A Family with Familial Hypobetalipoproteinemia Caused by a c.1468C>T in APOB

Abstract

We herein report the first family of Japanese individuals with familial hypobetalipoproteinemia caused by the c.1468C>T mutation in apolipoprotein B (APOB). A 13-year-old boy with extremely low levels of low-density lipoprotein (LDL) cholesterol (24 mg/dL) was referred to our hospital. The patient had no secondary causes of hypobetalipoproteinemia. His father and grandmother also exhibited low LDL cholesterol levels. A genetic analysis confirmed that they all had this variant in APOB (c.1468C>T). None of the patients exhibited atherosclerotic cardiovascular diseases or any other complications associated with low LDL cholesterol levels, including fatty liver, neurocognitive disorders, and cerebral hemorrhaging.

Introduction

Currently, the heritable primary causes of hypobetalipoproteinemia (HBL) are classified into two groups: Class 1, referred to as familial HBL (FHBL) caused by lipoprotein assembly and secretion defects; and Class 2, referred to as FHBL caused by enhanced lipoprotein catabolism (1). Class 1 includes abetalipoproteinemia, FHBL caused by loss-of-function (LOF) variants in apolipoprotein B (APOB) (previously called FHBL1), and chylomicron retention disease caused by LOF variants in secretion-associated Ras-related GTPase 1 B (SAR1B). Class 2 includes familial-combined hypolipidemia caused by LOF variants in angiopoietin-like 3 (ANGPTL3) (previously called FHBL2) and FHBL caused by LOF variants in proprotein convertase subtilisin/kexin type 9 (PCSK9) (previously called FHBL3).

Notably, most of these FHBL-associated genes have emerged as targets for low-density lipoprotein (LDL)-lowering therapies, mimicking these “human knockouts” (2-4). FHBL is mostly caused by protein-truncating variants in APOB (5). However, some missense variants in APOB are also likely to cause FHBL (6,7).

We herein report a family with FHBL caused by a pathogenic variant (c.1468C>T) that has been considered a missense variant (p.Arg490Trp) but has recently been reported to be a splicing variant (p.Arg490Serfs12) in APOB, among whom no disadvantages were observed. This study provides new insights into APOB inhibitors, which are currently associated with fatty liver disease.

Case Report

The proband (III-2) was referred to our hospital for a further examination of his low LDL cholesterol level, which had been identified at a local annual health checkup for the prevention of lifestyle-related diseases performed at his junior high school. The LDL cholesterol level of the proband (III-2) was 24 mg/dL (Fig. 1). The patient did not exhibit any secondary causes of HBL, including hyperthyroidism, bleeding, or any malignancies. Ultrasonography revealed no fatty liver. Through cascade screening, we found that the patient's father and grandmother also had HBL, suggesting a dominant pattern of inheritance (Table). Agarose gel electrophoresis and an ultracentrifugation analysis revealed a typical pattern of HBL in the proband (III-2), whereas a normal pattern was observed in his older sister (III-1) (Fig. 2) (8). None of the family members exhibited fat-soluble vitamin deficiency, atherosclerotic cardiovascular disease, or any other complications typically associated with low LDL cholesterol levels, including fatty liver, neurocognitive disorder, or cerebral hemorrhaging.

Figure 1.

Family tree. Black indicates the mutation status of c.1468C>T in APOB. HDL: high-density lipoprotein, LDL: low-density lipoprotein

Table.

Characteristics of the Study Subjects.

Subject (gender)		I-1 (female)		II-1 (male)		II-2 (female)		III-1 (female)		III-2 (male)
APOB c.1468C>T status		M/W		M/W		W/W		W/W		M/W
Age (yr)		77		49		51		19		13
Total cholesterol (mg/dL) (142-219)		130		122		230		181		84
Triglyceride (mg/dL) (40-149)		95		120		96		42		61
HDL cholesterol (mg/dL) (40-90)		59		50		52		60		45
LDL cholesterol (mg/dL) (65-139)		42		41		137		100		24
Apolipoprotein AI (mg/dL) (119-155)		134		123		121		134		118
Apolipoprotein AII (mg/dL) (25.9-35.7)		31.2		30.2		25.2		40.8		33.5
Apolipoprotein B (mg/dL) (73-109)		40		39		94		82		31
Apolipoprotein CII (mg/dL) (1.8-4.6)		2.8		3.1		2.2		2.4		2.9
Apolipoprotein CIII (mg/dL) (5.8-10.0)		6.8		6.7		2.3		5.9		6.1
Apolipoprotein E (mg/dL) (2.7-4.3)		3.8		2.9		2.4		2.8		3.1
Apolipoprotein E phenotype		E3/E2		E3/E3		E3/E3		E3/E3		E3/E3
CETP (μg/mL)		2.2		2.3		2.0		3.6		2.1
Vitamin A (IU/L) (27.2-102.7)		43.8		50.5		78.9		79.9		44.4
25-OH Vitamin D (ng/mL) (>30)		30.7		42.4		50.5		44.4		31.6
Vitamin E (IU/L) (0.75-1.41)		0.88		1.04		0.96		1.02		0.84
AST (IU/L) (13-30)		24		30		26		17		26
ALT (IU/L) (10-30)		30		24		24		12		14
γ-GTP (IU/L) (13-64)		16		10		33		16		13

Figure 2.

Agarose gel electrophoresis and an ultracentrifugation analysis of the proband and his older sister. (A) The older sister (III-1). (B) The proband (III-2). FC: fecal calprotectin, HDL-C: high-density lipoprotein cholesterol, IDL-C: intermediate-density lipoprotein cholesterol, LDL: low-density lipoprotein cholesterol, PL: phospholipid, TC: total cholesterol, TG: thyroglobulin, VLDL-C: very-low-density lipoprotein cholesterol

Through panel sequencing (9), we identified an extremely rare variant (allele frequency=0.00000796 in gnomAD Exomes) (10) of APOB [(NM_000384.3):c.1468C>T] in the proband (III-2), his father (II-1), and his grandmother (I-1). This variant is also classified as “pathogenic” as an FHBL in ClinVar, and as “likely pathogenic” according to the American College of Medical Genetics and Genomics criteria (11).

Systemic atherosclerosis in the proband and his father was assessed using carotid ultrasonography and brachial-ankle pulse wave velocity. Neither patient had any plaque in their carotid arteries, and their arterial stiffness was within the normal range according to age.

All blood examinations were performed during fasting. The proband's father did not have any siblings, but his grandmother had two younger brothers with unknown phenotypes. Written informed consent was obtained from all participants (or their guardians).

Discussion

Recently, the concept of FHBL has changed, such that it is now classified into two groups: Class 1 and Class 2, as described above (1). LOF variants in APOB are the most common cause of FHBL, resulting in the formation of truncated forms of APOB, which may or may not be secreted into the plasma (12). This situation is sometimes associated with fatty liver, as truncated APOB may not be secreted in the plasma. In accordance with this finding where FHBL is associated with fatty liver, mipomersen, an antisense oligonucleotide for APOB, causes fatty liver (2). Furthermore, several missense variants of APOB have been shown to cause FHBL.

One potential mechanism underlying FHBL is inhibition of APOB lipidation via increased binding to MTTP (13). The mechanism causing FHBL with the current variant (c.1468C>T) was previously attributed to a putative missense mutation (p.Arg490Trp), which would increase its binding to MTTP, thereby prolonging endoplasmic reticulum retention (7). However, a recent study demonstrated that this variant is actually a truncating variant, where the activation of a cryptic donor splice site leads to a premature stop codon (p.Arg490Serfs12) (14). FHBL caused by this particular variant (c.1468C>T) has been described as a major cause of Lebanese FHBL (15), although the present family appears to be the first Japanese family. However, some patients with FHBL caused by this variant exhibit fatty liver, while others do not, and the differences between these patients remain unclear. Accordingly, the accumulation of clinical and genetic data is needed to clarify the detailed mechanisms of pathogenicity of fatty liver associated with FHBL.

In conclusion, a family was diagnosed with FHBL caused by the c.1468C>T mutation in APOB, but an apparently “healthy” phenotype was observed. The accumulation of data obtained from such rare individuals may help elucidate the roles of APOB in humans.

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

Financial Support

This work was supported by a grant from the Ministry of Health, Labour and Welfare of Japan (Sciences Research Grant for Research on Rare and Intractable Diseases).