The First Japanese Case of Familial Hypercholesterolemia Caused by an apolipoprotein E (APOE) p.Leu167del Mutation

Abstract

We herein report the first Japanese case of familial hypercholesterolemia (FH) caused by a specific mutation in apolipoprotein E (APOE) [c.500_502delTCC (p.Leu167del)]. The proband was a 38-year-old man diagnosed with FH based on the clinical findings. Genetic testing revealed a rare pathogenic variant in APOE but no relevant mutation in any “FH genes,” including low-density lipoprotein (LDL) receptor, proprotein convertase subtilisin/kexin type 9, apolipoprotein B, and LDL receptor adaptor protein 1. His LDL cholesterol level was well controlled by the introduction of statins, ezetimibe, and proprotein convertase subtilisin/kexin type 9 inhibitors. Cascade and reverse cascade screening identified his son and father as also having FH caused by this particular mutation.

Introduction

Familial hypercholesterolemia (FH) is one of the most common inherited disorders characterized caused by genetic mutations in so-called “FH genes” associated with low-density lipoprotein (LDL) metabolism, such as LDL receptor (LDLR), apolipoprotein B (APOB), proprotein convertase subtilisin/kexin type 9 (PCSK9), or LDLR adaptor protein 1 (LDLRAP1) (1). It has been shown that the mutation detection rate among patients who fulfilled clinical criteria of FH was around 60%. In other words, as many as 40% of patients with clinical FH do not have any pathogenic mutations in the FH genes listed above (2).

We and others have shown that some of these patients without pathogenic variants in FH genes can be explained by polygenic FH caused by the accumulation of common genetic variations that can raise LDL cholesterol levels substantially (3,4), and sitosterolemia, which is a phenocopy of FH caused by rare genetic mutations in the ATP-binding cassette subfamily G member 5 (ABCG5) or ATP-binding cassette subfamily G member 8 (ABCG8) (5). There is another situation where a particular in-frame three base-pair deletion genetic mutation in apolipoprotein E (APOE) [c.500_502delTCC (p.Leu167del)] has been shown to be a rather rare cause of FH (6,7). Interestingly, the pathogenicity of this mutation has been confirmed by several segregation and functional analyses (8). In addition, we used an APOE knockout mouse model as an experimental model of FH and premature atherosclerosis (9). Therefore, it is not surprising that loss-of-function mutations in humans can lead to the FH phenotype. However, no prior data exists regarding patients with FH caused by this pathogenic mutation in APOE among the Japanese population.

We herein report the first Japanese family with FH caused by a specific mutation in APOE [c.500_502delTCC (p.Leu167del)]. The proband patient's LDL cholesterol level was well controlled by the introduction of statins, ezetimibe, and PCSK9 inhibitors. Cascade and reverse cascade screening identified his son and father as patients with FH caused by this particular mutation.

Case Report

A 38-years-old man was referred to our hospital with hyper-LDL cholesterolemia (256 mg/dL). He had had hyper-LDL cholesterolemia pointed out at least since he was 20 years old. However, he did not start treatment to reduce his LDL level because he had no symptoms at that time. In addition, he did not undergo any medical checkups until he was 40 years old, because he did not experience any symptoms. When he underwent his first medical checkup at 38 years old, his total cholesterol level was 321 mg/dL, and his LDL cholesterol level was 256 mg/dL. X-ray of the Achilles tendon revealed thickening of both tendons (Fig. 1). Based on his LDL cholesterol level and Achilles tendon thickness, he was diagnosed with FH (10) and referred to our hospital for a further examination. He had a family history of dyslipidemia in his father and son.

Figure 1.

X-ray of the Achilles tendon of the proband. (A) Before treatment (at 38 years old). (B) After treatment (at 40 years old).

Carotid ultrasonography revealed minor stenotic lesions in both carotid arteries. The maximum dose of statins (rosuvastatin 20 mg) and ezetimibe was introduced immediately after his referral to our hospital, and a PCSK9 inhibitor (evolocumab 140 mg every 2 weeks) was also introduced. His LDL cholesterol level reduced to 56 mg/dL after the introduction of these combination therapies. In parallel with drug titrations, genetic testing was performed to investigate the background of this situation. We found a heterozygous variant in APOE [c.500_502delTCC (p.Leu167del)], while there was no relevant mutation in “FH genes,” including LDLR, PCSK9, APOB, and LDLRAP1. Importantly, this mutation in APOE is considered a pathogenic variant of FH (6-8). During this titration of combination therapy, we observed regression of cutaneous and Achilles tendon xanthomas within two years (Fig. 1). The patient did not exhibit any apparent side effects, including hepatic, fatty liver, or kidney dysfunction.

Cascade screening and reverse cascade screening based on genetic testing identified the same pathogenic mutation in APOE in his father and son (Fig. 2). According to these results, LDL cholesterol-lowering therapy for his father was intensified to add a PCSK9 inhibitor (evolocumab 140 mg every 2 weeks) to meet the treatment target of clinical guidelines (LDL cholesterol ＜100 mg/dL), and his LDL cholesterol was lowered to approximately 60 mg/dL. His son, who was 8 years old, also had elevated LDL cholesterol levels (190 mg/dL), and we discussed how and when to treat him. We came to the agreement that statins would be introduced when he reached 10 years old, according to the JAS guideline treatment target of ＜100 mg/dL (11). We also confirmed the genotype-phenotype correlation between this particular mutation and hyper-LDL cholesterolemia in his family (Table). We found that individuals with this mutation exhibited hyper-LDL cholesterolemia and elevated apolipoprotein B levels that were compatible with the diagnosis of FH. Interestingly, we did not find any significant differences in the APOE levels.

Figure 2.

Family tree. The black arrow indicates the proband. The black color shows the status of c.500_502delTCC (p.Leu167del) (APOE). The numbers indicate the total cholesterol, triglyceride, HDL cholesterol, and LDL cholesterol levels. APOE: apolipoprotein E, HDL: high-density lipoprotein, LDL: low-density lipoprotein

Table.

Characteristics of the Study Subjects.

Subject (gender)	I.1 (woman)	I.2* (man)	I.3 (woman)	II.1 (man)	II.2 (woman)	III.1 (man)
APOE c.500_502delTCC (p.Leu167del) status	W/W	M/W	W/W	M/W	W/W	M/W
Age (yr)	74	72	72	38	37	8
Total cholesterol (mg/dL) (142-219)	201	186	224	321	188	268
Triglyceride (mg/dL) (40-149)	134	120	166	102	144	98
HDL cholesterol (mg/dL) (40-90)	65	45	50	42	66	56
LDL cholesterol (mg/dL) (65-139)	126	116	144	256	96	190
Apolipoprotein AI (mg/dL) (119-155)	137	120	130	101	139	130
Apolipoprotein AII (mg/dL) (25.9-35.7)	30.4	24.6	29.9	23.3	33.2	39.6
Apolipoprotein B (mg/dL) (73-109)	101	96	112	168	84	91
Apolipoprotein CII (mg/dL) (1.8-4.6)	2.8	2.6	3.4	2.4	3.8	2.4
Apolipoprotein CIII (mg/dL) (5.8-10.0)	6.8	5.5	6.7	4.6	6.6	5.9
Apolipoprotein E (mg/dL) (2.7-4.3)	3.4	3.5	4.1	3.6	3.1	4.1

W: wild type, M: mutation, APOE: apolipoprotein E, HDL: high-density lipoprotein, LDL: low-density lipoprotein

Discussion

We herein report the first Japanese case of FH caused by a specific mutation in APOE [c.500_502delTCC (p.Leu167del)]. Although this patient did not exhibit any pathogenic variants in so-called “FH genes”, we were able to accurately diagnose him with FH genetically via comprehensive genetic testing, leading to his optimal medical management, as well as cascade and reverse cascade screening.

Genetic testing for FH typically includes LDLR, PCSK9, APOB, and LDLRAP1, the so-called “FH genes.” It is true that the causal associations between these FH genes and the FH phenotype are quite robust. However, there are still substantial numbers of patients clinically diagnosed with FH who register as “negative” on genetic testing for these FH genes. There are several potential explanations for this discrepancy in the literature. First, genetic testing that is currently being used, especially using short-lead next-generation sequencing (NGS), may miss structural genetic variations. In fact, up to 10% of the genetic causes of FH are structural genetic variations in LDLR (12). Therefore, it is important to note how genetic testing is conducted. In the present case, we used short-lead NGS for FH and other associated genes. However, we assessed the presence of such structural genetic variations using software that can identify structural genetic variations via differences in depth, as described elsewhere (13). Accordingly, in the present case, there was little chance of missing such a situation. Second, polygenic FH appears to exist in addition to monogenic FH (3,4). In the present case, we confirmed perfect genotype-phenotype correlations among at least three generations, although we did not assess polygenic risk scores in this family. Third, there are some patients clinically diagnosed with FH who actually have sitosterolemia, due to the similar phenotypes of these diseases. In the present family, we performed genetic testing on ABCG5 and ABCG8 and found no relevant mutations in these genes.

It is important to note that the proband realized the risk of hyper-LDL cholesterolemia only after the genetic diagnosis of FH. In addition, a genetic diagnosis leads to cascade as well as reverse cascade screening. As a result, his father and son were genetically diagnosed with FH, which led to their optimal medical management.

APOE has been described as playing an important role in LDL metabolism (14). Some common and rare genetic variations are associated with elevation of LDL cholesterol, such as APOE4 and APOE7 (15,16). However, the effect sizes of these variations were not sufficiently large to cause FH. This particular in-frame three-base-pair deletion genetic mutation in APOE [c.500_502delTCC (p.Leu167del)], conversely, has been shown to be a cause of FH in different families. To our knowledge, this is the first Japanese family with FH caused by this mutation, strengthening the fact that this is a causal mutation of FH.

The authors state that they have no Conflict of Interest (COI).