Managing recalcitrant hypercholesterolemia in patients on current best standard of care: efficacy and safety of novel pharmacotherapies

Transcription

1 Clinical Lipidology ISSN: (Print) (Online) Journal homepage: Managing recalcitrant hypercholesterolemia in patients on current best standard of care: efficacy and safety of novel pharmacotherapies Amirhossein Sahebkar & Gerald F Watts To cite this article: Amirhossein Sahebkar & Gerald F Watts (2014) Managing recalcitrant hypercholesterolemia in patients on current best standard of care: efficacy and safety of novel pharmacotherapies, Clinical Lipidology, 9:2, To link to this article: Copyright 2014 Future Medicine Ltd Published online: 18 Jan Submit your article to this journal Article views: 33 View related articles View Crossmark data Citing articles: 1 View citing articles Full Terms & Conditions of access and use can be found at

2 Review Clinical Lipidology Managing recalcitrant hypercholesterolemia in patients on current best standard of care: efficacy and safety of novel pharmacotherapies Residual cardiovascular risk in patients with familial hypercholesterolemia (FH) on high-intensity statin therapy with or without ezetimibe and bile acid sequestrants can potentially be addressed by using agents that can further reduce plasma LDLcholesterol (LDL-C). Two monoclonal antibodies (mabs; evolocumab and alirocumab) against PCSK9, an antisense oligonucleotide against ApoB100 mrna (mipomersen; Kynmaro ) and a synthetic small-molecule inhibitor of MTTP (lomitapide; Juxtapid ) are novel lipid-modifying agents that have been shown in Phase II/III studies in FH patients to lower plasma ApoB and LDL-C with a superior efficiency compared with statin monotherapy. Moreover, these agents can modulate plasma levels of other lipid indices, such as triglycerides, HDL-cholesterol (HDL-C) and lipoprotein(a), that are not substantially affected by statins. The new agents act through increased hepatic clearance of ApoB-containing lipoproteins (PCSK9 mabs) or reduced assembly and secretion of VLDL (mipomersen and lomitapide). Anacetrapib and evacetrapib are newer inhibitors of CETP and can enhance the catabolism of ApoB, accompanied by marked elevations in plasma HDL-C. Mipomersen has been approved by the US FDA, and lomitapide by both the FDA and EMA, as adjunctive to intensive statin therapy and a fat-modified heart-healthy diet in patients with homozygous FH; their use should be accompanied by a careful monitoring of hepatic function owing to the increased risk of hepatic steatosis. PCSK9 mabs and newer CETP inhibitors are still under investigation but their use in clinical trials has been reported to be safe and well tolerated. Amirhossein Sahebkar 1,2 & Gerald F Watts*,3 1 Biotechnology Research Center, Mashhad University of Medical Sciences, Mashhad, Iran 2 Cardiovascular Research Center, Neurogenic Inflammation Research Center, Mashhad University of Medical Sciences, Mashhad, Iran 3 Lipid Disorders Clinic, Cardiovascular Medicine, Royal Perth Hospital & Metabolic Research Centre, School of Medicine & Pharmacology, University of Western Australia, Perth, Australia *Author for correspondence: Tel.: Fax: gerald.watts@uwa.edu.au Keywords: antisense oligonucleotide cardiovascular disease CETP cholesteryl ester transfer protein familial and nonamilial hypercholesterolemia microsomal triglyceride transfer protein MTTP PCSK9 proprotein convertase subtilisin/kexin type 9 The current best standard of care in familial hypercholesterolemia (FH) includes lifestyle modification (adherence to a fat-modified healthy heart diet, smoking cessation and physical activity) plus pharmacotherapy. Reduction of LDL-cholesterol (LDL-C) is the primary target in the management of FH. Plasma ApoB and non-hdl-cholesterol (non-hdl-c) are emerging lipid targets that reflect the total number and cholesterol content of atherogenic particles, respectively, and have been found to be better predictors of coronary heart disease (CHD) compared with LDL-C [1]. Statins are the mainstay of pharmacotherapy for reducing plasma ApoB and LDL-C in FH patients, and may be combined with ezetimibe, bile acid sequestrants or fibrates to intensify LDL-C reduction [2 6]. Combination of statins with other agents such as niacin and probucol is also practiced but the role of these agents in reducing cardiovascular (CV) events is controversial. As the most potent class of hypocholesterolemic agents currently available, statins can typically lower plasma ApoB by 20 40% and LDL-C by 30 50% [7]. However, this LDL-lowering efficacy is not sufficient to reach the lipid targets of ApoB <80 g/l, LDL-C <70 mg/dl and non-hdl-c <100 mg/dl. What is more, recent findings suggest that there is apparently part of /CLP Future Medicine Ltd Clin. Lipidol. (2014) 9(2), ISSN

3 Review Sahebkar & Watts no lower boundary for LDL-C concentrations [8,9]. Hence, it is likely that more stringent target levels for lipids be recommended by future guidelines, and this highlights the need for stronger lipid-modifying agents [7]. The LDL-lowering efficiency of statins can be further argued in patients with FH, as these patients have no or nonfunctional LDL receptors (LDLRs) and thus may not respond properly to the action of statins [10]. Even in patients who achieve recommended LDL-C target by statin therapy, a considerable proportion carry a residual CV risk (defined as the additional coronary risk of the patient despite attaining LDL-C goal by intensive statin therapy) that can be addressed by further reduction of LDL-ApoB [6,11 17]. Moreover, residual CV risk can also be accounted for by elevated plasma concentrations of triglycerides (TG) and reduced HDL-C, two factors that are only modestly affected by statin therapy. Another limitation associated with statin use is the potential of these drugs to induce new-onset diabetes, although the clinical relevance of this adverse effect is yet to be defined given the CV benefits of statins that can outweigh this risk [18]. Finally, statin therapy is associated with dose-dependent muscle-related adverse events that can range between mild episodes of myalgia to life-threatening myotoxicities such as rhabdomyolysis [19]. Since statin-related myopathies are dose dependent, they are more frequent in patients with severe dyslipidemias who are candidates for intensive statin therapy, necessitating alternative approaches for stain-intolerant subjects. We present an update of the clinical trial findings on novel lipid-modifying agents that provide adjunctive therapies to statins for the management of FH patients at high residual CV risk as well as patients intolerant of statins. Literature search PubMed and Scopus databases were searched to retrieve clinical trials on the efficacy and/or safety of novel LDL-C-lowering agents including PCSK9 inhibitors, mipomersen (Kynmaro ), lomitapide (Juxtapid ) and CETP inhibitors. PCSK9 inhibitors Mechanism of action The best-described physiological function of PCSK9 is acceleration of hepatic degradation of LDLR via direct binding and targeting the receptor to the lysosome [20]. Gain-of-function variants of the PCSK9 gene are associated with extremely high plasma LDL-C levels and FH phenotype [21]. Inhibition of PCSK9 at the mrna or protein levels has emerged as a promising strategy to lower LDL-C (Figure 1) [22,23]. The efficacy of fully humanized neutralizing monoclonal antibodies (mabs), evolocumab (formerly known as AMG 145) and alirocumab (formerly known as SAR236553/REGN727), has been investigated in several trials. These fully humanized mabs contain 100% human protein with no risk of immunogenicity, as opposed to chimeric and humanized mabs, which contain 33% and 5 10% mouse sequences, respecitvely, and thus a risk of immune reaction. Findings in FH patients In a Phase I trial, evolocumab (140 mg; three biweekly subcutaneous [sc.] injections) was administered to hetero zygous FH (HeFH) patients. The findings revealed reductions in ApoB, LDL-C, total cholesterol and Lp(a) by 47, 65, 42 and 50%, respectively [25]. In a single-arm trial, patients with homozygous FH (HoFH) were treated with evolocumab (420 mg; sc.) every 4 weeks (for at least 12 weeks) followed by 420 mg every 2 weeks for an additional 12 weeks. The results indicated reductions in ApoB, LDL-C, TG and Lp(a) by 13 16, 13 17, 4 6 and 12 21%, respectively, and elevations in ApoA-I and HDL-C by up to 4% [26]. RUTHERFORD trial was another Phase II study that evaluated the lipid-modifying effects of evolocumab (350 or 420 mg sc. every 4 weeks for a period of 12 weeks) in patients with HeFH refractory to statin therapy. The efficacy of evolocumab in reducing ApoB, LDL-C, non-hdl-c, total cholesterol, VLDL-C and TG were reported to be 35 46, 44 56, 42 53, 37 45, 25 35, and 23 31%, respectively. There were also increases in plasma ApoA-I ( %) and HDL-C (7 8%) following treatment with evolocumab [27]. In HeFH patients taking atorvastatin, there were significant reductions in ApoB (34 45%), LDL-C (41 58%), total cholesterol (28 37%), TG (15 36%) and Lp(a) (14 18%), and significant elevations in ApoA-I (17%) and HDL-C (23%) following treatment with alirocumab (three sc. injections of either 50, 100 or 150 mg at days 1, 29 and 43) [28]. In another study on HeFH patients on high-dose statin with or without ezetimibe, treatment with alirocumab ( mg sc. every 4 weeks or 150 mg sc. every 2 weeks over 12 weeks) resulted in reductions in ApoB (15 44%), LDL-C (18 57%), non-hdl-c (16 47%) and total cholesterol (10 36%) but not TG and Lp(a). There were also significant elevations in ApoA-I (by up to 4%) and HDL-C (by up to 10%) [29]. 222 Clin. Lipidol. (2014) 9(2) Findings in non-fh patients The lipid-modifying effects of evolocumab have been shown in several trials and in different populations. In a Phase I trial, ranging doses of evolocumab were administered to healthy subjects (7 420 mg sc. or 21 or 420 mg intravenously [iv.]; single dose), hyperfuture science group

4 Managing recalcitrant hypercholesterolemia in patients on current best standard of care Review Liver mabs against PCSK9 Hepatocyte Secretion Autocatalysis sirna against PCSK9 PCSK9 synthesis Nucleus Lysosomal degradation LDL receptor synthesis LDL receptor LDL cholesterol PCSK9 cleaved PCSK9 uncleaved Figure 1. PCSK9 accelerates hepatic degradation of LDL receptor via direct binding and targeting the receptor to lysosome. Due to its pivotal role in LDL metabolism, inhibition of PCSK9 has emerged as a promising strategy to lower LDL-cholesterol. sirnas and mabs have been developed to block mrna transcription and receptor binding activity of PCSK9, respectively. mab: Monoclonal antibody. Reproduced with permission from [24]. Rightslink license number: cholesterolemic patients on low- to moderate-dose statin therapy (14 or 35 mg once weekly, six doses; 140 or 280 mg every 2 weeks, three doses; 420 mg every 4 weeks, two doses) and hypercholesterolemic patients on high-dose statin therapy (140 mg; three biweekly sc. injections) [25]. The findings revealed significant reductions in plasma lipid concentrations by evolocumab in all groups: in healthy subjects, there were significant reductions in plasma ApoB (6 55%), LDL-C (6 64%) and total cholesterol (3 43%); in hypercholesterolemic patients on low-to-moderatedose statins, there were significant reductions in plasma ApoB (15 54%), LDL-C (22 75%), total cholesterol (14 45%) and Lp(a) (16 50%); and in hypercholesterolemic patients on high-dose statin therapy, there were reductions in ApoB, LDL-C, total cholesterol and Lp(a) by 50 and 62%, 35 and 43%, 35 and 42%, and 43 and 50%, respectively [25]. In the Phase II GAUSS trial in statin-intolerant patients, the effects of 12-week treatment with evolocumab (420 mg sc. every 4 weeks) plus ezetimibe (10 mg/day) was compared with those of ezetimibe (10 mg/day) monotherapy. There were significant reductions in ApoB (37%), LDL-C (47%), non-hdl-c (45%), total cholesterol (34%), VLDL-cholesterol (VLDL-C; 25%) and Lp(a) (21%), and elevations in ApoA-I (10%) and HDL-C (13%) in the group receiving evolocumab ezetimibe combination versus ezetimibe monotherapy [30]. In the same trial, comparison of monotherapy with escalated doses of evolocumab ( mg sc. every 4 weeks) with ezetimibe monotherapy (10 mg/day) revealed a significantly greater effect of the former treatment in reducing ApoB (22 30%), LDL-C (26 36%), non- HDL-C (25 34%), total cholesterol (19 27%) and Lp(a) (12 18%), and elevating ApoA-I (7 9%) and HDL-C (7 8%) [30]. In the LAPLACE-TIMI 57 trial in hypercholesterolemic patients on a stable dose of statin, administration of evolocumab at ranging doses ( mg sc. every 2 weeks or mg sc. every 4 weeks over a period of 12 weeks) caused significant reductions in ApoB (34 56%), LDL-C (42 66%), non-hdl-c (38 61%), total cholesterol (28 48%), VLDL-C (21 44%) and TG (13 34%), and increased ApoA-I (by up to 5%) and HDL-C (2 8%) [31]. The efficacy of evolocumab in treatment-naive hypercholesterolemic subjects (who have not taken lipid-lowering medications before) was tested in the MENDEL trial. The administration protocol comprised evolocumab at 223

5 Review Sahebkar & Watts a dose of mg sc. every 2 weeks or mg sc. every 4 weeks over a period of 12 weeks. The results revealed significant reductions in ApoB (32 44%), LDL-C (37 52%), non-hdl-c (35 47%), total cholesterol (25 35%), VLDL-C (10 36%), TG (2 12%) and Lp(a) (11 29%), and elevations in ApoA-I and HDL-C by up to 10% [32]. The OSLER study is the only Phase III trial completed with evolocumab. This was a 52-week trial assigning hypercholesterolemic patients from the abovementioned trials (GAUSS, LAPLACE- TIMI 57, MENDEL and RUTHERFORD) to either evolocumab (420 mg sc. every 4 weeks) plus standard of care, or standard of care alone. The results revealed reductions in ApoB (38%), LDL-C (50%), non-hdl- C (44%), total cholesterol (34%), VLDL-C (18%), TG (8%) and Lp(a) (21%), while there were increases in ApoA-I (4%) and HDL-C (5%) [33]. In two Phase I studies, the efficacy of alirocumab was tested in healthy subjects and non-fh hypercholesterolemic patients. In healthy subjects, administration of a single dose of alirocumab via the iv. ( mg/kg) or sc. ( mg) route caused a dose-dependent 28 65% and 32 46% reduction in plasma LDL-C, respectively [28]. There were also significant lipid-modifying effects after treatment with multiple doses of alirocumab (three injections of either 50, 100 or 150 mg sc. administered at days 1, 29 and 43 of the study) in non-fh hypercholesterolemic patients on atorvastatin. The magnitude of reductions in ApoB, LDL-C, total cholesterol and Lp(a) were 30 48, 38 65, and 30%, respectively, while there was no effect on TG. The increase in plasma ApoA-I and HDL-C were also reported to be 11 and 8 14%, respectively. Finally, in non-fh hypercholesterolemic patients who were not taking atorvastatin, three injections of alirocumab (150 mg) was found to reduce ApoB (45%), LDL-C (57%), total cholesterol (43%) and Lp(a) (44%), while having no effect on TG, ApoA-I and HDL-C [28]. Later, the efficacy of alirocumab (dose range: mg sc. every 2 weeks or mg sc. every 4 weeks over a period of 12 weeks) was tested in a Phase II study in patients with primary hypercholesterolemia on stable statin therapy. There were significant reductions in ApoB (29 58%), LDL-C (35 67%), non-hdl-c (32 60%), total cholesterol (21 43%), TG (15 29%) and Lp(a) (8 29%), and elevations in HDL-C (5 9%) but not ApoA-I [31]. In another trial in patients with the same characteristics, the efficacy of alirocumab (80 mg/week sc. for a period of 8 weeks) was evaluated in combination with low- or high-dose atorvastatin compared with high-dose atorvastatin monotherapy. The results indicated that addition of alirocumab to atorvastatin (either low or high dose) has a superior efficacy compared with high-dose statin monotherapy. Reported reduction rates in ApoB, LDL-C, non-hdl- C, total cholesterol, TG and Lp(a) were 46, 56, 42, 30, 13 and 28%, respectively. There was also a 10% elevation in plasma HDL-C but the changes in ApoA-I concentrations were not found to be significant [34]. Given the promising abovementioned lipid-modifying effects of PCSK9 mabs in Phase II/III trials, two outcome trials are currently underway to assess the impact of alirocumab (ODYSSEY Outcomes; NCT ) and evolocumab (FOURIER; NCT ) are in patients with acute coronary syndrome (primary outcome measure: occurrence of composite end point of CHD death, nonfatal myocardial infarction [MI], fatal and nonfatal ischemic stroke, and unstable angina requiring hospitalization) and clinically evident CV disease (primary outcome measure: the primary end point is the time to CV death, MI, hospitalization for unstable angina, stroke or coronary revascularization; whichever occurs first), respectively. Safety Both evolocumab and alirocumab have been reported to be safe and well tolerated, with no report of causing hepatic transaminase elevation or hepatotoxicity. Nonserious injection site reactions, rash and hypersensitivity have been reported as the most common adverse reactions following the use of PCSK9 mabs. In the GAUSS trial, the incidence of myalgia in statin-intolerant subjects receiving evolocumab was reported to range between 3.0 and 16% with different doses, and reach 20% in the group receiving evolocumab ezetimibe combination [30]. RN316 RN316 (also known as PF ) is a humanized PCSK9 mab that has recently been tested for its efficacy and safety in clinical trials. In a Phase I study, multiple iv. doses of RN316 ( mg/kg/week) were administered to primary hypercholesterolemic subjects for a period of 4 weeks. Significant reductions in plasma LDL-C (by up to 90%) and TG, and elevation of HDL-C concentrations were reported [35]. In a later Phase II trial, hypercholesterolemic subjects receiving high-to-maximal doses of statins were assigned to RN316 (dose range: mg/kg; iv.) every 4 weeks for a period of 12 weeks. RN316 was found to significantly lower plasma LDL-C at higher doses, and increase HDL-C concentrations, but none of the tested doses did significantly decrease plasma TG concentrations [36]. In both of the aforementioned trials, treatment with RN316 was reported to be well tolerated and adverse events were infrequent, mild and transient. A Phase III trial (NCT ) is currently in progress evaluating the efficacy of RN316 in 224 Clin. Lipidol. (2014) 9(2)

6 Managing recalcitrant hypercholesterolemia in patients on current best standard of care Review reducing the occurrence of major CV events (including hospitalization for unstable angina needing urgent revascularization; a composite end point of CV death, nonfatal MI, and nonfatal stroke; and a composite end point of all-cause death, nonfatal MI, and nonfatal stroke, and hospitalization for unstable angina needing urgent revascularization) in high-risk patients. ALN-PCS ALN-PCS is a sirna that targets PCSK9 mrna and has been shown in preclinical studies to efficiently reduce plasma PCSK9 and LDL-C. The efficacy and safety of ALN-PCS has been investigated by a Phase I trial in healthy subjects with raised cholesterol who were not on lipid-lowering therapy. ALN-PCS was administered at a single iv. dose of 0.015, 0.045, 0.090, 0.150, or mg/kg, and a dose-dependent reduction in plasma LDL-C was observed ranging from 6 to 40%, which was accompanied by corresponding reductions in plasma PCSK9 by 31 70% [37]. No serious adverse event was reported from ALN-PCS after 6 months of follow-up in the only conducted trial with this agent [37]. Mipomersen Mechanism of action Mipomersen is an antisense inhibitor of ApoB100 mrna translation, and acts by binding to the complementary sequence within the target mrna and subsequently causing degradation of target mrna via activation of RNaseH [38]. Downregulation of ApoB100 expression by mipomersen results in decreased secretion of VLDL from liver and a reduction in plasma levels of all ApoB-containing lipoproteins (Figure 2). Findings in FH patients The efficacy of mipomersen has been demonstrated in both HeFH and HoFH patients. In HeFH patients on stable lipid-lowering therapy, mipomersen can lower ApoB (7 32%), LDL-C (11 34%), non-hdl- C (7 30%), total cholesterol (7 25%), TG (9%) and Lp(a) (28%), but has no significant effect on ApoA- I and HDL-C. These results were obtained by studies administering mipomersen at escalated doses of mg (eight sc. injections during 6 weeks), or a fixed dose of 200 mg/week over 13 weeks [40]. In HoFH patients on aggressive lipid-lowering therapy, weekly administration of mipomersen (200 mg weekly; sc.) for a period of 26 weeks resulted in reductions in ApoB, LDL-C, non-hdl-c, total cholesterol, VLDL- C, TG and Lp(a) by 24, 21, 22, 19, 17, 15 and 23%, respectively. There was also a significant elevation in plasma HDL-C by 11%, yet the effect on ApoA-I was not significant [41]. Findings in non-fh patients In a Phase I trial, healthy subjects with mild hypercholesterolemia were injected with a single sc. dose of mg mipomersen, followed by a 4-week multiple-dosing regimen with the same assigned dose (comprising three alternate-day iv. infusions in the first week followed by three weekly sc. doses). Mipomersen dose-dependently reduced ApoB, LDL- C, total cholesterol, VLDL-C and TG concentrations by up to 47, 43, 40, 60 and 43%, respectively [42]. In hypercholesterolemic subjects receiving stable statin therapy, seven sc. doses of mg mipomersen over 5 weeks reduced ApoB, LDL-C, non-hdl-c, total cholesterol and TG by up to 52, 49, 49, 38 and 41%, respectively. However, no effect on HDL-C and VLDL-C was reported [43]. In subjects with untreated mild-to-moderate hyperlipidemia, multiple-dose treatment with ranging doses of mipomersen ( mg/week; sc.) for 26 weeks caused marked and dosedependent reductions in plasma ApoB, LDL-C, non- HDL-C, VLDL-C, TG and Lp(a) levels by up to 71, 71, 66, 61, 58 and 49%, respectively [44]. In high-risk statin-intolerant patients, treatment with mipomersen (200 mg/week; sc.) for a period of 26 weeks reduced ApoB (42%), LDL-C (45%), non-hdl-c (44%), total cholesterol (35%), VLDL-C (27%), TG (22%) and Lp(a) (27%), while ApoA-I and HDL-C levels remained unchanged [45]. In a Phase III study in patients with severe non-fh at high CV risk despite receiving maximally tolerated lipid-lowering regimen, weekly treatment with mipomersen (200 mg; sc.) for a period of 26 weeks resulted in reduction of ApoB, LDL-C, non-hdl-c, total cholesterol, VLDL-C, TG and Lp(a) by 33, 33, 33, 23, 14, 14 and 26%, respectively, but plasma levels of ApoA-I were reduced and HDL-C levels were not affected [46]. Safety The most common adverse events following mipomersen use are self-limiting injection site reactions and systemic reactions including fever, nausea and malaise. The main safety concern with mipomersen is increased hepatic TG accumulation and development of steatosis, which is due to decreased efflux of TG from liver. Increased hepatic TG by mipomersen is usually accompanied by a rise in plasma levels of transaminases. Although hepatic fat accumulation following mipomersen use is reversible, and its progression into frank fibrosis has not been demonstrated, a cautionary policy called Risk Evaluation and Mitigation Strategy (REMS) has been launched by the manufacturer and FDA to provide prescribers and pharmacists with training and certification

7 Review Sahebkar & Watts Hepatocyte Nucleus Mipomersen (antisense RNA) Mipomersen Mipomersen RNase H ApoB mrna ApoB ApoB DNA LDL Figure 2. Mechanism of action of mipomersen. (A) Mipomersen is a single-stranded antisense RNA designed to correspond to the AopB gene as a target for gene silencing. (B) mirna antisense duplex is formed: once mipomersen reaches the hepatocyte, it penetrates the cell membrane and nucleus (unclearmechanism) and binds to ApoB mrna with a high degree of fidelity. (C) RNaseH cleaves mrna: after hybridization with target mrna, the mipomersen mrna duplex is recognized and cleaved by endogenous RNaseH, en enzyme involved in endogenous DNA replication/repair. (D) Cleavage of ApoB mrna results in decreased ApoB synthesis and decreased LDL. Reproduced with permission from [39]. Rightslink license number: Lomitapide Mechanism of action MTTP is an enzyme involved in the assembly and secretion of ApoB-containing lipoproteins, and is localized in the endoplasmic reticulum of enterocytes and hepatocytes [47]. Patients with genetic deficiency of functional MTTP, known as abetalipoproteinemia, have extremely low levels of plasma VLDL and LDL [48]. Lomitapide, a small, synthetic molecule, is the most advanced MTTP inhibitor and reduces the secretion of chylomicrons and VLDL into circulation, which, in turn, cause reduced formation of LDL particles (Figure 3). Findings in FH patients Lomitapide was tested for the first time in a small uncontrolled trial in HoFH patients who were assigned to different doses of the agent (0.03, 0.1, 0.3 and 1.0 mg/kg/day; orally), each for a 4-week period. Analysis of lipids indicated dose-dependent reductions in ApoB, LDL-C, total cholesterol and TG by up to 56, 51, 58 and 65%, respectively, with no significant elevation in ApoA-I or HDL-C [50]. In an uncontrolled Phase III trial in HoFH patients, lomitapide was administered orally at a starting doses of 5 mg/day and was escalated (in 4-week intervals) to a maximum dose of 60 mg/day. After completion of the 26-week treatment period, plasma ApoB, LDL-C, non-hdl-c, total cholesterol, VLDL-C, TG and Lp(a) were found to be decreased by 43 49, 38 50, 39 50, 35 46, 28 45, and 15 19%, respectively, but there was no positive effect on either ApoA-I or HDL-C [51]. Findings in non-fh patients Escalated doses of lomitapide (5 10 mg/day; orally) with or without ezetimibe have been tested in patients with moderate hypercholesterolemia for a period of 12 weeks. The results indicated a greater efficacy of lomitapide versus ezetimibe, which was equivalent to 9, 10, 10, 11 and 20% for ApoB, LDL-C, non-hdl-c, total cholesterol and Lp(a), respectively. There were also a 226 Clin. Lipidol. (2014) 9(2)

8 Managing recalcitrant hypercholesterolemia in patients on current best standard of care Review greater efficacy of lomitapide versus ezetimibe in elevating ApoA-I and HDL-C, by 10 and 12%, respectively. The lipid-modifying effects were augmented when lomitapide was administered in combination with ezetimibe. However, in none of the treatment protocols tested was there any significant change in plasma TG levels [52]. Safety The most common adverse events associated with lomitapide use are gastrointestinal complications, which can be minimized by dose titration and adherence to lowfat diet. As with mipomersen, the use of lomitapide is concerned by the likelihood of hepatic fat accumulation and subsequent development of steatosis and hepatotoxicity. Elevation of hepatic transaminases has been reported in approximately 30% of subjects receiving lomitapide [51]. Therefore, plasma transaminase levels should be regularly monitored during treatment with lomitapide. Lomitapide is available only through the REMS program and is administered by certified centers and physicians that are trained to monitor drug-related adverse events cautiously. CETP inhibitors Mechanism of action CETP plays a pivotal role in lipoprotein metabolism by facilitating the bidirectional exchange of cholesteryl ester in HDL for TG in ApoB-containing particles, mainly VLDL and LDL. The net effect of this exchange is increased formation of small dense LDL particles [53]. Hence, inhibition of CETP is being sought as a viable approach for concomitant reduction of plasma LDL-C and elevation of HDL-C (Figure 4). Early CETP inhibitors were discontinued from further development owing to the off-target toxicity and subsequent increase in mortality (torcetrapib) and futility (dalcetrapib). However, newer agents (anacetrapib and evacetrapib) have been shown to be more efficacious than dalcetrapib and lack the off-target toxicities of torcetrapib in the early trials [54]. Findings in FH patients To date, there has been no published result on the lipidmodifying effects of new CETP inhibitors in patients with FH. However, there is an ongoing Phase III trial (NCT ) investigating the safety and efficacy of 12-week treatment with anacetrapib (10 mg/day) as an add-on to ongoing statin therapy in HeFH patients. The results of this study are awaited with interest. MTTP X MTTP X apob48 MTTP inhibitor apob100 MTTP inhibitor VLDL Chylomicron Enterocyte Hepatocyte Figure 3. MTTP facilitates the transfer of triglycerides to ApoB48 (in enterocytes) and ApoB100 (in hepatocytes), thereby mediating formation of chylomicrons and VLDL particles, respectively. Inhibition of MTTP reduces hepatic secretion of VLDL-ApoB which, in turn, reduces LDL formation. Reproduced with permission from [49]. Rightslink license number: Findings in non-fh patients In a Phase I study, subjects with primary hypercholesterolemia were assigned to escalated doses ( mg/day; orally) of anacetrapib for a period of 28 days. The results indicated a significant reduction in plasma ApoB (by up to 27%), LDL-C (by up to 41%) and an elevation of ApoA-I (by up to 47%) and HDL-C (by up to 130%) [55]. In patients with or at high risk of CHD, anacetrapib (100 mg/day orally) was administered for a period of 18 months. The results of this long-term study revealed significant reductions in ApoB (18 21%), LDL-C (36 40%), non-hdl-c (29 32%) and Lp(a) (36 39%), while ApoA-I and HDL-C were increased by and %, respectively [56]. In another trial in subjects with primary hypercholesterolemia or mixed dyslipidemia, the effects of anacetrapib was investigated both as monotherapy and in combifuture science group 227

9 Review Sahebkar & Watts ApoA-I Cholesterol-poor HDL SR-BI C Liver CE CE CE TRL C Macrophage CE CE Cholesterol-rich HDL TG TG CETP CETP inhibitors HDL remodeling Figure 4. Role of CETP in the metabolism of HDL. Newly secreted ApoA-I binds to phospholipids and forms C-poor HDL particles that subsequently become lipidated by C molecules effluxed from macrophages. After esterification, CE content of HDL is exchanged for triglycerides from TRLs, a reaction that is facilitated by CETP. Residual CE would be removed through the interaction of HDL with hepatic SR-BI. C: Cholesterol; CE: Cholesteryl ester; TRL: Triglyceride-rich lipoprotein. Reproduced with permission from [49]. Rightslink license number: nation with atorvastatin. As monotherapy, 8 weeks of treatment with anacetrapib at an oral daily dose of mg reduced ApoB and LDL-C by up to 32 and 39%, respectively, and dramatically increased HDL-C by 133%. Similar efficacy in altering plasma LDL-C and HDL-C was observed upon combination of anacetrapib with atorvastatin [57]. In none of the abovementioned studies was there a significant change in plasma TG no total cholesterol concentrations. In Japanese dyslipidemic patients, 8-week treatment with anacetrapib ( mg/day; orally) was found to reduce plasma LDL-C by 12 32% and increase HDL-C concentrations by %. In the same study, combination therapy with anacetrapib ( mg/day) plus atorvastatin (10 mg/day) was found to be associated with greater reductions in plasma LDL-C and elevations in HDL-C compared with atorvastatin monotherapy [58]. In the DEFINE trial, anacetrapib was administered to patients with or at high risk of CHD at a dose of 100 mg/day (orally) for 72 weeks. Anacetrapib was reported to decrease plasma ApoB (10%), LDL-C (19%) and non-hdl-c (18%), and increase HDL-C (73%) and ApoA-I (24%) concentrations [59]. Another CETP inhibitor, evacetrapib, has been shown to exert significant lipid-modifying effects in dyslipidemic patients both as monotherapy or combination therapy with statins. Monotehrapy with evacetrapib ( mg/day orally) for 12 weeks resulted in dose-dependent reductions in LDL-C (18 40%) and TG (12 20%), and elevation of HDL-C by % [60]. Evacetrapib (100 mg/day) in combination with different statins (simvastatin, atorvastatin or rosuvastatin) was reported to reduce LDL-C (11 14%) and increase HDL-C (78 88%), but had no superior effect on TG compared with statin monotherapy [60]. Safety Anacetrapib and evacetrapib have been reported to have a good safety and tolerability profile and lack the serious adverse events seen with torcetrapib, namely imbalances in serum electrolytes and aldosterone, and increased blood pressure. However, there are preclinical and clinical data indicating accumulation of anacetrapib in adipose tissue, a phenomenon that can prolong the elimination of drug from the body [61]. The clinical significance of this phenomenon deserves further investigation. There is an ongoing 76-week worldwide multicenter randomized controlled trial investigating the tolerability and efficacy of anacetrapib on top of statin therapy in patients with CHD or a CHD-equivalent disease (NCT ). The long-term efficacy of evacetrapib in reducing CV end points is currently under investigation by the ACCELERATE trial in patients with a high risk of vascular disease (NCT ). 228 Clin. Lipidol. (2014) 9(2)

10 Managing recalcitrant hypercholesterolemia in patients on current best standard of care Review Conclusion & future perspective Clinical use of the agents described here in this review been shown to be associated with significant decrements in plasma ApoB-containing lipoproteins (Table 1) via either blocking the endogenous synthesis (mipomersen) or hepatic efflux (lomitapide) of ApoB, or enhancement of hepatic clearance of ApoB from plasma (PCSK9 mabs and CETP inhibitors) [62,63]. This ApoB- and LDL-lowering efficacy might have special implications for the management of FH patients, who are at a very high risk of premature CHD despite receiving high-intensity pharmacotherapy according to the best standard of care guidelines [2,7,9,64 65], or those patients who are intolerant of statins due to the development of myopathic and hepatotoxic adverse reactions. While the efficiency of PCSK9 mabs, mipomersen and lomitapide has been demonstrated in FH patients, there has been no completed trial with new CETP inhibitors in FH, but one trial is underway (NCT ). An additional effect of PCSK9 mabs, mipomersen and lomitapide is reduction of plasma Lp(a). Lp(a) possesses thrombotic, antifibrinolytic and atherogenic properties and has a higher affinity to the arterial subendothelial space components [66] and oxidized phospholipids [67] compared with other atherogenic particles, thus serving as a strong predictor of CV disease [68]. In FH patients, elevated Lp(a) significantly increases the risk of premature CHD irrespective of LDL-C levels [69 71]. Treatment of hyperlipoproteinemia(a) is challenging owing to the lack of an effective and well-tolerated treatment. Treatment with mipomersen and lomitapide can reduce plasma TG by a greater extent compared with statins. TG-rich lipoprotein remnants have strong atherogenic properties owing to their capacity to penetrate and remain in the subendothelial space, and enhancement of foam cell formation, vascular inflammation and oxidative stress [72 74]. Hence, hyportriglyceridemic effects of these agents may contribute to further reduction in CV risk in statin-treated patients, even in those attaining LDL-C goals. Benefits of combination therapy with statins and novel hypolipidemic agents described here appear to exceed a simple additive effect. This particularly applies to PCSK9 inhibitors as elevation of PCSK9 by statins is an important reason for the limited LDL-C-lowering efficacy of stains at higher doses. Dose escalation studies have indicated that the LDL-C-lowering effect of statins is increased by only 6% upon dose doubling, a phenomenon known as the rule of 6% [75]. Therefore, coadministration with PCSK9 inhibitors not only exerts its own reducing effect but also potentiates the efficacy of statin therapy. Future studies are warranted to explore whether antidyslipidemic effects of PCSK9 mabs, mipomersen, Table 1. Summary of changes in plasma concentrations of lipids, lipoproteins and ApoB following treatment with new lipid-modifying agents. Agent Mechanism of action Populations studied Change (%) ApoB LDL-C Non-HDL-C VLDL-C TC TG HDL-C Lp(a) Evolocumab mab against PCSK9 Healthy subjects, primary hypercholesterolemia HeFH, HoFH Alirocumab mab against PCSK9 Healthy subjects, primary N hypercholesterolemia, HeFH Mipomersen ApoB ASO Healthy subjects, statin-intolerant N subjects, primary hypercholesterolemia, HeFH, HoFH Lomitapide MTTP inhibitor Primary hypercholesterolemia, HoFH N N N Anacetrapib CETP inhibitor Primary hypercholesterolemia mixed dyslipidemia, low HDL-C, patients at high risk of CHD N Evacetrapib CETP inhibitor Nonfamilial dyslipidemia N N N N ASO: Antisense oligonucleotide; CHD: Coronary heart disease; HDL-C: HDL-cholesterol; HeFH: Heterozygous familial hypercholesterolemia; HoFH: Homozygous familial hypercholesterolemia; LDL-C: LDL-cholesterol; Lp(a): Lipoprotein(a); mab: Monoclonal antibody; N: Not reported or not significant; TC: Total cholesterol; TG: Triglycerides; VLDL-C: VLDL-cholesterol

11 Review Sahebkar & Watts lomitapide and CETP inhibitors could reduce the incidence of CV end points. The FDA has approved the use of mipomersen and lomitapide, the latter also approved by EMA, for use as adjunctive to statin therapy in patients with HoFH. Mipomersen and lomitapide are likely to be superior to LDL apheresis in terms of cost, availability and patient compliance and acceptability, and preliminary data show the efficacy of these agents would change between patients on and off apheresis [51], but whether these agents can fully replace apheresis therapy needs to be answered by future studies. The broader application of mipomersen and lomitapide in hyperlipidemias is impeded by the associated hepatic adverse events associated with the use of these agents. However, PCSK9 mabs and newer CETP inhibitors may find wider applications in the management of lipid disorders beyond FH such as familial combined hyperlipidemia, familial hyperlipoproteinemia(a) and diabetic dyslipidemia, as well as in statin-intolerant or statin-resistant patients at high risk of CV disease [30,45]. Financial & competing interests disclosure GF Watts has received lecture fees and has served as an advisor to Sanofi-Aventis, Genzyme and Amgen Corporation. The authors have no other relevant affiliations or financial involvement with any organization or entity with a financial interest in or financial conflict with the subject matter or materials discussed in the manuscript apart from those disclosed. No writing assistance was utilized in the production of this manuscript Executive summary Background Elevated plasma levels of ApoB-containing lipoproteins are associated with an increased risk of premature coronary heart disease in patients with familial hypercholesterolemia (FH). Statins can typically lower plasma ApoB by 20 40% and LDL-cholesterol (LDL-C) by 30 50%, which is not sufficient to reach the lipid targets of ApoB <80 g/l, LDL-C <70 mg/dl and non-hdl-c <100 mg/dl. Even in patients who achieve the recommended LDL-C target by statin therapy, a considerable proportion carry a residual cardiovascular risk that can be addressed by further reduction of LDL-ApoB. PCSK9 inhibitors Inhibition of PCSK9 at the mrna or protein levels is an efficacious strategy to lower LDL by increasing the density of LDL receptors on the hepatocyte surface. Evolocumab reduces plasma ApoB and LDL-C in FH patients by up to 47 and 65%, respectively, and in non-fh patients, by up to 56 and 75%, respectively. Alirocumab reduces plasma ApoB and LDL-C in FH patients by up to 45 and 58%, respectively, and in non-fh patients, by up to 58 and 67%, respectively. ALN-PCS reduces plasma LDL-C in non-fh hypercholesterolemic subjects by up to 40%. Mipomersen Mipomersen acts through neutralization of ApoB100 mrna and reduces hepatic VLDL secretion, resulting in a reduction in the plasma levels of all ApoB-containing lipoproteins. Evolocumab reduces plasma ApoB and LDL-C in FH patients by up to 32 and 34%, respectively, and in non-fh patients, by up to 71%. Lomitapide Inhibition of MTTP by lomitapide reduces the assembly and secretion of VLDL, which in turn causes a reduction in plasma levels of all ApoB-containing lipoproteins. Lomitapide has been shown to reduce plasma ApoB and LDL-C by 56 and 51%, respectively. Lomitapide has been shown to be superior to ezetimibe in non-fh subjects, causing additionl reductions in ApoB (9%) and LDL-C (10%) when added to ezetimibe. CETP inhibitors Inhibition of CETP results in the reduction of plasma LDL-C and elevation of HDL-C. Anacetrapib and evacetrapib are new CETP inhibitors that have been shown to reduce ApoB and LDL-C by up to 27 and 41%, respectively, in non-fh subjects. The impact of anacetrapib in FH patients is being investigated in the Phase III trial NCT Conclusion & future perspective Residual cardiovascular risk in FH patients can be addressed by new ApoB-lowering agents including PCSK9 and CETP inhibitors, mipomersen and lomitapide. PCSK9 inhibitors, mipomersen and lomitapide can lower lipoprotein(a), which has strong atherothrombotic properties. Treatment with mipomersen and lomitapide can reduce plasma triglycerides, and CETP inhibitors can increase plasma HDL-C by greater efficacies compared with statins. Mipomersen and lomitapide have been approved as adjunctive to statin therapy in patients with homozygous FH, but their broader application is limited due to the potential to increase hepatic fat accumulation. 230 Clin. Lipidol. (2014) 9(2)

12 Managing recalcitrant hypercholesterolemia in patients on current best standard of care Review References Papers of special note have been highlighted as: of interest; of considerable interest 1 Pischon T, Girman CJ, Sacks FM, Rifai N, Stampfer MJ, Rimm EB. Non-high-density lipoprotein cholesterol and apolipoprotein B in the prediction of coronary heart disease in men. Circulation 112(22), (2005). 2 Watts GF, Gidding S, Wierzbicki AS et al. Integrated guidance on the care of familial hypercholesterolaemia from the International FH Foundation. Int. J. Cardiol. 171(3), (2014). A comprehensive review describing the latest guidelines for the identification and management of familial hypercholesterolemia (FH). 3 Nordestgaard BG, Chapman MJ, Humphries SE et al. Familial hypercholesterolaemia is underdiagnosed and undertreated in the general population: guidance for clinicians to prevent coronary heart disease: Consensus Statement of the European Atherosclerosis Society. Eur. Heart J. 34(45), (2013). A comprehensive review describing the latest guidelines for the identification and management of FH. 4 WHO. Familial hypercholesterolaemia: report of a WHO consultation. WHO, Paris, France (1997). 5 Fahed AC, Nemer GM. Familial hypercholesterolemia: the lipids or the genes? Nutr. Metab. (Lond.) 8(1), 23 (2011). 6 Neefjes LA, Ten Kate GJ, Rossi A et al. CT coronary plaque burden in asymptomatic patients with familial hypercholesterolaemia. Heart 97(14), (2011). 7 Jones P, Kafonek S, Laurora I, Hunninghake D. Comparative dose efficacy study of atorvastatin versus simvastatin, pravastatin, lovastatin, and fluvastatin in patients with hypercholesterolemia (the CURVES study). Am. J. Cardiol. 81(5), (1998). 8 Wiviott SD, Cannon CP, Morrow DA, Ray KK, Pfeffer MA, Braunwald E. Can low-density lipoprotein be too low? The safety and efficacy of achieving very low low-density lipoprotein with intensive statin therapy: a PROVE IT-TIMI 22 substudy. J. Am. Coll. Cardiol. 46(8), (2005). 9 Hsia J, MacFadyen JG, Monyak J, Ridker PM. Cardiovascular event reduction and adverse events among subjects attaining low-density lipoprotein cholesterol <50 mg/dl with rosuvastatin. The JUPITER trial (Justification for the Use of Statins in Prevention: an Intervention Trial Evaluating Rosuvastatin). J. Am. Coll. Cardiol. 57(16), (2011). 10 Choumerianou DM, Dedoussis GV. Familial hypercholesterolemia and response to statin therapy according to LDLR genetic background. Clin. Chem. Lab. Med. 43(8), (2005). 11 Pijlman AH, Huijgen R, Verhagen SN et al. Evaluation of cholesterol lowering treatment of patients with familial hypercholesterolemia: a large cross-sectional study in The Netherlands. Atherosclerosis 209(1), (2010). 12 Versmissen J, Oosterveer DM, Yazdanpanah M et al. Efficacy of statins in familial hypercholesterolaemia: a long term cohort study. BMJ 337, a2423 (2008). 13 Neil A, Cooper J, Betteridge J et al. Reductions in all-cause, cancer, and coronary mortality in statin-treated patients with heterozygous familial hypercholesterolaemia: a prospective registry study. Eur. Heart J. 29(21), (2008). 14 Harada-Shiba M, Sugisawa T, Makino H et al. Impact of statin treatment on the clinical fate of heterozygous familial hypercholesterolemia. J. Atheroscler. Thromb. 17(7), (2010). 15 Raal FJ, Pilcher GJ, Panz VR et al. Reduction in mortality in subjects with homozygous familial hypercholesterolemia associated with advances in lipid-lowering therapy. Circulation 124(20), (2011). 16 Elis A, Zhou R, Stein EA. Effect of lipid-lowering treatment on natural history of heterozygous familial hypercholesterolemia in past three decades. Am. J. Cardiol. 108(2), (2011). 17 Fruchart JC, Sacks F, Hermans MP et al. The Residual Risk Reduction Initiative: a call to action to reduce residual vascular risk in patients with dyslipidemia. Am. J. Cardiol. 102(10 Suppl.), 1K 34K (2008). 18 Bell DS, Dinicolantonio JJ, O Keefe JH. Is statin-induced diabetes clinically relevant? A comprehensive review of the literature. Diabetes Obes. Metab. doi: /dom (2013) (Epub ahead of print). 19 Harper CR, Jacobson TA. The broad spectrum of statin myopathy: from myalgia to rhabdomyolysis. Curr. Opin. Lipidol. 18(4), (2007). 20 Artenstein AW, Opal SM. Proprotein convertases in health and disease. N. Engl. J. Med. 365(26), (2011). 21 Abifadel M, Varret M, Rabes JP et al. Mutations in PCSK9 cause autosomal dominant hypercholesterolemia. Nat. Genet. 34(2), (2003). 22 Dowdall M. Highlighting the future potential of PCSK9- targeted therapeutics. Clin. Lipidol. 7(6), (2012). 23 Rhainds D, Arsenault BJ, Tardif JC. PCSK9 inhibition and LDL cholesterol lowering: the biology of an attractive therapeutic target and critical review of the latest clinical trials. Clin. Lipidol. 7(6), (2012). 24 Garber K. Biologics inch toward cholesterol-lowering market. Nat. Biotechnol. 30(4), (2012). 25 Dias CS, Shaywitz AJ, Wasserman SM et al. Effects of AMG 145 on low-density lipoprotein cholesterol levels: results from 2 randomized, double-blind, placebo-controlled, ascending-dose Phase 1 studies in healthy volunteers and hypercholesterolemic subjects on statins. J. Am. Coll. Cardiol. 60(19), (2012). 26 Stein EA, Honarpour N, Wasserman SM, Xu F, Scott R, Raal FJ. Effect of the proprotein convertase subtilisin/kexin 9 monoclonal antibody, AMG 145, in homozygous familial hypercholesterolemia. Circulation 128(19), (2013). 27 Raal F, Scott R, Somaratne R et al. Low-density lipoprotein cholesterol-lowering effects of AMG 145, a monoclonal antibody to proprotein convertase subtilisin/kexin type 9 serine protease in patients with heterozygous familial hypercholesterolemia: the Reduction of LDL-C with PCSK9 Inhibition in Heterozygous Familial Hypercholesterolemia 231

13 Review Sahebkar & Watts Disorder (RUTHERFORD) randomized trial. Circulation 126(20), (2012). 28 Stein EA, Mellis S, Yancopoulos GD et al. Effect of a monoclonal antibody to PCSK9 on LDL cholesterol. N. Engl. J. Med. 366(12), (2012). 29 Stein EA, Gipe D, Bergeron J et al. Effect of a monoclonal antibody to PCSK9, REGN727/SAR236553, to reduce low-density lipoprotein cholesterol in patients with heterozygous familial hypercholesterolaemia on stable statin dose with or without ezetimibe therapy: a Phase 2 randomised controlled trial. Lancet 380(9836), (2012). 30 Sullivan D, Olsson AG, Scott R et al. Effect of a monoclonal antibody to PCSK9 on low-density lipoprotein cholesterol levels in statin-intolerant patients: the GAUSS randomized trial. JAMA 308(23), (2012). 31 Giugliano RP, Desai NR, Kohli P et al. Efficacy, safety, and tolerability of a monoclonal antibody to proprotein convertase subtilisin/kexin type 9 in combination with a statin in patients with hypercholesterolaemia (LAPLACE- TIMI 57): a randomised, placebo-controlled, dose-ranging, Phase 2 study. Lancet 380(9858), (2012). 32 Koren MJ, Scott R, Kim JB et al. Efficacy, safety, and tolerability of a monoclonal antibody to proprotein convertase subtilisin/kexin type 9 as monotherapy in patients with hypercholesterolaemia (MENDEL): a randomised, double-blind, placebo-controlled, Phase 2 study. Lancet 380(9858), (2012). 33 Koren MJ, Giugliano RP, Raal FJ et al. Efficacy and safety of longer-term administration of evolocumab (AMG 145) in patients with hypercholesterolemia: 52-week results from the open-label study of long-term evaluation against LDL-C (OSLER) randomized trial. Circulation 129(2), (2013). A Phase III trial with evolocumab. 34 Roth EM, Mckenney JM, Hanotin C, Asset G, Stein EA. Atorvastatin with or without an antibody to PCSK9 in primary hypercholesterolemia. N. Engl. J. Med. 367(20), (2012). 35 AHA. Effects of 12 Weeks of Treatment with RN316 (PF ), a Humanized IgG2Δa Monoclonal Antibody Binding Proprotein Convertase Subtilisin Kexin Type 9, in Hypercholesterolemic Subjects on High and Maximal Dose Statins. ucm_ pdf 36 Gumbiner B, Udata C, Joh T et al. The effects of multiple dose administration of RN316 (PF ), a humanized IgG2a monoclonal antibody binding proprotein convertase subtilisin kexin type 9, in hypercholesterolemic subjects. Circulation 126, A13524 (2012). 37 Fitzgerald K, Frank-Kamenetsky M, Shulga-Morskaya S et al. Effect of an RNA interference drug on the synthesis of proprotein convertase subtilisin/kexin type 9 (PCSK9) and the concentration of serum LDL cholesterol in healthy volunteers: a randomised, single-blind, placebo-controlled, Phase 1 trial. Lancet 383(9911), (2014). 38 Crooke RM, Baker BF, Wedel M. Cardiovascular Therapeutic Applications in Antisense Drug Technology; Principles, Strategies and Applications (2nd Edition). CRC Press, FL, USA, (2007). 39 Kohli P, Cannon CP. A new approach to managing the statin-intolerant patient? Eur. Heart J. 33(9), (2012). 40 Akdim F, Visser ME, Tribble DL et al. Effect of mipomersen, an apolipoprotein B synthesis inhibitor, on low-density lipoprotein cholesterol in patients with familial hypercholesterolemia. Am. J. Cardiol. 105(10), (2010). 41 Raal FJ, Santos RD, Blom DJ et al. Mipomersen, an apolipoprotein B synthesis inhibitor, for lowering of LDL cholesterol concentrations in patients with homozygous familial hypercholesterolaemia: a randomised, double-blind, placebo-controlled trial. Lancet 375(9719), (2010). 42 Kastelein JJ, Wedel MK, Baker BF et al. Potent reduction of apolipoprotein B and low-density lipoprotein cholesterol by short-term administration of an antisense inhibitor of apolipoprotein B. Circulation 114(16), (2006). 43 Akdim F, Stroes ES, Sijbrands EJ et al. Efficacy and safety of mipomersen, an antisense inhibitor of apolipoprotein B, in hypercholesterolemic subjects receiving stable statin therapy. J. Am. Coll. Cardiol. 55(15), (2010). 44 Akdim F, Tribble DL, Flaim JD et al. Efficacy of apolipoprotein B synthesis inhibition in subjects with mild-to-moderate hyperlipidaemia. Eur. Heart J. 32(21), (2011). 45 Visser ME, Wagener G, Baker BF et al. Mipomersen, an apolipoprotein B synthesis inhibitor, lowers low-density lipoprotein cholesterol in high-risk statin-intolerant patients: a randomized, double-blind, placebo-controlled trial. Eur. Heart J. 33(9), (2012). 46 Thomas GS, Cromwell WC, Ali S, Chin W, Flaim JD, Davidson M. Mipomersen, an apolipoprotein B synthesis inhibitor, reduces atherogenic lipoproteins in patients with severe hypercholesterolemia at high cardiovascular risk: a randomized, double-blind, placebo-controlled trial. J. Am. Coll. Cardiol. 62(23), (2013). A Phase III trial with evolocumab. 47 Hussain MM, Shi J, Dreizen P. Microsomal triglyceride transfer protein and its role in ApoB-lipoprotein assembly. J. Lipid Res. 44(1), (2003). 48 Sankatsing RR, Fouchier SW, De Haan S et al. Hepatic and cardiovascular consequences of familial hypobetalipoproteinemia. Arterioscler. Thromb. Vasc. Biol. 25(9), (2005). 49 Joy TR. Novel therapeutic agents for lowering low density lipoprotein cholesterol. Pharmacol. Ther. 135(1), (2012). 50 Cuchel M, Bloedon LT, Szapary PO et al. Inhibition of microsomal triglyceride transfer protein in familial hypercholesterolemia. N. Engl. J. Med. 356(2), (2007). 51 Cuchel M, Meagher EA, Du Toit Theron H et al. Efficacy and safety of a microsomal triglyceride transfer 232 Clin. Lipidol. (2014) 9(2)