Disadvantages of Current LDL-cholesterol Lowering and the Role of PCSK9 Inhibitors

    Authors

    • Sanja Matijević — University of Rijeka School of Medicine, University Hospital Centre Rijeka, Rijeka, Croatia
    • — University of Rijeka School of Medicine, University Hospital Centre Rijeka, Rijeka, Croatia —
    • — University of Rijeka School of Medicine, University Hospital Centre Rijeka, Rijeka, Croatia —
    • Teodora Zaninović — University of Rijeka School of Medicine, University Hospital Centre Rijeka, Rijeka, Croatia
    • — University of Rijeka School of Medicine, University Hospital Centre Rijeka, Rijeka, Croatia —
    • Željka Rubeša — University of Rijeka School of Medicine, University Hospital Centre Rijeka, Rijeka, Croatia
    • — University of Rijeka School of Medicine, University Hospital Centre Rijeka, Rijeka, Croatia —
    • — University of Rijeka School of Medicine, University Hospital Centre Rijeka, Rijeka, Croatia —

    Abstract

    LDL cholesterol (LDL-C) is a strong independent cardiovascular (CV) risk factor that can be easily influenced. Today, statins are the therapy of choice for the achievement of target LDL-C values. Although controlled clinical trials have demonstrated their effectiveness and safety, in practice we are often met with statin intolerance as well as a failure to achieve target LDL-C values in a significant portion of the patients despite maximal doses. In patients with high and very high CV risk, other antilipemic pharmacotherapy is often also insufficient. PCSK9 inhibitors (PCSK9-I) are new revolutionary drugs with a potent effect on LDL-C. The rapid development of PCSK9-I began in 2003 with the discovery of a PCSK9 gene mutation in patients with familial hypercholesterolemia. The product of this gene, proprotein convertase subtilisin/kexin type 9 (PCSK9), has an important role in the expression of LDL receptors and cholesterol metabolism. Animal models demonstrated that inactivation of the PCSK9 gene lowers LDL-C with regression of atherosclerotic changes in the aorta. Heterozygotes and homozygotes with the inactivation mutation of PCSK9 have lower LDL-C values and lower incidence of atherosclerosis. Among the various groups of PCSK9-I, monoclonal antibodies saw strong development (alirocumab, evolocumab, bococizumab). Phase 3 clinical trials on familial and primary hypercholesterolemia with statin intolerance or resistance have demonstrated a strong positive effect of alirocumab and evolocumab on LDL-C values (a reduction of 60%), with high safety and good tolerability. The OSLER study on evolocumab also demonstrated positive effects on CV outcomes. Multiple clinical trials on PCSK9-I are currently monitoring their effect on CV morbidity and mortality. Positive results from these studies would confirm the great potential of PCSK9-I for better prevention and treatment of CV diseases.

    Keywords

    cardiovascular diseases, dyslipidemia, LDL-cholesterol, statins, PCSK9 inhibitors

    DOI

    https://doi.org/10.15836/ccar2016.330

    Full Text

    ## Introduction LDL cholesterol (LDL-C) is a strong independent cardiovascular (CV) risk factor that can be influenced. Achieving target LDL-C values is one of the fundamental goals of primary and secondary CV disease prevention (1, 2). Clinical trials have demonstrated that a reduction in LDL-C values of 1 mmol/L reduces the risk of unwanted CV outcomes by 22% (3, 4). According to current European Society of Cardiology guidelines, target LDL-C values are determined by the total CV risk, and are 3.0 mmol/L (34). Combining ezetimibe with bile acid sequestrants achieved better effects on LDL-C without a significant increase in side effects in comparison with bile acid sequestrants monotherapy (35). The limitations of this combined treatment are the high incidence of side effects of bile acid sequestrants (constipation, diarrhea, bloating, high serum triglycerides) and repeated drug administration, which reduces patient cooperation and causes frequent treatment interruptions (30). Fibrates and niacin have a larger effect on so-called non-HDL cholesterol, while their effect on LDL-C is much weaker. Fibrates reduced LDL-C by 13-35%, and the side effect profile is similar to that of statins (primarily myopathy), making them unsuitable for the treatment of patients with statin intolerance (36, 37). In combined treatment with statins in patients with statin resistance, they provide a significant added reduction in LDL-C, but many patients still do not reach target values (5, 38). Niacin is a well-known drug with beneficial antilipemic effects but poor tolerance. To reduce side effects, new forms of the drug have been studied – niacin extended release (ER-niacin) and the combination of niacin with laropiprant (the AIM-HIGH and HPS 2-THRIVE studies) (39, 40). Although a reduction in LDL-C was demonstrated, there was no reduction in unwanted CV events, and new serious side effects were present along with those already known. Thus, the European Medicines Agency (EMA) recently recommended withdrawing these drugs from the European market. New drugs with beneficial effects on LDL-C, mipomersen and lomitapide, have been recently approved by the American Food and Drug Administration (FDA) for the treatment of the homozygous hereditary hypercholesterolemia (**Figure 1**). Mipomersen is a specific oligonucleotide that binds to ApoB messenger RNA and inhibits its translation, i.e. the synthesis of ApoB (the primary structural apoprotein for LDL-C and other atherogenic lipoproteins), and consequently also VLDL particles, which finally results in lower LDL-C values. Lomitapide is an oral inhibitor for the microsomal triglyceride transfer protein (MTP) and an important link in the production of VLDL in the liver (41). Although clinical trials showed a significant reduction in LDL-C values, their effect on CV outcomes has not yet been examined. Due to common side effects (increased buildup of fats in the liver and severe skin reactions at the injection site for mipomersen), it is not likely that these drugs will ever be approved for the treatment of patients with statin intolerance and resistance (42, 43). Figure 1. New pharmacological options for the treatment of dyslipidaemias. *PCSK9: proprotein convertase subtilisin/kexin type 9, **CETP: cholesteryl ester transfer protein, ***MTP: microsomal triglyceride transfer protein. ## Proprotein convertase subtilisin/kexin type 9 (PCSK9) PCSK9 inhibitors are new revolutionary drugs for the treatment of elevated LDL-C values. The therapeutic use of PCSK9 inhibition was preceded by the discovery of a PCSK9 gene mutation in patients with the dominant form of hereditary hypercholesterolemia in 2003 (44). It has been demonstrated that the mutation of the PCSK9 gene is present in 10-25% of cases of heterozygous hereditary hypercholesterolemia, resulting in increased activity of the gene and elevated concentrations of PCSK9 in the liver and in peripheral blood (7, 45). PCSK9 is an enzyme from the proteinase K similar proteins, belonging to the secretory subtilisin group. It is primarily synthetized in hepatocytes and secernated in the liver, where it reaches the highest concentrations (46, 47). The key role of PCSK9 is the regulation of LDL-R expression in the liver. Binding the N-terminal end of the PCSK9 molecule to LDL-R leads to it being internalized in the cell and broken down with lysosomes (**Figure 2**). The result is a reduction in the amount of cell LDL-R, lower clearance, and higher plasma LDL-C concentration (48). Extrahepatic effects of PCSK9 include increased secretion of chylomicrons and regulation of cholesterol absorption in enterocytes (47). Studies using animal models demonstrated a strong effect of PCSK9 on the progression or regression of atherosclerotic changes (49). The results of these studies suggest that PCSK9 does not only affect the metabolism of cholesterol but is also involved in glucose metabolism, hepatocyte regeneration, and susceptibility to hepatitis C virus infection (50-53). Figure 2. Regulation of hepatic LDL receptor expression and mechanism of LDL cholesterol reduction by PCSK9 inhibition (adapted from Shimada YJ et al94). While the PCSK9 gene is more active in patients with familial hypercholesterolemia, clinical studies have shown that a mutation that results in reduced activity of the gene is accompanied by a reduction in LDL-C levels of 11-28% and a significantly lower incidence of CV events, lower carotid intima-media thickness, and less common peripheral arterial disease (54-59). These discoveries were the basis for the idea of applying pharmacological PCSK9 inhibition in patients with statin intolerance or resistance, and later in those with hereditary or primary hypercholesterolemia and in all high-risk CV patients who do not achieve target LDL-C values despite optimal therapy. The results of clinical trials have so far been very promising. As opposed to previously mentioned mipomersen and lomitapide, PCSK9 inhibitors did not show significant side effects, while achieving stronger LDL-C reduction. Multiple ways of inhibiting PCSK9 have been studied. While the first phase of clinical trials with antisense oligonucleotides has been discontinued (60), trial results for PCSK9 inhibition with monoclonal antibodies or small interfering RNA have been very promising. Small interfering RNA binds to PCSK9 messenger RNA and prevent its translation and the production of PCSK9. After successful pre-clinical animal model experiments, the beneficial effect of small interfering RNA has been demonstrated in the first phase of clinical trials, where a 70% reduction in free plasma PCSK9 and a 40% reduction in LDL-C were achieved (61, 62). The most recent focus is monoclonal inhibition of the PCSK9 molecule. Alirocumab and evolocumab are human monoclonal antibodies that bind PCSK9, increase the number of LDL-Rs, and reduce LDL-C concentrations (**Figure 2**). Bococizumab is a recombinant humanized mouse antibody against PCSK9. Multiple phase 3 clinical trials with alirocumab (ODYSSEY Mono, ODYSSEY COMBO I, ODYSSEY COMBO II) and evolocumab (DECARTES, LAPLACE-2, GAUSS-2, MENDEL-2, RUTHERFORD-2, OSLER-2, TESLA Part B) have been completed, while such trials with bococizumab are still under way (63, 64). ## Alirocumab The effectiveness of alirocumab was studied in over 6000 study participants with primary hypercholesterolemia that had statin intolerance or did not achieve target LDL-C values despite maximum tolerable hypolipidemic agent doses. ODYSSEY-MONO was the first study on alirocumab. The effectiveness and safety of alirocumab monotherapy was studied, in comparison with ezetimibe, in patients with hypercholesterolemia and moderate CV risk (10-year risk of death from CV events of 1-5% according to the SCORE tool) who had not been previously treated with statins or other hypolipidemic agents. In 103 patients with LDL-C values of 2.6-4.9 mmol/L, alirocumab led to a significant reduction in LDL-C values in comparison with ezetimibe. In most participants, the lower alirocumab dose of 75 mg once per two weeks was sufficient to reduce baseline LDL-C by 50% (65). The studies ODYSSEY COMBO I (311 patients) and ODYSSEY COMBO II (707 patients) consisted of participants with hypercholesterolemia and high CV risk that did not achieve target LDL-C values despite treatment with maximum tolerable statin doses (with or without additional hypolipidemic therapy). The effectiveness of alirocumab in comparison with ezetimibe was studied. Both studies demonstrated significantly better effects of alirocumab on LDL-C values and other studied lipid parameters. In the ODYSSEY COMBO II, 24 weeks of treatment with alirocumab achieved a reduction of LDL-C values of 50.6%, while ezetimibe reduced LDL-C by 20.7% (p<0.0001). Furthermore, significantly more participants treated with alirocumab achieved target LDL-C values (<1.8 mmol/L) in comparison with participants treated with ezetimibe (77.0% vs. 45.6%, p<0.0001) (66, 67). In the ODYSSEY-OPTIONS study, adding alirocumab to a statin resulted in significantly larger reduction of LDL-C values than adding ezetimibe, doubling the statin dose, or changing to the more potent rosuvastatin (68, 69). The studies ODYSSEY FH I and ODYSSEY FH II examined the effectiveness and safety of alirocumab in patients with heterozygous familial hypercholesterolemia. Target LDL-C values were defined based on CV risk: in patients with no diagnosed CV disease as <2.6 mmol/L, and as <1.8 mmol/L in those with already diagnosed CV diseases. At 24-week follow-up, 72.2% of participants using alirocumab achieved target LDL-C values in FH I and 81.4% in FH II (p<0.0001) (70, 71). The beneficial effect on LDL-C, along with good tolerance of alirocumab, was still present at 78-week follow-up (72). ODYSSEY LONG TERM, currently the broadest study on alirocumab, assessed the long-term safety and tolerability of alirocumab. It encompassed as many as 2341 high-risk patients (patients with heterozygous familial hypercholesterolemia with or without manifest CV diseases and patients with primary hypercholesterolemia and coronary heart disease) who did not achieve target LDL-C values despite maximum tolerable hypolipidemic therapy, with 44% of the participants taking maximum safe doses of statins. After 24 weeks of treatment, alirocumab achieved a high reduction in LDL-C values of 61.9%, as well as reduction of other significant lipid risk parameters – ApoB by 54.0%, non-HDL cholesterol by 52.3%, and lipoprotein(a) by 25.6%, along with an increase in HDL-C of 4.6% (p<0.0001). Target LDL-C values of <2.6 mmol/L were achieved in 76% of participants taking alirocumab, in comparison with just 2% of participants in the control group, and the target value of <1.8 mmol/L was achieved by as much as 81% of participants taking alirocumab in comparison with 9% in the control group (73). ## Evolocumab Evolocumab has proven its high effectiveness and safety in a number of clinical trials. Although the effect of the drug was similar when taken every two and every four weeks (74), larger fluctuations in LDL-C values were present in four-week doses (75). The MENDEL study included a total of 406 participants with hypercholesterolemia and statin intolerance randomized into subgroups taking evolocumab in doses of 70, 105, and 140 mg every 2 weeks, subgroups taking 280, 350, and 420 mg every 4 weeks, and taking ezetimibe. Evolocumab was shown to be greatly superior, and the strongest effect was achieved with the dose of 140 mg every 2 weeks, with no significant side effects (76). The MENDEL-2 study assessed the effectiveness, safety, and tolerability of evolocumab in comparison with ezetimibe and placebo in participants with hypercholesterolemia and high CV risk. Evolocumab reduced LDL-C values by 55-57% more in comparison with placebo and 38-40% more in comparison with ezetimibe (p<0.0001 in both cases) (77). The LAPLACE-TIMI study randomized 57 patients with hypercholesterolemia who were treated with statins into subgroups taking different doses of evolocumab in different intervals (every two and every four weeks). A significant reduction in LDL-C values was achieved after 12 weeks, depending on the evolocumab dose (42-66% when taken every two weeks and 41-50% when taken every four weeks) (78). The LAPLACE-2 study compared evolocumab at different doses and intervals (140 mg every two weeks and 420 once per month) with placebo and ezetimibe in 1896 participants with primary hypercholesterolemia and mixed dyslipidemia that were treated with moderate and high doses of statins. It was found that adding evolocumab to statin treatment reduces LDL-C values significantly more (by 66-75%, depending on the dose) after 10 to 12 weeks in comparison with ezetimibe or placebo (79). The DESCARTES study with evolocumab randomized participants into four groups based on CV risk and initial LDL-C values, consisting of participants receiving no additional hypolipidemic therapy, those that in addition to evolocumab also received atorvastatin 10 mg or atorvastatin 80 mg, and participants that received combination therapy consisting of atorvastatin 80 mg and ezetimibe 10 mg in addition to evolocumab. The strong beneficial effects of evolocumab were demonstrated in all four groups. In addition to the beneficial effect on LDL-C values, the study also found a significant reduction in other lipid CV risk factors – apolipoprotein B, lipoprotein(a), and triglycerides (80). The GAUSS study examined the effectiveness and tolerability of evolocumab in participants with statin intolerance due to myalgia or myopathy. Evolocumab doses of 280 mg, 350 mg, and 420 mg were tested, as well as evolocumab 420 mg in combination with ezetimibe 10 mg. The effect of evolocumab and the combination treatment was compared with ezetimibe 10 mg. At all doses, evolocumab achieved a significant reduction of LDL-C (40-65%, depending on the dose), with myalgia being present in only 7.4% participants using evolocumab, 20% of participants taking combination evolocumab and ezetimibe treatment, and 3.1% of participants receiving ezetimibe and placebo (81). The GAUSS-2 study also included participants with hypercholesterolemia and statin intolerance (82, 83). The study participants received evolocumab 140 mg every two weeks or 420 mg once per month, while the control group was given ezetimibe. Evolocumab led to a more significant reduction in LDL-C values with a lower incidence of muscle side effects in comparison with ezetimibe (12% for evolocumab, 23% for ezetimibe). The RUTHERFORD study demonstrated the high effectiveness of evolocumab in patients with the heterozygous hereditary hypercholesterolemia (167 participants) and high LDL-C despite taking the maximum tolerable statin dose (84). In the study RUTHERFORD-2, a third of the participants took maximum tolerable statin doses, and two thirds received ezetimibe in addition to statins. The participants were randomized into groups that received evolocumab 140 mg every two weeks, evolocumab 420 mg once per month, and the control group taking placebo. In both groups on evolocumab, significant reduction in LDL-C values was achieved after 12 weeks (59.2% for 140 mg and 61.3% for 420 mg doses) (p<0.0001) (85). The TESLA study demonstrated the positive effect of evolocumab in patients with homozygous familial hypercholesterolemia treated with maximum tolerable statin doses and other hypolipidemic therapy, with evolocumab treatment achieving an additional LDL-C reduction of 31% after 12 weeks in comparison with placebo, with no significant side effects (86). The OSLER study examined the effect of evolocumab on LDL-C values as well as CV outcomes. A total of 4465 participants were randomized in groups taking 140 mg evolocumab every 2 weeks and 420 mg once per month. During 11.1 months of follow-up, evolocumab reduced LDL-C values by 61% (from an average of 3.1 mmol/L to 1.2 mmol/L, p<0.001). Combined cardiovascular outcomes (mortality, acute coronary syndrome, heart failure, stroke, or TIA) were 2.18% after one year in the control group, and 0.95% in the evolocumab group (HR 0.47; 95% CI 0.28-0.78; p=0.003). Before randomization, most patients were receiving statins (69.7% in the evolocumab group and 70.9% in the control group receiving placebo). (87) The GLAGOV study with evolocumab will assess the effects of low LDL-C concentrations on volume regression of atherosclerotic plaque in patients with already diagnosed coronary heart disease (88). ## Bococizumab Phase 2b clinical trials with bococizumab achieved significant reduction of LDL-C values in participants with hypercholesterolemia treated with statins. The trials included 354 participants receiving doses of bococizumab 50 mg, 100 mg, and 150 mg every two weeks and doses of 200 mg and 300 mg once per month over the course of 12 weeks. Bococizumab achieved a significant beneficial effect on LDL-C values in comparison with placebo at all doses and intervals, with the strongest reduction in LDL-C being achieved by doses of 150 mg every two weeks and 300 mg once per month (89). Phase 3 clinical trials with bococizumab began in October 2013. They include two studies which primarily assess CV outcomes (SPIRE-I and SPIRE-II) and multiple studies assessing the effects of bococizumab on various lipid parameters, consisting of participants with hypercholesterolemia and high CV risk who were resistant to statins. The primary aim of the SPIRE-I study is to examine whether LDL-C reduction below values recommended in current guidelines has an added beneficial effect on CV outcomes in high-risk participants with initial LDL-C values of 1.81 to 2.59 mmol/L (63). The SPIRE-II study will examine the effectiveness and safety of bococizumab in high-risk patients who have not achieved the target LDL-C value of <2.59mmol/L despite statin therapy or in those with statin intolerance (64). Both of these studies use a starting bococizumab dose of 150 mg every two weeks. ## Additional effects of PCSK9 inhibitors In addition to a beneficial effect on LDL-C, PCSK9 inhibitors have also showed other beneficial hypolipidemic effects. Elevated lipoprotein(a) is an independent CV risk factor in patients treated with statins with low LDL-C (90). PCSK9 inhibitors reduced Apo(a) by approximately 30%, implying possible additional cardioprotective effects beyond those achieved by influencing LDL-C values. ## PCSK9 inhibitor safety The abovementioned studies also assessed the safety of PCSK9 inhibitors in addition to their effectiveness. There was no significant difference in the incidence of mild and more severe side effects between the assessed monoclonal antibodies and comparative drugs or placebo (79, 80). Less than 2% of participants receiving PCSK9 inhibitors presented with aminotransferase elevations of three or more times the upper limit of normal values. CK elevation was equally rare (91). The most common PCSK9 inhibitor side effects were nasopharyngitis, upper respiratory tract infections, flu-like symptoms, and back pain. Reactions at the subcutaneous site of drug injection were present in 2% of the study participants on alirocumab and 4% of those on evolocumab (92). The incidence of treatment termination due to side effects did not differ significantly between PCSK9 inhibitors (alirocumab and evolocumab) and comparison drugs (2-10%) (93). ## Conclusion LDL-C values associated with the most positive CV outcomes in persons with high and very high risk are very low (<2.6 and <1.8 mmol/L). Achieving low values is a great challenge, and currently patients often fail to achieve them with hypolipidemic therapy. The discovery of new regulatory mechanisms involved in lipoprotein metabolism has opened the way to new pharmacological approaches for the treatment of elevated LDL-C. Discovering the role of PSCK9 in LDL-R expression and LDL-C clearance has spurred the rapid development of new groups of drugs (94). Of the several groups of PCSK9 inhibitors, monoclonal antibodies have seen the most development. Alirocumab, evolocumab, and bococizumab reduce LDL-C values by 50-70%, with good tolerance and high safety. A great challenge in their practical application will undoubtedly be their price, and also in part their subcutaneous application. A beneficial effect on CV outcomes has been found in the OSLER study with evolocumab and is being assessed in a series of clinical trials. The results of these studies will certainly give a clearer picture of the clinical effectiveness and cost-effectiveness of PCSK9 inhibitors in chosen groups of patients.

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    Cardiologia Croatica
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    Disadvantages of Current LDL-cholesterol Lowering and the Role of PCSK9 Inhibitors

    Review Article
    Issue8
    Published
    Pages330-342
    PDF via DOIhttps://doi.org/10.15836/ccar2016.330
    cardiovascular diseases
    dyslipidemia
    LDL-cholesterol
    statins
    PCSK9 inhibitors

    Authors

    Sanja MatijevićUniversity of Rijeka School of Medicine, University Hospital Centre Rijeka, Rijeka, Croatia
    RončevićORCIDUniversity of Rijeka School of Medicine, University Hospital Centre Rijeka, Rijeka, Croatia
    David GobićORCIDUniversity of Rijeka School of Medicine, University Hospital Centre Rijeka, Rijeka, Croatia
    Teodora ZaninovićUniversity of Rijeka School of Medicine, University Hospital Centre Rijeka, Rijeka, Croatia
    JurjevićORCIDUniversity of Rijeka School of Medicine, University Hospital Centre Rijeka, Rijeka, Croatia
    Željka RubešaUniversity of Rijeka School of Medicine, University Hospital Centre Rijeka, Rijeka, Croatia
    MiculinićORCIDUniversity of Rijeka School of Medicine, University Hospital Centre Rijeka, Rijeka, Croatia
    Luka Zaputović*ORCIDUniversity of Rijeka School of Medicine, University Hospital Centre Rijeka, Rijeka, Croatia

    *Correspondence email: luka.zaputovic@medri.uniri.hr

    Abstract

    LDL cholesterol (LDL-C) is a strong independent cardiovascular (CV) risk factor that can be easily influenced. Today, statins are the therapy of choice for the achievement of target LDL-C values. Although controlled clinical trials have demonstrated their effectiveness and safety, in practice we are often met with statin intolerance as well as a failure to achieve target LDL-C values in a significant portion of the patients despite maximal doses. In patients with high and very high CV risk, other antilipemic pharmacotherapy is often also insufficient. PCSK9 inhibitors (PCSK9-I) are new revolutionary drugs with a potent effect on LDL-C. The rapid development of PCSK9-I began in 2003 with the discovery of a PCSK9 gene mutation in patients with familial hypercholesterolemia. The product of this gene, proprotein convertase subtilisin/kexin type 9 (PCSK9), has an important role in the expression of LDL receptors and cholesterol metabolism. Animal models demonstrated that inactivation of the PCSK9 gene lowers LDL-C with regression of atherosclerotic changes in the aorta. Heterozygotes and homozygotes with the inactivation mutation of PCSK9 have lower LDL-C values and lower incidence of atherosclerosis. Among the various groups of PCSK9-I, monoclonal antibodies saw strong development (alirocumab, evolocumab, bococizumab). Phase 3 clinical trials on familial and primary hypercholesterolemia with statin intolerance or resistance have demonstrated a strong positive effect of alirocumab and evolocumab on LDL-C values (a reduction of 60%), with high safety and good tolerability. The OSLER study on evolocumab also demonstrated positive effects on CV outcomes. Multiple clinical trials on PCSK9-I are currently monitoring their effect on CV morbidity and mortality. Positive results from these studies would confirm the great potential of PCSK9-I for better prevention and treatment of CV diseases.

    Full Text

    Introduction

    LDL cholesterol (LDL-C) is a strong independent cardiovascular (CV) risk factor that can be influenced. Achieving target LDL-C values is one of the fundamental goals of primary and secondary CV disease prevention (1, 2). Clinical trials have demonstrated that a reduction in LDL-C values of 1 mmol/L reduces the risk of unwanted CV outcomes by 22% (3, 4). According to current European Society of Cardiology guidelines, target LDL-C values are determined by the total CV risk, and are <1.8 mmol/L in persons with very high risk and <2.6 mmol/L for persons with high risk (5, 6). Patients with hereditary hypercholesterolemia are a population with especially high risk, since the diseases manifests already in their twenties in homozygous patients and in the forties in heterozygous patients (7–9). The treatment of choice for elevated LDL-C are the well-known statins. It was statins that demonstrated a strong association between elevated LDL-C and unwanted CV events in numerous clinical trials and meta-analyses (10). Although they were shown to be safe and effective in randomized controlled clinical trials (1-5% total muscle side effects), in everyday clinical practice we are often faced with the issue of statin intolerance and resistance to statin treatment (11).

    Statin intolerance

    The definition of statin intolerance has still not been agreed upon. There are several modern definitions (EMA, NLA, Canadian Working Group Consensus), all of which include general statin intolerance or intolerance to full treatment doses due to the appearance of various side effects (12–14). Most common are muscle side effects, ranging from asymptomatic elevation of creatine kinase (CK) levels to myalgia, myopathy, myositis, and rhabdomyolysis as the most severe and very rare muscle side effect (15). The causes of statin intolerance are not fully known, but it has been shown that patients have individual, genetically determined susceptibility to statins, dependent on gene polymorphism and proteins involved in the metabolism or transport of statins to hepatocytes (CYP 450, OATP) (16, 17). It is also well known that statin tolerance depends on the doses and the simultaneous application of drugs that affect statin metabolism (elevating plasma statin levels – gemfibrozil, itraconazole) (18–20) and that muscle side effects are more rare for some statins (pravastatin, fluvastatin, and pitavastatin) due to different metabolic pathways (21, 22). Recent studies indicate the possibility of achieving statin tolerance in most patients with statin intolerance through dose reduction and intermittent drug application (23, 24). Lower statin doses in the treatment of patients with an already diagnosed CV disease will rarely achieve target LDL-C values. The discrepancy in the incidence of muscle side effects in clinical and post-clinical trials is the result of strict inclusion criteria. Patients with already diagnosed muscle diseases, history of myalgia, elevated CK values, and those taking drugs that affect statin metabolism were not included in the clinical trials. Furthermore, most randomized statin studies did not include a detailed assessment of milder muscle side effects. The focus was on rhabdomyolysis, which is, as already mentioned, very rare.

    The second most common side effects are rare liver side effects that manifest in 3% of patients receiving high statin doses and in less than 1% of patients taking moderate or low doses (25). Statins are considered to be affecting the liver when aminotransferase levels increase by three or more times above the upper limit of normal levels after commencement of statin treatment. This is usually a transient effect (26). Severe liver damage caused by statins is rare and unpredictable. Newer studies have demonstrated that non-alcoholic fatty liver disease (non-alcoholic steatohepatitis) is not a contraindication for statin treatment (27, 28). On the contrary, statins led to aminotransferase reduction in a portion of these patients with mild to moderate elevation of liver enzymes.

    A rare statin side effect is newly developed diabetes type 2, especially in persons with previously impaired glucose tolerance. However, a reduction in total CV risk was achieved in these patients as well (29).

    Statin resistance

    Statin resistance is the inability to achieve target LDL-C values despite treatment with the maximum statin therapeutic dose. The inability to prevent atherosclerotic changes and reduce unwanted CV events using statins can also be considered statin resistance. This should be differentiated from the much more common pseudo-resistance, which is the consequence of the patient’s lack of treatment compliance. Not achieving target LDL-C values is often a result of unjustified prescription of insufficient doses or less effective statins (30). Like statin intolerance, statin resistance is mostly determined by genetic factors. Research has shown that the polymorphism of numerous genes causes different individual responses to statins and affects the pharmacokinetics and pharmacodynamics of the medication (31). In addition to genetic factors, other acquired factors have been shown to affect individual response to statins. Smokers and patients with arterial hypertension have weaker response than non-smokers and persons with normal arterial pressure (32). The inflammatory cytokine IL-1 beta affects feedback regulation of LDL receptors (LDL-R), reducing its expression, so a larger statin dose is required under inflammatory conditions to achieve the same effect on LDL-C values. Reduced LDL-R expression is also caused by hypothyroidism and treatment with amiodarone, regardless of the thyroid status of the patient (33).

    Other hypolipidemics with positive effects on LDL-cholesterol

    Other hypolipidemics available for achieving target LDL-C values in patents with statin resistance or intolerance are ezetimibe, bile acid sequestrants, fibrates, and niacin. Ezetimibe is a selective inhibitor of cholesterol absorption in the small intestine mucosa. In comparison with statins, it has a significantly weaker effect on LDL-C. In clinical trials on patients with familial hypercholesterolemia, ezetimibe monotherapy reduced LDL-C by a humble 7-11%. In combination with statins, the target LDL-C reduction of 3.0 mmol/L (34). Combining ezetimibe with bile acid sequestrants achieved better effects on LDL-C without a significant increase in side effects in comparison with bile acid sequestrants monotherapy (35). The limitations of this combined treatment are the high incidence of side effects of bile acid sequestrants (constipation, diarrhea, bloating, high serum triglycerides) and repeated drug administration, which reduces patient cooperation and causes frequent treatment interruptions (30).

    Fibrates and niacin have a larger effect on so-called non-HDL cholesterol, while their effect on LDL-C is much weaker. Fibrates reduced LDL-C by 13-35%, and the side effect profile is similar to that of statins (primarily myopathy), making them unsuitable for the treatment of patients with statin intolerance (36, 37). In combined treatment with statins in patients with statin resistance, they provide a significant added reduction in LDL-C, but many patients still do not reach target values (5, 38).

    Niacin is a well-known drug with beneficial antilipemic effects but poor tolerance. To reduce side effects, new forms of the drug have been studied – niacin extended release (ER-niacin) and the combination of niacin with laropiprant (the AIM-HIGH and HPS 2-THRIVE studies) (39, 40). Although a reduction in LDL-C was demonstrated, there was no reduction in unwanted CV events, and new serious side effects were present along with those already known. Thus, the European Medicines Agency (EMA) recently recommended withdrawing these drugs from the European market.

    New drugs with beneficial effects on LDL-C, mipomersen and lomitapide, have been recently approved by the American Food and Drug Administration (FDA) for the treatment of the homozygous hereditary hypercholesterolemia (Figure 1). Mipomersen is a specific oligonucleotide that binds to ApoB messenger RNA and inhibits its translation, i.e. the synthesis of ApoB (the primary structural apoprotein for LDL-C and other atherogenic lipoproteins), and consequently also VLDL particles, which finally results in lower LDL-C values. Lomitapide is an oral inhibitor for the microsomal triglyceride transfer protein (MTP) and an important link in the production of VLDL in the liver (41). Although clinical trials showed a significant reduction in LDL-C values, their effect on CV outcomes has not yet been examined. Due to common side effects (increased buildup of fats in the liver and severe skin reactions at the injection site for mipomersen), it is not likely that these drugs will ever be approved for the treatment of patients with statin intolerance and resistance (42, 43).

    Figure 1. New pharmacological options for the treatment of dyslipidaemias. PCSK9: proprotein convertase subtilisin/kexin type 9, CETP: cholesteryl ester transfer protein, **MTP: microsomal triglyceride transfer protein.

    Proprotein convertase subtilisin/kexin type 9 (PCSK9)

    PCSK9 inhibitors are new revolutionary drugs for the treatment of elevated LDL-C values. The therapeutic use of PCSK9 inhibition was preceded by the discovery of a PCSK9 gene mutation in patients with the dominant form of hereditary hypercholesterolemia in 2003 (44). It has been demonstrated that the mutation of the PCSK9 gene is present in 10-25% of cases of heterozygous hereditary hypercholesterolemia, resulting in increased activity of the gene and elevated concentrations of PCSK9 in the liver and in peripheral blood (7, 45).

    PCSK9 is an enzyme from the proteinase K similar proteins, belonging to the secretory subtilisin group. It is primarily synthetized in hepatocytes and secernated in the liver, where it reaches the highest concentrations (46, 47).

    The key role of PCSK9 is the regulation of LDL-R expression in the liver. Binding the N-terminal end of the PCSK9 molecule to LDL-R leads to it being internalized in the cell and broken down with lysosomes (Figure 2). The result is a reduction in the amount of cell LDL-R, lower clearance, and higher plasma LDL-C concentration (48). Extrahepatic effects of PCSK9 include increased secretion of chylomicrons and regulation of cholesterol absorption in enterocytes (47). Studies using animal models demonstrated a strong effect of PCSK9 on the progression or regression of atherosclerotic changes (49). The results of these studies suggest that PCSK9 does not only affect the metabolism of cholesterol but is also involved in glucose metabolism, hepatocyte regeneration, and susceptibility to hepatitis C virus infection (50–53).

    Figure 2. Regulation of hepatic LDL receptor expression and mechanism of LDL cholesterol reduction by PCSK9 inhibition (adapted from Shimada YJ et al94).

    While the PCSK9 gene is more active in patients with familial hypercholesterolemia, clinical studies have shown that a mutation that results in reduced activity of the gene is accompanied by a reduction in LDL-C levels of 11-28% and a significantly lower incidence of CV events, lower carotid intima-media thickness, and less common peripheral arterial disease (54–59).

    These discoveries were the basis for the idea of applying pharmacological PCSK9 inhibition in patients with statin intolerance or resistance, and later in those with hereditary or primary hypercholesterolemia and in all high-risk CV patients who do not achieve target LDL-C values despite optimal therapy. The results of clinical trials have so far been very promising. As opposed to previously mentioned mipomersen and lomitapide, PCSK9 inhibitors did not show significant side effects, while achieving stronger LDL-C reduction. Multiple ways of inhibiting PCSK9 have been studied. While the first phase of clinical trials with antisense oligonucleotides has been discontinued (60), trial results for PCSK9 inhibition with monoclonal antibodies or small interfering RNA have been very promising. Small interfering RNA binds to PCSK9 messenger RNA and prevent its translation and the production of PCSK9. After successful pre-clinical animal model experiments, the beneficial effect of small interfering RNA has been demonstrated in the first phase of clinical trials, where a 70% reduction in free plasma PCSK9 and a 40% reduction in LDL-C were achieved (61, 62).

    The most recent focus is monoclonal inhibition of the PCSK9 molecule. Alirocumab and evolocumab are human monoclonal antibodies that bind PCSK9, increase the number of LDL-Rs, and reduce LDL-C concentrations (Figure 2). Bococizumab is a recombinant humanized mouse antibody against PCSK9. Multiple phase 3 clinical trials with alirocumab (ODYSSEY Mono, ODYSSEY COMBO I, ODYSSEY COMBO II) and evolocumab (DECARTES, LAPLACE-2, GAUSS-2, MENDEL-2, RUTHERFORD-2, OSLER-2, TESLA Part B) have been completed, while such trials with bococizumab are still under way (63, 64).

    Alirocumab

    The effectiveness of alirocumab was studied in over 6000 study participants with primary hypercholesterolemia that had statin intolerance or did not achieve target LDL-C values despite maximum tolerable hypolipidemic agent doses.

    ODYSSEY-MONO was the first study on alirocumab. The effectiveness and safety of alirocumab monotherapy was studied, in comparison with ezetimibe, in patients with hypercholesterolemia and moderate CV risk (10-year risk of death from CV events of 1-5% according to the SCORE tool) who had not been previously treated with statins or other hypolipidemic agents. In 103 patients with LDL-C values of 2.6-4.9 mmol/L, alirocumab led to a significant reduction in LDL-C values in comparison with ezetimibe. In most participants, the lower alirocumab dose of 75 mg once per two weeks was sufficient to reduce baseline LDL-C by 50% (65).

    The studies ODYSSEY COMBO I (311 patients) and ODYSSEY COMBO II (707 patients) consisted of participants with hypercholesterolemia and high CV risk that did not achieve target LDL-C values despite treatment with maximum tolerable statin doses (with or without additional hypolipidemic therapy). The effectiveness of alirocumab in comparison with ezetimibe was studied. Both studies demonstrated significantly better effects of alirocumab on LDL-C values and other studied lipid parameters. In the ODYSSEY COMBO II, 24 weeks of treatment with alirocumab achieved a reduction of LDL-C values of 50.6%, while ezetimibe reduced LDL-C by 20.7% (p<0.0001). Furthermore, significantly more participants treated with alirocumab achieved target LDL-C values (<1.8 mmol/L) in comparison with participants treated with ezetimibe (77.0% vs. 45.6%, p<0.0001) (66, 67). In the ODYSSEY-OPTIONS study, adding alirocumab to a statin resulted in significantly larger reduction of LDL-C values than adding ezetimibe, doubling the statin dose, or changing to the more potent rosuvastatin (68, 69).

    The studies ODYSSEY FH I and ODYSSEY FH II examined the effectiveness and safety of alirocumab in patients with heterozygous familial hypercholesterolemia. Target LDL-C values were defined based on CV risk: in patients with no diagnosed CV disease as <2.6 mmol/L, and as <1.8 mmol/L in those with already diagnosed CV diseases. At 24-week follow-up, 72.2% of participants using alirocumab achieved target LDL-C values in FH I and 81.4% in FH II (p<0.0001) (70, 71). The beneficial effect on LDL-C, along with good tolerance of alirocumab, was still present at 78-week follow-up (72).

    ODYSSEY LONG TERM, currently the broadest study on alirocumab, assessed the long-term safety and tolerability of alirocumab. It encompassed as many as 2341 high-risk patients (patients with heterozygous familial hypercholesterolemia with or without manifest CV diseases and patients with primary hypercholesterolemia and coronary heart disease) who did not achieve target LDL-C values despite maximum tolerable hypolipidemic therapy, with 44% of the participants taking maximum safe doses of statins. After 24 weeks of treatment, alirocumab achieved a high reduction in LDL-C values of 61.9%, as well as reduction of other significant lipid risk parameters – ApoB by 54.0%, non-HDL cholesterol by 52.3%, and lipoprotein(a) by 25.6%, along with an increase in HDL-C of 4.6% (p<0.0001). Target LDL-C values of <2.6 mmol/L were achieved in 76% of participants taking alirocumab, in comparison with just 2% of participants in the control group, and the target value of <1.8 mmol/L was achieved by as much as 81% of participants taking alirocumab in comparison with 9% in the control group (73).

    Evolocumab

    Evolocumab has proven its high effectiveness and safety in a number of clinical trials. Although the effect of the drug was similar when taken every two and every four weeks (74), larger fluctuations in LDL-C values were present in four-week doses (75). The MENDEL study included a total of 406 participants with hypercholesterolemia and statin intolerance randomized into subgroups taking evolocumab in doses of 70, 105, and 140 mg every 2 weeks, subgroups taking 280, 350, and 420 mg every 4 weeks, and taking ezetimibe. Evolocumab was shown to be greatly superior, and the strongest effect was achieved with the dose of 140 mg every 2 weeks, with no significant side effects (76). The MENDEL-2 study assessed the effectiveness, safety, and tolerability of evolocumab in comparison with ezetimibe and placebo in participants with hypercholesterolemia and high CV risk. Evolocumab reduced LDL-C values by 55-57% more in comparison with placebo and 38-40% more in comparison with ezetimibe (p<0.0001 in both cases) (77). The LAPLACE-TIMI study randomized 57 patients with hypercholesterolemia who were treated with statins into subgroups taking different doses of evolocumab in different intervals (every two and every four weeks). A significant reduction in LDL-C values was achieved after 12 weeks, depending on the evolocumab dose (42-66% when taken every two weeks and 41-50% when taken every four weeks) (78). The LAPLACE-2 study compared evolocumab at different doses and intervals (140 mg every two weeks and 420 once per month) with placebo and ezetimibe in 1896 participants with primary hypercholesterolemia and mixed dyslipidemia that were treated with moderate and high doses of statins. It was found that adding evolocumab to statin treatment reduces LDL-C values significantly more (by 66-75%, depending on the dose) after 10 to 12 weeks in comparison with ezetimibe or placebo (79).

    The DESCARTES study with evolocumab randomized participants into four groups based on CV risk and initial LDL-C values, consisting of participants receiving no additional hypolipidemic therapy, those that in addition to evolocumab also received atorvastatin 10 mg or atorvastatin 80 mg, and participants that received combination therapy consisting of atorvastatin 80 mg and ezetimibe 10 mg in addition to evolocumab. The strong beneficial effects of evolocumab were demonstrated in all four groups. In addition to the beneficial effect on LDL-C values, the study also found a significant reduction in other lipid CV risk factors – apolipoprotein B, lipoprotein(a), and triglycerides (80).

    The GAUSS study examined the effectiveness and tolerability of evolocumab in participants with statin intolerance due to myalgia or myopathy. Evolocumab doses of 280 mg, 350 mg, and 420 mg were tested, as well as evolocumab 420 mg in combination with ezetimibe 10 mg. The effect of evolocumab and the combination treatment was compared with ezetimibe 10 mg. At all doses, evolocumab achieved a significant reduction of LDL-C (40-65%, depending on the dose), with myalgia being present in only 7.4% participants using evolocumab, 20% of participants taking combination evolocumab and ezetimibe treatment, and 3.1% of participants receiving ezetimibe and placebo (81). The GAUSS-2 study also included participants with hypercholesterolemia and statin intolerance (82, 83). The study participants received evolocumab 140 mg every two weeks or 420 mg once per month, while the control group was given ezetimibe. Evolocumab led to a more significant reduction in LDL-C values with a lower incidence of muscle side effects in comparison with ezetimibe (12% for evolocumab, 23% for ezetimibe).

    The RUTHERFORD study demonstrated the high effectiveness of evolocumab in patients with the heterozygous hereditary hypercholesterolemia (167 participants) and high LDL-C despite taking the maximum tolerable statin dose (84). In the study RUTHERFORD-2, a third of the participants took maximum tolerable statin doses, and two thirds received ezetimibe in addition to statins. The participants were randomized into groups that received evolocumab 140 mg every two weeks, evolocumab 420 mg once per month, and the control group taking placebo. In both groups on evolocumab, significant reduction in LDL-C values was achieved after 12 weeks (59.2% for 140 mg and 61.3% for 420 mg doses) (p<0.0001) (85).

    The TESLA study demonstrated the positive effect of evolocumab in patients with homozygous familial hypercholesterolemia treated with maximum tolerable statin doses and other hypolipidemic therapy, with evolocumab treatment achieving an additional LDL-C reduction of 31% after 12 weeks in comparison with placebo, with no significant side effects (86). The OSLER study examined the effect of evolocumab on LDL-C values as well as CV outcomes. A total of 4465 participants were randomized in groups taking 140 mg evolocumab every 2 weeks and 420 mg once per month. During 11.1 months of follow-up, evolocumab reduced LDL-C values by 61% (from an average of 3.1 mmol/L to 1.2 mmol/L, p<0.001). Combined cardiovascular outcomes (mortality, acute coronary syndrome, heart failure, stroke, or TIA) were 2.18% after one year in the control group, and 0.95% in the evolocumab group (HR 0.47; 95% CI 0.28-0.78; p=0.003). Before randomization, most patients were receiving statins (69.7% in the evolocumab group and 70.9% in the control group receiving placebo). (87)

    The GLAGOV study with evolocumab will assess the effects of low LDL-C concentrations on volume regression of atherosclerotic plaque in patients with already diagnosed coronary heart disease (88).

    Bococizumab

    Phase 2b clinical trials with bococizumab achieved significant reduction of LDL-C values in participants with hypercholesterolemia treated with statins. The trials included 354 participants receiving doses of bococizumab 50 mg, 100 mg, and 150 mg every two weeks and doses of 200 mg and 300 mg once per month over the course of 12 weeks. Bococizumab achieved a significant beneficial effect on LDL-C values in comparison with placebo at all doses and intervals, with the strongest reduction in LDL-C being achieved by doses of 150 mg every two weeks and 300 mg once per month (89). Phase 3 clinical trials with bococizumab began in October 2013. They include two studies which primarily assess CV outcomes (SPIRE-I and SPIRE-II) and multiple studies assessing the effects of bococizumab on various lipid parameters, consisting of participants with hypercholesterolemia and high CV risk who were resistant to statins. The primary aim of the SPIRE-I study is to examine whether LDL-C reduction below values recommended in current guidelines has an added beneficial effect on CV outcomes in high-risk participants with initial LDL-C values of 1.81 to 2.59 mmol/L (63). The SPIRE-II study will examine the effectiveness and safety of bococizumab in high-risk patients who have not achieved the target LDL-C value of <2.59mmol/L despite statin therapy or in those with statin intolerance (64). Both of these studies use a starting bococizumab dose of 150 mg every two weeks.

    Additional effects of PCSK9 inhibitors

    In addition to a beneficial effect on LDL-C, PCSK9 inhibitors have also showed other beneficial hypolipidemic effects. Elevated lipoprotein(a) is an independent CV risk factor in patients treated with statins with low LDL-C (90). PCSK9 inhibitors reduced Apo(a) by approximately 30%, implying possible additional cardioprotective effects beyond those achieved by influencing LDL-C values.

    PCSK9 inhibitor safety

    The abovementioned studies also assessed the safety of PCSK9 inhibitors in addition to their effectiveness. There was no significant difference in the incidence of mild and more severe side effects between the assessed monoclonal antibodies and comparative drugs or placebo (79, 80). Less than 2% of participants receiving PCSK9 inhibitors presented with aminotransferase elevations of three or more times the upper limit of normal values. CK elevation was equally rare (91).

    The most common PCSK9 inhibitor side effects were nasopharyngitis, upper respiratory tract infections, flu-like symptoms, and back pain. Reactions at the subcutaneous site of drug injection were present in 2% of the study participants on alirocumab and 4% of those on evolocumab (92). The incidence of treatment termination due to side effects did not differ significantly between PCSK9 inhibitors (alirocumab and evolocumab) and comparison drugs (2-10%) (93).

    Conclusion

    LDL-C values associated with the most positive CV outcomes in persons with high and very high risk are very low (<2.6 and <1.8 mmol/L). Achieving low values is a great challenge, and currently patients often fail to achieve them with hypolipidemic therapy. The discovery of new regulatory mechanisms involved in lipoprotein metabolism has opened the way to new pharmacological approaches for the treatment of elevated LDL-C.

    Discovering the role of PSCK9 in LDL-R expression and LDL-C clearance has spurred the rapid development of new groups of drugs (94). Of the several groups of PCSK9 inhibitors, monoclonal antibodies have seen the most development. Alirocumab, evolocumab, and bococizumab reduce LDL-C values by 50-70%, with good tolerance and high safety. A great challenge in their practical application will undoubtedly be their price, and also in part their subcutaneous application. A beneficial effect on CV outcomes has been found in the OSLER study with evolocumab and is being assessed in a series of clinical trials. The results of these studies will certainly give a clearer picture of the clinical effectiveness and cost-effectiveness of PCSK9 inhibitors in chosen groups of patients.

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