Endothelial lipase antibody for the treatment of cardiovascular diseases

Abstract

The present disclosure provides methods of administering antibodies and antigen-binding fragments thereof that specifically bind to human endothelial lipase (EL) to a subject in need thereof, for example, a subject with cardiovascular disease.

Description

Technical Field

[0001] [Cross - Reference to Related Applications] This application claims priority to U.S. Provisional Application No. 63 / 104,410, filed October 22, 2020; U.S. Provisional Application No. 62 / 940,164, filed November 25, 2019; and U.S. Provisional Application No. 62 / 932,257, filed November 7, 2019. The entire disclosure content of these U.S. provisional applications is incorporated herein by reference.

[0002] 1. Field The present disclosure generally relates to methods of using antibodies and antigen - binding fragments thereof that specifically bind to human endothelial lipase (EL) for the treatment of diseases or disorders such as cardiovascular diseases and cardiovascular disorders. Advantageous uses are provided.

Background Art

[0003] 2. Background Endothelial lipase (EL) is a circulating phospholipase that has been identified as a member of the triglyceride lipase family. EL has both phospholipase activity and triglyceride lipase activity and hydrolyzes high - density lipoprotein (HDL) more efficiently than other lipoproteins. EL is thought to play an important role in the regulation of plasma HDL cholesterol (HDL - C) levels. EL causes destabilization of HDL particles and rapid clearance by the kidney by hydrolyzing HDL phospholipids.

[0004] An increase in plasma EL concentration is associated not only with a worsening of the lipoprotein - lipid profile along with increases in plasma triglyceride and apolipoprotein B concentrations, but also with smaller low - density lipoprotein particle size (Non - Patent Document 1). Among individuals with elevated plasma EL concentration, an increase in inflammatory cytokine concentration and an increased prevalence of metabolic syndrome have also been observed (Non - Patent Document 1). Considering these and other factors, EL is thought to play an important role in cardiovascular diseases (Non - Patent Document 1).

[0005] Despite the effectiveness of current therapies for cardiovascular diseases such as high - potency statins, a significant residual risk of major cardiovascular (CV) events remains in patients with acute coronary syndrome (ACS). Most myocardial infarctions (MIs) occur in patients with normal low - density lipoprotein (LDL) levels, and despite treatment with high - dose high - potency statins, PCSK9 inhibitors, and / or ezetimibe after MI, the residual risk of a second CV event remains high. For example, in the IMPROVE - IT study (ezetimibe + statin), there was a 33% risk of CV events over 7 years of follow - up. Furthermore, in ODYSSEY (PCSK9 inhibitor + statin), there was a 9.5% residual risk over 2.8 years.

[0006] Low HDL - C levels have been identified as predictors of atherosclerotic CV events and risk factors for coronary heart disease (CHD). The hypothesis has also been proposed that HDL particle size and number may be useful clinical markers of HDL and related diseases.

[0007] Several attempts have been made to pharmacologically increase HDL levels using various mechanisms of action. In particular, four trials of cholesterol - ester transfer protein (CETP) inhibitors have been completed. CETP inhibitors increase HDL cholesterol, but in three of the four trials, CV outcomes were not improved, and in one trial, while CV events decreased, the effect was modest, with only a 9% relative risk reduction. This approach has been criticized because CETP inhibition causes blockade of LDL receptor - mediated reverse cholesterol transport.

[0008] Humans with partial and complete loss-of-function mutations in the gene encoding EL exhibit an increase in HDL-C, an increase in cholesterol efflux capacity (CEC), and a tendency to reduce CV risk. Therefore, neutralization of EL is a promising therapeutic mechanism. However, currently, there are no approved therapies that target EL or sufficiently reduce CV risk. Accordingly, there is a need for methods of effectively treating diseases and disorders, such as CV diseases and CV disorders, using anti-EL antibodies and antibody fragments thereof.

Prior Art Documents

Non-Patent Documents

[0009]

Non-Patent Document 1

Summary of the Invention

[0010] 3. Summary Provided herein are methods of treating cardiovascular disease in a subject. In certain embodiments, the method comprises administering to the subject from about 100 mg to about 350 mg of an antibody or antigen-binding fragment thereof that specifically binds to human endothelial lipase (EL).

[0011] Provided herein are methods of reducing atherosclerosis in a subject. In certain embodiments, the method comprises administering to the subject from about 100 mg to about 350 mg of an antibody or antigen-binding fragment thereof that specifically binds to human EL.

[0012] Provided herein is a method for treating cardiovascular disease or reducing atherosclerosis in a subject. In certain embodiments, the method comprises administering to the subject an antibody or antigen-binding fragment thereof that specifically binds to EL, wherein administration of the antibody or antigen-binding fragment thereof (a) increases high-density lipoprotein cholesterol (HDL-C) in the subject, (b) increases the number of high-density lipoprotein (HDL) particles in the subject, (c) increases the size of HDL particles in the subject, (d) increases the HDL phospholipids in the subject, (e) increases ApoA1 in the subject, and / or (f) increases the cholesterol efflux capacity (CEC) in the subject. In certain embodiments, administration reduces the risk of cardiovascular death, non-fatal myocardial infarction (MI), non-fatal stroke, and / or coronary artery revascularization in a subject having a history of acute coronary syndrome (ACS). In certain embodiments, administration prevents secondary cardiovascular events in the subject. In certain embodiments, administration reduces the risk of major adverse cardiovascular events (MACE) in the subject. In certain embodiments, administration increases the HDL phospholipids in the subject, (e) increases ApoA1 in the subject, and / or (f) increases the cholesterol efflux capacity (CEC) in the subject. In certain embodiments, administration reduces the risk of cardiovascular death, non-fatal myocardial infarction (MI), non-fatal stroke, and / or coronary artery revascularization in a subject having a history of acute coronary syndrome (ACS). In certain embodiments, administration prevents secondary cardiovascular events in the subject. In certain embodiments, administration reduces the risk of major adverse cardiovascular events (MACE) in the subject.

[0013] Provided herein is a method for reducing the risk of cardiovascular death, non-fatal myocardial infarction (MI), non-fatal stroke, and / or coronary artery revascularization in a subject having a history of acute coronary syndrome (ACS). In certain embodiments, the method comprises administering to the subject an antibody or antigen-binding fragment thereof that specifically binds to human EL.

[0014] Provided herein is a method for preventing secondary cardiovascular events in a subject. In certain embodiments, the method comprises administering to the subject an antibody or antigen-binding fragment thereof that specifically binds to human EL.

[0015] Provided herein is a method for reducing the risk of major adverse cardiovascular events (MACE) in a subject. In certain embodiments, the method comprises administering to the subject an antibody or antigen-binding fragment thereof that specifically binds to human EL.

[0016] In certain embodiments of the present disclosure, administration of an antibody or an antigen-binding fragment thereof increases (a) high-density lipoprotein cholesterol (HDL-C) in a subject, (b) the number of high-density lipoprotein (HDL) particles in a subject, (c) the size of HDL particles in a subject, (d) HDL phospholipids in a subject, (e) ApoA1 in a subject, and / or (f) cholesterol efflux capacity (CEC) in a subject.

[0017] Provided herein are methods of increasing HDL-C, the number of HDL particles, the size of HDL particles, HDL phospholipids, ApoA1, and / or CEC in a subject. In certain embodiments, the method comprises administering to the subject an antibody that specifically binds to EL or an antigen-binding fragment thereof.

[0018] Certain embodiments of the present disclosure include administering an antibody or antigen-binding fragment thereof in an amount of about 100 mg to about 350 mg. Certain embodiments of the present disclosure include administering an antibody or antigen-binding fragment thereof in an amount of about 100 mg, about 110 mg, about 120 mg, about 125 mg, about 130 mg, about 140 mg, about 150 mg, about 160 mg, about 170 mg, about 175 mg, about 180 mg, about 190 mg, about 200 mg, about 210 mg, about 220 mg, about 225 mg, about 230 mg, about 240 mg, about 250 mg, about 260 mg, about 270 mg, about 275 mg, about 280 mg, about 290 mg, about 300 mg, about 310 mg, about 320 mg, about 325 mg, about 330 mg, about 340 mg, or about 350 mg. Certain embodiments of the present disclosure include administering an antibody or antigen-binding fragment thereof in an amount of about 125 mg. Certain embodiments of the present disclosure include administering an antibody or antigen-binding fragment thereof in an amount of 125 mg. Certain embodiments of the present disclosure include administering an antibody or antigen-binding fragment thereof in an amount of about 250 mg. Certain embodiments of the present disclosure include administering an antibody or antigen-binding fragment thereof in an amount of 250 mg. Certain embodiments of the present disclosure include administering an antibody or antigen-binding fragment thereof in an amount of about 200 mg. Certain embodiments of the present disclosure include administering an antibody or antigen-binding fragment thereof in an amount of 200 mg to 250 mg.

[0019] In certain embodiments of the present disclosure, the antibody or antigen-binding fragment thereof is administered once per month. In certain embodiments of the present disclosure, the antibody or antigen-binding fragment thereof is administered once per month for at least 3 months. In certain embodiments of the present disclosure, the antibody or antigen-binding fragment thereof is administered once per month for at least 12 months or at least 24 months.

[0020] In certain embodiments of the present disclosure, the antibody or antigen-binding fragment thereof is administered parenterally. In certain embodiments of the present disclosure, the antibody or antigen-binding fragment thereof is administered subcutaneously.

[0021] In certain embodiments of the present disclosure, an antibody or an antigen-binding fragment thereof is administered via an attached prefilled syringe (APFS) or an autoinjector.

[0022] In certain embodiments of the present disclosure, administration of an antibody or an antigen-binding fragment thereof inhibits EL in a subject for 30 days.

[0023] In certain embodiments of the present disclosure, administration of an antibody or an antigen-binding fragment thereof increases HDL-C in a subject by at least 30%. In certain embodiments of the present disclosure, administration of an antibody or an antigen-binding fragment thereof increases HDL-C in a subject by at least 35%. In certain embodiments of the present disclosure, administration of an antibody or an antigen-binding fragment thereof increases HDL-C in a subject by at least 40%. In certain embodiments of the present disclosure, administration of an antibody or an antigen-binding fragment thereof increases HDL-C in a subject within 30 days from the first administration. In certain embodiments of the present disclosure, administration of an antibody or an antigen-binding fragment thereof increases HDL-C in a subject within 90 days from the first administration.

[0024] In certain embodiments of the present disclosure, administration of an antibody or an antigen-binding fragment thereof increases ApoA1 in a subject by at least 30%. In certain embodiments of the present disclosure, administration of an antibody or an antigen-binding fragment thereof increases ApoA1 in a subject by at least 35%. In certain embodiments of the present disclosure, administration of an antibody or an antigen-binding fragment thereof increases ApoA1 in a subject within 30 days from the first administration. In certain embodiments of the present disclosure, administration of an antibody or an antigen-binding fragment thereof increases ApoA1 in a subject within 90 days from the first administration.

[0025] In certain embodiments of the present disclosure, administration of an antibody or an antigen-binding fragment thereof increases non-ABCA1 cholesterol efflux capacity in a subject by at least 30%. In certain embodiments of the present disclosure, administration of an antibody or an antigen-binding fragment thereof increases non-ABCA1 cholesterol efflux capacity in a subject by at least 35%. In certain embodiments of the present disclosure, administration of an antibody or an antigen-binding fragment thereof increases non-ABCA1 cholesterol efflux capacity in a subject within 30 days from the first administration. In certain embodiments of the present disclosure, administration of an antibody or an antigen-binding fragment thereof increases non-ABCA1 cholesterol efflux capacity in a subject within 90 days from the first administration.

[0026] In certain embodiments of the present disclosure, administration of an antibody or an antigen-binding fragment thereof increases the number of HDL particles in a subject by at least 5% (e.g., as measured using NMR). In certain embodiments of the present disclosure, administration of an antibody or an antigen-binding fragment thereof increases the number of HDL particles in a subject by at least 8% (e.g., as measured using NMR). In certain embodiments of the present disclosure, administration of an antibody or an antigen-binding fragment thereof increases the number of HDL particles in a subject within 30 days from the first administration. In certain embodiments of the present disclosure, administration of an antibody or an antigen-binding fragment thereof increases the number of HDL particles in a subject within 90 days from the first administration.

[0027] In certain embodiments of the present disclosure, administration of an antibody or an antigen-binding fragment thereof increases the HDL particle size in a subject by at least 3%. In certain embodiments of the present disclosure, administration of an antibody or an antigen-binding fragment thereof increases the HDL particle size in a subject by at least 5%. In certain embodiments of the present disclosure, administration of an antibody or an antigen-binding fragment thereof increases the HDL particle size in a subject within 30 days from the first administration. In certain embodiments of the present disclosure, administration of an antibody or an antigen-binding fragment thereof increases the HDL particle size in a subject within 90 days from the first administration.

[0028] In certain embodiments of the present disclosure, administration of an antibody or an antigen-binding fragment thereof increases HDL phospholipids in a subject by at least 50%. In certain embodiments of the present disclosure, administration of an antibody or an antigen-binding fragment thereof increases HDL phospholipids in a subject within 30 days from the first administration. In certain embodiments of the present disclosure, administration of an antibody or an antigen-binding fragment thereof increases HDL phospholipids in a subject within 90 days from the first administration.

[0029] In certain embodiments of the present disclosure, administration of an antibody or an antigen-binding fragment thereof increases plasma phosphatidylinositol (PI) levels in a subject by at least 100% or at least 250%. In certain embodiments of the present disclosure, the increase in plasma PI levels is an increase in the levels of PI(14:2 / 20:0), PI(14:2 / 22:0), PI(14:2 / 22:1), PI(14:2 / 22:2), PI(16:0 / 16:1), PI(16:0 / 18:0), PI(16:0 / 18:2), PI(16:0 / 20:2), PI(16:0 / 20:3), PI(16:0 / 20:4), PI(16:0 / 22:4), PI(16:1 / 18:0), PI(16:1 / 18:1), PI(18:0 / 18:0), PI(18:0 / 18:1), PI(18:0 / 18:2), PI(18:0 / 18:3), PI(18:0 / 20:2), PI(18:0 / 20:3), PI(18:0 / 20:4), PI(18:0 / 22:4), PI(18:0 / 22:5), PI(18:0 / 22:6), PI(18:1 / 16:0), PI(18:1 / 18:1), PI(18:1 / 18:2), PI(18:1 / 20:2), PI(18:1 / 20:3), PI(18:1 / 20:4), and / or PI(18:2 / 18:2). In certain embodiments of the present disclosure, administration of an antibody or an antigen-binding fragment thereof increases plasma phosphatidylinositol (PI) levels in a subject within 90 days from the first administration.

[0030] In certain embodiments of the present disclosure, the subject has a cardiovascular disease. In certain embodiments of the present disclosure, the cardiovascular disease is coronary artery disease, coronary heart disease (CHD), chronic arterial disease, cerebrovascular disease, atherosclerotic cardiovascular disease, or peripheral arterial disease.

[0031] In certain embodiments of the present disclosure, the subject has stable coronary artery disease or stable coronary heart disease. In certain embodiments of the present disclosure, the subject has a history of acute coronary syndrome (ACS).

[0032] In certain embodiments of the present disclosure, the subject is receiving statin therapy. In certain embodiments of the present disclosure, the subject is not receiving statin therapy.

[0033] In certain embodiments of the present disclosure, the subject has a triglyceride level of 500 mg / dL or less before administration. In certain embodiments of the present disclosure, the subject has LDL-C of 100 mg / dL or less before administration.

[0034] In certain embodiments of the present disclosure, the subject is human.

[0035] In certain embodiments of the present disclosure, the antibody or its antigen-binding fragment neutralizes EL activity.

[0036] In certain embodiments of the present disclosure, the antibody or its antigen-binding fragment is associated with a reduced effector function. In certain embodiments of the present disclosure, the antibody or its antigen-binding fragment does not have antibody-dependent cell-mediated cytotoxicity (ADCC) activity. In certain embodiments of the present disclosure, the antibody does not have complement-dependent cytotoxicity (CDC) activity.

[0037] In certain embodiments of the present disclosure, the antibody binds to cynomolgus EL.

[0038] In certain embodiments of the present disclosure, the antibody or antigen-binding fragment thereof competitively inhibits the binding of an antibody comprising a VH comprising the amino acid sequence shown in SEQ ID NO: 7 and a VL comprising the amino acid sequence shown in SEQ ID NO: 8 to EL. In certain embodiments of the present disclosure, the antibody or antigen-binding fragment thereof binds to the same epitope of EL as an antibody comprising a VH comprising the amino acid sequence shown in SEQ ID NO: 7 and a VL comprising the amino acid sequence shown in SEQ ID NO: 8.

[0039] In certain embodiments of the present disclosure, the antibody or antigen-binding fragment thereof comprises the heavy chain variable region (VH) complementarity determining regions (CDRs) 1, VH CDR2, VH CDR3, light chain variable region (VL) CDR1, VL CDR2, and VL CDR3 of the sequence of MEDI5884. In certain embodiments of the present disclosure, the CDRs are CDRs according to the Kabat definition, CDRs according to the Chothia definition, or CDRs according to the AbM definition. In certain embodiments of the present disclosure, the antibody or antigen-binding fragment thereof comprises a VH CDR1 comprising the amino acid sequence of SEQ ID NO: 1, a VH CDR2 comprising the amino acid sequence of SEQ ID NO: 2, a VH CDR3 comprising the amino acid sequence of SEQ ID NO: 3, a VL CDR1 comprising the amino acid sequence of SEQ ID NO: 4, a VL CDR2 comprising the amino acid sequence of SEQ ID NO: 5, and a VL CDR3 comprising the amino acid sequence of SEQ ID NO: 6.

[0040] In certain embodiments of the present disclosure, the antibody or antigen-binding fragment thereof comprises a VH comprising the amino acid sequence shown in SEQ ID NO: 7 and / or a VL comprising the amino acid sequence shown in SEQ ID NO: 8.

[0041] In certain embodiments of the present disclosure, the antibody or antigen-binding fragment comprises an IgG heavy chain constant region. In certain embodiments of the present disclosure, the IgG heavy chain constant region is an IgG4 heavy chain constant region. In certain embodiments of the present disclosure, the IgG4 heavy chain constant region is an IgG4P heavy chain constant region. In certain embodiments of the present disclosure, the antibody or antigen-binding fragment thereof comprises a kappa light chain constant region.

[0042] In certain aspects of the present disclosure, the antibody or antigen-binding fragment comprises a heavy chain constant region and / or a light chain constant region. In certain aspects of the present disclosure, the heavy chain constant region is a human IgG4P heavy chain constant region and / or the light chain constant region is a human IgGκ light chain constant region.

[0043] In certain aspects of the present disclosure, the antibody or antigen-binding fragment thereof is a humanized antibody or antigen-binding fragment thereof.

[0044] In certain aspects of the present disclosure, the antibody or antigen-binding fragment thereof comprises a heavy chain constant region comprising the amino acid sequence shown in SEQ ID NO: 1 2 and / or a light chain constant region comprising the amino acid sequence shown in SEQ ID NO: 13.

[0045] In certain aspects of the present disclosure, the antibody comprises a heavy chain comprising the amino acid sequence shown in SEQ ID NO: 9 and a light chain comprising the amino acid sequence shown in SEQ ID NO: 10.

[0046] In certain aspects of the present disclosure, the antibody or antigen-binding fragment thereof is a full-length antibody.

[0047] In certain aspects of the present disclosure, the antibody or antigen-binding fragment thereof is an antigen-binding fragment. In certain aspects of the present disclosure, the antigen-binding fragment is Fab, Fab’, F(ab’) 2 , single-chain Fv (scFv), disulfide-linked Fv, V-NAR domain, IgNar, intrabody, IgGΔCH2, minibody, F(ab’) 3 , tetrabody, tribody, diabody, single-domain antibody, DVD-Ig, Fcab, mAb 2 , (scFv) 2 , or scFv-Fc.

[0048] Provided herein is a method of treating cardiovascular disease in a subject. In certain embodiments, the method comprises subcutaneously administering 250 mg of an antibody or an antigen-binding fragment thereof to the subject once a month, wherein the antibody or antigen-binding fragment specifically binds to human EL and comprises a VH comprising the amino acid sequence set forth in SEQ ID NO: 7 and a VL comprising the amino acid sequence set forth in SEQ ID NO: 8. In certain embodiments, the method comprises subcutaneously administering about 200 mg of an antibody or an antigen-binding fragment thereof to the subject once a month, wherein the antibody or antigen-binding fragment specifically binds to human EL and comprises a VH comprising the amino acid sequence set forth in SEQ ID NO: 7 and a VL comprising the amino acid sequence set forth in SEQ ID NO: 8. In certain embodiments, the method comprises subcutaneously administering 200 mg to 250 mg of an antibody or an antigen-binding fragment thereof to the subject once a month, wherein the antibody or antigen-binding fragment specifically binds to human EL and comprises a VH comprising the amino acid sequence set forth in SEQ ID NO: 7 and a VL comprising the amino acid sequence set forth in SEQ ID NO: 8. Provided herein is a method of reducing the risk of cardiovascular death, non-fatal myocardial infarction (MI), non-fatal stroke, and / or coronary artery revascularization in a subject with a history of acute coronary syndrome (ACS). In certain embodiments, the method comprises subcutaneously administering 250 mg of an antibody or an antigen-binding fragment thereof to the subject once a month, wherein the antibody or antigen-binding fragment specifically binds to human EL and comprises a VH comprising the amino acid sequence set forth in SEQ ID NO: 7 and a VL comprising the amino acid sequence set forth in SEQ ID NO: 8. In certain embodiments, the method comprises subcutaneously administering about 200 mg of an antibody or an antigen-binding fragment thereof to the subject once a month, wherein the antibody or antigen-binding fragment specifically binds to human EL and comprises a VH comprising the amino acid sequence set forth in SEQ ID NO: 7 and a VL comprising the amino acid sequence set forth in SEQ ID NO: 8.In certain aspects, the method comprises subcutaneously administering to a subject, once per month, 200 mg to 250 mg of an antibody or antigen-binding fragment thereof, wherein the antibody or antigen-binding fragment specifically binds to human EL and comprises a VH comprising the amino acid sequence set forth in SEQ ID NO:7 and a VL comprising the amino acid sequence set forth in SEQ ID NO:8.

[0049] In certain embodiments of the disclosure, the antibody comprises the amino acid sequence of a heavy chain constant region set forth in SEQ ID NO:9 and the amino acid sequence of a light chain constant region set forth in SEQ ID NO:10.

[0050] In certain embodiments of the present disclosure, the method further comprises administering an inhibitor of proprotein convertase subtilisin / kexin type 9 (PCSK9). In certain embodiments, the administration of the antibody or antigen-binding fragment thereof that specifically binds to human EL and the administration of the inhibitor of PCSK9 are simultaneous. In certain embodiments, the administration of the antibody or antigen-binding fragment thereof that specifically binds to human EL and the administration of the inhibitor of PCSK9 are simultaneous. The antibody or antigen-binding fragment thereof that specifically binds to human EL and the inhibitor of PCSK9 are administered in separate pharmaceutical compositions. In certain embodiments, the administration of the antibody or antigen-binding fragment thereof that specifically binds to human EL and the administration of the inhibitor of PCSK9 are sequential.

[0051] In certain embodiments, the inhibitor of PCSK9 is an anti-PCSK9 antibody or an antigen-binding fragment thereof. In certain embodiments, the inhibitor of PCSK9 is HS9, evolocumab, alirocumab, or bococizumab.

[0052] 4. Brief description of the drawings [Brief description of the drawings]

[0053]

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Mode for Carrying Out the Invention

[0054] 5. Detailed Description Provided herein is a method of administering an antibody (e.g., a monoclonal antibody) and antigen-binding fragment thereof that specifically binds to endothelial lipase (EL, e.g., human EL). Administration of the anti-EL antibody and antigen-binding fragment thereof can treat, for example, cardiovascular disease in a subject. The anti-EL antibody or antigen-binding fragment thereof can increase high density lipoprotein cholesterol (HDL-C) in a subject, increase the number of HDL particles, increase the size of HDL particles, increase HDL phospholipids, increase ApoA1, and / or increase cholesterol efflux capacity. In some aspects of the present disclosure, about 100 mg to about 350 mg (e.g., about 250 mg) of the antibody or antigen-binding fragment thereof is administered to a subject, and for example, the administration is performed about once a month (QM).

[0055] 5.1 Terms As used herein, the terms "endothelial lipase" or "EL" refer to mammalian EL polypeptides, including but not limited to native EL polypeptides and isoforms of EL polypeptides. "EL" encompasses not only full-length unprocessed EL polypeptides but also forms of EL polypeptides resulting from intracellular processing. As used herein, the term "human EL" refers to a polypeptide comprising the amino acid sequence of SEQ ID NO: 11. "EL polynucleotide", "EL nucleotide", or "EL nucleic acid" refers to a polynucleotide encoding EL.

[0056] The term "antibody" means an immunoglobulin molecule that recognizes a target, such as a protein, polypeptide, peptide, carbohydrate, polynucleotide, lipid, or combination thereof, via at least one antigen recognition site within the variable region of the immunoglobulin molecule and specifically binds to that target. As used herein, the term "antibody" refers to intact polyclonal antibodies, intact monoclonal antibodies, chimeric antibodies, humanized antibodies, human antibodies, fusion proteins comprising antibodies, and any other modified immunoglobulin molecules, so long as the antibody exhibits the desired biological activity. Antibodies can be any of the five major classes of immunoglobulins, IgA, IgD, IgE, IgG, and IgM, or their subclasses (isotypes) based on the individuality of their heavy chain constant domains, called alpha, delta, epsilon, gamma, and mu, respectively (e.g., IgG1, IgG2, IgG3, IgG4, IgA1, and IgA2). Different classes of immunoglobulins have different well-known subunit structures and three-dimensional conformations. Antibodies can be naked antibodies or conjugated to other molecules such as toxins, radioisotopes, etc.

[0057] The term "antibody fragment" refers to a part of an intact antibody. The "antigen-binding fragment", "antigen-binding domain", or "antigen-binding region" refers to the part of an intact antibody that binds to an antigen. An antigen-binding fragment may include the antigen recognition site of an intact antibody (e.g., complementarity-determining regions (CDRs) sufficient to specifically bind to an antigen). Examples of antigen-binding fragments of an antibody include, but are not limited to, Fab fragment, Fab' fragment, F(ab') 2 fragment, and Fv fragment, linear antibodies, and single-chain antibodies. Antigen-binding fragments of an antibody may be derived from any animal species such as rodents (e.g., mouse, rat, or hamster), and humans, or may be artificially produced.

[0058] The terms "anti-EL antibody", "EL antibody", and "antibody that binds to EL" refer to an antibody that can specifically bind to EL with sufficient affinity such that the antibody is useful as a diagnostic agent and / or therapeutic agent in the targeting of EL. As used herein, the terms "specifically binds", "immunologically specifically binds", "immunologically specifically recognizes", and "specifically recognizes" are similar terms in the context of an antibody or its antigen-binding fragment. These terms indicate that the antibody or its antigen-binding fragment binds to an epitope via its antigen-binding domain and that the binding involves some complementarity between the antigen-binding domain and the epitope. Thus, an antibody that "specifically binds" to human EL (SEQ ID NO: 11) may also bind to EL from other species (e.g., cynomolgus monkey) and / or EL proteins produced from other human alleles, but the degree of binding to an unrelated non-EL protein (e.g., other lipases such as hepatic lipase or lipoprotein lipase) is, for example, less than about 10% of the binding of the antibody to EL as measured by an in vitro neutralization assay.

[0059] A "monoclonal" antibody or antigen-binding fragment thereof refers to a homogeneous population of antibodies or antigen-binding fragments that are involved in the highly specific binding to a single antigenic determinant or epitope. This is typically in contrast to polyclonal antibodies, which contain different antibodies directed against different antigenic determinants. The term "monoclonal" antibody or antigen-binding fragment thereof refers to both intact and full-length monoclonal antibodies, as well as antibody fragments (Fab, Fab’, F(ab’) 2 , Fv, etc.), single-chain (scFv) variants, fusion proteins containing antibody portions, and any other modified immunoglobulin molecules containing antigen recognition sites. Furthermore, a "monoclonal" antibody or antigen-binding fragment thereof refers to such antibodies and antigen-binding fragments produced by several methods including, but not limited to, hybridomas, phage selection, recombinant expression, and transgenic animals.

[0060] As used herein, the terms "variable region" or "variable domain" are used interchangeably and are common in the art. The variable region typically refers to a part of the antibody, generally part of the light or heavy chain, typically the amino-terminal approximately 110-120 or 110-125 amino acids of the mature heavy chain and approximately 90-115 amino acids of the mature light chain, which differ in sequence between antibodies and are used for the binding and specificity of a particular antibody to a particular antigen. The sequence variability is concentrated in regions called complementarity-determining regions (CDRs), but the The more highly conserved regions within the variable domain are referred to as the framework region (FR). Without wishing to be bound by a particular mechanism or theory, it is believed that the CDRs of the light and heavy chains are primarily responsible for the interaction and specificity of the antibody with the antigen. In some aspects of the present disclosure, the variable region is a human variable region. In some aspects of the present disclosure, the variable region comprises rodent or mouse CDRs, and a human framework region (FR). In certain aspects of the present disclosure, the variable region is a variable region of a primate (e.g., non-human primate). In some aspects of the present disclosure, the variable region comprises rodent or mouse CDRs, and a primate (e.g., non-human primate) framework region (FR).

[0061] The terms "VL" and "VL domain" are used interchangeably to refer to the variable light chain region of an antibody.

[0062] The terms "VH" and "VH domain" are used interchangeably to refer to the variable heavy chain region of an antibody.

[0063] The term "Kabat numbering" and similar terms are recognized in the art and refer to systems for numbering amino acid residues in the variable regions of the heavy and light chains of an antibody, or antigen-binding fragments thereof. In some embodiments, CDRs can be identified according to the Kabat numbering system (see, e.g., Kabat EA & Wu TT (1971) Ann NY Acad Sci 190: 382-391, and Kabat EA et al., (1991) Sequences of Proteins of Immunological Interest, Fifth Edition, U.S. Department of Health and Human Services, NIH Publication No. 91-3242). Using the Kabat numbering system, CDRs within an antibody heavy chain molecule are typically present at amino acid positions 31-35 (CDR1), amino acid positions 50-65 (CDR2), and amino acid positions 95-102 (CDR3), which may optionally include one or two additional amino acids (referred to as 35A and 35B in the Kabat numbering scheme) following 35. Using the Kabat numbering system, CDRs within an antibody light chain molecule are typically present at amino acid positions 24-34 (CDR1), amino acid positions 50-56 (CDR2), and amino acid positions 89-97 (CDR3). In some embodiments of the present disclosure, the CDRs of the antibodies described herein were determined according to the Kabat numbering scheme.

[0064] Instead, Chothia refers to the position of structural loops (Chothia and Lesk, J. Mol. Biol. 196:901-917 (1987)). The end of the Chothia CDR-H1 loop when numbered using the Kabat numbering rules varies between H32 and H34 depending on the length of the loop (this is because the Kabat numbering scheme places the insertions at H35A and H35B; if neither 35A nor 35B is present, the loop ends at 32; if only 35A is present, the loop ends at 33; if both 35A and 35B are present, the loop ends at 34). The AbM hypervariable regions represent a compromise between the Kabat CDRs and the Chothia structural loops and are used in Oxford Molecular's AbM antibody modeling software.

[0065] TIFF0007692411000001.tif49170

[0066] As used herein, the terms "constant region" and "constant domain" are used interchangeably and have a common meaning in the art. The constant region is the carboxyl-terminal portion of the light and / or heavy chains that is not directly involved in antibody binding to an antigen, for example, an antibody portion, but can exhibit various effector functions such as interaction with Fc receptors. The constant regions of immunoglobulin molecules generally have a more conserved amino acid sequence compared to the immunoglobulin variable domains.

[0067] As used herein, the term "heavy chain," when used in reference to an antibody, refers to any different class based on the amino acid sequence of the constant domain, for example, the classes IgA, IgD, IgE, IgG, and IgM (this includes subclasses of IgG, for example, IgG 1 , IgG 2 , IgG 3 and IgG 4which may refer to alpha (α), delta (δ), epsilon (ε), gamma (γ), and mu (μ), each giving rise to an antibody that includes

[0068] As used herein, the term "light chain," when used in reference to an antibody, may refer to any different type, e.g., kappa (κ) or lambda (λ), based on the amino acid sequence of the constant domain. Light chain amino acid sequences are well known in the art. In some aspects of the present disclosure, the light chain is a human light chain.

[0069] The term "MEDI5884" refers to an anti-EL antibody that includes the heavy chain of SEQ ID NO: 9 and the light chain of SEQ ID NO: 10. MEDI5884 is also referred to as "S6F1-4P" and includes the heavy chain variable region of the h55A1-S6 antibody and the light chain variable region of the h55A1-F1 antibody, as disclosed in U.S. Patent Application Publication No. 2017 / 0260290, which is hereby incorporated by reference in its entirety.

[0070] The term "chimeric" antibody or antigen-binding fragment thereof refers to an antibody or antigen-binding fragment thereof whose amino acid sequence is derived from two or more species. Typically, the variable regions of both the light and heavy chains correspond to the variable regions of an antibody or antigen-binding fragment thereof from a mammalian species (e.g., mouse, rat, rabbit, etc.) having the desired specificity, affinity, and potency, while the constant regions are homologous to the sequences of an antibody or antigen-binding fragment thereof from another species (usually human) to avoid eliciting an immune response in that species.

[0071] The term "humanized" antibody or antigen-binding fragment thereof refers to a specific immunoglobulin chain, chimeric immunoglobulin, or fragment thereof that contains minimal non-human (e.g., murine) sequences in the form of a non-human (e.g., murine) antibody or antigen-binding fragment. Typically, a humanized antibody or antigen-binding fragment thereof is a human immunoglobulin in which the residues from the complementarity-determining regions (CDRs) have been replaced by residues from the CDRs of a non-human species (e.g., mouse, rat, rabbit, hamster) having the desired specificity, affinity, and capacity ("CDR-grafted") (Jones et al., Nature 321:522-525 (1986), Riechmann et al., Nature 332:323-327 (1988), Verhoeyen et al., Science 239:1534-1536 (1988)). In some cases, certain Fv framework region (FR) residues of the human immunoglobulin are replaced by the corresponding residues in an antibody or fragment derived from a non-human species having the desired specificity, affinity, and capacity. The humanized antibody or antigen-binding fragment thereof can be further modified by replacement of additional residues either in the Fv framework region and / or within the non-human CDR residues to refine and optimize the specificity, affinity, and / or capacity of the antibody or antigen-binding fragment. Generally, a humanized antibody or antigen-binding fragment thereof will contain a variable domain that includes all or substantially all of the CDR regions corresponding to the non-human immunoglobulin, but all or substantially all of the FR regions are those of the consensus sequence of the human immunoglobulin. A humanized antibody or antigen-binding fragment thereof can also include the constant region or constant domains (Fc) of the immunoglobulin, typically at least a portion of the constant region or constant domains (Fc) of the human immunoglobulin. Examples of methods used to produce humanized antibodies are U.S. Patent No. 5,225,539, Roguska et al., Proc. Natl. Acad. Sci., USA, 91(3):969-973 (1994), and Roguska et al., Protein It is described in Eng. 9(10):895-904 (1996). In some aspects of the present disclosure, a "humanized antibody" is a resurfaced antibody.

[0072] The term "human" antibody or antigen-binding fragment thereof means an antibody or antigen-binding fragment thereof having an amino acid sequence derived from the human immunoglobulin locus, where such antibody or antigen-binding fragment is made using any technique known in the art. This definition of a human antibody or antigen-binding fragment thereof includes intact or full-length antibodies and fragments thereof.

[0073] "Binding affinity" generally refers to the total strength of non-covalent interactions between a single binding site of a molecule (e.g., an antibody or antigen-binding fragment thereof) and its binding partner (e.g., an antigen). Unless otherwise indicated, as used herein, "binding affinity" refers to the intrinsic binding affinity that reflects the 1:1 interaction between members of a binding pair (e.g., an antibody or antigen-binding fragment thereof and an antigen). The affinity of molecule X for its partner Y can generally be represented by the dissociation constant (K D ). Affinity can be measured and / or represented in many ways known in the art, including, but not limited to, the equilibrium dissociation constant (K D ) and the equilibrium association constant (K A ). K D is calculated from the quotient of k off / k on , while K A is calculated from the quotient of k on / k off . k on refers to the association rate constant of an antibody or antigen-binding fragment thereof for an antigen, for example, and k off refers to the dissociation of an antibody or antigen-binding fragment thereof from an antigen, for example. k on and k off can be determined by techniques known to those of skill in the art, such as BIAcore™ or KinExA.

[0074] As used herein, "epitope" is a term in the art and refers to the localized region of an antigen to which an antibody or an antigen-binding fragment thereof can specifically bind. An epitope may be, for example, adjacent amino acids of a polypeptide (linear or contiguous epitopes), or an epitope may be, for example, derived together from two or more non-adjacent regions of a polypeptide(s) (conformational, non-linear, discontinuous, or non-adjacent epitopes). In some aspects of the present disclosure, the epitope to which an antibody or an antigen-binding fragment thereof specifically binds can be identified, for example, by NMR spectroscopy, X-ray crystallographic studies, ELISA assays, hydrogen / deuterium exchange combined with mass spectrometry (e.g., liquid chromatography electrospray mass spectrometry), array-based oligopeptide scanning assays, and / or mutagenesis mapping (e.g., site-directed mutagenesis mapping). For X-ray crystallographic studies, crystallization can be performed using any of the methods known in the art (e.g., such as Giege R et al., (1994) Acta Crystallogr D Biol Crystallogr 50(Pt 4): 339-350, McPherson A (1990) Eur J Biochem 189: 1-23, Chayen NE (1997) Structure 5: 1269-1274, McPherson A (1976) J Biol Chem 251: 6300-6303). Antibody / its antigen-binding fragment:antigen crystals can be studied using well-known X-ray diffraction techniques, such as X-PLOR (Yale University, 1992, distributed by Molecular Simulations, Inc.; e.g., Meth Enzymol (1985) volumes 114 & 115, eds Wyckoff See HW et al., U.S. Patent Application Publication No. 2004 / 0014194), and can be refined using computer software such as BUSTER (Bricogne G (1993) Acta Crystallogr D Biol Crystallogr 49(Pt 1): 37-60, Bricogne G (1997) Meth Enzymol 276A: 361-423, ed Carter CW, Roversi P et al., (2000) Acta Crystallogr D Biol Crystallogr 56(Pt 10): 1316-1323). Mutagenesis mapping studies can be performed using any method known to those skilled in the art. For example, for descriptions of mutagenesis techniques, including alanine scanning mutagenesis techniques, see Champe M et al., (1995) J Biol Chem 270: 1388-1394, and Cunningham BC & Wells JA (1989) Science 244: 1081-1085.

[0075] An antibody that "binds to the same epitope" as a reference antibody refers to an antibody that binds to the same amino acid residues as the reference antibody. The ability of an antibody to bind to the same epitope as a reference antibody can be determined by hydrogen / deuterium exchange assays (see Coales et al. Rapid Commun. Mass Spectrom. 2009; 23:639-647), or by X-ray crystallography studies.

[0076] An antibody is said to "competitively inhibit" the binding of a reference antibody to a given epitope if it preferentially binds to that epitope or an overlapping epitope to such an extent that it blocks the binding of the reference antibody to the epitope to some degree. Competitive inhibition can be determined by any method known in the art, such as a competitive ELISA assay. An antibody can be said to competitively inhibit the binding of a reference antibody to a given epitope by at least 90%, at least 80%, at least 70%, at least 60%, or at least 50%.

[0077] As used herein, an "inhibitor of PCSK9" is an agent that blocks the interaction between PCSK9 and the LDL receptor.

[0078] An "isolated" polypeptide, antibody, polynucleotide, vector, cell, or composition is a polypeptide, antibody, polynucleotide, vector, cell, or composition in a form not found in nature. Isolated polypeptides, antibodies, polynucleotides, vectors, cells, or compositions include those that have been purified to the extent that they are no longer in their natural form. In some embodiments of the present disclosure, the isolated antibody, polynucleotide, vector, cell, or composition is substantially pure. As used herein, "substantially pure" means that the substance is at least 50% pure (i.e., free of contaminants), at least 90% pure, at least 95% pure, at least 98% pure, or at least 99% pure.

[0079] The terms "polypeptide," "peptide," and "protein" are used interchangeably herein and refer to a polymer of amino acids of any length. The polymer can be linear or branched, can contain modified amino acids, and can be interrupted by non-amino acids. The term also includes amino acid polymers that have been modified naturally or by any other manipulation or modification, such as disulfide bond formation, glycosylation, lipidation, acetylation, phosphorylation, or conjugation to a labeling component. Also included within the definition are polypeptides containing, for example, one or more analogs of amino acids (including, for example, non-natural amino acids, etc.) and other modifications known in the art. Since the polypeptides of the present disclosure are antibody-based, in some embodiments of the present disclosure, the polypeptide can exist as a single chain or associated chains.

[0080] As used herein, the term "host cell" can be any type of cell, e.g., a primary cell, a cell in culture, or a cell derived from a cell line. In some aspects of the present disclosure, the term "host cell" refers to a cell transfected with a nucleic acid molecule and the progeny or potential progeny of such a cell. The progeny of such a cell may not be identical to the parent cell transfected with the nucleic acid molecule, e.g., due to mutations that may occur in subsequent generations or environmental influences, or integration of the nucleic acid molecule into the host cell genome.

[0081] The term "pharmaceutical formulation" refers to a formulation in a form that enables the biological activity of the active ingredient to be effective and that does not contain additional ingredients that are unacceptably toxic to the subject to whom the formulation is administered. The formulation may be sterile.

[0082] The terms "administer", "administering", "administration", etc., as used herein, refer to methods (e.g., intravenous administration) that can be used to enable delivery of a drug, e.g., an anti-EL antibody or an antigen-binding fragment thereof, to a desired biological site of action. Administration techniques that can be used with the agents and methods described herein can be found, e.g., in Goodman and Gilman, The Pharmacological Basis of Therapeutics, current edition, Pergamon; and Remington's, Pharmaceutical Sciences, current edition, Mack Publishing Co., Easton, Pa.

[0083] As used herein, the terms "subject" and "patient" are used interchangeably. The subject can be an animal. In some aspects of the present disclosure, the subject is a mammal such as a non-human animal (e.g., a cow, pig, horse, cat, dog, rat, mouse, monkey or other primate, etc.). In some aspects of the present disclosure, the subject is a cynomolgus monkey. In some aspects of the present disclosure, the subject is a human.

[0084] The term "therapeutically effective amount" refers to the amount of a drug, such as an anti-EL antibody or an antigen-binding fragment thereof, that is effective in treating a disease or disorder in a subject. The terms "treat," "treatment," "treating," "alleviate," and "alleviating" refer to therapeutic measures that cure, retard, reduce the symptoms of, and / or halt the progression of a pathological condition or disorder. Thus, subjects in need of treatment include those already diagnosed with or suspected of having the disorder. Administration "in combination with" one or more additional therapeutic agents includes simultaneous (co-administration) or sequential administration in any order.

[0085] As used in this disclosure and the claims, the singular forms "a," "an," and "the" specifying a quantity do not exclude the plural unless the context clearly dictates otherwise.

[0086] It is to be understood that whenever aspects of this disclosure are described herein with the word "comprising," similar aspects described in terms of "consisting of" and / or "consisting essentially of" are also provided.

[0087] As used herein, the term "or" is understood to be inclusive unless specifically stated otherwise or otherwise apparent from the context. The term "and / or" as used in phrases such as "A and / or B" herein is intended to include both "A and B," "A or B," "A," and "B." Similarly, the term "and / or" as used in phrases such as "A, B, and / or C" is intended to include A, B, and C; A, B, or C; A or C; A or B; B or C; A and C; A and B; B and C; A (alone); B (alone); and C (alone).

[0088] As used herein, the terms "about" and "approximately," when used to modify a numerical value or numerical range, indicate that a deviation of up to 5% to 10% above and 5% to 10% below the value or range is also within the intended meaning of the specified value or range.

[0089] Any composition or method provided herein can be combined with one or more of any of the other compositions and methods provided herein.

[0090] 5.2 Therapeutic methods using anti-EL antibodies or antigen-binding fragments thereof Provided herein is a method of administering to a subject in need thereof an anti-EL antibody or antigen-binding fragment thereof described herein, or a pharmaceutical composition thereof described herein.

[0091] As shown herein, administration of an anti-EL antibody or antigen-binding fragment thereof, or a pharmaceutical composition thereof, can treat cardiovascular diseases in a subject (e.g., a human subject). The cardiovascular disease can be, for example, coronary artery disease, coronary heart disease, cerebrovascular disease, or peripheral artery disease.

[0092] Administration of an anti-EL antibody or antigen-binding fragment thereof, or a pharmaceutical composition thereof, can reduce or prevent atherosclerosis in a subject (e.g., a human subject).

[0093] Administration of an anti-EL antibody or antigen-binding fragment thereof, or a pharmaceutical composition thereof, can prevent or reduce the risk of secondary cardiovascular events in a subject (e.g., a human subject).

[0094] Administration of an anti-EL antibody or an antigen-binding fragment thereof, or a pharmaceutical composition thereof, can reduce the risk of cardiovascular death, non-fatal myocardial infarction (MI), non-fatal stroke, coronary artery revascularization, or any combination thereof in a subject (e.g., a human subject). The subject can be, for example, a subject with a history of acute coronary syndrome (ACS).

[0095] Administration of an anti-EL antibody or an antigen-binding fragment thereof, or a pharmaceutical composition thereof, can prevent or reduce the risk of major adverse cardiovascular events (MACE) in a subject (e.g., a human subject).

[0096] Administration of an anti-EL antibody or an antigen-binding fragment thereof, or a pharmaceutical composition thereof, can (i) increase high-density lipoprotein cholesterol (HDL-C), (ii) increase the number of high-density lipoprotein (HDL) particles, (iii) increase HDL particle size, (iv) increase HDL phospholipids, (v) increase ApoA1, (vi) increase cholesterol efflux capacity (CEC), or (vii) be any combination thereof.

[0097] Administration of an anti-EL antibody or an antigen-binding fragment thereof, or a pharmaceutical composition thereof, can also increase plasma phosphatidylinositol (PI) levels.

[0098] In some embodiments, administration of an anti-EL antibody or an antigen-binding fragment thereof inhibits EL.

[0099] In some embodiments, administration of an anti-EL antibody or an antigen-binding fragment thereof in a subject Increases HDL-C. Administration of an anti-EL antibody or an antigen-binding fragment thereof can increase HDL-C by, for example, at least 30%, at least 35%, or at least 40%. Thus, administration of an anti-EL antibody or an antigen-binding fragment thereof can increase HDL-C by about 30% to about 125%, about 30% to about 100%, about 30% to about 75%, about 30% to about 50%, about 30% to about 45%, or about 30% to about 40%. The increase in HDL-C can occur within 30 days, within 60 days, or within 90 days from the first administration.

[0100] In some embodiments, administration of an anti-EL antibody or an antigen-binding fragment thereof increases the number of HDL particles in a subject. Administration of an anti-EL antibody or an antigen-binding fragment thereof can increase the number of HDL particles by, for example, at least 5%, at least 8%, at least 10%, or at least 15% (e.g., as measured using NMR). Thus, administration of an anti-EL antibody or an antigen-binding fragment thereof can increase the number of HDL particles by about 5% to about 20%, or about 5% to about 18% (e.g., as measured using NMR). The increase in the number of HDL particles can occur within 30 days, within 60 days, or within 90 days from the first administration.

[0101] In some embodiments, administration of an anti-EL antibody or an antigen-binding fragment thereof increases the HDL particle size in a subject. Administration of an anti-EL antibody or an antigen-binding fragment thereof can increase the HDL particle size by, for example, at least 3%, or at least 5%. Thus, administration of an anti-EL antibody or an antigen-binding fragment thereof can increase the HDL particle size by about 3% to about 10%, or about 3% to about 7%. The increase in HDL particle size can occur within 30 days, within 60 days, or within 90 days from the first administration.

[0102] In some embodiments, administration of an anti-EL antibody or an antigen-binding fragment thereof increases HDL phospholipids in a subject. Administration of the anti-EL antibody or an antigen-binding fragment thereof can increase HDL phospholipids, for example, by at least 50%. The increase in HDL phospholipids can occur within 30 days, within 60 days, or within 90 days from the first administration.

[0103] In some embodiments, administration of an anti-EL antibody or an antigen-binding fragment thereof increases apolipoprotein A1 (apoA1) in a subject. Administration of the anti-EL antibody or an antigen-binding fragment thereof can increase apoA1, for example, by at least 30%. Thus, administration of the anti-EL antibody or an antigen-binding fragment thereof can increase apoA1 by about 30% to about 40%, or about 30% to about 35%. The increase in apoA1 can occur within 30 days, within 60 days, or within 90 days from the first administration.

[0104] In some embodiments, administration of an anti-EL antibody or an antigen-binding fragment thereof increases cholesterol efflux capacity (CEC) in a subject. CEC can be measured using, for example, the method described in Thacker et al., Journal of Lipid Research 56: 1282-1295 (2015). Administration of the anti-EL antibody or an antigen-binding fragment thereof can increase the cholesterol efflux capacity of non-ATP-binding cassette transporter A1 (ABCA1), for example, by at least 30%, or at least 35%. Thus, administration of the anti-EL antibody or an antigen-binding fragment thereof can increase the non-ABCA1 cholesterol efflux capacity by about 30% to about 40%, or about 30% to about 35%.

[0105] In some embodiments, administration of an anti-EL antibody or an antigen-binding fragment thereof increases plasma phosphatidylinositol (PI) levels in a subject. In some embodiments Moreover, the increase in PI level is an increase in the levels of PI(14:2 / 20:0), PI(14:2 / 22:0), PI(14:2 / 22:1), PI(14:2 / 22:2), PI(16:0 / 16:1), PI(16:0 / 18:0), PI(16:0 / 18:2), PI(16:0 / 20:2), PI(16:0 / 20:3), PI(16:0 / 20:4), PI(16:0 / 22:4), PI(16:1 / 18:0), PI(16:1 / 18:1), PI(18:0 / 18:0), PI(18:0 / 18:1), PI(18:0 / 18:2), PI(18:0 / 18:3), PI(18:0 / 20:2), PI(18:0 / 20:3), PI(18:0 / 20:4), PI(18:0 / 22:4), PI(18:0 / 22:5), PI(18:0 / 22:6), PI(18:1 / 16:0), PI(18:1 / 18:1), PI(18:1 / 18:2), PI(18:1 / 20:2), PI(18:1 / 20:3), PI(18:1 / 20:4), and / or PI(18:2 / 18:2). In some embodiments, the increase in PI level is an increase in the levels of PI(14:2 / 22:2), PI(16:0 / 16:1), PI(16:0 / 18:2), PI(16:0 / 20:3), PI(16:0 / 20:4), PI(18:0 / 18:1), PI(18:0 / 18:2), PI(18:0 / 20:2), PI(18:0 / 20:3), PI(18:0 / 20:4), PI(18:0 / 22:6), and / or PI(18:1 / 16:0). Administration of an anti-EL antibody or an antigen-binding fragment thereof can increase plasma PI levels by, for example, at least several hundred percent for more abundant PI species and by several hundred percent to several thousand percent from baseline for less abundant PI species. Thus, administration of an anti-EL antibody or an antigen-binding fragment thereof can increase plasma PI by about 100% to 1000% depending on the PI species.In some cases, administration of an anti-EL antibody or an antibody-binding fragment thereof increases the levels of at least 10 plasma PI species, for example, by at least 100% or 100% - 1000%. In some cases, administration of an anti-EL antibody or an antigen-binding fragment thereof increases the levels of at least 12 plasma PI species (e.g., PI(14:2 / 22:2), PI(16:0 / 16:1), PI(16:0 / 18:2), PI(16:0 / 20:3), PI(16:0 / 20:4), PI(18:0 / 18:1), PI(18:0 / 18:2), PI(18:0 / 20:2), PI(18:0 / 20:3), PI(18:0 / 20:4), PI(18:0 / 22:6), and PI(18:1 / 16:0)), for example, by at least 100% or 100% - 1000%. In some cases, administration of an anti-EL antibody or an antigen-binding fragment thereof increases the levels of at least 30 plasma PI species (e.g., PI(14:2 / 20:0), PI(14:2 / 22:0), PI(14:2 / 22:1), PI(14:2 / 22:2), PI(16:0 / 16:1), PI(16:0 / 18:0), PI(16:0 / 18:2), PI(16:0 / 20:2), PI(16:0 / 20:3), PI(16:0 / 20:4), PI(16:0 / 22:4), PI(16:1 / 18:0), PI(16:1 / 18:1), PI(18:0 / 18:0), PI(18:0 / 18:1), PI(18:0 / 18:2), PI(18:0 / 18:3), PI(18:0 / 20:2), PI(18:0 / 20:3), PI(18:0 / 20:4), PI(18:0 / 22:4), PI(18:0 / 22:5), PI(18:0 / 22:6), PI(18:1 / 16:0), PI(18:1 / 18:1), PI(18:1 / 18:2), PI(18:1 / 20:2), PI(18:1 / 20:3), PI(18:1 / 20:4), and PI(18:2 / 18:2)), for example, by at least 100% or 100% - 1000%.

[0106] In some cases, administration of the anti-EL antibody or an antibody binding fragment thereof increases the levels of at least 10 plasma PI species by at least 250% or 250% to 1000%. In some cases, administration of the anti-EL antibody or an antigen binding fragment thereof increases the levels of at least 12 plasma PI species (e.g., PI(14:2 / 22:2), PI(16:0 / 16:1), PI(16:0 / 18:2), PI(16:0 / 20:3), PI(16:0 / 20:4), PI(18:0 / 18:1), PI(18:0 / 18:2), PI(18:0 / 20:2), PI(18:0 / 20:3), PI(18:0 / 20:4), PI(18:0 / 22:6), and PI(18:1 / 16:0)) by at least 250% or 250% to 1000%. In some cases, administration of the anti-EL antibody or an antigen binding fragment thereof increases the levels of at least 30 plasma PI species (e.g., PI(14:2 / 20:0), PI(14:2 / 22:0), PI(14:2 / 22:1), PI(14:2 / 22:2), PI(16:0 / 16:1), PI(16:0 / 18:0), PI(16:0 / 18:2), PI(16:0 / 20:2), PI(16:0 / 20:3), PI(16:0 / 20:4), PI(16:0 / 22:4), PI(16:1 / 18:0), PI(16:1 / 18:1), PI(18:0 / 18:0), PI(18:0 / 18:1), PI(18:0 / 18:2), PI(18:0 / 18:3), PI(18:0 / 20:2), PI(18:0 / 20:3), PI(18:0 / 20:4), PI(18:0 / 22:4), PI(18:0 / 22:5), PI(18:0 / 22:6), PI(18:1 / 16:0), PI(18:1 / 18:1), PI(18:1 / 18:2), PI(18:1 / 20:2), PI(18:1 / 20:3), PI(18:1 / 20:4), and PI(18:2 / 18:2)) by at least 250% or 250% to 1000%.

[0107] The increase in plasma PI can occur within 90 days of the first administration.

[0108] In some embodiments, the anti-EL antibody or its antigen-binding fragment, or a pharmaceutical composition thereof, is administered at a dose of about 100 mg to about 350 mg. In some embodiments, the anti-EL antibody or its antigen-binding fragment, or a pharmaceutical composition thereof, is administered at a dose of about 100 mg to about 250 mg. In some embodiments, the anti-EL antibody or its antigen-binding fragment, or a pharmaceutical composition thereof, is administered at a dose of about 100 mg to about 200 mg. The dose of the antibody or its antigen-binding fragment, or a pharmaceutical composition thereof, can be administered parenterally, such as subcutaneously. The dose of the antibody or its antigen-binding fragment, or a pharmaceutical composition thereof, can be administered using an attached prefilled syringe (APFS) or an autoinjector. The dose of the antibody or its antigen-binding fragment, or a pharmaceutical composition thereof, can be administered about once a month.

[0109] In some embodiments, the anti-EL antibody or its antigen-binding fragment, or a pharmaceutical composition thereof, is administered at a dose of about 200 mg to about 350 mg. In some embodiments, the anti-EL antibody or its antigen-binding fragment, or a pharmaceutical composition thereof, is administered at a dose of about 200 mg to about 300 mg. In some embodiments, the anti-EL antibody or its antigen-binding fragment, or a pharmaceutical composition thereof, is administered at a dose of about 200 mg to about 250 mg. The dose of the antibody or its antigen-binding fragment, or a pharmaceutical composition thereof, can be administered parenterally, such as subcutaneously. The dose of the antibody or its antigen-binding fragment, or a pharmaceutical composition thereof, can be administered using an attached prefilled syringe (APFS) or an autoinjector. The dose of the antibody or its antigen-binding fragment, or a pharmaceutical composition thereof, can be administered about once a month.

[0110] In some embodiments, the anti-EL antibody or its antigen-binding fragment, or a pharmaceutical composition thereof, is administered at a dose of about 250 mg to about 300 mg. In some embodiments, the anti-EL antibody or its antigen-binding fragment, or a pharmaceutical composition thereof, is administered at a dose of about 250 mg to about 350 mg. The dose of the antibody or its antigen-binding fragment, or a pharmaceutical composition thereof, can be administered parenterally, for example, subcutaneously. The dose of the antibody or its antigen-binding fragment, or a pharmaceutical composition thereof, can be administered using an attached prefilled syringe (APFS) or an autoinjector. The antibody or its antigen-binding fragment, or a pharmaceutical composition thereof, can be administered about once a month.

[0111] In some embodiments, the anti-EL antibody or an antigen-binding fragment thereof, or a pharmaceutical composition thereof, is administered at a dose of about 100 mg. In some embodiments, the anti-EL antibody or an antigen-binding fragment thereof, or a pharmaceutical composition thereof, is administered at a dose of about 110 mg. In some embodiments, the anti-EL antibody or an antigen-binding fragment thereof, or a pharmaceutical composition thereof, is administered at a dose of about 120 mg. In some embodiments, the anti-EL antibody or an antigen-binding fragment thereof, or a pharmaceutical composition thereof, is administered at a dose of about 125 mg. In some embodiments, the anti-EL antibody or an antigen-binding fragment thereof, or a pharmaceutical composition thereof, is administered at a dose of about 130 mg. In some embodiments, the anti-EL antibody or an antigen-binding fragment thereof, or a pharmaceutical composition thereof, is administered at a dose of about 140 mg. In some embodiments, the anti-EL antibody or an antigen-binding fragment thereof, or a pharmaceutical composition thereof, is administered at a dose of about 150 mg. In some embodiments, the anti-EL antibody or an antigen-binding fragment thereof, or a pharmaceutical composition thereof, is administered at a dose of about 160 mg. In some embodiments, the anti-EL antibody or an antigen-binding fragment thereof, or a pharmaceutical composition thereof, is administered at a dose of about 170 mg. In some embodiments, the anti-EL antibody or an antigen-binding fragment thereof, or a pharmaceutical composition thereof, is administered at a dose of about 175 mg. In some embodiments, the anti-EL antibody or an antigen-binding fragment thereof, or a pharmaceutical composition thereof, is administered at a dose of about 180 mg. In some embodiments, the anti-EL antibody or an antigen-binding fragment thereof, or a pharmaceutical composition thereof, is administered at a dose of about 190 mg. In some embodiments, the anti-EL antibody or an antigen-binding fragment thereof, or a pharmaceutical composition thereof, is administered at a dose of about 200 mg. In some embodiments, the anti-EL antibody or an antigen-binding fragment thereof, or a pharmaceutical composition thereof, is administered at a dose of about 210 mg. In some embodiments, the anti-EL antibody or an antigen-binding fragment thereof, or a pharmaceutical composition thereof, is administered at a dose of about 220 mg. In some embodiments, the anti-EL antibody or an antigen-binding fragment thereof, or a pharmaceutical composition thereof, is administered at a dose of about 225 mg.In some embodiments, the anti-EL antibody or antigen-binding fragment thereof, or a pharmaceutical composition thereof, is administered at a dose of about 230 mg. In some embodiments, the anti-EL antibody or antigen-binding fragment thereof, or a pharmaceutical composition thereof, is administered at a dose of about 240 mg. In some embodiments, the anti-EL antibody or antigen-binding fragment thereof, or a pharmaceutical composition thereof, is administered at a dose of about 250 mg. In some embodiments, the anti-EL antibody or antigen-binding fragment thereof, or a pharmaceutical composition thereof, is administered at a dose of about 260 mg. In some embodiments, the anti-EL antibody or antigen-binding fragment thereof, or a pharmaceutical composition thereof, is administered at a dose of about 270 mg. In some embodiments, the anti-EL antibody or antigen-binding fragment thereof, or a pharmaceutical composition thereof, is administered at a dose of about 275 mg. In some embodiments, the anti-EL antibody or antigen-binding fragment thereof, or a pharmaceutical composition thereof, is administered at a dose of about 280 mg. In some embodiments, the anti-EL antibody or antigen-binding fragment thereof, or a pharmaceutical composition thereof, is administered at a dose of about 290 mg. In some embodiments, the anti-EL antibody or antigen-binding fragment thereof, or a pharmaceutical composition thereof, is administered at a dose of about 300 mg. In some embodiments, the anti-EL antibody or antigen-binding fragment thereof, or a pharmaceutical composition thereof, is administered at a dose of about 310 mg. In some embodiments, the anti-EL antibody or antigen-binding fragment thereof, or a pharmaceutical composition thereof, is administered at a dose of about 320 mg. In some embodiments, the anti-EL antibody or antigen-binding fragment thereof, or a pharmaceutical composition thereof, is administered at a dose of about 325 mg. In some embodiments, the anti-EL antibody or antigen-binding fragment thereof, or a pharmaceutical composition thereof, is administered at a dose of about 330 mg. In some embodiments, the anti-EL antibody or antigen-binding fragment thereof, or a pharmaceutical composition thereof, is administered at a dose of about 340 mg. In some embodiments, the anti-EL antibody or... The antibody or its antigen-binding fragment, or their pharmaceutical composition, is administered at a dose of about 350 mg. The dose of the antibody or its antigen-binding fragment, or their pharmaceutical composition, can be administered parenterally, such as subcutaneously. The dose of the antibody or its antigen-binding fragment, or their pharmaceutical composition, can be administered using an attached prefilled syringe (APFS) or an autoinjector. The dose of the antibody or its antigen-binding fragment, or their pharmaceutical composition, can be administered about once a month.

[0112] In some embodiments, the anti-EL antibody or its antigen-binding fragment, or their pharmaceutical composition, is administered at a dose of about 125 mg or 125 mg. The dose of about 125 mg or 125 mg of the antibody or its antigen-binding fragment, or their pharmaceutical composition, can be administered parenterally, such as subcutaneously. The dose of about 125 mg or 125 mg of the antibody or its antigen-binding fragment, or their pharmaceutical composition, can be administered using an attached prefilled syringe (APFS) or an autoinjector.

[0113] In some embodiments, the anti-EL antibody or its antigen-binding fragment, or their pharmaceutical composition, is administered at a dose of about 250 mg or 250 mg. The dose of about 250 mg or 250 mg of the antibody or its antigen-binding fragment, or their pharmaceutical composition, can be administered parenterally, such as subcutaneously. The dose of about 250 mg or 250 mg of the antibody or its antigen-binding fragment, or their pharmaceutical composition, can be administered using an attached prefilled syringe (APFS) or an autoinjector.

[0114] In some embodiments, the anti-EL antibody or its antigen-binding fragment, or a pharmaceutical composition thereof, is administered about once a month or once a month. The antibody or its antigen-binding fragment, or a pharmaceutical composition thereof, administered about once a month or once a month can be administered parenterally, for example, subcutaneously. The antibody or its antigen-binding fragment, or a pharmaceutical composition thereof, administered about once a month or once a month can be administered using an attached prefilled syringe (APFS) or an autoinjector.

[0115] In some embodiments, the anti-EL antibody or its antigen-binding fragment, or a pharmaceutical composition thereof, is administered about once a month at a dose of about 125 mg. The dose of about 125 mg administered about once a month can be administered parenterally, for example, subcutaneously. The dose of about 125 mg administered about once a month can be administered using an attached prefilled syringe (APFS) or an autoinjector.

[0116] In some embodiments, the anti-EL antibody or its antigen-binding fragment, or a pharmaceutical composition thereof, is administered about once a month at a dose of about 250 mg. The dose of about 250 mg administered about once a month can be administered parenterally, for example, subcutaneously. The dose of about 250 mg administered about once a month can be administered using an attached prefilled syringe (APFS) or an autoinjector.

[0117] In some embodiments, the anti-EL antibody or its antigen-binding fragment, or a pharmaceutical composition thereof, is administered once a month at a dose of 125 mg. The dose of 125 mg administered once a month can be administered parenterally, for example, subcutaneously. The dose of about 125 mg administered about once a month can be administered using an attached prefilled syringe (APFS) or an autoinjector.

[0118] In some embodiments, the anti-EL antibody or its antigen-binding fragment, or a pharmaceutical composition thereof, is administered once a month at a dose of 250 mg. The 250 mg dose administered once a month can be administered parenterally, such as subcutaneously. The approximately 250 mg dose administered about once a month can be administered using an attached prefilled syringe (APFS) or an autoinjector.

[0119] According to the methods provided herein, the anti-EL antibody or its antigen-binding fragment, or a pharmaceutical composition comprising the anti-EL antibody or its antigen-binding fragment, can be administered parenterally. In some embodiments of the present disclosure, the anti-EL antibody or its antigen-binding fragment, or a pharmaceutical composition comprising the anti-EL antibody or its antigen-binding fragment, is administered subcutaneously.

[0120] In some embodiments of the present disclosure, the anti-EL antibody or its antigen-binding fragment is administered for at least 3 months (e.g., about once a month). In some embodiments of the present disclosure, the anti-EL antibody or its antigen-binding fragment is administered for at least 12 months (e.g., about once a month). In some embodiments of the present disclosure, the anti-EL antibody or its antigen-binding fragment is administered for at least 24 months (e.g., about once a month).

[0121] In some embodiments of the present disclosure, the present disclosure relates to the anti-EL antibody or its antigen-binding fragment, or a pharmaceutical composition provided herein, which is used as a pharmaceutical administered at about 100 mg to about 350 mg or about 200 mg to about 350 mg (e.g., 250 mg). In some embodiments of the present disclosure, the present disclosure relates to the anti-EL antibody or its antigen-binding fragment, or a pharmaceutical composition provided herein, which is used as a pharmaceutical administered at about 100 mg to about 350 mg or about 200 mg to about 350 mg (e.g., about 250 mg).

[0122] In some aspects of the present disclosure, the present disclosure relates to an anti-EL antibody or an antigen-binding fragment thereof, or a pharmaceutical composition provided herein, which is used as a medicament administered approximately once a month at about 100 mg to about 350 mg or about 200 mg to about 350 mg (e.g., 250 mg). In some aspects of the present disclosure, the present disclosure relates to an anti-EL antibody or an antigen-binding fragment thereof, or a pharmaceutical composition provided herein, which is used as a medicament administered once a month at about 100 mg to about 350 mg or about 200 mg to about 350 mg (e.g., about 250 mg).

[0123] According to the methods provided herein, an anti-EL antibody or an antigen-binding fragment thereof, or a pharmaceutical composition comprising an anti-EL antibody or an antigen-binding fragment thereof can be administered in combination with an inhibitor of PCSK9. Inhibitors of PCSK9 are disclosed, for example, in Chaudhary et al., World J. Cardiol. 9: 76-91 (2017) and Chodorge et al., Sci. Rep. 8: 17545 (2018) (each of which is incorporated herein by reference in its entirety). An inhibitor of PCSK9 can be, for example, an antibody or an antigen-binding fragment that binds to PCSK9. Examples of antibodies or antigen-binding fragments that inhibit PCSK9 include HS9, alirocumab, evolocumab, and bococizumab.

[0124] The HS9 antibody (in the context of a GLP-1 fusion protein called MEDI4166) is disclosed in Chodorge et al., Sci. Rep. 8: 17545 (2018) and PCT International Publication No. WO 2015 / 127273 (each of which is incorporated herein by reference in its entirety). The HS9 antibody comprises the following variable heavy chain sequence and variable light chain sequence: HS9 variable heavy chain sequence: QVQLVQSGAEVKKPGASVKVSCKASGYTFTSYYMHWVRQAPGQGLEWMGEISPSGGSTSYNQKFQGRVTMTRDTSTSTVYMELSSLRSEDTAVYYCARERPLYASDLWGQGTTVTVSS (SEQ ID NO: 14) HS9 variable light chain sequence: DIQMTQSPSSLSASVGDRVTITCQASQDVKTAVAWYQQKPGKAPKLLIYSASYRYTGVPSRFSGSGSGTDFTFTISSLQPEDIATYYCQQRYSLWRTFGQGTKLEIK (SEQ ID NO: 15).

[0125] In some aspects of the present disclosure, an antibody that inhibits PCSK9 or an antigen-binding fragment thereof comprises the variable heavy chain sequence of SEQ ID NO: 14. In some aspects of the present disclosure, an antibody that inhibits PCSK9 or an antigen-binding fragment thereof comprises the variable light chain sequence of SEQ ID NO: 15. In some aspects of the present disclosure, an antibody that inhibits PCSK9 or an antigen-binding fragment thereof comprises the variable heavy chain sequence of SEQ ID NO: 14 and the variable light chain sequence of SEQ ID NO: 15.

[0126] In some aspects of the present disclosure, an antibody that inhibits PCSK9 comprises a human IgG1 heavy chain. In some aspects of the present disclosure, an antibody that inhibits PCSK9 comprises a human IgG1 heavy chain containing the triple mutation L234F / L235E / P331S (“IgG1-TM”). In some aspects of the present disclosure, an antibody that inhibits PCSK9 comprises a human kappa light chain. In some aspects of the present disclosure, an antibody that inhibits PCSK9 comprises a) an IgG1-TM heavy chain containing the variable heavy chain sequence of SEQ ID NO: 14 and b) a kappa light chain containing the variable light chain sequence of SEQ ID NO: 15.

[0127] In some aspects of the present disclosure, an inhibitor of PCSK9 can promote the uptake of LDL-C in HepG2 cells treated with recombinant PCSK9 (e.g., as disclosed in Chodorge et al., Sci. Rep. 8: 17545 (2018)).

[0128] As shown herein, an inhibitor of PCSK9 can be administered simultaneously (in the same pharmaceutical composition or in separate pharmaceutical compositions) or sequentially with an anti-EL antibody or an antigen-binding fragment thereof.

[0129] 5.3 EL Antibodies and Antigen-Binding Fragments Thereof Provided herein is a method of treating cardiovascular disease in a subject (e.g., a human subject), the method comprising administering to the subject an antibody (e.g., a monoclonal antibody such as a chimeric antibody, a humanized antibody, or a human antibody) that specifically binds to EL (e.g., human EL) and an antigen-binding fragment thereof. Exemplary EL antibodies and antigen-binding fragments thereof that can be used in the methods provided herein are known in the art. The amino acid sequence for human EL is known in the art and is provided herein as the sequence of SEQ ID NO: 11 for the mature form of the protein (lacking the leader sequence).

[0130] Mature human EL (lacking the leader sequence): SPVPFGPEGRLEDKLHKPKATQTEVKPSVRFNLRTSKDPEHEGCYLSVGHSQPLEDCSFNMTAKTFFIIHGWTMSGIFENWLHKLVSALHTREKDANVVVVDWLPLAHQLYTDAVNNTRVVGHSIARMLDWLQEKDDFSLGNVHLIGYSLGAHVAGYAGNFVKGTVGRITGLDPAGPMFEGADIHKRLSPDDADFVDVLHTYTRSFGLSIGIQMPVGHIDIYPNGGDFQPGCGLNDVLGSIAYGTITEVVKCEHERAVHLFVDSLVNQDKPSFAFQCTDSNRFKKGICLSCRKNRCNSIGYNAKKMRNKRNSKMYLKTRAGMPFRVYHYQMKIHVFSYKNMGEIEPTFYVTLYGTNADSQTLPLEIVERIEQNATNTFLVYTEEDLGDLLKIQLTWEGASQSWYNLWKEFRSYLSQPRNPGRELNIRRIRVKSGETQRKLTFCTEDPENTSISPGRELWFRKCRDGWRMKNETSPTVELP (SEQ ID NO: 11).

[0131] In some aspects of the present disclosure, the antibody or antigen-binding fragment thereof used in the methods described herein specifically binds to human EL. In some aspects of the present disclosure, the antibody or antigen-binding fragment thereof used in the methods described herein specifically binds to human EL and cynomolgus monkey EL.

[0132] In some aspects of the present disclosure, the anti-EL antibody or antigen-binding fragment thereof binds to human EL with an equilibrium dissociation constant (KD) of about 4.06 nM (e.g., measured using surface plasmon resonance). In some aspects of the present disclosure, the anti-EL antibody or its antigen-binding f The fragment binds to cynomolgus EL with a KD of about 1.56 nM (e.g., measured using surface plasmon resonance). In some aspects of the present disclosure, the anti-EL antibody or antigen-binding fragment thereof binds to human EL with a KD constant of about 4.06 nM and binds to cynomolgus EL with a KD of about 1.56 nM (e.g., measured using surface plasmon resonance).

[0133] In some aspects of the present disclosure, the anti-EL antibody or antigen-binding fragment thereof can neutralize or inhibit EL.

[0134] The ability of the anti-EL antibody or antigen-binding fragment thereof to neutralize EL can be determined using the following protocol: Conditioned medium is incubated at 37 °C for 2 hours at concentrations ranging from 1000 nM to 31.6 pM in half-area 96-well microplates containing assay buffer (20 mM Tris-HCl, 150 mM NaCl, 4 mM CaCl 2 , 0.5% BSA) and HDL (e.g., human HDL) in the presence or absence of the anti-EL antibody or antigen-binding fragment thereof. Following this incubation, free fatty acid release is measured in each well using the NEFA-HR(2) assay kit (Wako Diagnostics, Mountain View, CA). Absorbance at both 550 nm and 660 nm is measured using a SpectraMax M5 plate reader. The value obtained by subtracting the 660 nm reading from the 550 nm reading is used for further analysis. The percent activity "Ax" at antibody concentration "x" is calculated using the following equation: Ax = [(Ex - V0) / (E0 - V0)] × 100 (where "Ex" is the average value of absorbance units in the presence of the enzyme when the inhibitor is included, and "E0" and "V0" are the respective average values of absorbance units in the absence of the inhibitor and in the absence of the enzyme. The 50% inhibitory concentration (IC50) is calculated and plotted using GraphPad Prism.

[0135] The same assay can be performed using VLDL (e.g., human VLDL) as a substrate instead of HDL (e.g., human HDL) to show that the anti-EL antibody does not neutralize hepatic lipase or lipoprotein lipase.

[0136] In some embodiments of the present disclosure, the anti-EL antibody or an antigen-binding fragment thereof has a half-maximal inhibitory concentration (IC 50 ) of about 1.3 nM against human EL. In some embodiments of the present disclosure, the anti-EL antibody or an antigen-binding fragment thereof has an IC 50 of about 1.7 nM against cynomolgus EL. In some embodiments of the present disclosure, the anti-EL antibody or an antigen-binding fragment thereof has an IC 50 of about 1.3 nM against human EL and an IC 50 of about 1.7 nM against cynomolgus EL.

[0137] In some embodiments of the present disclosure, the anti-EL antibody or an antigen-binding fragment thereof does not inhibit very low density lipoprotein (VLDL) lipolysis mediated by either hepatic lipase or lipoprotein lipase.

[0138] In some embodiments of the present disclosure, the anti-EL antibody or an antigen-binding fragment thereof does not block the exchange of cholesterol from HDL particles to LDL particles.

[0139] In some embodiments of the present disclosure, the anti-EL antibody or an antigen-binding fragment thereof increases the delivery of low density lipoprotein (LDL) to the low density lipoprotein receptor (LDLR).

[0140] In some embodiments of the present disclosure, the antibody or its antigen-binding fragment used in the methods described herein specifically binds to human EL and comprises the six CDRs of the MEDI5884 antibody listed as shown in Tables 1 and 2.

[0141] [Table 1]

[0142] [Table 2]

[0143] In some aspects of the present disclosure, the antibody or antigen-binding fragment thereof used in the methods described herein specifically binds to human EL and comprises the VH of the MEDI5884 antibody listed in Table 3.

[0144] [Table 3]

[0145] In some aspects of the present disclosure, the antibody or antigen-binding fragment thereof used in the methods described herein specifically binds to human EL and comprises the VL of the MEDI5884 antibody listed in Table 4.

[0146] [Table 4]

[0147] In some aspects of the present disclosure, the antibody or antigen-binding fragment thereof used in the methods described herein specifically binds to human EL and comprises the VH and VL of the MEDI5884 antibody listed in Tables 3 and 4.

[0148] In some aspects of the present disclosure, the antibody or antigen-binding fragment thereof used in the methods described herein specifically binds to human EL and comprises the heavy chain sequence of the MEDI5884 antibody listed in Table 5.

[0149] [Table 5]

[0150] In some aspects of the present disclosure, the antibody or antigen-binding fragment thereof used in the methods described herein specifically binds to human EL and comprises the light chain sequence of the MEDI5884 antibody listed in Table 6.

[0151] [Table 6]

[0152] In some aspects of the present disclosure, the antibody or antigen-binding fragment used in the methods described herein specifically binds to human EL and comprises the heavy chain sequence and the light chain sequence of the MEDI5884 antibody listed in Tables 5 and 6.

[0153] In some aspects of the present disclosure, the antibodies or antigen-binding fragments thereof used in the methods described herein are described by their VL domain alone or their VH domain alone, or by their three VL CDRs alone or their three VH CDRs alone. For example, see Rader C et al., (1998) PNAS 95: 8910-8915 (incorporated herein by reference in its entirety), which describes humanizing a murine anti-αvβ3 antibody by identifying the complementary light or heavy chain from a human light chain library or heavy chain library, respectively, thereby obtaining a humanized antibody variant having an affinity equal to or higher than that of the original antibody. Also see Clackson T et al., (1991) Nature 352: 624-628 (incorporated herein by reference in its entirety), which describes a method of generating an antibody that specifically binds to a particular antigen by using a particular VL domain (or VH domain) and screening a library for the complementary VH domain or (VL domain); screening generated 14 new partners for a particular VH domain and 13 new partners for a particular VL domain, which were strong binders as determined by ELISA. Also see Kim SJ & Hong HJ, (2007) J Microbiol 45: 572-577 (incorporated herein by reference in its entirety), which describes a method of generating an antibody that specifically binds to a particular antigen by using a particular VH domain and screening a library for the complementary VL domain (e.g., a human VL library); the selected VL domain could then be used to direct the selection of additional complementary (e.g., human) VH domains.

[0154] In some embodiments, the CDRs of an antibody or antigen-binding fragment thereof can be determined according to the Chothia numbering scheme that refers to the positions of the structural loops of immunoglobulins (e.g., see Chothia C & Lesk AM, (1987), J Mol Biol 196: 901-917, Al-Lazik ani B et al., (1997) J Mol Biol 273: 927-948, Chothia C et al., (1992) J Mol Biol 227: 799-817, Tramontano A et al., (1990) J Mol Biol 215(1): 175-82, and U.S. Patent No. 7,709,226). Typically, when using the Kabat numbering rules, the Chothia CDR-H1 loop is present at amino acids 26 to 32, 33, or 34 of the heavy chain, the Chothia CDR-H2 loop is present at amino acids 52 to 56 of the heavy chain, and the Chothia CDR-H3 loop is present at amino acids 95 to 102 of the heavy chain, while the Chothia CDR-L1 loop is present at amino acids 24 to 34 of the light chain, the Chothia CDR-L2 loop is present at amino acids 50 to 56 of the light chain, and the Chothia CDR-L3 loop is present at amino acids 89 to 97 of the light chain. The end of the Chothia CDR-H1 loop when numbered using the Kabat numbering rules varies between H32 and H34 depending on the length of the loop (this is because the Kabat numbering scheme places insertions at H35A and H35B; when neither 35A nor 35B is present, the loop ends at 32; when only 35A is present, the loop ends at 33; when both 35A and 35B are present, the loop ends at 34).

[0155] In some embodiments, provided herein are methods of administering an antibody and antigen-binding fragment thereof that specifically binds to EL (e.g., human EL) and comprises the Chothia VH CDR and VL CDR of the MEDI5884 antibody listed in Tables 3 and 4. In some embodiments of the present disclosure, provided herein are methods of administering an antibody or antigen-binding fragment thereof that specifically binds to EL (e.g., human EL) and comprises one or more CDRs, wherein the Chothia CDR and Kabat CDR have the same amino acid sequence. In some embodiments of the present disclosure, provided herein are methods of administering an antibody and antigen-binding fragment thereof that specifically binds to EL (e.g., human EL) and comprises a combination of Kabat CDR and Chothia CDR.

[0156] In some embodiments of the present disclosure, the CDRs of an antibody or antigen-binding fragment thereof can be determined according to the IMGT numbering system described in Lefranc M-P, (1999) The Immunologist 7: 132-136 and Lefranc M-P et al., (1999) Nucleic Acids Res 27: 209-212. According to the IMGT numbering scheme, VH-CDR1 is at positions 26-35, VH-CDR2 is at positions 51-57, VH-CDR3 is at positions 93-102, VL-CDR1 is at positions 27-32, VL-CDR2 is at positions 50-52, and VL-CDR3 is at positions 89-97. In some embodiments of the present disclosure, provided herein are methods of administering an antibody and antigen-binding fragment thereof that specifically binds to EL (e.g., human EL) and comprises the IMGT VH CDR and VL CDR of the MEDI5884 antibody listed in Tables 3 and 4, such as those described in Lefranc M-P (1999) and Lefranc M-P (1999) above.

[0157] In some embodiments, the CDRs of an antibody or antigen-binding fragment thereof are from MacCallum RM It can be determined according to et al., (1996) J Mol Biol 262: 732-745. For example, see "Protein Sequence and Structure Analysis of Antibody Variable Domains" by Martin A. in Antibody Engineering, edited by Kontermann and Duebel, Chapter 31, pages 422-439, Springer-Verlag, Berlin (2001). In some aspects of the present disclosure, provided herein is a method of administering an antibody or an antigen-binding fragment thereof that specifically binds to EL (e.g., human EL) and contains the VH CDR and VL CDR of the MEDI5884 antibody listed in Tables 3 and 4 determined by the method in MacCallum RM et al.

[0158] In some aspects, the CDRs of the antibody or its antigen-binding fragment correspond to an intermediate between the Kabat CDR and the Chothia structural loop and can be determined according to the AbM numbering scheme that refers to the AbM hypervariable regions used by the AbM antibody modeling software of Oxford Molecular (Oxford Molecular Group, Inc.). In some aspects of the present disclosure, provided herein is a method of administering an antibody or an antigen-binding fragment thereof that specifically binds to EL (e.g., human EL) and contains the VH CDR and VL CDR of the MEDI5884 antibody listed in Tables 3 and 4 determined by the AbM numbering scheme. In some aspects of the present disclosure, provided herein is a method of administering an antibody or an antigen-binding fragment thereof that specifically binds to EL (e.g., human EL) and contains the VH CDR and VL CDR of the MEDI5884 antibody listed in Tables 3 and 4 determined by the AbM numbering scheme.

[0159] In some aspects of the present disclosure, provided herein is a method of administering an antibody comprising a heavy chain and a light chain.

[0160] With respect to the heavy chain, in some aspects of the present disclosure, the heavy chain is a gamma heavy chain. The constant region of the human IgG4P heavy chain may contain the following amino acid sequence: ASTKGPSVFPLAPCSRSTSESTAALGCLVKDYFPEPVTVSWNSGALTSGVHTFPAVLQSSGLYSLSSVVTVPSSSLGTKTYTCNVDHKPSNTKVDKRVESKYGPPCPPCPAPEFLGGPSVFLFPPKPKDTLMISRTPEVTCVVVDVSQEDPEVQFNWYVDGVEVHNAKTKPREEQFNSTYRVVSVLTVLHQDWLNGKEYKCKVSNKGLPSSIEKTISKAKGQPREPQVYTLPPSQEEMTKNQVSLTCLVKGFYPSDIAVEWESNGQPENNYKTTPPVLDSDGSFFLYSRLTVDKSRWQEGNVFSCSVMHEALHNHYTQKSLSLSLGK (SEQ ID NO: 12).

[0161] In some aspects of the present disclosure, an antibody that immunospecifically binds to an EL (e.g., human EL) used in the methods described herein comprises a heavy chain whose amino acid sequence of the VH domain comprises the CDR amino acid sequences shown in Table 1, wherein the constant region of the heavy chain comprises a human gamma (γ) heavy chain constant region, e.g., the amino acid sequence of human IgG4P.

[0162] In some aspects of the present disclosure, an antibody that immunospecifically binds to an EL (e.g., human EL) used in the methods described herein comprises a heavy chain whose amino acid sequence of the VH domain comprises the amino acid sequences shown in Table 3, wherein the constant region of the heavy chain comprises a human gamma (γ) heavy chain constant region, e.g., the amino acid sequence of human IgG4P.

[0163] Regarding the light chain, in some aspects of the present disclosure, the light chain of the antibody described herein is a kappa light chain. The constant region of the human C kappa light chain may comprise the following amino acid sequence: RTVAAPSVFIFPPSDEQLKSGTASVVCLLNNFYPREAKVQWKVDNALQSGNSQESVTEQDSKDSTYSLSSTLTLSKADYEKHKVYACEVTHQGLSSPVTKSFNRGEC (SEQ ID NO: 13).

[0164] In some aspects of the present disclosure, an antibody that immunospecifically binds to an EL (e.g., human EL) used in the methods described herein comprises a light chain whose amino acid sequence of the VL domain comprises the CDR amino acid sequences shown in Table 2, wherein the constant region of the light chain comprises the amino acid sequence of the human C kappa light chain constant region.

[0165] In some aspects of the present disclosure, an antibody that immunospecifically binds to an EL (e.g., human EL) used in the methods described herein comprises a light chain whose amino acid sequence of the VL domain comprises the sequence shown in Table 4, wherein the constant region of the light chain comprises the amino acid sequence of the human C kappa light chain constant region.

[0166] In some aspects of the present disclosure, an antibody that immunospecifically binds to an EL (e.g., human EL) used in the methods described herein comprises a VH domain and a VL domain that comprise the amino acid sequences of any VH domain and VL domain described herein, wherein the constant region comprises the amino acid sequence of the constant region of an IgG (e.g., human IgG) immunoglobulin molecule. In some aspects of the present disclosure, an antibody that immunospecifically binds to an EL (e.g., human EL) used in the methods described herein comprises a VH domain and a VL do main that comprise the amino acid sequences of any VH domain and VL domain described herein, wherein the constant region comprises the amino acid sequence of the constant region of an IgG4P kappa (e.g., human IgG4P kappa) immunoglobulin molecule.

[0167] As shown herein, an antibody or antigen-binding fragment thereof that immunospecifically binds to an EL (e.g., human EL) used in the methods described herein can have reduced effector function, for example, compared to an antibody or antigen-binding fragment having a wild-type IgG1 sequence. The reduction in effector function can be due to, for example, the sequence of the constant region of the antibody or its antigen-binding fragment.

[0168] As shown herein, an antibody or antigen-binding fragment thereof that immunospecifically binds to an EL (e.g., human EL) used in the methods described herein may lack CDC activity and / or ADCC activity, for example, as a result of the sequence of the constant region.

[0169] In some aspects of the disclosure, an antibody or antigen-binding fragment thereof described herein that immunospecifically binds to an EL (e.g., human EL) comprises a heavy chain and a light chain, wherein (i) the heavy chain comprises a VH domain comprising the amino acid sequences of VH CDR1, VL CDR2, and VL CDR3 of the MEDI5884 antibody listed in Table 1, (ii) the light chain comprises a VL domain comprising the amino acid sequences of VL CDR1, VH CDR2, and VH CDR3 of the MEDI5884 antibody listed in Table 2, (iii) the heavy chain further comprises a constant heavy chain domain comprising the amino acid sequence of the constant domain of the human IgG4P heavy chain, and (iv) the light chain further comprises a constant light chain domain comprising the amino acid sequence of the constant domain of the human kappa light chain.

[0170] In some aspects of the disclosure, an antibody or antigen-binding fragment thereof described herein that immunospecifically binds to an EL (e.g., human EL) comprises a heavy chain and a light chain, wherein (i) the heavy chain comprises a VH domain comprising the amino acid sequence of the VH domain of the MEDI5884 antibody listed in Table 3, (ii) the light chain comprises a VL domain comprising the amino acid sequence of the VL domain of the MEDI5884 antibody listed in Table 4, (iii) the heavy chain further comprises a constant heavy chain domain comprising the amino acid sequence of the constant domain of the human IgG4P heavy chain, and (iv) the light chain further comprises a constant light chain domain comprising the amino acid sequence of the constant domain of the human kappa light chain.

[0171] In certain aspects, an antigen-binding fragment described herein that immunospecifically binds to an EL (e.g., human EL) is selected from the group consisting of Fab, Fab’, F(ab’) 2 , and scFv, wherein Fab, Fab’, F(ab’) 2, or scFv, comprises the heavy chain variable region sequence and the light chain variable region sequence of the anti-EL antibody or antigen-binding fragment thereof described herein. Fab, Fab’, F(ab’) 2 , or scFv, can be produced by any technique known to those skilled in the art. In some embodiments of the present disclosure, Fab, Fab’ 2 , or scFv, further comprises a moiety that extends the half-life of the antibody in vivo. This moiety is also referred to as a "half-life extension moiety". Any moiety known to those skilled in the art that extends the half-life of Fab, Fab’ 2 , or scFv in vivo can be used. For example, the half-life extension moiety can include an Fc region, a polymer, albumin, or an albumin-binding protein or albumin-binding compound. Examples of polymers include natural or synthetic, optionally substituted linear or branched polyalkylene, polyalkenylene, polyoxylalkylene, polysaccharide, polyethylene glycol, polypropylene glycol, polyvinyl alcohol, methoxypolyethylene glycol, lactose, amylose, dextran, glycogen, or derivatives thereof. Examples of substituents include one or more hydroxy groups, methyl groups, or methoxy groups. In some embodiments of the present disclosure, Fab, Fab’ 2 , or scFv can be modified by adding one or more C-terminal amino acids to attach a half-life extension moiety. In some embodiments of the present disclosure, the half-life extension moiety is polyethylene glycol or human serum albumin. In some embodiments of the present disclosure, Fab, Fab’ 2 , or scFv is fused to an Fc region.

[0172] 5.4 Pharmaceutical Compositions In this specification, provided is a method of administering a composition comprising an anti-EL antibody or an antigen-binding fragment thereof having a desired purity in a physiologically acceptable carrier, additive, or stabilizer (Remington's Pharmaceutical Sciences (1990) Mack Publishing Co. (Easton, Pennsylvania)). The acceptable carrier, additive, or stabilizer is non-toxic to the recipient at the dosages and concentrations used (see, for example, Gennaro, Remington: The Science and Practice of Pharmacy with Facts and Comparisons: Drugfacts Plus, 20th Edition (2003), Ansel et al., Pharmaceutical Dosage Forms and Drug Delivery Systems, 7th Edition, Lippencott Williams and Wilkins (2004), Kibbe et al., Handbook of Pharmaceutical Excipients, 3rd Edition, Pharmaceutical Press (2000)). The composition used for in vivo administration can be sterile. This can be readily achieved, for example, by filtration through a sterile filtration membrane.

[0173] In some aspects of the present disclosure, provided is a method of administering a pharmaceutical composition comprising: (i) an isolated antibody or an antigen-binding fragment thereof that specifically binds to human EL and comprises (a) the sequences of the heavy-chain variable region (VH) complementarity-determining regions (CDR) 1, VH CDR2, VH CDR3, and the light-chain variable region (VL) CDR1, CDR2, and CDR3 of SEQ ID NOs: 1 to 6, respectively, (b) a variable heavy-chain region comprising the amino acid sequence of SEQ ID NO: 7 and / or a variable light-chain region comprising the amino acid sequence of SEQ ID NO: 8, or (c) a heavy chain comprising the amino acid sequence of SEQ ID NO: 9 and / or a light chain comprising the amino acid sequence of SEQ ID NO: 10; and (ii) a pharmaceutically acceptable additive.

[0174] In some aspects of the present disclosure, a pharmaceutical composition comprising an isolated antibody or antigen-binding fragment thereof that specifically binds to human EL also comprises an inhibitor of PCSK9. In some aspects of the present disclosure, a pharmaceutical composition comprising an isolated antibody or antigen-binding fragment thereof that specifically binds to human EL is administered in combination with an inhibitor of PCSK9.

[0175] 5.5 Antibody Generation and Polynucleotides Antibodies and antigen-binding fragments thereof that immunospecifically bind to EL (e.g., human EL) can be generated by any method known in the art for the synthesis of antibodies and antigen-binding fragments thereof, e.g., by chemical synthesis or recombinant expression techniques. The methods described herein use conventional techniques in molecular biology, microbiology, genetic analysis, recombinant DNA, organic chemistry, biochemistry, PCR, oligonucleotide synthesis and modification, nucleic acid hybridization, and related fields within the skill of the art, unless otherwise indicated. These techniques are described, for example, in the references cited herein and are well explained in the literature. For example, Sambrook J et al., (2001) Molecular Cloning: A Laboratory Manual, Cold Spring Harbor Laboratory Press, Cold Spring Harbor, NY, Ausubel FM et al., Current Protocols in Molecular See Biology, John Wiley & Sons (1987 and annual updates), Current Protocols in Immunology, John Wiley & Sons (1987 and annual updates), Gait (ed.) (1984) Oligonucleotide Synthesis: A Practical Approach, IRL Press, Eckstein (ed.) (1991) Oligonucleotides and Analogues: A Practical Approach, IRL Press, Birren B et al., (ed.) (1999) Genome Analysis: A Laboratory Manual, Cold Spring Harbor Laboratory Press.

[0176] In some embodiments, provided herein is a method of administering an anti-EL antibody or antigen-binding fragment thereof, or a pharmaceutical composition comprising such an antibody or fragment, wherein the antibody or fragment is produced by recombinant expression of a polynucleotide comprising a nucleotide sequence in a host cell.

[0177] In some embodiments, the anti-EL antibody or antigen-binding fragment administered according to the methods provided herein is encoded by a polynucleotide encoding an anti-EL antibody or antigen-binding fragment or domain thereof that has been optimized, for example, by codon / RNA optimization, replacement with a heterologous signal sequence, and removal of mRNA destabilizing elements. Methods for generating a nucleic acid encoding an anti-EL antibody or antigen-binding fragment or domain thereof (e.g., a heavy chain, a light chain, a VH domain, or a VL domain) optimized for recombinant expression by introducing codon changes (e.g., codon changes that encode the same amino acid due to the degeneracy of the genetic code) and / or removing inhibitory regions within the mRNA can be carried out, for example, by adapting the optimization methods described in U.S. Patent No. 5,965,726, U.S. Patent No. 6,174,666, U.S. Patent No. 6,291,664, U.S. Patent No. 6,414,132, and U.S. Patent No. 6,794,498, as appropriate.

[0178] The polynucleotide can be, for example, in the form of RNA or DNA. DNA includes cDNA, genomic DNA, and synthetic DNA. The DNA can be double-stranded or single-stranded. In the case of single-stranded DNA, it can be a coding strand or a non-coding (antisense) strand. In some embodiments of the present disclosure, the polynucleotide is cDNA or DNA lacking one or more introns. In some embodiments of the present disclosure, the polynucleotide is a non-naturally occurring polynucleotide. In some embodiments of the present disclosure, the polynucleotide is recombinantly produced. In some embodiments of the present disclosure, the polynucleotide is isolated. In some embodiments of the present disclosure, the polynucleotide is substantially pure. In some embodiments of the present disclosure, the polynucleotide is purified from natural components.

[0179] In some embodiments, a vector (e.g., an expression vector) comprises a nucleotide sequence encoding an anti-EL antibody and its antigen-binding fragment or domain thereof for recombinant expression in a host cell, preferably a mammalian cell. In some embodiments, a cell, e.g., a host cell, comprises such a vector that recombinantly expresses an anti-EL antibody or its antigen-binding fragment (e.g., a human antibody or a humanized antibody or its antigen-binding fragment) described herein. Thus, a method for generating an antibody or its antigen-binding fragment described herein may include expressing such an antibody or its antigen-binding fragment in a host cell.

[0180] An expression vector can be introduced into a cell (e.g., a host cell) by conventional techniques and then the resulting cell can be cultured by conventional techniques to generate an antibody or its antigen-binding fragment (e.g., the 6 CDRs of MEDI5884, VH, VL, VH and VL, heavy chain, light chain, or an antibody or its antigen-binding fragment comprising a heavy chain and a light chain) or its domain (e.g., VH, VL, VH and VL, heavy chain, or light chain of MEDI5884) described herein.

[0181] In some embodiments of the present disclosure, an anti-EL antibody or its antigen-binding fragment (e.g., an antibody or its antigen-binding fragment comprising the CDRs of MEDI5884) is administered according to the methods provided herein and is generated in a host cell. In some embodiments of the present disclosure, the host cell is a CHO cell.

[0182] In some aspects of the present disclosure, the antibodies or antigen-binding fragments thereof administered according to the methods provided herein are isolated or purified. Generally, an isolated antibody or antigen-binding fragment thereof is substantially free of other antibodies or antigen-binding fragments thereof having different antigen specificities. For example, in some aspects of the present disclosure, preparations of the antibodies or antigen-binding fragments thereof described herein are substantially free of cellular material and / or chemical precursors.

[0183] The following examples are provided by way of illustration and not by way of limitation.

Examples

[0184] 6. Examples The examples in this section (i.e., Section 6) are provided by way of illustration and not by way of limitation.

[0185] 6.1 Example 1: Nonclinical Pharmacology of MEDI5884 From nonclinical in vivo pharmacology studies, it was revealed that administration of single subcutaneous (SC) doses of MEDI5884 (0.5 mg / kg, 6 mg / kg, or 30 mg / kg) in normal male cynomolgus monkeys increased plasma HDL-C in a dose-dependent manner. The increases in HDL-C from baseline to maximum effect for the 0.5 mg / kg, 6 mg / kg, and 30 mg / kg doses were 63 ± 14 mg / dL to 96 ± 29 mg / dL, 60 ± 3.8 mg / dL to 111 ± 5.6 mg / dL, and 54 ± 7.4 mg / dL to 122 ± 17 mg / dL, respectively. Smaller increases were also observed in total cholesterol (TC), low-density lipoprotein cholesterol (LDL-C), and non-HDL-C (TC minus HDL-C).

[0186] Apolipoprotein A1 (ApoA1), the major lipoprotein component of HDL, also increased by 75 ± 5.5% at a dose of 30 mg / kg. Since a dose-dependent increase in serum phospholipids was observed, this indicates that MEDI5884 inhibited the hydrolysis of phospholipids contained in HDL particles.

[0187] Increases in ATP-binding cassette transporter A1 (ABCA1), cholesterol efflux capacity (CEC), overall cholesterol efflux, and the total number of HDL particles were observed at the highest dose (30 mg / kg). Increases in the average number of total low-density lipoprotein (LDL) particles and large low-density lipoprotein (LDL) particles were also observed. Cholesterol efflux was measured using HDL obtained from animals in the MEDI5884 treatment groups at 0, 0.5, 1, 2, 3, 7, and 14 days after dosing at 0.5 mg / kg and 30 mg / kg. ABCA1 efflux temporarily decreased below baseline (days 0.5 and 1) after treatment with 30 mg / kg of MEDI5884 but exceeded the baseline level by 6%, 26%, and 114% on days 3, 7, and 14, respectively. Similarly, overall efflux temporarily decreased below baseline (-6%) on day 0.5 and increased to 6%, 5%, 41%, and 53% on days 2, 3, 7, and 14, respectively.

[0188] HDL particles were characterized using NMR spectroscopy in samples obtained from cynomolgus monkeys treated with MEDI5884. Treatment with 30 mg / kg of MEDI5884 increased the total number of HDL particles by up to 49 ± 6.3%, and the number of both small and large particles increased by up to 182 ± 85% and 104 ± 10%, respectively. Medium-sized HDL particles decreased by up to 48 ± 14% on day 2 and returned to baseline by day 49, suggesting speciation or interconversion of HDL particles when a new equilibrium is established. It increased. For the HDL size, from the baseline to the plateau level for the doses of 0.5 mg / kg, 6 mg / kg, and 30 mg / kg, it increased from 10.0 ± 0.033 nm to 10.9 ± 0.17 nm, from 10 ± 0.23 nm to 10.7 ± 0.12 nm, and from 9.9 ± 0.033 nm to 10.7 ± 0.13 nm, respectively.

[0189] These data support, for example, the use of MEDI5884 in human patients for the prevention of secondary cardiovascular events.

[0190] 6.2 Example 2: Non-Clinical Pharmacokinetics and Safety of MEDI5884 MEDI5884 was administered to cynomolgus monkeys in a single-dose non-Good Laboratory Practice (GLP) PK / pharmacodynamics (PD) study and a two-repeat-dose GLP toxicity study. The SC administration of MEDI5884 at a single-dose of up to 30 mg / kg was well tolerated. No unexpected deaths, adverse clinical findings, injection site reactions, or adverse effects on body weight were seen. After repeated SC administration of MEDI5884 at 10 mg / kg, 30 mg / kg, or 100 mg / kg per dose for 1 month or 6 months (administered once every 2 weeks), all animals survived until the scheduled sacrifice. No MEDI5884-related changes were observed in clinical observations, ophthalmic evaluations, body weight, behavior, neurophysiology, respiratory rate, injection site irritation scoring, heart rate, electrocardiogram (ECG), or blood pressure. Treatment-related findings were limited to minimal to moderate pharmacologically mediated increases in TC, HDL-C, LDL-C, and phospholipids at all dose levels. By microscopy, the presence of minimal to moderate perivascular lymphocyte / mixed leukocyte infiltration was observed at the SC injection sites of a few MEDI5884-treated animals, and no treatment-related histopathological findings were observed in other tissues. Based on these results, the no-observed-adverse-effect level (NOAEL) was determined to be 100 mg / kg per dose.

[0191] 6.3 Example 3: Phase 1 Clinical Evaluation of MEDI5884 MEDI5884 was evaluated in a Phase 1, first-in-human, blinded, placebo-controlled, single ascending dose (SAD) study to assess the safety, PK, and PD of subcutaneously (SC) administered MEDI5884 in healthy subjects not on statin therapy. Subjects were randomized in a 3:1 ratio to receive 30 mg, 100 mg, 300 mg, or 600 mg of MEDI5884 or placebo. The initial cohorts were 6 MEDI5884 subjects and 2 placebo subjects per dose level. These cohorts were then replicated using subjects of Japanese ancestry to provide data to support the conduct of clinical studies in Japan. A total of 64 subjects were enrolled at one facility in the United States (US). The follow-up period varied by cohort from 28 days to 90 days post-dose.

[0192] Subjects received a single injection of MEDI5884 (or placebo) at the 30 mg and 100 mg doses, three SC injections (100 mg each) at the 300 mg dose, or six SC injections (100 mg each) at the 600 mg dose.

[0193] Subjects The median (SD) age of enrolled subjects was 35.9 (9) years, 93.9% were male, 53.1% were Asian, 28.1% were White, 14.1% were Black or African American, and 4.7% reported multiple ethnicities, and 90.6% were not of Hispanic ethnicity.

[0194] Pharmacokinetics After single SC administration of MEDI5884 at doses of 30 mg, 100 mg, 300 mg, and 600 mg, MEDI5884 exhibited non-linear PK, probably due to target-mediated drug disposition, with Cmax and AUC values exceeding dose proportionality. PK parameters are summarized in Table 7.

[0195]

Table 7

[0196] The mean PK profiles observed between Japanese Americans and the general US population (Caucasian cohort) almost overlapped. Higher exposure was observed in Japanese American subjects at a dose of 300 mg, which was probably due to the difference in body weight between Japanese American and general US populations. A moderate effect of body weight on the PK profile was observed in both Caucasian and Japanese American subjects. The CL / F at a dose of 600 mg was 0.378 L / day and 0.254 L / day in the general US population (Caucasian cohort) and Japanese American population, respectively. The data suggest that there are no substantial ethnic differences in the PK of MEDI5884.

[0197] Safety and immunogenicity Safety data were evaluated in 64 subjects including 48 subjects administered MEDI5884 (4 dose cohorts of 12 subjects each [30 mg, 100 mg, 300 mg, or 600 mg]; 24 had Japanese ancestry) and 16 subjects administered placebo (4 dose cohorts of 4 subjects each; 8 had Japanese ancestry). No treatment-emergent adverse events (TEAEs), treatment-emergent serious adverse events (TESAEs), or deaths leading to withdrawal from the study were seen. TEAEs occurred at similar rates in subjects administered MEDI5884 (16 / 48, 33.3%) or placebo (5 / 16, 31.3%).

[0198] There were no positive ADA results in subjects in the general US population, but a positive anti-drug antibody (ADA) response was detected in 6 out of 32 Japanese American subjects, including 1 placebo recipient. No AE was reported to be associated with ADA. In some Japanese American subjects with positive ADA, lower exposure was observed compared to other Japanese American subjects without ADA. The exposure in these ADA-positive subjects was pooled in the Within the range of exposure of the subjects at the same dose within the group, no effect on PD or safety was observed. In summary, ADA was rare, not associated with the effects on adverse events (AE) or PD, and had no clinically relevant effect on PK.

[0199] Pharmacodynamics The overall baseline lipid levels were similar between placebo subjects and those treated with MEDI5884.

[0200] In these healthy subjects not receiving statin therapy, a significant increase in HDL-C was observed after administration of MEDI5884. The mean (SD) percent change from baseline in HDL-C on day 28 was 4.1% (17.4), 42.0% (26.9), 39.7% (22.5), and 49.8% (17.3) in subjects receiving MEDI5884 at 30 mg, 100 mg, 300 mg, or 600 mg, respectively, compared with 15.9% (16.8) in the placebo case.

[0201] An increase in apoA1 was also observed. In the pooled population, small increases in LDL-C and apoB were observed. These increases did not appear to be dose-related and mainly occurred with a delay after administration. No clear effect on triglyceride levels was observed.

[0202] 6.4 Example 4: Phase 2a Clinical Evaluation of MEDI5884 MEDI5884 was also evaluated in a phase 2a, randomized, double-blind, placebo-controlled, parallel-design study to assess the safety, PK, PD, and immunogenicity of MEDI5884 in subjects with stable coronary heart disease (CHD) who were receiving concomitant high-intensity statin therapy and had triglyceride levels of 500 mg / dL or less and LDL-C of 100 mg / dL or less.

[0203] A total of 132 subjects received 3 monthly SC administrations of placebo or MEDI5884 at doses of 50 mg, 100 mg, 200 mg, 350 mg, and 500 mg.

[0204] Subject This study mainly enrolled male subjects (87.0%) of white race (90.8%), and the median age at enrollment was 67 years. The overall baseline characteristics were similar between the placebo subjects and the subjects treated with MEDI5884. The subjects who received a total of three doses of the investigational drug were as follows: 22 out of 23 (95.7%) in the placebo group, and 18 out of 20 (90%), 22 out of 24 (91.7%), 20 out of 22 (90.9%), 20 out of 21 (95.2%), and 21 out of 22 (95.5%) in the MEDI5584 50mg group, MEDI5584 100mg group, MEDI5584 200mg group, MEDI5584 350mg group, and MEDI5584 500mg group, respectively. One subject in the MEDI5884 50mg group was included as having completed treatment even though he had only received two doses because he missed the visit for dose 2.

[0205] Pharmacokinetics An interim PK analysis was performed based on the data up to day 111. The mean MEDI5884 concentration-time profiles after three monthly SC administrations of MEDI5884 are shown in Figure 1A for each dose cohort. The PK parameters based on non-compartmental analysis are summarized in Table 8.

[0206]

Table 8

[0207] MEDI5884 showed non-linear PK, probably due to target-mediated drug disposition, but linear-range PK was observed 30 days after dosing at the MEDI5884 doses of 350mg and 500mg. Cmax and AUC were approximately supra-dose proportionality. Large inter-subject variability and slight drug accumulation were observed. The mean estimated CL / F at 500mg was 0.389 L / day.

[0208] Safety and Immunogenicity Death or related TESAE was not seen, and AE was almost balanced between the MEDI5884 treatment group (59 out of 109 [54.1%]) and the placebo group (17 out of 23 [73.9%]), independent of the dose of MEDI5884, and represented events predicted to occur in the enrolled population. Injection site reactions by self-report occurred in 15 out of 109 (14%) subjects treated with MEDI5884 and 3 out of 23 (13%) subjects treated with placebo. Injection site reactions reported by the study responsible physicians occurred in 9 out of 109 (8%) subjects treated with MEDI5884 and 3 out of 23 (13%) subjects treated with placebo. These reactions were mild to moderate in severity. Eight subjects discontinued dosing (1 placebo recipient and 1 MEDI5884 recipient withdrew consent for dosing, and the other 6 withdrew for laboratory observations mandated by the protocol [some were recorded as AE, 4 had elevated apoB recorded, and 2 had elevated triglycerides recorded]).

[0209] ADA was rare, had low titers, was not associated with AE, and had no effect on PK. ADA was similar in placebo recipients and MEDI5884 recipients. Therefore, the observed ADA may represent false positives.

[0210] Pharmacodynamic effects Overall baseline lipid levels were similar between placebo subjects and those treated with MEDI5884.

[0211] Target engagement of MEDI5884, which is the suppression of EL, was shown to be dose-dependent (Figure 1B). In particular, the amount of hEL bound by MEDI5884 in human plasma was measured using a Meso Scale Diagnostics (MSD)-based immunoassay platform It was measured. Briefly, the wells on a 96-well plate were coated with MEDI5884 and incubated overnight at 4°C. The next day, the wells were washed with PBS containing 0.05% Tween20 washing buffer and blocked with I-Block buffer (Applied Biosystems) for 1 hour at room temperature. The plate was washed. Then, recombinant EL protein standard (Origene Technologies) and human plasma samples were added to the corresponding wells and incubated for 1 hour at room temperature. After washing, biotinylated EL detection antibody (Origene Technologies) was added to the corresponding wells and incubated for 1 hour at room temperature. After washing the plate, streptavidin-sulfo-TAG antibody (MSD) was added to the corresponding wells and incubated for 1 hour at room temperature. After washing, the wells were incubated with the read buffer (MSD). The plate was read using a MESO Sector S 600 plate reader, and the data were analyzed using the MSD Discovery Workbench software analysis program.

[0212] In these patients, a dose-dependent increase in HDL-C from baseline was observed in the MEDI5884 treatment group. The mean (SD) percent change from baseline in HDL-C on day 91 (using the last observation carried forward (LOCF) imputation method) was 2.88% (14.86), 21.82% (30.66), 34.41% (34.89), 43.29% (31.09), and 48.31% (25.63) in subjects administered MEDI5884 at 50 mg, 100 mg, 200 mg, 350 mg, or 500 mg, respectively, compared to -3.01% (13.60) in the placebo case. Figure 2 and Tables 9A and 9B show the observed (non-LOCF) percent change from baseline in HDL-C over a 90-day period and the observed (non-LOCF) change from baseline in HDL-C on day 91.

[0213] [Table 9A]

[0214]

Table 9B

[0215] A dose-dependent increase from baseline in HDL particle number and HDL particle size (Table 10) was also observed in the MEDI5884 treatment group relative to the placebo group.

[0216]

Table 10

[0217] A dose-dependent increase from baseline in apoA1 (Figure 3 and Tables 11A and 11B) and high-density lipoprotein phospholipid (HDL-PL) (Figure 4 and Table 12) was also observed in the MEDI5884 treatment group relative to the placebo group. In particular, Figure 3 and Tables 11A and 11B show the observed (non-LOCF) percent change from baseline in ApoA1 over a 90-day period, and the observed (non-LOCF) change from baseline in ApoA1 on Day 91. The mean (SD) percent change from baseline in ApoA1 on Day 91 (using the LOCF imputation method) was 1.32% (14.81), 15.88% (19.66), 24.82% (21.92), 36.26% (27.36), and 36.85% (18.03) in subjects administered MEDI5884 at 50 mg, 100 mg, 200 mg, 350 mg, or 500 mg, respectively, versus 1.42% (11.20) for placebo.

[0218]

Table 11A

[0219]

Table 11B

[0220]

Table 12

[0221] A dose-dependent increase from baseline in ABCA1-mediated and total cholesterol efflux was also observed in the MEDI5884 treatment group compared to the placebo group. The effect of MEDI5884 on non-ABCA1 cholesterol efflux is shown in Figure 5.

[0222] In the MEDI5884 treatment group, a slight increase in triglycerides, LDL-C, and apoB was also observed. In the lower-dose groups, no clear dose-dependent relationship was seen in the increase in triglycerides, LDL-C, and apoB in the MEDI5884 treatment group. The change in apoB was significant only at the 500 mg dose. Table 13 shows the apoB levels by dose observed on Day 91.

[0223]

Table 13

[0224] In three subjects treated with MEDI5884, triglycerides increased to near or above 1000 mg / dL. These subjects had additional risk factors for hypertriglyceridemia. A dose-dependent effect of MEDI5884 on triglyceride levels was not observed.

[0225] At a dose of 200 mg of MEDI5884, on Day 91, compared to placebo (-1.6 [-3.0] mg / dL for HDL-C, -2.6 [-1.0] mg / dL for LDL-C, and -0.5 [0.0] mg / dL for apoB), mean (median) changes from baseline of 15.2 (11.0) mg / dL in HDL-C, 6.1 (5.0) mg / dL in LDL-C (direct), and 2 (-1.0) mg / dL in apoB were induced.

[0226] The estimated parameters of the main PK / PD models are summarized in Table 14.

[0227]

Table 14

[0228] 6.5 Example 5: Phase 2B Clinical Evaluation of MEDI5884 The data analysis discussed above was performed, and a monthly dose of 250 mg of SC MEDI5884 was selected for further evaluation.

[0229] More specifically, by examining the data in detail, it was revealed that administration of five doses of MEDI5884 showed a clear dose-dependent increase in exposure in patients (Figure 1A), which led to dose-dependent target engagement (i.e., inhibition of EL levels) (Figure 1B). The EL level at a 200 mg dose was not consistently inhibited over approximately 30 days, whereas the EL level at a 350 mg dose maintained complete inhibition during the 30-day dosing interval. Considering the monthly dosing interval, the optimal dose seemed to be between 200 mg and 35 0 mg of the studied doses.

[0230] Biomarkers such as HDL-C, ApoA1, and HDL-PL increased in a dose-dependent manner after inhibiting EL activity (Figures 2 - 4). The time course of the biomarkers indicated that the effective dose should exceed 200 mg. Safety biomarkers within the pathway such as LDL-C, ApoB, and TG increased with a less distinct dose-dependence compared to the efficacy biomarkers. Since the increase in ApoB and TG at the 500 mg dose was identified as a concern, this indicates that a dose less than the 500 mg dose should be selected as the optimal dose.

[0231] The multiple comparison procedure modeling (MCP Mod) method was applied to evaluate the relationship between the area under the effect curve (AUEC) of HDL-C, ApoA1, HDL-PL, LDL, ApoB, and TG with respect to the dose during the period from day 60 to day 90. The desired biomarkers of HDL-C level, ApoA1 level, and HDL-PL level reached a maximum with increasing dose (Figures 2-4), enabling the estimation of the dose (ED 90 ) that achieves 90% of the maximum biomarker level. The estimated ED 90 was 205 mg, 270 mg, and 265 mg, respectively (Figure 6). The undesired biomarkers of LDL, ApoB, and TG did not reach a plateau but showed the following trends: (1) the LDL level continued to increase within the range of 50 mg to 500 mg, (2) the ApoB level was independent of the dose except at 500 mg, and (3) the TG value remained constant regardless of the administered dose. Thus, the results from the MCP Mod approach support that monthly administration of 250 mg has a high likelihood of achieving 90% of the maximum efficacy of the desired biomarkers without causing high levels of undesired biomarkers.

[0232] A mathematical model was developed to describe the pharmacokinetics (PK) of MEDI5884 and the biomarker profiles of HDL and ApoA1 after administration of MEDI5884. The PK model part used a two-compartment PK model with parallel linear and non-linear elimination pathways, while the PD model part for biomarker modeling followed a typical indirect response model involving inhibition of the elimination pathway of each biomarker that brought about an increase in biomarker level after administration (Figure 7). This model does not include HDL-P, but changes in HDL-P are indirectly addressed by the evaluation of HDL-C and ApoA1. An increase in HDL-C without an increase in ApoA1 means larger HDL particles but not more particle numbers.

[0233] When analyzing PK data simultaneously from a wide dosage range (50 mg to 500 mg), MEDI5884 showed non-linear PK due to target-mediated drug disappearance that is likely to saturate at low dosages. Therefore, for the simulation of the PK profile at 250 mg, which is an interpolation between the PK profile observed after a 200 mg dose and the PK profile observed after a 350 mg dose, it is reasonable to assume that PK is linear at high dosages.

[0234] The observed PK / PD data from the Phase 2a study of MEDI5884 in subjects with CHD receiving high-intensity statin therapy were well characterized by this model. Although large inter-subject variability was observed in PK, HDL, and ApoA1 after dosing, a clear relationship was seen between MEDI5884 exposure and the corresponding increases in HDL-C and ApoA1. Using the model-estimated 50% inhibitory concentration (IC 50 ) values, IC 90 was calculated (i.e., 3.03 μg / mL and 3.43 μg / mL for HDL-C and ApoA1, respectively). The estimated median trough concentration after QM at 250 mg was approximately 3.46 μg / mL based on the following simulation, which is close to the target exposure of IC 90 for HDL-C and ApoA1. The estimated time courses of MEDI5884, HDL, and ApoA1 after administration of a monthly dose of 250 mg of MEDI5884 are shown in Figure 8.

[0235] In summary, these data suggest that a monthly dose of 250 mg of MEDI5884 shows: (i) linear PK and target engagement (EL inhibition) over 30 days after dosing, (ii) median trough levels exceeding the IC90 for maximum increases in HDL-C and ApoA1 based on PK / PD modeling, and (iii) minimal unwanted increases in LDL-C and apoB based on a multiple comparison procedure modeling approach.

[0236] Therefore, these data support the evaluation of MEDI5884 in a Phase 2b, randomized, double-blind, placebo-controlled study in adults with a history of myocardial infarction (MI) receiving high-intensity statin therapy. In these studies, MEDI5884 (250 mg) or placebo was administered once monthly for 24 months at a dose of 250 mg to demonstrate that MEDI5884 can reduce the rates of cardiovascular death, MI, stroke, and coronary artery revascularization.

[0237] 6.6 Example 6: Inhibition of EL and PCSK9 in Cynomolgus Monkeys A combined pharmacological study was conducted to evaluate the effect of inhibition of proprotein convertase subtilisin / kexin type 9 (PCSK9) on lipoprotein metabolism after MEDI5884 treatment. The study protocol is shown in Figure 9. To better mimic a virtual patient population already on LDL-C lowering medication, healthy cynomolgus monkeys were first treated for 4 weeks starting on day 0 by weekly subcutaneous injection of a PCSK9 neutralizing monoclonal antibody (mAb) (10 mg / kg, n = 8) or vehicle to establish a low LDL-C baseline. The PCSK9 monoclonal antibody used was HS9 (containing the VH and VL sequences of SEQ ID NO: 14 and SEQ ID NO: 15, respectively). The HS9 antibody (in the context of the GLP-1 fusion protein called MEDI4166) is disclosed in Chodorge et al., Sci. Rep. 8: 17545 (2018) and PCT International Publication No. WO 2015 / 127273 (each incorporated herein by reference in its entirety).

[0238] Subsequently, four animals from each group were administered subcutaneous doses of MEDI5884 (10 mg / kg, n = 4) or vehicle (n = 4) on day 28 (vertical dashed lines in Figures 10 and 11) and day 42. LDL-C, HDL-C, ApoB, and ApoA1 were measured in plasma samples collected at the indicated time points. Total extraction and ABCA1 extraction were also evaluated.

[0239] An increase in LDL-C was observed after EL neutralization. Compared to vehicle-treated animals, PCSK9 inhibition reduced LDL-C by 75% (Figure 10, left graph) and maintained the level of that reduction during the 4-week induction period, but had no apparent effect on HDL-C (Figure 10, right graph). In vehicle-treated animals during the induction period, MEDI5884 caused an increase in both HDL-C and LDL-C. When added in addition to the PCSK9 inhibitor during the second 4-week period, MEDI5884 maintained the ability to increase HDL-C to a similar extent, but the magnitude of the LDL-C increase was significantly blunted, indicating that LDL particles continued to be taken up by LDL receptors (Figure 10). The patterns of LDL-C and HDL-C were comparable to the respective changes in ApoB and ApoA1 (Figure 11). The effects on total efflux and ABCA1 efflux are shown in Figure 12. In particular, the increase in efflux observed in relation to the administration of MEDI5884 (including the administration of MEDI5884 + HS9) generally reflected the observed increase in HDL-C.

[0240] These results demonstrate evidence of cholesterol uptake by LDL receptors and indicate that the increase in LDL-C observed with MEDI5884 treatment in monkeys can be mitigated by a mechanism that upregulates LDL receptors. These results further indicate that MEDI5884 can be administered in combination with a PCSK9 inhibitor. This combination therapy can utilize the combined action of two complementary mechanisms that both target different aspects of cholesterol reverse transport.

[0241] 6.7 Example 7: Effect of escalating doses of MEDI5884 on plasma phosphatidylinositol (PI) levels in subjects with stable coronary artery disease The effect of MEDI5884 on plasma phosphatidylinositol (PI) levels in patients with stable coronary artery disease was quantified. For quantification, a high-throughput multiplex method combining hydrophilic interaction chromatography (HILIC) separation and multiple reaction monitoring (MRM) in negative mode was used. A total of 31 endogenous PI species were monitored.

[0242] Reagents All lipid standards were purchased from Avanti Polar Lipids (Alabaster, AL). Three PI standards were used in this study: PI(12:0 / 13:0) was used as the internal standard (IS), and PI(17:0 / 14:1) and PI(21:0 / 22:6) were used as surrogate analytes. HPLC grade water was purchased from Honeywell (Charlotte, NC). HPLC grade isopropanol (IPA), acetonitrile (ACN), and ammonia were all purchased from Sigma-Aldrich (St. Louis, MO). Ammonium acetate and bovine serum albumin (BSA) were purchased from MilliporeSigma (Burlington, MA). PBS was purchased from Lonza BioWhittaker (Morristown, NJ). Special glass-coated 96-well extraction plates and glass vials with PTFE (polytetrafluoroethylene) liners were obtained from Thermo-Fisher (Waltham, MA). Human plasma (pooled plasma and individual plasma) was purchased from BioIVT (Westbury, NY).

[0243] Extraction procedure for plasma samples The internal standard (IS) was spiked into isopropanol (IPA) at 60 nM (final concentration) to prepare an IS-IPA solution. One pooled lot of human plasma was used as quality control (QC), and two levels of low QC (LQC) (8-fold diluted with 40 mg / mL bovine serum albumin (BSA) in PBS) and high QC (HQC) (undiluted plasma) were prepared. All test samples were diluted 8-fold with 40 mg / mL BSA. 8-fold dilutions of both HQC samples and test samples were prepared using an Agilent Bravo automated liquid handling system (Santa Clara, CA) equipped with a Series III 96 LT disposable tip head, an automated liquid handling platform. According to a pre-specified plate map, 20 μL of LQC, HQC, or test sample was transferred to a separate extraction plate and then precipitated with 180 μL of IS-IPA using the above-mentioned automated device. The samples were shaken vigorously on an IKA MTS 2 / 4 digital microtiter shaker (Wilmington, NC) for approximately 10 minutes (900 rpm to 1200 rpm).

[0244] The samples were then centrifuged at 2500 g for 5 minutes. Automated liquid handling was used again to transfer 20 μL of plasma extracted with IS-IPA to 140 μL of reconstitution solution (90.25% (v / v) acetonitrile (ACN), 9.75% (v / v) water, 10 mM ammonium acetate). The samples were then shaken at 600 rpm for 5 minutes.

[0245] Hydrophilic interaction chromatography separation Separation by chromatography was carried out on a Nexera X2 UHPLC system (Shimadzu Corporation, Kyoto, Japan) with an Acquity ultra-performance liquid chromatography (UPLC) BEH (ethylene-bridged hybrid) hydrophilic interaction chromatography (HILIC) column (130 Å, 1.7 μm, 2.1 mm × 100 mm, Waters, Milford, Massachusetts). The mobile phases used were mobile phase A (MPA) (5% water, 95% ACN, v / v) and mobile phase B (MPB) (50% water, 50% ACN, v / v), both containing 10 mM ammonium acetate (pH 8.0 - 8.5). For each sample or QC, 10 μL of the reconstituted IPA extract was injected onto the column. Separation was carried out at 37 °C at a flow rate of 0.5 mL / min. The separation gradient was 5% → 13% MPA over 4 minutes, followed by a 2-minute wash period and a 4-minute equilibration period.

[0246] Mass spectrometry Detection by mass spectrometry was achieved using a 6500+ quadrupole ion trap (QTRAP) mass spectrometer (Sciex, Framingham, Massachusetts) operating in negative electrospray ionization (ESI) multiple reaction monitoring (MRM) mode. The MS conditions were adjusted and the retention times were defined using the synthetic reference standard PI species listed above. The most intense signal intensity structurally characteristic MRM transitions in negative ESI mode that enabled the identification of the two acyl chains were selected during the adjustment of the synthetic reference standards (surrogate analyte and IS). Subsequently, structurally similar MRMs that enabled the identification of the two acyl chains of each endogenous PI species were predicted using LipidView (Sciex, Redwood Shores, California) and are shown in Table 15. After adjusting the synthetic reference standards, the same source parameters were used for all PI species. Details regarding the acquisition method can be found in Table 16.

[0247]

Table 15

[0248] [Table 16]

[0249] Data collection, analysis and reporting After acquisition, samples from each acquisition batch were analyzed using the MultiQuant software. The samples were analyzed according to a specified quantification method (.qmethod). Details of the quantification method can be found in Table 17 for component and outlier settings. Integration and regression settings were optimized for each individual batch based on the peaks of interest generated. However, identical integration and regression parameters were applied to all samples within the same batch. Peak areas of the internal standard and peak areas of endogenous PI species were calculated for QC and unknown samples within a batch. Peak area ratios of endogenous PI species were calculated based on peak area integration in MultiQuant. MultiQuant quantification result files (.qsession) were then exported to .txt files and further data analysis was performed using Excel (Microsoft Office 2016) and Spotfire (TIBCO™ Spotfire™ Analyst 7.9.2 HF-011 build version 7.9.2.0.12). To assess the linearity of the instrument response and measurement precision for each endogenous PI species, the following were evaluated: HQC / LQC ratio, % difference of HQC / LQC ratio from the nominal value, and %CV for HQC and LQC. Most species had %CV values ​​of 30% or less for HQC and LQC values, and the % difference of HQC / LQC ratio from the nominal value was within 70%-130%.

[0250] [Table 17] TIFF0007692411000022.tif238170

[0251] result A total of 978 plasma samples from subjects with stable coronary artery disease were tested as described above. The samples were analyzed in a total of 15 assays. For all PI species, all 15 assays met the acceptance criteria, except for PI(16:0 / 16:0) which had consistently unacceptable variability and HQC / LQC ratios. There were no assays considered invalid.

[0252] Clinical samples were also obtained and analyzed from healthy volunteers administered MEDI5884. Some variability was observed between batches. To correct for batch - to - batch variability and explore the possibility of bridging data between the clinical study on CAD patients and the clinical study on healthy volunteers, the SERRF normalization algorithm (Fan S., et al. Anal Chem Mar 5;91 5:3590 - 6 (2019)) was evaluated and found to perform better than other normalization methods evaluated for this dataset. According to the results shown in FIGS. 13A - 13C, FIGS. 14A - 14C, FIG. 15, and FIG. 16, it was demonstrated that PI levels were significantly higher in healthy subjects across all PI species compared to MEDI5884 - untreated CAD patients. This difference was determined to be statistically significant by a two - sided unequal - variance t - test (p < 0.0005 for 12 PI species). In the placebo arms of both clinical studies, PI levels remained relatively stable over a period of several months, and lower levels were maintained in CAD patients compared to healthy volunteers for the 12 PI species examined.

[0253] Levels of various PI species over time relative to the day of visit in healthy patients and patients with CAD are shown in FIGS. 13A - 13C and FIGS. 14A - 14C. A comparison of the median PI species levels in healthy volunteers and CAD patients on day 21 is shown in FIG. 15, and a comparison of the median PI species levels over the full day is shown in FIG. 16. As shown in FIGS. 13A - 13C, FIGS. 14A - 14C, FIG. 15, and FIG. 16, after three monthly subcutaneous (SC) administrations of MEDI5884, most plasma PI species increased in a dose - dependent manner relative to placebo. The duration of the increase in plasma PI appeared to correlate with MEDI5884 exposure. For most PI species, the increase in PI levels reached saturation at the 350 mg dose level of MEDI5884, and no further increase in PI relative to baseline occurred at the 500 mg dose level of MEDI5884. However, at the 200 mg dose level of MEDI5884, on day 91, the increase in PI approached the saturation levels observed in the higher - dose cohorts. The percent change from baseline for each PI species varied from approximately 1000% for the less abundant species to approximately 100% - 200% for the more abundant species for both CAD subjects and healthy volunteer subjects (see FIGS. 17A - 17E, FIG. 18, and FIGS. 19A - 19E; Table 18, which summarizes data obtained for all PI species at the indicated doses and days of visit, and Tables 19A - 19D, which show data obtained for individual PI species). The average change across all PI species for CAD patients reached a maximum increase of approximately 250% - 300% at doses of 200 mg and above.

[0254] [Table 18] TIFF0007692411000024.tif35170

[0255] [Table 19A]

[0256] [Table 19B]

[0257]

Table 19C

[0258]

Table 19D

[0259] The present invention is not limited in scope by the specific embodiments described herein. Indeed, various modifications of the invention will become apparent to those skilled in the art in addition to those described. Such modifications are intended to be included within the scope of the appended claims. Item 1 A method for treating cardiovascular disease in a subject, comprising administering to the subject from about 100 mg to about 350 mg of an antibody or an antigen-binding fragment thereof that specifically binds to human endothelial lipase (EL). Item 2 A method for reducing atherosclerosis in a subject, comprising administering to the subject from about 100 mg to about 350 mg of an antibody or an antigen-binding fragment thereof that specifically binds to human EL. Item 3 A method for treating cardiovascular disease or reducing atherosclerosis in a subject, comprising administering to the subject an antibody or an antigen-binding fragment thereof that specifically binds to EL, wherein administration of the antibody or its antigen-binding fragment (a) increases high-density lipoprotein cholesterol (HDL-C) in the subject, (b) increases the number of high-density lipoprotein (HDL) particles in the subject, (c) increases the size of HDL particles in the subject, (d) increases HDL phospholipids in the subject, (e) increases apolipoprotein A1 (ApoA1) in the subject, and / or (f) increases cholesterol efflux capacity (CEC) in the subject. Item 4 The method according to item 3, wherein the administration reduces the risk of cardiovascular death, non-fatal myocardial infarction (MI), non-fatal stroke, and / or coronary artery revascularization in a subject with a history of acute coronary syndrome (ACS). Item 5 The method according to item 3 or 4, wherein the administration prevents secondary cardiovascular events in the subject. Item 6 The method according to any one of items 3 to 5, wherein the administration reduces the risk of major cardiovascular events (MACE) in the subject. Item 7 A method for reducing the risk of cardiovascular death, non-fatal myocardial infarction (MI), non-fatal stroke, and / or coronary artery revascularization in a subject with a history of acute coronary syndrome (ACS), comprising administering to the subject an antibody or an antigen-binding fragment thereof that specifically binds to human EL. Item 8 A method for preventing secondary cardiovascular events in a subject, the method comprising administering to the subject an antibody that specifically binds to human EL or an antigen-binding fragment thereof. Item 9 A method for reducing the risk of major cardiovascular events (MACE) in a subject, the method comprising administering to the subject an antibody that specifically binds to human EL or an antigen-binding fragment thereof. Item 10 The administration of the antibody or an antigen-binding fragment thereof (a) increases high-density lipoprotein cholesterol (HDL-C) in the subject, (b) increases the number of high-density lipoprotein (HDL) particles in the subject, (c) increases the HDL particle size in the subject, (d) increases HDL phospholipids in the subject, (e) increases ApoA1 in the subject, and / or (f) increases cholesterol efflux capacity (CEC) in the subject, the method according to any one of Items 7 to 9. Item 11 A method for increasing HDL-C, the number of HDL particles, HDL particle size, HDL phospholipids, ApoA1, and / or CEC in a subject, the method comprising administering to the subject an antibody that specifically binds to EL or an antigen-binding fragment thereof. Item 12 The method according to any one of Items 3 to 10, comprising administering about 100 mg to about 350 mg of the antibody or an antigen-binding fragment thereof. Item 13 The method according to any one of Items 1 to 12, comprising administering about 100 mg, about 110 mg, about 120 mg, about 125 mg, about 130 mg, about 140 mg, about 150 mg, about 160 mg, about 170 mg, about 175 mg, about 180 mg, about 190 mg, about 200 mg, about 210 mg, about 220 mg, about 225 mg, about 230 mg, about 240 mg, about 250 mg, about 260 mg, about 270 mg, about 275 mg, about 280 mg, about 290 mg, about 300 mg, about 310 mg, about 320 mg, about 325 mg, about 330 mg, about 340 mg, or about 350 mg of the antibody or an antigen-binding fragment thereof. Item 14 The method according to any one of Items 1 to 12, comprising administering about 250 mg of the antibody or an antigen-binding fragment thereof. Item 15 The method according to any one of Items 1 to 12, comprising administering 250 mg of the antibody or an antigen-binding fragment thereof. Item 16 The method according to any one of claims 1 to 12, comprising administering about 200 mg of said antibody or antigen-binding fragment thereof. Claim 17 The method according to any one of claims 1 to 12, comprising administering 200 mg to 250 mg of said antibody or antigen-binding fragment thereof. Claim 18 The method according to any one of claims 1 to 12, comprising administering about 125 mg of said antibody or antigen-binding fragment thereof. Claim 19 The method according to any one of claims 1 to 12, comprising administering 125 mg of said antibody or antigen-binding fragment thereof. Claim 20 The method according to any one of claims 1 to 19, wherein said antibody or antigen-binding fragment thereof is administered once a month. Claim 21 The method according to claim 20, wherein said antibody or antigen-binding fragment thereof is administered once a month for at least 3 months. Claim 22 The method according to claim 21, wherein said antibody or antigen-binding fragment thereof is administered once a month for at least 12 months or at least 24 months. Claim 23 The method according to any one of claims 1 to 22, wherein said antibody or antigen-binding fragment thereof is administered parenterally. Claim 24 The method according to claim 23, wherein said antibody or antigen-binding fragment thereof is administered subcutaneously. Claim 25 The method according to any one of claims 1 to 24, wherein said antibody or antigen-binding fragment thereof is administered via an attached prefilled syringe (APFS) or autoinjector. Claim 26 The method according to any one of claims 1 to 25, wherein administration of said antibody or antigen-binding fragment thereof inhibits EL in said subject for 30 days. Claim 27 The method according to any one of claims 1 to 26, wherein administration of said antibody or antigen-binding fragment thereof increases HDL-C in said subject by at least 30%. Claim 28 The method according to claim 27, wherein administration of said antibody or antigen-binding fragment thereof increases HDL-C in said subject by at least 35%. Claim 29 The method according to claim 28, wherein administration of said antibody or antigen-binding fragment thereof increases HDL-C in said subject by at least 40%. Claim 30 The method according to any one of claims 26 to 29, wherein administration of said antibody or antigen-binding fragment thereof increases HDL-C in said subject within 30 days from the first administration. Claim 31 The method according to any one of items 26 to 29, wherein administration of said antibody or antigen-binding fragment thereof increases HDL-C in said subject within 90 days from the first administration. Item 32 The method according to any one of items 1 to 31, wherein administration of said antibody or antigen-binding fragment thereof increases ApoA1 in said subject by at least 30%. Item 33 The method according to item 32, wherein administration of said antibody or antigen-binding fragment thereof increases ApoA1 in said subject by at least 35%. Item 34 The method according to item 32 or 33, wherein administration of said antibody or antigen-binding fragment thereof increases ApoA1 in said subject within 30 days from the first administration. Item 35 The method according to item 32 or 33, wherein administration of said antibody or antigen-binding fragment thereof increases ApoA1 in said subject within 90 days from the first administration. Item 36 The method according to any one of items 1 to 35, wherein administration of said antibody or antigen-binding fragment thereof increases the cholesterol efflux capacity of non-ATP-binding cassette transporter A1 (ABCA1) in said subject by at least 30%. Item 37 The method according to item 36, wherein administration of said antibody or antigen-binding fragment thereof increases the cholesterol efflux capacity of non-ABCA1 in said subject by at least 35%. Item 38 The method according to item 36 or 37, wherein administration of said antibody or antigen-binding fragment thereof increases the cholesterol efflux capacity of non-ABCA1 in said subject within 30 days from the first administration. Item 39 The method according to item 36 or 37, wherein administration of said antibody or antigen-binding fragment thereof increases the cholesterol efflux capacity of non-ABCA1 in said subject within 90 days from the first administration. Item 40 The method according to any one of items 1 to 39, wherein administration of said antibody or antigen-binding fragment thereof increases the number of HDL particles in said subject by at least 5%. Item 41 The method according to item 40, wherein administration of said antibody or antigen-binding fragment thereof increases the number of HDL particles in said subject by at least 8%. Item 42 The method according to item 40 or 41, wherein administration of said antibody or antigen-binding fragment thereof increases the number of HDL particles in said subject within 30 days from the first administration. Item 43 The method according to item 40 or 41, wherein administration of said antibody or antigen-binding fragment thereof increases the number of HDL particles in said subject within 90 days from the first administration. Item 44 The method according to any one of Items 1 to 43, wherein administration of the antibody or its antigen-binding fragment increases the HDL particle size in the subject by at least 3%. Item 45 The method according to Item 44, wherein administration of the antibody or its antigen-binding fragment increases the HDL particle size in the subject by at least 5%. Item 46 The method according to Item 44 or 45, wherein administration of the antibody or its antigen-binding fragment increases the HDL particle size in the subject within 30 days from the first administration. Item 47 The method according to Item 44 or 45, wherein administration of the antibody or its antigen-binding fragment increases the HDL particle size in the subject within 90 days from the first administration. Item 48 The method according to any one of Items 1 to 47, wherein administration of the antibody or its antigen-binding fragment increases the HDL phospholipid in the subject by at least 50%. Item 49 The method according to Item 47, wherein administration of the antibody or its antigen-binding fragment increases the HDL phospholipid in the subject within 30 days from the first administration. Item 50 The method according to Item 47, wherein administration of the antibody or its antigen-binding fragment increases the HDL phospholipid in the subject within 90 days from the first administration. Item 51 The method according to any one of Items 1 to 50, wherein administration of the antibody or its antigen-binding fragment increases the plasma phosphatidylinositol (PI) level in the subject by at least 100% or at least 250%. Item 52 The increase in the plasma PI level in the method according to item 51 is an increase in the PI(14:2 / 20:0) level, PI(14:2 / 22:0) level, PI(14:2 / 22:1) level, PI(14:2 / 22:2) level, PI(16:0 / 16:1) level, PI(16:0 / 18:0) level, PI(16:0 / 18:2) level, PI(16:0 / 20:2) level, PI(16:0 / 20:3) level, PI(16:0 / 20:4) level, PI(16:0 / 22:4) level, PI(16:1 / 18:0) level, PI(16:1 / 18:1) level, PI(18:0 / 18:0) level, PI(18:0 / 18:1) level, PI(18:0 / 18:2) level, PI(18:0 / 18:3) level, PI(18:0 / 20:2) level, PI(18:0 / 20:3) level, PI(18:0 / 20:4) level, PI(18:0 / 22:4) level, PI(18:0 / 22:5) level, PI(18:0 / 22:6) level, PI(18:1 / 16:0) level, PI(18:1 / 18:1) level, PI(18:1 / 18:2) level, PI(18:1 / 20:2) level, PI(18:1 / 20:3) level, PI(18:1 / 20:4) level, and / or PI(18:2 / 18:2) level. Item 53 The administration of the antibody or its antigen-binding fragment in the method according to item 51 or 52 increases the plasma phosphatidylinositol (PI) level in the subject within 90 days from the first administration. Item 54 The subject in the method according to any one of items 2 to 53 has a cardiovascular disease. Item 55 The cardiovascular disease in the method according to item 1 or 54 is coronary artery disease, coronary heart disease, chronic arterial disease, cerebrovascular disease, atherosclerotic cardiovascular disease, or peripheral arterial disease. Item 56 The subject in the method according to any one of items 1 to 53 has stable coronary artery disease or stable coronary heart disease. Item 57 The subject in the method according to any one of items 1 to 53 has a history of acute coronary syndrome (ACS). Item 58 The subject in the method according to any one of items 1 to 57 is receiving statin therapy. Item 59 The subject in the method according to any one of items 1 to 57 is not receiving statin therapy. Item 60 The method according to any one of items 1 to 59, wherein the subject has a triglyceride level of 500 mg / dL or less before the administration. Item 61 The method according to any one of items 1 to 60, wherein the subject has LDL-C of 100 mg / dL or less before the administration. Item 62 The method according to any one of items 1 to 61, wherein the subject is human. Item 63 The method according to any one of items 1 to 62, wherein the antibody or its antigen-binding fragment neutralizes EL activity. Item 64 The method according to any one of items 1 to 63, wherein the antibody or its antigen-binding fragment is accompanied by a decrease in effector function. Item 65 The method according to any one of items 1 to 64, wherein the antibody or its antigen-binding fragment does not have antibody-dependent cell-mediated cytotoxicity (ADCC) activity. Item 66 The method according to any one of items 1 to 65, wherein the antibody does not have complement-dependent cytotoxicity (CDC) activity. Item 67 The method according to any one of items 1 to 66, wherein the antibody binds to cynomolgus EL. Item 68 The method according to any one of items 1 to 67, wherein the antibody or its antigen-binding fragment competitively inhibits the binding of an antibody containing a VH comprising the amino acid sequence shown in SEQ ID NO: 7 and a VL comprising the amino acid sequence shown in SEQ ID NO: 8 to EL. Item 69 The method according to any one of items 1 to 68, wherein the antibody or its antigen-binding fragment binds to the same EL epitope as an antibody containing a VH comprising the amino acid sequence shown in SEQ ID NO: 7 and a VL comprising the amino acid sequence shown in SEQ ID NO: 8. Item 70 The method according to any one of items 1 to 69, wherein the antibody or its antigen-binding fragment comprises CDR1, VH CDR2, VH CDR3 of the heavy chain variable region (VH) of the sequence of MEDI5884, CDR1, VL CDR2, and VL CDR3 of the light chain variable region (VL). Item 71 The method according to item 70, wherein the CDR is a CDR according to the Kabat definition, a CDR according to the Chothia definition, or a CDR according to the AbM definition. Item 72 The antibody or antigen-binding fragment thereof according to any one of items 1 to 69, comprising a VH CDR1 containing the amino acid sequence of SEQ ID NO: 1, a VH CDR2 containing the amino acid sequence of SEQ ID NO: 2, a VH CDR3 containing the amino acid sequence of SEQ ID NO: 3, a VL CDR1 containing the amino acid sequence of SEQ ID NO: 4, a VL CDR2 containing the amino acid sequence of SEQ ID NO: 5, and a VL CDR3 containing the amino acid sequence of SEQ ID NO: 6. Item 73 The antibody or antigen-binding fragment thereof according to any one of items 1 to 72, comprising a VH containing the amino acid sequence shown in SEQ ID NO: 7 and / or a VL containing the amino acid sequence shown in SEQ ID NO: 8. Item 74 The antibody or antigen-binding fragment thereof according to any one of items 1 to 76, comprising an IgG heavy chain constant region. Item 75 The method according to item 74, wherein the IgG heavy chain constant region is an IgG4 heavy chain constant region. Item 76 The method according to item 75, wherein the IgG4 heavy chain constant region is an IgG4P heavy chain constant region. Item 77 The antibody or antigen-binding fragment thereof according to any one of items 1 to 76, comprising a kappa light chain constant region. Item 78 The antibody or antigen-binding fragment thereof according to any one of items 1 to 76, comprising a heavy chain constant region and / or a light chain constant region. Item 79 The method according to item 78, wherein the heavy chain constant region is a human IgG4P heavy chain constant region and / or the light chain constant region is a human IgGκ light chain constant region. Item 80 The antibody or antigen-binding fragment thereof according to any one of items 1 to 79, which is a humanized antibody or antigen-binding fragment thereof. Item 81 The antibody or antigen-binding fragment thereof according to any one of items 1 to 80, comprising a heavy chain constant region containing the amino acid sequence shown in SEQ ID NO: 12 and / or a light chain constant region containing the amino acid sequence shown in SEQ ID NO: 13. Item 82 The antibody according to any one of items 1 to 81, comprising a heavy chain containing the amino acid sequence shown in SEQ ID NO: 9 and a light chain containing the amino acid sequence shown in SEQ ID NO: 10. Item 83 The antibody or antigen-binding fragment thereof according to any one of items 1 to 82, which is a full-length antibody. Item 84 The antibody or antigen-binding fragment thereof according to any one of items 1 to 80, which is an antigen-binding fragment. Item 85 The antigen-binding fragment is Fab, Fab’, F(ab’) 2 , a single-chain antibody Fv (scFv), disulfide-linked Fv, V-NAR domain, IgNar, intrabody, IgGΔCH2, minibody, F(ab’) 3 , tetrabody, tribody, diabody, single-domain antibody, DVD-Ig, Fcab, mAb 2 , (scFv) 2 , or the method according to item 84, comprising scFv-Fc Item 86 A method for treating cardiovascular disease in a subject, comprising subcutaneously administering 250 mg of an antibody or an antigen-binding fragment thereof to the subject once a month, wherein the antibody or antigen-binding fragment specifically binds to human EL and comprises a VH comprising the amino acid sequence shown in SEQ ID NO: 7 and a VL comprising the amino acid sequence shown in SEQ ID NO: 8 Item 87 A method for reducing the risk of cardiovascular death, non-fatal myocardial infarction (MI), non-fatal stroke, and / or coronary artery revascularization in a subject with a history of acute coronary syndrome (ACS), comprising subcutaneously administering 250 mg of an antibody or an antigen-binding fragment thereof to the subject once a month, wherein the antibody or antigen-binding fragment specifically binds to human EL and comprises a VH comprising the amino acid sequence shown in SEQ ID NO: 7 and a VL comprising the amino acid sequence shown in SEQ ID NO: 8 Item 88 A method for treating cardiovascular disease in a subject, comprising subcutaneously administering about 200 mg of an antibody or an antigen-binding fragment thereof to the subject once a month, wherein the antibody or antigen-binding fragment specifically binds to human EL and comprises a VH comprising the amino acid sequence shown in SEQ ID NO: 7 and a VL comprising the amino acid sequence shown in SEQ ID NO: 8 Item 89 A method for reducing the risk of cardiovascular death, non-fatal myocardial infarction (MI), non-fatal stroke, and / or coronary artery revascularization in a subject with a history of acute coronary syndrome (ACS), comprising subcutaneously administering about 200 mg of an antibody or an antigen-binding fragment thereof to the subject once a month, wherein the antibody or antigen-binding fragment specifically binds to human EL and comprises a VH comprising the amino acid sequence shown in SEQ ID NO: 7 and a VL comprising the amino acid sequence shown in SEQ ID NO: 8 Item 90 A method for treating cardiovascular disease in a subject, comprising subcutaneously administering 200 mg to 250 mg of an antibody or an antigen-binding fragment thereof to the subject once a month, wherein the antibody or antigen-binding fragment specifically binds to human EL and comprises a VH comprising the amino acid sequence shown in SEQ ID NO: 7 and a VL comprising the amino acid sequence shown in SEQ ID NO: 8. Item 91 A method for reducing the risk of cardiovascular death, non-fatal myocardial infarction (MI), non-fatal stroke, and / or coronary artery revascularization in a subject with a history of acute coronary syndrome (ACS), comprising subcutaneously administering 200 mg to 250 mg of an antibody or an antigen-binding fragment thereof to the subject once a month, wherein the antibody or antigen-binding fragment specifically binds to human EL and comprises a VH comprising the amino acid sequence shown in SEQ ID NO: 7 and a VL comprising the amino acid sequence shown in SEQ ID NO: 8. Item 92 The method according to any one of Items 86 to 91, wherein the antibody comprises the amino acid sequence of the heavy chain constant region shown in SEQ ID NO: 9 and the amino acid sequence of the light chain constant region shown in SEQ ID NO: 10. Item 93 The method according to any one of Items 1 to 92, further comprising administering an inhibitor of PCSK9. Item 94 The method according to Item 93, wherein the administration of the antibody or antigen-binding fragment thereof that specifically binds to human EL and the administration of the inhibitor of PCSK9 are simultaneous. Item 95 The method according to Item 94, wherein the antibody or antigen-binding fragment thereof that specifically binds to human EL and the inhibitor of PCSK9 are administered in separate pharmaceutical compositions. Item 96 The method according to Item 93, wherein the administration of the antibody or antigen-binding fragment thereof that specifically binds to human EL and the administration of the inhibitor of PCSK9 are sequential. Item 97 The method according to any one of Items 93 to 96, wherein the inhibitor of PCSK9 is an anti-PCSK9 antibody or an antigen-binding fragment thereof. Item 98 The method according to Item 97, wherein the inhibitor of PCSK9 is inclisiran, evolocumab, alirocumab, or bococizumab.

[0260] All references (e.g., publications, patents, or patent applications) cited in this specification are hereby incorporated by reference in their entirety for all purposes to the same extent as if each individual reference (e.g., publication, patent, or patent application) were specifically and individually indicated and made a part of this specification for all purposes.

[0261] Other embodiments are within the scope of the following claims.

Claims

1. A composition for treating cardiovascular disease or reducing atherosclerosis in a subject, wherein the composition contains 50 mg to 500 mg of an anti-endothelial lipase (EL) antibody having a VH CDR1 with the amino acid sequence of SEQ ID NO: 1, a VH CDR2 with the amino acid sequence of SEQ ID NO: 2, a VH CDR3 with the amino acid sequence of SEQ ID NO: 3, a VL CDR1 with the amino acid sequence of SEQ ID NO: 4, a VL CDR2 with the amino acid sequence of SEQ ID NO: 5, and a VL CDR3 with the amino acid sequence of SEQ ID NO: 6, wherein the composition is administered once a month, wherein the composition (a) increases high-density lipoprotein cholesterol (HDL-C) in the subject, (b) increases the number of high-density lipoprotein (HDL) particles in the subject, (c) increases the HDL particle size in the subject, (d) increases HDL phospholipids in the subject, (e) increases apolipoprotein A1 (ApoA1) in the subject, (f) increases cholesterol efflux capacity (CEC) in the subject, and / or (g) increases plasma phosphatidylinositol (PI) in the subject, composition.

2. The composition according to claim 1, which reduces the risk of cardiovascular death, non-fatal myocardial infarction (MI), non-fatal stroke, peripheral artery disease, atherosclerosis, and / or coronary artery revascularization in a subject with a history of acute coronary syndrome (ACS).

3. The composition according to claim 1 or 2, which prevents secondary cardiovascular events in the subject.

4. The composition according to any one of claims 1 to 3, which reduces the risk of major adverse cardiovascular events (MACE) in the subject.

5. The composition according to any one of claims 1 to 4, which contains 200 mg to 500 mg of the anti-EL antibody.

6. The method according to any one of claims 1 to 5, wherein the composition is administered parenterally.

7. The method according to any one of claims 1 to 6, wherein the composition is administered subcutaneously.

8. The method according to any one of claims 1 to 7, wherein the composition is administered via an attached prefilled syringe (APFS) or an autoinjector.

9. The composition according to any one of claims 1 to 8, which increases HDL-C in the subject by at least 30%.

10. The composition according to any one of claims 1 to 9, which increases ApoA1 in the subject by at least 30%.

11. The composition according to any one of claims 1 to 10, which increases the number of HDL particles in the subject by at least 5%.

12. The composition according to any one of claims 1 to 11, which increases the HDL particle size in the subject by at least 3%.

13. The composition according to any one of claims 1 to 12, which increases HDL phospholipids in the subject by at least 50%.

14. The composition according to any one of claims 1 to 13, wherein administration of the composition increases the non-ABCA1 cholesterol efflux capacity in the subject by at least 35%.

15. The composition increases the plasma phosphatidylinositol (PI) level in the subject, and the increase in the plasma PI level is an increase in the level of PI(14:2 / 20:0), PI(14:2 / 22:0), PI(14:2 / 22:1), PI(14:2 / 22:2), PI(16:0 / 16:1), PI(16:0 / 18:0), PI(16:0 / 18:2), PI(16:0 / 20:2), PI(16:0 / 20:3), PI(16:0 / 20:4), PI(16:0 / 22:4), PI(16:1 / 18:0), PI(16:1 / 18:1), PI(18:0 / 18:0), PI(18:0 / 18:1), PI(18:0 / 18:2), PI(18:0 / 18:3), PI(18:0 / 20:2), PI(18:0 / 20:3), PI(18:0 / 20:4), PI(18:0 / 22:4), PI(18:0 / 22:5), PI(18:0 / 22:6), PI(18:1 / 16:0), PI(18:1 / 18:1), PI(18:1 / 18:2), PI(18:1 / 20:2), PI(18:1 / 20:3), PI(18:1 / 20:4), and / or PI(18:2 / 18:2). The composition according to any one of claims 1 to 14.

16. The cardiovascular disease is selected from coronary artery disease, coronary heart disease, chronic arterial disease, cerebrovascular disease, atherosclerotic cardiovascular disease, and peripheral arterial disease. The composition according to any one of claims 1 to 15.

17. The composition according to any one of claims 1 to 16 for treating peripheral arterial disease.

18. The subject is receiving statin therapy. The composition according to any one of claims 1 to 17.

19. The subject has a triglyceride level of 500 mg / dL or less before the administration. The composition according to any one of claims 1 to 18.

20. The anti-EL antibody comprises a VH containing the amino acid sequence shown in SEQ ID NO: 7 and / or a VL containing the amino acid sequence shown in SEQ ID NO:

8. The composition according to any one of claims 1 to 19.

21. The composition according to any one of claims 1 to 20, further characterized in that the composition and the inhibitor of PCSK9 are administered simultaneously or sequentially in succession.

22. The inhibitor of PCSK9 is an anti-PCSK9 antibody or an antigen-binding fragment thereof, and the anti-PCSK9 antibody or an antigen-binding fragment thereof is selected from HS9, evolocumab, alirocumab, and bococizumab, the composition according to claim 21.

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