Elguindy, A

Yacoub, MH

Global Cardiology Science and Practice

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PubMed

Spiral - Imperial College Digital Repository

OPEN ACCESS EditorialThe discovery of PCSK9 inhibitors:A tale of creativity and multifacetedtranslational researchAhmed Elguindy1, Magdi H Yacoub2,*Creativity is defined as producing ostensibly out of nothing, something of Beauty, Order orSignificance, Peter Medawar (1915–1987)Creativity is just connecting the dots, Steve Jobs (1955–2011)Atherosclerosis, obesity, andmetabolic syndromeare closely linked and constitute, arguably, themostmenacing three conditions to modern society. Thus urgent, concerted efforts at several levels, utilisingmodern tools, are required to tackle them. Translational research is a rapidly expanding branch ofscience, dedicated to rapid delivery of discoveries from the bench to the bedside. The growingimportance of this specialty in our field is evidenced by the establishment of the International Society ofCardiovascular Translational Research and its dedicated Journal by Nabil Dib and colleagues.1The fundamental discovery of the relationship between, lipids, cholesterol and atherosclerosis wasmade in the 1960s.2–4 This led to the discovery of lipid lowering drugs, including statins, whichconstitute one of the blockbuster drugs of modern times. In spite of that, the toll of the three conditionsmentioned above continued almost unabated, particularly in patients with familial hypercholestero-lemia (FH) in whom LDL apheresis and/or liver transplantation are needed in many instances to achieveadequate reduction of LDL-cholesterol levels.5,6 The landmark discovery of the role of the LDL receptorsin lipid clearance by Goldstein and Brown (Figures 1 and 2)7,8 earned them the Nobel Prize, and openedthe door to further translational research which led to several further discoveries, including that ofProprotein Convertase Subtilisin/Kexin type 9 (PCSK9). In addition to establishing the role of the LDLreceptors in clearing LDL-C from the circulation, and that FH is due to a mutation in the LDL receptorgene, Brown and Goldstein described three new cellular processes, including receptor-mediatedendocytosis (paving the way for further research into the mechanisms of vesicle transport andcellular trafficking; work for which Rothman, Schekman and Sudhof have recently been awardedthe Nobel Prize in Medicine), receptor recycling, and feedback regulation of receptors, which ledto discovery of hydroxymethylglutaryl coenzyme A (HMG-CoA) reductase and statins.Several years later, the discovery of a major regulatory pathway controlling the number andfunction of the LDL receptors was made by a large number of researchers working in differentCite this article as: Elguindy A, Yacoub MH. The discovery of PCSK9 inhibitors: A tale of creativityand multifaceted translational research, Global Cardiology Science and Practice 2013:39http://dx.doi.org/10.5339/gcsp.2013.39http://dx.doi.org/10.5339/gcsp.2013.39Submitted: 1 December 2013Accepted: 2 December 2013ª 2013 Elguindy, Yacoub, licenseeBloomsbury Qatar FoundationJournals. This is an open accessarticle distributed under the termsof the Creative CommonsAttribution license CC BY 3.0, whichpermits unrestricted use,distribution and reproduction in anymedium, provided the original workis properly cited.1Aswan Heart Centre, Aswan, Egypt2Qatar Cardiovascular Research Centre,Doha, Qatar*Email: m.yacoub@imperial.ac.ukLDL ReceptorsLDLProteinCholesterylLinoleateLDL BindingAmino Acids1. HMG CoAReductaseACATLDL Receptors2.3.Internal-izationLysosomalHydrolysisRegulatoryActionsCholesterylOleateCholesterolFigure 1. LDL receptor pathway of mammalian cells (from Goldstein J and Brown S. The LDL Receptor.Arterioscler Thromb Vasc Biol. 2009;29:431–438).fields of translational research. This was started by the massive contribution of Nabil Seidah’sgroup from Montreal (Figure 3). While working on the chemistry and functions of a family ofproteins termed proprotein convertases, they identified PCSK9 encoded by a gene on chromosome 1(Figure 4). Collaborative work with a French group showed that a gain of function mutation in thatgene was responsible for FH in a French family.9 This was followed by similar findings from Oslo.10Figure 2. Joseph L. Goldstein and Michael S. Brown.Figure 3. Nabil Seidah.ERSKI-1GolgiPC5BFurinFurin TGNPC5APC7PACE4ConstitutiveLDLRRegulatedPC1PC2SGFurinHSPGPCSK9PC5BCell surfacePACE4PCSK9PC5AFolding andautocalyticprocessing ofproproteinconvertasesPC7PC7EndosomeFigure 4. Subcellular localization of proprotein convertases. Upon exiting the endoplasmic reticulum (ER), mostof the basic amino acid-specific proprotein convertases traverse the Golgi apparatus towards the trans-Golginetwork (TGN). The activated membrane-bound subtilisin kexin isozyme 1 (SKI1) is mostly concentrated in thecis- andmedial-Golgi, from where it is then sent to lysosomes for degradation, and does not normally reach thecell surface. Proprotein convertase subtilisin kexin 9 (PCSK9) is secreted from the TGN directly into the mediumas an enzymatically inactive non-covalent complex of the protease and its prosegment. Upon binding to the low-density lipoprotein receptor (LDLR) at the cell surface, the PCSK9–LDLR complex is internalized into endosomesand then sent to lysosomes for degradation. (from Seidah N and Prat A. The biology and therapeutic targeting ofproprotein convertases. Nat Rev Drug Discov. 2012 May;11(5):367–83).Page 344 of 347Elguindy and Yacoub. Global Cardiology Science and Practice 2013:39In contrast, findings from the Dallas Heart Study, showed that loss of function mutations in PCSK9gene in a subset of Afro-Americans were associated with very low cholesterol levels and markedlyreduced incidence of cardiovascular disease.11 The mechanisms involved are schematicallyshown in Figure 5.RecyclingendosomeEndosome EndosomeLysosomesABD374S127PCSK9 contact residuesLDL receptor contact residuesPCSK9ProdomainProdomain extension across active siteCatalytic domainCarboxyl-terminal domainLDL receptorLDL ReceptorLDLa repeats (LDL binding)EGF-like repeatsYWTD motifs (β propeller)LDLPCSK9Figure 5. A: Model of PCSK9-mediated sorting of LDL receptors to lysosomes. The EGFa domain of the LDLreceptor is required for proper sorting of the LDL receptor back to the cell surface. The EGFa domain may containa sorting signal that interacts with an endosomal protein (green star), directing the LDL receptor back to the cellsurface on recycling endosomes (green arrows). Binding of PCSK9 might interfere with that signal, preventingthe LDL receptor from returning to the cell surface. Alternatively, PCSK9 could contain a distinct sorting signal(red star) that results in the sorting of the PCSK9-LDL receptor complex (red arrows) to lysosomes. The gain-of-function mutation involving S127 of PCSK9 may enhance the sorting of the PCSK9-LDL receptor complex tolysosomes. B: The structure of the LDL receptor and PCSK9 at endosomal pH. The LDL receptor is folded backupon itself at low pH; however, the face of the EGFa domain that binds PCSK9 is exposed. The LDL receptor-binding site on PCSK9 is at the apex of a roughly triangular structure formed by the tripartite domain structure ofPCSK9. The D374 residue that is altered in gain-of-function PCSK mutants is located within the apical LDLreceptor-binding site, whereas the S127 residue is quite distant from the binding interface. S127 mutations donot affect binding of PCSK9 to the LDL receptor. Gain-of-function mutations affecting residue 127 may reduce LDLreceptors by enhancing the sorting of LDL receptors to lysosomes, rather than by affecting the strength ofPCSK9-LDL receptor interactions. (figure and accompanying legend from Peterson A, Fong L and YoungS. PCSK9 function and Physiology. J Lipid Res. 2008 June; 49(6): 1152–1156).Page 345 of 347Elguindy and Yacoub. Global Cardiology Science and Practice 2013:39THE RUSH TO THE CLINICNot surprisingly, the above mentioned exciting findings was followed by the development of severalstrategies to inhibit the protein or RNA using monoclonal antibodies (Figure 6) or RNA interferencedrugs respectively,12–19 with extremely promising results in Phase 1 and 2 trials. Inhibition ofPCSK9-mediated degradation of LDL receptors by an epidermal growth factor-like repeat A (EGF-A)peptide has also been demonstrated in a mouse model.20THE FUTUREThe search for other means of inhibiting PCSK9 continues. Increased expression of PCSK9 has recentlybeen shown to be a key mechanism by which human resistin – an adipose tissue-derived adipokine –downregulates hepatocyte LDL receptor expression,21 hence increasing the risk of atheroscleroticcardiovascular disease.22,23 This interesting link opens the door for future research into resistininhibition as another strategy for inhibiting PCSK9; possibly with added pleotropic effects.24Such drugs are expected to have a major effect on cardiovascular health, and represent a triumphfor translational research. The story stands as a prime example of creativity as defined by the lateSir Peter Medawar, with a methodology that follows Steve Jobs’ approach.25REFERENCES[1] Hall J, Dib N. The path from pre-clinical research to FDA approval. J Cardiovasc Transl Res. 2008;1(2):93–94.[2] Stamler J. The problem of elevated blood cholesterol. Am J Public Health Nations Health. 1960;50(3 Pt 2):14–19.[3] Kannel WB, Dawber TR, Friedman GD, Glennon WE, McNamara PM. Risk factors in coronary heart disease. Anevaluation of several serum lipids as predictors of coronary heart disease; the framingham study. Ann Intern Med.1964;61:888–899.[4] Kannel WB, Castelli WP, McNamara PM. Serum lipid fractions and risk of coronary heart disease. The Framinghamstudy. Minn Med. 1969;52:1225–1230.[5] Hovingh GK, Davidson MH, Kastelein JJP, O’Connor AM. Diagnosis and treatment of familial hypercholesterolaemia.Eur Heart J. 2013;34(13):962–971.150100500150100500150100500120PCSK9% LDL-C80060040020001008060402000 14Anti-KLH antibody, 3 mg/kgmAb1, 3mg/kgAnti-KLH antibody, 3 mg/kgmAb1, 3mg/kgAnti-KLH antibody, 3 mg/kgmAb1, 3mg/kg7Time following injection (days)ACBDTime following injection (days)Time following injection (days)Time following injection (days)Serum Total Cholesterol(% of predose)Serum HDL Cholesterol(% of predose)Serum LDL Cholesterol(% of predose)Serum LDL Cholesterol(% of predose)Unbound PCSK9(ng/ml)**** **** ********* ***210 147 21 0 147 210 147 21Figure 6. Total cholesterol, LDL-C and HDL-C levels after injection of a neutralizing monoclonal antibody againstPCSK9 compared to a control antibody against keyhole limpet hemocyanin (KLH). Results are expressed asmean ^ SEM. *p , 0.05; **p , 0.01; ***p , 0.001 vs. anti-KLH control antibody at the same time point, n ¼ 4per group. (from Chan J. et al. A proprotein convertase subtilisin/kexin type 9 neutralizing antibody reducesserum cholesterol in mice and nonhuman primates. Proc Natl Acad Sci U S A. 2009 Jun 16;106(24):9820–5).Page 346 of 347Elguindy and Yacoub. Global Cardiology Science and Practice 2013:39[6] Ibrahim M, El-Hamamsy I, Barbir M, Yacoub MH. Translational lessons from a case of combined heart and livertransplantation for familial hypercholesterolemia 20 years post-operatively. J Cardiovasc Transl Res.2012;5(3):351–358.[7] Goldstein JL, Brown MS. Familial hypercholesterolemia: identification of a defect in the regulation of 3-hydroxy-3-methylglutaryl coenzyme A reductase activity associated with overproduction of cholesterol. Proc Natl Acad Sci U S A.1973;70:2804–2808.[8] Goldstein JL, Brown MS. Binding and degradation of low density lipoproteins by cultured human fibroblasts.Comparison of cells from a normal subject and from a patient with homozygous familial hypercholesterolemia.J Biol Chem. 1974;249:5153–5162.[9] Abifadel M, Varret M, Rabés JP, Allard D, Ouguerram K, Devillers M, Cruaud C, Benjannet S, Wickham L, Erlich D, Derré A,Villéger L, Farnier M, Beucler I, Bruckert E, Chambaz J, Chanu B, Lecerf JM, Luc G, Moulin P, Weissenbach J, Prat A,Krempf M, Junien C, Seidah NG, Boileau C. Mutations in PCSK9 cause autosomal dominant hypercholesterolemia.Nat Genet. 2003;34:154–156.[10] Leren TP. Mutations in the PCSK9 gene in Norwegian subjects with autosomal dominant hypercholesterolemia. ClinGenet. 2004;65:419–422.[11] Cohen J, Pertsemlidis A, Kotowski IK, Graham R, Garcia CK, Hobbs HH. Low LDL cholesterol in individuals of Africandescent resulting from frequent nonsense mutations in PCSK9. 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The discovery of PCSK9 inhibitors: A tale of creativity and multifaceted translational research.

Ahmed Elguindy

Magdi H. Yacoub

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The discovery of PCSK9 inhibitors: A tale of creativity and multifaceted translational research

The discovery of PCSK9 inhibitors: A tale of creativity and multifaceted translational research.

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