Dyslipidemia, and specifically elevated low-density lipoprotein (LDL) cholesterol, is one of the most important cardiovascular risk factors. Statins are considered first line therapy for the primary and secondary prevention of cardiovascular disease. However, statins may not be adequate treatment for elevated circulating LDL levels and are ineffective in certain familial hypercholesterolemias. The discovery of proprotein convertase subtilisin/kexin type 9 (PCSK9), a regulatory protein that affects LDL receptors, offers a new alternative for these patients. Moreover, gain-of-function PCSK9 mutations were discovered to be the root cause of familial autosomal dominant hypercholesterolemia. Inhibition of PSCK9 reduces plasma LDL levels, even in patients for whom statins are ineffective or not tolerated. Alirocumab and evolocumab, human monoclonal antibodies that inhibit PCSK9, have been approved to lower LDL levels. While there are drawbacks to these treatments, including adverse events, administration by subcutaneous injection, and high cost, these drugs are indicated for the treatment of atherosclerotic cardiovascular disease and familial hypercholesterolemia as adjunct to diet and maximally tolerated statin therapy. PCSK9 inhibitors may work synergistically with statins to lower LDL. Novel approaches to PCSK9 inhibition are currently in development with the aim of providing safe and effective treatment options to decrease cardiovascular event burden, ideally at lower cost and with oral bioavailability.

Introduction

Discovered in 2003, proprotein convertase subtilisin/kexin type 9 (PCSK9) is a positive regulator of plasma low-density lipoprotein (LDL) [1]. It behaves as a chaperone to increase degradation of the LDL receptor. The primary source of PCSK9 is the liver, but it is also expressed in the small intestine, kidney, and brain. In humans, circulating PCSK9 levels correlate positively with serum total and LDL concentration [2]. The clinical importance of PCSK9 lies in the development of inhibitors for use in the treatment of hypercholesterolemia and dyslipidemia and for high cardiovascular risk persons unable to tolerate statins. PCSK9 inhibitors inactivate the molecule which decreases LDL receptor degradation, leading to increased hepatocyte surface accumulation and consequent enhanced LDL removal from the bloodstream.

PCSK9 processing

The PCSK9 gene is located on chromosome 1p32.3 and contains 12 exons and 11 introns.

PCSK9 is a secreted protein and it circulates in human plasma. It is synthesized in the endoplasmic reticulum as a 72-kDa 692 amino acid preproPCSK9 protein, comprises a signal peptide (amino acids 1–30), a prodomain (amino acids 31–152), a catalytic domain (amino acids 153–404), a hinge region (amino acids 405–452), and a cysteine/histidine-rich domain (amino acids 453–692) (Figure 1A). Then, its signal peptide is cleaved in the endoplasmic reticulum, followed by activation through autocatalytic cleavage at position 152. This intramolecular autocleavage is the only enzyme activity of PCSK9 and it generates two moieties, a 14-kDa propeptide (to amino acid 152) and a glycosylated 63-kDa mature enzyme [3]. The propeptide and enzyme form a tightly bound heterodimer that moves through the Golgi apparatus and is secreted in an inactive state [4]. It then binds to the LDL receptor, leading to endocytosis and degradation of the LDL receptor in the phagolysosome [5]. Interestingly, binding to the LDL receptor does not directly activate PCSK9 as a protease [6]. The protease activity of PCSK9 is limited to its growth and secretion, not LDL receptor degradation. PCSK9 loses protease function since its prodomain remains noncovalently bound to the catalytic domain during the maturation process [7].

Processing of PCSK9 (A) and its primary structure (NCBI access number Q8NBP7) and missense mutations (B)

Figure 1
Processing of PCSK9 (A) and its primary structure (NCBI access number Q8NBP7) and missense mutations (B)

(A) The mature form of pre-pro-PCSK9 contains a signal peptide (SP, colored in green), a prodomain (colored in red), a catalytic domain (colored in blue), and a cysteine/histidine-rich domain (colored in brown). Between the catalytic and cysteine/histidine-rich domains, there is a short hinge domain (colored in black). (B) Naturally occurring mutations—R46L, S127R, D374Y, E670G, and C679X—are indicated as black dots in the primary sequence. In the original reports, R46L was named based on the pro-PCSK9 primary sequence while the others were based on the pre-pro-PCSK9 primary sequence. Shown in (B), it is Arg66 mutated to Leu. We retain the name of R46L to be consistent with the original report.

Figure 1
Processing of PCSK9 (A) and its primary structure (NCBI access number Q8NBP7) and missense mutations (B)

(A) The mature form of pre-pro-PCSK9 contains a signal peptide (SP, colored in green), a prodomain (colored in red), a catalytic domain (colored in blue), and a cysteine/histidine-rich domain (colored in brown). Between the catalytic and cysteine/histidine-rich domains, there is a short hinge domain (colored in black). (B) Naturally occurring mutations—R46L, S127R, D374Y, E670G, and C679X—are indicated as black dots in the primary sequence. In the original reports, R46L was named based on the pro-PCSK9 primary sequence while the others were based on the pre-pro-PCSK9 primary sequence. Shown in (B), it is Arg66 mutated to Leu. We retain the name of R46L to be consistent with the original report.

Mechanism and regulation

PCSK9 expression is primarily regulated at the transcriptional level by sterol-regulatory element-binding protein (SREBP)-2, a membrane-bound transcription factor that regulates multiple genes involved in cholesterol homeostasis in cells, including the LDL receptor [8,9]. A sterol-regulatory element (SRE), which forms the target site for SREBPs, is located in the proximal promoter region approximately 330 bp upstream of the translation start site and functions as an important cis-element regulating PCSK9 transcription. Hepatocyte nuclear factor-1α (HNF1α), a liver-enriched transcription factor, up-regulates PCSK9 by binding to a highly conserved HNF1 binding site in the PCSK9 promoter region [10].

The major function of PCSK9 is degradation of the LDL receptor. The PCSK9 catalytic domain binds to the LDL receptor through the epidermal growth factor (EGF)-A domain of the LDL receptor. Binding between PCSK9 and the LDL receptor is augmented at the acidic pH that exists in the endosome-lysosomal compartment, which inhibits recycling of the LDL receptor to the cell membrane and instead targets the PCSK9–LDLR complex for degradation in the lysosome [11]. PCSK9 complexed with the LDL receptor prevents endocytic recycling of the receptor, leading to lysosomal degradation of both proteins [12]. The C-terminal domain of PCSK9 is essential to induce degradation of the LDL receptor [13]. However, the exact mechanism remains unclear [14]. Studies have revealed a mechanism where direct binding or association between the receptor and protein may be involved shown through coprecipitation of PCSK9 and the LDL receptor after PCSK9 is added exogenously to HepG2 human hepatoma cells [15]. It has been determined that the LDL receptor is not degraded at the cell surface but intracellularly even when clathrin-coated pit- mediated endocytosis is blocked [16].

There are two PCKS9-mediated mechanisms for targeting the LDL receptor for degradation: pre- and postsecretion of PCSK9 (Figure 2). Thus, binding and formation of the PCKS9-LDL receptor complex can occur either intracellularly or at the cell surface. Presecretory, intracellular PCSK9 binds and directs the nascent LDL receptor from the trans-Golgi apparatus to the lysosomes for degradation [17]. Postsecretory PCSK9 can act as a secreted ligand that regulates the LDL receptor by forming a complex with the receptor itself. Extracellular targeting predominates in the liver. In the slower extracellular pathway, PCSK9 travels in the blood until binding to EGF-A domains on hepatic tissue and undergoing clathrin-mediated endocytosis into a lysosome [7]. Internalization and degradation of the LDL receptor on the cell surface via PCSK9 requires a phosphotyrosine binding protein called adaptor protein autosomal recessive hypercholesterolemia (ARH), specifically in lymphocytes and hepatocytes [15–16,18,19]. This adaptor binds to a motif in the cytoplasmic tail of the LDL receptor and directs the clustering of the LDL receptor into clathrin-coated pits. In contrast, when PCSK9 associates with the LDL receptor within the cell, ARH is not needed for degradation to occur [20].

Effect of PCSK9 and its inhibitors on cholesterol metabolism

Figure 2
Effect of PCSK9 and its inhibitors on cholesterol metabolism

Transcription of the PCSK9 gene is regulated by sterols through a sterol-regulatory element (SRE) motif embedded within the gene promoter. PCSK9 gene transcription is also regulated by the hepatocyte nuclear factor (HNF)1α through a highly conserved HNF1 binding site within the PCSK9 gene promoter region. Binding of the transcription factor SREBP-2 and hepatocyte nuclear factor-1α to their respective response elements SRE and HNF1 up-regulates PCSK9 expression (dashed green arrow). The PCSK9 protein is then secreted by exocytosis and circulates in the blood. The extracellular PCSK9 recognizes and binds to the EGF-A domain of the surface LDL receptor, which leads to the internalization and degradation of the LDL receptor with the aid of ARH, as shown in dashed black arrows. On the other hand, the newly synthesized PCSK9 binds and guides the LDL receptor to the lysosome for degradation, which occurs intracellularly as shown in dashed red arrows. In the presence of PCSK9 inhibitors, the extracellular route is blocked; the surface LDL receptor can go through the normal recycling route and return to the plasma membrane, as shown in dashed purple arrows.

Figure 2
Effect of PCSK9 and its inhibitors on cholesterol metabolism

Transcription of the PCSK9 gene is regulated by sterols through a sterol-regulatory element (SRE) motif embedded within the gene promoter. PCSK9 gene transcription is also regulated by the hepatocyte nuclear factor (HNF)1α through a highly conserved HNF1 binding site within the PCSK9 gene promoter region. Binding of the transcription factor SREBP-2 and hepatocyte nuclear factor-1α to their respective response elements SRE and HNF1 up-regulates PCSK9 expression (dashed green arrow). The PCSK9 protein is then secreted by exocytosis and circulates in the blood. The extracellular PCSK9 recognizes and binds to the EGF-A domain of the surface LDL receptor, which leads to the internalization and degradation of the LDL receptor with the aid of ARH, as shown in dashed black arrows. On the other hand, the newly synthesized PCSK9 binds and guides the LDL receptor to the lysosome for degradation, which occurs intracellularly as shown in dashed red arrows. In the presence of PCSK9 inhibitors, the extracellular route is blocked; the surface LDL receptor can go through the normal recycling route and return to the plasma membrane, as shown in dashed purple arrows.

Although it is well established that the major function of PCSK9 is the degradation of LDL receptors, emerging research suggests that the protein may play an additional role in cholesterol metabolism via apolipoprotein(apo)B. ApoB, and specifically apoB100, is a positively charged protein component of LDL, which promotes the binding of LDL to LDL receptors, facilitating receptor-mediated endocytosis of LDL [21]. In large quantities, the positive charge of apoB100 binds via electrostatic interaction to the negatively charged proteoglycans found in the extracellular matrix of the arterial intima, trapping the circulating LDL. Retention of LDL in the extracellular matrix can lead to LDL oxidation which enables LDL internalization by scavenger receptors and foam cell formation. Elevated apoB has been clinically linked to the development and progression of atherosclerosis [22,23]. A study conducted by Sun et al. [24] showed that PCSK9 can bind to amino acid sequences within the N-terminal region of apoB and that overexpression of PCSK9 in cultured mouse primary hepatocytes induced apob100 biosynthesis. Not only did PCSK9 increase apoB100 production, but also reduced degradation of apoB in autophagosome in mouse liver. As such, PCSK9 promotes apoB production and secretion which, in turn, may increase circulating LDL and contribute to hypercholesterolemia and atherosclerosis. In a later study, Sun and colleagues [25] showed that deletion of PCSK9 in the LDL receptor deficient murine model could reduce atherosclerosis through a non-LDL receptor-dependent mechanism. Instead, these mice exhibited reduced apoB100 leading to less atherogenic lipoprotein particles and less endothelial activation.

Human manifestations of PCSK9 mutations

Naturally occurring mutations in the human PCSK9 gene confer gain-of-function or loss-of-function activity on the LDL receptor (Figure 1B). In humans, gain-of-function mutations in PCSK9 cause reduced LDL receptor levels in hepatocytes leading to familial hypercholesterolemia. As proof-of-concept, it has been shown that adenoviral vector-mediated PCSK9 overexpression accelerates the degradation of the LDL receptor in HepG2 human hepatoma cells [26]. Decreasing the number of LDL receptors on the cell membrane reduces LDL cholesterol uptake, which leads to higher LDL cholesterol in the blood. In contrast, loss-of-function mutations reduce LDL cholesterol levels via increased hepatic uptake and correspondingly lower cardiovascular event rates [27].

A number of specific mutations in the PCSK9 gene have been characterized and identified as the cause of familial hypercholesterolemia in carrier families [28]. The D374Y variant is a gain-of function missense mutation that strengthens PCSK9 binding to the LDL receptor leading to accelerated atherosclerosis and premature coronary heart disease [29,30]. These patients have particularly severe dyslipidemia as do those with the S127R [31]. Unlike D374Y, the S127R mutant does not have a markedly increased affinity for the LDL receptor, but exhibits delayed autocatalytic cleavage and secretion into the extracellular space [5,32]. The mechanism by which S127R results in hypercholesterolemia is not fully understood, but the prolonged stay at the cell surface of this variant may increase binding and degradation of the LDL receptor at the cell surface [3].

E670G (rs505151), a common single nucleotide polymorphism in the PCSK9 gene that results in the substitution of glutamate for a glycine residue at position 670 l in the cysteine-rich C-terminal domain in exon 12, is associated with increased LDL cholesterol and atherosclerosis risk, but studies have been somewhat inconsistent, depending on population [33,34].

Humans with the single PCSK9 point mutation R46L have a replacement of arginine with leucine at position 46 in exon 1 and show loss of function with a concomitant lower concentration of circulating LDL of 9–15% and a 28–47% reduction in incidence of coronary heart disease versus noncarriers [35,36]. Patients harboring this variant have reduced plasma levels of PCSK9 [37]. The R46L mutation exhibits slower trafficking from the trans-Golgi network with delayed exit leading to decreased targeting and degradation of the LDL receptor and enhanced LDL clearance [38]. Nuclear magnetic resonance spectroscopic analysis of PCSK9 R46L carriers versus noncarriers revealed lower levels of apoB, LDL, very-low density lipoprotein, and lipoprotein(a) in carriers [35].

The C679X loss-of-function mutation with a stop codon replacing cysteine 679 results in hypocholesterolemia and reduces cardiovascular risk by a dramatic 88% [37,39]. The C679X form found most commonly in African Americans and 100-fold less frequently in European Americans leads to a truncated protein missing 14 amino acids at the C terminal end and instead of being secreted, this protein is retained in the endoplasmic reticulum [37,40].

Current medications: alirocumab and evolocumab

Currently there are two FDA approved drugs on the market: alirocumab (Praluent®; also known as SAR236553/REGN727; Sanofi/Regeneron) and evolocumab, (Repatha®; Amgen), both fully human monoclonal antibodies to PCSK9 that were approved in 2015 (Table 1) [41]. By inhibiting the interaction between PCSK9 and the LDL receptor, these monoclonal antibodies neutralize PCSK9, leading to an increase in the number of LDL receptors and, ultimately, enhancing uptake of LDL particles (Figure 2). Alirocumab is approved as an adjunct to diet in persons with hypercholesterolemia who are already on maximally tolerated statin therapy and require further lowering of LDL [42]. Alirocumab significantly reduces LDL by up to 60% both as monotherapy and when combined with other lipid-lowering drugs in cases of heterozygous familial hypercholesterolemia and in dyslipidemic individuals with nonfamilial elevated cholesterol [43]. When 75 mg of alirocumab was injected subcutaneously into healthy subjects age 18–45 years with LDL exceeding 95 mg/dl not on other lipid-lowering therapies, a near complete loss of free PCSK9 that was unbound to antibody was achieved between day 3 and 4 [44]. At day 14 and 15, LDL levels reached maximal reduction [44,45]. The median half-life at steady state is 17–20 days in patients receiving alirocumab by subcutaneous injection (75 or 150 mg every 2 weeks) and this is reduced to 12 days when administered with a statin [46,47].

Table 1
PCSK9 inhibitors are used to reduce LDL cholesterol levels
DeveloperGeneric nameTrade nameOther nameChemical featuresRoute of administrationReferences
Sanofi/ Regeneron Alirocumab Praluent SAR236553; REGN727 Human monoclonal antibody to PCSK9 Subcutaneous injection Reducing CV mortality by 48% up to 80 weeks [46,47
Amgen Evolocumab Repatha AMG145 Human monoclonal antibody to PCSK9 Subcutaneous injection Reducing CV mortality by 21–27% in up to 26 months [66
Pfizer Bococizumab  PF-04950615 Partially mouse monoclonal antibody to PCSK9 Subcutaneous injection Development halted at phase 3 due to immunogenicity, low efficacy, and injection site reactions [118,119
The Medicines Company Inclisiran – ALN-PCSSC siRNA for hepatocyte-specific PCSK9 Subcutaneous injection Currently in phase 2 and phase 3 trials [121] (https://clinicaltrials.gov/ct2/show/NCT03060577; https://clinicaltrials.gov/ct2/show/NCT03400800
Santaris Pharma A/S – – SPC5001 Antisense oligonucleotide for PCSK9 mRNA Subcutaneous injection Successful in nonhuman primate studies; human trials halted due to renal tubular toxicity [123–125
Affiris AG – – AT04A Virus-like vaccine for PCSK9 Subcutaneous injection Successful in animal studies; currently in phase 1 [126,127] (https://clinicaltrials.gov/ct2/show/NCT02508896
Shifa Biomedical Corporation – – – Small molecule Oral administration Currently in the discovery stage [128–131
DeveloperGeneric nameTrade nameOther nameChemical featuresRoute of administrationReferences
Sanofi/ Regeneron Alirocumab Praluent SAR236553; REGN727 Human monoclonal antibody to PCSK9 Subcutaneous injection Reducing CV mortality by 48% up to 80 weeks [46,47
Amgen Evolocumab Repatha AMG145 Human monoclonal antibody to PCSK9 Subcutaneous injection Reducing CV mortality by 21–27% in up to 26 months [66
Pfizer Bococizumab  PF-04950615 Partially mouse monoclonal antibody to PCSK9 Subcutaneous injection Development halted at phase 3 due to immunogenicity, low efficacy, and injection site reactions [118,119
The Medicines Company Inclisiran – ALN-PCSSC siRNA for hepatocyte-specific PCSK9 Subcutaneous injection Currently in phase 2 and phase 3 trials [121] (https://clinicaltrials.gov/ct2/show/NCT03060577; https://clinicaltrials.gov/ct2/show/NCT03400800
Santaris Pharma A/S – – SPC5001 Antisense oligonucleotide for PCSK9 mRNA Subcutaneous injection Successful in nonhuman primate studies; human trials halted due to renal tubular toxicity [123–125
Affiris AG – – AT04A Virus-like vaccine for PCSK9 Subcutaneous injection Successful in animal studies; currently in phase 1 [126,127] (https://clinicaltrials.gov/ct2/show/NCT02508896
Shifa Biomedical Corporation – – – Small molecule Oral administration Currently in the discovery stage [128–131

Two FDA approved PCSK9 inhibitors, alirocrumab and evolocumbab, are human monoclonal antibodies to PCSK9.

Several experimental drugs, including those with novel chemical features, are in varying stages of drug discovery and development;

CV, cardiovascular.

Evolocumab is also administered subcutaneously. When given with statins or alone, it is shown to reduce high LDL cholesterol levels from 54% to 80%, apo-B100 from 31% to 61%, and lipoprotein (a) from 12% to 36%, in a dose-dependent manner [48,49].

The Monoclonal Antibody Against PCSK9 to Reduce Elevated LDL-C in Subjects Currently Not Receiving Drug Therapy for Easing Lipid Levels-2 (MENDEL-2) trial compared evolocumab with placebo or ezetimibe, a potent inhibitor of intestinal cholesterol absorption, in male and female subjects age 18–80 years with primary hypercholesterolemia not taking statins [50]. Both drugs had minimal adverse effects. For alirocumab, injection site reactions, myalgia, neurocognitive events, and ophthalmologic events were more common when compared with placebo groups [51] while for evolocumab, nonspecific adverse events such as arthralgia, headache, limb pain, fatigue, and neurocognitive events were reported more frequently than in placebo groups [52]. Not surprisingly, both drugs are contraindicated in patients who have shown hypersensitivity reactions [53,54].

Both drugs have varying indications of use based on FDA approval. Alirocumab is indicated as an adjunct to diet and maximally tolerated statin therapy for the treatment of adults with heterozygous familial hypercholesterolemia or clinical atherosclerotic cardiovascular disease, who require additional lowering of LDL cholesterol [53]. Evolocumab is approved for use in addition to diet and maximally tolerated statin therapy in adult patients with heterozygous familial hypercholesterolemia, homozygous familial hypercholesterolemia, or clinical atherosclerotic cardiovascular disease, such as heart attacks or strokes, who require additional lowering of LDL cholesterol [54]. In order for these drugs to be effective, the patient must have at least some LDL receptor function and therefore, they are ineffective in cases of homozygous null LDL receptor gene.

There is the possibility for expansion of the current indications for PCSK9 inhibitors. A study conducted by 34 clinicians/scientists defined three groups who may be considered for therapy: 1(a). Adults with established cardiovascular disease and LDL cholesterol ≥100 mg/dl while on lifestyle modifications and maximally tolerated hypolipidemic treatment, i.e. high-intensity statin + ezetimibe; 1(b). adults with diabetes and established cardiovascular disease or chronic kidney disease or target organ damage and LDL cholesterol ≥100 mg/dl while on lifestyle modifications and maximally tolerated hypolipidemic treatment, i.e. high-intensity statin + ezetimibe. 2. Adults with familial hypercholesterolemia (FH) without established cardiovascular disease and LDL cholesterol ≥130 mg/dl while on lifestyle modifications and maximally tolerated hypolipidemic treatment, i.e. high-intensity statin + ezetimibe (evolocumab is also indicated in children above 12 years with homozygous FH). 3. Adults at high or very high cardiovascular risk who are statin intolerant and have an LDL cholesterol ≥100 and ≥130 mg/dl respectively, while on any tolerated hypolipidemic treatment [55].

Efficacy of PCSK9 inhibitors

Although PCSK9 inhibitors lower LDL, their effectiveness in reducing cardiovascular mortality and overall mortality are more meaningful measures of their value in patient care. A number of clinical studies have evaluated these critical endpoints [56].

In the Further Cardiovascular Outcomes Research with PCSK9 Inhibition in Subjects with Elevated Risk (FOURIER) study (NCT01764633), a randomized, double-blind, placebo-controlled trial of evolocumab on a background of statin therapy in more than 27,000 persons age 40–85 years with established atherosclerotic cardiovascular disease, evolocumab reduced the rate of cardiovascular events by 21 to 27% over a mean follow-up duration of 26 months. However, no effect was detected on cardiovascular mortality during the short follow-up period. The adverse event rate did not differ between study arms [57].

Similarly, in the global phase 3 ODYSSEY LONG TERM trial, a randomized, double-blind, placebo-controlled trial of alirocumab on 2341 patients with coronary heart disease and/or hypercholesterolemia whilst on maximally tolerated statin therapy, the rate of major adverse cardiovascular events was 48% lower in the alirocumab group than that of the placebo controlled group during the 80-week follow-up. Adverse cardiovascular events were defined as a composite of death from nonfatal myocardial infarction, ischemic stroke, and coronary heart conditions requiring hospitalization. The adverse event rate did not differ between study arms when hospitalization from congestive heart failure and coronary revascularization were considered. Effects on individual mortality were not found among subjects in both groups [58].

Using LDL cholesterol as the main indicator of cardiovascular risk fails to account for other lipid molecules and their role in cardiovascular events. In addition to hypercholesterolemia, hypertriglyceridemia which includes measurement of LDL, high-density lipoprotein (HDL), non-HDL and apo-B lipoproteins, also contributes to cardiovascular mortality risk [59–62]. Non-HDL levels are considered of importance as a therapeutic target alongside LDL levels by the National Lipid Association [63], while the American College of Cardiology Consensus Decision Pathway have implemented targeting non-HDL cholesterol levels in high risk patients [64]. Non-HDL cholesterol includes all studied atheromatous lipid particles and is computed as the difference between total cholesterol and HDL cholesterol [65]. In a cohort study with 5794 patients, LDL and non-HDL cholesterol levels predicted cardiovascular events in patients with triglyceride levels <200 mg/dl, but LDL cholesterol did not predict cardiovascular risk in patients with TG >200 mg/dl, while non-HDL cholesterol remained a good risk indicator [66].

The effects of alirocumab on non-HDL and apo-B levels have shown significant impact in an analysis of ten pooled phase 3 ODYSSEY trials, with eight of the studies including alirocumab given to patients utilizing statins and two studies that included patients administered alirocumab without concomitant statin use. In these studies, a total of 4983 patients with a broad range of hypercholesterolemia were split into four groups according to alirocumab dosage, concomitant statin therapy use and a control group utilizing either placebos or ezetimibe. In all four pools, alirocumab allowed between 72 and 92% of patients to achieve the target non-HDL level of <100 mg/dl over the course of 24 weeks [67]. In a poststudy analysis of the phase 3 trials, major cardiovascular events including cardiovascular mortality, nonfatal myocardial infarction, ischemic stroke, and unstable angina requiring hospitalization occurred in 1.8% in the alirocumab group versus 2.6% in the placebo group and 2.8% in the alirocumab group versus 1.5% in the ezetimibe group [68]. These studies show that PCSK9 inhibitors may be an effective option in reducing the risk of cardiovascular events in conditions characterized by imbalances among LDL cholesterol, non-HDL cholesterol and apo-B levels, such as those found in obese patients, diabetics and patients with metabolic syndrome.

A meta-analysis that consisted of 10,159 patients with hypercholesterolemia from 24 various phase 2 and 3 randomized, controlled trials compared the results of all-cause and cardiovascular mortality in PCSK9 inhibitor treatment groups with non-PCSK9 treatment groups. PCSK9 treatments included varying dosages of evolocumab and alirocumab while non-PCSK9 treatments included placebos, ezetimibe, and statin therapy. While there was a statistically significant reduction in all-cause mortality in the PCSK9 inhibitor treatment groups (0.31% mortality rate) versus the non-PCSK9 inhibitor treatment groups (0.53% mortality rate), the reduction in cardiovascular mortality in the PCSK9 inhibitor treatment groups (0.19% mortality rate) versus the non-PCSK9 inhibitor treatment groups (0.33% mortality rate) was deemed statistically insignificant. Data on effects of alirocumab and evolocumab on myocardial infarction and unstable angina were also analyzed from 10 of the 24 randomized, controlled trials. From a total of 5195 patients, there was a statistically significant reduction in the PCSK9 inhibitor treatment groups (0.58% MI rate) versus the non-PCSK9 inhibitor treatment groups (1.00% MI rate). Analysis on unstable angina did not statistically differ among the two study arms [69]. Although showing promising reductions in mortality, the drawback of these results is based on their derivation from study data as opposed to patient trial data.

In March 2018, results were released for the ongoing ODYSSEY OUTCOMES trial which tested the hypothesis that alirocumab reduces cardiovascular morbidity and, for the first time, mortality in patients with acute coronary syndrome and who have high atherogenic lipoprotein levels despite therapy with maximally tolerated statins. A total of 18,924 patients from 57 countries who had acute coronary syndrome within the last 12 months and enduring LDL levels at or above 70 mg/dl and non-HDL levels at or above 100 mg/dl despite treatment with intensive statins were randomized into a placebo group and a group receiving 75 or 150 mg biweekly subcutaneous injection of alirocumab. Treatment follow-up after 2.8 years showed that the primary efficacy outcome of major adverse cardiac events including myocardial infarction, coronary heart disease, unstable angina, or ischemic stroke requiring hospitalization was 9.5% in the alirocumab group as compared with 11.1% in the placebo group with significant reductions in the alirocumab group versus placebo for each component of major adverse cardiac events.

Secondary efficacy endpoints showed all cause-death to be significantly lower by 15% in the alirocumab group at 3.5% versus 4.1% in the placebo group. However, there was no significant difference in coronary heart disease deaths and cardiovascular deaths. There was a significant reduction in all-cause mortality which may be an even greater preserver of life in the long term, but the lack of significance for coronary heart disease deaths and cardiovascular deaths may need to be looked into to validate overall efficacy. All endpoints were reduced in patients with baseline levels of LDL of 100 mg/dl or higher; alirocumab reduced major adverse cardiac events by 24% and all-cause mortality by 29%. This demonstrates that PCSK9 inhibitors may be of greatest clinical benefit for this population. Safety evaluation showed only minor local injection site reactions; the occurrence was greater in the alirocumab than the placebo group (3.1% versus 2.1%). The mortality outcomes of this trial along with the outcomes of the previous FOURIER trial which showed reductions in both LDL levels and cardiovascular events in patients administered evolocumab are promising for protecting life in high risk patients [70].

It should be emphasized that only the preliminary results are available and the very modest clinical impact relative to the high cost of alirocumab presents a difficult decision as to whether the expense is justified and whether these drugs might be reserved for specific subgroups shown to achieve the most benefit. Much more data are needed to resolve this issue. Patient trials have yet to show definitive effects of PCSK9 inhibitors on cardiovascular mortality, which if proven would greatly increase the influence of the PCSK9 inhibitors in the medical realm.

PCSK9 inhibitors versus other nonstatin lipid lowering strategies

When statin treatment is poorly tolerated or inadequate to control serum lipids, alternative strategies may be employed. In the past, possibilities included the addition of fenofibrate, ezetimibe, niacin, or omega 3 fatty acids. However, niacin, fibrates, or both are no longer recommended by the American College of Cardiology and the U.S. Food and Drug Administration has withdrawn approval for coadministration of extended release niacin and delayed release fibrates in combination with a statin [71]. This withdrawal was prompted by the Atherothrombosis Intervention in Metabolic Syndrome with Low HDL/High Triglycerides: Impact on Global Health Outcomes (AIM-HIGH) and Heart Protection Study 2: Treatment of HDL to Reduce the Incidence of Vascular Events (HPS2-THRIVE) study outcomes. AIM-HIGH found no additional clinical benefit in over 3,000 statin-treated patients with stable atherosclerotic cardiovascular disease, low HDL and high triglycerides given extended release niacin over 36 months [72]. HPS2-THRIVE investigated the combination of extended‐release niacin and laropiprant (to reduce niacin-induced flushing) versus placebo on the primary outcome of major vascular event, comprising nonfatal myocardial infarction, death from coronary causes, stroke, and arterial revascularization in 25,673 high-risk patients with previous cardiovascular disease. The extended‐release niacin‐laropiprant group attained modest LDL lowering and higher HDL, but there was no difference in incidence of major vascular events and the extended‐release niacin‐laropiprant group had a higher incidence of adverse events, including gastrointestinal bleeding, infections, musculoskeletal events, and skin- and diabetes-related events [73,74].

The association between lowering LDL cholesterol and cardiovascular risk reduction among statin and nonstatin therapies including PCSK9 inhibitors was explored in a systemic review and meta-analysis from 49 randomized clinical trials that lasted at least 6 months and consisted of 312,175 participants [75]. Four single interventions among control and noncontrol groups were tested: statins, nonstatin therapies that decrease LDL cholesterol via decreasing intrahepatic cholesterol leading to LDL receptor up-regulation (diet, bile-sequestrants, ezetimibe etc.), therapies that do not up-regulate LDL receptor expression (fibrates, niacin etc.), and PCSK9 inhibitors which recycle more LDL receptors to the cell membrane and thus increase LDL clearance [76]. Relative risk (RR) of major vascular events defined as a composite of cardiovascular death, acute myocardial infarction, or other acute coronary pathology was measured per each 1-mmol/l reduction in LDL cholesterol levels. PCSK9 inhibitors exhibited the highest risk reduction in major vascular events with a RR of 0.49 (51% relative reduction) as compared to statins with an RR of 0.77 (23% relative reduction), interventions that up-regulate LDL receptor expression with a RR of 0.75 (25% relative reduction), and therapies that do not increase LDL receptor expression with the lowest risk reductions. Limitations to this analysis were that the major vascular events measured were not the same for each trial and trial level data was assessed as opposed to patient level data, which would take into account patients’ baseline cardiovascular risk and give insight into absolute risk reduction in patients. Furthermore, the majority of the trials (25 of 49) tested statins while only two trials tested PCSK9 inhibitors.

ODYSSEYDM-DYSLIIDEMIA, a randomized multicenter study of persons with type 2 diabetes and mixed dyslipidemia needing treatment beyond statins to control non-HDL cholesterol, compared alirocumab to usual care options: fenofibrate, ezetimibe, omega 3 fatty acid formulation, nicotinic acid, or no additional therapy. After 24 weeks, alirocumab was superior to usual care in reducing non-HDL cholesterol, ApoB, Lp(a) total cholesterol, and LDL. Alirocumab did not demonstrate an advantage in lowering triglycerides. The study was of short duration and not designed to determine whether alirocumab would yield better clinical outcomes [77].

The data available for decision-making regarding use of PCSK9 inhibitors versus alternatives are incomplete. Thus, further studies are warranted to give more definitive guidance as to which drug regimens are best for patients based on patient clinical characteristics and risk factors.

PCSK9 further clinical uses

PCSK9 inhibition may be efficacious in reducing cardiovascular risk in conditions other than hypercholesterolemia. For example, it may be a useful treatment for patients with elevated lipoprotein [Lp] (a) [78]. Lp(a) is a particle with composition similar to LDL, but containing one molecule of apo B100 and an additional protein, apo (a), attached via a disulfide bond [79,80]. High Lp(a) is a cardiovascular risk factor in multiple diseases including calcific aortic valve stenosis [81] and myocardial infarction [82,83].

While statins appear to have a neutral effect on Lp(a), a study of human carriers of the PCSK9 R46L loss-of-function mutation found reduced Lp(a) in these subjects accompanied by reduced risk of aortic valve stenosis and MI [84]. This suggests the use of PCSK9 inhibitors as a therapeutic approach to aortic stenosis. A phase 2 clinical study of 631 patients with hypercholesterolemia receiving statin therapy was conducted in which subjects were randomized into three different dosing groups of AMG145, a fully human, monoclonal, immunoglobulin G2 antibody to PCSK9, versus a placebo group. Comparatively, AMG145 showed greater reduction of Lp(a) for all three dosing groups versus the placebo. Approximately 70, 105, and 140 mg given every 2 weeks reduced Lp(a) at 12 weeks by 18%, 32%, and 32%, respectively while 280, 350, and 420 mg given every 4 weeks reduced Lp(a) by 18%, 23%, and 23%, respectively suggesting an additional benefit outside of LDL cholesterol reduction with the use of PCSK9 inhibition as an add-on therapy with statins. Further study was suggested on long-term clinical outcomes and safety for reducing LDL cholesterol [85].

Peripheral Artery Disease (PAD) indicates vascular obstructive conditions that are usually associated with concomitant coronary and cerebrovascular diseases [86,87]. PAD puts patients at an increased risk for cardiovascular morbidity and mortality. PAD can also be referred to as atherosclerotic occlusive disease of the extremities and includes defining symptoms of intermittent claudication, which is leg discomfort/dysfunction from lack of blood supply to the lower extremities and an ankle brachial index (ABI) <0.90 [86]. Patients with PAD suffer morbidity from major adverse limb events ranging from acute limb ischemia to limb amputation [87]. PCSK9 inhibitors have been shown to reduce cardiovascular events in the FOURIER study, suggesting their use in treating PAD. The FOURIER trial, a 2.2-year randomized trial of evolocumab versus placebo, followed 27,564 patients on statin therapy with atherosclerotic conditions. A subgroup from the trial consisting of 3642 patients had PAD based on eligibility criteria of intermittent lower limb claudication, an ankle brachial index <0.85, a history of a peripheral artery revascularization procedure, or atherosclerotic disease associated amputation.

In comparison with placebo treated patients, those treated with evolocumab showed greater reduction in major cardiovascular events, or the composite of cardiovascular death, MI, stroke, hospitalization for unstable angina, or coronary revascularization by 21% and reduced the composite of cardiovascular death, MI, or stroke by 27%. Evolocumab also reduced major adverse limb events by 42%; reduced levels of LDL-C down to 10 mg/dl from evolocumab treatment linearly correlated with a significantly lower statistical risk of major adverse limb events such as acute limb ischemia and limb amputation. This study demonstrated that PCSK9 inhibitors may serve as beneficial add-on options to reduce major adverse cardiovascular events in patients with PAD. The FOURIER trial was one of the few studies that explored the connection between lowered LDL levels and decreased major adverse limb events in PAD. Further long-term non-subgroup studies and nonrandomized data collection were suggested in order to evaluate efficacy and safety results [87].

Statins and PCSK9 inhibitors

Since the introduction of the lipid hypothesis which suggests decreasing serum cholesterol and specifically LDL significantly reduces coronary heart disease [88], scientists and physicians have been pursuing new ways to reduce blood cholesterol. 3-Hydroxy-3-methylglutarylcoenzyme A (HMG-CoA) reductase inhibitors or statins are one of the most frequently prescribed medications worldwide and are used for lipid reduction in patients with hyperlipidemia and cardiovascular disease [89]. They are generally very well tolerated, but do have dose-dependent adverse effects including myositis and myalgias which develop in 10–15% of patients [89]. Less common adverse effects include hepatotoxicity, peripheral neuropathy, impaired myocardial contractility, autoimmune diseases [89], and drug-induced liver injury [90]. Patients on high intensity statin plans tend to discontinue treatment after their first refill due to the adverse effects, specifically myopathy [91]. The use of statins in elderly persons above age 75–80 years is controversial and may not be of benefit [92,93]. Traditionally, physicians have prescribed statin monotherapy for cardiovascular events as research failed to demonstrate that the addition of other lipid modifying medications would result in better outcomes. However, the use of PCSK9 inhibitors in combination with statins is now seen with increasing frequency [94]. Many times, patients may exhibit reduced LDL-C with statins, but fail to reach the desired LDL-C levels. A multinational survey showed that 70% of very-high-risk patients receiving statin monotherapy in community practices did not reach the LDL-C level of <1.8 mmol/l (<70 mg/dl) [95].

Statins reduce LDL in serum and up-regulate LDL receptor expression by inhibiting HMG-CoA reductase, the rate-limiting enzyme in cholesterol biosynthesis [96,97]. However, this mechanism also leads to the increased expression of PCSK9 [98]. Thus, the presence of PCSK9 will decrease the overall response to statin therapy, suggesting the use of PCSK9 inhibitor add-on therapy with statins to better treat hyperlipidemia and cardiovascular disease [99–101].

Statin intolerance

The inability to tolerate statin therapy mainly due to muscle pain is called statin intolerance, which is a common clinical concern affecting adherence and benefit with statin use [102,103]. A phase 2, 12-week, randomized, double-blind, placebo- and ezetimibe-controlled, dose-ranging study was conducted to test the efficacy of evolocumab in patients with statin intolerance. The 160 patients were all intolerant to at least one statin and were randomized to evolocumab alone at 280, 350 or 420 mg, evolocumab 420 mg combined with ezetimibe 10 mg or ezetimibe 10 mg with placebo. At week 12, mean changes in LDL cholesterol levels from baseline were −67 mg/dl for the evolocumab 280 mg group; −70 mg/dl for the evolocumab 350 mg group; −91 mg/dl for the evolocumab 420 mg group; and −110 mg/dl for the 420 mg evolocumab + 10 mg ezetimibe group compared with −14 mg/dl for the placebo/ezetimibe group. Thus, subcutaneous injection of evolocumab significantly reduced LDL levels suggesting potential use of evolocumab for statin intolerant patients. Adverse effects were seen, the most common being myalgia occurring in 19 patients as well as 4 other serious adverse events. Overall, short-term tolerability was consistent with other studies of PCSK9 inhibitor antibodies [104].

Further study of evolocumab in statin intolerant hypercholesterolemic subjects was conducted within the GAUSS-2 (Goal Achievement after Utilizing an Anti-PCSK9 Antibody in Statin Intolerant Subjects) trial, a 12-week, double-blind randomized study that compared reduction of LDL by evolocumab and ezetimibe in 307 patients [105]. Evolocumab reduced LDL from baseline by 53% to 56% versus 37% to 39% reduction with ezetimibe. Patients did encounter muscle-related side effects. However, only 12% of evolocumab-treated patients compared with 23% of ezetimibe-treated patients presented with muscle symptoms. PCSK9 inhibitors show promise as therapy for statin-intolerant patients compared with currently prescribed medication for a population with largely unmet clinical need and for high-risk patients with elevated cholesterol who are statin intolerant [106].

Current studies on the safety profile of PCSK9 inhibitors in relation to statin and other nonstatin LDL cholesterol lowering therapies point favorably to PCSK9 inhibitor use, yet are limited in number. The GAUSS-3 (Goal Achievement After Utilizing an Anti-PCSK9 Antibody in Statin Intolerant Subjects 3) study was a two-stage randomized clinical trial which consisted of 500 subjects intolerant to statins due to adverse related events. All subjects had a washout phase, consisting of an initial 4-week period without any treatment for LDL cholesterol. Then, in the first stage, Phase A, patients were randomized in a 1:1 ratio to receive either atorvastatin (20 mg/daily) or placebo for 10 weeks followed by a 2-week washout period, then crossover for another 10 weeks of the alternate regimen. A total of 209 out of 492 or 42.6% of patients reported muscle-related adverse effects with atorvastatin therapy whilst 26.5% of patients experienced these effects with placebo [107]. These results showed that muscle-related adverse effects cannot be solely related to statins. A total of 218 patients who were intolerant to statins continued on to Phase B, which consisted of a 24-week randomized trial testing nonstatin therapy; 73 patients received ezetimibe and subcutaneous placebo and 145 patients received evolocumab plus oral placebo. Results showed 28.8% of patients experienced muscle-related adverse effects with ezetimibe therapy versus 20.7% in the evolocumab group. Phase B showed that evolocumab may be of great benefit for patients experiencing muscle-related adverse effects from statin use [107]. Since this was such a small subgroup of patients, further clinical trials are needed with a larger sample size and direct comparison of efficacy of evolocumab versus ezetimibe in statin intolerant patients.

Disadvantages of PCSK9 inhibitors

Both FDA approved PCSK9 inhibitors, alirocumab and evolocumab, display similar adverse effects that have been documented in many clinical trials. Patient data collected from various studies evaluating the efficacy of alirocumab in a total of 2476 patients showed nasopharyingitis, cough, upper respiratory tract infections, musculoskeletal pain, injection site reaction, diarrhea, and myalgia as some of the most common side effects. Alirocumab cessation in many trials was due to allergic reactions and raised liver enzymes [108]. Similarly, data combined from 5416 patients enrolled in clinical trials that evaluated evolocumab showed the most common adverse effects to be nasopharynigitis, upper respiratory tract infections, arthralgia and nausea. Cardiac effects were also noted in 2.4% of patients, and included heart palpitations, angina pectoris and arrhythmias [108,109]. Liver enzymes, alanine aminotransferase and aspartate transferase, were elevated in some patients administered evolocumab [110]. Both alirocumab and evolocumab are contraindicated in patients with hypersensitivity reactions including hypersensitivity vasculitis and extreme drug-induced allergic reactions [108].

Neurocognitive effects including delirium, dementia and amnesia-like symptoms have been reported in clinical trials using both alirocumab and evolucumab [108,110]. Meta-analysis done on a total of 11 studies consisting of 10,656 patients, included data from the ODYSSEY LONG TERM (Long-Term Safety and Tolerability of Alirocumab in High Cardiovascular Risk Patients With Hypercholesterolemia Not Adequately Controlled With Their Lipid Modifying Therapy) [58,110] and OSLER (Open-Label Study of 12 Early Phase 2–3 Trials) [110,111]. ODYSSEY LONG TERM, a Phase 3 clinical trial, evaluated alirocumab efficacy while OSLER was an extended study of evolocumab efficacy and included OSLER-1 and OSLER-2 for patients in phase 2 and phase 3 trials respectively [110]. Data of the aforementioned studies showed a 0.8% incidence of neurocognitive events such as delirium, attention disturbances, dementia, and amnesia in patients administered PCSK9 inhibitors and a 0.5% incidence of neurocognitive events in patients not taking PCSK9 inhibitors. Meta-analysis showed no difference in neurocognitive incidence between the two groups [110]. However, many of these symptoms were noted without any cognitive assessments administered and without information on the baseline cognitive status of patients [108,110]. Studies dedicated to neurocognitive effects of PCKS9 inhibitors, such as the EBBINGHAUS study, have shown no significant difference in cognitive function in patients administered evolocumab or placebo as well as statin therapy over a 19-month period [112]. Long-term trials on this topic will give more insight into these concerning findings.

The clinical lab trials conducted on PCSK9 inhibitors may show some promising results, but there are issues regarding practical use of these drugs in the real world, specifically cost and frequency of administration. The cost of PCSK9 inhibitors is over 100 times higher than the widely used statins [113]. It should be noted, however, that the budget impact of PCSK9 inhibitors when added to statin therapy overall and specifically to reduce cardiovascular events when compared with other lipid lowering biologics is small [113,114]. Although costs remain substantial in the near future, the long-term cost will gradually decrease over time if generic versions become available. Furthermore, both FDA-approved PCSK9 inhibitors are administered via subcutaneous injections. Recommended dosages for evolocumab include subcutaneous injections of 120 mg every 2 weeks or 420 mg monthly for patients with dyslipidemia and a 420-mg subcutaneous injection for patients with homozygous familial hypercholesterolemia [115,116]. Recommended starting dosage for alirocumab is 75 mg subcutaneously every 2 weeks [117]. The frequency of administration and route via injection not only increases costs, but also hinders some patients from remaining complaint. Oral versions of PCSK9 inhibitors are currently under development and may be a way to decrease costs and may be a more tolerable option for patients [130].

New strategies to inhibit PCSK9

Currently, the only two FDA approved PCSK9 inhibitors are monoclonal antibodies. A partially mouse monoclonal antibody to PCSK9, bococizumab, was in phase 3 trials when Pfizer discontinued studies due to immunogenicity, low efficacy, and injection site reactions [118,119]. Overall, monoclonal antibodies currently available have shortcomings including adverse events, need for infusion, and high cost, leaving the door open for new strategies of PCSK9 inhibition.

Alternative PCSK9 inhibitors are at various stages of the drug development process (Table 1). They include small interfering RNA (siRNA), vaccines, antisense oligonucleotides, and small molecule inhibitors [120]. Among these, siRNAs have had the most success with PCSK9 inhibition. The appeal of siRNAs is that they are more selective than monoclonal antibodies, which may cut down on off-target adverse events compared with the current PCSK9 inhibitory therapies. siRNAs have successfully silenced hepatocyte-specific PCSK9 with few adverse events in the phase 2 ORION-1 clinical trial. With this chemically synthesized siRNA, called inclisiran or ALN-PCSSC, the hepatocyte specificity is expected to minimize the off-target effects. Subsequently, the siRNA may be better tolerated and have a better safety profile than the existing monoclonal antibodies [121,122]. Inclisiran is currently in second phase 2 where it is being tested against the current available treatment, evolocumab (https://clinicaltrials.gov/ct2/show/NCT03060577), and in phase 3 against a placebo to determine safety and efficacy in the ORION-11 trial, a double-blind, randomized study in 1500 patients with atherosclerotic cardiovascular disease or atherosclerotic cardiovascular disease-risk equivalents and elevated LDL cholesterol (https://clinicaltrials.gov/ct2/show/NCT03400800).

Antisense oligonucleotide methodology is a strategy separate from, but related to the siRNA approach. Antisense oligonucleotides, chemically modified nucleic acid sequences that suppress gene expression by binding complementary RNA, have been used to target PCSK9 mRNA to inhibit its synthesis. Antisense agents have been successful in nonhuman primates [123]. However, no successful human trials have been conducted to date. A human study in The Netherlands of a 14-mer oligonucleotide had to be halted due to renal tubular toxicity [124]. Despite the roadblocks, researchers are still pursuing antisense oligonucleotides in the discovery and preclinical stages [125].

AFFITOPE® is a virus-like particle that elicits an antibody response and it has been applied against PCSK9 to create AT04A, an anti-PCSK9 AFFITOPE-based vaccine [126,127]. If successful, this therapy would not require frequent administration. This is more cost effective and comfortable for patients when compared with monoclonal antibodies which need to be administered every 2–4 weeks. Animal studies for AFFITOPE have been favorable and the drug is currently in phase 1 clinical trials (https://clinicaltrials.gov/ct2/show/NCT02508896).

Small molecule PCSK9 inhibitors are still in the discovery stage as researchers are continuing to identify effective drug targets. The PCSK9 binding sites are large and flat which make it difficult to develop a molecule that specifically binds to that site. Small molecules are also prone to off-target side effects because they are free in the systemic circulation [125]. Despite the potential drawbacks, there is a great deal of interest in creating small molecule PCSK9 inhibitors because they can be orally administered which is more convenient and comfortable for patients compared with injections [125]. Thus far, a few possible targets have been identified. Du et al. [128] discovered that the prodomain and C-terminal domain can be viable targets for PCSK9 inhibition and that the isolated C-terminal domain can prevent PCSK9-mediated LDL receptor degradation. Zhang et al. [129] identified Pep2-8, a PCSK9-binding peptide that is the smallest effective PCSK9 inhibitor to be discovered. Perhaps the most promising small molecule inhibitors were generated by Elshourbagy et al. [130,131] who has synthesized antagonists active at the nanomolar level. The small molecules they created interfere with the PCSK9/LDL receptor interaction. They have good oral bioavailability and are likely to be suitable for oral administration. These are in development with Shifa Biomedical Corporation.

Conclusions

PCSK9 has the ability to bind to LDL receptors on the surface of hepatocytes and promote degradation of these receptors within the liver. When this degradation activity is disabled by pharmacologic inhibition, LDL receptors remain intact and LDL levels decline. This has opened the door for the treatment of familial hypercholesterolemia and for novel approaches in treating hyperlipidemia. PCSK9 inhibition may also lessen atherogenic properties of lipoprotein particles by reducing apoB100. In recent years, PCSK9 inhibiting therapies have become a reality with two monoclonal antibodies approved by the FDA in 2015 and more drugs in the pipeline; an impressive feat given that PCSK9 was only discovered in 2003. Emerging monoclonal antibodies, siRNAs, antisense oligonucleotides, vaccines, and small molecules may further bridge the gap in treatment that currently exists for familial hypercholesterolemia, hyperlipidemia, and statin-intolerant persons with high cardiovascular risk. Further, as these drugs are explored, new indications for their use are likely (Table 2). There is the possibility of expansion to include use in dyslipidemic states related to diabetes, renal insufficiency and peripheral artery disease and to those with multiple cardiovascular risk factors [132–135].

Table 2
Current and possible future indications for PCSK9 inhibitors
Generic nameCurrent indicationsPossible future indicationsClinical trials
Alirocumab [41–44,53• Heterozygous familial hypercholesterolemia in adults
• Clinical atherosclerotic cardiovascular disease in adults who require additional lowering of LDL-C in adults 
• Atherosclerosis
• Hyperlipidemia
• Peripheral arterial disease
• Coronary heart disease
• Myocardial infarction
• Type 2 diabetes mellitus
• Dyslipidemia secondary to nephrotic syndrome
• Hemodialysis
• Peritoneal dialysis 
Ongoing or planned clinical trials registered with Clinicaltrials.gov for possible future indications
(https://clinicaltrials.gov/ct2/results?cond=&term=+alirocumab&cntry=&state=&city=&dist=
Evolocumab [48,49,54• Heterozygous familial hypercholesterolemia in adults
• Clinical atherosclerotic cardiovascular disease in adults who require additional lowering of LDL-C in adults 
• Atherosclerotic cardiovascular disease
• Hyperlipidemia
• Mixed dyslipidemia
• Peripheral arterial disease
• Type 2 diabetes mellitus
• Dyslipidemia associated with type 2 diabetes mellitus
• Percutaneous coronary intervention 
Ongoing or planned clinical trials registered with Clinicaltrials.gov for possible future indications
(https://clinicaltrials.gov/ct2/results?cond=&term=Evolocumab&cntry=&state=&city=&dist=
Inclisiran [122None • Atherosclerotic cardiovascular disease
• Symptomatic atherosclerosis
• Hyperlipidemia
• Type 2 diabetes
• Familial hypercholesterolemia
• Renal impairment 
Ongoing or planned clinical trials registered with Clinicaltrials.gov for possible future indications
(https://clinicaltrials.gov/ct2/results?cond=&term=Inclisiran&cntry=&state=&city=&dist=
AT04A [126,127None • Hypercholesterolemia Recently completed phase 1, results pending
(https://clinicaltrials.gov/ct2/show/NCT02508896?term=AT04A&rank=1
Generic nameCurrent indicationsPossible future indicationsClinical trials
Alirocumab [41–44,53• Heterozygous familial hypercholesterolemia in adults
• Clinical atherosclerotic cardiovascular disease in adults who require additional lowering of LDL-C in adults 
• Atherosclerosis
• Hyperlipidemia
• Peripheral arterial disease
• Coronary heart disease
• Myocardial infarction
• Type 2 diabetes mellitus
• Dyslipidemia secondary to nephrotic syndrome
• Hemodialysis
• Peritoneal dialysis 
Ongoing or planned clinical trials registered with Clinicaltrials.gov for possible future indications
(https://clinicaltrials.gov/ct2/results?cond=&term=+alirocumab&cntry=&state=&city=&dist=
Evolocumab [48,49,54• Heterozygous familial hypercholesterolemia in adults
• Clinical atherosclerotic cardiovascular disease in adults who require additional lowering of LDL-C in adults 
• Atherosclerotic cardiovascular disease
• Hyperlipidemia
• Mixed dyslipidemia
• Peripheral arterial disease
• Type 2 diabetes mellitus
• Dyslipidemia associated with type 2 diabetes mellitus
• Percutaneous coronary intervention 
Ongoing or planned clinical trials registered with Clinicaltrials.gov for possible future indications
(https://clinicaltrials.gov/ct2/results?cond=&term=Evolocumab&cntry=&state=&city=&dist=
Inclisiran [122None • Atherosclerotic cardiovascular disease
• Symptomatic atherosclerosis
• Hyperlipidemia
• Type 2 diabetes
• Familial hypercholesterolemia
• Renal impairment 
Ongoing or planned clinical trials registered with Clinicaltrials.gov for possible future indications
(https://clinicaltrials.gov/ct2/results?cond=&term=Inclisiran&cntry=&state=&city=&dist=
AT04A [126,127None • Hypercholesterolemia Recently completed phase 1, results pending
(https://clinicaltrials.gov/ct2/show/NCT02508896?term=AT04A&rank=1

Two PCSK9 inhibitors, alirocrumab and evolocumbab, are FDA approved. Two other drugs, inclisiran and AT04A, have not been approved by the FDA and are currently in clinical trials.

The availability of drugs to inhibit PCSK9 is a welcome discovery as statins, the most commonly used modality for lowering LDL, are largely ineffective for familial hypercholesterolemia and may not adequately reverse dyslipidemia in patients with atherosclerotic cardiovascular disease. Whether PCSK9 inhibitor-induced reductions in LDL will translate into fewer cardiovascular and cerebrovascular events and lower mortality must be documented in future large-scale studies. This is a time of rapid advances in which we are exploring the place of PCSK9 inhibitors in cardiovascular risk reduction while also acknowledging the possibility that newer, less costly approaches targeting other aspects of cholesterol synthesis, metabolism or disposal may supplant these drugs or that their price may be brought down which would lower the barrier to their use.

Clinical perspectives

  • PCSK9, a protein that regulates LDL receptor degradation, is a novel target for the treatment of hypercholesterolemia not well-controlled on statins or other therapies.

  • Currently available PCSK9 inhibitors, monoclonal antibodies that prevent binding of PCSK9 to the LDL receptor, are effective in reducing LDL levels, but their high cost and mechanism of administration by subcutaneous injection are disadvantages.

  • New therapies based on antisense technologies that target PCSK9 are on the horizon and may fill the gap in treatment for underserved hypercholesterolemic patients.

Acknowledgments

We thank Janet and Robert Buescher for their generous support. We thank Ms. Tanya Patterson-Stanley for getting this group of authors together.

Funding

This work was supported by American Heart Association Grant [16GRNT26430041] and by the Elizabeth Daniell Research Fund.

Competing Interests

The authors declare that there are no competing interests associated with the manuscript.

Author Contribution

All authors contributed to the writing and editing of this manuscript.

Abbreviations

     
  • ABI

    ankle brachial index

  •  
  • apo-

    apolipoprotein

  •  
  • EGF

    epidermal growth factor

  •  
  • FH

    familial hypercholesterolemia

  •  
  • HDL

    high-density lipoprotein

  •  
  • HMG-CoA

    3-hydroxy-3-methylglutarylcoenzyme A

  •  
  • HNF1α

    hepatocyte nuclear factor-1α

  •  
  • LDL

    low-density lipoprotein

  •  
  • Lp

    lipoprotein

  •  
  • PAD

    peripheral artery disease

  •  
  • PCSK9

    proprotein convertase subtilisin/kexin type 9

  •  
  • siRNA

    small interfering RNA

  •  
  • SRE

    sterol-regulatory element

  •  
  • SREBP

    sterol-regulatory element-binding protein

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