Madrid IJ, Power MD, Lieberburg I, van Duinen SG, Bots GT, Luyendijk W, Frangione B. Mutation of the Alzheimer's disease amyloid gene in hereditary cerebral hemorrhage, Dutch type. Science. 1990 Jun 1;248(4959):1124-6. PubMed.
5. Van Broeckhoven C, Haan J, Bakker E, Hardy JA, Van Hul W, Wehnert A, Vegter-Van der Vlis M, Roos RA. Amyloid beta protein precursor gene and hereditary cerebral hemorrhage with amyloidosis (Dutch). Science. 1990 Jun 1;248(4959):1120-2. PubMed.
6. Fernandez-Madrid I, Levy E, Marder K, Frangione B. Codon 618 variant of Alzheimer amyloid gene associated with inherited cerebral hemorrhage. Ann Neurol. 1991 Nov;30(5):730-3. PubMed.
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9. Wattendorff AR, Frangione B, Luyendijk W, Bots GT. Hereditary cerebral haemorrhage with amyloidosis, Dutch type (HCHWA-D): clinicopathological studies. J Neurol Neurosurg Psychiatry. 1995 Jun;58(6):699-705. PubMed.
10. Natté R, Maat-Schieman ML, Haan J, Bornebroek M, Roos RA, van Duinen SG. Dementia in hereditary cerebral hemorrhage with amyloidosis-Dutch type is associated with cerebral amyloid angiopathy but is independent of plaques and neurofibrillary tangles. Ann Neurol. 2001 Dec;50(6):765-72. PubMed.
11. Cajamarca SA, Norris EH, van der Weerd L, Strickland S, Ahn HJ. Cerebral amyloid angiopathy-linked β-amyloid mutations promote cerebral fibrin deposits via increased binding affinity for fibrinogen. Proc Natl Acad Sci U S A. 2020 Jun 23;117(25):14482-14492. Epub 2020 Jun 9 PubMed.
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14. Miravalle L, Tokuda T, Chiarle R, Giaccone G, Bugiani O, Tagliavini F, Frangione B, Ghiso J. Substitutions at codon 22 of Alzheimer's abeta peptide induce diverse conformational changes and apoptotic effects in human cerebral endothelial cells. J Biol Chem. 2000 Sep 1;275(35):27110-6. PubMed.
15. Murakami K, Irie K, Morimoto A, Ohigashi H, Shindo M, Nagao M, Shimizu T, Shirasawa T. Neurotoxicity and physicochemical properties of Abeta mutant peptides from cerebral amyloid angiopathy: implication for the pathogenesis of cerebral amyloid angiopathy and Alzheimer's disease. J Biol Chem. 2003 Nov 14;278(46):46179-87. Epub 2003 Aug 27 PubMed.
16. Grand Moursel L, van Roon-Mom WM, Kiełbasa SM, Mei H, Buermans HP, van der Graaf LM, Hettne KM, de Meijer EJ, van Duinen SG, Laros JF, van Buchem MA, 't Hoen PA, van der Maarel SM, van der Weerd L. Brain Transcriptomic Analysis of Hereditary Cerebral Hemorrhage With Amyloidosis-Dutch Type. Front Aging Neurosci. 2018;10:102. Epub 2018 Apr 13 PubMed.
17. Wisniewski T, Ghiso J, Frangione B. Peptides homologous to the amyloid protein of Alzheimer's disease containing a glutamine for glutamic acid substitution have accelerated amyloid fibril formation. Biochem Biophys Res Commun. 1991 Nov 14;180(3):1528. PubMed.
18. Fawzi NL, Kohlstedt KL, Okabe Y, Head-Gordon T. Protofibril assemblies of the arctic, Dutch, and Flemish mutants of the Alzheimer's Abeta1-40 peptide. Biophys J. 2008 Mar 15;94(6):2007-16. Epub 2007 Nov 21 PubMed.
19. Hatami A, Monjazeb S, Milton S, Glabe CG. Familial Alzheimer's Disease Mutations within the Amyloid Precursor Protein Alter the Aggregation and Conformation of the Amyloid-β Peptide. J Biol Chem. 2017 Feb 24;292(8):3172-3185. Epub 2017 Jan 3 PubMed.
20. Yang X, Meisl G, Frohm B, Thulin E, Knowles TP, Linse S. On the role of sidechain size and charge in the aggregation of Aβ42 with familial mutations. Proc Natl Acad Sci U S A. 2018 Jun 26;115(26):E5849-E5858. Epub 2018 Jun 12 PubMed.
21. Illes-Toth E, Meisl G, Rempel DL, Knowles TP, Gross ML. Pulsed Hydrogen-Deuterium Exchange Reveals Altered Structures and Mechanisms in the Aggregation of Familial Alzheimer's Disease Mutants. ACS Chem Neurosci. 2021 Jun 2;12(11):1972-1982. Epub 2021 May 14 PubMed.
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24. Massi F, Peng JW, Lee JP, Straub JE. Simulation study of the structure and dynamics of the Alzheimer's amyloid peptide congener in solution. Biophys J. 2001 Jan;80(1):31-44. PubMed.
25. Massi F, Straub JE. Structural and dynamical analysis of the hydration of the Alzheimer's beta-amyloid peptide. J Comput Chem. 2003 Jan 30;24(2):143-53. PubMed.
26. Bitan G, Vollers SS, Teplow DB. Elucidation of primary structure elements controlling early amyloid beta-protein oligomerization. J Biol Chem. 2003 Sep 12;278(37):34882-9. Epub 2003 Jul 2 PubMed.
27. He KC, Chen YR, Liang CT, Huang SJ, Tzeng CY, Chang CF, Huang SJ, Huang HB, Lin TH. Conformational Characterization of Native and L17A/F19A-Substituted Dutch-Type β-Amyloid Peptides. Int J Mol Sci. 2020 Apr 7;21(7) PubMed.
28. Kassler K, Horn AH, Sticht H. Effect of pathogenic mutations on the structure and dynamics of Alzheimer's A beta 42-amyloid oligomers. J Mol Model. 2010 May;16(5):1011-20. Epub 2009 Nov 12 PubMed.
29. Davidson DS, Kraus JA, Montgomery JM, Lemkul JA. Effects of Familial Alzheimer's Disease Mutations on the Folding Free Energy and Dipole-Dipole Interactions of the Amyloid β-Peptide. J Phys Chem B. 2022 Oct 6;126(39):7552-7566. Epub 2022 Sep 23 PubMed.
30. McKnelly KJ, Kreutzer AG, Howitz WJ, Haduong K, Yoo S, Hart C, Nowick JS. Effects of Familial Alzheimer's Disease Mutations on the Assembly of a β-Hairpin Peptide Derived from Aβ16-36. Biochemistry. 2022 Mar 15;61(6):446-454. Epub 2022 Feb 25 PubMed.
31. Kim M, Bezprozvanny I. Analysis of Non-Amyloidogenic Mutations in APP Supports Loss of Function Hypothesis of Alzheimer's Disease. Int J Mol Sci. 2023 Jan 20;24(3) PubMed.
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33. Monro OR, Mackic JB, Yamada S, Segal MB, Ghiso J, Maurer C, Calero M, Frangione B, Zlokovic BV. Substitution at codon 22 reduces clearance of Alzheimer's amyloid-beta peptide from the cerebrospinal fluid and prevents its transport from the central nervous system into blood. Neurobiol Aging. 2002;23(3):405-12. PubMed.
34. Tsubuki S, Takaki Y, Saido TC. Dutch, Flemish, Italian, and Arctic mutations of APP and resistance of Abeta to physiologically relevant proteolytic degradation. Lancet. 2003 Jun 7;361(9373):1957-8. PubMed.
35. Hernandez-Guillamon M, Mawhirt S, Fossati S, Blais S, Pares M, Penalba A, Boada M, Couraud PO, Neubert TA, Montaner J, Ghiso J, Rostagno A. Matrix metalloproteinase 2 (MMP-2) degrades soluble vasculotropic amyloid-beta E22Q and L34V mutants, delaying their toxicity for human brain microvascular endothelial cells. J Biol Chem. 2010 Aug 27;285(35):27144-27158. Epub 2010 Jun 24 PubMed.
36. Ohshima Y, Taguchi K, Mizuta I, Tanaka M, Tomiyama T, Kametani F, Yabe-Nishimura C, Mizuno T, Tokuda T. Mutations in the β-amyloid precursor protein in familial Alzheimer's disease increase Aβ oligomer production in cellular models. Heliyon. 2018 Jan;4(1):e00511. Epub 2018 Feb 1 PubMed.
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Further Reading