Figure 2 - available via license: Creative Commons Attribution 4.0 International
Content may be subject to copyright.
Schematic drawing of plasminogen and apo(a) and their evolutionary relationship. (A) Plasminogen contains seven domains comprising the N-terminal peptide domain (NTP), five kringle domains (K1-K5) and the C-terminal serine protease domain (SP). In apo(a), the protease domain has been changed by mutation in critical residues, destroying its plasmin activity and the kringle K5 of plasminogen was retained as a single copy (KV), while K1-K3 were lost. K4 of plasminogen has differentiated into 10 subtypes of KIV in apo(a) (KIV-1 to KIV-10). Kringle KIV-2 exists in multiple copies and can range from 2 to > 40 copies. (B) Lp(a) consists of an LDL particle to which apo(a) is covalently linked by a disulfide bond. The LDL particle is composed of a central lipid core and a single molecule of apoB. The disulfide bond is formed between apoB and KIV-9 of apo(a).
Source publication
Increased plasma concentrations of lipoprotein(a) (Lp(a)) are associated with an increased risk for cardiovascular disease. Lp(a) is composed of apolipoprotein(a) (apo(a)) covalently bound to apolipoprotein B of low-density lipoprotein (LDL). Many of apo(a)’s potential pathological properties, such as inhibition of plasmin generation, have been att...
Contexts in source publication
Context 1
... interacts with lysine residues in for example fibrin and cell membrane proteins (13). Apo(a) is the result of duplication of the PLA gene encoding plasminogen. The C-terminal kringles 4 and 5 (K4 and K5) and the protease domain of plasminogen have been transferred to apo(a), while the N-terminal kringle 1-3 (K1-K3) of plasminogen have been lost (Fig. 2). In apo(a), the protease domain has lost its catalytic activity by mutation and kringle 4 (K4) has differentiated into 10 subtypes named KIV-1 to KIV-10 (35). Kringle 5 (K5) of plasminogen has been retained as a single copy, named KV in apo(a). Kringle KIV-2 exists in multiple copies ranging from 2 to > 40 in different individuals ...
Context 2
... lost (Fig. 2). In apo(a), the protease domain has lost its catalytic activity by mutation and kringle 4 (K4) has differentiated into 10 subtypes named KIV-1 to KIV-10 (35). Kringle 5 (K5) of plasminogen has been retained as a single copy, named KV in apo(a). Kringle KIV-2 exists in multiple copies ranging from 2 to > 40 in different individuals (Fig. 2). The size of the apo(a) isoform is inversely correlated to the plasma concentration of Lp(a) and therefore also the risk of cardiovascular disease ...
Context 3
... effects of Lp(a) (49). For this reason, we set up an SPR assay to monitor the interaction between fl_apo(a) and fibrin in real time. Fibrin was immobilized on the biosensor surface and the response monitored when fl_apo(a) in solution was injected with and without compound. The apo(a):fibrin interaction was inhibited by TX with a Ki of 30 µM ( Fig. 5 and S2A). We then tested the apo(a) kringle selective compounds, AZ-05 KIV-10 and AZ-06 KIV-7 up to a concentration of 200 µM (Fig. 5). AZ-06 KIV-7 reached a maximum of 30 % inhibition at 25 µM but lost effect at higher concentrations and at 200 µM no inhibition was observed. AZ-05 KIV-10 , on the other hand, showed an increase in inhibition ...
Context 4
... LDL (37-39). To mimic this crucial step in Lp(a) formation we developed an Lp(a) assembly assay using SPR that allowed us to monitor the binding of apo(a) to apoB in a native immobilized LDL particle in real time. Biotinylated LDL particles were attached to the chip surface, fl_apo(a) was injected, and the binding of fl_apo(a) to LDL recorded ( Fig. S2B). Addition of TX to fl_apo(a) before injection inhibited the formation of the non-covalent apo(a):LDL complex with a Ki of 30 µM (Fig. 5). No binding was observed between fl_apo(a) and a reference surface with LDL particles where all surface exposed lysines had been modified by acetylation. These experiments support a specific ...
Context 5
... particles are formed by apo(a) binding to LDL (Fig. 2B) and our SPR results demonstrated that TX and AZ-05 KIV-10 prevented apo(a) binding to LDL (Fig. 5B). Xu (53) showed by scanning force microscopy that apo(a) binds to the LDL sphere at two distant sites and that, occasionally, apo(a) could bridge two LDL particles, bringing them together. We therefore hypothesized that addition of ...
Context 6
... management facility. Upon assay start 98 μL of apo(a) mix were added to all wells to give a final highest compound concentration of 200 μM and an apo(a) concentration as defined under the specified assay. The apo(a) concentration was selected based on the approximate Kd values of apo(a) for fibrin and LDL (ca 50 nM and ca 200 nM respectively, Fig. S2). In both SPR assays a report point, reflecting the binding of apo(a) in response units (RU), was recorded upon sample injection. The response of the control channel was subtracted from the response of the sample ...
Similar publications
![Lipoprotein(a) [Lp(a)] structure. Created with BioRender.com. Reprinted...](publication/354713370/figure/fig1/AS:11431281210505582@1702048236287/Lipoproteina-Lpa-structure-Created-with-BioRendercom-Reprinted-from-Ref-2_Q320.jpg)
Lipoprotein(a) (Lp(a)) is a low density lipoprotein particle that is associated with poor cardiovascular prognosis due to pro-atherogenic, pro-thrombotic, pro-inflammatory and pro-oxidative properties. Traditional lipid-lowering therapy does not provide a sufficient Lp(a) reduction. For PCSK9 inhibitors a small reduction of Lp(a) levels could be sh...
Citations
... Apo(a) exhibits significant sequence homology with plasminogen however, unlike plasminogen it is not an active protease 2 . Apo(a) contains a high degree of variants in its polypeptide chain length due to the variable number of kringle domains 3 . An estimated 20-25% of the world's population is believed to have elevated Lp(a) levels 4 There is evidence that inflammatory conditions such as hypothyroidism, growth hormone therapy, kidney disease 8 and pregnancy increase levels of Lp(a) 5,6,7,8 . ...
Lipoprotein(a) (Lp(a)) is a well-recognized causal risk factor for atherosclerotic cardiovascular disease (ASCVD) and calcific aortic valve stenosis. There are ongoing challenges with screening and management in primary and secondary prevention; however, future recommendations for clinical practice await the outcomes of clinical trials that are in progress.
... As a proof of principle, the in vitro assembly of Lp(a) is inhibited by ε-aminocaproic acid and tranexamic acid, which bind to the lysine-binding domain of apo(a) K IV 7, despite their low binding affinities for K IV 7 as measured by the equilibrium dissociation constant (K d ) (230 µM and 63 µM, respectively) 24 . Similarly, low-affinity but selective K IV 7 and K IV 10 small-molecule binders have been described that reduce the formation of Lp(a) in vitro with micromolar potency 25 . ...
Lipoprotein(a) (Lp(a)), an independent, causal cardiovascular risk factor, is a lipoprotein particle that is formed by the interaction of a low-density lipoprotein (LDL) particle and apolipoprotein(a) (apo(a))1,2. Apo(a) first binds to lysine residues of apolipoprotein B-100 (apoB-100) on LDL through the Kringle IV (KIV) 7 and 8 domains, before a disulfide bond forms between apo(a) and apoB-100 to create Lp(a) (refs. 3–7). Here we show that the first step of Lp(a) formation can be inhibited through small-molecule interactions with apo(a) KIV7–8. We identify compounds that bind to apo(a) KIV7–8, and, through chemical optimization and further application of multivalency, we create compounds with subnanomolar potency that inhibit the formation of Lp(a). Oral doses of prototype compounds and a potent, multivalent disruptor, LY3473329 (muvalaplin), reduced the levels of Lp(a) in transgenic mice and in cynomolgus monkeys. Although multivalent molecules bind to the Kringle domains of rat plasminogen and reduce plasmin activity, species-selective differences in plasminogen sequences suggest that inhibitor molecules will reduce the levels of Lp(a), but not those of plasminogen, in humans. These data support the clinical development of LY3473329—which is already in phase 2 studies—as a potent and specific orally administered agent for reducing the levels of Lp(a).
... Similar functional analogies in the binding of Lp(a) to cellular receptors for plasminogen or to proteins with affinity for plasminogen have been demonstrated recently [89,90]. Interestingly, small-molecule inhibitors have been developed, which target the lysine-binding site of KIV 10 and are able to attenuate the pathophysiological effects of Lp(a) [91]. ...
Lipoprotein(a) is an apolipoprotein B100-containing low-density lipoprotein-like particle that is rich in cholesterol, and is associated with a second major protein, apolipoprotein(a). Apolipoprotein(a) possesses structural similarity to plasminogen but lacks fibrinolytic activity. As a consequence of its composite structure, lipoprotein(a) may: (1) elicit a prothrombotic/antifibrinolytic action favouring clot stability; and (2) enhance atherosclerosis progression via its propensity for retention in the arterial intima, with deposition of its cholesterol load at sites of plaque formation. Equally, lipoprotein(a) may induce inflammation and calcification in the aortic leaflet valve interstitium, leading to calcific aortic valve stenosis. Experimental, epidemiological and genetic evidence support the contention that elevated concentrations of lipoprotein(a) are causally related to atherothrombotic risk and equally to calcific aortic valve stenosis. The plasma concentration of lipoprotein(a) is principally determined by genetic factors, is not influenced by dietary habits, remains essentially constant over the lifetime of a given individual and is the most powerful variable for prediction of lipoprotein(a)-associated cardiovascular risk. However, major interindividual variations (up to 1000-fold) are characteristic of lipoprotein(a) concentrations. In this context, lipoprotein(a) assays, although currently insufficiently standardized, are of considerable interest, not only in stratifying cardiovascular risk, but equally in the clinical follow-up of patients treated with novel lipid-lowering therapies targeted at lipoprotein(a) (e.g. antiapolipoprotein(a) antisense oligonucleotides and small interfering ribonucleic acids) that markedly reduce circulating lipoprotein(a) concentrations. We recommend that lipoprotein(a) be measured once in subjects at high cardiovascular risk with premature coronary heart disease, in familial hypercholesterolaemia, in those with a family history of coronary heart disease and in those with recurrent coronary heart disease despite lipid-lowering treatment. Because of its clinical relevance, the cost of lipoprotein(a) testing should be covered by social security and health authorities.
... 112 Structural work on the binding properties of Kringle domains has led to the development of AZ-005 which inhibits the function of K-IV10 domain and this small molecule inhibitor may proceed to human trials. 113 ...
Lipoprotein(a) forms a subfraction of the lipid profile and is characterized by the addition of apolipprotein(a) (apo(a)) to apoB100 derived particles. Its levels are mostly genetically determined inversely related to the number of protein domain (kringle) repeats in apo(a). In epidemiological studies, it shows consistent association with cardiovascular disease (CVD) and most recently with extent of aortic stenosis. Issues with standardizing the measurement of Lp(a) are being resolved and consensus statements favor its measurement in patients at high risk of, or with family histories of CVD events. Major lipid-lowering therapies such as statin, fibrates, and ezetimibe have little effect on Lp(a) levels. Therapies such as niacin or cholesterol ester transfer protein (CETP) inhibitors lower Lp(a) as well as reducing other lipid-related risk factors but have failed to clearly reduce CVD events. Proprotein convertase subtilisin kexin-9 (PCSK9) inhibitors reduce cholesterol and Lp(a) as well as reducing CVD events. New antisense therapies specifically targeting apo(a) and hence Lp(a) have greater and more specific effects and will help clarify the extent to which intervention in Lp(a) levels will reduce CVD events.
Purpose of review
To review the development of oral agents to lower Lp(a) levels as an approach to reducing cardiovascular risk, with a focus on recent advances in the field.
Recent findings
Extensive evidence implicates Lp(a) in the causal pathway of atherosclerotic cardiovascular disease and calcific aortic stenosis. There are currently no therapies approved for lowering of Lp(a). The majority of recent therapeutic advances have focused on development of injectable agents that target RNA and inhibit synthesis of apo(a). Muvalaplin is the first, orally administered, small molecule inhibitor of Lp(a), which acts by disrupting binding of apo(a) and apoB, in clinical development. Nonhuman primate and early human studies have demonstrated the ability of muvalaplin to produce dose-dependent lowering of Lp(a). Ongoing clinical trials will evaluate the impact of muvalaplin in high cardiovascular risk and will ultimately need to determine whether this strategy lowers the rate of cardiovascular events.
Summary
Muvalaplin is the first oral agent, developed to lower Lp(a) levels. The ability of muvalaplin to reduce cardiovascular risk remains to be investigated, in order to determine whether it will be a useful agent for the prevention of cardiovascular disease.
Elevated plasma levels of lipoprotein(a) (Lp(a)) are a prevalent, independent, and causal risk factor for atherosclerotic cardiovascular disease and calcific aortic valve disease. Lp(a) consists of a lipoprotein particle resembling low density lipoprotein and the covalently-attached glycoprotein apolipoprotein(a) (apo(a)). Novel therapeutics that specifically and potently lower Lp(a) levels are currently in advanced stages of clinical development, including in large, phase 3 cardiovascular outcomes trials. However, fundamental unanswered questions remain concerning some key aspects of Lp(a) biosynthesis and catabolism as well as the true pathogenic mechanisms of the particle. In this review, we describe the salient biochemical features of Lp(a) and apo(a) and how they underlie the disease-causing potential of Lp(a), the factors that determine plasma Lp(a) concentrations, and the mechanism of action of Lp(a)-lowering drugs.
ADAMTS13, a disintegrin and metalloprotease with a thrombospondin type 1 motif, member 13, regulates the length of Von Willebrand factor (VWF) multimers and their platelet-binding activity. ADAMTS13 is constitutively secreted as an active protease and is not inhibited by circulating protease inhibitors. Therefore, the mechanisms that regulate ADAMTS13 protease activity are unknown. We performed an unbiased proteomics screen to identify ligands of ADAMTS13 by optimizing the application of BioID to plasma. Plasma BioID identified 5 plasma proteins significantly labeled by the ADAMTS13-birA* fusion, including VWF and plasminogen. Glu-plasminogen, Lys-plasminogen, mini-plasminogen, and apo(a) bound ADAMTS13 with high affinity, whereas micro-plasminogen did not. None of the plasminogen variants or apo(a) bound to a C-terminal truncation variant of ADAMTS13 (MDTCS). The binding of plasminogen to ADAMTS13 was attenuated by tranexamic acid or ε-aminocaproic acid, and tranexamic acid protected ADAMTS13 from plasmin degradation. These data demonstrate that plasminogen is an important ligand of ADAMTS13 in plasma by binding to the C-terminus of ADAMTS13. Plasmin proteolytically degrades ADAMTS13 in a lysine-dependent manner, which may contribute to its regulation. Adapting BioID to identify protein-interaction networks in plasma provides a powerful new tool to study protease regulation in the cardiovascular system.
Cardiovascular diseases (CVDs) are the leading cause of death globally, attributed to a complex etiology involving metabolic, genetic, and protein-related factors. Lipoprotein(a) (Lp(a)), identified as a genetic risk factor, exhibits elevated levels linked to an increased risk of cardiovascular diseases. The lipoprotein(a) kringle domains have recently been identified as a potential target for the treatment of CVDs, in this study we utilized a fragment-based drug design approach to design a novel, potent, and safe inhibitor for lipoprotein(a) kringle domain. With the use of fragment library (61,600 fragments) screening, combined with analyses such as MM/GBSA, molecular dynamics simulation (MD), and principal component analysis, we successfully identified molecules effective against the kringle domains of Lipoprotein(a). The hybridization process (Breed) of the best fragments generated a novel 249 hybrid molecules, among them 77 exhibiting superior binding affinity (≤ -7 kcal/mol) compared to control AZ-02 (-6.9 kcal/mol), Importantly, the top ten molecules displayed high similarity to the control AZ-02. Among the top ten molecules, BR1 exhibited the best docking energy (-11.85 kcal/mol ), and higher stability within the protein LBS site, demonstrating the capability to counteract the pathophysiological effects of lipoprotein(a) [Lp(a)]. Additionally, principal component analysis (PCA) highlighted a similar trend of motion during the binding of BR1 and the control compound (AZ-02), limiting protein mobility and reducing conformational space. Moreover, ADMET analysis indicated favorable drug-like properties, with BR1 showing minimal violations of Lipinski’s rules. Overall, the identified compounds hold promise as potential therapeutics, addressing a critical need in cardiovascular medicine. Further preclinical and clinical evaluations are needed to validate their efficacy and safety, potentially ushering in a new era of targeted therapies for CVDs.
Elevated plasma concentrations of lipoprotein(a) (Lp(a)) are a causal risk factor for the development of atherothrombotic disorders including coronary heart disease. However, the pathological mechanisms underlying this causal relationship remain incompletely defined. Lp(a) consists of a lipoprotein particle in which apolipoproteinB100 is covalently linked to the unique glycoprotein apolipoprotein(a) (apo(a)). The remarkable homology between apo(a) and the fibrinolytic proenzyme plasminogen strongly suggests an antifibrinolytic role: apo(a) contains a strong lysine binding site and can block the sites on fibrin and cellular receptors required for plasminogen activation, but itself lacks proteolytic activity. While numerous in vitro and animal model studies indicate that apo(a) can inhibit plasminogen activation and fibrinolysis, this activity may not be preserved in Lp(a). Moreover, elevated Lp(a) does not reduce the efficacy of thrombolytic therapy and is not a risk factor for some non-atherosclerotic thrombotic disorders such as venous thromboembolism. Accordingly, different prothrombotic mechanisms for Lp(a) must be contemplated. Evidence exists that Lp(a) binds to and inactivates tissue factor pathway inhibitor and stimulates expression of tissue factor by monocytes. Moreover, some studies have shown that Lp(a) promotes platelet activation and aggregation, at least in response to some agonists. Lp(a) alters the structure of the fibrin network to make it less permeable and more resistant to lysis. Finally, Lp(a) may promote the development of a vulnerable plaque phenotype that is more prone to rupture and hence the precipitation of atherothrombotic events. Further study, especially in animal models of thrombosis, is required to clarify the prothrombotic effects of Lp(a).
