Overview

Clinical Phenotype: Alzheimer's Disease, Multiple Conditions
Reference Assembly: GRCh37/hg19
Position: Chr19:45412013 C>T
Transcript: NM_000041; ENSG00000130203
dbSNP ID: rs121918393
Coding/Non-Coding: Coding
DNA Change: Substitution
Expected RNA Consequence: Substitution
Expected Protein Consequence: Missense
Codon Change: CGC to TGC
Reference Isoform: APOE Isoform 1
Genomic Region: Exon 4

Findings

This variant was examined in a study of dementia, including Alzheimer’s disease (AD), and was proposed as being possibly protective (Rasmussen et al., 2020). In two cohorts totaling more than 100,000 individuals from Copenhagen, Denmark, the variant was found in 22 individuals, 12 who were 60 years old or older, none of whom had dementia (3.7 percent of non-carriers in this age bracket had AD). In addition, carriers of the variant had high levels of ApoE in blood, a finding that suggested low AD risk based on an analyses of nine APOE variants, including G145D, showing that low ApoE in plasma was associated with increased AD risk, whereas high levels were associated with reduced risk, after adjustment for the common APOE2/E3/E4 alleles. The authors noted these features suggested a similarity between R154C and the protective allele R176C (APOE2). Interestingly, a subsequent study of the same Danish cohorts revealed yet another similarity with APOE2: an association with increased risk of age-related macular degeneration (aHR, 4.52; 95% CI, 1.13-18.13; Rasmussen et al., 2022).

Of note, another variant at this same position, R154S (Christchurch), has been tied to protection against autosomal dominant AD and APOE4-driven brain pathologies. 

Non-Neurological Conditions

R154C has been associated with elevated lipids in blood, hyperlipidemia, and more specifically the risk for hyperlipoproteinemia type III (HLPP3), also known as familial dysbetalipoproteinemia, a hyperlipidemic condition that can lead to early onset atherosclerosis and heart disease. The mutation was first identified in a 39-year-old Canadian man diagnosed with HLPP3 (Walden et al., 1994). He was one of 22 patients whose APOE proteins migrated on an isoelectric focusing gel to the position of the R176C (APOE2) allele, the most common cause of HLPP3 when present in homozygous form. Restriction isotyping, followed by DNA sequencing, however, revealed that he had only one copy of APOE2, while the other APOE allele harbored the R154C mutation on an APOE3 backbone. The mutation was also found in the proband’s father, who appeared to be homozygous for APOE3, as well as in his two brothers who, like the proband, carried an APOE2 allele. The father’s plasma lipid profile, however, was not consistent with HLPP3. Furthermore, although both brothers had the β form of very low-density lipoprotein (β-VLDL), a particularly atherogenic lipoprotein particle characteristic of HLPP3, they were not hyperlipidemic and did not fulfill the criteria for HLPP3 diagnosis.

Subsequent studies have also found variability in the conditions associated with this variant. For example, while in one German family the mutation appeared to have a late-onset dominant effect, with the proband and her father suffering from severe HLPP3 (Feussner et al., 1996a), another study reported that none of four German carriers had HLPP3 (März et al., 1998). In this latter study, two of the carriers had elevated triglyceride levels, but their VLDL levels and β-VLDL levels were normal, resulting in a diagnosis of a different type of lipoprotein disease, HLPP4. Other studies also indicate the mutation is not always associated with HLPP3 (Hubácek et al., 2000, Hubácek et al., 2002, Hubácek et al., 2008).  Sometimes carriers have no detectable abnormality in their plasma lipid profiles, even within the same family of an affected carrier (Hubácek et al., 2000). A compilation of several studies reported that 22 of 39 carriers have been diagnosed with HLPP3 (Koopal et al., 2017).

In most cases, the mutation appears to be associated with some disruption of lipid metabolism, even when HLPP3 is not present. A study of 12 Czech mutation carriers, for example, found that, except for one individual, all carriers had elevated plasma lipids, or were being treated for hyperlipidemia (Hubácek et al., 2009). Moreover, in 22 Danish carriers, including two who were on lipid-lowering therapy, average plasma triglyceride levels were high (16 percent higher than non-carriers), as were ApoE (67 percent higher) and remnant cholesterol (50 percent higher), while both low-density lipoprotein (LDL) cholesterol and high-density lipoprotein (HDL) cholesterol levels were low (16 and 12 percent lower than non-carriers, respectively)  (Rasmussen et al., 2020, Rasmussen et al., 2023). The latter study also found that individuals with high levels of ApoE, triglycerides, and remnant cholesterol who carried rare APOE variants, such as R154C, were at higher risk of peripheral arterial disease. 

In addition, other conditions have been tied to R154C. Heart disease, for example, has been found with and without associated plasma lipid abnormalities. Two of three Czech patients carrying the mutation had elevated plasma lipid levels and a family history of coronary artery disease (Vrablík et al., 2003). Two other Czech individuals, however, had premature myocardial infarction, without dyslipidemia (Hubácek et al., 2009). Also, a potential connection with body weight has been noted. Hubácek and colleagues observed that nearly all of the carriers they studied, over a dozen, were overweight (Hubácek et al., 2008, Hubácek et al., 2009).

The global frequency of this mutation as reported in gnomAD was 0.00009—including 14 carriers of which seven were of South Asian ancestry and six of non-Finnish European ancestry (gnomAD v2.1.1, May 2022). Frequencies may be higher in some subgroups, however. For example, in a Czech population study of approximately 1,000 individuals, the mutation was found in 1.2 percent of all participants (Hubácek et al., 2008). Indeed, this high incidence has raised questions about the variant’s role in dyslipidemia (Evans et al., 2013).

The variant has been found in carriers of all three common APOE isoforms, APOE2, APOE3, and APOE4 (e.g., Walden et al., 1994; Feussner et al., 1996a; Rasmussen et al., 2020).

Biological Effect

This mutation, located in the receptor-binding region of ApoE, appears to reduce binding to LDL receptors. An indirect assay to test the binding of carrier VLDL particles to  receptors on the surface of cultured mouse macrophages showed the mutant’s binding is reduced relative to ApoE3 (33 percent), although not to the extent of that of ApoE2 (Walden et al., 1994). Consistent with these findings, experiments using human skin fibroblasts and labeled ApoE particles—recombinant mutant ApoE loaded onto either synthetic lipid vesicles or VLDL particles from ApoE-deficient individuals—indicated LDL receptor binding was only 14 percent that of ApoE3 (März et al., 1998). Interestingly, an artificial substitution at this same site, R154A, substantially reduced binding of ApoE4 to the microglial leukocyte immunoglobulin-like receptor B3 (LilrB3), a receptor that binds to ApoE4 more strongly than to ApoE3 or ApoE2 and activates pro-inflammatory pathways (Zhou et al., 2023).

Although R154 has been predicted to be one of eight amino acids lining the shallow groove that binds and makes direct contact with the sulfo groups of heparan sulfate proteoglycans (Libeu et al., 2001), the effect of R154C on heparin affinity is unclear. One group reported heparin binding to be 61 percent that of ApoE3 (März et al., 1998), while another reported no detectable alteration (Feussner et al., 1996a).

This mutation alters an amino acid that is highly conserved in mammals (Feussner et al., 1996a; Frieden, 2015). The replacement of a basic arginine by a neutral cysteine may affect ionic interactions with negatively charged amino acids of the LDL receptor-binding domain. Moreover, computer modeling suggested an effect on ApoE three-dimensional structure (Feussner et al., 1996b).

This variant's PHRED-scaled CADD score, which integrates diverse information in silico, was above 20, suggesting a deleterious effect (CADD v.1.6, May 2022).

Note on nomenclature

Some papers include APOE2 in the name of this variant because its isoelectric migration is very similar to that of R176C (APOE2).

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References

Mutations Citations

1. APOE R176C (ApoE2)
2. APOE R154S (Christchurch)

Paper Citations

1. Rasmussen KL, Tybjaerg-Hansen A, Nordestgaard BG, Frikke-Schmidt R. APOE and dementia - resequencing and genotyping in 105,597 individuals. Alzheimers Dement. 2020 Dec;16(12):1624-1637. Epub 2020 Aug 18 PubMed.
2. Rasmussen KL, Tybjærg-Hansen A, Nordestgaard BG, Frikke-Schmidt R. Associations of Alzheimer Disease-Protective APOE Variants With Age-Related Macular Degeneration. JAMA Ophthalmol. 2023 Jan 1;141(1):13-21. PubMed.
3. Walden CC, Huff MW, Leiter LA, Connelly PW, Hegele RA. Detection of a new apolipoprotein-E mutation in type III hyperlipidemia using deoxyribonucleic acid restriction isotyping. J Clin Endocrinol Metab. 1994 Mar;78(3):699-704. PubMed.
4. Feussner G, Albanese M, Mann WA, Valencia A, Schuster H. Apolipoprotein E2 (Arg-136-->Cys), a variant of apolipoprotein E associated with late-onset dominance of type III hyperlipoproteinaemia. Eur J Clin Invest. 1996 Jan;26(1):13-23. PubMed.
5. März W, Hoffmann MM, Scharnagl H, Fisher E, Chen M, Nauck M, Feussner G, Wieland H. Apolipoprotein E2 (Arg136 --> Cys) mutation in the receptor binding domain of apoE is not associated with dominant type III hyperlipoproteinemia. J Lipid Res. 1998 Mar;39(3):658-69. PubMed.
6. Hubácek JA, Pitha J, Stávek P, Schmitz G, Poledne R. Variable expression of hypercholesterolemia in Apolipoprotein E2* (Arg136 --> Cys) heterozygotes. Physiol Res. 2000;49(3):307-14. PubMed.
7. Hubacek JA, Pitha J, Skodová Z, Poledne R. Rare variant of apolipoprotein E (Arg136-->Cys) in a subject with normal lipid values. Physiol Res. 2002;51(1):107-8. PubMed.
8. Hubacek JA, Adamková V, Stavek P, Kubinova R, Peasey A, Pikhart H, Marmot M, Bobak M. Apolipoprotein E Arg136 --> Cys mutation and hyperlipidemia in a large central European population sample. Clin Chim Acta. 2008 Feb;388(1-2):217-8. PubMed.
9. Koopal C, Marais AD, Westerink J, Visseren FL. Autosomal dominant familial dysbetalipoproteinemia: A pathophysiological framework and practical approach to diagnosis and therapy. J Clin Lipidol. 2017 Jan - Feb;11(1):12-23.e1. Epub 2016 Oct 13 PubMed.
10. Hubácek JA, Poledne R, Pitha J, Aschermann M, Skalická H, Stanek V. Apolipoprotein E Arg136 --> Cys in individuals with premature myocardial infarction. Folia Biol (Praha). 2009;55(3):116-8. PubMed.
11. Rasmussen KL, Luo J, Nordestgaard BG, Tybjærg-Hansen A, Frikke-Schmidt R. APOE and vascular disease: Sequencing and genotyping in general population cohorts. Atherosclerosis. 2023 Nov;385:117218. Epub 2023 Aug 9 PubMed.
12. Vrablík M, Horínek A, Ceska R, Stulc T, Kvasnicka T. Familial dysbetalipoproteinemia in three patients with apoE 2*(Arg136-->Cys) gene variant. Physiol Res. 2003;52(5):647-50. PubMed.
13. Evans D, Beil FU, Aberle J. Resequencing the APOE gene reveals that rare mutations are not significant contributory factors in the development of type III hyperlipidemia. J Clin Lipidol. 2013 Nov-Dec;7(6):671-4. Epub 2013 May 25 PubMed.
14. Zhou J, Wang Y, Huang G, Yang M, Zhu Y, Jin C, Jing D, Ji K, Shi Y. LilrB3 is a putative cell surface receptor of APOE4. Cell Res. 2023 Feb;33(2):116-130. Epub 2023 Jan 2 PubMed.
15. Libeu CP, Lund-Katz S, Phillips MC, Wehrli S, Hernáiz MJ, Capila I, Linhardt RJ, Raffaï RL, Newhouse YM, Zhou F, Weisgraber KH. New insights into the heparan sulfate proteoglycan-binding activity of apolipoprotein E. J Biol Chem. 2001 Oct 19;276(42):39138-44. Epub 2001 Aug 10 PubMed.
16. Frieden C. ApoE: the role of conserved residues in defining function. Protein Sci. 2015 Jan;24(1):138-44. Epub 2014 Dec 9 PubMed.
17. Feussner G, Albanese M, Valencia A. Three-dimensional structure of the LDL receptor-binding domain of the human apolipoprotein E2 (Arg136-->Cys) variant. Atherosclerosis. 1996 Oct 25;126(2):177-84. PubMed.

Further Reading