PCSK9 inhibition-mediated reduction in Lp(a) with evolocumab: an analysis of 10 clinical trials and the LDL receptor's role
2016; Elsevier BV; Volume: 57; Issue: 6 Linguagem: Inglês
10.1194/jlr.p065334
ISSN1539-7262
AutoresFrederick J. Raal, Robert P. Giugliano, Marc S. Sabatine, Michael J. Koren, Dirk Blom, Nabil G. Seidah, Narimon Honarpour, Armando Lira, Allen Xue, P. Chiruvolu, Simon Jackson, Mei Di, Matthew Peach, Ransi Somaratne, Scott M. Wasserman, Robert C. Scott, Evan A. Stein,
Tópico(s)Lipoproteins and Cardiovascular Health
ResumoLipoprotein (a) [Lp(a)] is independently associated with CVD risk. Evolocumab, a monoclonal antibody (mAb) to proprotein convertase subtilisin/kexin type 9 (PCSK9), decreases Lp(a). The potential mechanisms were assessed. A pooled analysis of Lp(a) and LDL cholesterol (LDL-C) in 3,278 patients from 10 clinical trials (eight phase 2/3; two extensions) was conducted. Within each parent study, biweekly and monthly doses of evolocumab statistically significantly reduced Lp(a) at week 12 versus control (P < 0.001 within each study); pooled median (quartile 1, quartile 3) percent reductions were 24.7% (40.0, 3.6) and 21.7% (39.9, 4.2), respectively. Reductions were maintained through week 52 of the open-label extension, and correlated with LDL-C reductions [with and without correction for Lp(a)-cholesterol] at both time points (P < 0.0001). The effect of LDL and LDL receptor (LDLR) availability on Lp(a) cell-association was measured in HepG2 cells: cell-associated LDL fluorescence was reversed by unlabeled LDL and Lp(a). Lp(a) cell-association was reduced by coincubation with LDL and PCSK9 and reversed by adding PCSK9 mAb. These studies support that reductions in Lp(a) with PCSK9 inhibition are partly due to increased LDLR-mediated uptake. In most situations, Lp(a) appears to compete poorly with LDL for LDLR binding and internalization, but when LDLR expression is increased with evolocumab, particularly in the setting of low circulating LDL, Lp(a) is reduced. Lipoprotein (a) [Lp(a)] is independently associated with CVD risk. Evolocumab, a monoclonal antibody (mAb) to proprotein convertase subtilisin/kexin type 9 (PCSK9), decreases Lp(a). The potential mechanisms were assessed. A pooled analysis of Lp(a) and LDL cholesterol (LDL-C) in 3,278 patients from 10 clinical trials (eight phase 2/3; two extensions) was conducted. Within each parent study, biweekly and monthly doses of evolocumab statistically significantly reduced Lp(a) at week 12 versus control (P < 0.001 within each study); pooled median (quartile 1, quartile 3) percent reductions were 24.7% (40.0, 3.6) and 21.7% (39.9, 4.2), respectively. Reductions were maintained through week 52 of the open-label extension, and correlated with LDL-C reductions [with and without correction for Lp(a)-cholesterol] at both time points (P < 0.0001). The effect of LDL and LDL receptor (LDLR) availability on Lp(a) cell-association was measured in HepG2 cells: cell-associated LDL fluorescence was reversed by unlabeled LDL and Lp(a). Lp(a) cell-association was reduced by coincubation with LDL and PCSK9 and reversed by adding PCSK9 mAb. These studies support that reductions in Lp(a) with PCSK9 inhibition are partly due to increased LDLR-mediated uptake. In most situations, Lp(a) appears to compete poorly with LDL for LDLR binding and internalization, but when LDLR expression is increased with evolocumab, particularly in the setting of low circulating LDL, Lp(a) is reduced. Lipoprotein (a) [Lp(a)] is an LDL-like particle consisting of hepatic synthesized apo(a), a plasminogen-like glycoprotein that is disulfide-linked to the apoB moiety of circulating LDL, most likely at the hepatocellular surface (1Utermann G. The mysteries of lipoprotein(a).Science. 1989; 246: 904-910Crossref PubMed Scopus (1098) Google Scholar). Lp(a) levels are highly variable, primarily genetically determined, and independently associated with CVD. Both epidemiological and genetic studies show that increased levels of Lp(a) are consistently and positively associated with CVD risk (2Erqou, S., S. Kaptoge, P. L. Perry, E. Di Angelantonio, A. Thompson, I. R. White, S. M. Marcovina, R. Collins, S. G. Thompson, and J. Danesh, ; Emerging Risk Factors Collaboration. 2009. Lipoprotein(a) concentration and the risk of coronary heart disease, stroke, and nonvascular mortality. JAMA. 302: 412–423.Google Scholar, 3B. G. Nordestgaard, M. J. Chapman, K. Ray, J. Borén, F. Andreotti, G. F. Watts, H. Ginsberg, P. Amarenco, A. Catapano, O. S. Descamps, ; European Atherosclerosis Society Consensus Panel. 2010. Lipoprotein(a) as a cardiovascular risk factor: current status. Eur. Heart J. 31: 2844–2853.Google Scholar, 4Clarke, R., J. F. Peden, J. C. Hopewell, T. Kyriakou, A. Goel, S. C. Heath, S. Parish, S. Barlera, M. G. Franzosi, S. Rust, ; PROCARDIS Consortium. 2009. Genetic variants associated with Lp(a) lipoprotein level and coronary disease. N. Engl. J. Med. 361: 2518–2528.Google Scholar). Statins, while effective at lowering LDL cholesterol (LDL-C), do not reduce Lp(a) levels and may even increase them slightly (5Hernández C. Francisco G. Ciudin A. Chacón P. Montoro B. Llaverias G. Blanco-Vaca F. Simó R. Effect of atorvastatin on lipoprotein (a) and interleukin-10: a randomized placebo-controlled trial.Diabetes Metab. 2011; 37: 124-130Crossref PubMed Scopus (28) Google Scholar, 6Gonbert S. Malinsky S. Sposito A.C. Laouenan H. Doucet C. Chapman M.J. Thillet J. Atorvastatin lowers lipoprotein(a) but not apolipoprotein(a) fragment levels in hypercholesterolemic subjects at high cardiovascular risk.Atherosclerosis. 2002; 164: 305-311Abstract Full Text Full Text PDF PubMed Scopus (66) Google Scholar, 7Crouse 3rd, J.R. Frohlich J. Ose L. Mercuri M. Tobert J.A. Effects of high doses of simvastatin and atorvastatin on high-density lipoprotein cholesterol and apolipoprotein A-I.Am. J. Cardiol. 1999; 83: 1476-1477, A7Abstract Full Text Full Text PDF PubMed Scopus (131) Google Scholar, 8Hunninghake D.B. Stein E.A. Mellies M.J. Effects of one year of treatment with pravastatin, an HMG-CoA reductase inhibitor, on lipoprotein a.J. Clin. Pharmacol. 1993; 33: 574-580Crossref PubMed Scopus (44) Google Scholar, 9Fieseler H.G. Armstrong V.W. Wieland E. Thiery J. Schütz E. Walli A.K. Seidel D. Serum Lp(a) concentrations are unaffected by treatment with the HMG-CoA reductase inhibitor Pravastatin: results of a 2-year investigation.Clin. Chim. Acta. 1991; 204: 291-300Crossref PubMed Scopus (39) Google Scholar, 10Irudayam J.B. Sivaraj R. Nirmala P. Effect of statins on lipoprotein (a) in dyslipidemic patients.Int. J. Basic Clin. Pharmacol. 2014; 3: 1024-1029Crossref Google Scholar, 11Romagnuolo R. Scipione C.A. Boffa M.B. Marcovina S.M. Seidah N.G. Koschinsky M.L. Lipoprotein(a) catabolism is regulated by proprotein convertase subtilisin/kexin type 9 through the low density lipoprotein receptor.J. Biol. Chem. 2015; 290: 11649-11662Abstract Full Text Full Text PDF PubMed Scopus (150) Google Scholar, 12Kostner G.M. Gavish D. Leopold B. Bolzano K. Weintraub M.S. Breslow J.L. HMG CoA reductase inhibitors lower LDL cholesterol without reducing Lp(a) levels.Circulation. 1989; 80: 1313-1319Crossref PubMed Scopus (285) Google Scholar, 13Yeang C. Hung M-Y. Byun Y-S. Clopton P. Yang X. Witztum J.L. Tsimikas S. Effect of therapeutic interventions on oxidized phospholipids on apolipoprotein B100 and lipoprotein(a).J. Clin. Lipidol. In press. 2016; Abstract Full Text Full Text PDF Scopus (70) Google Scholar). In addition, the reduction of LDL-C with statins has been shown to markedly decrease or attenuate the CVD relationship with Lp(a) (14Berg K. Dahlén G. Christophersen B. Cook T. Kjekshus J. Pedersen T. Lp(a) lipoprotein level predicts survival and major coronary events in the Scandinavian Simvastatin Survival Study.Clin. Genet. 1997; 52: 254-261Crossref PubMed Scopus (130) Google Scholar). Other lipid-lowering drug therapies that do not primarily lower LDL-C by upregulating LDL receptor (LDLR) activity (e.g., bile acid sequestrants, ezetimibe) have no reported effect on Lp(a), while moderate reductions are seen with drugs that inhibit the production of LDL-C or its precursor [e.g., niacin (13Yeang C. Hung M-Y. Byun Y-S. Clopton P. Yang X. Witztum J.L. Tsimikas S. Effect of therapeutic interventions on oxidized phospholipids on apolipoprotein B100 and lipoprotein(a).J. Clin. Lipidol. In press. 2016; Abstract Full Text Full Text PDF Scopus (70) Google Scholar), mipomersen, lomitapide]. Yet Lp(a) is robustly and consistently reduced by treatment with proprotein convertase subtilisin/kexin type 9 (PCSK9) antibodies, which reduce LDL-C via upregulation of LDLR activity (15Raal F.J. Giugliano R.P. Sabatine M.S. Koren M.J. Langslet G. Bays H. Blom D. Eriksson M. Dent R. Wasserman S.M. et al.Reduction in lipoprotein(a) with PCSK9 monoclonal antibody evolocumab (AMG 145): a pooled analysis of more than 1,300 patients in 4 phase II trials.J. Am. Coll. Cardiol. 2014; 63: 1278-1288Crossref PubMed Scopus (289) Google Scholar, 16Gaudet D. Kereiakes D.J. McKenney J.M. Roth E.M. Hanotin C. Gipe D. Du Y. Ferrand A.C. Ginsberg H.N. Stein E.A. Effect of alirocumab, a monoclonal proprotein convertase subtilisin/kexin 9 antibody, on lipoprotein(a) concentrations (a pooled analysis of 150 mg every two weeks dosing from phase 2 trials).Am. J. Cardiol. 2014; 114: 711-715Abstract Full Text Full Text PDF PubMed Scopus (162) Google Scholar, 17Dias C.S. Shaywitz A.J. Wasserman S.M. Smith B.P. Gao B. Stolman D.S. Crispino C.P. Smirnakis K.V. Emery M.G. Colbert A. 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. 2012; 60: 1888-1898Crossref PubMed Scopus (229) Google Scholar, 18Sullivan D. Olsson A.G. Scott R. Kim J.B. Xue A. Gebski V. Wasserman S.M. Stein E.A. Effect of a monoclonal antibody to PCSK9 on low-density lipoprotein cholesterol levels in statin-intolerant patients: the GAUSS randomized trial.JAMA. 2012; 308: 2497-2506Crossref PubMed Scopus (403) Google Scholar, 19Koren M.J. Scott R. Kim J.B. Knusel B. Liu T. Lei L. Bolognese M. Wasserman S.M. 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. 2012; 380: 1995-2006Abstract Full Text Full Text PDF PubMed Scopus (313) Google Scholar, 20Stein E.A. Mellis S. Yancopoulos G.D. Stahl N. Logan D. Smith W.B. Lisbon E. Gutierrez M. Webb C. Wu R. et al.Effect of a monoclonal antibody to PCSK9 on LDL cholesterol.N. Engl. J. Med. 2012; 366: 1108-1118Crossref PubMed Scopus (577) Google Scholar, 21Giugliano, R. P., N. R. Desai, P. Kohli, W. J. Rogers, R. Somaratne, F. Huang, T. Liu, S. Mohanavelu, E. B. Hoffman, S. T. McDonald, ; LAPLACE-TIMI 57 Investigators. 2012. 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: 2007–2017.Google Scholar). The mechanism(s) by which PCSK9 inhibition reduces Lp(a) has not yet been determined, but potential explanations include decreased apoB synthesis (11Romagnuolo R. Scipione C.A. Boffa M.B. Marcovina S.M. Seidah N.G. Koschinsky M.L. Lipoprotein(a) catabolism is regulated by proprotein convertase subtilisin/kexin type 9 through the low density lipoprotein receptor.J. Biol. Chem. 2015; 290: 11649-11662Abstract Full Text Full Text PDF PubMed Scopus (150) Google Scholar), decreased Lp(a) synthesis, reduced availability of apoB-containing lipoproteins (LDL) for linkage to Lp(a), enhanced Lp(a) uptake and clearance by the LDLR (11Romagnuolo R. Scipione C.A. Boffa M.B. Marcovina S.M. Seidah N.G. Koschinsky M.L. Lipoprotein(a) catabolism is regulated by proprotein convertase subtilisin/kexin type 9 through the low density lipoprotein receptor.J. Biol. Chem. 2015; 290: 11649-11662Abstract Full Text Full Text PDF PubMed Scopus (150) Google Scholar, 22Sun H. Samarghandi A. Zhang N. Yao Z. Xiong M. Teng B.B. Proprotein convertase subtilisin/kexin type 9 interacts with apolipoprotein B and prevents its intracellular degradation, irrespective of the low-density lipoprotein receptor.Arterioscler. Thromb. Vasc. Biol. 2012; 32: 1585-1595Crossref PubMed Scopus (126) Google Scholar), or other hepatic receptors in the setting of low LDL-C levels (22Sun H. Samarghandi A. Zhang N. Yao Z. Xiong M. Teng B.B. Proprotein convertase subtilisin/kexin type 9 interacts with apolipoprotein B and prevents its intracellular degradation, irrespective of the low-density lipoprotein receptor.Arterioscler. Thromb. Vasc. Biol. 2012; 32: 1585-1595Crossref PubMed Scopus (126) Google Scholar). The Program to Reduce LDL-C and Cardiovascular Outcomes Following Inhibition of PCSK9 in Different Populations (PROFICIO) is a pooled analysis of 3,278 patients on various background lipid-lowering therapies from eight parent phase 2 and phase 3 studies, and two extension studies of evolocumab, a fully human monoclonal antibody (mAb) to PCSK9 (18Sullivan D. Olsson A.G. Scott R. Kim J.B. Xue A. Gebski V. Wasserman S.M. Stein E.A. Effect of a monoclonal antibody to PCSK9 on low-density lipoprotein cholesterol levels in statin-intolerant patients: the GAUSS randomized trial.JAMA. 2012; 308: 2497-2506Crossref PubMed Scopus (403) Google Scholar, 19Koren M.J. Scott R. Kim J.B. Knusel B. Liu T. Lei L. Bolognese M. Wasserman S.M. 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. 2012; 380: 1995-2006Abstract Full Text Full Text PDF PubMed Scopus (313) Google Scholar, 21Giugliano, R. P., N. R. Desai, P. Kohli, W. J. Rogers, R. Somaratne, F. Huang, T. Liu, S. Mohanavelu, E. B. Hoffman, S. T. McDonald, ; LAPLACE-TIMI 57 Investigators. 2012. 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: 2007–2017.Google Scholar, 23Koren, M. J., P. Lundqvist, M. Bolognese, J. M. Neutel, M. L. Monsalvo, J. Yang, J. B. Kim, R. Scott, S. M. Wasserman, and H. Bays, ; MENDEL-2 Investigators. 2014. Anti-PCSK9 monotherapy for hypercholesterolemia: the MENDEL-2 randomized, controlled phase III clinical trial of evolocumab. J. Am. Coll. Cardiol. 63: 2531–2540.Google Scholar, 24J. G. Robinson, B. S. Nedergaard, W. J. Rogers, J. Fialkow, J. M. Neutel, D. Ramstad, R. Somaratne, J. C. Legg, P. Nelson, R. Scott, ; LAPLACE-2 Investigators. 2014. Effect of evolocumab or ezetimibe added to moderate- or high-intensity statin therapy on LDL-C lowering in patients with hypercholesterolemia: the LAPLACE-2 randomized clinical trial. JAMA. 311: 1870–1882.Google Scholar, 25Raal F. Scott R. Somaratne R. Bridges I. Li G. Wasserman S.M. Stein E.A. 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 Disorder (RUTHERFORD) randomized trial.Circulation. 2012; 126: 2408-2417Crossref PubMed Scopus (461) Google Scholar, 26Raal, F. J., E. A. Stein, R. Dufour, T. Turner, F. Civeira, L. Burgess, G. Langslet, R. Scott, A. G. Olsson, D. Sullivan, ; RUTHERFORD-2 Investigators. 2015. PCSK9 inhibition with evolocumab (AMG 145) in heterozygous familial hypercholesterolaemia (RUTHERFORD-2): a randomised, double-blind, placebo-controlled trial. Lancet. 385: 331–340.Google Scholar, 27Stroes, E., D. Colquhoun, D. Sullivan, F. Civeira, R. S. Rosenson, G. F. Watts, E. Bruckert, L. Cho, R. Dent, B. Knusel, ; GAUSS-2 Investigators. 2014. Anti-PCSK9 antibody effectively lowers cholesterol in patients with statin intolerance: the GAUSS-2 randomized, placebo-controlled phase 3 clinical trial of evolocumab. J. Am. Coll. Cardiol. 63: 2541–2548.Google Scholar, 28Koren, M. J., R. P. Giugliano, F. J. Raal, D. Sullivan, M. Bolognese, G. Langslet, F. Civeira, R. Somaratne, P. Nelson, T. Liu, ; OSLER Investigators. 2014. 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: 234–243.Google Scholar). In this analysis, we (1Utermann G. The mysteries of lipoprotein(a).Science. 1989; 246: 904-910Crossref PubMed Scopus (1098) Google Scholar) evaluated the relationship between posttreatment LDL-C levels with and without correction for Lp(a)-cholesterol [Lp(a)-C] and Lp(a) reductions, and (2Erqou, S., S. Kaptoge, P. L. Perry, E. Di Angelantonio, A. Thompson, I. R. White, S. M. Marcovina, R. Collins, S. G. Thompson, and J. Danesh, ; Emerging Risk Factors Collaboration. 2009. Lipoprotein(a) concentration and the risk of coronary heart disease, stroke, and nonvascular mortality. JAMA. 302: 412–423.Google Scholar) determined whether increased LDLR activity could be responsible for clearance of Lp(a) by examining whether this potential effect could be modulated in vitro. Data from eight randomized placebo-controlled blinded phase 2 and 3 clinical trials [MENDEL-1 (19Koren M.J. Scott R. Kim J.B. Knusel B. Liu T. Lei L. Bolognese M. Wasserman S.M. 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. 2012; 380: 1995-2006Abstract Full Text Full Text PDF PubMed Scopus (313) Google Scholar), MENDEL-2 (23Koren, M. J., P. Lundqvist, M. Bolognese, J. M. Neutel, M. L. Monsalvo, J. Yang, J. B. Kim, R. Scott, S. M. Wasserman, and H. Bays, ; MENDEL-2 Investigators. 2014. Anti-PCSK9 monotherapy for hypercholesterolemia: the MENDEL-2 randomized, controlled phase III clinical trial of evolocumab. J. Am. Coll. Cardiol. 63: 2531–2540.Google Scholar), LAPLACE-TIMI 57 (21Giugliano, R. P., N. R. Desai, P. Kohli, W. J. Rogers, R. Somaratne, F. Huang, T. Liu, S. Mohanavelu, E. B. Hoffman, S. T. McDonald, ; LAPLACE-TIMI 57 Investigators. 2012. 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: 2007–2017.Google Scholar), LAPLACE-2 (24J. G. Robinson, B. S. Nedergaard, W. J. Rogers, J. Fialkow, J. M. Neutel, D. Ramstad, R. Somaratne, J. C. Legg, P. Nelson, R. Scott, ; LAPLACE-2 Investigators. 2014. Effect of evolocumab or ezetimibe added to moderate- or high-intensity statin therapy on LDL-C lowering in patients with hypercholesterolemia: the LAPLACE-2 randomized clinical trial. JAMA. 311: 1870–1882.Google Scholar), RUTHERFORD-1 (25Raal F. Scott R. Somaratne R. Bridges I. Li G. Wasserman S.M. Stein E.A. 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 Disorder (RUTHERFORD) randomized trial.Circulation. 2012; 126: 2408-2417Crossref PubMed Scopus (461) Google Scholar), RUTHERFORD-2 (26Raal, F. J., E. A. Stein, R. Dufour, T. Turner, F. Civeira, L. Burgess, G. Langslet, R. Scott, A. G. Olsson, D. Sullivan, ; RUTHERFORD-2 Investigators. 2015. PCSK9 inhibition with evolocumab (AMG 145) in heterozygous familial hypercholesterolaemia (RUTHERFORD-2): a randomised, double-blind, placebo-controlled trial. Lancet. 385: 331–340.Google Scholar), GAUSS-1 (18Sullivan D. Olsson A.G. Scott R. Kim J.B. Xue A. Gebski V. Wasserman S.M. Stein E.A. Effect of a monoclonal antibody to PCSK9 on low-density lipoprotein cholesterol levels in statin-intolerant patients: the GAUSS randomized trial.JAMA. 2012; 308: 2497-2506Crossref PubMed Scopus (403) Google Scholar), GAUSS-2 (27Stroes, E., D. Colquhoun, D. Sullivan, F. Civeira, R. S. Rosenson, G. F. Watts, E. Bruckert, L. Cho, R. Dent, B. Knusel, ; GAUSS-2 Investigators. 2014. Anti-PCSK9 antibody effectively lowers cholesterol in patients with statin intolerance: the GAUSS-2 randomized, placebo-controlled phase 3 clinical trial of evolocumab. J. Am. Coll. Cardiol. 63: 2541–2548.Google Scholar)] and two open-label extension trials [OSLER-1 (28Koren, M. J., R. P. Giugliano, F. J. Raal, D. Sullivan, M. Bolognese, G. Langslet, F. Civeira, R. Somaratne, P. Nelson, T. Liu, ; OSLER Investigators. 2014. 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: 234–243.Google Scholar), OSLER-2] were used in this analysis. Patients who completed one of the phase 2 or 3 clinical trials and agreed to participate in the open-label extension studies were rerandomized to standard of care (SOC) or SOC plus evolocumab (supplementary Table 1, supplementary Fig. 1), and informed consent was obtained from all patients prior to their participation in the studies. All studies were approved by independent ethics committees and institutional review boards, and all were conducted in accordance with applicable country regulations and the Declaration of Helsinki. Endpoints for this analysis included the percent change from baseline to parent study week 12 and extension study week 52 in Lp(a) and LDL-C and the potential relationship between these changes. Lp(a) was measured by the same standardized isoform-independent method throughout the studies. In the phase 2 studies, LDL-C was determined by both Friedewald formula and ultracentrifugation. In the phase 3 studies LDL-C was calculated using the Friedewald formula, but was measured by ultracentrifugation using the same blood sample if LDL-C by Friedewald was 400 mg/dl; LDL-C was measured by ultracentrifugation using the same blood sample. Patients in the control group included those randomized to receive placebo injections every 2 weeks (Q2W) or monthly (QM), placebo plus ezetimibe, ezetimibe alone, or SOC. Because LDL-C, as measured by ultracentrifugation and calculated by Friedewald formula, includes cholesterol in the form of Lp(a)-C, we corrected for Lp(a)-C by multiplying Lp(a) mass by a factor of 0.3, as approximately 30% of Lp(a) mass is cholesterol (29Seman L.J. Breckenridge W.C. Isolation and partial characterization of apolipoprotein (a) from human lipoprotein (a).Biochem. Cell Biol. 1986; 64: 999-1009Crossref PubMed Scopus (67) Google Scholar). Analyses to compare the efficacy of evolocumab versus control for Lp(a) and LDL-C reductions were performed using the ANCOVA model in each dosing regimen (Q2W or QM). Multiplicity was controlled in the phase 3 studies, but not the phase 2 studies. Correlations between Lp(a) and LDL-C, corrected and uncorrected for Lp(a)-C for all patients, were assessed using Spearman's correlation coefficient for patients with Lp(a) ≥5 nmol/l (the lower limit of detection) at parent study week 12. All analyses were done with SAS/STAT version 9.2 software (SAS Institute, Cary, NC). Fully human anti-PCSK9 mAbs (mAb1, mAb2, mAb3, and mAb4) were all of the IgG2 subclass, and were generated as previously described (30Chan J.C. Piper D.E. Cao Q. Liu D. King C. Wang W. Tang J. Liu Q. Higbee J. Xia Z. et al.A proprotein convertase subtilisin/kexin type 9 neutralizing antibody reduces serum cholesterol in mice and nonhuman primates.Proc. Natl. Acad. Sci. USA. 2009; 106: 9820-9825Crossref PubMed Scopus (330) Google Scholar, 31Henne K.R. Ason B. Howard M. Wang W. Sun J. Higbee J. Tang J. Matsuda K.C. Xu R. Zhou L. et al.Anti-PCSK9 antibody pharmacokinetics and low-density lipoprotein-cholesterol pharmacodynamics in nonhuman primates are antigen affinity-dependent and exhibit limited sensitivity to neonatal Fc receptor-binding enhancement.J. Pharmacol. Exp. Ther. 2015; 353: 119-131Crossref PubMed Scopus (23) Google Scholar). The IgG2 control mAb utilized for in vivo studies was a human anti-keyhole limpet hemocyanin antibody (Amgen Inc., Thousand Oaks, CA). The anti-PCSK9 mAbs differed in binding affinity at neutral and acidic pH, leading to differences in half-life and duration of LDL-C lowering (mAb1 < mAb3 < mAb4 < mAb2) (31Henne K.R. Ason B. Howard M. Wang W. Sun J. Higbee J. Tang J. Matsuda K.C. Xu R. Zhou L. et al.Anti-PCSK9 antibody pharmacokinetics and low-density lipoprotein-cholesterol pharmacodynamics in nonhuman primates are antigen affinity-dependent and exhibit limited sensitivity to neonatal Fc receptor-binding enhancement.J. Pharmacol. Exp. Ther. 2015; 353: 119-131Crossref PubMed Scopus (23) Google Scholar). Fully human anti-LDLR antibodies were generated by immunizing XenoMouse with human LDLR extracellular domain and human EGFa:EGFb-Fc and selecting high-affinity antibodies binding at the EGFa:EGFb domain region (data not shown). All procedures involving laboratory animals were approved by the institutional animal care and use committee at an Association for Assessment and Accreditation of Laboratory Animal Care International-accredited facility, and adhered to the National Research Council's Guide for the Care and Use of Laboratory Animals. Data were generated in three separate single-dose studies in cynomolgus monkeys (Macaca fascicularis) at Valley Biosystems (West Sacramento, CA). Details of these studies are provided in the supplement. The specificity of anti-LDLR antibodies, mAbA and mAbB (Amgen Inc.), for the LDLR was determined using ELISAs. Details of this analysis are provided in the supplement. Anti-LDLR antibodies and PCSK9 were also tested for the ability to modulate expression of other cell receptors. Lysates of HepG2 cells grown in a lipoprotein-deficient serum (LPDS) and treated with PCSK9 with or without mAb1, or with mAbB were analyzed using Western blots probed with antibodies to the VLDL receptor (VLDLR) (Thermo Scientific, Rockford, IL), scavenger receptor class B member 1 (SR-B1) (Thermo Scientific), LDLR-related protein 1 (LRP1) (Thermo Scientific), and cluster of differentiation 36 (CD36) (R&D Systems, Minneapolis, MN). Because PCSK9 inhibition has been shown to increase expression of the LDLR and increase hepatic uptake of LDL (32Dietschy J.M. Turley S.D. Cholesterol metabolism in the brain.Curr. Opin. Lipidol. 2001; 12: 105-112Crossref PubMed Scopus (728) Google Scholar), studies to ascertain a potential connection between the LDLR and cellular Lp(a) association were performed. The majority of LDL is taken up by the LDLR, although ∼10–20% can associate with cells via other receptors or potential non-receptor-mediated pathways (32Dietschy J.M. Turley S.D. Cholesterol metabolism in the brain.Curr. Opin. Lipidol. 2001; 12: 105-112Crossref PubMed Scopus (728) Google Scholar); therefore, fluorescently labeled LDL uptake by HepG2 cells was used as a surrogate assay of LDLR expression and activity. Fluorescently labeled LDL (LDL BODIPY; Life Technologies, Carlsbad, CA) was mixed with increasing concentrations of unlabeled LDL (Sigma-Aldrich, St. Louis, MO), Lp(a) (Lee Biosolutions), HDL (Sigma-Aldrich), or PBS (control), and was then incubated with HepG2 cells seeded in 96-well culture plates (Costar; Corning Life Sciences, Tewksbury, MA) for 3 h at 37°C. After five washes with PBS, cell-associated relative fluorescence units were measured using a plate reader (Safire; Tecan Systems, Inc., San Jose, CA). Cell association with Lp(a) was determined by Western blot using anti-human Lp(a) mAb (EPR6474; Abcam PLC, Cambridge, MA) of HepG2 whole cell lysate. Cells were incubated with purified Lp(a) (Lee Biosolutions) for 4 h at 37°C. In the competition studies, 10 μg/ml of Lp(a) was mixed with 1–40 μg/ml of LDL or HDL (Sigma-Aldrich) and incubated with HepG2 cells for 4 h in 6-well plates at 37°C. Cells were washed five times with PBS and lysed. Protein (40 μg/lane) was resolved on 3–8% tris-acetate SDS-PAGE gels (Life Technologies) for Lp(a) and 4–12% bis-tris SDS-PAGE gels (Life Technologies) for LDLR. Proteins were transferred to nitrocellulose membranes and probed with anti-Lp(a) antibody (1:10,000 dilution), anti-LDLR antibody (1:5,000 dilution; Abcam), or β-actin control (anti-actin antibody, 1:2,000 dilution; Cell Signaling Technology, Inc., Danvers, MA). The role of LDLR in Lp(a) cell association was also assessed by manipulating the level of LDLR on HepG2 cells. LDLR expression was increased by culturing HepG2 cells overnight in medium containing 10% LPDS or maintained at normal levels by culturing in medium containing 10% FBS. LDLR levels were reduced by incubating the cells with 25 or 50 μg/ml of recombinant human PCSK9 (Amgen Inc.) in the presence or absence of 200 μg/ml mAb1. Cell LDLR levels were also downregulated using the anti-LDLR antibodies, mAbA and mAbB (Amgen Inc.), at 50 μg/ml. In each experiment, cells were incubated with 10 μg/ml Lp(a) for 4 h at 37°C. Protein lysates were analyzed using Western blots and probed with antibodies to Lp(a), LDLR, and β-actin as described above. A total of 3,278 patients received at least one dose of study drug and were included in the analyses (supplementary Table 2). The mean (SD) age was 57.7 (11.2) years; most patients were white (91.6%), half were women (50.1%), and most were treated with statins (69.6%). Baseline median [first and third quartiles (Q1, Q3)] Lp(a) was 37.0 (11.0, 148.0) nmol/l; mean (SD) uncorrected LDL-C was 134.3 (47.9) mg/dl; when corrected for Lp(a)-C, LDL-C was 122.9 (48.9) mg/dl. In the pooled dataset, evolocumab (140 mg Q2W and 420 mg QM) resulted in mean (95% confidence interval) percent reductions in uncorre
Referência(s)