A method for treating atherosclerotic cardiovascular disease with an LPA-targeted RNAi construct.

Abstract

To provide a method for treating atherosclerotic cardiovascular disease using an LPA-targeted RNAi construct. [Solution] The present invention relates to a method for treating or preventing atherosclerotic cardiovascular disease and other conditions associated with elevated levels of lipoprotein (a) (Lp(a)) using RNAi constructs that target the LPA gene encoding apolipoprotein (a), a component of Lp(a) particles. In particular, the present invention relates to a method for lowering serum Lp(a) levels and reducing the risk of cardiovascular events in patients with elevated Lp(a) levels, comprising administering an LPA-targeted RNAi construct according to a specific dosing schedule. A pharmaceutical composition comprising an LPA-targeted RNAi construct used in the method is also disclosed.

Description

[Technical Field]

[0001] Cross-reference of related applications This application claims the benefits of U.S. Provisional Patent Application No. 63 / 110,309, filed November 5, 2020, which is incorporated herein by reference in its entirety.

[0002] Description of electronically submitted text files This application is electronically filed in ASCII format and includes a sequence listing which is incorporated herein by reference in its entirety. A computer-readable copy of the sequence listing, prepared on November 1, 2021, is named A-2694-WO-PCT_ST25 and is 3.5 kilobytes in size.

[0003] The present invention relates to pharmaceutical compositions and methods for treating atherosclerotic cardiovascular disease and other conditions associated with elevated lipoprotein (a) (Lp(a)). In particular, the present invention relates to a method for reducing serum Lp(a) levels and lowering the risk of cardiovascular events, such as cardiovascular death, myocardial infarction, stroke, and coronary artery regeneration, by administering an LPA-targeted RNAi construct according to a specific dosing regimen in patients with elevated Lp(a). [Background technology]

[0004] Atherosclerotic cardiovascular disease is extremely common and remains the leading cause of death worldwide, despite the widespread use of low-density lipoprotein (LDL) lowering therapy. While LDL lowering therapy reduces the risk of major cardiac events, the remaining cardiovascular risks faced by some patients with low LDL levels suggest other mechanisms of cardiovascular pathology. Over the past few decades, compelling evidence from epidemiological studies and meta-analyses, Mendelian randomized analyses, and genome-wide association studies has shown that as serum Lp(a) levels increase, the risk of coronary artery disease and atherosclerosis-related disorders increases (Clarke et al., N.Engl.J.Med., Vol.361:2518-2528, 2009; Kamstrup et al., JAMA, Vol.301:2331-2339, 2009; Nordestgaard et al. al.,European Heart Journal,Vol.31:2844-2853,2010;Helgadottir et al.,J.Am.Coll.Cardiol,Vol.60:722-729,2012;Thanassoulis et al.,J.Am.Coll.Cardiol.,Vol.55:2491-2498,2010;Kamstrup et al. al.,J.Am.Coll. Cardiol.,Vol.63:470-477,2014;Kral et al.,Journal of Cardiology,Vol.118:656-661,2016;Thanassoulis et al.,J.Lipid Res.,Vol.57:917-924,2016;Tsimikas et al. (al., J. Am. Coll. Cardiol., Vol. 69: 692-711, 2017). In particular, the relationship between Lp(a) levels and coronary artery disease, myocardial infarction, stroke, peripheral vascular disease, and aortic stenosis has been described in several genetic and observational studies (reviewed in Schmidt et al., J. Lipid Res., Vol. 57: 1339-1359, 2016). This risk relationship is continuum, and it is known that the effect increases proportionally with higher Lp(a) levels. This association persists even after adjusting for other lipid parameters (Emerging Risk Factors Collaboration, JAMA, Vol. 302: 412-423, 2009).

[0005] Lp(a) is a low-density lipoprotein consisting of LDL particles and the glycoprotein apolipoprotein (a) (apo(a)) linked to apolipoprotein B of the LDL particles by disulfide bonds (Schmidt et al., op. cit.). apo(a) is encoded by the LPA gene and is expressed almost exclusively in primates, including humans. apo(a) shows homology to plasminogen and exists in various isoforms due to intragenetic size polymorphism resulting from a variable number of Kringle IV2 (KIV-2) domain repeats (see Kronenberg and Utermann, J.Intern.Med., Vol.273:6-30, 2013). An inverse correlation has been observed between the size of the apo(a) isoform and the plasma level of Lp(a) particles (Sandholzer et al., Hum.Genet., Vol.86;607-614, 1991). Lp(a) contains pro-inflammatory oxidized phospholipids that contribute to its atherogenic effect (Tsimikas et al., J.Am.Coll.Cardiol., Vol.63:1724-1734, 2014).

[0006] High plasma Lp(a) concentrations are genetically defined, maintained at stable levels, cannot be regulated by lifestyle changes (diet, exercise, or other environmental factors), and are not effectively regulated by any of the lipid-lowering drugs currently available. Currently, there are no approved therapies that demonstrate a reduction in the risk of cardiovascular events by lowering Lp(a). Lp(a) can be gently lowered (by about 20-30%) with the proprotein convertase subtilisin / kexin type 9 (PCSK9) inhibitor, niacin, or mipomersen (Santos et al., Arterioscler. Thromb. Vasc. Biol., Vol.35:689-699, 2015; Yeang et al., Curr. Opin. Lipidol., Vol.26:169-178, 2015; and Landray et al., N.Engl. J.Med., Vol.371:203-212, 2014). Apheresis is effective in lowering Lp(a), but is currently only available in a few countries with limited access (Julius, J. Cardiovasc. Dev. Dis., Vol.5:27-37, 2018). In addition, apheresis is an invasive and extremely expensive procedure that requires frequent hospital visits, making it unsuitable as a long-term treatment for patients requiring lifelong care (Khan et al., Eur. Heart J., Vol.38:1561-1569, 2017; Roeseler et al., Arterioscler.Thromb.Vasc.Biol., Vol.36:2019-2027, 2016; Leebmann et al., Circulation, Vol.128:2567-2576, 2013; Safarova et al., Atheroscler.Suppl., Vol.14:93-99, 2013).

[0007] Antisense oligonucleotides targeting apo(a) messenger RNA transcripts (AKCEA-APO(a)-LRx; also known as ISIS 681257 and TQJ230) have been developed and are currently undergoing clinical trials (reviewed in Graham et al., J Lipid Res., Vol.57:340-351, 2016). In healthy subjects with baseline Lp(a) levels of 75 nmol / L or higher, administration of 10 mg, 20 mg, or 40 mg on days 1, 3, 5, 8, 15, and 22 has been reported to reduce Lp(a) concentrations by an average of 66%, 80%, and 92% on day 36, and by an average of 39%, 53%, and 58% on day 113 (Viney et al., Lancet, Vol.388:2239-2253, 2016). In subsequent Phase 2 studies, AKCEA-APO(a)-LRx reduced Lp(a) levels by an average of 35%, 56%, and 72% at week 25 in patients with established cardiovascular disease and baseline Lp(a) levels of 150 nmol / L or higher, when administered once every four weeks at doses of 20 mg, 40 mg, or 60 mg (Tsimikas et al., New England Journal of Medicine, Vol.382:244-255, 2020). A Phase 3 cardiovascular outcomes trial is ongoing, with the molecule administered monthly at a dose of 80 mg (ClinicalTrials.gov identifier NCT04023552). However, there is still a need in this technology for new therapeutic agents that can rapidly and persistently lower Lp(a) levels, enabling low-dose, low-frequency administration plans for the treatment and prevention of atherosclerotic cardiovascular disease. [Prior art documents] [Non-patent literature]

[0008] [Non-Patent Document 1] Clarke et al.,N.Engl.J.Med.,Vol.361:2518-2528,2009

Non-Patent Document 2

Non-Patent Document 3

Non-Patent Document 4

Non-Patent Document 5

Non-Patent Document 6

Non-Patent Document 7

Non-Patent Document 8

Non-Patent Document 9

Non-Patent Document 10

Non-Patent Document 11

Non-Patent Document 12

Summary of the Invention

Means for Solving the Problem

[0009] The present invention is, in part, based on the identification of an LPA-targeting RNAi construct, particularly inclisiran, for effectively reducing circulating Lp(a) levels for the treatment of atherosclerotic cardiovascular disease. Thus, in some embodiments, the present invention provides a method of reducing serum or plasma Lp(a) levels in a patient who needs to reduce serum or plasma Lp(a) levels, the method comprising administering to the patient an LPA RNAi construct described herein at a dose of about 9 mg to about 675 mg at an administration interval of at least 8 weeks. In some such embodiments, the patient to whom the LPA RNAi construct is administered is diagnosed with or at risk of developing a cardiovascular disease, such as coronary artery disease, carotid artery disease, peripheral artery disease, myocardial infarction, cerebrovascular disease, stroke, aortic valve stenosis, stable or unstable angina, atrial fibrillation, heart failure, hyperlipidemia, heterozygous familial hypercholesterolemia, or homozygous familial hypercholesterolemia. The patient may have a history or family history of myocardial infarction and / or may be diagnosed with acute coronary syndrome. In other embodiments, the patient to whom the LPA RNAi construct is administered is diagnosed with chronic kidney disease.

[0010] In certain embodiments, the present invention provides a method for treating, alleviating, or preventing atherosclerosis in patients who require treatment, alleviation, or prevention of atherosclerosis, or for treating, alleviating, or preventing cardiovascular disease in patients who require treatment, alleviation, or prevention of cardiovascular disease. In such embodiments, the method comprises administering to a patient a dose of about 9 mg to about 675 mg of an LPA RNAi construct described herein at dosing intervals of at least 8 weeks. Cardiovascular diseases that may be treated, remitted, alleviated, or prevented by the method of the present invention include coronary artery disease, carotid artery disease, peripheral artery disease, myocardial infarction, cerebrovascular disease, stroke, aortic stenosis, stable or unstable angina, atrial fibrillation, heart failure, hyperlipidemia, heterozygous familial hypercholesterolemia, or homozygous familial hypercholesterolemia.

[0011] The present invention also includes a method of reducing the risk of cardiovascular events in patients with atherosclerotic cardiovascular disease. In some embodiments, the method comprises administering to a patient an LPA RNAi construct described herein at a dose of about 9 mg to about 675 mg at an administration interval of at least 8 weeks. The cardiovascular event can be a major cardiovascular event, such as cardiovascular death, non-fatal myocardial infarction, non-fatal stroke, or hospitalization for unstable angina. In some embodiments, the cardiovascular event can be a major adverse limb event, such as acute limb ischemia, major amputation surgery, or peripheral vascular revascularization for ischemia. In certain embodiments, the cardiovascular event is cardiovascular death, myocardial infarction, stroke, and / or coronary artery revascularization. The patient with atherosclerotic cardiovascular disease to whom the LPA RNAi construct is to be administered may have a history of coronary artery revascularization, a history of coronary artery bypass grafting, a diagnosis of coronary artery disease, a diagnosis of atherosclerotic cerebrovascular disease, a diagnosis of peripheral artery disease, and / or a history of myocardial infarction. In one embodiment, the patient to whom the LPA RNAi construct is to be administered has recently experienced a myocardial infarction event, for example, the patient has experienced a myocardial infarction within 1 year before the first administration of the LPA RNAi construct. In another embodiment, the patient to whom the LPA RNAi construct is to be administered is hospitalized for acute coronary syndrome or unstable angina.

[0012] The patient to whom the LPA RNAi construct is to be administered according to the method of the present invention has an elevated serum or plasma level of Lp(a). In some embodiments, the patient has a serum or plasma Lp(a) level of about 70 nmol / L or higher before the first administration of the LPA RNAi construct. In other embodiments, the patient has a serum or plasma Lp(a) level of about 150 nmol / L or higher before the first administration of the LPA RNAi construct. In certain embodiments, the patient has a serum or plasma Lp(a) level of about 175 nmol / L or higher before the first administration of the LPA RNAi construct. In certain other embodiments, the patient has a serum or plasma Lp(a) level of about 200 nmol / L or higher before the first administration of the LPA RNAi construct.

[0013] In some embodiments of the method of the present invention, the patient to whom the LPA RNAi construct is administered is undergoing lipid-lowering therapy, for example, to lower the patient's LDL-LC levels. Lipid-lowering therapy may include PCSK9 inhibitors, e.g., PCSK9 antagonist monoclonal antibodies (e.g., evolocumab, alirocumab), statins (e.g., atorvastatin, cerivastatin, flavastatin, lovastatin, mevastatin, pitavastatin, pravastatin, rosuvastatin, simvastatin), cholesterol absorption inhibitors (e.g., ezetimibe), bempedoic acid, nicotinic acid (e.g., niacin), fibrinic acid (e.g., gemfibrozil, fenofibrate), bile acid adsorbents (e.g., cholestyramine, colestipol, coleseveram), LDL apheresis, or a combination thereof. In these and other embodiments, the patient may have a serum LDL-C level of approximately 100 mg / dL or less or approximately 70 mg / dL or less prior to the first administration of the LPA RNAi construct.

[0014] In certain embodiments of the method of the present invention, a fixed dose of the LPA RNAi construct is administered to the patient once every 12 weeks or once every 3 months. In some such embodiments, the fixed dose may be about 10 mg to about 225 mg, about 75 mg to about 225 mg, about 50 mg to about 100 mg, or about 150 mg to about 225 mg. In one embodiment, LPA The RNAi construct is administered to the patient at a fixed dose of approximately 10 mg once every 12 weeks or once every 3 months. In another embodiment, the LPA RNAi construct is administered to the patient at a fixed dose of approximately 75 mg once every 12 weeks or once every 3 months. In yet another embodiment, the LPA RNAi construct is administered to the patient at a fixed dose of approximately 150 mg once every 12 weeks or once every 3 months. In yet another embodiment, the LPA RNAi construct is administered to the patient at a fixed dose of approximately 225 mg once every 12 weeks or once every 3 months.

[0015] In certain other embodiments of the method of the present invention, a fixed dose of the LPA RNAi construct is administered to the patient once every 24 weeks or once every 6 months. In some such embodiments, the fixed dose may be about 225 mg to about 675 mg, about 225 mg to about 450 mg, or about 200 mg to about 300 mg. In some embodiments, the LPA RNAi construct is administered to the patient in a fixed dose of about 225 mg once every 24 weeks or once every 6 months. In other embodiments, the LPA RNAi construct is administered to the patient in a fixed dose of about 300 mg once every 24 weeks or once every 6 months. In certain embodiments, the LPA RNAi construct is administered to the patient in a fixed dose of about 450 mg once every 24 weeks or once every 6 months. In certain other embodiments, the LPA RNAi construct is administered to the patient in a fixed dose of about 675 mg once every 24 weeks or once every 6 months.

[0016] Administration of an LPA RNAi construct to a patient according to the method of the present invention substantially lowers the patient's plasma or serum Lp(a) levels over a long period. For example, in some embodiments of the method of the present invention, administration of an LPA RNAi construct lowers the patient's serum or plasma Lp(a) levels by more than 80% for at least 12 weeks, at least 16 weeks, or at least 24 weeks compared to the patient's baseline serum or plasma Lp(a) levels. In other embodiments of the method of the present invention, administration of an LPA RNAi construct lowers the patient's serum or plasma Lp(a) levels by more than 90% for at least 12 weeks, at least 16 weeks, or at least 24 weeks compared to the patient's baseline serum or plasma Lp(a) levels. In certain embodiments, administration of an LPA RNAi construct to a patient according to the method of the present invention lowers the patient's plasma or serum Lp(a) levels to about 100 nmol / L or less. In some embodiments, administration of an LPA RNAi construct to a patient according to the method of the present invention lowers the patient's plasma or serum Lp(a) levels to about 75 nmol / L or less. In another embodiment, administration of the LPA RNAi construct to a patient according to the method of the present invention reduces the patient's plasma or serum Lp(a) level to about 50 nmol / L or less.

[0017] In any embodiment of the method disclosed herein, the LPA administered to the patient The RNAi construct may be a double-stranded RNA molecule, such as an siRNA molecule, comprising a sense strand and an antisense strand, wherein the antisense strand contains a region having a sequence complementary to the LPA mRNA sequence. Preferably, the sense strand contains a sequence sufficiently complementary to the antisense strand to form a double-stranded region of about 15 to about 30 base pairs in length. In certain embodiments of the method of the present invention, the LPA RNAi construct administered to a patient comprises a sense strand and an antisense strand, each having a length of about 19 to about 23 nucleotides, wherein the antisense strand contains a sequence complementary to the LPA mRNA sequence, and the sense strand contains a sequence complementary to the antisense strand. In such one embodiment, the sense strand and antisense strand of the LPA RNAi construct may each be 21 nucleotides in length and can hybridize to form a 21-base-pair-long double-stranded region such that the RNAi construct has two blunt ends. In another such embodiment, the sense strand and antisense strand of the LPA RNAi construct may each be 19 nucleotides long and hybridize with each other to form a 19-base-pair-long double-stranded region such that the RNAi construct has two blunt ends.

[0018] In some embodiments of the method of the present invention, the LPA RNAi construct administered to a patient further comprises a targeting moiety containing an asialoglycoprotein receptor ligand, the targeting moiety being covalently bonded to the sense strand, for example, to the 5' end of the sense strand. The targeting moiety may include a trivalent GalNAc moiety, for example, a moiety having the structure of Structure 1 described herein. In certain embodiments, the LPA RNAi construct administered to a patient according to the method of the present invention comprises a sense strand containing the sequence of SEQ ID NO: 1 and an antisense strand containing the sequence of SEQ ID NO: 2. In some embodiments, the LPA RNAi construct comprises a sense strand containing or derived from the sequence of SEQ ID NO: 3, and an antisense strand containing or derived from the sequence of SEQ ID NO: 4. In certain preferred embodiments, the sense strand and / or antisense strand of the LPA RNAi construct comprises one or more modified nucleotides. In such embodiments, the LPA RNAi construct comprises a sense strand containing or derived from a sequence of modified nucleotides according to SEQ ID NO: 5, and an antisense strand containing or derived from a sequence of modified nucleotides according to SEQ ID NO: 6. In a preferred embodiment, the LPA RNAi construct administered to the patient according to the method of the present invention is olpasilan.

[0019] The present invention also provides a pharmaceutical composition comprising an LPA RNAi construct, e.g., olpasilan, for use in the methods of the present invention described herein. The pharmaceutical composition may comprise one or more pharmaceutically acceptable diluents, carriers, or excipients. In certain embodiments, the pharmaceutical composition comprises an LPA RNAi construct (e.g., olpasilan), potassium phosphate buffer, and sodium chloride, wherein the composition has a pH of about 6.6 to about 7.0, preferably about 6.8. Any pharmaceutical composition described herein may be incorporated into an injection device, e.g., a pre-filled syringe, an autoinjector, an injection pump, an on-body injector, and an injection pen, for administration to a patient (e.g., subcutaneous administration) according to the methods described herein. In some embodiments, administration of an LPA RNAi construct (e.g., olpasilan), or a pharmaceutical composition comprising an LPA RNAi construct (e.g., olpasilan), to a patient according to the methods of the present invention is by subcutaneous injection. In such embodiments, the injection volume is about 2 mL or less, or about 1 mL or less, e.g., about 1 mL.

[0020] The use of LPA RNAi constructs for the preparation of drugs for administration in any of the methods disclosed herein, or in accordance with any of the methods disclosed herein, is specifically intended. For example, the present invention includes an LPA RNAi construct used in a method for treating, alleviating, or preventing atherosclerosis or cardiovascular disease in a patient who needs to treat, alleviate, or prevent atherosclerosis or cardiovascular disease, the method comprising administering the LPA RNAi construct to the patient in doses of about 9 mg to about 675 mg at dosing intervals of at least 8 weeks. The present invention also includes an LPA RNAi construct used in a method for lowering serum or plasma Lp(a) levels in a patient, the method comprising administering the LPA RNAi construct to the patient in doses of about 9 mg to about 675 mg at dosing intervals of at least 8 weeks. In certain embodiments, the present invention provides an LPA RNAi construct used in a method for reducing the risk of cardiovascular events in patients with atherosclerotic cardiovascular disease, the method comprising administering the LPA RNAi construct to the patient in doses of about 9 mg to about 675 mg at intervals of at least 8 weeks.

[0021] Furthermore, the present invention encompasses the use of LPA RNAi constructs in the preparation of agents for treating, alleviating, or preventing atherosclerosis or cardiovascular disease in patients who require treatment, alleviation, or prevention of atherosclerosis or cardiovascular disease, the agents being administered at doses of approximately 9 mg to approximately 675 mg at intervals of at least 8 weeks, or formulated for administration at such doses. In some embodiments, the present invention provides the use of LPA RNAi constructs in the preparation of agents for lowering serum or plasma Lp(a) levels in patients, the agents being administered at doses of approximately 9 mg to approximately 675 mg at intervals of at least 8 weeks, or formulated for administration at such doses. In other embodiments, the present invention provides the use of an LPA RNAi construct in the preparation of a drug for reducing the risk of cardiovascular events in patients with atherosclerotic cardiovascular disease, the drug being administered at doses of approximately 9 mg to approximately 675 mg at intervals of at least 8 weeks, or being formulated for administration at such doses. [Brief explanation of the drawing]

[0022] [Figure 1] The structure of the LPA RNAi construct, orpasilan, is schematically shown. The upper strand, arranged from 5' to 3', is the sense strand (SEQ ID NO: 5), and the lower strand, arranged from 3' to 5', is the antisense strand (SEQ ID NO: 6). Black circles represent nucleotides with 2'-O-methyl modification, white circles represent nucleotides with 2'-deoxy-2'-fluoro modification, and gray circles represent deoxyadenosine nucleotides linked to adjacent nucleotides via 3'-3' bonds (i.e., inverted). Gray lines connecting the circles represent phosphodiester bonds, while black lines connecting the circles represent phosphorothioate bonds. The trivalent GalNAc moiety with the shown structure is represented by R1 and is covalently bonded to the 5' end of the sense strand via a phosphorothioate bond. [Figure 2]These line graphs show the percentage change from baseline in plasma Lp(a) levels in human subjects after a single subcutaneous dose of placebo or olpasilan at the indicated dose, across each of the seven cohorts (1–7) throughout the study days. Baseline values ​​were the mean of screening Lp(a) and day 1 predose Lp(a) levels. If only one value was available, that value was used as the baseline. [Figure 3-1] For subjects with a baseline Lp(a) level of ≥150 nmol / L, the predicted baseline Lp(a) levels are shown as percentages for quarterly (Q3M) administration of olpasilan at doses of 10 mg (Figure 3A), 30 mg (Figure 3B), 50 mg (Figure 3C), 75 mg (Figure 3D), 150 mg (Figure 3E), and 225 mg (Figure 3F). The horizontal line in each graph represents an 80% reduction in Lp(a) level from baseline. The predicted Lp(a) levels are based on PK / PD model simulations for 10,000 subjects. Predicted data are shown as median values ​​(solid lines), with the 95% prediction interval represented by shading. The black circles in Figure 3D represent observational data from Cohort 7 described in Example 1. [Figure 3-2] For subjects with a baseline Lp(a) level of ≥150 nmol / L, the predicted baseline Lp(a) levels are shown as percentages for quarterly (Q3M) administration of olpasilan at doses of 10 mg (Figure 3A), 30 mg (Figure 3B), 50 mg (Figure 3C), 75 mg (Figure 3D), 150 mg (Figure 3E), and 225 mg (Figure 3F). The horizontal line in each graph represents an 80% reduction in Lp(a) level from baseline. The predicted Lp(a) levels are based on PK / PD model simulations for 10,000 subjects. Predicted data are shown as median values ​​(solid lines), with the 95% prediction interval represented by shading. The black circles in Figure 3D represent observational data from Cohort 7 described in Example 1. [Figure 4-1]For subjects with a baseline Lp(a) level of ≥150 nmol / L, the predicted baseline Lp(a) levels for twice-yearly (Q6M) administration of olpasilan at doses of 10 mg (Figure 4A), 75 mg (Figure 4B), 150 mg (Figure 4C), 225 mg (Figure 4D), 450 mg (Figure 4E), and 675 mg (Figure 4F) are shown as percentages. The horizontal line in each graph represents an 80% reduction in the Lp(a) level from baseline. The predicted Lp(a) levels are based on PK / PD model simulations for 10,000 subjects. Predicted data are shown as median values ​​(solid lines), and the 95% prediction interval is represented by shading. The black circles in Figure 4B represent observational data from Cohort 7 described in Example 1. [Figure 4-2] For subjects with a baseline Lp(a) level of ≥150 nmol / L, the predicted baseline Lp(a) levels for twice-yearly (Q6M) administration of olpasilan at doses of 10 mg (Figure 4A), 75 mg (Figure 4B), 150 mg (Figure 4C), 225 mg (Figure 4D), 450 mg (Figure 4E), and 675 mg (Figure 4F) are shown as percentages. The horizontal line in each graph represents an 80% reduction in the Lp(a) level from baseline. The predicted Lp(a) levels are based on PK / PD model simulations for 10,000 subjects. Predicted data are shown as median values ​​(solid lines), and the 95% prediction interval is represented by shading. The black circles in Figure 4B represent observational data from Cohort 7 described in Example 1. [Modes for carrying out the invention]

[0023] Lp(a) has been reported to be a risk factor for various forms of cardiovascular disease, including myocardial infarction, stroke, peripheral artery disease, and aortic stenosis. Lp(a) levels are genetically determined and, unlike LDL cholesterol (LDL-C) levels, cannot be altered by diet, exercise, or other lifestyle changes. Currently, there are no approved therapies that selectively target apo(a) to substantially lower Lp(a) levels. This invention provides a novel administration scheme for an RNAi construct that targets mRNA transcribed from the LPA gene encoding the apo(a) protein for sustained suppression of Lp(a) levels for the treatment or prevention of atherosclerosis and associated cardiovascular symptoms. A specific LPA-targeting RNAi construct, olpasilan, was observed to reduce Lp(a) levels by 71% to 96% after a single dose in human subjects with baseline Lp(a) levels ≥70 nmol / L, with a maximum percentage reduction of >90%. The effect persisted for more than 6 months with single doses of 9 mg or higher (see Example 1). Specifically, a single dose of as low as 9 mg of olpasilan reduced Lp(a) levels by more than 80% for more than 3 months in human subjects, while single doses of 75 mg and 225 mg of olpasilan suppressed Lp(a) levels by more than 80% for more than 6 months. Furthermore, olpasilan was well-tolerated at these doses, and there were no serious treatment-related adverse events (see Example 1). This robust and sustained suppression of Lp(a) within this dosage range was unexpected. This is because, based on allometric scaling of olpasilan doses evaluated in cynomolgus monkeys, it was predicted that an eight-fold higher dose (e.g., 75 mg vs. 9 mg) would be required to achieve an 80% reduction in Lp(a) over one month.

[0024] Furthermore, the depth and duration of Lp(a) suppression by olpasilan in human subjects were surprising, considering the results reported in human subjects with other nucleic acid therapies targeting apo(a). AKCEA-APO(a)-LRx, an antisense oligonucleotide targeting apo(a), has been reported to reduce Lp(a) levels by 35% to 80% in human subjects 6 months after treatment. However, achieving 80% and 72% reductions in Lp(a) levels, respectively, required a weekly dose of 20 mg or a monthly dose of 60 mg of AKCEA-APO(a)-LRx (see Tsimikas et al., New England Journal of Medicine, Vol.382:244-255, 2020). In contrast, as described herein, a single dose of olpasilan has enabled administration of olpasilan at lower doses and longer dosing intervals, for example, once every three months or once every six months, by resulting in a reduction of more than 80% of Lp(a) levels for longer than six months. Thus, the method of the present invention achieves considerable improvements in human treatment for atherosclerotic cardiovascular disease, such as improved patient adherence, reduced drug costs, and reduced volume and frequency of injections. Accordingly, in certain embodiments, the present invention provides a method for treating, preventing, or reducing the risk of developing cardiovascular disease in a patient, comprising administering an effective amount of LPA RNAi construct to the patient according to a specific dosing schedule described herein.

[0025] Atherosclerosis is a disease in which plaques composed of fatty substances, cholesterol, calcium, fibrin, and cellular waste products form in various arteries throughout the body. Over time, the plaques harden and narrow the lumen of the arteries, restricting blood flow to organs and tissues in the body. Atherosclerosis can lead to the development of several other diseases, such as cardiovascular disease, cerebrovascular disease, or chronic kidney disease, depending on the specific arteries affected by the accumulation of atherosclerotic plaques. For example, coronary artery disease can occur if plaque forms in the coronary arteries and partially blocks blood flow to the heart, which can lead to angina and myocardial infarction. The formation of atherosclerotic plaques in the carotid arteries, which supply oxygen-rich blood to the brain, can lead to carotid artery disease, and if blood flow is reduced or blocked, it can cause a transient ischemic attack or stroke. When plaques form in the major arteries supplying blood to the limbs and pelvis, peripheral artery disease may develop, potentially leading to abdominal aortic aneurysms and limb ischemia (causing paralysis and pain). When atherosclerotic plaques accumulate in the renal arteries, chronic kidney disease may develop, leading to a decline in renal function over time, which can result in renal failure. Lp(a) is an atherogenic lipoprotein. Elevated levels of Lp(a) have been associated with an increased risk of coronary artery disease, peripheral artery disease, myocardial infarction, and stroke, in particular. The methods of the present invention are useful in treating, mitigating, or preventing atherosclerosis in patients by lowering circulating Lp(a) levels. Therefore, in some embodiments, the present invention provides a method for treating, mitigating, or preventing atherosclerosis in patients, comprising administering an effective amount of an LPA RNAi construct to a patient according to one of the dosing regimens described herein. In one embodiment, the present invention comprises the use of any of the LPA RNAi constructs described herein for the preparation of a drug for treating, alleviating, or preventing atherosclerosis in a patient who needs to do so, the drug being administered according to any of the dosing regimens described herein or being formulated for administration according to such regimens.In another embodiment, the present invention provides an LPA RNAi construct used in a method for treating, alleviating, or preventing atherosclerosis in a patient who requires such treatment, alleviation, or prevention, the method comprising administering the LPA RNAi construct in accordance with any of the administration schedules described herein.

[0026] In certain embodiments, the present invention also provides a method for treating, alleviating, relieving, or preventing cardiovascular disease in a patient who needs to do so, comprising administering an effective amount of an LPA RNAi construct to the patient according to one of the dosing regimens described herein. In some embodiments, the present invention comprises the use of one of the LPA RNAi constructs described herein for the preparation of a drug for treating, alleviating, or preventing cardiovascular disease in a patient who needs to do so, the drug being administered according to one of the dosing regimens described herein or being formulated for such administration. In other embodiments, the present invention provides an LPA RNAi construct used in a method for treating, alleviating, or preventing cardiovascular disease in a patient who needs to do so, for example, one of the LPA RNAi constructs described herein, the method comprising administering the LPA RNAi construct according to one of the dosing regimens described herein. Cardiovascular disease is a class of diseases and conditions affecting the blood vessels or heart, and includes, but is not limited to, myocardial infarction, heart failure, transient ischemic attack, stroke (ischemic and hemorrhagic), atherosclerosis, coronary artery disease, peripheral vascular disease (e.g., peripheral artery disease), aneurysms (e.g., abdominal aortic aneurysm), carotid artery disease, cerebrovascular disease, stable or unstable angina, atrial fibrillation, hyperlipidemia, familial hypercholesterolemia (heterozygous and homozygous), unstable plaque, and aortic stenosis. Therefore, in certain embodiments, patients to be treated according to the methods of the present invention are diagnosed with cardiovascular disease and / or at risk of developing it. Patients at risk of developing cardiovascular disease may have a family history of cardiovascular disease and / or may have one or more risk factors for cardiovascular disease. Such risk factors include, but are not limited to, hypertension, elevated levels of non-HDL cholesterol, elevated levels of triglycerides, diabetes mellitus, obesity, or smoking.The diagnosis of atherosclerosis and cardiovascular disease can be made using various methods known to those skilled in the art, including one or more of the following: a patient's health examination and family history, the patient's risk factors, physical examination, blood tests measuring various biomarkers such as lipid concentrations (e.g., LDL-C, triglycerides, Lp(a), glycated hemoglobin A1C, C-reactive protein, apolipoprotein B, cardiac troponin T, etc.), electrocardiogram, echocardiogram, stress test, chest X-ray, computed tomography (CT) scan (e.g., cardiac CT scan), and angiography.

[0027] In some embodiments, the cardiovascular disease to be treated, alleviated, remitted, or prevented according to the methods of the present invention is coronary artery disease. Signs and symptoms of coronary artery disease may include chest pain (e.g., angina), shortness of breath, myocardial infarction, stenosis of one or more coronary arteries, pain or discomfort in the arm or shoulder, weakness, dizziness, nausea, and a history of coronary artery bypass and / or percutaneous coronary intervention. In relevant embodiments, the cardiovascular disease to be treated, alleviated, remitted, or prevented according to the methods of the present invention is myocardial infarction.

[0028] In other embodiments, cardiovascular diseases that are treated, mitigated, remitted, or prevented according to the methods of the present invention are coronary artery diseases, particularly atherosclerotic cerebrovascular diseases. Cerebrovascular diseases refer to disorders in which areas of the brain are temporarily or permanently affected by ischemia or hemorrhage due to dysfunction or complications of one or more cerebral blood vessels. Examples of cerebrovascular diseases, but not limited to, include transient ischemic attacks, strokes (ischemic or hemorrhagic), carotid artery stenosis, vertebral artery stenosis, intracranial artery stenosis, aneurysms, and vascular malformations. Signs and symptoms of cerebrovascular disease may include dizziness, nausea, vomiting, very severe headache, confusion, disorientation, memory loss, paralysis or weakness of the arms, legs, or face (especially unilaterally), abnormal or slurred speech, difficulty understanding, loss of visual field or difficulty seeing, loss of balance, coordination, or ability to walk, carotid stenosis, and a history of transient ischemic attacks and / or carotid vascular regeneration. In one embodiment, the cardiovascular disease to be treated, alleviated, remitted, or prevented according to the method of the present invention is stroke.

[0029] In certain other embodiments, the cardiovascular disease treated or prevented according to the methods of the present invention is peripheral artery disease. Signs and symptoms of peripheral artery disease may include pain or muscle spasms in the leg or arm during walking (claudication), paralysis or weakness of the leg, coldness of the lower leg or foot, pain in the toes, unhealed foot or leg, discoloration of the leg, hair loss or slower hair growth of the foot and leg, slower growth of toenails, shiny skin of the leg, no or weak pulse in the leg or foot, ankle-brachial index ≤ 0.90, and a history of abdominal aortic aneurysm, abdominal aortic treatment (percutaneous or surgical), and / or peripheral artery vascular regeneration (percutaneous or surgical).

[0030] In some embodiments, administration of an LPA RNAi construct according to the method of the present invention is for the treatment of atherosclerosis and other cardiovascular diseases and symptoms. As used herein, the terms “treatment” or “to treat” mean the application or administration of an LPA RNAi construct to a patient who has or has been diagnosed with atherosclerosis or other cardiovascular disease, has signs of atherosclerosis or other cardiovascular disease, is at risk of developing atherosclerosis or other cardiovascular disease, or has a predisposition to atherosclerosis or other cardiovascular disease, one or more symptoms of atherosclerosis or other cardiovascular disease, is at risk of developing atherosclerosis or other cardiovascular disease, or has a tendency to develop atherosclerosis or other cardiovascular disease, for the purpose of treating, curing, alleviating, reducing, modifying, relieving, or improving atherosclerosis or other cardiovascular disease. The term “treatment” encompasses any improvement in the patient’s condition, including slowing or halting the progression of atherosclerosis or other cardiovascular disease, reducing the number or severity of signs of atherosclerosis or other cardiovascular disease, or increasing the frequency or length of periods in the patient that are free from signs of atherosclerosis or other cardiovascular disease. As used herein, the term “patient” refers to a mammal, including humans, and may be used interchangeably with the term “subject.” In a preferred embodiment, the patient is a human patient.

[0031] In certain preferred embodiments, administration of an LPA RNAi construct to a patient according to any of the methods of the present invention reduces the circulating Lp(a) level or concentration (e.g., serum or plasma Lp(a) level / concentration) in the patient compared to the circulating Lp(a) level (e.g., baseline Lp(a) level / concentration) in the patient before administration of the LPA RNAi construct, or compared to the circulating Lp(a) level / concentration in the patient not receiving the LPA RNAi construct. Accordingly, in some embodiments, the present invention provides a method for reducing serum or plasma Lp(a) levels (or concentrations) in a patient who requires such reduction, comprising administering an LPA RNAi construct to the patient according to any of the dosing schedules described herein. In one embodiment, the present invention includes the use of any of the LPA RNAi constructs described herein for the preparation of a drug for reducing serum or plasma Lp(a) levels (or concentrations) in a patient who requires such reduction, the drug being administered according to any of the dosing schedules described herein or formulated for administration according to such a schedule. In another embodiment, the present invention provides an LPA RNAi construct used in a method for lowering serum or plasma Lp(a) levels (or concentrations) in a patient who requires such reduction, for example, any of the LPA RNAi constructs described herein, the method comprising administering the LPA RNAi construct according to any of the administration plans described herein. In some embodiments, the patient who requires a reduction in serum or plasma Lp(a) levels (or concentrations) is a patient diagnosed with or at risk of cardiovascular disease, for example, any of the cardiovascular diseases described above. In some such embodiments, the cardiovascular disease is coronary artery disease, carotid artery disease, peripheral artery disease, myocardial infarction, cerebrovascular disease, stroke, aortic stenosis, stable or unstable angina, atrial fibrillation, heart failure, hyperlipidemia, heterozygous familial hypercholesterolemia, or homozygous familial hypercholesterolemia. In a particular embodiment, the patient who requires a reduction in serum or plasma Lp(a) levels (or concentrations) has a history of myocardial infarction or a family history of myocardial infarction.

[0032] In certain embodiments, patients requiring a reduction in serum or plasma Lp(a) levels (or concentrations) are diagnosed with acute coronary syndrome. Acute coronary syndrome refers to symptoms associated with a sudden reduction in blood flow to the heart, often caused by the breakdown of atherosclerotic plaques and partial or complete thrombosis of the coronary arteries. Examples of acute coronary syndrome include acute myocardial ischemia or infarction, such as non-ST-elevation myocardial infarction (NSTEMI) and ST-elevation myocardial infarction (STEMI), as well as unstable angina. Even if acute coronary syndrome does not initially result in an infarct, it is a sign of a high risk of infarction and must be diagnosed and treated promptly. The signs and symptoms of acute coronary syndrome typically begin abruptly and may include chest pain (angina) or discomfort, pain radiating from the chest to the shoulder, arm, upper abdomen, back, neck, or jaw, nausea or vomiting, indigestion, shortness of breath, sudden heavy sweating, dizziness, lightheadedness, or fainting, unusual or inexplicable fatigue, and feelings of restlessness or anxiety.

[0033] In certain other embodiments, patients requiring a reduction in serum or plasma Lp(a) levels (or concentrations) are diagnosed with chronic kidney disease. Chronic kidney disease generally refers to progressive damage to and loss of function of the kidneys. Chronic kidney disease worsens over time, and patients may be at increased risk of other cardiovascular diseases. In one embodiment, a patient to be treated according to the method of the present invention has stage 3 chronic kidney disease. The stage of kidney disease is determined by the estimated glomerular filtration rate (eGFR), which is a value based on the amount of creatinine in the blood. Stage 3 chronic kidney disease is approximately 30 mL / min / 1.73 m 2 Approximately 59 mL / min / 1.73 m 2 Characterized by an eGFR of approximately 15 mL / min / 1.73m², the patient may have some initial symptoms, such as swelling in the hands and feet, back pain, and urination, to a greater or lesser degree than normal. Additionally, patients with stage 3 chronic kidney disease may have other health-related problems, such as hypertension, anemia, and bone disease. In another embodiment, the patient to be treated according to the method of the present invention has stage 4 chronic kidney disease. Patients with stage 4 chronic kidney disease have an eGFR of approximately 15 mL / min / 1.73m². 2Approximately 29 mL / min / 1.73 m 2 Typically, this manifests as more or less symptomatic swelling of the hands and feet, back pain, and urination than normal.

[0034] In some embodiments, administration of an LPA RNAi construct to a patient according to the method of the present invention reduces the patient's serum or plasma Lp(a) level (or concentration) by at least about 50%, at least about 55%, at least about 60%, at least about 65%, at least about 70%, at least about 75%, at least about 80%, at least about 85%, at least about 90%, at least about 91%, at least about 92%, at least about 93%, at least about 94%, or at least about 95% compared to the patient's serum or plasma Lp(a) level (or concentration) before administration of the RNAi construct (e.g., baseline Lp(a) level or concentration), or compared to the patient's serum or plasma Lp(a) level (or concentration) not receiving the RNAi construct. In these and other embodiments, after administration of the LPA RNAi construct (e.g., a single dose of the LPA RNAi construct), circulating Lp(a) levels or concentrations are reduced in the patient for at least 4 weeks, at least 6 weeks, at least 8 weeks, at least 10 weeks, at least 12 weeks, at least 14 weeks, at least 16 weeks, at least 18 weeks, at least 20 weeks, at least 22 weeks, at least 24 weeks, at least 26 weeks, at least 28 weeks, at least 30 weeks, at least 32 weeks, at least 36 weeks, or at least 48 weeks.

[0035] In one embodiment of the method of the present invention, administration of an LPA RNAi construct (e.g., a single dose of the LPA RNAi construct) reduces the patient's serum or plasma Lp(a) level (or concentration) by more than 50% for at least 12 weeks compared to the patient's baseline serum or plasma Lp(a) level (or concentration). The baseline serum or plasma Lp(a) level (or concentration) refers to the patient's serum or plasma Lp(a) level (or concentration) before administration of the LPA RNAi construct (i.e., pretreatment level or concentration). The baseline level / concentration may be a single measurement taken before the patient receives the LPA RNAi construct, or it may be the average of two or more measurements taken before the patient receives the LPA RNAi construct. In another embodiment of the method of the present invention, administration of an LPA RNAi construct (e.g., a single dose of an LPA RNAi construct) reduces the patient's serum or plasma Lp(a) level (or concentration) by more than 50% for at least 24 weeks compared to the patient's baseline serum or plasma Lp(a) level (or concentration). In yet another embodiment of the method of the present invention, administration of an LPA RNAi construct (e.g., a single dose of an LPA RNAi construct) reduces the patient's serum or plasma Lp(a) level (or concentration) by more than 80% for at least 12 weeks compared to the patient's baseline serum or plasma Lp(a) level (or concentration). In yet another embodiment of the method of the present invention, administration of an LPA RNAi construct (e.g., a single dose of an LPA RNAi construct) reduces the patient's serum or plasma Lp(a) level (or concentration) by more than 80% for at least 24 weeks compared to the patient's baseline serum or plasma Lp(a) level (or concentration). In a particular embodiment of the method of the present invention, administration of an LPA RNAi construct (e.g., a single dose of an LPA RNAi construct) reduces the patient's serum or plasma Lp(a) level (or concentration) by more than 80% for at least 32 weeks compared to the patient's baseline serum or plasma Lp(a) level (or concentration).In some embodiments of the method of the present invention, administration of an LPA RNAi construct (e.g., a single dose of an LPA RNAi construct) reduces the patient's serum or plasma Lp(a) level (or concentration) by more than 90% for at least 12 weeks compared to the patient's baseline serum or plasma Lp(a) level (or concentration). In other embodiments of the method of the present invention, administration of an LPA RNAi construct (e.g., a single dose of an LPA RNAi construct) reduces the patient's serum or plasma Lp(a) level (or concentration) by more than 90% for at least 16 weeks compared to the patient's baseline serum or plasma Lp(a) level (or concentration).

[0036] In certain embodiments, administration of an LPA RNAi construct to a patient according to the method of the present invention reduces the absolute Lp(a) level (or concentration) in the patient's serum or plasma to about 150 nmol / L or less, about 125 nmol / L or less, about 100 nmol / L or less, about 75 nmol / L or less, about 70 nmol / L or less, about 65 nmol / L or less, about 60 nmol / L or less, about 55 nmol / L or less, about 50 nmol / L, about 45 nmol / L or less, about 40 nmol / L or less, about 35 nmol / L or less, or about 30 nmol / L or less. In one embodiment, administration of an LPA RNAi construct to a patient according to the method of the present invention reduces the absolute Lp(a) level (or concentration) in the patient's serum or plasma to about 125 nmol / L or less. In another embodiment, administration of an LPA RNAi construct to a patient according to the method of the present invention reduces the absolute Lp(a) level (or concentration) in the patient's serum or plasma to about 100 nmol / L or less. In yet another embodiment, administration of an LPA RNAi construct to a patient according to the method of the present invention reduces the absolute Lp(a) level (or concentration) in the patient's serum or plasma to about 75 nmol / L or less. In yet another embodiment, administration of an LPA RNAi construct to a patient according to the method of the present invention reduces the absolute Lp(a) level (or concentration) in the patient's serum or plasma to about 50 nmol / L or less.

[0037] While there is a preference for measuring Lp(a) levels / concentrations in units of particle concentration (e.g., nmol / L) (see, e.g., Wilson et al., Journal of Clinical Lipidology, Vol. 13; 374-392, 2019), Lp(a) levels may also be measured in units of mass concentration (e.g., mg / dL). In such embodiments, administration of an LPA RNAi construct to a patient according to the method of the present invention can reduce the Lp(a) level (or concentration) in the patient's serum or plasma to about 100 mg / dL or less, about 90 mg / dL or less, about 80 mg / dL or less, about 70 mg / dL or less, about 60 mg / dL or less, about 50 mg / dL or less, about 45 mg / dL or less, about 40 mg / dL or less, about 35 mg / dL or less, about 30 mg / dL or less, about 25 mg / dL or less, about 20 mg / dL or less, or about 15 mg / dL or less.

[0038] Lp(a) levels in plasma or serum samples can be measured using commercially available kits, such as the Lp(a) ELISA assay kit from Mercodia AB (Uppsala, Sweden), the Lp(a) immunoturbidimetric assay from Randox Laboratories Ltd. (Crumlin, United Kingdom), or the Tina-quant® Lp(a) Gen.2 assay from F. Hoffmann-La Roche Ltd. (Basel, Switzerland), or other methods known in the art, such as those described in Marcovina and Albers, J. Lipid Res., Vol. 57: 526-537, 2016. In certain embodiments, Lp(a) levels are measured using a turbidimetric immunoassay normalized to detect and quantify Lp(a) particles independent of apo(a) isoform size. In these and other embodiments, the assays used to measure Lp(a) levels are normalized to nmol / L against the IFCC reference material SRM2B (Marcovina et al., Clin. Chem., Vol. 46: 1946-1967, 2000).

[0039] As previously described, elevated levels of circulating Lp(a) are associated with an increased risk of cardiovascular disease. Therefore, the methods of the present invention are also useful in reducing the risk of cardiovascular events in patients with elevated serum or plasma levels of Lp(a). Accordingly, in certain embodiments, the present invention provides a method for reducing the risk of cardiovascular events in patients with atherosclerotic cardiovascular disease, comprising administering an effective amount of an LPA RNAi construct to a patient according to one of the dosing regimens described herein. In one embodiment, the present invention comprises the use of one of the LPA RNAi constructs described herein for the preparation of a drug for reducing the risk of cardiovascular events in patients with atherosclerotic cardiovascular disease, the drug being administered according to one of the dosing regimens described herein or being formulated for administration according to such regimen. In another embodiment, the present invention provides an LPA RNAi construct, such as any of the LPA RNAi constructs described herein, for use in a method to reduce the risk of cardiovascular events in patients with atherosclerotic cardiovascular disease, the method comprising administering the LPA RNAi construct according to any of the administration plans described herein.

[0040] In some embodiments, a cardiovascular event is one or more of the following: cardiovascular death, myocardial infarction, stroke (e.g., ischemic stroke), coronary artery regeneration, hospitalization for unstable angina, hospitalization for heart failure, peripheral artery regeneration, acute limb ischemia, transient ischemic attack, major limb amputation for ischemia, cerebral artery regeneration (all resulting in death). In certain embodiments, a cardiovascular event is cardiovascular death, myocardial infarction, stroke (e.g., ischemic stroke), and / or coronary artery regeneration. In some such embodiments, a cardiovascular event is cardiovascular death, myocardial infarction, and / or coronary artery regeneration. In other such embodiments, a cardiovascular event is myocardial infarction and / or coronary artery regeneration. In other embodiments, a cardiovascular event is a major cardiovascular event selected from cardiovascular death, non-fatal myocardial infarction, non-fatal stroke, and hospitalization for unstable angina. In yet another embodiment, the cardiovascular event is a major adverse limb event selected from acute limb ischemia, major amputation, and peripheral vascular regeneration due to ischemia. In one embodiment, the cardiovascular event is cardiovascular death. In another embodiment, the cardiovascular event is a non-fatal myocardial infarction. In yet another embodiment, the cardiovascular event is a non-fatal stroke (e.g., ischemic stroke). In yet another embodiment, the cardiovascular event is coronary artery regeneration.

[0041] In certain embodiments, patients administered with an LPA RNAi construct according to the method of the present invention have a relative risk reduction of at least 15%, at least 20%, at least 25%, or at least 30% for any of the cardiovascular events described above, compared to patients who do not receive the LPA RNAi construct. In one embodiment, patients administered with an LPA RNAi construct according to the method of the present invention have a relative risk reduction of about 15% to about 25% for any one of cardiovascular death, myocardial infarction, and ischemic stroke, compared to patients who do not receive the LPA RNAi construct. In another embodiment, patients administered with an LPA RNAi construct according to the method of the present invention have a relative risk reduction of about 20% to about 30% for any one of cardiovascular death, myocardial infarction, and ischemic stroke, compared to patients who do not receive the LPA RNAi construct.

[0042] In certain other embodiments, patients administered with an LPA RNAi construct according to the method of the present invention have an absolute risk reduction of at least 1.5%, at least 1.8%, at least 2.0%, at least 2.2%, at least 2.5%, at least 2.8%, at least 3.0%, at least 3.2%, or at least 3.5% for any of the cardiovascular events described above. In one embodiment, patients administered with an LPA RNAi construct according to the method of the present invention have an absolute risk reduction of about 1.5% to about 3.0% for any one of cardiovascular death, myocardial infarction, and ischemic stroke. In another embodiment, patients administered with an LPA RNAi construct according to the method of the present invention have an absolute risk reduction of about 2.0% to about 3.5% for any one of cardiovascular death, myocardial infarction, and ischemic stroke. In yet another embodiment, patients administered an LPA RNAi construct according to the method of the present invention exhibit an absolute risk reduction of approximately 2.0% to approximately 3.0% for any one of the following: cardiovascular death, myocardial infarction, and ischemic stroke.

[0043] In any of the embodiments described above, patients administered the LPA RNAi construct according to the method of the present invention to reduce cardiovascular events may have a history of coronary artery regeneration, a history of coronary artery bypass grafting, a diagnosis of coronary artery disease, a diagnosis of atherosclerotic cerebrovascular disease, a diagnosis of peripheral artery disease, and / or a history of myocardial infarction. In certain embodiments, patients administered the LPA RNAi construct according to the method of the present invention to reduce cardiovascular events have experienced myocardial infarction. For example, in some such embodiments, patients administered the LPA RNAi construct according to the method of the present invention to reduce cardiovascular events have experienced myocardial infarction within one, two, three, four, or five years of receiving the first dose of the LPA RNAi construct. In one such embodiment, patients administered the LPA RNAi construct according to the method of the present invention to reduce cardiovascular events have experienced myocardial infarction within one year of receiving the first dose of the LPA RNAi construct. In certain other embodiments, patients administered the LPA RNAi construct according to the method of the present invention to reduce cardiovascular events are hospitalized or have recently been hospitalized for acute coronary syndrome or unstable angina.

[0044] In certain preferred embodiments, patients to whom the LPA RNAi construct is administered according to the method of the present invention are patients with elevated circulating levels or concentrations of Lp(a) (e.g., elevated serum or plasma levels / concentrations of Lp(a)). Patients to whom the LPA RNAi construct is administered according to the method of the present invention may have baseline circulating Lp(a) levels or concentrations of approximately 50 nmol / L or higher, approximately 55 nmol / L or higher, approximately 60 nmol / L or higher, approximately 65 nmol / L or higher, approximately 70 nmol / L or higher, approximately 75 nmol / L or higher, approximately 100 nmol / L or higher, approximately 125 nmol / L or higher, approximately 150 nmol / L or higher, approximately 175 nmol / L or higher, approximately 200 nmol / L or higher, approximately 225 nmol / L or higher, or approximately 250 nmol / L or higher. In one embodiment, if the patient's serum or plasma Lp(a) level (or concentration) prior to the first administration of the LPA RNAi construct is about 70 nmol / L or higher, the LPA RNAi construct is administered according to the method of the present invention. In another embodiment, if the patient's serum or plasma Lp(a) level (or concentration) prior to the first administration of the LPA RNAi construct is about 100 nmol / L or higher, the LPA RNAi construct is administered according to the method of the present invention. In yet another embodiment, if the patient's serum or plasma Lp(a) level (or concentration) prior to the first administration of the LPA RNAi construct is about 125 nmol / L or higher, the LPA RNAi construct is administered according to the method of the present invention. In yet another embodiment, if the patient's serum or plasma Lp(a) level (or concentration) prior to the first administration of the LPA RNAi construct is about 150 nmol / L or higher, the LPA RNAi construct is administered according to the method of the present invention. In some embodiments, the patient is administered the LPA RNAi construct according to the method of the present invention if their serum or plasma Lp(a) level (or concentration) prior to the first administration of the LPA RNAi construct is about 175 nmol / L or higher. In other embodiments, the patient is administered the LPA RNAi construct according to the method of the present invention if their serum or plasma Lp(a) level (or concentration) prior to the first administration of the LPA RNAi construct is about 200 nmol / L or higher.In certain other embodiments, the patient is administered the LPA RNAi construct according to the method of the present invention if their serum or plasma Lp(a) level (or concentration) prior to the first administration of the LPA RNAi construct is approximately 225 nmol / L or higher.

[0045] In less preferred embodiments where the circulating Lp(a) level (or concentration) is measured in mass concentration units, patients to whom the LPA RNAi construct is administered according to the method of the present invention may have a circulating Lp(a) level (or concentration) of approximately 30 mg / dL or higher, approximately 35 mg / dL or higher, approximately 40 mg / dL or higher, approximately 45 mg / dL or higher, approximately 50 mg / dL or higher, approximately 55 mg / dL or higher, approximately 60 mg / dL or higher, approximately 65 mg / dL or higher, approximately 70 mg / dL or higher, approximately 75 mg / dL or higher, approximately 90 mg / dL or higher, or approximately 100 mg / dL or higher. In one embodiment, a patient is administered the RNAi construct of the present invention if their serum or plasma Lp(a) level (or concentration) prior to the first administration of the LPA RNAi construct is approximately 50 mg / dL or higher. In another embodiment, the patient is administered the RNAi construct of the present invention if their serum or plasma Lp(a) level (or concentration) prior to the first administration of the LPA RNAi construct is approximately 60 mg / dL or higher. In yet another embodiment, the patient is administered the RNAi construct of the present invention if their serum or plasma Lp(a) level (or concentration) prior to the first administration of the LPA RNAi construct is approximately 70 mg / dL or higher. In yet another embodiment, the patient is administered the RNAi construct of the present invention if their serum or plasma Lp(a) level (or concentration) prior to the first administration of the LPA RNAi construct is approximately 90 mg / dL or higher.

[0046] As discussed above, Lp(a) levels (or concentrations) in plasma or serum samples can be measured using commercially available kits, such as the Lp(a) ELISA assay kit from Mercodia AB (Uppsala, Sweden), the Lp(a) immunoturbidimetric assay from Randox Laboratories Ltd. (Crumlin, United Kingdom), or the Tina-quant® Lp(a) Gen.2 assay from F. Hoffmann-La Roche Ltd. (Basel, Switzerland), or other methods known in the art, such as those described in Marcovina and Albers, J. Lipid Res., Vol. 57: 526-537, 2016. In certain embodiments, Lp(a) levels are measured using a turbidimetric immunoassay normalized to detect and quantify Lp(a) particles independent of apo(a) isoform size. In these and other embodiments, the assays used to measure Lp(a) levels are normalized to nmol / L against the IFCC reference material SRM2B (Marcovina et al., Clin. Chem., Vol. 46: 1946-1967, 2000).

[0047] In some embodiments, patients receiving the LPA RNAi construct according to the method of the present invention have serum low-density lipoprotein cholesterol (LDL-C) levels that are within the normal range or can be adjusted to the normal range through treatment with one or more lipid-reducing therapies. For example, in one embodiment, patients receiving the LPA RNAi construct according to the method of the present invention have serum LDL-C levels of about 100 mg / dL or less prior to the first administration of the LPA RNAi construct. In another embodiment, patients receiving the LPA RNAi construct according to the method of the present invention have serum LDL-C levels of about 70 mg / dL or less prior to the first administration of the LPA RNAi construct. In related embodiments, patients receiving the LPA RNAi construct according to the method of the present invention are undergoing one or more lipid-reducing therapies. Lipid-lowering therapies include, but are not limited to, PCSK9 inhibitors, e.g., PCSK9 antagonist monoclonal antibodies (e.g., evolocumab, alirocumab) and PCSK9-targeted siRNAs (e.g., inclisilan), statins (e.g., atorvastatin, cerivastatin, flavastatin, lovastatin, mevastatin, pitavastatin, pravastatin, rosuvastatin, simvastatin), cholesterol absorption inhibitors (e.g., ezetimibe), bempedoic acid, nicotinic acid (e.g., niacin), fibrinic acid (e.g., gemfibrozil, fenofibrate), bile acid adsorbents (e.g., cholestyramine, colestipol, coleseveram), LDL apheresis, or combinations thereof. In certain embodiments, patients to be administered the LPA RNAi construct according to the method of the present invention are receiving lipid-reducing therapy selected from the group consisting of PCSK9 antagonist monoclonal antibodies, statins, ezetimibe, bempedoic acid, or a combination thereof.

[0048] In certain embodiments, patients to be administered the LPA RNAi construct according to the method of the present invention have a serum triglyceride level of less than approximately 500 mg / dL prior to the first administration of the LPA RNAi construct. For example, a patient's baseline serum triglyceride level (e.g., prior to the first administration of the LPA RNAi construct) may be less than approximately 400 mg / dL, less than approximately 375 mg / dL, less than approximately 350 mg / dL, less than approximately 325 mg / dL, less than approximately 300 mg / dL, less than approximately 275 mg / dL, less than approximately 250 mg / dL, less than approximately 225 mg / dL, less than approximately 200 mg / dL, less than approximately 175 mg / dL, or less than approximately 150 mg / dL. In one embodiment, patients to be administered the LPA RNAi construct according to the method of the present invention have a serum triglyceride level of less than approximately 400 mg / dL prior to the first administration of the LPA RNAi construct. In another embodiment, patients to be administered the LPA RNAi construct according to the method of the present invention have a serum triglyceride level of approximately 50 mg / dL to approximately 400 mg / dL prior to the first administration of the LPA RNAi construct. In yet another embodiment, patients to be administered the LPA RNAi construct according to the method of the present invention have a serum triglyceride level of approximately 150 mg / dL to approximately 375 mg / dL prior to the first administration of the LPA RNAi construct.

[0049] Measurements of LDL-C, triglycerides, total cholesterol, high-density lipoprotein cholesterol (HDL-C), very low-density lipoprotein cholesterol (VLDL-C), and other lipid biomarkers, such as apolipoprotein A1 and apolipoprotein B, can be measured using a standard lipid panel with patient-derived blood samples. In some embodiments, the patient fasts for at least 9 hours, preferably 12 hours, before the sample is collected. Therefore, the levels / concentrations of the aforementioned lipid biomarkers (e.g., LDL-C, triglycerides) may be at fasting levels.

[0050] In some embodiments, patients who are administered the LPA RNAi construct according to the method of the present invention do not exhibit type 2 diabetes with untreated or poorly controlled glycated hemoglobin A1C levels. For example, patients who are administered the LPA RNAi construct according to the method of the present invention have baseline (e.g., before the first administration of the LPA RNAi construct) glycated hemoglobin A1C levels of less than 10.0%, less than 9.5%, less than 9.0%, less than 8.5%, less than 8.0%, less than 7.5%, less than 7.0%, less than 6.5%, less than 6.0%, or less than 5.5%. In one embodiment, patients who are administered the LPA RNAi construct according to the method of the present invention have a glycated hemoglobin A1C level of less than 8.5% before the first administration of the LPA RNAi construct. In another embodiment, patients to be administered the LPA RNAi construct according to the method of the present invention have a glycated hemoglobin A1C level of less than approximately 7.0% prior to the first administration of the LPA RNAi construct.

[0051] In other embodiments, patients to be administered the LPA RNAi construct according to the method of the present invention do not exhibit poorly controlled systolic and / or diastolic blood pressure. For example, patients to be administered the LPA RNAi construct according to the method of the present invention have a mean resting systolic blood pressure at baseline (e.g., before the first administration of the LPA RNAi construct) of less than approximately 180 mmHg, less than approximately 160 mmHg, less than approximately 140 mmHg, less than approximately 135 mmHg, less than approximately 130 mmHg, less than approximately 125 mmHg, or less than approximately 120 mmHg, and a mean resting diastolic blood pressure at baseline (e.g., before the first administration of the LPA RNAi construct) of less than approximately 120 mmHg, less than approximately 110 mmHg, less than approximately 100 mmHg, less than approximately 85 mmHg, less than approximately 90 mmHg, or less than approximately 80 mmHg. In one embodiment, a patient to be administered the LPA RNAi construct according to the method of the present invention has a mean systolic blood pressure at rest less than approximately 180 mmHg and a mean diastolic blood pressure less than approximately 110 mmHg prior to the first administration of the LPA RNAi construct. In another embodiment, a patient to be administered the LPA RNAi construct according to the method of the present invention has a mean systolic blood pressure at rest less than approximately 160 mmHg and a mean diastolic blood pressure less than approximately 100 mmHg prior to the first administration of the LPA RNAi construct. In yet another embodiment, a patient to be administered the LPA RNAi construct according to the method of the present invention has a mean systolic blood pressure at rest less than approximately 140 mmHg and a mean diastolic blood pressure less than approximately 90 mmHg prior to the first administration of the LPA RNAi construct.

[0052] In some embodiments, patients who are administered the LPA RNAi construct according to the method of the present invention do not show signs of severe renal failure. Therefore, in certain embodiments, patients who are administered the LPA RNAi construct according to the method of the present invention have a baseline eGFR (e.g., before the first administration of the LPA RNAi construct) of at least about 30 mL / min / 1.73m². 2 at least approximately 45 mL / min / 1.73 m 2 at least approximately 60 mL / min / 1.73 m 2, at least about 75 mL / min / 1.73 m 2 , or at least about 90 mL / min / 1.73 m 2 is. In certain embodiments, the patient to whom the LPA RNAi construct will be administered according to the method of the present invention has an eGFR of about 30 mL / min / 1.73 m 2 or more prior to the first administration of the LPA RNAi construct.

[0053] In other embodiments, the patient to whom the LPA RNAi construct will be administered according to the method of the present invention has no signs of active liver disease or liver dysfunction. Active liver disease can be determined by measuring one or more biomarkers of liver function, such as those included in a liver function test or liver panel, including albumin, alkaline phosphatase (ALP), alanine transaminase (ALT), aspartate aminotransferase (AST), gamma-glutamyl transpeptidase (GGT), bilirubin, and lactate dehydrogenase (LD). In certain embodiments, the patient to whom the LPA RNAi construct will be administered according to the method of the present invention has an ALT level at baseline (e.g., prior to the first administration of the LPA RNAi construct) that is no more than 3 times the upper limit of normal (ULN). In related embodiments, the patient to whom the LPA RNAi construct will be administered according to the method of the present invention has an ALT level at baseline (e.g., prior to the first administration of the LPA RNAi construct) that is no more than 3 times the ULN. In these and other embodiments, the patient to whom the LPA RNAi construct will be administered according to the method of the present invention has a total bilirubin level at baseline (e.g., prior to the first administration of the LPA RNAi construct) that is no more than 2 times the ULN. In some embodiments, the patient to whom the LPA RNAi construct will be administered according to the method of the present invention has, at baseline (e.g., prior to the first administration of the LPA RNAi construct): (i) a serum AST level of less than about 170 units / L, (ii) a serum AST level of less than about 150 units / L, and / or (iii) a total bilirubin level of less than about 2.0 mg / dL.

[0054] In one embodiment, the method of the present invention comprises administering an effective dose of an LPA RNAi construct to a patient. “Effective dose” means an amount sufficient to treat, alleviate or induce remission of a cardiovascular disease or one or more symptoms of a cardiovascular disease, in particular a condition or symptom associated with a cardiovascular disease, or otherwise prevent, hinder, delay or halt in any way the progression of a cardiovascular disease or any other undesirable symptom associated with a cardiovascular disease. Alternatively, an effective dose may mean an amount sufficient to reduce the occurrence or severity of sequelae resulting from a cardiovascular disease. For example, in some embodiments, an effective dose of an LPA RNAi construct is an amount sufficient to reduce the occurrence or severity of cardiovascular events in a patient with atherosclerosis or other cardiovascular disease, such as myocardial infarction, stroke, or vascular regeneration of the coronary, cerebral, or peripheral arteries.

[0055] In certain embodiments of the method of the present invention, the LPA RNAi construct is administered to the patient in a fixed dose. “Fixed dose” refers to a dose administered to all patients regardless of patient-specific factors, such as body weight. Therefore, the fixed dose is not adjusted on a patient-by-patient basis based on the patient's body weight. In some embodiments of the method of the present invention, the LPA RNAi construct may be administered to the patient in a fixed dose of approximately 9 mg to approximately 675 mg at dosing intervals of at least 8 weeks. For example, the fixed dose of the LPA RNAi construct may be approximately 9 mg, 10 mg, 15 mg, 30 mg, 50 mg, 60 mg, 70 mg, 75 mg, 80 mg, 90 mg, 100 mg, 125 mg, 150 mg, 175 mg, 200 mg, 225 mg, 250 mg, 300 mg, 350 mg, 400 mg, 450 mg, 500 mg, 550 mg, 600 mg, 650 mg, or 675 mg, and the dose is administered at intervals of at least 8 weeks. Furthermore, a range between any and all of these endpoints is intended, and for example, the fixed dose of the LPA RNAi construct administered to the patient in the method of the present invention may be about 10 mg to about 225 mg, about 50 mg to about 100 mg, about 150 mg to about 225 mg, about 225 mg to about 675 mg, about 75 mg to about 150 mg, about 225 mg to about 450 mg, about 75 mg to about 225 mg, about 10 mg to about 75 mg, or about 200 mg to about 300 mg, and the dose is administered at intervals of at least 8 weeks.

[0056] Any of the doses of the LPA RNAi constructs described herein is preferably administered at intervals of at least 8 weeks (i.e., the dose is not administered to the patient more frequently than once every 8 weeks (or once every 2 months)). For example, the administration interval may be about 8 weeks, about 12 weeks, about 16 weeks, about 20 weeks, about 24 weeks, about 28 weeks, or about 32 weeks. In certain embodiments, the administration interval is about 12 weeks, for example, a fixed dose of the LPA RNAi construct is administered to the patient once every 12 weeks (or once every 3 months). In certain other embodiments, the administration interval is about 24 weeks, for example, a fixed dose of the LPA RNAi construct is administered to the patient once every 24 weeks (or once every 6 months).

[0057] A fixed dose of the LPA RNAi construct may be administered at each dosing interval as a single bolus (e.g., by a single subcutaneous injection) or as two or more consecutive bolus injections (e.g., two or more subcutaneous injections). In some embodiments, the entire fixed dose of the LPA RNAi construct is administered to the patient at each dosing interval by a single bolus injection, for example, using a pre-filled syringe or injection device as further described herein. For example, a fixed dose of 225 mg of the LPA RNAi construct may be administered to the patient at each dosing interval (e.g., once every 12 weeks) as a single bolus injection of 225 mg, possibly using an autoinjector, pen injector, or pre-filled syringe containing a 225 mg dose. In other embodiments, the entire fixed dose of the LPA RNAi construct is administered to the patient as two or more consecutive bolus injections. As an example, a fixed dose of 225 mg of the LPA RNAi construct may be administered to the patient at each dosing interval (e.g., once every 12 weeks) by three consecutive injections of 75 mg each, and possibly by three injection devices (e.g., autoinjectors, pen injectors, or pre-filled syringes) each containing a 75 mg dose. Consecutive injections given within a single day are considered a single dose within the context of the present invention. In other words, as an example, a fixed dose of 225 mg administered once every 12 weeks may be administered to the patient as a single bolus injection of 225 mg once every 12 weeks, or as three consecutive bolus injections of 75 mg each administered to the patient within a single day, once every 12 weeks.

[0058] In certain embodiments of the method of the present invention, the fixed dose of the LPA RNAi construct described herein is administered once every 12 weeks or once every 3 months. In some such embodiments, the method of the present invention includes administering the LPA RNAi construct to a patient at a fixed dose of about 10 mg to about 225 mg once every 12 weeks or once every 3 months. In other embodiments, the method of the present invention includes administering the LPA RNAi construct to a patient at a fixed dose of about 50 mg to about 100 mg once every 12 weeks or once every 3 months. In yet another embodiment, the method of the present invention includes administering the LPA RNAi construct to a patient at a fixed dose of about 75 mg to about 225 mg once every 12 weeks or once every 3 months. In yet another embodiment, the method of the present invention includes administering the LPA RNAi construct to a patient at a fixed dose of about 150 mg to about 225 mg once every 12 weeks or once every 3 months. In one embodiment, the method of the present invention includes administering the LPA RNAi construct to a patient at a fixed dose of approximately 10 mg once every 12 weeks or once every 3 months. In another embodiment, the method of the present invention includes administering the LPA RNAi construct to a patient at a fixed dose of approximately 30 mg once every 12 weeks or once every 3 months. In yet another embodiment, the method of the present invention includes administering the LPA RNAi construct to a patient at a fixed dose of approximately 75 mg once every 12 weeks or once every 3 months. In yet another embodiment, the method of the present invention includes administering the LPA RNAi construct to a patient at a fixed dose of approximately 100 mg once every 12 weeks or once every 3 months. In yet another embodiment, the method of the present invention includes administering the LPA RNAi construct to a patient at a fixed dose of approximately 125 mg once every 12 weeks or once every 3 months. In another embodiment, the method of the present invention includes administering the LPA RNAi construct to a patient at a fixed dose of approximately 175 mg once every 12 weeks or once every 3 months. In another embodiment, the method of the present invention includes administering the LPA RNAi construct to a patient at a fixed dose of approximately 200 mg once every 12 weeks or once every 3 months.In yet another embodiment, the method of the present invention includes administering the LPA RNAi construct to a patient at a fixed dose of approximately 225 mg once every 12 weeks or once every 3 months.

[0059] In certain other embodiments of the method of the present invention, the fixed dose of the LPA RNAi construct described herein is administered once every 24 weeks or once every 6 months. In some such embodiments, the method of the present invention includes administering the LPA RNAi construct to a patient at a fixed dose of about 225 mg to about 675 mg once every 24 weeks or once every 6 months. In other embodiments, the method of the present invention includes administering the LPA RNAi construct to a patient at a fixed dose of about 225 mg to about 450 mg once every 24 weeks or once every 6 months. In yet another embodiment, the method of the present invention includes administering the LPA RNAi construct to a patient at a fixed dose of about 200 mg to about 300 mg once every 24 weeks or once every 6 months. In one embodiment, the method of the present invention includes administering the LPA RNAi construct to a patient at a fixed dose of about 225 mg once every 24 weeks or once every 6 months. In another embodiment, the method of the present invention includes administering the LPA RNAi construct to the patient at a fixed dose of approximately 300 mg once every 24 weeks or once every 6 months. In yet another embodiment, the method of the present invention includes administering the LPA RNAi construct to the patient at a fixed dose of approximately 450 mg once every 24 weeks or once every 6 months. In yet another embodiment, the method of the present invention includes administering the LPA RNAi construct to the patient at a fixed dose of approximately 675 mg once every 24 weeks or once every 6 months.

[0060] In some embodiments of the method of the present invention, the LPA RNAi construct is administered to the patient over a set of treatment periods. The “treatment period” begins with the administration of the first dose of the LPA RNAi construct and ends with the administration of the final dose of the LPA RNAi construct. Treatment periods are approximately 12 to 240 weeks, approximately 24 to 144 weeks, approximately 3 months to 60 months, approximately 6 months to 48 months, for example, approximately 12 weeks, approximately 24 weeks, approximately 36 weeks, approximately 48 weeks, approximately 60 weeks, approximately 72 weeks, approximately 84 weeks, approximately 96 weeks, approximately 108 weeks, approximately 120 weeks, approximately 132 weeks, approximately 144 weeks, approximately 156 weeks, approximately 168 weeks, approximately 180 weeks, approximately 192 weeks, approximately 20 The treatment period may be 4 weeks, approximately 216 weeks, approximately 228 weeks, approximately 240 weeks, approximately 3 months, approximately 6 months, approximately 9 months, approximately 12 months, approximately 15 months, approximately 18 months, approximately 21 months, approximately 24 months, approximately 27 months, approximately 30 months, approximately 33 months, approximately 36 months, approximately 39 months, approximately 42 months, approximately 45 months, approximately 48 months, approximately 51 months, approximately 54 months, approximately 57 months, or approximately 60 months. In some embodiments, the treatment period is approximately 48 weeks. In other embodiments, the treatment period is approximately 192 weeks. In yet another embodiment, the treatment period is approximately 12 months. In yet another embodiment, the treatment period is approximately 48 months. In certain embodiments, the treatment period may be longer than 240 weeks or 60 months; for example, the treatment period may be more than 5 years, for example, 6, 7, 8, 9, or 10 years or more. In one specific embodiment, the LPA RNAi construct is administered for a treatment period of at least about 36 weeks, resulting in a statistically significant percentage reduction in serum or plasma Lp(a) levels from baseline compared to subjects who do not receive the LPA RNAi construct. In another specific embodiment, the LPA RNAi construct is administered for a treatment period of at least about 48 weeks, resulting in a statistically significant percentage reduction in serum or plasma Lp(a) levels from baseline compared to subjects who do not receive the LPA RNAi construct.

[0061] The methods described herein involve administering an LPA RNAi construct to a patient. As used herein, the term “LPA RNAi construct” refers to an agent comprising an RNA molecule that, when introduced into cells, can downregulate the expression of the LPA gene via an RNA interference mechanism. RNA interference is a process in which a nucleic acid molecule induces sequence-specific cleavage and degradation of a target RNA molecule (e.g., messenger RNA or mRNA molecule), for example, via the RNA-induced silencing complex (RISC) pathway. In some embodiments, the LPA RNAi construct comprises a double-stranded RNA molecule containing two antiparallel strands of consecutive nucleotides that are sufficiently complementary to each other to hybridize to form a double-stranded region. “Hybridizing” typically refers to the pairing of complementary polynucleotides via hydrogen bonds between complementary bases in two polynucleotides (e.g., Watson-Crick, Hoogsteen, or reverse Hoogsteen hydrogen bonds). A strand containing a region having a sequence substantially complementary to the target LPA sequence (e.g., target LPA mRNA) is referred to as the “antisense strand.” The “sense strand” refers to a strand containing a region substantially complementary to a region of the antisense strand. In some embodiments, the sense strand may contain a region having a sequence substantially identical to the target sequence.

[0062] Double-stranded RNA molecules may include chemical modifications to ribonucleotides, such as modifications to ribose sugars, bases, or other skeletal components of ribonucleotides, as described herein or known in the art. Any modifications used on double-stranded RNA molecules (e.g., siRNA, shRNA, etc.) are encompassed by the term “double-stranded RNA” for the purposes of this disclosure.

[0063] As used herein, a first sequence is "complementary" to a second sequence if, under certain conditions such as physiological conditions, a polynucleotide containing the first sequence can hybridize to a polynucleotide containing the second sequence to form a double-stranded region. Other such conditions may include moderate or stringent hybridization conditions known to those skilled in the art. A first sequence is considered 100% complementary to a second sequence if a polynucleotide containing the first sequence forms base pairs with a polynucleotide containing the second sequence without any mismatches over the entire length of one or both nucleotide sequences. A sequence is "substantially complementary" to a target sequence if it is at least about 80%, 85%, 90%, 95%, 96%, 97%, 98%, or 99% complementary to the target sequence. The complementarity percentage can be calculated by dividing the number of bases in the first sequence that are complementary to the bases at corresponding positions in the second sequence or target sequence by the total length of the first sequence. Furthermore, if two sequences hybridize, and there are 5, 4, 3, or 2 or fewer mismatches across the entire 30-base-pair double-stranded region, one sequence can be said to be substantially complementary to the other. Generally, if any nucleotide overhangs exist as defined herein, the sequence of such overhangs is not considered when determining the degree of complementarity between the two sequences. For example, a 21-nucleotide sense strand and a 21-nucleotide antisense strand that hybridize to form a 19-base-pair double-stranded region with a 2-nucleotide overhang at the 3' end of each strand would be considered perfectly complementary when the term is used herein.

[0064] In some embodiments, a region of the antisense strand contains a sequence that is substantially or completely complementary to the region of the target LPA RNA sequence (e.g., LPA mRNA). In such embodiments, the sense strand may contain a sequence that is completely complementary to the sequence of the antisense strand. In other such embodiments, the sense strand may contain a sequence that is substantially complementary to the sequence of the antisense strand, for example, a sequence having 1, 2, 3, 4, or 5 mismatches in the double-stranded region formed by the sense and antisense strands. In certain embodiments, it is preferable that any mismatches occur within the terminal region (e.g., within 6, 5, 4, 3, or 2 nucleotides of the 5' and / or 3' ends of the strand). In one embodiment, any mismatches in the double-stranded region formed by the sense and antisense strands occur within 6, 5, 4, 3, or 2 nucleotides of the 5' end of the antisense strand.

[0065] In certain embodiments, the sense and antisense strands of a double-stranded RNA may be two separate molecules that hybridize to form a double-stranded region, but otherwise remain separate. Such a double-stranded RNA molecule formed from two separate strands is referred to as "small interfering RNA" or "short interfering RNA" (siRNA). Therefore, in some embodiments, the LPA RNAi construct used in the method of the present invention includes siRNA.

[0066] In other embodiments, the sense and antisense strands that hybridize to form a double-stranded region may be part of a single RNA molecule, i.e., the sense and antisense strands are part of the self-complementary region of the single RNA molecule. In such cases, the single RNA molecule includes a double-stranded region (also called a stem region) and a loop region. The 3' end of the sense strand is ligated to the 5' end of the antisense strand by an adjacent sequence of unpaired nucleotides, thereby forming a loop region. The loop region is typically long enough to allow the RNA molecule itself to refold so that the antisense strand can base-pair with the sense strand to form a double-stranded or stem region. The loop region may contain about 3 to about 25, about 5 to about 15, or about 8 to about 12 unpaired nucleotides. Such RNA molecules having at least partially self-complementary regions are referred to as "small hairpin RNA" (shRNA). In certain embodiments, the LPA RNAi construct used in the method of the present invention includes shRNA. The length of a single, at least partially self-complementary RNA molecule may be approximately 40 to 100 nucleotides, approximately 45 to 85 nucleotides, or approximately 50 to 60 nucleotides, and may include double-stranded regions and loop regions having the lengths described herein, respectively.

[0067] The LPA RNAi construct used in the method of the present invention comprises a sense strand and an antisense strand, the antisense strand comprising a region having a sequence substantially or completely complementary to the LPA messenger RNA (mRNA) sequence. As used herein, “LPA mRNA sequence” refers to a messenger RNA sequence that includes allelic variants and splice variants encoding the apo(a) protein, including variants or isoforms of the apo(a) protein from any species (e.g., non-human primates, humans). The LPA gene (also known as AK38, APOA, and LP) encodes the apo(a) protein, which is the main component of low-density lipoprotein particles known as lipoprotein(a) or Lp(a). In humans, the LPA gene is found on chromosome 6 at locus 6q25.3–q26. The LPA gene exhibits a high degree of polymorphism due to differences in the number of copies of the Kringle IV2 (KIV-2) domain between individuals, which can range from 2 to over 40 copies (see, for example, Kronenberg and Utermann, J.Intern.Med., Vol.273:6-30, 2013).

[0068] Furthermore, an LPA mRNA sequence can be a transcript sequence represented as its complementary DNA (cDNA) sequence. A cDNA sequence refers to an mRNA transcript sequence represented as DNA bases (e.g., guanine, adenine, thymine, and cytosine) rather than RNA bases (e.g., guanine, adenine, uracil, and cytosine). Therefore, the antisense strand of the LPA RNAi construct used in the method of the present invention may include a region having a sequence that is substantially or completely complementary to the target LPA mRNA sequence or LPA cDNA sequence. Any LPA mRNA or cDNA sequence selected from the NCBI reference sequences NM_005577.4 (human), XM_015448520.1 (cynomolgus monkey), XM_028847001.1 (rhesus monkey), XM_024357489.1 (chimpanzee), and XM_031012244.1 (gorilla) may or may not be used as the LPA mRNA or cDNA sequence. In certain embodiments, the LPA mRNA sequence is a human transcript listed in the NCBI database as the reference sequence NM_005577.4.

[0069] The sense strand of an LPA RNAi construct typically contains a sequence sufficiently complementary to the antisense strand sequence so that, under physiological conditions, the two strands hybridize to form a double-stranded region. A “double-stranded region” refers to a region within two complementary or substantially complementary polynucleotides that form a double helix between them by base pairing, either through Watson-Crick base pairing or other hydrogen bonding interactions. The double-stranded region of an LPA RNAi construct should be long enough to allow the LPA RNAi construct to enter the RNA interference pathway, for example, by binding to the Dicer enzyme and / or RISC complex. For example, in some embodiments, the double-stranded region is about 15 to about 30 base pairs long. Furthermore, other lengths of the double-stranded region within this range are suitable, for example, about 15 to about 28 base pairs, about 15 to about 26 base pairs, about 15 to about 24 base pairs, about 15 to about 22 base pairs, about 17 to about 28 base pairs, about 17 to about 26 base pairs, about 17 to about 24 base pairs, about 17 to about 23 base pairs, about 17 to about 21 base pairs, about 19 to about 25 base pairs, about 19 to about 23 base pairs, or about 19 to about 21 base pairs. In a particular embodiment, the double-stranded region is about 17 to about 26 base pairs long. In another embodiment, the double-stranded region is about 19 to about 21 base pairs long. In one embodiment, the double-stranded region is about 19 base pairs long. In yet another embodiment, the double-stranded region is about 21 base pairs long.

[0070] In embodiments where the sense and antisense strands are two separate molecules (e.g., the LPA RNAi construct includes siRNA), the sense and antisense strands do not need to be the same length as the double-stranded region. For example, one or both strands may be longer than the double-stranded region and have one or more unpaired nucleotides or mismatches located on the sides of the double-stranded region. Thus, in some embodiments, the RNAi construct includes at least one nucleotide overhang. As used herein, “nucleotide overhang” refers to an unpaired nucleotide or nucleotide that extends beyond the double-stranded region at the end of a strand. Nucleotide overhangs are typically created when the 3' end of one strand extends beyond the 5' end of the other strand, or when the 5' end of one strand extends beyond the 3' end of the other strand. Nucleotide overhangs are generally 1–6 nucleotides, 1–5 nucleotides, 1–4 nucleotides, 1–3 nucleotides, 2–6 nucleotides, 2–5 nucleotides, or 2–4 nucleotides. In some embodiments, the nucleotide overhang contains 1, 2, 3, 4, 5, or 6 nucleotides. In a particular embodiment, the nucleotide overhang contains 1 to 4 nucleotides. In a particular embodiment, the nucleotide overhang contains 2 nucleotides. In a particular other embodiment, the nucleotide overhang contains a single nucleotide. If the nucleotide overhang is located within the antisense strand, the nucleotides within the overhang may be complementary to the target gene sequence, may form a mismatch with the target gene sequence, or may contain any other sequence (e.g., polypyrimidine sequences or polypurine sequences, e.g., UU, TT, AA, GG, etc.).

[0071] Nucleotide overhangs may be present at the 5' or 3' ends of one or both strands. For example, in one embodiment, the LPA RNAi construct includes nucleotide overhangs at the 5' and 3' ends of the antisense strand. In another embodiment, LPA The RNAi construct includes nucleotide overhangs at the 5' and 3' ends of the sense strand. In some embodiments, the LPA RNAi construct includes nucleotide overhangs at the 5' end of the sense strand and the 5' end of the antisense strand. In other embodiments, the LPA RNAi construct includes nucleotide overhangs at the 3' end of the sense strand and the 3' end of the antisense strand.

[0072] RNAi constructs may contain a nucleotide overhang at one end of a double-stranded RNA molecule and a blunt end at the other. “Blunt end” means that the sense and antisense strands are fully base-paired at the ends of the molecule and there are no unpaired nucleotides extending beyond the double-stranded region. In some embodiments, the LPA RNAi construct contains a nucleotide overhang at the 3' end of the sense strand and blunt ends at the 5' end of the sense strand and the 3' end of the antisense strand. In other embodiments, the LPA RNAi construct contains a nucleotide overhang at the 3' end of the antisense strand and blunt ends at the 5' end of the antisense strand and the 3' end of the sense strand. In certain embodiments, the LPA RNAi construct contains blunt ends at both ends of the double-stranded RNA molecule. In such embodiments, the sense and antisense strands are of equal length, and the double-stranded region is of equal length to the sense and antisense strands (i.e., the molecule is double-stranded over its entire length).

[0073] The sense strand and antisense strand in the LPA RNAi construct used in the method of the present invention may each independently be about 15 to about 30 nucleotides long, about 19 to about 30 nucleotides long, about 18 to about 28 nucleotides long, about 19 to about 27 nucleotides long, about 19 to about 25 nucleotides long, about 19 to about 23 nucleotides long, about 19 to about 21 nucleotides long, about 21 to about 25 nucleotides long, or about 21 to about 23 nucleotides long. In certain embodiments, the sense strand and antisense strand are each independently about 18, about 19, about 20, about 21, about 22, about 23, about 24, or about 25 nucleotides long. In some embodiments, the sense strand and antisense strand form a double-stranded region that is the same length but shorter than these strands, such that the LPA RNAi construct has two nucleotide overhangs. For example, in one embodiment, the LPA RNAi construct includes (i) a sense strand and an antisense strand, each 21 nucleotides long, (ii) a 19-base-pair-long double-stranded region, and (iii) nucleotide overhangs of two unpaired nucleotides at both the 3' end of the sense strand and the 3' end of the antisense strand. In another embodiment, the LPA RNAi construct includes (i) a sense strand and an antisense strand, each 23 nucleotides long, (ii) a 21-base-pair-long double-stranded region, and (iii) nucleotide overhangs of two unpaired nucleotides at both the 3' end of the sense strand and the 3' end of the antisense strand. In yet another embodiment, the sense strand and antisense strand have the same length and form a double-stranded region such that there are no nucleotide overhangs at either end of the double-stranded molecule along their entire length. In a particular embodiment, the LPA RNAi construct used in the method of the present invention is blunt-ended and includes (i) a sense strand and an antisense strand, each 21 nucleotides long, and (ii) a 21-base-pair-long double-stranded region. In another specific embodiment, the LPA RNAi construct used in the method of the present invention is blunt-ended and comprises (i) a sense strand and an antisense strand, each 19 nucleotides long, and (ii) a double-stranded region 19 base pairs long.

[0074] In other embodiments, the sense strand or antisense strand is longer than the other strand, and the two strands form a double-stranded region having a length equal to the length of the shorter strand, such that the LPA RNAi construct includes at least one nucleotide overhang. For example, in one embodiment, the LPA RNAi construct used in the method of the present invention includes (i) a 19-nucleotide sense strand, (ii) a 21-nucleotide antisense strand, (iii) a 19-base-pair-length double-stranded region, and (iv) a nucleotide overhang of two unpaired nucleotides at the 3' end of the antisense strand. In another embodiment, the LPA RNAi construct used in the method of the present invention includes (i) a 21-nucleotide sense strand, (ii) a 23-nucleotide antisense strand, (iii) a 21-base-pair-length double-stranded region, and (iv) a nucleotide overhang of two unpaired nucleotides at the 3' end of the antisense strand.

[0075] The LPA RNAi construct used in the method of the present invention may contain one or more modified nucleotides. “Modified nucleotide” refers to a nucleotide having one or more chemical modifications to a nucleoside, nucleic acid base, pentose ring, or phosphate group. As used herein, modified nucleotides do not include ribonucleotides containing adenosine monophosphate, guanosine monophosphate, uridine monophosphate, and cytidine monophosphate. However, the LPA RNAi construct may include combinations of modified nucleotides and ribonucleotides. Incorporation of modified nucleotides into one or both strands of a double-stranded RNA molecule can improve the in vivo stability of the RNA molecule, for example, by reducing the molecule's sensitivity to nucleases and other degradation processes. Furthermore, the effectiveness of the LPA RNAi construct in reducing LPA gene expression can be enhanced by the incorporation of modified nucleotides.

[0076] In certain embodiments, the modified nucleotide has a ribose sugar modification. Such sugar modifications may include modifications at the 2' and / or 5' positions of the pentose ring, as well as bicyclic sugar modifications. A 2'-modified nucleotide refers to a nucleotide having a pentose ring with a substituent other than OH at the 2' position. Such 2' modifications include 2'H (e.g., deoxyribonucleotide), 2'-O-alkyl (e.g., O-C1~C1), and 10 Or O-C1~C 10 Examples of substituted alkyl groups include, but are not limited to, 2'-O-allyl (O-CH2CH=CH2), 2'-C-allyl, 2'-deoxy-2'-fluoro (also called 2'-F or 2'-fluoro), 2'-O-methyl (-OCH3), 2'-O-methoxyethyl (O-(CH2)2OCH3), 2'-OCF3, 2'-O(CH2)2SCH3, 2'-O-aminoalkyl, 2'-amino (e.g., NH2), 2'-O-ethylamine, and 2'-azide. Modifications at the 5' position of the pentose ring include, but are not limited to, 5'-methyl (R or S); 5'-vinyl, and 5'-methoxy.

[0077] "Bicyclic sugar modification" refers to the modification of a pentose ring in which two atoms of the ring are linked by a bridge to form a second ring, thereby creating a bicyclic sugar structure. In some embodiments, the bicyclic sugar modification includes a bridge between the 4' and 2' carbon atoms of the pentose ring. A nucleotide containing a sugar moiety having a bicyclic sugar modification is referred to herein as a bicyclic nucleic acid or BNA. Examples of bicyclic sugar modifications include α-L-methyleneoxy(4'-CH2-O-2') bicyclic nucleic acid (BNA); β-D-methyleneoxy(4'-CH2-O-2')BNA (also called locked nucleic acid or LNA); ethyleneoxy(4'-(CH2)2-O-2')BNA; aminooxy(4'-CH2-ON(R)-2')BNA; oxyamino(4'-CH2-N(R)-O-2')BNA; methyl(methyleneoxy)(4'-CH(CH3)-O-2')BNA (constructor Examples include, but are not limited to, constrained ethyl (also known as cEt); methylene-thio(4'-CH2-S-2')BNA; methylene-amino(4'-CH2-N(R)-2')BNA; methyl carbon ring(4'-CH2-CH(CH3)-2')BNA; propylene carbon ring(4'-(CH2)3-2')BNA; and methoxy(ethyleneoxy)(4'-CH(CH2OMe)-O-2')BNA (also known as constrained MOE or cMOE). These and other glycosylated nucleotides that can be incorporated into the LPA RNAi construct used in the methods of the present invention are described in U.S. Patent No. 9,181,551, U.S. Patent Application Publication No. 2016 / 0122761, and Deleavey and Damha, Chemistry and Biology, Vol. 19:937-954, 2012.

[0078] In some embodiments, the LPA RNAi construct comprises one or more 2'-fluoro-modified nucleotides, 2'-O-methyl-modified nucleotides, 2'-O-methoxyethyl-modified nucleotides, 2'-O-alkyl-modified nucleotides, 2'-O-allyl-modified nucleotides, bicyclic nucleic acids (BNAs), deoxyribonucleotides, or combinations thereof. In certain embodiments, the LPA RNAi construct comprises one or more 2'-fluoro-modified nucleotides, 2'-O-methyl-modified nucleotides, 2'-O-methoxyethyl-modified nucleotides, deoxyribonucleotides, or combinations thereof. In a particular embodiment, the LPA RNAi construct used in the method of the present invention comprises one or more 2'-fluoro-modified nucleotides, 2'-O-methyl-modified nucleotides, deoxyribonucleotides, or combinations thereof. In some such embodiments, the deoxyribonucleotide may be a terminal nucleotide at the 3' and / or 5' ends of the sense strand or antisense strand. In embodiments where the deoxyribonucleotide is the terminal nucleotide, the terminal nucleotide may be an inverted nucleotide, i.e., it may be linked to an adjacent nucleotide via a 3'-3' nucleotide bond (if it is at the 3' end of the chain) or a 5'-5' nucleotide bond (if it is at the 5' end of the chain), rather than via a natural 3'-5' nucleotide bond.

[0079] Both the sense and antisense strands of the LPA RNAi construct may contain one or more modified nucleotides. For example, in some embodiments, the sense strand contains one, two, three, four, five, six, seven, eight, nine, or more modified nucleotides. In certain embodiments, all nucleotides in the sense strand are modified nucleotides. In some embodiments, the antisense strand contains one, two, three, four, five, six, seven, eight, nine, or more modified nucleotides. In other embodiments, all nucleotides in the antisense strand are modified nucleotides. In certain other embodiments, all nucleotides in the sense strand and all nucleotides in the antisense strand are modified nucleotides. In these and other embodiments, the modified nucleotides may be 2'-fluoromodified nucleotides, 2'-O-methylmodified nucleotides, or a combination thereof.

[0080] In certain embodiments, the modified nucleotides incorporated into one or both strands of the LPA RNAi construct used in the method of the present invention have modifications to nucleic acid bases (also referred to herein as “bases”). “Modified nucleic acid bases” or “modified bases” refers to bases other than the naturally occurring purine bases adenine (A) and guanine (G), and the pyrimidine bases thymine (T), cytosine (C), and uracil (U). Modified nucleic acid bases can be synthetic or naturally occurring modifications, and include universal bases, 5-methylcytosine (5-me-C), 5-hydroxymethylcytosine, xanthine (X), hypoxanthine (I), 2-aminoadenine, 6-methyladenine, 6-methylguanine, and other alkyl derivatives of adenine and guanine, 2-propyl and other alkyl derivatives of adenine and guanine, 2-thiouracil, 2-thiothymine, and 2-thiocytosine, 5-halouracil and cytosine, 5-propynyluracil and cytosine, and 6-azoura. Examples include, but are not limited to, syl, cytosine, and thymine, 5-uracil (pseudouracil), 4-thiouracil, 8-halo, 8-amino, 8-thiol, 8-thioalkyl, 8-hydroxyl, and other 8-substituted adenines and guanines, 5-halo, especially 5-bromo, 5-trifluoromethyl, and other 5-substituted uracils and cytosines, 7-methylguanine and 7-methyladenine, 8-azaguanine and 8-azaadenine, 7-deazaguanine and 7-deazaadenine, and 3-deazaguanine and 3-deazaadenine.

[0081] In some embodiments, the modified base is a universal base. A “universal base” refers to a base analog that indiscriminately forms base pairs with all native bases in RNA and DNA without altering the double helix structure of the resulting double-stranded region. Universal bases are known to those skilled in the art and include, but are not limited to, inosine, C-phenyl, C-naphthyl, and other aromatic derivatives, azole carboxamides, and nitroazole derivatives such as 3-nitropyrrole, 4-nitroindole, 5-nitroindole, and 6-nitroindole.

[0082] Other suitable modified bases that can be incorporated into LPA RNAi constructs are those described in Herdewijn, Antisense Nucleic Acid Drug Dev., Vol.10:297-310, 2000 and Peacock et al., J.Org.Chem., Vol.76:7295-7300, 2011. Those skilled in the art are well aware that guanine, cytosine, adenine, thymine, and uracil can be substituted with other nucleic acid bases, such as the modified nucleic acid bases mentioned above (without substantially altering the base-pairing properties of polynucleotides containing such substituted nucleic acid bases).

[0083] In some embodiments, the sense and antisense strands of the LPA RNAi construct used in the method of the present invention may contain one or more debasalized nucleotides. A “debasalized nucleotide” or “debasalized nucleoside” is a nucleotide or nucleoside lacking a nucleic acid base at the 1' position of a ribose sugar. In certain embodiments, the debasalized nucleotide is incorporated at the ends of the sense and / or antisense strands of the RNAi construct. In one embodiment, the sense strand contains a debasalized nucleotide as a terminal nucleotide at its 3' end, its 5' end, or both its 3' and 5' ends. In another embodiment, the antisense strand contains a debasalized nucleotide as a terminal nucleotide at its 3' end, its 5' end, or both its 3' and 5' ends. In embodiments where the debasalized nucleotide is a terminal nucleotide, the terminal nucleotide may be an inverted nucleotide, i.e., it may be linked to an adjacent nucleotide via a 3'-3' nucleotide bond (if it is at the 3' end of the chain) or a 5'-5' nucleotide bond (if it is at the 5' end of the chain), rather than a natural 3'-5' nucleotide bond. The debasalized nucleotide may also contain sugar modifications, such as any of the sugar modifications described above. In certain embodiments, the debasalized nucleotide may contain a 2' modification, such as a 2'-fluoro modification, a 2'-O-methyl modification, or a 2'-H (deoxy) modification. In one embodiment, the debasalized nucleotide contains a 2'-O-methyl modification. In another embodiment, the debasalized nucleotide contains a 2'-H modification (i.e., a deoxy-debasalized nucleotide).

[0084] Furthermore, the LPA RNAi construct used in the method of the present invention may contain one or more modified nucleotide bonds. As used herein, the term "modified nucleotide bond" refers to nucleotide bonds other than the natural 3'-5' phosphodiester bond. In some embodiments, the modified nucleotide bond is a phosphorus-containing nucleotide bond, such as phosphotriesters, aminoalkyl phosphotriesters, alkylphosphonates (e.g., methylphosphonate, 3'-alkylenephosphonate), phosphinates, phosphoramidates (e.g., 3'-aminophosphoramidate and aminoalkylphosphoramidate), phosphorothioate (P=S), chiral phosphorothioate, phosphorodithioate, thionophosphoramidate, thionoalkylphosphonate, thionoalkylphosphotriester, and boranophosphate. In one embodiment, the modified nucleotide bond is a 2'-5' phosphodiester bond. In other embodiments, the modified nucleotide bond is a phosphorus-free nucleotide bond and may be referred to as a modified nucleoside bond. Examples of phosphorus-free bonds include, but are not limited to, morpholino bonds (partially formed from the sugar portion of a nucleoside); siloxane bonds (-O-Si(H)2-O-); sulfide, sulfoxide, and sulfone bonds; formacetyl and thioformacetyl bonds; alkene-containing skeletons; sulfamate skeletons; methylenemethylimino (-CH2-N(CH3)-O-CH2-) and methylenehydrazino bonds; sulfonate and sulfonamide bonds; amide bonds; and other mixtures of N, O, S, and CH2 constituent parts. In one embodiment, the modified nucleoside bond is a peptide-based bond (e.g., aminoethylglycine) for creating peptide nucleic acids or PNAs, such as those described in U.S. Patent No. 5,539,082, U.S. Patent No. 5,714,331, and U.S. Patent No. 5,719,262.Other suitable modified nucleotide-nucleotide and nucleoside-nucleotide bonds that may be used in LPA RNAi constructs are described in U.S. Patent No. 6,693,187, U.S. Patent No. 9,181,551, U.S. Patent Application Publication No. 2016 / 0122761, and Deleavey and Damha, Chemistry and Biology, Vol. 19:937-954, 2012.

[0085] In certain embodiments, the LPA RNAi construct used in the method of the present invention includes one or more phosphorothioate nucleotide interbonds. These phosphorothioate nucleotide interbonds may be present in the sense strand, antisense strand, or both strands of the LPA RNAi construct. For example, in some embodiments, the sense strand includes one, two, three, four, five, six, seven, eight or more phosphorothioate nucleotide interbonds. In other embodiments, the antisense strand includes one, two, three, four, five, six, seven, eight or more phosphorothioate nucleotide interbonds. In yet another embodiment, both strands include one, two, three, four, five, six, seven, eight or more phosphorothioate nucleotide interbonds. The LPA RNAi construct may include one or more phosphorothioate nucleotide interbonds at the 3' end, 5' end, or both the 3' and 5' ends of the sense strand, antisense strand, or both strands. For example, in certain embodiments, the LPA RNAi construct contains about 1 to about 6 or more (e.g., about 1, 2, 3, 4, 5, or 6 or more) consecutive phosphorothioate internucleotide bonds at the 3' ends of the sense strand, antisense strand, or both strands. In other embodiments, the LPA RNAi construct contains about 1 to about 6 or more (e.g., about 1, 2, 3, 4, 5, or 6 or more) consecutive phosphorothioate internucleotide bonds at the 5' ends of the sense strand, antisense strand, or both strands. In any embodiment in which one or both strands contain one or more phosphorothioate internucleotide bonds, the remaining internucleotide bonds in the strands may be native 3'-5' phosphodiester bonds. For example, in some embodiments, each internucleotide bond in the sense strand and antisense strand is selected from phosphodiesters and phosphorothioate, and at least one internucleotide bond is a phosphorothioate.

[0086] In some embodiments of the RNAi construct of the present invention, the 5' ends of the sense strand, antisense strand, or both the antisense strand and the sense strand include a phosphate moiety. As used herein, the term "phosphate moiety" refers to a terminal phosphate group including an unmodified phosphate (-OP=O)(OH)OH) and a modified phosphate. Examples of modified phosphates include phosphates in which one or more of the O and OH groups are substituted with H, O, S, N(R), or alkyl (where R is H, an amino protecting group, or an unsubstituted or substituted alkyl). Examples of phosphate moieties include, but are not limited to, 5'-monophosphates; 5'-diphosphates; 5'-triphosphates; 5'-guanosine caps (7-methylated or unmethylated); 5'-adenosine caps; or any other modified or unmodified nucleotide cap structures; 5'-monothiophosphates (phosphorothioate); 5'-monodithiophosphates (phosphorodithioate); 5'-alpha-thiotriphosphates; 5'-gamma-thiotriphosphates; 5'-phosphoamidates; 5'-vinyl phosphates; 5'-alkylphosphonates (e.g., alkyl=methyl, ethyl, isopropyl, propyl, etc.); 5'-cyclopropylphosphonates; and 5'-alkyletherphosphonates (e.g., alkylether=methoxymethyl, ethoxymethyl, etc.).

[0087] Modified nucleotides that can be incorporated into LPA RNAi constructs suitable for use in the methods of the present invention may have a number of chemical modifications as described herein. For example, a modified nucleotide may have modifications to a ribose sugar and modifications to a nucleic acid base. For example, a modified nucleotide may include a 2' sugar modification (e.g., 2'-fluoro or 2'-O-methyl) and a modified base (e.g., 5-methylcytosine or pseudouracil). In other embodiments, a modified nucleotide may include a sugar modification combined with a 5' phosphate modification that, when incorporated into a polynucleotide, results in inter-modified nucleotide or internucleoside bonding. For example, in some embodiments, a modified nucleotide may include a 2'-fluoro modification, a 2'-O-methyl modification, or a bicyclic sugar modification and a sugar modification such as a 5' phosphorothioate group. Thus, in some embodiments, one or both strands of the RNAi construct of the present invention may include a combination of a 2' modified nucleotide or BNA and a phosphorothioate nucleotide bond. In certain embodiments, both the sense strand and antisense strand of the RNAi construct of the present invention include a combination of 2'-fluoromodified nucleotides, 2'-O-methylmodified nucleotides, and phosphorothioate nucleotide internucleotide bonds.

[0088] The LPA gene is primarily expressed in the liver. Therefore, in certain embodiments, LPA It is desirable to deliver the RNAi construct specifically to liver cells. Therefore, in some embodiments, the LPA RNAi construct used in the method of the present invention may include a targeting moiety that directs the LPA RNAi construct specifically to liver cells (e.g., hepatocytes) using various approaches, as will be described in more detail below. In certain embodiments, the LPA RNAi construct includes a targeting moiety containing a ligand that binds to a surface-expressed asialoglycoprotein receptor (ASGR) or its components (e.g., ASGR1, ASGR2).

[0089] In some embodiments, the LPA RNAi construct can be specifically targeted to the liver by using ligands that bind to or interact with proteins expressed on the surface of hepatocytes. For example, in certain embodiments, the ligand may include an antigen-binding protein (e.g., an antibody or its binding fragment (e.g., Fab, scFv)) that specifically binds to receptors expressed on hepatocytes, such as the asialoglycoprotein receptor and the LDL receptor. In one particular embodiment, the ligand includes an antibody or its binding fragment that specifically binds to ASGR1 and / or ASGR2. In another embodiment, the ligand includes a Fab fragment of an antibody that specifically binds to ASGR1 and / or ASGR2. The "Fab fragment" consists of one immunoglobulin light chain (i.e., a light chain variable region (VL) and a constant region (CL)) and one immunoglobulin heavy chain (CH1 region and variable region (VH)). In another embodiment, the ligand includes a single-chain variable antibody fragment (scFv fragment) of an antibody that specifically binds to ASGR1 and / or ASGR2. The “scFv fragment” comprises a VH region and a VL region of an antibody, which are located within a single polypeptide chain and, optionally, include a peptide linker between the VH and VL regions that allows Fv to form a desired structure for antigen binding. Exemplary antibodies and their conjugated fragments that specifically bind to ASGR1 and can be used as asiaroglycoprotein receptors in the targeting portion of LPA RNAi constructs used in the methods of the present invention are described in International Publication No. 2017 / 058944 (which is incorporated herein by reference in its entirety). Other antibodies or their conjugated fragments that specifically bind to ASGR1, LDL receptors, or other liver surface-expressed proteins and are suitable for use as targeting portions in LPA RNAi constructs are commercially available.

[0090] In certain embodiments, the targeted moiety includes carbohydrates. “Carbohydrate” refers to a compound composed of one or more monosaccharide units (which may be linear, branched, or cyclic) having at least six carbon atoms, each carbon atom bonded to an oxygen, nitrogen, or sulfur atom. Examples of carbohydrates include, but are not limited to, sugars (e.g., monosaccharides, disaccharides, trisaccharides, tetrasaccharides, and oligosaccharides containing about 4, 5, 6, 7, 8, or 9 monosaccharide units) and polysaccharides, such as starch, glycogen, cellulose, and polysaccharide gums. In some embodiments, the carbohydrates incorporated into the targeted moiety are monosaccharides selected from pentoses, hexoses, or heptoses, as well as disaccharides and trisaccharides containing such monosaccharide units. In other embodiments, the carbohydrates incorporated into the targeted moiety are amino sugars such as galactosamine, glucosamine, N-acetylgalactosamine, and N-acetylglucosamine.

[0091] In some embodiments, the targeting moiety comprises an asialoglycoprotein receptor ligand containing glucose, galactose, galactosamine, glucosamine, N-acetylglucosamine, N-acetyl-galactosamine, or any derivative thereof. In certain embodiments, the asialoglycoprotein receptor ligand comprises N-acetyl-galactosamine (GalNAc) or a derivative thereof. Ligands containing glucose, galactose, and GalNAc are particularly effective in directing the compound to hepatocytes because such ligands bind to ASGR expressed on the surface of hepatocytes. See, for example, D'Souza and Devarajan, J. Control Release, Vol. 203: 126-139, 2015. Examples of GalNAc-containing or galactose-containing ligands that can be incorporated into the targeting portion of the LPA RNAi construct used in the method of the present invention are described in U.S. Patent No. 7,491,805; U.S. Patent No. 8,106,022; U.S. Patent No. 8,877,917; and U.S. Patent No. 10,246,709; U.S. Patent Publication No. 20030130186; and International Publication No. 2013166155, all of which are incorporated herein by reference in their entirety.

[0092] In certain embodiments, the targeted moiety within the LPA RNAi construct includes a polyhydric carbohydrate moiety. As used herein, “polyhydric carbohydrate moiety” refers to a moiety comprising two or more carbohydrate units capable of independently binding to or interacting with other molecules. For example, a polyhydric carbohydrate moiety includes two or more binding domains composed of carbohydrates, capable of binding to two or more different molecules or two or more different sites on the same molecule. The valence of a carbohydrate moiety indicates the number of individual binding domains within the carbohydrate moiety. For example, the terms “monovalent,” “divalent,” “trivalent,” and “tetravalent” with respect to carbohydrate moieties refer to carbohydrate moieties having one, two, three, and four binding domains, respectively. A polyhydric carbohydrate moiety may include a polyhydric lactose moiety, a polyhydric galactose moiety, a polyhydric glucose moiety, a polyhydric N-acetyl-galactosamine moiety, a polyhydric N-acetyl-glucosamine moiety, a polyhydric mannose moiety, or a polyhydric fucose moiety. In some embodiments, the targeted moiety includes a polyhydric galactose moiety. In other embodiments, the targeted moiety includes a polyhydric N-acetyl-galactosamine moiety. In these and other embodiments, the polyhydric carbohydrate moiety may be dihydric, trihydric, or tetrahydric. In such embodiments, the polyhydric carbohydrate moiety may be bivalent or trivalent. In one particular embodiment, the polyhydric N-acetyl-galactosamine moiety is trihydric or tetrahydric. In another particular embodiment, the polyhydric galactose moiety is trihydric or tetrahydric.

[0093] The targeting moiety can be directly or indirectly bound to or conjugated to the RNA molecule of the LPA RNAi construct. For example, in some embodiments, the targeting moiety is directly covalently bound to the sense or antisense strand of the LPA RNAi construct. In other embodiments, the targeting moiety is covalently bound to the sense or antisense strand of the LPA RNAi construct via a linker. The targeting moiety can be bound to the nucleic acid base, sugar moiety, or internucleotide bond of the polynucleotide (e.g., sense or antisense strand) of the LPA RNAi construct used in the method of the present invention. Conjugation or binding to purine nucleic acid bases or their derivatives can occur at any position, including intra-ring and extra-ring atoms. In certain embodiments, the targeting moiety is bound to positions 2, 6, 7, or 8 of the purine nucleic acid base. Conjugation or binding to pyrimidine nucleic acid bases or their derivatives can also occur at any position. In some embodiments, positions 2, 5, and 6 of the pyrimidine nucleic acid base can be bound to the targeting moiety. Conjugation or binding to the sugar moiety of a nucleotide can occur at any carbon atom. Exemplary carbon atoms of the sugar moiety that can be bound to the targeting moiety include the 2', 3', and 5' carbon atoms. In addition, in debasalized nucleotides, for example, the 1' position can be bound to the targeting moiety. Furthermore, internucleotide bonds can support the binding of the targeting moiety. For phosphorus-containing bonds (e.g., phosphodiesters, phosphorothioates, phosphorodithioates, phosphoramidates, etc.), the targeting moiety can be bound directly to the phosphorus atom, or to the O, N, or S atoms bonded to the phosphorus atom. For amine-containing or amide-containing nucleoside bonds (e.g., PNA), the targeting moiety can be bound to the nitrogen atom of the amine or amide, or to an adjacent carbon atom.

[0094] In some embodiments, the targeting moiety may be bound to the 3' or 5' end of either the sense strand or the antisense strand. In certain preferred embodiments, the targeting moiety is covalently bound to the 5' end of the sense strand. In such embodiments, the targeting moiety is bound to the 5' terminal nucleotide of the sense strand. In these, and other embodiments, the targeting moiety is bound at the 5' position of the 5' terminal nucleotide of the sense strand. In embodiments where the inverted debasalized nucleotide or inverted deoxyribonucleotide is the 5' terminal nucleotide of the sense strand and is linked to an adjacent nucleotide via a 5'-5' nucleotide bond, the targeting moiety may be bound at the 3' position of the inverted debasalized nucleotide or inverted deoxyribonucleotide. In other embodiments, the targeting moiety is covalently bound to the 3' end of the sense strand. For example, in some embodiments, the targeting moiety is bound to the 3' terminal nucleotide of the sense strand. In certain such embodiments, the targeting moiety is bound at the 3' position of the 3' terminal nucleotide of the sense strand. In embodiments where the inverted debasalized nucleotide or inverted deoxyribonucleotide is the 3' terminal nucleotide of the sense strand and is linked to an adjacent nucleotide via a 3'-3' nucleotide bond, the targeting moiety may be attached at the 5' position of the inverted debasalized nucleotide or inverted deoxyribonucleotide. In alternative embodiments, the targeting moiety is attached near the 3' end of the sense strand, but before one or more terminal nucleotides (i.e., before terminal 1, 2, 3, or 4). In some embodiments, the targeting moiety is attached at the 2' position of the sugar of the 3' terminal nucleotide of the sense strand. In other embodiments, the targeting moiety is attached at the 2' position of the sugar of the 5' terminal nucleotide of the sense strand.

[0095] In certain embodiments, the targeting moiety is linked to the sense or antisense strand via a linker. A "linker" is an atom or atomic group that covalently bonds a ligand to a polynucleotide component of the LPA RNAi construct. Linkers can be about 1 to about 30 atomic lengths, about 2 to about 28 atomic lengths, about 3 to about 26 atomic lengths, about 4 to about 24 atomic lengths, about 6 to about 20 atomic lengths, about 7 to about 20 atomic lengths, about 8 to about 20 atomic lengths, about 8 to about 18 atomic lengths, about 10 to about 18 atomic lengths, and about 12 to about 18 atomic lengths. In some embodiments, the linker may include a bifunctional linking moiety, which generally includes an alkyl moiety having two functional groups. One of the functional groups is selected to bind to the compound of interest (e.g., the sense or antisense strand of the RNAi construct), and the other is selected to essentially bind to any selected group, such as the targeting moiety or its components as described herein. In certain embodiments, the linker comprises a chain structure or oligomer consisting of repeating units such as ethylene glycol units or amino acid units. Examples of functional groups typically used in the difunctional bond portion include, but are not limited to, electrophiles for reacting with nucleophilic groups and nucleophiles for reacting with electrophilic groups. In some embodiments, the difunctional bond portion may be amino, hydroxyl, carboxylic acid, thiol, or unsaturated (e.g., double or triple bonds).

[0096] Linkers that can be used to attach the targeting portion to the sense or antisense strand in the LPA RNAi construct used in the method of the present invention include pyrrolidine, 8-amino-3,6-dioxaoctanoic acid, succinimidyl 4-(N-maleimidomethyl)cyclohexane-1-carboxylate, 6-aminohexanoic acid, and substituted C1-C. 10 Alkyl, substituted, or unsubstituted C2-C 10 Alkenyl, or substituted or unsubstituted C2-C 10Examples of alkynyls include, but are not limited to, alkynyl substituents. Preferred substituents for such linkers include, but are not limited to, hydroxyl, amino, alkoxy, carboxy, benzyl, phenyl, nitro, thiol, thioalkoxy, halogen, alkyl, aryl, alkenyl, and alkynyl substituents. Other types of linkers suitable for linking the targeting moiety to the sense or antisense strand in the LPA RNAi construct used in the method of the present invention are known in the art and include the linkers described in U.S. Patent No. 7,723,509; U.S. Patent No. 8,017,762; U.S. Patent No. 8,828,956; U.S. Patent No. 8,877,917; and U.S. Patent No. 9,181,551.

[0097] In certain embodiments, the targeting moiety covalently bound to the sense or antisense strand of the LPA RNAi construct used in the method of the present invention includes a GalNAc moiety, such as a polyvalent GalNAc moiety. In some embodiments, the polyvalent GalNAc moiety is a trivalent GalNAc moiety bound to the 3' end of the sense strand. In other embodiments, the polyvalent GalNAc moiety is a trivalent GalNAc moiety bound to the 5' end of the sense strand. In yet another embodiment, the polyvalent GalNAc moiety is a tetravalent GalNAc moiety bound to the 3' end of the sense strand. In yet another embodiment, the polyvalent GalNAc moiety is a tetravalent GalNAc moiety bound to the 5' end of the sense strand.

[0098] In a particular embodiment, the LPA RNAi construct used in the method of the present invention includes a targeting moiety having the following structure [Structure 1]. [ka] In a preferred embodiment, the targeting moiety having this structure is covalently bonded to the 5' end of the sense chain via a phosphorothioate or phosphate diester bond.

[0099] In a particular embodiment, an LPA RNAi construct suitable for use in the method of the present invention is: a sense strand and an antisense strand, each about 19 to about 23 nucleotides in length, wherein the antisense strand contains a sequence complementary to the LPA mRNA sequence, and the sense strand contains a sequence complementary to the sequence of the antisense strand; A targeting moiety containing an asialoglycoprotein receptor ligand, wherein the targeting moiety is covalently bound to the 5' end of the sense chain. This includes. In some embodiments, the LPA RNAi construct has two blunt ends. For example, in some such embodiments, the sense strand and the antisense strand are each 21 nucleotides long and hybridize with each other to form a 21-base pair long double-stranded region. In other such embodiments, the sense strand and the antisense strand are each 19 nucleotides long and hybridize with each other to form a 19-base pair long double-stranded region. In other embodiments, the LPA RNAi construct has two nucleotide overhangs. In one such embodiment, the LPA RNAi construct includes (i) a sense strand and an antisense strand, each 21 nucleotides long, (ii) a 19-base pair long double-stranded region, and (iii) nucleotide overhangs of two unpaired nucleotides at both the 3' end of the sense strand and the 3' end of the antisense strand.

[0100] In some embodiments, the targeted portion includes a trivalent chromosome GalNAc moiety, for example, any of the trivalent chromosome GalNAc moieties described in U.S. Patent No. 10,246,709 (which is incorporated herein by reference in its entirety). In one preferred embodiment, the targeted portion has the structure of Structure 1 described above.

[0101] In certain embodiments, the antisense strand of the LPA RNAi construct includes sequences substantially complementary or fully complementary to nucleotides 2706-2726, nucleotides 2697-2726, or nucleotides 2708-2725 of the human LPA mRNA transcript shown in NCBI reference sequence NM_005577.4. In such embodiments, the LPA RNAi construct may include a sense strand that is substantially complementary or fully complementary to the antisense strand targeting this region. Thus, in these embodiments, the sense strand may include sequences identical to nucleotides 2706-2726, nucleotides 2697-2726, or nucleotides 2725-2708 of the human LPA mRNA transcript shown in NCBI reference sequence NM_005577.4.

[0102] In some embodiments, the sense strand of the LPA RNAi construct used in the method of the present invention contains the sequence 5'-GCCCCUUAUUGUUAUACG-3' (SEQ ID NO: 1). In related embodiments, the antisense strand of the LPA RNAi construct used in the method of the present invention contains the sequence 5'-CGUAUAACAAUAAGGGGC-3' (SEQ ID NO: 2).

[0103] Examples of LPA RNAi constructs suitable for use in the methods of the present invention are described in International Publication No. 2017 / 059223 (which is incorporated herein by reference in its entirety). The double-stranded AD03851, AD03853, and AD03536 described in International Publication No. 2017 / 059223 are particularly useful in the methods of the present invention. In certain preferred embodiments, the LPA RNAi construct used in the methods of the present invention comprises a sense strand containing or derived from the sequence 5'-CAGCCCCUUAUUGUUAUACGA-3' (SEQ ID NO: 3) and an antisense strand containing or derived from the sequence 5'-UCGUAUAACAAUAAGGGGCUG-3' (SEQ ID NO: 4). In related embodiments, the LPA used in the methods of the present invention The RNAi construct includes a sense strand containing or comprising a sequence of modified nucleotides following the sequence 5'-csagccccuUfAfUfuguuauacgs(invdA)-3' (SEQ ID NO: 5), and an antisense strand containing or comprising a sequence of modified nucleotides following the sequence 5'-usCfsgUfaUfaacaaUfaAfgGfgGfcsUfsg-3' (SEQ ID NO: 6), where a, g, c, and u are 2'-O -methyladenosine, 2'-O-methylguanosine, 2'-O-methylcytidine, and 2'-O-methyluridine; Af, Gf, Cf, and Uf are 2'-deoxy-2'-fluoro("2'-fluoro")adenosine, 2'-fluoroguanosine, 2'-fluorocytidine, and 2'-fluorouridine, respectively; invdA is an inverted deoxyadenosine (3'-3' linked nucleotide), and s is a phosphorothioate bond. In some such embodiments, the targeting moiety having the structure of Structure 1 described herein is covalently bonded to the 5' end of the sense strand via a phosphorothioate bond.

[0104] In another embodiment, the LPA RNAi construct used in the method of the present invention comprises a sense strand containing the sequence 5'-GCCCCUUAUUGUUAUACGAUU-3' (SEQ ID NO: 7) and an antisense strand containing the sequence 5'-UCGUAUAACAAUAAGGGGCUU-3' (SEQ ID NO: 8). In a related embodiment, the LPA RNAi construct used in the method of the present invention comprises a sense strand containing or derived from a sequence of modified nucleotides following the sequence 5'-gsccccuUfAfUfuguuauacgauus(invAb)-3' (SEQ ID NO: 9), and an antisense strand containing or derived from a sequence of modified nucleotides following the sequence 5'-usCfsgUfaUfaacaaUfaAfgGfgGfcsusu-3' (SEQ ID NO: 10), where a, g, c, and u are 2'- These are O-methyladenosine, 2'-O-methylguanosine, 2'-O-methylcytidine, and 2'-O-methyluridine; Af, Gf, Cf, and Uf are 2'-deoxy-2'-fluoro("2'-fluoro")adenosine, 2'-fluoroguanosine, 2'-fluorocytidine, and 2'-fluorouridine, respectively; invAb is an inverted debasal nucleotide (3'-3' linked nucleotide), and s is a phosphorothioate bond. In some such embodiments, the targeting moiety having the structure of Structure 1 described herein is covalently bonded to the 5' end of the sense strand via a phosphorothioate bond.In other related embodiments, the LPA RNAi construct used in the method of the present invention comprises a sense strand comprising or derived from a sequence of modified nucleotides following the sequence 5'-(invAb)GfcCfcCfuUfAfUfuGfuUfaUfaCfgausu(invAb)-3' (SEQ ID NO: 11), and an antisense strand comprising or derived from a sequence of modified nucleotides following the sequence 5'-usCfsgsUfaUfaAfCfAfauaAfgGfgGfcusu-3' (SEQ ID NO: 12), where a, g, c, and u are 2'-O-methyladenosine, 2 These are '-O-methylguanosine, 2'-O-methylcytidine, and 2'-O-methyluridine; Af, Gf, Cf, and Uf are 2'-deoxy-2'-fluoro("2'-fluoro")adenosine, 2'-fluoroguanosine, 2'-fluorocytidine, and 2'-fluorouridine, respectively; invAb is an inverted debasal nucleotide (a 5'-5' linked nucleotide if it is on the 5' end of the chain, or a 3'-3' linked nucleotide if it is on the 3' end of the chain), and s is a phosphorothioate bond. In some of these embodiments, the targeting moiety having the structure of Structure 1 described herein is covalently bonded to the 5' end of the sense chain via a phosphodiester bond.

[0105] In a particular preferred embodiment, the LPA RNAi construct administered to a patient according to the method of the present invention is olpasilan. The structure of olpasilan is schematically shown in Figure 1 and is described in International Publication No. 2017 / 059223, where olpasilan is represented as double-stranded no. AD03851. Orpasilan is a double-stranded siRNA molecule comprising two separate strands, a sense strand and an antisense strand (each 21 nucleotides long). The nucleic acid base sequences of the sense strand and antisense strand are perfectly complementary to each other and hybridize to form a 21-base-pair-long double helix. The nucleotide sequences for the sense strand and antisense strand of olpasilan are shown in SEQ ID NOs: 3 and SEQ ID NOs: 4, respectively. Both the sense strand and antisense strand of olpasilan consist of modified nucleotides, and the modification sequences for each strand are shown in SEQ ID NOs: 5 (sense strand) and SEQ ID NOs: 6 (antisense strand). The trivalent GalNAc moiety having the structure of Structure 1 (and represented as R1 in Figure 1) is covalently bonded to the 5' end of the sense chain of orpasilane by a phosphorothioate bond. The term orpasilane refers to the free acid of the compound shown in Figure 1, and its pharmaceutically acceptable salts, such as the sodium salt.

[0106] The LPA RNAi constructs used in the method of the present invention can be readily prepared using techniques known in the art, for example, by conventional solid-phase nucleic acid synthesis. The polynucleotides of the RNAi constructs can be assembled using standard nucleotides or nucleoside precursors (e.g., phosphoramidites) in a suitable nucleic acid synthesizer. Automated nucleic acid synthesizers are commercially available from several vendors, including the DNA / RNA synthesizer from Applied Biosystems (Foster City, CA), the MerMade synthesizer from BioAutomation (Irving, TX), and the OligoPilot synthesizer from GE Healthcare Life Sciences (Pittsburgh, PA). Exemplary methods for synthesizing LPA RNAi constructs and selecting targeting moieties are described in the examples of International Publication No. 2017 / 059223 and U.S. Patent No. 10,246,709, both of which are incorporated herein by reference in their entirety.

[0107] Oligonucleotides can be synthesized via phosphoramidite chemistry by using a 2'-silyl protecting group in combination with dimethoxytrityl (DMT), which is acid-unstable at the 5' position of the ribonucleoside. The final deprotection conditions are known not to significantly degrade the RNA product. All synthesis can be carried out on a large, medium, or small scale using any automated or manual synthesizer. Furthermore, synthesis can be performed in multi-well plates, columns, or glass slides.

[0108] The 2'-O-silyl group can be removed by exposure to fluoride ions, which may be any source of fluoride ions, such as salts containing fluoride ions paired with inorganic counterions, such as cesium fluoride and potassium fluoride, or salts containing fluoride ions paired with organic counterions, such as tetraalkylammonium fluoride. Crown ether catalysts may be used in combination with inorganic fluorides in the deprotection reaction. Preferred sources of fluoride ions include tetrabutylammonium fluoride or aminohydrofluorides (e.g., aqueous HF in a dipolar aproton solvent, such as dimethylformamide, in combination with triethylamine).

[0109] The stability of tryesters with respect to fluorides can be modified by selecting the protecting group used for tryesters with phosphite and tryesters. Methyl protection of phosphotryesters or tryesters with phosphite can stabilize the binding to fluoride ions and improve process yield.

[0110] Since ribonucleosides have a reactive 2'-hydroxyl substituent, it is sometimes desirable to protect the reactive 2' position in the RNA with a protecting group that is orthogonal to the 5'-O-dimethoxytrityl protecting group (for example, a protecting group that is stable to acid treatment). Silyl protecting groups satisfy this condition and can be easily removed in the final fluoride deprotection step, thereby minimizing RNA degradation.

[0111] Tetrazole catalysts can be used in standard phosphoramidite coupling reactions. Preferred catalysts include, for example, tetrazole, S-ethyl-tetrazole, benzylthiotetrazole, and p-nitrophenyltetrazole.

[0112] As can be understood by those skilled in the art, further methods for synthesizing the LPA RNAi constructs described herein will be apparent to them. In addition, various synthetic steps can be performed in alternative order or sequence to obtain the desired compounds. Other synthetic chemical transformations, protecting groups (e.g., hydroxyl, amino, and others present in bases), and methodologies for protecting and deprotecting RNAi constructs that are useful in the synthesis of RNAi constructs are known in the art, such as those described in R. Larock, Comprehensive Organic Transformations, VCH Publishers (1989); TW. Greene and PG. Wüts, Protective Groups in Organic Synthesis, 2nd ed., John Wiley and Sons (1991); L. Fieser and M. Fieser, Fieser and Fieser's Reagents for Organic Synthesis, John Wiley and Sons (1994); and L. Paquette, ed., Encyclopedia of Reagents for Organic Synthesis, John Wiley and Sons (1995), and subsequent editions thereof. Furthermore, custom synthesis of RNAi agents is available from several private vendors, including Agilent Technologies (Santa Clara, CA), Nitto Denko Avecia (Milford, MA), Dharmacon, Inc. (Lafayette, CO), AxoLabs GmbH (Kulmbach, Germany), and Ambion, Inc. (Foster City, CA).

[0113] LPA RNAi constructs are typically administered to patients in a pharmaceutical composition that may contain a pharmaceutically acceptable carrier, excipient, or diluent. Therefore, the present invention also includes pharmaceutical compositions and formulations comprising an LPA RNAi construct and a pharmaceutically acceptable carrier, excipient, or diluent, as used in the methods of the present invention described herein. For clinical use, the pharmaceutical compositions and formulations will be prepared in a form appropriate to the intended use. Generally, this will involve the preparation of compositions that are essentially free not only from pyrogens but also from other impurities that may be harmful to humans or animals.

[0114] The terms "pharmaceutically acceptable" or "pharmacologically acceptable" refer to molecular entities and compositions that do not cause adverse, allergic, or other undesirable reactions when administered to animals or humans. As used herein, "pharmaceutically acceptable carriers, excipients, or diluents" include solvents, buffers, solutions, dispersions, coatings, antimicrobial and antifungal agents, isotonic agents, and absorption retarders that are acceptable for use in the formulation of pharmaceuticals, such as medicines, suitable for human administration. The use of such media and agents with pharmaceutically active substances is well known in the art. Any conventional media or agent is intended for use in therapeutic compositions unless it is incompatible with the LPA RNAi constructs described herein. Additionally, auxiliary active ingredients may be incorporated into compositions, provided they do not inactivate the LPA RNAi constructs in the composition.

[0115] The compositions and methods for formulating pharmaceutical compositions depend on several criteria, including but not limited to the route of administration, the type and degree of the disease or disorder to be treated, or the dose to be administered. In some embodiments, the pharmaceutical composition is formulated based on the intended route of delivery. For example, in certain embodiments, the pharmaceutical composition is formulated for parenteral delivery. Parenteral delivery forms include intravenous, intra-arterial, subcutaneous, intrathecal, intraperitoneal, or intramuscular injection or infusion. In one embodiment, the pharmaceutical composition is formulated for intravenous delivery. In another embodiment, the pharmaceutical composition is formulated for subcutaneous delivery. In some embodiments, the pharmaceutical composition contains an effective amount of an LPA RNAi construct. The effective amount of the LPA RNAi construct, in particular olpasilan, may be any of the doses described herein.

[0116] Administration of a pharmaceutical composition containing an LPA RNAi construct according to the method of the present invention may be via any common route, as long as the target tissue is available through that route. Such routes include, but are not limited to, parenteral (e.g., subcutaneous, intramuscular, intraperitoneal, or intravenous), oral, nasal, buccal, intradermal, transdermal, and sublingual routes, or direct injection into liver tissue or delivery via the hepatic portal vein. In some embodiments of the method of the present invention, the LPA RNAi construct, or a pharmaceutical composition containing an LPA RNAi construct, is administered parenterally to the patient. For example, in certain embodiments, the LPA RNAi construct, or a pharmaceutical composition containing an LPA RNAi construct, is administered intravenously. In other embodiments, the LPA RNAi construct, or a pharmaceutical composition containing an LPA RNAi pharmaceutical composition, is administered subcutaneously, for example, by subcutaneous injection. In such embodiments, the subcutaneous injection volume is about 2 mL or less, for example, about 2 mL, about 1.8 mL, about 1.7 mL, about 1.6 mL, about 1.5 mL, about 1.4 mL, about 1.3 mL, about 1.2 mL, about 1.1 mL, about 1 mL, about 0.9 mL, about 0.8 mL, about 0.7 mL, about 0.6 mL, or about 0.5 mL. In one embodiment, the subcutaneous injection volume is about 1 mL or less. In another embodiment, the subcutaneous injection volume is about 1 mL. In yet another embodiment, the subcutaneous injection volume is about 1.5 mL.

[0117] In embodiments where the pharmaceutical composition is administered by parenteral injection, the pharmaceutical composition may be administered to the patient by syringe. In some embodiments, the syringe is pre-filled with the pharmaceutical composition. In other embodiments where the pharmaceutical composition is administered to the patient by parenteral injection, for example subcutaneous injection, the pharmaceutical composition is administered by an injection device, including, but not limited to, a self-administering device. Such devices are commercially available and include, but are not limited to, autoinjectors, dosing pens, microinjection pumps, on-body injectors, and pre-filled syringes. Exemplary devices for administering a pharmaceutical composition containing an effective amount of an LPA RNAi construct (e.g., olpasilan) according to the method of the present invention include autoinjectors (e.g., SureClick®, EverGentle®, Avanti®, DosePro®, Molly®, and Leva®), pen injection devices (e.g., Madie® pen injector, DCP® pen injector, BD Vystra® disposable pen, BD® reusable pen), and pre-filled syringes (BD Examples include Sterifill®, BD Hypak®, and Baxter pre-filled syringes. In some embodiments, a pharmaceutical composition containing an effective amount of an LPA RNAi construct (e.g., olpasilan) is administered to the patient by a pre-filled syringe. In other embodiments, a pharmaceutical composition containing an effective amount of an LPA RNAi construct (e.g., olpasilan) is administered to the patient by an autoinjector. In certain such embodiments, the injection volume of the syringe, autoinjector, or other injection device is about 2 mL or less, for example, about 2 mL, about 1.8 mL, about 1.7 mL, about 1.6 mL, about 1.5 mL, about 1.4 mL, about 1.3 mL, about 1.2 mL, about 1.1 mL, about 1 mL, about 0.9 mL, about 0.8 mL, about 0.7 mL, about 0.6 mL, or about 0.5 mL. In one embodiment, the injection volume of the syringe, autoinjector, or other injection device is about 1 mL or less. In another embodiment, the injection volume of the syringe, auto-injector, or other injection device is approximately 1 mL. In yet another embodiment, the injection volume of the syringe, auto-injector, or other injection device is approximately 1.5 mL.

[0118] Colloidal dispersion systems such as polymer complexes, nanocapsules, microspheres, beads, and lipid-based systems, including oil-in-water emulsions, micelles, mixed micelles, and liposomes, can be used as delivery vehicles for the LPA RNAi constructs of the present invention. Commercially available lipid emulsions suitable for delivering the nucleic acids of the present invention include Intralipid® (Baxter International Inc.), Liposyn® (Abbott Pharmaceuticals), Liposyn® II (Hospira), Liposyn® III (Hospira), Nutrilipid (B. Braun Medical Inc.), and other similar lipid emulsions. Liposomes (i.e., artificial membrane vesicles) are preferred colloidal systems used as in vivo delivery vehicles. The LPA RNAi constructs may be encapsulated within liposomes or complexed with liposomes, particularly cationic liposomes. In addition, LPA RNAi constructs can complex with lipids, particularly cationic lipids. Suitable lipids and liposomes include neutral (e.g., dioleoylphosphatidylethanolamine (DOPE), dimyristoylphosphatidylcholine (DMPC), and dipalmitoylphosphatidylcholine (DPPC), distearoylphosphatidylcholine), negative (e.g., dimyristoylphosphatidylglycerol (DMPG)), and cationic (e.g., dioleoyltetramethylaminopropyl (DOTAP) and dioleoylphosphatidylethanolamine (DOTMA)). The preparation and use of such colloidal dispersions are well known in this art. Furthermore, exemplary formulations are disclosed in U.S. Patent Nos. 5,981,505, 6,217,900, 6,383,512, 5,783,565, 7,202,227, 6,379,965, 6,127,170, 5,837,533, 6,747,014, and in International Publication No. 03 / 093449.

[0119] In some embodiments, the LPA RNAi construct is completely encapsulated within a lipid formulation to form, for example, SNALP or other nucleic acid-lipid particles. As used herein, the term "SNALP" refers to stable nucleic acid-lipid particles. SNALP typically contains cationic lipids, non-cationic lipids, and lipids that prevent particle aggregation (e.g., PEG-lipid conjugates). SNALP is extremely useful for systemic administration because it exhibits a long circulating lifetime after intravenous injection and accumulates at distal sites (e.g., sites physically separated from the administration site). Nucleic acid-lipid particles typically have an average diameter of about 50 nm to 150 nm, 60 nm to 130 nm, 70 nm to 110 nm, or 70 nm to 90 nm and are substantially non-toxic. In addition, when present within nucleic acid-lipid particles, the nucleic acids are resistant to degradation by nucleases in aqueous solutions. Nucleic acid-lipid particles and methods for preparing them are disclosed, for example, in U.S. Patent No. 5,976,567, U.S. Patent No. 5,981,501, U.S. Patent No. 6,534,484, U.S. Patent No. 6,586,410, U.S. Patent No. 6,815,432, and International Publication No. 96 / 40964.

[0120] Examples of pharmaceutical compositions containing LPA RNAi constructs suitable for injection include sterile aqueous solutions or dispersions, and sterile powders for the immediate preparation of sterile solutions or dispersions for injection. Generally, these preparations are sterile and fluid enough to be easily injected. The preparations must be stable under manufacturing and storage conditions and protected from contamination by microorganisms such as bacteria and fungi. Suitable solvents or dispersion media may include, for example, water, ethanol, polyols (e.g., glycerol, propylene glycol, liquid polyethylene glycol, etc.), suitable mixtures thereof, and vegetable oils. For example, the particle size required for dispersions can be maintained by the use of coatings such as lecithin, and appropriate fluidity can be maintained by the use of surfactants. Prevention of microbial action can be achieved by various antimicrobial and antifungal agents, such as parabens, chlorobutanol, phenol, sorbic acid, thimerosal, etc. In many cases, it is preferable to include isotonic agents, such as sugars or sodium chloride. Sustained absorption of an injectable composition can be achieved by using absorption-delaying agents, such as aluminum monostearate and gelatin, in the composition.

[0121] Sterile injectable solutions can be prepared by incorporating an appropriate amount of the active compound into a solvent, along with any other optional components as needed (e.g., those listed above), and then sterilizing by filtration. Generally, dispersion systems are prepared by incorporating various sterilized active ingredients into a sterile vehicle containing a basic dispersion medium and other desired components, such as those listed above. For sterile powders for the preparation of sterile injectable solutions, preferred preparation methods include vacuum drying and freeze-drying techniques, from which a powder of the active ingredient plus any additional desired components is obtained from a pre-sterilized filtered solution.

[0122] The compositions used in the method of the present invention can generally be formulated in neutral or salt form. Examples of pharmaceutically acceptable salts include acid addition salts (formed with a free amino group) derived from inorganic acids (e.g., hydrochloric acid or phosphoric acid) or organic acids (e.g., acetic acid, oxalic acid, tartaric acid, mandelic acid, etc.). Salts formed with a free carboxyl group can be derived from inorganic bases (e.g., sodium, potassium, ammonium, calcium, or ferric hydroxide) or organic bases (e.g., isopropylamine, trimethylamine, histidine, procaine, etc.). Sodium salts of the LPA RNAi construct are particularly useful for therapeutic administration to human subjects. Therefore, in certain preferred embodiments, LPA RNAi constructs, particularly orpasilane, are in the form of a sodium salt. In other embodiments, the LPA RNAi construct (e.g., orpasilane) is in the form of a potassium salt.

[0123] For parenteral administration in aqueous solutions, for example, the solution is usually appropriately buffered, and the liquid diluent is first isotonic with, for example, sufficient saline or glucose. Such aqueous solutions can be used, for example, for intravenous, intramuscular, subcutaneous, and intraperitoneal administration. Particularly in consideration of this disclosure, it is preferable to use sterile aqueous media as known to those skilled in the art. As an example, a single dose may be dissolved in 1 ml of isotonic NaCl solution and added to 1000 ml of subcutaneous injection solution, or injected into a candidate site for injection or administration (see, for example, “Remington's Pharmaceutical Sciences” 15th Edition, pages 1035-1038 and 1570-1580). For human administration, the preparation must meet the standards of sterility, pyrogenicity, general safety, and purity required by FDA standards. In certain embodiments, the pharmaceutical composition used in the method of the present invention comprises or consists of sterile saline and an LPA RNAi construct described herein (e.g., olpasilan). In other embodiments, the pharmaceutical composition used in the method of the present invention comprises or consists of an LPA RNAi construct described herein (e.g., olpasilane) and sterile water (e.g., water for injection, WFI). In yet another embodiment, the pharmaceutical composition used in the method of the present invention comprises or consists of an LPA RNAi construct described herein (e.g., olpasilane) and phosphate-buffered saline (PBS).

[0124] In certain embodiments, a pharmaceutical composition useful for treating, relieving, preventing, or reducing the risk of cardiovascular disease according to the method of the present invention comprises an effective amount of an LPA RNAi construct (e.g., olpasilan), potassium phosphate buffer, and sodium chloride. In some such embodiments, the pharmaceutical composition comprises about 10 mg / mL to about 200 mg / mL of an LPA RNAi construct (e.g., olpasilan), about 5 mM to about 30 mM potassium phosphate, and about 20 mM to about 160 mM sodium chloride. In other embodiments, the pharmaceutical composition comprises about 65 mg / mL to about 85 mg / mL of an LPA RNAi construct (e.g., olpasilan), about 15 mM to about 25 mM potassium phosphate, and about 70 mM to about 90 mM sodium chloride. In yet another embodiment, the pharmaceutical composition comprises about 140 mg / ml to about 160 mg / ml of an LPA RNAi construct (e.g., olpasilane), about 15 mM to about 25 mM potassium phosphate, and about 30 mM to about 50 mM sodium chloride. The pH of any of these pharmaceutical compositions may be in the range of about 6.4 to about 7.2 (e.g., pH of about 6.4, about 6.6, about 6.8, about 7.0, or about 7.2).

[0125] In some embodiments, the pharmaceutical composition to be administered according to the method of the present invention contains about 10 mg / ml of LPA RNAi construct (e.g., olpasilan), about 5 mM to about 15 mM potassium phosphate, and about 135 mM to about 155 mM sodium chloride at pH 6.8 ± 0.2. In one embodiment, the pharmaceutical composition contains about 10 mg / ml of LPA RNAi construct (e.g., olpasilan), about 10 mM potassium phosphate, and about 145 mM sodium chloride at pH 6.8. In other embodiments, the pharmaceutical composition to be administered according to the method of the present invention contains about 75 mg / ml of LPA RNAi construct (e.g., olpasilan), about 15 mM to about 25 mM potassium phosphate, and about 70 mM to about 90 mM sodium chloride at pH 6.8 ± 0.2. In one such embodiment, the pharmaceutical composition contains about 75 mg / ml of LPA RNAi construct (e.g., olpasilan), about 20 mM potassium phosphate, and about 80 mM sodium chloride at pH 6.8. In a particular embodiment, the pharmaceutical composition to be administered according to the method of the present invention contains about 150 mg / ml of LPA RNAi construct (e.g., olpasilan), about 15 mM to about 25 mM potassium phosphate, and about 30 mM to about 50 mM sodium chloride at pH 6.8 ± 0.2. In a particular embodiment, the pharmaceutical composition contains about 150 mg / ml of LPA RNAi construct (e.g., olpasilan), about 20 mM potassium phosphate, and about 40 mM sodium chloride at pH 6.8.

[0126] Any LPA RNAi construct described herein may be incorporated into any of the above-described pharmaceutical compositions and administered to a patient according to the method of the present invention. In a particular embodiment, the LPA RNAi construct incorporated into any of the above-described pharmaceutical compositions and administered to a patient according to the method of the present invention is olpasilan.

[0127] In some embodiments, the pharmaceutical composition of the present invention is packaged in or stored within a device for administration, such as any of the above-described injection devices (e.g., pre-filled syringes, auto-injectors, injection pumps, on-body injectors, and injection pens). Devices for aerosolized or powdered formulations include, but are not limited to, inhalers, ventilators, and aspirators. Therefore, the present invention includes an administration device containing the pharmaceutical composition of the present invention for treating or preventing one or more of the disorders or diseases described herein.

[0128] The following examples, including the experiments conducted and the results achieved, are provided for illustrative purposes only and should not be construed as limiting the scope of the appended claims. [Examples]

[0129] Example 1. Phase 1 randomized, double-blind, placebo-controlled, single-dose escalation study to evaluate the safety, tolerability, pharmacokinetics, and pharmacodynamics of olpasilan in subjects with elevated plasma lipoprotein(a) levels. Mendel's studies, and recent epidemiological randomized controlled trials, have established lipoprotein(α) (Lp(α)) as a strong causative risk factor for myocardial infarction and other atherosclerotic complications. Currently, there are no approved drugs that selectively target Lp(α) and demonstrate a reduction in cardiovascular events. Orpasilan (also known as AMG 890) is an siRNA designed to reduce Lp(α) production by targeting mRNA transcribed from the LPA gene. The structure of orpasilan is shown in Figure 1.

[0130] This Phase 1 study was a randomized, double-blind, placebo-controlled single-dose escalation study conducted at eight locations in the United States and Australia in subjects with elevated plasma lipoprotein(a). Approximately 80 subjects were planned for enrollment in nine single-dose escalation cohorts; in each cohort, subjects were randomized in a 3:1 ratio to receive either olpasiran or placebo.

[0131] Eligible participants were non-fertile women and men, both aged 18–60 years (inclusive) for cohorts 1–5 and 18–65 years (inclusive) for cohorts 6–9. For cohorts 1–5, plasma Lp(a) concentrations were ≥70 nmol / L and ≤199 nmol / L at screening; for cohorts 6–9, plasma Lp(a) concentrations were ≥200 nmol / L at screening; and for cohorts 6–9, at least six participants in each cohort had a stable statin dose for at least six weeks at the time of enrollment. At the time of randomization, there were no clinically significant abnormalities in the participants' medical history.

[0132] After providing informed consent, subjects were screened against eligibility criteria over 28 days and admitted to the research facility on day -1. Following completion of pre-administration procedures, subjects received the study drug (olpasilan or placebo). Subjects in cohorts 1-5 stayed at the research facility from day -1 to day 4 and returned to the facility for evaluation until the end of the study. Subjects in cohorts 1-5 received single subcutaneous doses of 3 mg, 9 mg, 30 mg, 75 mg, and 225 mg, respectively; and subjects in cohorts 6-9 received 9 mg, 75 mg, 225 mg, and 675 mg, respectively (see Table 1 below).

[0133] [Table 1]

[0134] For cohorts 1-5 and cohort 9, the first two enrolled subjects in each cohort were randomized and administered either olpasilan or placebo in a 1:1 ratio (sentinel pair) at the same study site on the same day in a blinded manner. The same dose was administered to the remaining cohort subjects only after the investigators deemed it safe and at least 24 hours after the sentinel pair administration. Enrollment in cohorts 1-5 was staggered. After the dosing regimen in the preceding cohort was found safe and moderately tolerable by the dose-level review team (DLRT), all subjects were administered to subsequent cohorts based on available safety data up to day 15 of the study. Enrollment in cohorts 5-7 began after the dosing regimen in cohort 4 was found safe and moderately tolerable by the DLRT based on available safety data up to day 15 of the study. Participants returned to the facility for post-treatment evaluation at day 113 for cohorts 1 and 2, and at day 225 for cohorts 3–7. Participants returned for follow-up until their Lp(a) concentration was at least 80% of baseline (approximately every two weeks for cohorts 1 and 2, and monthly for cohorts 3–7). Blood and urine samples were collected throughout the study to evaluate olpasiran pharmacokinetics (PK) and pharmacodynamics (PD). Safety variables were also assessed periodically.

[0135] The primary endpoints were safety and tolerability as measured by adverse events (TEAEs) caused by the procedure, safety laboratory analyses, vital signs, and electrocardiogram (ECG). The secondary endpoint was the maximum observed concentration (C max ), the time point at which the maximum observed concentration was reached (t maxOrpasiran PK parameters included, but were not limited to, ), and area under the concentration-time curve (AUC); and PD parameters included the change and percentage change in plasma Lp(a) levels at each scheduled visit. The baseline value for Lp(a) was defined as the mean of screening and one day prior to administration. If for any reason only one value was available, that value was used as the baseline. The exploratory endpoints included the percentage change in low-density lipoprotein cholesterol (LDL-C) and total apolipoprotein B (ApoB) at each scheduled visit.

[0136] Sixty-four subjects were enrolled in the study and administered either olpasilan or placebo (Cohorts 1-5: olpasilan, n=30, doses: 3mg, 9mg, 30mg, 75mg, 225mg; placebo, n=10; Cohorts 6-7: olpasilan, n=18, doses: 9mg and 75mg; placebo, n=6). In Cohorts 1-5, the subjects who received olpasilan (n=30) had a mean (SD) age of 43.9 (13.5) years, 30.0% were female, 63.3% were Hispanic or Latino, 30.0% were Black or African American, and 70.0% were White. In cohorts 1-5, the subjects who received placebo (n=10) had a mean (SD) age of 46.3 (8.5) years, 30.0% were female, 50.0% were Hispanic or Latino, 30.0% were Black or African American, and 70.0% were white. In cohorts 6 and 7, the subjects who received olpasilan (n=18) had a mean (SD) age of 52.7 (9.4) years, 33.3% were female, 27.8% were Hispanic or Latino, and 88.9% were white. In cohorts 6 and 7, the subjects who received placebo (n=6) had a mean (SD) age of 57.8 (5.8) years, 66.7% were female, 33.3% were of Hispanic or Latin American descent, and 83.3% were white. 67% of all subjects enrolled in cohorts 6 and 7 (n=24) used statins at baseline. Subjects had few comorbidities. No lipid-modulating drugs were used in cohorts 1-5, but a significant proportion of subjects in cohorts 6 and 7 took statins and / or ezetimibe. Median (Q1, Q3) baseline Lp(a) concentrations were 124 nmol / L (104, 137) in subjects who received placebo in cohorts 1-5, and 122 nmol / L (97, 146) in subjects who received olpasilan in cohorts 1-5.The median (Q1, Q3) baseline Lp(a) concentration was 272 nmol / L (233, 307) in the placebo-receiving subjects in cohorts 6 and 7, and 253 nmol / L (224, 334) in the olpasilan-receiving subjects in cohorts 6 and 7.

[0137] Orpasilan appeared to be well-tolerated. There were no serious adverse events related to the procedure. One placebo participant experienced a serious noncardiac chest pain, which was considered unrelated to the procedure. In cohorts 1–5, the most common TEAE was upper respiratory tract infection (10% placebo, 13% orpasilan). See Table 2 below. In cohorts 6–7, the most common TEAEs were headache (50% placebo, 28% orpasilan) and upper respiratory tract infection (17% placebo, 17% orpasilan) (Table 2). Only one participant in the study experienced an injection site reaction. There was no apparent dose-response relationship with the frequency of adverse events. No clinically relevant changes in liver function, platelet or coagulation parameters, or renal function were observed.

[0138] [Table 2]

[0139] Lp(a) suppression occurred in a dose-response manner. As shown in Figure 2, in cohorts 1–5, a single dose of olpasilan effectively reduced mean Lp(a) levels by 71–96% (dose-based) from baseline at day 43 and by 80–94% at day 113 (cohorts 2–5). In cohorts 6 and 7, a single dose of olpasilan effectively reduced mean Lp(a) levels by 75% and 89%, respectively, from baseline at day 43 and by 61% and 80%, respectively, at day 113 (Figure 2). A sharp drop in Lp(a) was observed from day 15, with maximum Lp(a) suppression observed between days 43 and 71. Lp(a) concentrations gradually recovered but remained well below placebo levels at day 225. Single doses of 9 mg or more were responsible for Lp(a) reductions that lasted for 3–6 months.

[0140] The pharmacokinetic parameters of olpasilan in each of the seven administration cohorts are shown in Table 3 below. After single doses of 3, 9, 30, 75, and 225 mg (cohorts 1-5), olpasilan was rapidly absorbed, with a geometric mean C max This occurred within 7.5 hours after administration. Geometric mean half-life (t 1 / 2 The AUC values ​​ranged from 3 to 8 hours, and the majority of olpasilan was cleared from the serum within 2 to 3 days. Systemic exposure increased approximately proportionally to the dose, up to a maximum of 225 mg. Orpasilan AUC exposure in subjects with baseline Lp(a) ≥ 200 nmol / L (cohorts 6 and 7) was approximately 18–33% lower than in subjects with baseline Lp(a) ≥ 70 to ≤ 199 nmol / L (cohorts 2 and 4).

[0141] [Table 3]

[0142] Results from the Phase 1 study demonstrated that a single dose of olpasilan was well-tolerated and significantly reduced Lp(a) in adults with elevated Lp(a), with an approximate observed median percentage reduction of >90%. The effect lasted for 3–6 months at doses of 9 mg or higher. In the high Lp(a) group (Lp(a) ≥ 70 to ≤ 199 nmol / L), doses of 75 mg and 225 mg were superimposable in terms of their effect on Lp(a) concentration; similarly, doses of 9 and 30 mg were superimposable. However, in the very high Lp(a) group (Lp(a) ≥ 200 nmol / L), doses of 9 and 75 mg showed a percentage reduction in Lp(a) suppression from baseline compared to the same doses in the high Lp(a) cohort.

[0143] The depth and duration of Lp(a) level suppression observed with this low single dose of olpasilan were significantly better than expected from planned human doses, based on studies of olpasilan in cynomolgus monkeys. Based on efficacy data for olpasilan in cynomolgus monkeys (see, e.g., Example 18 in International Publication No. 2017 / 059223), a planned human dose of 75 mg was predicted to reduce Lp(a) levels by approximately 80% for at least one month. Notably, as previously mentioned, a single dose as low as 9 mg of olpasilan reduced Lp(a) levels by more than 80% for more than three months in human subjects. Single doses of 75 mg and 225 mg of olpasilan suppressed Lp(a) levels by more than 80% for more than six months. Therefore, olpasilan can be administered to human patients requiring Lp(a) reduction at lower doses and longer dosing intervals (up to once every 6 months). Such dosing regimens offer several different benefits, such as improved patient loyalty, reduced drug costs, and reduced volume and frequency of injections.

[0144] Example 2. Orpasiran PK / PD model for designing a dosage plan for optimal Lp(a) reduction. Based on the Phase 1 data described in Example 1, we developed a mathematical model to characterize the pharmacokinetics of olpasilan and Lp(a) suppression in healthy volunteers with elevated plasma Lp(a) levels (≥70 nmol / L to <200 nmol / L for cohorts 1-5, and ≥200 nmol / L for cohorts 6-7). The pharmacokinetics (PK) of olpasilan were described using a PK model that includes first-order absorption from the subcutaneous administration site into the circulatory system, distribution to the liver via asialoglycoprotein receptor (ASGPR) absorption, recycling of olpasilan from the liver back into the circulatory system, and removal via systemic circulation and degradation from the liver. We found that serum exposure to olpasilan correlated with baseline Lp(a). Therefore, the function of regulating olpasilan bioavailability by baseline Lp(a) was also included in the model. By describing the suppression of Lp(a) from baseline using a PK / PD model, the model-predicted olpasilan liver concentration promoted LPA mRNA degradation, leading to decreased Lp(a) production and suppression during periods of sufficient olpasilan concentration in the liver. The relationship between olpasilan liver concentration and LPA mRNA degradation was described using E. max The model was developed using a model. Changes in LPA mRNA concentration were estimated based on the degree of Lp(a) suppression. Baseline Lp(a) levels indicated the Lp(a) synthesis and degradation rates. Higher baseline values ​​were associated with greater synthesis rates.

[0145] During model development and for clinical dosing plan simulations, the following assumptions were made: a) Simulations for Phase II dose selection were performed for a target population with a baseline Lp(a) level of ≥150 nmol / L. However, model parameters were estimated from subjects with baseline Lp(a) values ​​of ≥70 nmol / L to <200 nmol / L, and ≥200 nmol / L. b) We assumed that the inter-subject and intra-subject variability after multiple doses for the Phase 2 population was the same as that estimated for the Phase 1 study subjects; and c) The continuation of Lp(a) suppression was based on the model predicted olpasilan PK / PD half-life in the liver, and the estimation of the response after multiple doses was based on the observed suppression from the Phase 1 study and the accumulation of effects at the end of the dosing interval.

[0146] This model was able to adequately predict a significant observed reduction in olpasilan exposure in subjects with higher baseline Lp(a) levels (≥150 nmol / L). This suggests that higher doses of olpasilan may be essential to achieve targeted Lp(a) suppression in this patient population. Simulations of this model were performed to explore Q3M and Q6M dosing schedules and to extrapolate predicted Lp(a) suppression after multiple doses of olpasilan. For each candidate dosing schedule, the proportion of subjects achieving targeted Lp(a) suppression (≥80% reduction from baseline) and the proportion of subjects achieving absolute Lp(a) value ≤50 nmol / L at the end of the dosing interval were calculated.

[0147] Figures 3A-3F show the predicted Lp(a) levels as a percentage of baseline for Q3M administration of olpasilan at doses of 10 mg, 30 mg, 50 mg, 75 mg, 150 mg, and 225 mg for subjects with a baseline Lp(a) level of ≥150 nmol / L. The model predicts that doses of 10 mg or higher will suppress Lp(a) levels by 80% or more throughout the entire 3-month dosing interval. Table 4 below shows the predicted percentage of subjects that achieve at least an 80% reduction from baseline in Lp(a) levels at different doses of olpasilan administered once every 3 months (Q3M administration) at each dosing interval, while Table 5 shows the predicted percentage of subjects that achieve an absolute Lp(a) level of 50 nmol / L or less with the same olpasilan dosing plan.

[0148] [Table 4]

[0149] [Table 5]

[0150] Model simulations based on Phase 1 data predict that a quarterly (Q3M) dose of 10 mg of olpasilan will result in approximately 42% of subjects with a baseline Lp(a) level of ≥150 nmol / L achieving at least an 80% reduction in Lp(a) from baseline by 12 months. A quarterly dose of 75 mg or more of olpasilan is predicted to achieve an 80% or greater Lp(a) reduction in at least 90% of subjects with a baseline Lp(a) level of ≥150 nmol / L as early as 3 months after receiving a single dose of olpasilan. A similar proportion of subjects is predicted to achieve absolute Lp(a) values ​​of ≥50 nmol / L with these same dosing regimens.

[0151] Furthermore, simulations were performed on a twice-yearly (Q6M) dosing plan model for olpasilan. Figures 4A-4F show the predicted Lp(a) levels as a percentage of baseline for Q6M olpasilan administration at doses of 10 mg, 75 mg, 150 mg, 225 mg, 450 mg, and 675 mg for subjects with a baseline Lp(a) level of ≥150 nmol / L. The model predicts that at least a dose of 75 mg will suppress the Lp(a) level by more than 80% throughout the entire 6-month dosing interval. Table 6 below shows the predicted percentage of subjects that achieve at least an 80% reduction from baseline in Lp(a) levels at different doses of olpasilan administered once every 6 months (Q6M administration) at each dosing interval, while Table 7 shows the predicted percentage of subjects that achieve an absolute Lp(a) level of 50 nmol / L or less with the same olpasilan dosing plan.

[0152] [Table 6]

[0153] [Table 7]

[0154] Modeling data show that a dose of at least 75 mg of olpasilan administered every six months (Q6M) is predicted to reduce Lp(a) levels by at least 80% from baseline in approximately 50% of subjects with baseline Lp(a) levels ≥ 150 nmol / L after only two doses (i.e., 12 months after treatment). The same proportion of patients are also predicted to achieve an absolute Lp(a) level of less than 50 nmol / L one year after treatment with 75 mg of olpasilan administered every six months. A dose of 225 mg of olpasilan administered every six months is predicted to suppress Lp(a) levels by at least 80% in approximately 76% of subjects one year after treatment, while doses of 450 mg or more administered every six months are predicted to suppress Lp(a) levels above this threshold in approximately 90% of subjects one year after treatment.

[0155] Based on recent Mendelian randomized studies, a reduction of 80% or more in Lp(a) levels from baseline is expected to result in clinically meaningful cardiovascular benefits in patients with atherosclerotic cardiovascular disease (Burgess et al., JAMA Cardiol., Vol.3:619-627, 2018; Lamina et al., JAMA Cardiol., Vol.4:575-579, 2019; and Madsen et al., Arterioscler. Thromb. Vasc. Biol., 40:255-266, 2020). Therefore, olpacilan PK / PD modeling focused on identifying olpacilan dosing regimens that could reduce Lp(a) levels significantly beyond this threshold. The results of the olpacilan PK / PD modeling, and the simulations described in this embodiment, demonstrate the following: A 10 mg dose administered once every 3 months or once every 12 weeks resulted in an ≥80% Lp(a) reduction in approximately half (42%) of subjects with a baseline Lp(a) level ≥150 nmol / L by 12 months, and a median Lp(a)% reduction from baseline in approximately 77% of subjects at 6 and 12 months; A dose of 75 mg administered once every 3 months or once every 12 weeks is expected to achieve a ≥80% reduction in Lp(a) from baseline within a range of 2-3 doses in the majority of subjects (94%), and approximately 90% of subjects are expected to achieve an absolute Lp(a) concentration of 50 nmol / L or less with this dosing regimen. A dose of 225 mg administered once every three months or once every 12 weeks is expected to achieve a reduction of ≥80% Lp(a) in 98% of subjects and reduce Lp(a) levels to an absolute concentration of 50 nmol / L or less in 96% of subjects. • For doses of 10 mg or more, administration every 3 months or every 12 weeks results in a suppression of less than 20% of baseline Lp(a) levels throughout the entire 3-month interval in the majority (≧90%) of subjects with baseline Lp(a) levels of 150 nmol / L or higher; and A dose of 225 mg administered once every 6 months or once every 24 weeks resulted in a median Lp(a) reduction from baseline in 88% of subjects, and approximately 74% achieved an absolute Lp(a) concentration of 50 nmol / L or less.

[0156] Example 3. A double-blind, randomized, placebo-controlled phase 2 study to evaluate the efficacy, safety, and tolerability of olpasilan in subjects with elevated lipoprotein (a) levels. The primary objective of this Phase II study is to evaluate the effect of subcutaneous administration of olpacilan every 12 weeks (Q12W) compared to placebo on the percentage change from baseline in Lp(a) levels at 36 weeks after treatment in patients with atherosclerotic cardiovascular disease and elevated Lp(a). A secondary objective of the study is to evaluate the effect of subcutaneous administration of olpacilan at Q12W compared to placebo on the percentage change from baseline in the following: (i) Lp(a) levels at 48 weeks after treatment, (ii) low-density lipoprotein cholesterol (LDL-C) levels at 36 and 48 weeks after treatment, and (iii) apolipoprotein B (ApoB) levels at 36 and 48 weeks after treatment, as well as characterization of the pharmacokinetic properties of olpacilan. Administration of olpacilan every 24 weeks (Q24W) will also be evaluated.

[0157] Approximately 240 participants will be randomized in a 1:1:1:1:1 ratio. Four arms will be treated with olpasilan and one arm with placebo. Randomization will be stratified by screening for Lp(a) ≤ 200 nmol / L vs > 200 nmol / L and by region (Japan vs. non-Japan). The study treatment period will be 48 weeks, with doses administered on day 1, week 12, week 24, and week 36. After week 48, safety follow-up will be continued for ≥ 40 weeks without further administration of olpasilan or placebo, until Lp(a) returns to 80% of baseline (whichever is later). Participants will continue standard care according to local guidelines (including stable lipid-changing therapy) during the treatment period and the long safety follow-up period.

[0158] After signing informed consent, subjects will enter a screening phase (up to 4 weeks), during which eligibility criteria will be evaluated. Eligible subjects are adults aged 18–80 years with atherosclerotic cardiovascular disease whose Lp(a) is >150 nmol / L during the screening period. Specifically, subjects will be enrolled in the study if they meet all of the following key inclusion criteria: • Ages 18-80 During screening by the central lab, Lp(a) > 150 nmol / L • Atherosclerotic cardiovascular disease based on one of the following: • History of coronary artery regeneration via percutaneous coronary intervention (PCI) or coronary artery bypass grafting (CABG); • Diagnosis of coronary artery disease, regardless of whether or not there has been a previous myocardial infarction; • Diagnosis of atherosclerotic cerebrovascular disease; or • Diagnosis of peripheral artery disease • For subjects not receiving lipid-modulation therapy (not required for eligibility), lipid-modulation therapy, including statin doses, must remain stable according to local guidelines for ≥ 4 weeks prior to screening and during screening. Participants will be excluded from the study if they meet any of the following important exclusion criteria: • Estimated glomerular filtration rate (eGFR) during screening <30 mL / min / 1.73 m 2 Severe renal failure is defined as • A history or clinical finding of hepatic dysfunction defined as aspartate aminotransferase (AST) or alanine aminotransferase (ALT) > 3 × upper limit of normal (ULN), or total bilirubin (TBL) > 2 × ULN during screening. • Malignant tumors within the past 5 years prior to the day before (excluding non-melanoma skin cancer, cervical carcinoma in situ, ductal carcinoma in situ, or stage 1 prostate cancer) • Poorly controlled hypertension on day 1 (defined as mean systolic blood pressure of ≥160 mmHg or mean diastolic blood pressure of ≥100 mmHg at rest) • Fasting triglycerides of ≥400 mg / dL (4.5 mmol / L) during screening • Type 1 diabetes or poorly controlled type 2 diabetes, as determined by a glycated hemoglobin (HbA1c) level of ≥8.5% as determined by the central laboratory at the time of screening.

[0159] Eligible participants for the study are those with a baseline Lp(a) >150 nmol / L. This threshold is based on available epidemiological data indicating that Lp(a) >125 nmol / L is considered elevated from general population data (Averna et al., Atheroscler Suppl., Vol.26:16-24, 2017; Nordestgaard and Langsted, J. Lipid Res., Vol.57:1953-75, 2016; Ohro-Melander, J Intern Med., Vol.278:433-46, 2015; Leebmann et al., Circulation, Vol.128:2567-2576, 2013). In addition, the enrollment population needs to have a higher baseline Lp(a) based on the degree of absolute Lp(a) reduction essential to demonstrate the corresponding effect on cardiovascular events. Therefore, the registration criterion of Lp(a) > 150 nmol / L generates a study population with a median Lp(a) of approximately 200 nmol / L, and enables the evaluation of the efficacy and safety of olpasilane in subjects with very high Lp(a) levels.

[0160] Eligible registered participants will be randomized in a 1:1:1:1:1 ratio to one of the following five treatment groups (each group containing approximately 48 participants): Group 1: 10mg Orpasilan Q12W Group 2: 75mg Orpasilan Q12W Group 3: 225mg Orpasilan Q12W Group 4: 225mg Orpasiran Q24W Group 5: Placebo Q12W As described in Example 2, these olpasilan dosing regimens are expected to suppress Lp(a) levels by at least 80% from baseline throughout the entire dosing interval (3 or 6 months) in human subjects with baseline Lp(a) levels >150 nmol / L. Orpasilan is administered by subcutaneous injection at doses of 10 mg, 75 mg, or 225 mg, depending on the assigned treatment group, once every 12 weeks (treatment groups 1-3) or once every 24 weeks (treatment group 4). Samples for evaluating serum Lp(a), LDL-C, and ApoB, as well as other clinical laboratory analytes, are collected from enrolled subjects during screening, before the administration of the first dose of olpasilan, and at weeks 12, 24, 36, and 48, and at various other time points during the study. Blood samples are collected to measure serum concentrations of olpasilan at various time points during the study to evaluate olpasilan pharmacokinetic parameters.

[0161] Screening for Lp(a) is performed in the central laboratory using a turbidimetric immunoassay that is standardized to detect and quantify Lp(a) particles independently of apo(a) isoform size, and is either approved or under investigation, such as the Tina-quant® lipoprotein(a) Gen.2 assay available from Roche Diagnostics. The assay has been recognized as effective for measuring Lp(a) in nmol / L in serum samples with a detection limit of 7 nmol / L and is standardized to nmol / L against the IFCC reference material SRM2B (Marcovina et al., Clin. Chem., Vol. 46: 1946-1967, 2000). Lipid panels and assays for other clinical analytes, such as ApoB, hemoglobin A1C, ALT, AST, and bilirubin, are performed in the central laboratory using standard methods.

[0162] A primary analysis will be conducted if all randomized subjects had the opportunity to complete the 36-week evaluation, or if they completed it early. The treatment duration analysis will be terminated if all subjects had the opportunity to complete the 48-week evaluation, or if they completed it early. The final analysis will be conducted after the last subject has completed long-term safety follow-up and has completed the study, or has completed it early. The primary endpoint (percentage change from baseline in Lp(a) at 36 weeks) will be compared between groups using a repeated measures linear effects model that includes treatment duration, stratification factors, scheduled visits, and the interaction of treatment with scheduled visits. Type I errors will be controlled for multiple comparisons between the active and placebo arms using the Hochberg procedure. Percentage changes from baseline in secondary endpoints, namely Lp(a) at 48 weeks, and ApoB and LDL-C at 36 and 48 weeks, will be analyzed similarly to the primary endpoint. Safety endpoints (e.g., adverse events caused by the treatment) will be summarized descriptively. Baseline Lp(a) is defined as the mean of two recent non-missing Lp(a) values ​​measured by the central laboratory before or on day 1 of the study. If for any reason only one value is available, that value will be used as the baseline.

[0163] A reduction of over 80% in Lp(a) from baseline was observed with a single dose of olpasilan that lasted for more than 3 months (see Example 1). This sustained reduction in Lp(a) is expected to result in clinically meaningful cardiovascular benefits by reducing the risk of cardiovascular events in patients with atherosclerotic cardiovascular disease. Recent Mendelian randomized studies suggest that in individuals with very high baseline Lp(a) concentrations, an 80%–90% reduction in Lp(a) is expected to translate into a clinically meaningful reduction in the risk of cardiovascular events (Burgess et al., JAMA Cardiol., Vol.3:619-627, 2018; Lamina et al.). (al., JAMA Cardiol., Vol.4:575-579, 2019; and Madsen et al., Arterioscler. Thromb. Vasc. Biol., 40:255-266, 2020). Therefore, the results of this study are expected to show that olpasilan, compared to placebo, at all doses tested, results in a significant percentage reduction in Lp(a) from baseline in subjects with elevated Lp(a) in atherosclerotic cardiovascular disease. In particular, olpasilan is expected to effectively reduce Lp(a) levels to below 50 nmol / L in the majority of subjects at a low dose of 10 mg administered once every 12 weeks. This is expected to result in a reduction in the risk of cardiovascular events in such subjects. Orpasilan administered at a dose of 75 mg once every 12 weeks is expected to be a particularly effective dosing regimen based on the PK / PD modeling described in Example 2.

[0164] All publications, patents, and patent applications discussed and cited herein are incorporated herein by reference in their entirety. The disclosed invention is not limited to the specific methodologies, protocols, and materials described herein, and these are subject to change. Furthermore, the terminology used herein is intended solely to describe specific embodiments and is not intended to limit the scope of the appended claims.

[0165] Those skilled in the art will recognize or confirm, through simple routine experimentation, many equivalents to the specific embodiments of the invention described herein. Such equivalents are intended to be encompassed by the following claims. The present invention provides, for example, the following items: (Item 1) A method for treating, mitigating, or preventing atherosclerosis in a patient who requires treatment, mitigation, or prevention of atherosclerosis, comprising administering an LPA RNAi construct to the patient at doses of approximately 9 mg to approximately 675 mg at intervals of at least 8 weeks, wherein the LPA RNAi construct comprises a sense strand containing the sequence of SEQ ID NO: 1, an antisense strand containing the sequence of SEQ ID NO: 2, and a targeting moiety containing an asialoglycoprotein receptor ligand, the targeting moiety being covalently bonded to the 5' end of the sense strand. (Item 2) A method for lowering serum or plasma Lp(a) levels in a patient requiring a reduction in serum or plasma Lp(a) levels, comprising administering an LPA RNAi construct to the patient at doses of approximately 9 mg to approximately 675 mg at intervals of at least 8 weeks, wherein the LPA RNAi construct comprises a sense strand containing the sequence of SEQ ID NO: 1, an antisense strand containing the sequence of SEQ ID NO: 2, and a targeting moiety containing an asialoglycoprotein receptor ligand, the targeting moiety being covalently bonded to the 5' end of the sense strand. (Item 3) The patient described above is diagnosed with or at risk of cardiovascular disease, as described in item 2. (Item 4) The method according to item 3, wherein the cardiovascular disease is coronary artery disease, carotid artery disease, peripheral artery disease, myocardial infarction, cerebrovascular disease, stroke, aortic stenosis, stable or unstable angina, atrial fibrillation, heart failure, hyperlipidemia, heterozygous familial hypercholesterolemia, or homozygous familial hypercholesterolemia. (Item 5) The patient is diagnosed with coronary artery disease, according to the method described in item 2. (Item 6) The patient in question has a history of myocardial infarction, as described in item 2. (Item 7) The patient is diagnosed with acute coronary artery syndrome, according to the method described in item 2. (Item 8) A method for treating, alleviating, or preventing cardiovascular disease in a patient who requires treatment, alleviation, or prevention of cardiovascular disease, comprising administering an LPA RNAi construct to the patient at doses of approximately 9 mg to approximately 675 mg at intervals of at least 8 weeks, wherein the LPA RNAi construct comprises a sense strand containing the sequence of SEQ ID NO: 1, an antisense strand containing the sequence of SEQ ID NO: 2, and a targeting moiety containing an asialoglycoprotein receptor ligand, the targeting moiety being covalently bonded to the 5' end of the sense strand. (Item 9) The method according to item 8, wherein the cardiovascular disease is coronary artery disease, carotid artery disease, peripheral artery disease, myocardial infarction, cerebrovascular disease, stroke, aortic stenosis, stable or unstable angina, atrial fibrillation, heart failure, hyperlipidemia, heterozygous familial hypercholesterolemia, or homozygous familial hypercholesterolemia. (Item 10) A method for reducing the risk of cardiovascular events in patients with atherosclerotic cardiovascular disease, comprising administering an LPA RNAi construct to the patient at doses of approximately 9 mg to approximately 675 mg at intervals of at least 8 weeks, wherein the LPA RNAi construct comprises a sense strand containing the sequence of SEQ ID NO: 1, an antisense strand containing the sequence of SEQ ID NO: 2, and a targeting moiety containing an asialoglycoprotein receptor ligand, the targeting moiety being covalently bonded to the 5' end of the sense strand. (Item 11) The method according to item 10, wherein the cardiovascular event is cardiovascular death, myocardial infarction, stroke, and / or coronary artery regeneration. (Item 12) The patient has a history of coronary artery regeneration, a history of coronary artery bypass grafting, a diagnosis of coronary artery disease, a diagnosis of atherosclerotic cerebrovascular disease, a diagnosis of peripheral artery disease, and / or a history of myocardial infarction, as described in item 10 or 11. (Item 13) The method according to any one of items 10-12, wherein the patient has experienced a myocardial infarction within one year prior to the first administration of the LPA RNAi construct. (Item 14) The patient is hospitalized for acute coronary syndrome or unstable angina, according to any one of items 10-12. (Item 15) The method according to any one of items 1 to 14, wherein the patient has a serum or plasma Lp(a) level of approximately 70 nmol / L or higher prior to the first administration of the LPA RNAi construct. (Item 16) The method according to any one of items 1 to 14, wherein the patient has a serum or plasma Lp(a) level of approximately 100 nmol / L or higher prior to the first administration of the LPA RNAi construct. (Item 17) The method according to any one of items 1 to 14, wherein the patient has a serum or plasma Lp(a) level of approximately 125 nmol / L or higher prior to the first administration of the LPA RNAi construct. (Item 18) The method according to any one of items 1 to 14, wherein the patient has a serum or plasma Lp(a) level of approximately 150 nmol / L or higher prior to the first administration of the LPA RNAi construct. (Item 19) The method according to any one of items 1 to 14, wherein the patient has a serum or plasma Lp(a) level of approximately 175 nmol / L or higher prior to the first administration of the LPA RNAi construct. (Item 20) The method according to any one of items 1 to 14, wherein the patient has a serum or plasma Lp(a) level of approximately 200 nmol / L or higher prior to the first administration of the LPA RNAi construct. (Item 21) The method according to any one of items 1 to 14, wherein the patient has a serum or plasma Lp(a) level of approximately 225 nmol / L or higher prior to the first administration of the LPA RNAi construct. (Item 22) The method according to any one of items 1 to 21, wherein the aforementioned administration interval is approximately 12 weeks. (Item 23) The method according to any one of items 1 to 21, wherein the aforementioned administration interval is approximately 24 weeks. (Item 24) The method according to any one of items 1 to 21, wherein the LPA RNAi construct is administered to the patient once every 12 weeks in a dose of approximately 10 mg to approximately 225 mg. (Item 25) The method according to item 24, wherein the LPA RNAi construct is administered to the patient once every 12 weeks at a dose of approximately 50 mg to approximately 100 mg. (Item 26) The method according to item 24, wherein the LPA RNAi construct is administered to the patient once every 12 weeks at a dose of approximately 150 mg to approximately 225 mg. (Item 27) The method according to item 24, wherein the LPA RNAi construct is administered to the patient once every 12 weeks at a dose of approximately 75 mg. (Item 28) The method according to item 24, wherein the LPA RNAi construct is administered to the patient once every 12 weeks at a dose of approximately 150 mg. (Item 29) The method according to item 24, wherein the LPA RNAi construct is administered to the patient once every 12 weeks at a dose of approximately 225 mg. (Item 30) The method according to any one of items 1 to 21, wherein the LPA RNAi construct is administered to the patient once every 24 weeks in a dose of approximately 225 mg to approximately 675 mg. (Item 31) The method according to item 30, wherein the LPA RNAi construct is administered to the patient once every 24 weeks at a dose of approximately 225 mg. (Item 32) The patient is receiving lipid-lowering therapy, according to any one of items 1 to 31. (Item 33) The lipid-lowering therapy described above is a statin, ezetimibe, a PCSK9 inhibitor, bempedoic acid, or a combination thereof, as described in item 32. (Item 34) The method according to any one of items 1 to 33, wherein the patient has a serum low-density lipoprotein cholesterol (LDL-C) level of approximately 100 mg / dL or less prior to the first administration of the LPA RNAi construct. (Item 35) The method according to any one of items 1 to 33, wherein the patient has a serum low-density lipoprotein cholesterol (LDL-C) level of approximately 70 mg / dL or less prior to the first administration of the LPA RNAi construct. (Item 36) The patient in question had an estimated glomerular filtration rate of approximately 30 mL / min / 1.73 m² prior to the first administration of the LPA RNAi construct. 2 The method described in any one of items 1 to 35 is as described above. (Item 37) The method according to any one of items 1 to 36, wherein the patient has a mean systolic blood pressure at rest less than approximately 160 mmHg and a mean diastolic blood pressure less than approximately 100 mmHg prior to the first administration of the LPA RNAi construct. (Item 38) The method according to any one of items 1 to 37, wherein the patient has a glycated hemoglobin A1c level of less than approximately 8.5% prior to the first administration of the LPA RNAi construct. (Item 39) The method according to any one of items 1 to 38, wherein the patient has a serum triglyceride level of less than approximately 400 mg / dL prior to the first administration of the LPA RNAi construct. (Item 40) The method according to any one of items 1 to 39, wherein the sense strand of the LPA RNAi construct comprises or consists of the sequence of SEQ ID NO: 3, and the antisense strand of the LPA RNAi construct comprises or consists of the sequence of SEQ ID NO: 4. (Item 41) The method according to any one of items 1 to 40, wherein the sense strand of the LPA RNAi construct comprises or consists of a sequence of modified nucleotides according to Sequence ID No. 5, and the antisense strand of the LPA RNAi construct comprises or consists of a sequence of modified nucleotides according to Sequence ID No. 6. (Item 42) The targeting portion of the LPA RNAi construct is: [ka] A method according to any one of items 1 to 41, having the structure described in item 1 to 41. (Item 43) The method according to any one of items 1 to 42, wherein the LPA RNAi construct is orpasilane. (Item 44) The method according to any one of items 1 to 43, wherein administration of the LPA RNAi construct reduces the serum or plasma Lp(a) level in the patient by more than 50% for at least 12 weeks compared to the patient's baseline serum or plasma Lp(a) level. (Item 45) The method according to any one of items 1 to 43, wherein administration of the LPA RNAi construct reduces the patient's serum or plasma Lp(a) level by more than 80% for at least 12 weeks compared to the patient's baseline serum or plasma Lp(a) level. (Item 46) The method according to any one of items 1 to 43, wherein administration of the LPA RNAi construct reduces the serum or plasma Lp(a) level in the patient by more than 90% for at least 12 weeks compared to the patient's baseline serum or plasma Lp(a) level. (Item 47) The method according to any one of items 1 to 43, wherein administration of the LPA RNAi construct reduces the serum or plasma Lp(a) level in the patient to approximately 100 nmol / L or less. (Item 48) The method according to any one of items 1 to 43, wherein administration of the LPA RNAi construct reduces the serum or plasma Lp(a) level in the patient to approximately 75 nmol / L or less. (Item 49) The method according to any one of items 1 to 43, wherein administration of the LPA RNAi construct reduces the serum or plasma Lp(a) level in the patient to approximately 50 nmol / L or less. (Item 50) The method according to any one of items 1 to 49, wherein the LPA RNAi construct is administered to the patient in a pharmaceutical composition containing potassium phosphate and sodium chloride. (Item 51) The method according to any one of items 1 to 50, wherein the LPA RNAi construct is administered to the patient by subcutaneous injection. (Item 52) The method according to item 51, wherein the volume of the injection is approximately 1 mL or less. (Item 53) An LPA RNAi construct for use in a method of treating, mitigating, or preventing atherosclerosis in a patient who requires treatment, mitigation, or prevention of atherosclerosis, the method comprising administering the LPA RNAi construct to the patient at doses of approximately 9 mg to approximately 675 mg at intervals of at least 8 weeks, wherein the LPA RNAi construct comprises a sense strand containing the sequence of SEQ ID NO: 1, an antisense strand containing the sequence of SEQ ID NO: 2, and a targeting moiety containing an asialoglycoprotein receptor ligand, the targeting moiety being covalently bonded to the 5' end of the sense strand, and LPA RNAi construct. (Item 54) An LPA RNAi construct for use in a method to lower serum or plasma Lp(a) levels in a patient requiring a reduction in serum or plasma Lp(a) levels, the method comprising administering the LPA RNAi construct to the patient at doses of approximately 9 mg to approximately 675 mg at intervals of at least 8 weeks, wherein the LPA RNAi construct comprises a sense strand containing the sequence of SEQ ID NO: 1, an antisense strand containing the sequence of SEQ ID NO: 2, and a targeting moiety containing an asialoglycoprotein receptor ligand, the targeting moiety being covalently bonded to the 5' end of the sense strand. (Item 55) The aforementioned patient is diagnosed with cardiovascular disease or is at risk thereof, and the LPA RNAi construct for use as described in item 54. (Item 56) The cardiovascular disease is coronary artery disease, carotid artery disease, peripheral artery disease, myocardial infarction, cerebrovascular disease, stroke, aortic stenosis, stable or unstable angina, atrial fibrillation, heart failure, hyperlipidemia, heterozygous familial hypercholesterolemia, or homozygous familial hypercholesterolemia, as described in item 55 of the LPA RNAi construct for use. (Item 57) The aforementioned patient is diagnosed with chronic kidney disease and is a candidate for use of the LPA RNAi construct described in item 54. (Item 58) The aforementioned patient has a history of myocardial infarction, and the LPA RNAi construct for use as described in item 54. (Item 59) The aforementioned patient is diagnosed with acute coronary syndrome, and LPA is used for the purposes described in item 54. RNAi construct. (Item 60) An LPA RNAi construct for use in a method of treating, mitigating, or preventing cardiovascular disease in a patient requiring treatment, mitigation, or prevention of cardiovascular disease, the method comprising administering the LPA RNAi construct to the patient at doses of approximately 9 mg to approximately 675 mg at intervals of at least 8 weeks, wherein the LPA RNAi construct comprises a sense strand containing the sequence of SEQ ID NO: 1, an antisense strand containing the sequence of SEQ ID NO: 2, and a targeting moiety containing an asialoglycoprotein receptor ligand, the targeting moiety being covalently bonded to the 5' end of the sense strand. (Item 61) The cardiovascular disease is coronary artery disease, carotid artery disease, peripheral artery disease, myocardial infarction, cerebrovascular disease, stroke, aortic stenosis, stable or unstable angina, atrial fibrillation, heart failure, hyperlipidemia, heterozygous familial hypercholesterolemia, or homozygous familial hypercholesterolemia, as described in item 60 of the LPA RNAi construct for use. (Item 62) An LPA RNAi construct used in a method to reduce the risk of cardiovascular events in patients with atherosclerotic cardiovascular disease, the method comprising administering the LPA RNAi construct to the patient at doses of approximately 9 mg to approximately 675 mg at intervals of at least 8 weeks, wherein the LPA RNAi construct comprises a sense strand containing the sequence of SEQ ID NO: 1, an antisense strand containing the sequence of SEQ ID NO: 2, and a targeting moiety containing an asialoglycoprotein receptor ligand, the targeting moiety being covalently bonded to the 5' end of the sense strand. (Item 63) The cardiovascular events are cardiovascular death, myocardial infarction, stroke, and / or coronary artery regeneration, as described in item 62 of the LPA RNAi construct for use. (Item 64) The patient has a history of coronary artery regeneration, a history of coronary artery bypass grafting, a diagnosis of coronary artery disease, a diagnosis of atherosclerotic cerebrovascular disease, a diagnosis of peripheral artery disease, and / or a history of myocardial infarction, and the LPA RNAi construct for use as described in item 62 or 63. (Item 65) The patient has experienced a myocardial infarction within one year prior to the first administration of the LPA RNAi construct, for use as described in any one of items 62-64. (Item 66) The patient is hospitalized for acute coronary syndrome or unstable angina, and the LPA RNAi construct for use as described in any one of items 62-64. (Item 67) The patient has a serum or plasma Lp(a) level of approximately 70 nmol / L or higher prior to the first administration of the LPA RNAi construct, and the LPA RNAi construct for use as described in any one of items 53 to 66. (Item 68) The patient has a serum or plasma Lp(a) level of approximately 100 nmol / L or higher prior to the first administration of the LPA RNAi construct, for use as described in any one of items 53 to 66. (Item 69) The patient is the LPA RNAi construct for use as described in any one of items 53 to 66, wherein the serum or plasma Lp(a) level prior to the first administration of the LPA RNAi construct is approximately 125 nmol / L or higher. (Item 70) The patient has a serum or plasma Lp(a) level of approximately 150 nmol / L or higher prior to the first administration of the LPA RNAi construct, for use as described in any one of items 53 to 66. (Item 71) The patient is the LPA RNAi construct for use as described in any one of items 53 to 66, wherein the serum or plasma Lp(a) level prior to the first administration of the LPA RNAi construct is approximately 175 nmol / L or higher. (Item 72) The patient has a serum or plasma Lp(a) level of approximately 200 nmol / L or higher prior to the first administration of the LPA RNAi construct, for use as described in any one of items 53 to 66. (Item 73) The patient is the LPA RNAi construct for use as described in any one of items 53 to 66, wherein the serum or plasma Lp(a) level prior to the first administration of the LPA RNAi construct is approximately 225 nmol / L or higher. (Item 74) The aforementioned administration interval is approximately 12 weeks, LPA RNAi construct for use as described in any one of items 53 to 73. (Item 75) The aforementioned administration interval is approximately 24 weeks, LPA RNAi construct for use as described in any one of items 53 to 73. (Item 76) The LPA RNAi construct is administered to the patient once every 12 weeks at a dose of approximately 10 mg to approximately 225 mg, for use as described in any one of items 53 to 73. RNAi construct. (Item 77) The LPA RNAi construct is administered to the patient once every 12 weeks at a dose of approximately 50 mg to approximately 100 mg, as described in item 76. (Item 78) The LPA RNAi construct described above is administered to the patient once every 12 weeks at a dose of approximately 150 mg to approximately 225 mg, as described in item 76. (Item 79) The LPA RNAi construct is administered to the patient once every 12 weeks at a dose of approximately 75 mg, as described in item 76. (Item 80) The LPA RNAi construct is administered to the patient once every 12 weeks at a dose of approximately 150 mg, as described in item 76. (Item 81) The LPA RNAi construct is administered to the patient once every 12 weeks at a dose of approximately 225 mg, as described in item 76. (Item 82) The LPA RNAi construct is administered to the patient once every 24 weeks at a dose of approximately 225 mg to approximately 675 mg, as described in any one of items 53 to 73. (Item 83) The LPA RNAi construct is administered to the patient once every 24 weeks at a dose of approximately 225 mg, as described in item 82. (Item 84) The patient is receiving lipid-lowering therapy and is an LPA RNAi construct for use as described in any one of items 53-83. (Item 85) The aforementioned lipid-lowering therapy is a statin, ezetimibe, a PCSK9 inhibitor, bempedoic acid, or a combination thereof, as described in item 84 of the LPA RNAi constructs for use. (Item 86) The patient has a serum low-density lipoprotein cholesterol (LDL-C) level of approximately 100 mg / dL or less prior to the first administration of the LPA RNAi construct, for use as described in any one of items 53 to 85. (Item 87) The patient has a serum low-density lipoprotein cholesterol (LDL-C) level of approximately 70 mg / dL or less prior to the first administration of the LPA RNAi construct, and the LPA RNAi construct for use as described in any one of items 53 to 85. (Item 88) The patient in question had an estimated glomerular filtration rate of approximately 30 mL / min / 1.73 m² prior to the first administration of the LPA RNAi construct. 2 The above describes the LPA RNAi constructs for use as described in any one of items 53-87. (Item 89) The patient has a mean systolic blood pressure at rest less than approximately 160 mmHg and a mean diastolic blood pressure less than approximately 100 mmHg prior to the first administration of the LPA RNAi construct, and the LPA RNAi construct for use as described in any one of items 53 to 88. (Item 90) The patient has a glycated hemoglobin A1c level of less than approximately 8.5% prior to the first administration of the LPA RNAi construct, for use as described in any one of items 53 to 89. (Item 91) The patient has a serum triglyceride level of less than approximately 400 mg / dL prior to the first administration of the LPA RNAi construct, for use as described in any one of items 53 to 90. (Item 92) An LPA RNAi construct for use as described in any one of items 53 to 91, wherein the sense strand of the LPA RNAi construct comprises or consists of the sequence of SEQ ID NO: 3, and the antisense strand of the LPA RNAi construct comprises or consists of the sequence of SEQ ID NO: 4. (Item 93) An LPA RNAi construct for use as described in any one of items 53 to 92, wherein the sense strand of the LPA RNAi construct comprises or consists of a sequence of modified nucleotides according to Sequence ID No. 5, and the antisense strand of the LPA RNAi construct comprises or consists of a sequence of modified nucleotides according to Sequence ID No. 6. (Item 94) The targeting portion of the LPA RNAi construct is: [ka] An LPA RNAi construct having the structure described in any one of items 53 to 93 for use as described in item 53 to 93. (Item 95) The LPA RNAi construct is olpasilane, an LPA RNAi construct for use as described in any one of items 53 to 94. (Item 96) An LPA RNAi construct for use according to any one of items 53 to 95, wherein administration of the LPA RNAi construct reduces the patient's serum or plasma Lp(a) level by more than 50% for at least 12 weeks compared to the patient's baseline serum or plasma Lp(a) level. (Item 97) An LPA RNAi construct for use according to any one of items 53 to 95, wherein administration of the LPA RNAi construct reduces the patient's serum or plasma Lp(a) level by more than 80% for at least 12 weeks compared to the patient's baseline serum or plasma Lp(a) level. (Item 98) An LPA RNAi construct for use according to any one of items 53 to 95, wherein administration of the LPA RNAi construct reduces the serum or plasma Lp(a) level in the patient by more than 90% for at least 12 weeks compared to the patient's baseline serum or plasma Lp(a) level. (Item 99) An LPA RNAi construct for use as described in any one of items 53 to 95, wherein administration of the LPA RNAi construct reduces the serum or plasma Lp(a) level in the patient to approximately 100 nmol / L or less. (Item 100) An LPA RNAi construct for use as described in any one of items 53 to 95, wherein administration of the LPA RNAi construct reduces the serum or plasma Lp(a) level in the patient to approximately 75 nmol / L or less. (Item 101) An LPA RNAi construct for use as described in any one of items 53 to 95, wherein administration of the LPA RNAi construct reduces the serum or plasma Lp(a) level in the patient to approximately 50 nmol / L or less. (Item 102) The LPA RNAi construct is administered to the patient in a pharmaceutical composition containing potassium phosphate and sodium chloride, as described in any one of items 53 to 101. (Item 103) The LPA RNAi construct is administered to the patient by subcutaneous injection, and is an LPA RNAi construct for use as described in any one of items 53 to 102. (Item 104) The volume of the injection is approximately 1 mL or less, LPA RNAi construct for use as described in item 103. (Item 105) Use of an LPA RNAi construct for the preparation of an agent for treating, mitigating, or preventing atherosclerosis in patients who require treatment, mitigation, or prevention of atherosclerosis, wherein the agent is administered at doses of approximately 9 mg to approximately 675 mg at intervals of at least 8 weeks, or is formulated for administration at such doses, and the LPA RNAi construct comprises a sense strand containing the sequence of SEQ ID NO: 1, an antisense strand containing the sequence of SEQ ID NO: 2, and a targeting moiety containing an asialoglycoprotein receptor ligand, wherein the targeting moiety is covalently bonded to the 5' end of the sense strand. (Item 106) Use of an LPA RNAi construct for the preparation of a drug for lowering serum or plasma Lp(a) levels in patients requiring a reduction in serum or plasma Lp(a) levels, wherein the drug is administered at doses of approximately 9 mg to approximately 675 mg at intervals of at least 8 weeks, or is formulated for administration at such doses, and the LPA RNAi construct comprises a sense strand containing the sequence of SEQ ID NO: 1, an antisense strand containing the sequence of SEQ ID NO: 2, and a targeting moiety containing an asialoglycoprotein receptor ligand, wherein the targeting moiety is covalently bonded to the 5' end of the sense strand. (Item 107) The aforementioned patient is diagnosed with or at risk of cardiovascular disease, as described in item 106. (Item 108) The cardiovascular disease is coronary artery disease, carotid artery disease, peripheral artery disease, myocardial infarction, cerebrovascular disease, stroke, aortic stenosis, stable or unstable angina, atrial fibrillation, heart failure, hyperlipidemia, heterozygous familial hypercholesterolemia, or homozygous familial hypercholesterolemia, as described in item 107. (Item 109) The aforementioned patient is diagnosed with chronic kidney disease, and the use described in item 106 applies. (Item 110) The patient in question has a history of myocardial infarction; use as described in item 106. (Item 111) The patient is diagnosed with acute coronary syndrome, as described in item 106. (Item 112) Use of an LPA RNAi construct for the preparation of a medicament for treating, alleviating or preventing cardiovascular disease in a patient who requires treatment, alleviation or prevention of cardiovascular disease, wherein the medicament is administered at a dose of about 9 mg to about 675 mg at an administration interval of at least 8 weeks or is formulated for administration at that dose, and the LPA RNAi construct comprises a sense strand comprising the sequence of SEQ ID NO: 1, an antisense strand comprising the sequence of SEQ ID NO: 2, and a targeting moiety comprising an asialoglycoprotein receptor ligand, wherein the targeting moiety is covalently attached to the 5' end of the sense strand. (Item 113) The use according to item 112, wherein the cardiovascular disease is coronary artery disease, carotid artery disease, peripheral artery disease, myocardial infarction, cerebrovascular disease, stroke, aortic valve stenosis, stable or unstable angina, atrial fibrillation, heart failure, hyperlipidemia, heterozygous familial hypercholesterolemia, or homozygous familial hypercholesterolemia. (Item 114) Use of an LPA RNAi construct for the preparation of a medicament for reducing the risk of cardiovascular events in patients with atherosclerotic cardiovascular disease, wherein the medicament is administered at a dose of about 9 mg to about 675 mg at an administration interval of at least 8 weeks or is formulated for administration at that dose, and the LPA RNAi construct comprises a sense strand comprising the sequence of SEQ ID NO: 1, an antisense strand comprising the sequence of SEQ ID NO: 2, and a targeting moiety comprising an asialoglycoprotein receptor ligand, wherein the targeting moiety is covalently attached to the 5' end of the sense strand. (Item 115) The use according to item 114, wherein the cardiovascular event is cardiovascular death, myocardial infarction, stroke, and / or coronary artery revascularization. (Item 116) The use according to item 114 or 115, wherein the patient has a history of coronary artery revascularization, a history of coronary artery bypass grafting, a diagnosis of coronary artery disease, a diagnosis of atherosclerotic cerebrovascular disease, a diagnosis of peripheral artery disease, and / or a history of myocardial infarction. (Item 117) The use according to any one of items 114 to 116, wherein the patient has experienced myocardial infarction within 1 year before the first administration of the drug. (Item 118) The use according to any one of items 114 to 116, wherein the patient is hospitalized due to acute coronary syndrome or unstable angina. (Item 119) The use according to any one of items 105 to 118, wherein the serum or plasma Lp(a) level of the patient before the first administration of the drug is about 70 nmol / L or higher. (Item 120) The use according to any one of items 105 to 118, wherein the serum or plasma Lp(a) level of the patient before the first administration of the drug is about 100 nmol / L or higher. (Item 121) The use according to any one of items 105 to 118, wherein the serum or plasma Lp(a) level of the patient before the first administration of the drug is about 125 nmol / L or higher. (Item 122) The use according to any one of items 105 to 118, wherein the serum or plasma Lp(a) level of the patient before the first administration of the drug is about 150 nmol / L or higher. (Item 123) The use according to any one of items 105 to 118, wherein the serum or plasma Lp(a) level of the patient before the first administration of the drug is about 175 nmol / L or higher. (Item 124) The use according to any one of items 105 to 118, wherein the serum or plasma Lp(a) level of the patient before the first administration of the drug is about 200 nmol / L or higher. (Item 125) The use according to any one of items 105 to 118, wherein the serum or plasma Lp(a) level of the patient before the first administration of the drug is about 225 nmol / L or higher. (Item 126) The use according to any one of items 105 to 125, wherein the dosing interval is about 12 weeks. (Item 127) The aforementioned administration interval is approximately 24 weeks, as described in any one of items 105 to 125. (Item 128) The drug is administered to the patient once every 12 weeks in a dose of approximately 10 mg to approximately 225 mg, or is formulated for administration at such doses, as described in any one of items 105 to 125. (Item 129) The drug is administered to the patient once every 12 weeks in a dose of approximately 50 mg to approximately 100 mg, or is formulated for administration at such a dose, as described in item 128. (Item 130) The drug is administered to the patient once every 12 weeks in a dose of approximately 150 mg to approximately 225 mg, or is formulated for administration at such a dose, as described in item 128. (Item 131) The drug is administered to the patient once every 12 weeks in a dose of approximately 75 mg, or is formulated for administration at such a dose, as described in item 128. (Item 132) The drug is administered to the patient once every 12 weeks at a dose of approximately 150 mg, or is formulated for administration at that dose, as described in item 128. (Item 133) The drug is administered to the patient once every 12 weeks at a dose of approximately 225 mg, or is formulated for administration at that dose, as described in item 128. (Item 134) The drug is administered to the patient once every 24 weeks in a dose of approximately 225 mg to approximately 675 mg, or is formulated for administration at such doses, as described in any one of items 105 to 125. (Item 135) The drug is administered to the patient once every 24 weeks at a dose of approximately 225 mg, or is formulated for administration at that dose, as described in item 134. (Item 136) The patient is receiving lipid-lowering therapy, and the use is as described in any one of items 105-135. (Item 137) The lipid-lowering therapy described above is a statin, ezetimibe, a PCSK9 inhibitor, bempedoic acid, or a combination thereof, as described in item 136. (Item 138) The use described in any one of items 105 to 137, wherein the patient has a serum low-density lipoprotein cholesterol (LDL-C) level of approximately 100 mg / dL or less prior to the first administration of the drug. (Item 139) The use described in any one of items 105-137, wherein the patient has a serum low-density lipoprotein cholesterol (LDL-C) level of approximately 70 mg / dL or less prior to the first administration of the drug. (Item 140) The patient in question had an estimated glomerular filtration rate of approximately 30 mL / min / 1.73 m² prior to the first administration of the drug. 2 The above applies to any use described in any one of items 105-139. (Item 141) The use described in any one of items 105 to 140, wherein the patient has a mean systolic blood pressure at rest less than approximately 160 mmHg and a mean diastolic blood pressure less than approximately 100 mmHg prior to the first administration of the drug. (Item 142) The use described in any one of items 105-141, wherein the patient has a glycated hemoglobin A1c level of less than approximately 8.5% prior to the first administration of the drug. (Item 143) The use described in any one of items 105-142, wherein the patient has a serum triglyceride level of less than approximately 400 mg / dL prior to the first administration of the drug. (Item 144) The use described in any one of items 105 to 143, wherein the sense strand of the LPA RNAi construct contains or comprises the sequence of SEQ ID NO: 3, and the antisense strand of the LPA RNAi construct contains or comprises the sequence of SEQ ID NO: 4. (Item 145) The sense strand of the LPA RNAi construct comprises or consists of a sequence of modified nucleotides according to SEQ ID NO: 5, and the antisense strand of the LPA RNAi construct comprises or consists of a sequence of modified nucleotides according to SEQ ID NO: 6, the use according to any one of items 105 to 144. (Item 146) The targeting portion of the LPA RNAi construct is:

Chemical formula

Claims

[Claim 1] The invention described in the present specification.

Images