Coronary Artery Plaque Regression by a PCSK9 Antibody and Rosuvastatin in Double-heterozygous Familial Hypercholesterolemia with an LDL Receptor Mutation and a PCSK9 V4I Mutation.

The low-density lipoprotein-cholesterol (LDL-C) level of a 38-year-old man diagnosed with acute coronary syndrome was 257 mg/dL. The administration of a proprotein convertase subtilisin-kexin type 9 (PCSK9) antibody in addition to rosuvastatin plus ezetimibe was initiated, reducing his LDL-C level to 37 mg/dL. A genetic analysis revealed both an LDL receptor (LDLR) mutation and a PCSK9 V4I mutation. Nine months after revascularization, intravascular ultrasound revealed plaque regression in the coronary arteries. LDLR/PCSK9 mutation carriers are prone to coronary artery disease. Intensive LDL-C lowering by including PCSK9 antibody was associated with coronary plaque regression, suggesting the expectation of prognosis improvement.

Introduction

Familial hypercholesterolemia (FH) is characterized by marked hypercholesterolemia since birth, and FH patients are known to develop early coronary artery diseases (CADs) (1). Therefore, the early diagnosis of this disease is important for the prognosis.

Evolocumab (AMG-145; Repatha™, Amgen, Thousand Oaks, USA) is a monoclonal antibody that inhibits proprotein convertase subtilisin-kexin type 9 (PCSK9) and lowers the low-density lipoprotein cholesterol (LDL-C) levels (2). The Further Cardiovascular Outcomes Research with PCSK9 Inhibition in Subjects with Elevated Risk (FOURIER) trial recently revealed that the inhibition of PCSK9 with evolocumab combined with statin therapy lowered LDL-C levels and reduced the risk of cardiovascular events (3). Furthermore, the Global Assessment of Plaque Regression with a PCSK9 Antibody as Measured by Intravascular Ultrasound (GLAGOV) Randomized Clinical Trial demonstrated that the addition of evolocumab to statin therapy produced greater LDL-C-lowering effects and atheroma regression than statin monotherapy (4).

The combination of an LDL receptor (LDLR) gene mutation and a PCSK9 V4I mutation in clinically diagnosed FH is a severe state and causes an individual to be prone to CAD (5). We herein report the first case of acute coronary syndrome in a double-heterozygous FH patient (LDLR/PCSK9) who received a PCSK9 antibody associated with coronary plaque regression.

Case Report

A 38-year-old man was admitted to the emergency department complaining of sudden left-sided chest compression. His cardiac risk factors were dyslipidemia, current smoking status and a family history. His father had suffered from myocardial infarction at 40 years of age, and his mother and older sister showed lipid abnormalities. His body height was 176 cm, and his body weight was 75 kg. Dyslipidemia had been noted at 18 years of age. His medications included atorvastatin 10 mg daily, but his adherence was poor.

An electrocardiogram (ECG) in the emergency department showed severe ST-segment elevation in leads II, III, and aVF and depression in leads V1-V6. His echocardiogram results showed hypokinesis in the posterior wall and septum in the left ventricle. All cardiac enzyme levels, including those of troponin-T, were normal. Considering these findings, he was diagnosed with ST elevation myocardial infarction and underwent emergent coronary angiography (CAG) because his symptoms persisted and his cardiac enzyme levels had increased over time. CAG revealed occlusion of the proximal right coronary artery (RCA) and the proximal left anterior descending coronary artery (LAD) (Fig. 1A and B). ECG and echocardiogram results showed that the culprit lesion was in the RCA, as indicated by the presence of hypokinesis in its posterior wall. Therefore, a 3.5×28-mm biolimus-eluting stent (BMX-J™; Terumo, Tokyo, Japan) and a 4.0×33-mm everolimus-eluting stent (Xience Alpine™; Abbott Vascular, Santa Clara, USA) were implanted in the RCA, generating excellent results on both angiography (Fig. 1C) and intravascular ultrasound (IVUS), without any stent edge dissection or incomplete dilatation.

Figure 1.

Coronary angiography. (A) Total occlusion of the mid right coronary artery (arrow). (B) Total occlusion of the proximal left anterior descending coronary artery (arrow). (C) Post-stent deployment, with excellent results.

At the emergency department, the patient's LDL-C, high-density lipoprotein-cholesterol (HDL-C) and triglyceride (TG) levels were 257, 38 and 378 mg/dL, respectively. These levels were measured using the direct method according to the protocol supplied by the manufacturer. This patient exhibited dyslipidemia at a young age and had a family history of dyslipidemia. According to his home doctor, the LDL-C level while untreated was 272 mg/dL. Therefore, the patient was suspected of having FH. In addition to his LDL-C level and family history, Achilles' tendon thickening and the presence of corneal rings were observed. Achilles tendon xanthoma (right 11 mm, left 13 mm) confirmed the diagnosis of FH according to both Japanese (6) and Western (7-9) diagnostic criteria. Lipid management combined with rosuvastatin 20 mg and ezetimibe 10 mg did not achieve the target level of the guidelines (10), so a human anti-PCSK9 monoclonal antibody, evolocumab, was introduced. After 2 months, the patient’s LDL-C, HDL-C and TG levels had improved to 37, 41 and 121 mg/dL, respectively, with rosuvastatin 10 mg daily and evolocumab 140 mg administered percutaneously every 2 weeks.

Detailed serial changes in the lipid parameters are shown in Fig. 2. The time of lipid measurement was just before evolocumab injection. PCSK9 antibody was initiated to achieve the target level of the guidelines. As the target level of the guideline was achieved by the introduction of PCSK9 antibody, the rosuvastatin dose was reduced and ezetimibe was withdrawn. Evolocumab is approved for subcutaneous injection at 140 mg once every 2 weeks or 420 mg once every 4 weeks; given that the effects of these approaches considered equivalent, injecting 140 mg subcutaneously once every 2 weeks is more economical.

Figure 2.

Clinical courses of the LDL cholesterol concentration levels (upper panel) and detailed serial changes in the lipid parameters (lower panel). LDL: low-density lipoprotein

To examine the phenotype of FH, we performed a genetic analysis as described previously (5). The genetic analysis revealed that the patient had an LDLR frameshift mutation (c.1655delT, p.I531TfsX15) and a PCSK9 V4I mutation. Figs. 3 and 4 show the DNA sequence data of exon 11 of LDLR gene and exon 1 of PCSK9 gene, respectively.

Figure 3.

DNA sequence data of exon 11 of LDLR gene. The arrowhead in the upper panel indicates the c.1655delT, p.I531TfsX15 mutation. (･･･ACACT･･･) instead of the normal subject data (･･･ACATCT･･･).

Figure 4.

DNA sequence data of exon 1 of the PCSK9 gene. The arrowhead indicates the c.10G>A, p.V4I mutation [GGGCACC(G/A)TCA] instead of the normal subject data (GGGCACCGTCA).

Three months after the initial percutaneous coronary intervention (PCI), the LAD lesion was successfully stented with a 3.0×33-mm everolimus-eluting stent (Xience Alpine™; Abbott Vascular) at a university hospital. No further stent thrombosis or stent restenosis had occurred at nine months of follow-up.

In intravascular studies, serial changes on IVUS (40-MHz IntrafocusⓇ WR, Terumo, at the acute phase; and 60-MHz AltaViewⓇ, Terumo, at the follow-up phase) demonstrated that the percent atheroma volume (PAV) (4, 11) was decreased at 9 months (41.6%, Fig. 5D) compared with immediately after PCI (45.3%, Fig. 5B). The detailed measurement results are shown in Table. Aggressive LDL-C-lowering therapy with PCSK9 antibody plus rosuvastatin resulted in coronary plaque regression. The patient's other family members were not genotyped.

Figure 5.

(A) Intravascular ultrasound (IVUS) images (arrowhead site in Fig. 1C) immediately after PCI. (B) IVUS images (arrowhead site in Fig. 1C) immediately after PCI. The yellow zones indicate plaque. (C) IVUS images (arrowhead site in Fig. 1C) nine months after PCI. (D) IVUS images (arrowhead site in Fig. 1C) nine months after PCI. The yellow zones indicate plaque. PCI: percutaneous coronary intervention

Table.

Detailed Measurement Results Obtained by Intravascular Ultrasounds (IVUSs).

Status	Lumen volume (mm3)	Vessel volume (mm3)	TAV (mm3)	Length (mm)	PAV (%)		ΔPAV (%)	ΔTAV (mm3)
initial IVUS	145	264	120	10	45.3
9 months IVUS	151	259	108	10	41.6		-3.7	-12

TAV: total atheroma volume, PAV: percent atheroma volume, ΔPAV: differences in PAV between initial and 9 month IVUS, ΔTAV: differences in TAV between initial and 9 month IVUS

The patient provided their consent for the publication of this study. The protocol for the genetic analysis was approved by the Ethics Review Committee of the National Cerebral and Cardiovascular Center (M17-56).

Discussion

The c.1655delT, p.I531TfsX15 mutation present in exon 11 of the LDLR gene detected in this report has already been reported (1). Because this mutation alters the reading frame of the codon, inducing a stop codon in the EGF precursor homology region of mature LDLR, it produces truncated LDLR protein lacking the EGF precursor homology region, the O-linked carbohydrate region, the membrane-spanning domain and the cytoplasmic-tail region. Although no functional assay has been reported for this mutation, the truncated protein is considered to lose its function as a cell surface LDLR protein (12). In addition, this mutation is defined as pathogenic in ClinVar (13). Therefore, the LDLR activity is considered to have been completely lost.

We recently reported that there were no changes in the levels of serum lipids, such as LDL-C, due to the presence or absence of V4I mutation in the PCSK9 gene in FH heterozygotes, but patients with both V4I mutation in the PCSK9 gene and a mutation in the LDLR gene showed a 30% increase in the serum untreated LDL-C level and frequency of coronary artery disease compared with patients with a mutation in the LDLR gene only (5). Therefore, in patients with a mutation in the LDLR gene, the PCSK9 V4I mutant may function as a modifier of the clinical manifestation. In addition, when we prepared a plasmid with a V4I mutation in the PCSK9 gene, introduced it into HEK 293 cells, and compared it with a wild-type (WT) plasmid-introduced strain, we confirmed that PCSK9 V4I mutant was expressed intracellularly and secreted outside the cell, and the expression of LDLR and the uptake of LDL into the cell were also the same as with the PCSK9 WT plasmid (Hori et al. in preparation). Therefore, we believe that the presence of only the V4I mutation in the PCSK9 gene does not affect the LDLR activity. In contrast, in patients with a truncated mutation in LDLR (resulting in half of the typical LDLR activity), overlapping of the V4I mutation in the PCSK9 gene may further reduce the LDLR activity. However, the mechanism has not been clarified.

Arteriosclerotic plaques can regress for a long time (14), according to large-scale clinical studies (15), thus verifying the close relationship between LDL-C levels and the incidence of atherosclerotic diseases and eliciting significant interest in the mechanism underlying this interaction. Furthermore, with IVUS, changes in coronary plaques over time can be observed easily. In addition, with the introduction of statins, which can significantly improve the lipid profile, plaque regression following drug intervention has been confirmed. Recently, with the advent of intravascular endoscopy and optical coherence tomography (OCT), aspects of plaque regression have gradually become apparent. We analyzed coronary plaque progression/regression using IVUS and reported that combination therapy with a statin and ezetimibe resulted in coronary plaque regression after 9 to 12 months of treatment (11).

FH is an autosomal-dominant inherited disorder that is caused by mutations in the LDLR and PCSK9 genes (16). In Japan, one FH homozygote patient is estimated per million, and one FH heterozygote patient is estimated per 500 (17). In 1973, Khachadurian et al. reported on families with a background of genetically dominant hypercholesterolemia and on those with recessive inheritance (18). In 1974, Brown and Goldstein found that genetically dominant hypercholesterolemia was caused by mutations in the LDLR gene, which became known as FH (19). It has become clear, however, that some patients with the FH phenotype do not exhibit mutations in the LDLR gene. A report in 1986 found that a mutation in the apolipoprotein B-100 (APOB) gene, which is a ligand for LDLR, resulted in a decreased binding affinity to LDLR and defective familial apoB-100 (FDB), producing the pathology of FH (20). Two frameshift mutations (the 5-bp insertion at codon 395 in exon 9 and the single-nucleotide deletion at codon 531 in exon 11) were detected in the mutant LDLR genes responsible for FH (12). In the present case, we detected the second of these two mutations by a genetic analysis. In 2003, Abifadel reported that PCSK9 is the third leading causative gene of FH (21). Conversely, autosomal recessive hypercholesterolemia (ARH) has been clarified as a genetic disease of the autosomal recessive genetic form (22, 23), and its pathogenic genes have been reported (24).

LDLR gene abnormalities have been confirmed in 53% of FH patients (5). In Japan, FH due to an APOB gene abnormality has not been described. The frequency of the E32K mutation in the PCSK9 gene has been reported to be approximately 5% in FH cases (5, 25, 26). In Western countries, the incidence of FH with a PCSK9 gene mutation is as low as 0% to 1.5% (27, 28). However, Japan has many cases of FH due to PCSK9 gene mutations. Indeed, in Japan, the V41 mutation of the PCSK9 gene is relatively frequent. It has been reported that the double heterozygote combined with an LDLR gene mutation results in a high LDL-C level (5). We recently reported that double heterozygotes are classified as homozygotes and that evolocumab was more effective in double heterozygote cases than in other homozygote cases in a study of 106 FH homozygote cases (29).

FH results in a higher LDL-C level than non-FH with hypercholesterolemia (30), and a person with an FH-causative gene mutation has a higher risk of CAD than does a person without the mutation, even if they have similar LDL-C levels (31). CAD is believed to develop when the lifetime cumulative LDL-C level exceeds a certain level in FH patients with delayed treatment initiation and in patients with other risk factors who require stronger lipid-lowering agents (7).

Although the effects of plaque regression on other phenotypes, such as ARH, should be investigated, it is more important to note the family history and to focus on physical findings, such as Achilles tendon hypertrophy, in patients with markedly high LDL-C levels.

In a meta-analysis of clinical test results using statin preparations, a linear correlation was reported between the LDL-C levels and plaque regression, and the degree of plaque regression reportedly increased as the LDL-C levels decreased (32, 33). In the present case, using evolocumab, we reduced the LDL-C levels from 257 to 37 mg/dL. Therefore, this regression of plaque might be a natural result.

A limitation of this study was that we were unable to observe the plaque volume and characteristics using OCT. Therefore, the plaque regression that was observed using IVUS may have been reasonable. Another limitation of this study is that plaque regression by statin therapy could not be ruled out. The use of PCSK9 antibody is allowed only in combination with statin therapy, according to the Japanese health insurance system. Finally, the IVUS images from two time points were acquired at different facilities, so our analysis may not have been appropriately conducted.

To our knowledge, this is the first report of a case of reduced LDL-C levels and plaque regression in a double-heterozygous FH patient with an LDLR mutation and a PCSK9 V4I mutation following treatment with PCSK9 antibody and statin.

Author's disclosure of potential Conflicts of Interest (COI).

Kenji Sakamoto: Research funding, Daiichi Sankyo. Koichi Kaikita: Honoraria, Bayer Yakuhin and Daiichi Sankyo; Research funding, Bayer Yakuhin, Daiichi Sankyo, Novartis Pharma and SBI Pharma. Mariko Harada-Shiba: Honoraria, Aegerion, Astellas Amgen, Astellas, Sanofi, Astrazeneca, JCR Pharm, Daiichi-Sankyo, MSD, Phizer, Torii, Shionogi and Tanabe-Mitsubishi; Research funding, Astellas, Kaneka Medics, MSD, Takeda, Asttellas Amgen and Aegerion. Kenichi Tsujita: Honoraria, Amgen Astellas BioPharma, Bayer Yakuhin, Daiichi Sankyo, MSD and Sanofi; Research funding, AstraZeneca, Astellas Pharma, Bayer Yakuhin, Boehringer Ingelheim Japan, Boston Scientific Japan, Chugai Pharmaceutical, Daiichi Sankyo, Eisai, Kowa Pharmaceutical, Mitsubishi Tanabe Pharma, MSD, Pfizer Japan, Sanofi, Shionogi and Takeda Pharmaceutical.