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Bertrand Cariou - One of the best experts on this subject based on the ideXlab platform.
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Inhibiting PCSK9 — biology beyond LDL control
Nature Reviews Endocrinology, 2019Co-Authors: Robert M. Stoekenbroek, Bertrand Cariou, Gilles Lambert, Gerard K HovinghAbstract:Clinical trials have unequivocally shown that inhibition of proprotein convertase subtilisin/kexin type 9 (PCSK9) efficaciously and safely prevents cardiovascular events by lowering levels of LDL cholesterol. PCSK9 in the circulation is derived mainly from the liver, but the protein is also expressed in the pancreas, the kidney, the intestine and the central nervous system. Although PCSK9 modulates cholesterol metabolism by regulating LDL receptor expression in the liver, in vitro and in vivo studies have suggested that PCSK9 is involved in various other physiological processes. Although therapeutic PCSK9 inhibition could theoretically have undesired effects by interfering with these non-cholesterol-related processes, studies of individuals with genetically determined reduced PCSK9 function and clinical trials of PCSK9 inhibitors have not revealed clinically meaningful adverse consequences of almost completely eradicating PCSK9 from the circulation. The clinical implications of PCSK9 functions beyond lipid metabolism in terms of wanted or unwanted effects of therapeutic PCSK9 inhibition therefore appear to be limited. The objective of this Review is to describe the physiological role of PCSK9 beyond the LDL receptor to provide a rational basis for monitoring the effects of PCSK9 inhibition as these drugs gain traction in the clinic. PCSK9 is expressed in several tissues other than the liver, including the pancreas, the kidney, the intestine and the brain. Although PCSK9 might be involved in various pathophysiological and physiological processes in different organ systems, the clinical implications for therapeutic PCSK9 inhibition seem to be limited. Clinical trials of PCSK9 inhibitors and studies of individuals with genetically determined reduced PCSK9 activity have provided reassurance regarding the safety of therapeutic PCSK9 inhibition. Proprotein convertase subtilisin/kexin type 9 (PCSK9) inhibitors prevent cardiovascular events by lowering levels of LDL cholesterol derived from the liver. However, PCSK9 is expressed in many other tissues, including the pancreas and central nervous system. This Review explores the functions of PCSK9 beyond the control of cholesterol levels.
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Inhibiting PCSK9 — biology beyond LDL control
Nature Reviews Endocrinology, 2018Co-Authors: Robert M. Stoekenbroek, Bertrand Cariou, Gilles Lambert, Gerard K HovinghAbstract:Clinical trials have unequivocally shown that inhibition of proprotein convertase subtilisin/kexin type 9 (PCSK9) efficaciously and safely prevents cardiovascular events by lowering levels of LDL cholesterol. PCSK9 in the circulation is derived mainly from the liver, but the protein is also expressed in the pancreas, the kidney, the intestine and the central nervous system. Although PCSK9 modulates cholesterol metabolism by regulating LDL receptor expression in the liver, in vitro and in vivo studies have suggested that PCSK9 is involved in various other physiological processes. Although therapeutic PCSK9 inhibition could theoretically have undesired effects by interfering with these non-cholesterol-related processes, studies of individuals with genetically determined reduced PCSK9 function and clinical trials of PCSK9 inhibitors have not revealed clinically meaningful adverse consequences of almost completely eradicating PCSK9 from the circulation. The clinical implications of PCSK9 functions beyond lipid metabolism in terms of wanted or unwanted effects of therapeutic PCSK9 inhibition therefore appear to be limited. The objective of this Review is to describe the physiological role of PCSK9 beyond the LDL receptor to provide a rational basis for monitoring the effects of PCSK9 inhibition as these drugs gain traction in the clinic.Proprotein convertase subtilisin/kexin type 9 (PCSK9) inhibitors prevent cardiovascular events by lowering levels of LDL cholesterol derived from the liver. However, PCSK9 is expressed in many other tissues, including the pancreas and central nervous system. This Review explores the functions of PCSK9 beyond the control of cholesterol levels.Key pointsPCSK9 is expressed in several tissues other than the liver, including the pancreas, the kidney, the intestine and the brain.Although PCSK9 might be involved in various pathophysiological and physiological processes in different organ systems, the clinical implications for therapeutic PCSK9 inhibition seem to be limited.Clinical trials of PCSK9 inhibitors and studies of individuals with genetically determined reduced PCSK9 activity have provided reassurance regarding the safety of therapeutic PCSK9 inhibition.
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Inhibiting PCSK9 — biology beyond LDL control
Nature reviews. Endocrinology, 2018Co-Authors: Robert M. Stoekenbroek, Bertrand Cariou, Gilles Lambert, Gerard K HovinghAbstract:Clinical trials have unequivocally shown that inhibition of proprotein convertase subtilisin/kexin type 9 (PCSK9) efficaciously and safely prevents cardiovascular events by lowering levels of LDL cholesterol. PCSK9 in the circulation is derived mainly from the liver, but the protein is also expressed in the pancreas, the kidney, the intestine and the central nervous system. Although PCSK9 modulates cholesterol metabolism by regulating LDL receptor expression in the liver, in vitro and in vivo studies have suggested that PCSK9 is involved in various other physiological processes. Although therapeutic PCSK9 inhibition could theoretically have undesired effects by interfering with these non-cholesterol-related processes, studies of individuals with genetically determined reduced PCSK9 function and clinical trials of PCSK9 inhibitors have not revealed clinically meaningful adverse consequences of almost completely eradicating PCSK9 from the circulation. The clinical implications of PCSK9 functions beyond lipid metabolism in terms of wanted or unwanted effects of therapeutic PCSK9 inhibition therefore appear to be limited. The objective of this Review is to describe the physiological role of PCSK9 beyond the LDL receptor to provide a rational basis for monitoring the effects of PCSK9 inhibition as these drugs gain traction in the clinic.
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homozygous familial hypercholesterolemia patients with identical mutations variably express the ldlr low density lipoprotein receptor highlights implications for the efficacy of evolocumab
Arteriosclerosis Thrombosis and Vascular Biology, 2017Co-Authors: Aurelie Thedrez, Bertrand Cariou, Dirk J Blom, Stephane Raminmangata, Valentin Blanchard, Mikael Croyal, Kevin Chemello, Brice Nativel, Matthieu Pichelin, Steeve BouraneAbstract:Objective— Evolocumab, a PCSK9 (proprotein convertase subtilisin kexin type 9)–neutralizing antibody, lowers low-density lipoprotein cholesterol (LDL-C) in homozygous familial hypercholesterolemic (HoFH) patients with reduced LDLR (low-density lipoprotein receptor) function. However, their individual responses are highly variable, even among carriers of identical LDLR genetic defects. We aimed to elucidate why HoFH patients variably respond to PCSK9 inhibition. Approach and Results— Lymphocytes were isolated from 22 HoFH patients enrolled in the TAUSSIG trial (Trial Assessing Long Term Use of PCSK9 Inhibition in Subjects With Genetic LDL Disorders). Ten patients were true homozygotes (FH1/FH1) and 5 identical compound heterozygotes (FH1/FH2). Lymphocytes were plated with or without mevastatin, recombinant PCSK9 (rPCSK9), or a PCSK9-neutralizing antibody. Cell surface LDLR expression was analyzed by flow cytometry. All HoFH lymphocytes had reduced cell surface LDLR expression compared with non-FH lymphocytes, for each treatment modality. Lymphocytes from FH1/FH2 patients (LDLR defective/negative) displayed the lowest LDLR expression levels followed by lymphocytes from FH1/FH1 patients (defective/defective). Mevastatin increased, whereas rPCSK9 reduced LDLR expression. The PCSK9-neutralizing antibody restored LDLR expression. Lymphocytes displaying higher LDLR expression levels were those isolated from patients presenting with lowest levels of LDL-C and apolipoprotein B, before and after 24 weeks of evolocumab treatment. These negative correlations remained significant in FH1/FH1 patients and appeared more pronounced when patients with apolipoprotein E3/E3 genotypes were analyzed separately. Significant positive correlations were found between the levels of LDLR expression and the percentage reduction in LDL-C on evolocumab treatment. Conclusions— Residual LDLR expression in HoFH is a major determinant of LDL-C levels and seems to drive their individual response to evolocumab.
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Abstract 19748: Mature Enterocytes Lose Their Ability to Secrete PCSK9
Circulation, 2016Co-Authors: François Moreau, Bertrand Cariou, Nabil G Seidah, Annik Prat, Damien Garcon, Claire Blanchard, Audrey Ayer, Xavier Prieur, Michel Neunlist, Cedric Le MayAbstract:Introduction: Proprotein Convertase Subtilisin Kexin of type 9 (PCSK9) promotes LDL receptor lysosomal degradation and is a key regulator of cholesterol metabolism. Beyond the liver, the small intestine is the second organ where PCSK9 is highly expressed, but its ability to secrete PCSK9 remains a matter of debate. Hypothesis: Our study aims to determine whether the small intestine and enterocytes are able to secrete PCSK9. Methods: In vivo , intestinal PCSK9 secretion was assessed by ELISA and western blot in wild-type (WT) and liver-specific PCSK9-deficient mice (L-PCSK9 KO). Ex vivo , local PCSK9 secretion was evaluated from human and murine intestinal explants mounted in Ussing chambers. In vitro , PCSK9 secretion was measured from human intestinal Caco-2 cells along their differentiation. Results: In vivo , PCSK9 concentrations were similar in tail and portal blood and mesenteric lymph from WT mice and undetectable in the portal blood from L-PCSK9 KO mice. Similarly, no local human and murine PCSK9 secretion was measured in Ussing Chambers. Interestingly, PCSK9 secretion was detected at significant level during the first 10 days of Caco-2 cells differentiation then dropped by more than 30 times to become null after 16 days. Despite no change in PCSK9 mRNA levels, intracellular PCSK9 protein content was reduced by 4 times between day 8 and 16 and was associated with a PCSK9 half-life reduction. While the cleavage and PCSK9 trafficking from ER to Golgi were not altered with differentiation, mature PCSK9 accumulated in the Golgi. 2D electrophoresis revealed that the PCSK9 isoelectric point was reduced at late stages of differentiation, suggesting a potential change in the PCSK9 phosphorylation status. Conclusions: We show that mature enterocyte lose their ability to secrete PCSK9. Underlying molecular mechanisms remain under investigation but involve at least a PCSK9 half-life decrease and a post-traductional modification altering the post-golgi trafficking of PCSK9.
Cedric Le May - One of the best experts on this subject based on the ideXlab platform.
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Impact of protease inhibitors on circulating PCSK9 levels in HIV-infected antiretroviral-naive patients from an ongoing prospective cohort
AIDS, 2017Co-Authors: Franck Boccara, Mathilde Ghislain, Laurence Meyer, Cecile Goujard, Cedric Le May, Corinne Vigouroux, Jean P. Bastard, Soraya Fellahi, Jacqueline Capeau, Ariel CohenAbstract:OBJECTIVE: The study aims to assess the association between proprotein convertase subtilisin/kexin type 9 (PCSK9), a major regulator of LDL cholesterol (LDL-C) homeostasis, and HIV-related dyslipidaemia in a cohort of HIV-positive (HIV+) patients under protease inhibitors. METHODS: Plasma PCSK9 levels were measured in 103 HIV+ patients before and after initiating protease inhibitor-based antiretroviral therapy (ART), and in 90 HIV-negative controls matched for age and sex. PCSK9 was measured by ELISA. HIV+ patients who were not virologically suppressed at follow-up or were on lipid-lowering therapy were excluded. RESULTS: In HIV+ (median age 36 years; 77.7% men), PCSK9 levels did not increase after protease inhibitor exposure (median 14 months) (279.5 ng/ml before, 289.6 ng/ml after; P = 0.49) and were significantly elevated versus controls at all timepoints (adjusted P value before and after: \textless0.05). After protease inhibitor initiation, total cholesterol, LDL-C and HDL cholesterol levels increased, but LDL-C remained lower versus controls. At baseline, PCSK9 levels were positively associated with immunodeficiency and the severity of HIV disease [HIV-1 viral load (P = 0.01), CD4 T-cell count \textless200/μl, P = 0.002], stage C HIV disease (P = 0.0002). In protease inhibitor-treated patients, PCSK9 levels were no longer associated with HIV-related factors but with total cholesterol (P = 0.0006), LDL-C (P = 0.01), HDL cholesterol (P = 0.01), triglycerides (P = 0.05) and glycaemia (P = 0.006). CONCLUSION: PSCK9 levels are elevated in HIV+ patients. In ART-naive patients, the relationship between PCSK9 levels and infection severity suggests an effect of HIV disease. After initiating protease inhibitor-containing ART in virologically suppressed patients, PCSK9 levels were associated with dyslipidaemia similar to controls.
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Abstract 19748: Mature Enterocytes Lose Their Ability to Secrete PCSK9
Circulation, 2016Co-Authors: François Moreau, Bertrand Cariou, Nabil G Seidah, Annik Prat, Damien Garcon, Claire Blanchard, Audrey Ayer, Xavier Prieur, Michel Neunlist, Cedric Le MayAbstract:Introduction: Proprotein Convertase Subtilisin Kexin of type 9 (PCSK9) promotes LDL receptor lysosomal degradation and is a key regulator of cholesterol metabolism. Beyond the liver, the small intestine is the second organ where PCSK9 is highly expressed, but its ability to secrete PCSK9 remains a matter of debate. Hypothesis: Our study aims to determine whether the small intestine and enterocytes are able to secrete PCSK9. Methods: In vivo , intestinal PCSK9 secretion was assessed by ELISA and western blot in wild-type (WT) and liver-specific PCSK9-deficient mice (L-PCSK9 KO). Ex vivo , local PCSK9 secretion was evaluated from human and murine intestinal explants mounted in Ussing chambers. In vitro , PCSK9 secretion was measured from human intestinal Caco-2 cells along their differentiation. Results: In vivo , PCSK9 concentrations were similar in tail and portal blood and mesenteric lymph from WT mice and undetectable in the portal blood from L-PCSK9 KO mice. Similarly, no local human and murine PCSK9 secretion was measured in Ussing Chambers. Interestingly, PCSK9 secretion was detected at significant level during the first 10 days of Caco-2 cells differentiation then dropped by more than 30 times to become null after 16 days. Despite no change in PCSK9 mRNA levels, intracellular PCSK9 protein content was reduced by 4 times between day 8 and 16 and was associated with a PCSK9 half-life reduction. While the cleavage and PCSK9 trafficking from ER to Golgi were not altered with differentiation, mature PCSK9 accumulated in the Golgi. 2D electrophoresis revealed that the PCSK9 isoelectric point was reduced at late stages of differentiation, suggesting a potential change in the PCSK9 phosphorylation status. Conclusions: We show that mature enterocyte lose their ability to secrete PCSK9. Underlying molecular mechanisms remain under investigation but involve at least a PCSK9 half-life decrease and a post-traductional modification altering the post-golgi trafficking of PCSK9.
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Role of PCSK9 beyond liver involvement
Current Opinion in Lipidology, 2015Co-Authors: Bertrand Cariou, Karim Si-tayeb, Cedric Le MayAbstract:PURPOSE OF REVIEW: Proprotein convertase subtilisin kexin type 9 (PCSK9) acts as an endogenous natural inhibitor of the LDL receptor pathway, by targeting the receptor to lysosomes for degradation. Beside the liver, PCSK9 is also expressed at significant levels in other tissues, where its function remains unclear. The current review focuses on the extrahepatic actions of PCSK9. RECENT FINDINGS: The generation of liver-specific PCSK9 knockout mice has clearly indicated that PCSK9 affects cholesterol homeostasis via its action on extrahepatic organs. PCSK9 is highly expressed in the intestine, where it controls the production of triglyceride-rich lipoproteins and the transintestinal cholesterol excretion. The role of PCSK9 in the endocrine pancreas and glucose homeostasis remains unclear because conflicting data exist concerning the metabolic phenotype of PCSK9-deficient mice. Sparse data suggest that PCSK9 might also play a role in kidneys, vascular smooth muscle cells, and neurons. SUMMARY: Based on the virtuous combination of genetic and pharmacological approaches, the major function of PCSK9 as a key regulator of hepatic LDL receptor metabolism had quickly emerged. Accumulating evidence indicates that intestinal PCSK9 is also involved in the modulation of lipid homeostasis. Additional studies are warranted to decipher the physiological function of PCSK9 in other extrahepatic tissues and thus to better assess the safety of PCSK9 inhibitors.
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Clinical aspects of PCSK9.
Atherosclerosis, 2011Co-Authors: Bertrand Cariou, Cedric Le May, Philippe CostetAbstract:Proprotein convertase subtilisin kexin type 9 (PCSK9) is a circulating protein that impairs LDL clearance by promoting the LDL receptor (LDLR) degradation. PCSK9 has emerged as a new pharmacological target for hypercholesterolemia, and different PCSK9 inhibitors are now evaluated in clinical trials. Here, we propose an overview of the clinical perspectives of PCSK9. First, we describe the clinical features of patients with PCSK9 mutations, and how these variations impact the cardiovascular risk. Then, we extensively discuss the potential role of circulating PCSK9 as a new biomarker of lipid metabolism. Indeed, many studies conducted in healthy and type 2 diabetic patients have tested the association of circulating PCSK9 with LDL-cholesterol as well as with multiple metabolic parameters. The overall picture of the clinical relevance of circulating PCSK9 is complicated by the effect of nutritional status and hypolipidemic drugs such as statins, fibrates, ezetimibe on plasma PCSK9 concentrations. Finally, we present a brief overview of the available therapeutic strategies to inhibit PCSK9.
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Proprotein convertase subtilisin kexin type 9 null mice are protected from postprandial triglyceridemia
Arteriosclerosis Thrombosis and Vascular Biology, 2009Co-Authors: Cedric Le May, Sanae Kourimate, Cedric Langhi, Maud Chetiveaux, Anne Jarry, Christine Comera, Xavier Collet, Folkert Kuipers, M Krempf, Bertrand CariouAbstract:Objectives— Proprotein convertase subtilisin kexin type 9 (PCSK9) is a natural inhibitor of the low-density lipoprotein receptor, and its deficiency in humans results in low plasma LDL-cholesterol and protection against cardiovascular disease. We explored whether PCSK9 expression impacts postprandial triglyceridemia, another important cardiovascular risk factor. Methods and Results— Real-time PCR and confocal microscopy were used to show that PCSK9 is expressed throughout the entire small intestine and in human enterocytes. On olive oil gavage, PCSK9-deficient mice showed a dramatically decreased postprandial triglyceridemia compared with their wild-type littermates. Lymph analysis revealed that intestinal TG output is not quantitatively modified by PCSK9 deletion. However, PCSK9−/− mice present with a significant reduction of lymphatic apoB secretion compared to PCSK9+/+ mice. Modulating PCSK9 expression in polarized CaCo-2 cells confirmed the relationship between PCSK9 and apoB secretion; PCSK9−/− mice consistently secrete larger TG-rich lipoprotein than wild-type littermates. Finally, kinetic studies showed that PCSK9-deficient mice have an increased ability to clear chylomicrons compared to wild-type littermates. Conclusion— These findings indicate that in addition to its effect on LDL-cholesterol, PCSK9 deficiency might protect against cardiovascular disease by reducing postprandial triglyceridemia. This study shows that PCSK9 is expressed throughout the entire small intestine and in human enterocytes. In mice, PCSK9 deficiency dramatically reduces postprandial triglyceridemia. This phenotype can be explained by qualitative modifications of intestinal chylomicron production and an enhanced hepatic catabolism.
Nabil G Seidah - One of the best experts on this subject based on the ideXlab platform.
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a single domain antibody against the cys and his rich domain of PCSK9 and evolocumab exhibit different inhibition mechanisms in humanized PCSK9 mice
Biological Chemistry, 2018Co-Authors: Rachid Essalmani, Elodie Weider, Jadwiga Marcinkiewicz, Ann Chamberland, Delia Susanresiga, Anna Roubtsova, Nabil G Seidah, Annik PratAbstract:: Proprotein convertase subtilisin/kexin type 9 (PCSK9) is a secreted protein that binds and escorts the low density lipoprotein receptor (LDLR) into the lysosomal degradation pathway. Prescribed monoclonal antibodies (mAbs) against PCSK9 prevent its binding to the LDLR, and result in ~60% lower LDL cholesterol (LDLc) levels. Although efficient, mAbs are expensive. Hence other PCSK9 inhibitors are needed. For screening purpose, we developed C57BL/6J mice expressing the human PCSK9 gene under the control of its own promoter, but lacking endogenous mouse PCSK9. All lines recapitulate the endogenous PCSK9 expression pattern. The Tg2 line that expresses physiological levels of human PCSK9 (hPCSK9) was selected to characterize the inhibitory properties of a previously reported single domain antibody (sdAb), PKF8-mFc, which binds the C-terminal domain of PCSK9. Upon intraveinous injection of 10 mg/kg, PKF8-mFc and the mAb evolocumab neutralized ~50% and 100% of the hPCSK9 impact on total cholesterol (TC) levels, respectively, but PKF8-mFc had a more sustained effect. PKF8-mFc barely affected hPCSK9 levels, whereas evolocumab promoted a 4-fold increase 3 days post-injection, suggesting very different inhibitory mechanisms. The present study also shows that the new transgenic mice are well suited to screen a variety of hPCSK9 inhibitors.
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Abstract 19748: Mature Enterocytes Lose Their Ability to Secrete PCSK9
Circulation, 2016Co-Authors: François Moreau, Bertrand Cariou, Nabil G Seidah, Annik Prat, Damien Garcon, Claire Blanchard, Audrey Ayer, Xavier Prieur, Michel Neunlist, Cedric Le MayAbstract:Introduction: Proprotein Convertase Subtilisin Kexin of type 9 (PCSK9) promotes LDL receptor lysosomal degradation and is a key regulator of cholesterol metabolism. Beyond the liver, the small intestine is the second organ where PCSK9 is highly expressed, but its ability to secrete PCSK9 remains a matter of debate. Hypothesis: Our study aims to determine whether the small intestine and enterocytes are able to secrete PCSK9. Methods: In vivo , intestinal PCSK9 secretion was assessed by ELISA and western blot in wild-type (WT) and liver-specific PCSK9-deficient mice (L-PCSK9 KO). Ex vivo , local PCSK9 secretion was evaluated from human and murine intestinal explants mounted in Ussing chambers. In vitro , PCSK9 secretion was measured from human intestinal Caco-2 cells along their differentiation. Results: In vivo , PCSK9 concentrations were similar in tail and portal blood and mesenteric lymph from WT mice and undetectable in the portal blood from L-PCSK9 KO mice. Similarly, no local human and murine PCSK9 secretion was measured in Ussing Chambers. Interestingly, PCSK9 secretion was detected at significant level during the first 10 days of Caco-2 cells differentiation then dropped by more than 30 times to become null after 16 days. Despite no change in PCSK9 mRNA levels, intracellular PCSK9 protein content was reduced by 4 times between day 8 and 16 and was associated with a PCSK9 half-life reduction. While the cleavage and PCSK9 trafficking from ER to Golgi were not altered with differentiation, mature PCSK9 accumulated in the Golgi. 2D electrophoresis revealed that the PCSK9 isoelectric point was reduced at late stages of differentiation, suggesting a potential change in the PCSK9 phosphorylation status. Conclusions: We show that mature enterocyte lose their ability to secrete PCSK9. Underlying molecular mechanisms remain under investigation but involve at least a PCSK9 half-life decrease and a post-traductional modification altering the post-golgi trafficking of PCSK9.
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annexin a2 reduces PCSK9 protein levels via a translational mechanism and interacts with the m1 and m2 domains of PCSK9
Journal of Biological Chemistry, 2014Co-Authors: Yascara Grisel Luna Saavedra, Nabil G Seidah, Josee Hamelin, Janice Mayne, Claude Lazure, Maryssa Canuel, Sophie Routhier, Roxane Desjardins, Robert DayAbstract:Annexin A2 (AnxA2) was reported to be an extracellular endogenous inhibitor of proprotein convertase subtilisin kexin type 9 (PCSK9) activity on cell-surface LDL receptor degradation. In this study, we investigated the effect of silencing the expression of AnxA2 and PCSK9 in HepG2 and Huh7 cells to better define the role of AnxA2 in PCSK9 regulation. AnxA2 knockdown in Huh7 cells significantly increased PCSK9 protein levels as opposed to AnxA2 knockdown in HepG2 cells. However, HepG2 cells overexpressing AnxA2 had lower levels of PCSK9 protein. Overall, our data revealed a plausible new role of AnxA2 in the reduction of PCSK9 protein levels via a translational mechanism. Moreover, the C-terminal Cys/His-rich domain of PCSK9 is crucial in the regulation of PCSK9 activity, and we demonstrated by far-Western blot assay that the M1 and M2 domains are necessary for the specific interaction of PCSK9's C-terminal Cys/His-rich domain and AnxA2. Finally, we produced and purified recombinant PCSK9 from humans and mice, which was characterized and used to perform 1,1'-dioctadecyl-3,3,3',3'-tetramethylindocarbocyanine perchlorate LDL cell-based assays on the stable knockdown HepG2 and Huh7 cells. We also demonstrated for the first time the equipotency of human and mouse PCSK9 R218S on human cells.
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proprotein convertase subtilisin kexin type 9 PCSK9 can mediate degradation of the low density lipoprotein receptor related protein 1 lrp 1
PLOS ONE, 2013Co-Authors: Maryssa Canuel, Annik Prat, Marie-claude Asselin, Xiaowei Sun, Eustache Paramithiotis, Nabil G SeidahAbstract:Elevated LDL-cholesterol (LDLc) levels are a major risk factor for cardiovascular disease and atherosclerosis. LDLc is cleared from circulation by the LDL receptor (LDLR). Proprotein convertase subtilisin/kexin 9 (PCSK9) enhances the degradation of the LDLR in endosomes/lysosomes, resulting in increased circulating LDLc. PCSK9 can also mediate the degradation of LDLR lacking its cytosolic tail, suggesting the presence of as yet undefined lysosomal-targeting factor(s). Herein, we confirm this, and also eliminate a role for the transmembrane-domain of the LDLR in mediating its PCSK9-induced internalization and degradation. Recent findings from our laboratory also suggest a role for PCSK9 in enhancing tumor metastasis. We show herein that while the LDLR is insensitive to PCSK9 in murine B16F1 melanoma cells, PCSK9 is able to induce degradation of the low density lipoprotein receptor-related protein 1 (LRP-1), suggesting distinct targeting mechanisms for these receptors. Furthermore, PCSK9 is still capable of acting upon the LDLR in CHO 13-5-1 cells lacking LRP-1. Conversely, PCSK9 also acts on LRP-1 in the absence of the LDLR in CHO-A7 cells, where re-introduction of the LDLR leads to reduced PCSK9-mediated degradation of LRP-1. Thus, while PCSK9 is capable of inducing degradation of LRP-1, the latter is not an essential factor for LDLR regulation, but the LDLR effectively competes with LRP-1 for PCSK9 activity. Identification of PCSK9 targets should allow a better understanding of the consequences of PCSK9 inhibition for lowering LDLc and tumor metastasis.
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circulating proprotein convertase subtilisin kexin 9 PCSK9 regulates vldlr protein and triglyceride accumulation in visceral adipose tissue
Arteriosclerosis Thrombosis and Vascular Biology, 2011Co-Authors: Anna Roubtsova, Jadwiga Marcinkiewicz, Ann Chamberland, Nabil G Seidah, Mercedes N Munkonda, Zuhier Awan, Claude Lazure, Katherine Cianflone, Annik PratAbstract:Objective— Proprotein convertase subtilisin/kexin 9 (PCSK9) promotes the degradation of the low-density lipoprotein receptor (LDLR), and its gene is the third locus implicated in familial hypercholesterolemia. Herein, we investigated the role of PCSK9 in adipose tissue metabolism. Methods and Results— At 6 months of age, PCSK9 −/− mice accumulated ≈80% more visceral adipose tissue than wild-type mice. This was associated with adipocyte hypertrophy and increased in vivo fatty acid uptake and ex vivo triglyceride synthesis. Moreover, adipocyte hypertrophy was also observed in PCSK9 −/− Ldlr −/− mice, indicating that the LDLR is not implicated. Rather, we show here by immunohistochemistry that PCSK9 −/− males and females exhibit 4- and ≈40-fold higher cell surface levels of very-low-density lipoprotein receptor (VLDLR) in perigonadal depots, respectively. Expression of PCSK9 in the liver of PCSK9 −/− females reestablished both circulating PCSK9 and normal VLDLR levels. In contrast, specific inactivation of PCSK9 in the liver of wild-type females led to ≈50-fold higher levels of perigonadal VLDLR. Conclusion— In vivo, endogenous PCSK9 regulates VLDLR protein levels in adipose tissue. This regulation is achieved by circulating PCSK9 that originates entirely in the liver. PCSK9 is thus pivotal in fat metabolism: it maintains high circulating cholesterol levels via hepatic LDLR degradation, but it also limits visceral adipogenesis likely via adipose VLDLR regulation.
Gilles Lambert - One of the best experts on this subject based on the ideXlab platform.
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Inhibiting PCSK9 — biology beyond LDL control
Nature Reviews Endocrinology, 2019Co-Authors: Robert M. Stoekenbroek, Bertrand Cariou, Gilles Lambert, Gerard K HovinghAbstract:Clinical trials have unequivocally shown that inhibition of proprotein convertase subtilisin/kexin type 9 (PCSK9) efficaciously and safely prevents cardiovascular events by lowering levels of LDL cholesterol. PCSK9 in the circulation is derived mainly from the liver, but the protein is also expressed in the pancreas, the kidney, the intestine and the central nervous system. Although PCSK9 modulates cholesterol metabolism by regulating LDL receptor expression in the liver, in vitro and in vivo studies have suggested that PCSK9 is involved in various other physiological processes. Although therapeutic PCSK9 inhibition could theoretically have undesired effects by interfering with these non-cholesterol-related processes, studies of individuals with genetically determined reduced PCSK9 function and clinical trials of PCSK9 inhibitors have not revealed clinically meaningful adverse consequences of almost completely eradicating PCSK9 from the circulation. The clinical implications of PCSK9 functions beyond lipid metabolism in terms of wanted or unwanted effects of therapeutic PCSK9 inhibition therefore appear to be limited. The objective of this Review is to describe the physiological role of PCSK9 beyond the LDL receptor to provide a rational basis for monitoring the effects of PCSK9 inhibition as these drugs gain traction in the clinic. PCSK9 is expressed in several tissues other than the liver, including the pancreas, the kidney, the intestine and the brain. Although PCSK9 might be involved in various pathophysiological and physiological processes in different organ systems, the clinical implications for therapeutic PCSK9 inhibition seem to be limited. Clinical trials of PCSK9 inhibitors and studies of individuals with genetically determined reduced PCSK9 activity have provided reassurance regarding the safety of therapeutic PCSK9 inhibition. Proprotein convertase subtilisin/kexin type 9 (PCSK9) inhibitors prevent cardiovascular events by lowering levels of LDL cholesterol derived from the liver. However, PCSK9 is expressed in many other tissues, including the pancreas and central nervous system. This Review explores the functions of PCSK9 beyond the control of cholesterol levels.
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Inhibiting PCSK9 — biology beyond LDL control
Nature Reviews Endocrinology, 2018Co-Authors: Robert M. Stoekenbroek, Bertrand Cariou, Gilles Lambert, Gerard K HovinghAbstract:Clinical trials have unequivocally shown that inhibition of proprotein convertase subtilisin/kexin type 9 (PCSK9) efficaciously and safely prevents cardiovascular events by lowering levels of LDL cholesterol. PCSK9 in the circulation is derived mainly from the liver, but the protein is also expressed in the pancreas, the kidney, the intestine and the central nervous system. Although PCSK9 modulates cholesterol metabolism by regulating LDL receptor expression in the liver, in vitro and in vivo studies have suggested that PCSK9 is involved in various other physiological processes. Although therapeutic PCSK9 inhibition could theoretically have undesired effects by interfering with these non-cholesterol-related processes, studies of individuals with genetically determined reduced PCSK9 function and clinical trials of PCSK9 inhibitors have not revealed clinically meaningful adverse consequences of almost completely eradicating PCSK9 from the circulation. The clinical implications of PCSK9 functions beyond lipid metabolism in terms of wanted or unwanted effects of therapeutic PCSK9 inhibition therefore appear to be limited. The objective of this Review is to describe the physiological role of PCSK9 beyond the LDL receptor to provide a rational basis for monitoring the effects of PCSK9 inhibition as these drugs gain traction in the clinic.Proprotein convertase subtilisin/kexin type 9 (PCSK9) inhibitors prevent cardiovascular events by lowering levels of LDL cholesterol derived from the liver. However, PCSK9 is expressed in many other tissues, including the pancreas and central nervous system. This Review explores the functions of PCSK9 beyond the control of cholesterol levels.Key pointsPCSK9 is expressed in several tissues other than the liver, including the pancreas, the kidney, the intestine and the brain.Although PCSK9 might be involved in various pathophysiological and physiological processes in different organ systems, the clinical implications for therapeutic PCSK9 inhibition seem to be limited.Clinical trials of PCSK9 inhibitors and studies of individuals with genetically determined reduced PCSK9 activity have provided reassurance regarding the safety of therapeutic PCSK9 inhibition.
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Inhibiting PCSK9 — biology beyond LDL control
Nature reviews. Endocrinology, 2018Co-Authors: Robert M. Stoekenbroek, Bertrand Cariou, Gilles Lambert, Gerard K HovinghAbstract:Clinical trials have unequivocally shown that inhibition of proprotein convertase subtilisin/kexin type 9 (PCSK9) efficaciously and safely prevents cardiovascular events by lowering levels of LDL cholesterol. PCSK9 in the circulation is derived mainly from the liver, but the protein is also expressed in the pancreas, the kidney, the intestine and the central nervous system. Although PCSK9 modulates cholesterol metabolism by regulating LDL receptor expression in the liver, in vitro and in vivo studies have suggested that PCSK9 is involved in various other physiological processes. Although therapeutic PCSK9 inhibition could theoretically have undesired effects by interfering with these non-cholesterol-related processes, studies of individuals with genetically determined reduced PCSK9 function and clinical trials of PCSK9 inhibitors have not revealed clinically meaningful adverse consequences of almost completely eradicating PCSK9 from the circulation. The clinical implications of PCSK9 functions beyond lipid metabolism in terms of wanted or unwanted effects of therapeutic PCSK9 inhibition therefore appear to be limited. The objective of this Review is to describe the physiological role of PCSK9 beyond the LDL receptor to provide a rational basis for monitoring the effects of PCSK9 inhibition as these drugs gain traction in the clinic.
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The PCSK9 decade.
Journal of lipid research, 2012Co-Authors: Gilles Lambert, Barbara Sjouke, Benjamin Choque, John J.p. Kastelein, Gerard K HovinghAbstract:PCSK9 proprotein convertase subtilisin/kexin type (PCSK9) is a crucial protein in LDL cholesterol (LDL-C) metabolism by virtue of its pivotal role in the degradation of the LDL receptor. In recent years, both in vitro and in vivo studies have greatly supplemented our understanding of the (patho)physiological role of PCSK9 in human biology. In the current review, we summarize studies published or in print before May 2012 concerning the physiological role of PCSK9 in cholesterol metabolism. Moreover, we briefly describe the clinical phenotypes encountered in carriers of mutations in the gene encoding PCSK9. As PCSK9 has emerged as a novel target for LDL-C lowering therapy, methods to inhibit PCSK9 will also be reviewed. Initial data from investigations of PCSK9 inhibition in humans are promising and indicate that PCSK9 inhibition may be a viable new therapeutic option for the treatment of dyslipidemia and associated cardiovascular diseases.
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fenofibrate concomitantly decreases serum proprotein convertase subtilisin kexin type 9 and very low density lipoprotein particle concentrations in statin treated type 2 diabetic patients
Diabetes Obesity and Metabolism, 2010Co-Authors: Dick C Chan, Gilles Lambert, Sandra J Hamilton, Kerryanne Rye, Gerard T Chew, Alicia J Jenkins, Gerald F WattsAbstract:Aim: Diabetic dyslipidaemia, characterized by hypertriglyceridaemia as a result of elevated serum very-low-density lipoprotein (VLDL) concentrations, contributes to the increased risk of cardiovascular disease (CVD) in type 2 diabetes (T2DM). Proprotein convertase subtilisin/kexin type 9 (PCSK9) may play a role in regulating VLDL metabolism. We investigated the effect of fenofibrate on serum PCSK9 and VLDL particle concentrations in T2DM patients already receiving statin therapy. Methods: In a double-blind randomized crossover study, 15 statin-treated T2DM patients (63 ± 8 years, body mass index (BMI) 29 ± 3 kg/m2) were treated with fenofibrate (145 mg/day) or matching placebo for 12 weeks. Serum PCSK9 concentrations were measured by immunoassay. VLDL particle concentration and size were determined by nuclear magnetic resonance spectroscopy. Results: Fenofibrate decreased serum triglycerides (−23%), VLDL-triglycerides (−51%), total cholesterol (−11%), LDL-cholesterol (−16%), apolipoprotein B-100 (−16%), apolipoprotein C-III (−20%) and PCSK9 (−13%) concentrations compared with placebo (p < 0.05). Fenofibrate also decreased serum concentrations of large (−45%), medium (−66%) and small VLDL (−67%) particles (p < 0.05), without altering VLDL particle size. Serum PCSK9 reduction correlated with decreases in total (r = 0.526, p = 0.044) and small (r = 0.629, p = 0.021) VLDL particle concentrations. Conclusions: Fenofibrate concomitantly decreased serum PCSK9 and VLDL particle concentrations in statin-treated T2DM patients. These findings support a mechanistic link between PCSK9 and VLDL metabolism, possibly through an effect of PSK9 on VLDL receptor degradation.
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a single domain antibody against the cys and his rich domain of PCSK9 and evolocumab exhibit different inhibition mechanisms in humanized PCSK9 mice
Biological Chemistry, 2018Co-Authors: Rachid Essalmani, Elodie Weider, Jadwiga Marcinkiewicz, Ann Chamberland, Delia Susanresiga, Anna Roubtsova, Nabil G Seidah, Annik PratAbstract:: Proprotein convertase subtilisin/kexin type 9 (PCSK9) is a secreted protein that binds and escorts the low density lipoprotein receptor (LDLR) into the lysosomal degradation pathway. Prescribed monoclonal antibodies (mAbs) against PCSK9 prevent its binding to the LDLR, and result in ~60% lower LDL cholesterol (LDLc) levels. Although efficient, mAbs are expensive. Hence other PCSK9 inhibitors are needed. For screening purpose, we developed C57BL/6J mice expressing the human PCSK9 gene under the control of its own promoter, but lacking endogenous mouse PCSK9. All lines recapitulate the endogenous PCSK9 expression pattern. The Tg2 line that expresses physiological levels of human PCSK9 (hPCSK9) was selected to characterize the inhibitory properties of a previously reported single domain antibody (sdAb), PKF8-mFc, which binds the C-terminal domain of PCSK9. Upon intraveinous injection of 10 mg/kg, PKF8-mFc and the mAb evolocumab neutralized ~50% and 100% of the hPCSK9 impact on total cholesterol (TC) levels, respectively, but PKF8-mFc had a more sustained effect. PKF8-mFc barely affected hPCSK9 levels, whereas evolocumab promoted a 4-fold increase 3 days post-injection, suggesting very different inhibitory mechanisms. The present study also shows that the new transgenic mice are well suited to screen a variety of hPCSK9 inhibitors.
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Abstract 19748: Mature Enterocytes Lose Their Ability to Secrete PCSK9
Circulation, 2016Co-Authors: François Moreau, Bertrand Cariou, Nabil G Seidah, Annik Prat, Damien Garcon, Claire Blanchard, Audrey Ayer, Xavier Prieur, Michel Neunlist, Cedric Le MayAbstract:Introduction: Proprotein Convertase Subtilisin Kexin of type 9 (PCSK9) promotes LDL receptor lysosomal degradation and is a key regulator of cholesterol metabolism. Beyond the liver, the small intestine is the second organ where PCSK9 is highly expressed, but its ability to secrete PCSK9 remains a matter of debate. Hypothesis: Our study aims to determine whether the small intestine and enterocytes are able to secrete PCSK9. Methods: In vivo , intestinal PCSK9 secretion was assessed by ELISA and western blot in wild-type (WT) and liver-specific PCSK9-deficient mice (L-PCSK9 KO). Ex vivo , local PCSK9 secretion was evaluated from human and murine intestinal explants mounted in Ussing chambers. In vitro , PCSK9 secretion was measured from human intestinal Caco-2 cells along their differentiation. Results: In vivo , PCSK9 concentrations were similar in tail and portal blood and mesenteric lymph from WT mice and undetectable in the portal blood from L-PCSK9 KO mice. Similarly, no local human and murine PCSK9 secretion was measured in Ussing Chambers. Interestingly, PCSK9 secretion was detected at significant level during the first 10 days of Caco-2 cells differentiation then dropped by more than 30 times to become null after 16 days. Despite no change in PCSK9 mRNA levels, intracellular PCSK9 protein content was reduced by 4 times between day 8 and 16 and was associated with a PCSK9 half-life reduction. While the cleavage and PCSK9 trafficking from ER to Golgi were not altered with differentiation, mature PCSK9 accumulated in the Golgi. 2D electrophoresis revealed that the PCSK9 isoelectric point was reduced at late stages of differentiation, suggesting a potential change in the PCSK9 phosphorylation status. Conclusions: We show that mature enterocyte lose their ability to secrete PCSK9. Underlying molecular mechanisms remain under investigation but involve at least a PCSK9 half-life decrease and a post-traductional modification altering the post-golgi trafficking of PCSK9.
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proprotein convertase subtilisin kexin type 9 PCSK9 can mediate degradation of the low density lipoprotein receptor related protein 1 lrp 1
PLOS ONE, 2013Co-Authors: Maryssa Canuel, Annik Prat, Marie-claude Asselin, Xiaowei Sun, Eustache Paramithiotis, Nabil G SeidahAbstract:Elevated LDL-cholesterol (LDLc) levels are a major risk factor for cardiovascular disease and atherosclerosis. LDLc is cleared from circulation by the LDL receptor (LDLR). Proprotein convertase subtilisin/kexin 9 (PCSK9) enhances the degradation of the LDLR in endosomes/lysosomes, resulting in increased circulating LDLc. PCSK9 can also mediate the degradation of LDLR lacking its cytosolic tail, suggesting the presence of as yet undefined lysosomal-targeting factor(s). Herein, we confirm this, and also eliminate a role for the transmembrane-domain of the LDLR in mediating its PCSK9-induced internalization and degradation. Recent findings from our laboratory also suggest a role for PCSK9 in enhancing tumor metastasis. We show herein that while the LDLR is insensitive to PCSK9 in murine B16F1 melanoma cells, PCSK9 is able to induce degradation of the low density lipoprotein receptor-related protein 1 (LRP-1), suggesting distinct targeting mechanisms for these receptors. Furthermore, PCSK9 is still capable of acting upon the LDLR in CHO 13-5-1 cells lacking LRP-1. Conversely, PCSK9 also acts on LRP-1 in the absence of the LDLR in CHO-A7 cells, where re-introduction of the LDLR leads to reduced PCSK9-mediated degradation of LRP-1. Thus, while PCSK9 is capable of inducing degradation of LRP-1, the latter is not an essential factor for LDLR regulation, but the LDLR effectively competes with LRP-1 for PCSK9 activity. Identification of PCSK9 targets should allow a better understanding of the consequences of PCSK9 inhibition for lowering LDLc and tumor metastasis.
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circulating proprotein convertase subtilisin kexin 9 PCSK9 regulates vldlr protein and triglyceride accumulation in visceral adipose tissue
Arteriosclerosis Thrombosis and Vascular Biology, 2011Co-Authors: Anna Roubtsova, Jadwiga Marcinkiewicz, Ann Chamberland, Nabil G Seidah, Mercedes N Munkonda, Zuhier Awan, Claude Lazure, Katherine Cianflone, Annik PratAbstract:Objective— Proprotein convertase subtilisin/kexin 9 (PCSK9) promotes the degradation of the low-density lipoprotein receptor (LDLR), and its gene is the third locus implicated in familial hypercholesterolemia. Herein, we investigated the role of PCSK9 in adipose tissue metabolism. Methods and Results— At 6 months of age, PCSK9 −/− mice accumulated ≈80% more visceral adipose tissue than wild-type mice. This was associated with adipocyte hypertrophy and increased in vivo fatty acid uptake and ex vivo triglyceride synthesis. Moreover, adipocyte hypertrophy was also observed in PCSK9 −/− Ldlr −/− mice, indicating that the LDLR is not implicated. Rather, we show here by immunohistochemistry that PCSK9 −/− males and females exhibit 4- and ≈40-fold higher cell surface levels of very-low-density lipoprotein receptor (VLDLR) in perigonadal depots, respectively. Expression of PCSK9 in the liver of PCSK9 −/− females reestablished both circulating PCSK9 and normal VLDLR levels. In contrast, specific inactivation of PCSK9 in the liver of wild-type females led to ≈50-fold higher levels of perigonadal VLDLR. Conclusion— In vivo, endogenous PCSK9 regulates VLDLR protein levels in adipose tissue. This regulation is achieved by circulating PCSK9 that originates entirely in the liver. PCSK9 is thus pivotal in fat metabolism: it maintains high circulating cholesterol levels via hepatic LDLR degradation, but it also limits visceral adipogenesis likely via adipose VLDLR regulation.
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PCSK9 impedes hepatitis C virus infection in vitro and modulates liver CD81 expression.
Hepatology, 2009Co-Authors: Patrick Labonté, Annik Prat, Syntia Begley, Carl Guévin, Marie-claude Asselin, Nasha Nassoury, Gaétan Mayer, Nabil G SeidahAbstract:Human PCSK9 is known to enhance the degradation of membrane-bound receptors such as the hepatocyte low-density lipoprotein receptor (LDLR), ApoER2, and very low-density lipoprotein receptor. Because the LDLR is suspected to be involved in hepatitis C virus (HCV) entry, we also tested whether PCSK9 can affect the levels of CD81, a major HCV receptor. Interestingly, stable expression of PCSK9 or a more active membrane-bound form of the protein (PCSK9-ACE2) resulted in a marked reduction in CD81 and LDLR expression. Therefore, we analyzed the antiviral effect of PCSK9 in vitro using the HCV genotype 2a (JFH1) virus. The results clearly demonstrated that cells expressing PCSK9 or PCSK9-ACE2, but not the ACE2 control protein, were resistant to HCV infection. Furthermore, addition of purified soluble PCSK9 to cell culture supernatant impeded HCV infection in a dose-dependent manner. As expected, HuH7 cells expressing PCSK9-ACE2 were also resistant to infection by HCV pseudoparticles. In addition, we showed that CD81 cell surface expression is modulated by PCSK9 in an LDLR-independent manner. Finally, in the liver of single PCSK9 and double (PCSK9 + Ldlr) knockout mice, both LDLR and/or CD81 protein expression levels were significantly reduced, but not those of transferrin and scavenger receptor class B type 1. Conclusion: Our results demonstrate an antiviral effect of the circulating liver PCSK9 on HCV in cells and show that PCSK9 down-regulates the level of mouse liver CD81 expression in vivo. Therefore, we propose that the plasma level and/or activity of PCSK9 may modulate HCV infectivity in humans.