METHODS FOR TREATING ATHEROSCLEROTIC CARDIOVASCULAR DISEASE WITH LPA-TARGETED RNAi CONSTRUCTS

Abstract

To provide methods for treating an atherosclerotic cardiovascular disease with LPA-targeted RNAi constructs.SOLUTION: The present invention relates to methods for treating or preventing an atherosclerotic cardiovascular disease and other conditions associated with elevated levels of lipoprotein (a) (Lp(a)) using RNAi constructs targeting the LPA gene, which encodes apolipoprotein(a) being a component of Lp(a) particles. In particular, the present invention relates to methods for reducing serum Lp(a) levels and reducing a risk of cardiovascular events in patients with elevated levels of Lp(a) comprising administering an LPA-targeted RNAi construct according to specific dosage regimens. Further, pharmaceutical compositions containing the LPA-targeted RNAi constructs for use in the methods are also disclosed.SELECTED DRAWING: None

Description

[Technical field]

[0001] CROSS-REFERENCE TO RELATED APPLICATIONS This application claims the benefit of U.S. Provisional Patent Application No. 63 / 110,309, filed November 5, 2020, which is incorporated by reference in its entirety.

[0002] Description of electronically submitted text files This application has been submitted electronically in ASCII format and contains a Sequence Listing, which is incorporated herein by reference in its entirety. A copy of the Sequence Listing in computer readable format, created on November 1, 2021, is titled 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 methods for lowering serum levels of Lp(a) and reducing the risk of cardiovascular events, such as cardiovascular death, myocardial infarction, stroke, and coronary revascularization, in patients with elevated Lp(a) by administering an LPA-targeting RNAi construct according to a specific dosing regimen. [Background technology]

[0004] Atherosclerotic cardiovascular disease is highly prevalent and remains the leading cause of mortality worldwide, despite widespread use of low-density lipoprotein (LDL)-lowering therapy. Although LDL-lowering therapy reduces the risk of major cardiac events, the residual cardiovascular risk faced by some patients with low LDL levels implies other mechanisms of cardiovascular pathology. Over the past few decades, compelling evidence from epidemiological studies and meta-analyses, Mendelian randomization analyses, and genome-wide association studies has demonstrated that increasing serum Lp(a) concentrations are associated with increased risk of coronary artery disease and atherosclerosis-related disorders (Clarke et al., N. Engl. J. Med., Vol. 361: 2518-2528, 2009; Kamstrup et al., JAMA, Vol. 301: 2331-2339, 2009; Nordestgaard et al., JAMA, Vol. 301: 2331-2339, 2009). 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). It is known that this risk relationship is continuous, with higher Lp(a) levels being proportionally more affected. This association persists even after adjustment for other lipid parameters (Emerging Risk Factors Collaboration, JAMA, Vol. 302: 412-423, 2009).

[0005] Lp(a) is a low-density lipoprotein composed of LDL particles and the glycoprotein apolipoprotein(a) (apo(a)) linked to apolipoprotein B of the LDL particle by a disulfide bond (Schmidt et al., supra). Apo(a) is encoded by the LPA gene and is expressed almost exclusively in primates, including humans. Apo(a) exhibits homology to plasminogen and exists in various isoforms due to size polymorphisms within the gene resulting from a variable number of kringle IV type 2 (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 apo(a) isoforms and plasma levels of Lp(a) particles (Sandholzer et al., Hum. Genet., Vol. 86; 607-614, 1991). Lp(a) contains proinflammatory oxidized phospholipids that contribute to its atherogenic effects (Tsimikas et al., J. Am. Coll. Cardiol., Vol. 63:1724-1734, 2014).

[0006] High plasma Lp(a) concentrations are genetically defined, maintained at a stable level, cannot be regulated by habitual changes (diet, exercise, or other environmental factors), and are not effectively regulated by any of the currently available lipid-lowering drugs. Currently, there are no approved therapies shown to reduce the risk of cardiovascular events by lowering Lp(a). Moderate reductions in Lp(a) (approximately 20-30%) have been observed with proprotein convertase subtilisin / kexin type 9 (PCSK9) inhibitors, 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 used in a few countries with limited access (Julius, J. Cardiovasc. Dev. Dis., Vol. 5: 27-37, 2018). Additionally, apheresis is an invasive and extremely expensive procedure requiring frequent clinic visits, making it unavailable as a long-term treatment for subjects requiring lifelong therapy (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 the apo(a) messenger RNA transcript (AKCEA-APO(a)-LRx; also known as ISIS 681257 and TQJ230) have been developed and are currently undergoing clinical investigation (reviewed in Graham et al., J Lipid Res., Vol. 57:340-351, 2016). When administered at doses of 10 mg, 20 mg, or 40 mg on days 1, 3, 5, 8, 15, and 22 to healthy subjects with baseline Lp(a) levels of 75 nmol / L or higher, the molecule was reported to lower Lp(a) concentrations by an average of 66%, 80%, and 92%, respectively, at day 36, and by an average of 39%, 53%, and 58%, respectively, at day 113 (Viney et al., Lancet, Vol. 388:2239-2253, 2016). In a subsequent Phase 2 study, 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 with baseline Lp(a) levels of 150 nmol / L or higher when administered to patients once every 4 weeks at doses of 20 mg, 40 mg, or 60 mg, respectively (Tsimikas et al., New England Journal of Medicine, Vol. 382: 244-255, 2020). A Phase 3 cardiovascular outcomes trial in which the molecule is administered monthly at a dose of 80 mg is ongoing (ClinicalTrials.gov identifier NCT04023552). However, there remains a need in the art for new therapeutic agents that potently lower Lp(a) levels for extended periods of time, allowing for lower-dose, less frequent dosing regimens 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

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Summary of the Invention

[0009] The present invention is based, in part, on the identification of a therapeutic regimen of LPA-targeting RNAi constructs, particularly olpaciran, for effectively lowering circulating Lp(a) levels for the treatment of atherosclerotic cardiovascular disease. Thus, in some embodiments, the present invention provides a method of lowering serum or plasma Lp(a) levels in a patient in need thereof, comprising administering to the patient an LPA RNAi construct as described herein at a dose of about 9 mg to about 675 mg, with an administration interval of at least 8 weeks. In some such embodiments, the patient to whom the LPA RNAi construct is administered has been diagnosed with or is at risk of developing cardiovascular disease, such as 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. 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, mitigating, or preventing atherosclerosis in a patient in need thereof, or for treating, mitigating, or preventing cardiovascular disease in a patient in need thereof. In such embodiments, the method comprises administering to the patient an LPA RNAi construct as described herein at a dose of about 9 mg to about 675 mg, with an administration interval of at least 8 weeks. The cardiovascular disease to be treated, ameliorated, mitigated, or prevented by the method of the present invention may 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 includes administering to a patient an LPA RNAi construct described herein at a dose of about 9 mg to about 675 mg with a dosing 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 revascularization for ischemia. In certain embodiments, the cardiovascular event is cardiovascular death, myocardial infarction, stroke, and / or coronary revascularization. The atherosclerotic cardiovascular disease patient to whom the LPA RNAi construct is administered may have a history of coronary 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 administered has recently experienced a myocardial infarction event, for example, the patient has experienced a myocardial infarction within one year prior to the first administration of the LPA RNAi construct. In another embodiment, the patient to whom the LPA RNAi construct is administered is hospitalized for acute coronary syndrome or unstable angina.

[0012] The patient to be administered the LPA RNAi construct according to the method of the present invention has elevated serum or plasma levels of Lp(a). In some embodiments, the patient has serum or plasma Lp(a) levels of about 70 nmol / L or more before the first administration of the LPA RNAi construct. In other embodiments, the patient has serum or plasma Lp(a) levels of about 150 nmol / L or more before the first administration of the LPA RNAi construct. In certain embodiments, the patient has serum or plasma Lp(a) levels of about 175 nmol / L or more before the first administration of the LPA RNAi construct. In certain other embodiments, the patient has serum or plasma Lp(a) levels of about 200 nmol / L or more 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 level. The lipid-lowering therapy can be a PCSK9 inhibitor, for example, a PCSK9 antagonist monoclonal antibody (e.g., evolocumab, alirocumab), a statin (e.g., atorvastatin, cerivastatin, fluvastatin, lovastatin, mevastatin, pitavastatin, pravastatin, rosuvastatin, simvastatin), a cholesterol absorption inhibitor (e.g., ezetimibe), bempedoic acid, nicotinic acid (e.g., niacin), fibric acid (e.g., gemfibrozil, fenofibrate), a bile acid adsorbent (e.g., cholestyramine, colestipol, colesevelam), LDL apheresis, or a combination thereof. In these and other embodiments, the patient may have a serum LDL-C level of about 100 mg / dL or less, or about 70 mg / dL or less, prior to the first administration of the LPA RNAi construct.

[0014] In certain embodiments of the methods of the present invention, a fixed dose of the LPA RNAi construct is administered to a patient once every 12 weeks or once every three months. In some such embodiments, the fixed dose can 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, the LPA The RNAi construct is administered to the patient at a fixed dose of about 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 about 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 about 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 about 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 can 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 once every 24 weeks or once every 6 months at a fixed dose of about 225 mg. In other embodiments, the LPA RNAi construct is administered to the patient once every 24 weeks or once every 6 months at a fixed dose of about 300 mg. In certain embodiments, the LPA RNAi construct is administered to the patient once every 24 weeks or once every 6 months at a fixed dose of about 450 mg. In certain other embodiments, the LPA RNAi construct is administered to the patient once every 24 weeks or once every 6 months at a fixed dose of about 675 mg.

[0016] The administration of the LPA RNAi construct to a patient according to the method of the present invention substantially reduces the patient's plasma or serum Lp(a) for an extended period of time. For example, in some embodiments of the method of the present invention, the administration of the LPA RNAi construct reduces the serum or plasma Lp(a) level in the patient 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) level. In other embodiments of the method of the present invention, the 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, at least 16 weeks, or at least 24 weeks, compared to the patient's baseline serum or plasma Lp(a) level. In certain embodiments, the 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 100 nmol / L or less. In some embodiments, the 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 75 nmol / L or less. In another embodiment, administration of an LPA RNAi construct to a patient according to the methods of the invention reduces the patient's plasma or serum Lp(a) levels to below about 50 nmol / L.

[0017] In any embodiment of the methods 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, the antisense strand comprising a region having a sequence complementary to the LPA mRNA sequence. Preferably, the sense strand comprises a sequence sufficiently complementary to that of the antisense strand to form a duplex region of about 15 to about 30 base pairs in length. In certain embodiments of the methods of the present invention, the LPA RNAi construct administered to the patient comprises a sense strand and an antisense strand, each of which is about 19 to about 23 nucleotides in length, the antisense strand comprises a sequence complementary to the LPA mRNA sequence, and the sense strand comprises a sequence complementary to that of the antisense strand. In one such embodiment, the sense and antisense strands of the LPA RNAi construct may each be 21 nucleotides in length and may hybridize to each other to form a duplex region of 21 base pairs in length, such that the RNAi construct has two blunt ends. In another such embodiment, the sense and antisense strands of the LPA RNAi construct can each be 19 nucleotides in length and can hybridize to each other to form a duplex region 19 base pairs long, 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 the patient further comprises a targeting moiety comprising an asialoglycoprotein receptor ligand, the targeting moiety being covalently linked to the sense strand, for example, to the 5' end of the sense strand. The targeting moiety may comprise 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 the patient according to the method of the present invention comprises a sense strand comprising the sequence of SEQ ID NO:1 and an antisense strand comprising the sequence of SEQ ID NO:2. In some embodiments, the LPA RNAi construct comprises a sense strand comprising or consisting of the sequence of SEQ ID NO:3, and an antisense strand comprising or consisting of 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 an embodiment, the LPA RNAi construct comprises a sense strand comprising or consisting of a sequence of modified nucleotides according to SEQ ID NO:5, and an antisense strand comprising or consisting of a sequence of modified nucleotides according to SEQ ID NO:6. In a preferred embodiment, the LPA RNAi construct administered to a patient according to the methods of the present invention is olpaciran.

[0019] The present invention also provides pharmaceutical compositions comprising an LPA RNAi construct, e.g., olpaciran, for use in the methods of the invention described herein. The pharmaceutical compositions may include one or more pharma- ceutically acceptable diluents, carriers, or excipients. In certain embodiments, the pharmaceutical compositions include an LPA RNAi construct (e.g., olpaciran), potassium phosphate buffer, and sodium chloride, and the composition has a pH of about 6.6 to about 7.0, preferably about 6.8. Any of the pharmaceutical compositions 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 (e.g., subcutaneous administration) to a patient according to the methods described herein. In some embodiments, administration of an LPA RNAi construct (e.g., olpaciran), or a pharmaceutical composition comprising an LPA RNAi construct (e.g., olpaciran), to a patient according to the methods of the 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] Use of the LPA RNAi construct in any of the methods disclosed herein or for preparation of a medicament for administration according to any of the methods disclosed herein is specifically contemplated. For example, the present invention includes an LPA RNAi construct for use in a method of treating, attenuating, or preventing atherosclerosis or cardiovascular disease in a patient in need thereof, the method comprising administering to the patient the LPA RNAi construct at a dose of about 9 mg to about 675 mg with an administration interval of at least 8 weeks. The present invention also includes an LPA RNAi construct for use in a method of reducing serum or plasma Lp(a) levels in a patient, the method comprising administering to the patient the LPA RNAi construct at a dose of about 9 mg to about 675 mg with an administration interval of at least 8 weeks. In certain embodiments, the present invention provides an LPA RNAi construct for use in a method for reducing the risk of a cardiovascular event in a patient with atherosclerotic cardiovascular disease, the method comprising administering to the patient the LPA RNAi construct at a dose of about 9 mg to about 675 mg with an administration interval of at least 8 weeks.

[0021] The invention also encompasses the use of an LPA RNAi construct in the preparation of a medicament for treating, attenuating, or preventing atherosclerosis or cardiovascular disease in a patient in need thereof, wherein the medicament is administered or formulated for administration at a dose of about 9 mg to about 675 mg with an administration interval of at least 8 weeks. In some embodiments, the invention provides the use of an LPA RNAi construct in the preparation of a medicament for reducing serum or plasma Lp(a) levels in a patient, wherein the medicament is administered or formulated for administration at a dose of about 9 mg to about 675 mg with an administration interval of at least 8 weeks. In another embodiment, the present invention provides the use of an LPA RNAi construct in the preparation of a medicament for reducing the risk of a cardiovascular event in a patient with atherosclerotic cardiovascular disease, wherein the medicament is administered or formulated for administration at a dose of about 9 mg to about 675 mg with a dosing interval of at least 8 weeks. [Brief description of the drawings]

[0022] [Figure 1] Schematic diagram of the structure of the LPA RNAi construct, Olpaciran. The top strand, listed in the 5' to 3' direction, is the sense strand (SEQ ID NO:5), and the bottom strand, listed in the 3' to 5' direction, is the antisense strand (SEQ ID NO:6). Black circles represent nucleotides with a 2'-O-methyl modification, white circles represent nucleotides with a 2'-deoxy-2'-fluoro modification, and grey circles represent deoxyadenosine nucleotides linked to adjacent nucleotides via a 3'-3' bond (i.e., inverted). Grey lines connecting the circles represent phosphodiester bonds, while black lines connecting the circles represent phosphorothioate bonds. A trivalent GalNAc moiety having the structure shown is represented by R1 and is covalently attached to the 5' end of the sense strand by a phosphorothioate bond. [Diagram 2]1 is a line graph showing the percent change from baseline in plasma Lp(a) levels in human subjects following a single subcutaneous dose of placebo or olpaciran at the indicated doses in each of cohorts 1-7 across study days. Baseline values ​​were the average 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] Predicted Lp(a) levels of baseline are shown as percentages for quarterly (Q3M) administration of olpaciran at doses of 10 mg (FIG. 3A), 30 mg (FIG. 3B), 50 mg (FIG. 3C), 75 mg (FIG. 3D), 150 mg (FIG. 3E), and 225 mg (FIG. 3F) for subjects with baseline Lp(a) levels ≧150 nmol / L. The horizontal line in each graph represents an 80% reduction in Lp(a) levels from baseline. 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 95% prediction intervals represented by shading. The filled circles in FIG. 3D represent observed data for cohort 7 described in Example 1. [Figure 3-2] Predicted Lp(a) levels of baseline are shown as percentages for quarterly (Q3M) administration of olpaciran at doses of 10 mg (FIG. 3A), 30 mg (FIG. 3B), 50 mg (FIG. 3C), 75 mg (FIG. 3D), 150 mg (FIG. 3E), and 225 mg (FIG. 3F) for subjects with baseline Lp(a) levels ≧150 nmol / L. The horizontal line in each graph represents an 80% reduction in Lp(a) levels from baseline. 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 95% prediction intervals represented by shading. The filled circles in FIG. 3D represent observed data for cohort 7 described in Example 1. [Figure 4-1]Predicted Lp(a) levels of baseline are shown as percentages for twice-yearly (Q6M) administration of olpaciran at doses of 10 mg (FIG. 4A), 75 mg (FIG. 4B), 150 mg (FIG. 4C), 225 mg (FIG. 4D), 450 mg (FIG. 4E), and 675 mg (FIG. 4F) for subjects with baseline Lp(a) levels ≧150 nmol / L. The horizontal line in each graph represents an 80% reduction in Lp(a) levels from baseline. 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 95% prediction intervals represented by shading. The filled circles in FIG. 4B represent observed data for cohort 7 described in Example 1. [Figure 4-2] Predicted Lp(a) levels of baseline are shown as percentages for twice-yearly (Q6M) administration of olpaciran at doses of 10 mg (FIG. 4A), 75 mg (FIG. 4B), 150 mg (FIG. 4C), 225 mg (FIG. 4D), 450 mg (FIG. 4E), and 675 mg (FIG. 4F) for subjects with baseline Lp(a) levels ≧150 nmol / L. The horizontal line in each graph represents an 80% reduction in Lp(a) levels from baseline. 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 95% prediction intervals represented by shading. The filled circles in FIG. 4B represent observed data for cohort 7 described in Example 1. DETAILED DESCRIPTION OF THE PREFERRED EMBODIMENTS

[0023] Lp(a) has been reported to be a causative risk factor for various forms of cardiovascular disease, including myocardial infarction, stroke, peripheral arterial 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 is no approved treatment that selectively targets apo(a) to substantially lower Lp(a) levels. The present invention provides a novel administration regimen of an RNAi construct that targets mRNA transcribed from the LPA gene encoding apo(a) protein to sustain the suppression of Lp(a) levels for the treatment or prevention of atherosclerosis and related cardiovascular conditions. A specific LPA-targeting RNAi construct, olpaciran, was observed to reduce Lp(a) concentrations by 71%-96% in human subjects with baseline Lp(a) levels of ≧70 nmol / L after a single dose, with a maximum percent reduction of >90%, with the effect persisting for more than 6 months at single doses of 9 mg or more (see Example 1). Specifically, a single dose of olpaciran as low as 9 mg reduced Lp(a) levels by more than 80% for more than 3 months in human subjects, while single olpaciran doses of 75 mg and 225 mg suppressed Lp(a) levels by more than 80% for more than 6 months. Additionally, olpaciran was well tolerated at these doses, with no serious treatment-related adverse events (see Example 1). The robust and sustained suppression of Lp(a) at this dosage range was unexpected. This is because, based on allometric scaling of olpaciran dose evaluated in cynomolgus monkeys, it was predicted that an 8-fold higher dose (e.g., 75 mg vs. 9 mg) would be required to produce an 80% reduction in Lp(a) over one month.

[0024] Additionally, the depth and duration of Lp(a) suppression in human subjects by olpaciran was surprising given the results reported in human subjects with other nucleic acid therapeutics 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 after 6 months of treatment. However, a weekly dose of 20 mg or a monthly dose of 60 mg of AKCEA-APO(a)-LRx was required to achieve an 80% and 72% reduction in Lp(a) levels, respectively (see Tsimikas et al., New England Journal of Medicine, Vol. 382: 244-255, 2020). In contrast, as described herein, a single dose of olpaciran reduces Lp(a) levels by more than 80% for more than 6 months, allowing olpaciran to be administered at lower doses and at longer dosing intervals, for example, once every 3 months or once every 6 months. Thus, the methods of the present invention provide significant improvements in human treatment of atherosclerotic cardiovascular disease, including, for example, improved patient adherence, reduced drug costs, and reduced injection volume and frequency. Thus, in certain embodiments, the present invention provides a method of treating, preventing, or reducing the risk of developing cardiovascular disease in a patient in need thereof, comprising administering to the patient an effective amount of an LPA RNAi construct according to a particular dosing regimen described herein.

[0025] Atherosclerosis is a disease in which plaque, composed of fatty substances, cholesterol, calcium, fibrin, and cellular waste products, builds up in various arteries in the body. Over time, the plaque hardens and narrows the lumen of the artery, thereby 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, based on the particular artery affected by atherosclerotic plaque buildup. For example, coronary artery disease occurs when plaque builds up in the coronary arteries and partially blocks blood flow to the heart, which can lead to angina and myocardial infarction. Atherosclerotic plaque buildup 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, can cause transient ischemic attacks or strokes. When plaques build up in the major arteries that supply blood to the limbs and pelvis, peripheral arterial disease can result, leading to abdominal aortic aneurysms and limb ischemia (causing paralysis and pain). When atherosclerotic plaques build up in the renal arteries, chronic kidney disease can develop and over time lead to reduced kidney function, which can result in renal failure. Lp(a) is an atherogenic lipoprotein. Its elevated levels have been associated with increased risk of, among others, coronary artery disease, peripheral arterial disease, myocardial infarction, and stroke. The methods of the present invention are useful for treating, attenuating, or preventing atherosclerosis in patients by lowering circulating Lp(a) levels. Thus, in some embodiments, the present invention provides a method of treating, attenuating, or preventing atherosclerosis in a patient in need thereof, comprising administering to the patient an effective amount of an LPA RNAi construct according to any of the dosing regimens described herein. In one embodiment, the invention includes the use of any of the LPA RNAi constructs described herein for the preparation of a medicament for treating, mitigating, or preventing atherosclerosis in a patient in need thereof, wherein the medicament is administered according to, or formulated for administration according to, any of the dosing regimens described herein.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 of treating, alleviating, or preventing atherosclerosis in a patient in need thereof, the method comprising administering the LPA RNAi construct according to any of the dosing regimens described herein.

[0026] In certain embodiments, the present invention also provides a method for treating, alleviating, ameliorating, or preventing cardiovascular disease in a patient in need thereof, comprising administering to the patient an effective amount of an LPA RNAi construct according to any of the dosage regimens described herein. In some embodiments, the present invention includes the use of any of the LPA RNAi constructs described herein for the preparation of a medicament for treating, alleviating, or preventing cardiovascular disease in a patient in need thereof, the medicament being administered or formulated for administration according to any of the dosage regimens described herein. In other embodiments, the present invention provides an LPA RNAi construct, such as any of the LPA RNAi constructs described herein, for use in a method for treating, alleviating, or preventing cardiovascular disease in a patient in need thereof, the method comprising administering an LPA RNAi construct according to any of the dosage regimens described herein. Cardiovascular disease is a class of diseases and conditions that affect blood vessels or the heart, including, but not limited to, myocardial infarction, heart failure, transient ischemic attack, stroke (ischemic and hemorrhagic), atherosclerosis, coronary artery disease, peripheral vascular disease (e.g. peripheral arterial disease), aneurysm (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. Thus, in certain embodiments, the patient to be treated according to the method of the present invention is diagnosed with or at risk of developing cardiovascular disease. 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, high blood pressure, elevated levels of non-HDL cholesterol, elevated levels of triglycerides, diabetes, obesity, or smoking.Diagnosis of atherosclerosis and cardiovascular disease can be made using a variety of methods known to those of skill in the art, which may include one or more of the following: a patient's health check and family history, the patient's risk factors, physical examinations, blood tests measuring various biomarkers such as lipid levels (e.g., LDL-C, triglycerides, Lp(a), glycated hemoglobin A1C, C-reactive protein, apolipoprotein B, cardiac troponin T, etc.), electrocardiograms, echocardiograms, stress tests, chest x-rays, computed tomography (CT) scans (e.g., cardiac CT scans), and angiograms.

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

[0028] In another embodiment, the cardiovascular disease that is treated, alleviated, ameliorated or prevented according to the method of the present invention is coronary artery disease, particularly atherosclerotic cerebrovascular disease.Cerebrovascular disease refers to the disorder that the brain area is temporarily or permanently affected by ischemia or hemorrhage due to one or more malfunctions or complications of cerebral blood vessels.Cerebrovascular disease includes, but is not limited to, transient ischemic attack, stroke (ischemic or hemorrhagic), carotid artery stenosis, vertebral artery stenosis, intracranial artery stenosis, aneurysm and vascular malformation. Signs and symptoms of cerebrovascular disease may include dizziness, nausea, vomiting, very severe headache, confusion, disorientation, memory loss, numbness or weakness of the arms, legs, or face (especially on one side), abnormal or slurred speech, difficulty in understanding, loss of vision or difficulty in seeing, loss of balance, coordination, or ability to walk, carotid stenosis, and a history of transient ischemic attacks and / or carotid revascularization. In one embodiment, the cardiovascular disease to be treated, alleviated, ameliorated, 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 method of the present invention is peripheral artery disease.The signs and symptoms of peripheral artery disease can include: pain or muscle cramps in legs or arms while walking (claudication), numbness or weakness of legs, cold legs or feet, pain in toes, non-healing legs or legs, change in leg color, hair loss or slower hair growth of legs and legs, slower growth of toenails, shiny skin of legs, no pulse or weak pulse of legs or feet, ankle-brachial index ≦ 0.90, and history of abdominal aortic aneurysm, abdominal aortic treatment (percutaneous or surgical), and / or peripheral artery revascularization (percutaneous or surgical).

[0030] In some embodiments, the administration of LPA RNAi construct according to the method of the present invention is for the treatment of atherosclerosis and other cardiovascular diseases and conditions.The term "treatment" or "treat" used herein refers to the application or administration of LPA RNAi construct to a patient who has or has been diagnosed with atherosclerosis or other cardiovascular diseases, has signs of atherosclerosis or other cardiovascular diseases, is at risk of developing atherosclerosis or other cardiovascular diseases, or is prone to atherosclerosis or other cardiovascular diseases, has one or more symptoms of atherosclerosis or other cardiovascular diseases, is at risk of developing atherosclerosis or other cardiovascular diseases, or is prone to atherosclerosis or other cardiovascular diseases, for the purpose of curing, curing, alleviating, alleviating, modifying, ameliorating, or improving atherosclerosis or other cardiovascular diseases. The term "treatment" includes any improvement in disease in a patient, including slowing or stopping the progression of atherosclerosis or other cardiovascular disease in the patient, reducing the number or severity of symptoms of atherosclerosis or other cardiovascular disease, or increasing the frequency or length of time during which the patient is free of symptoms of atherosclerosis or other cardiovascular disease. As used herein, the term "patient" refers to a mammal, including a human, 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 circulating Lp(a) levels or concentrations (e.g., serum or plasma Lp(a) levels / concentrations) in the patient compared to circulating Lp(a) levels (e.g., baseline Lp(a) levels / concentrations) in the patient prior to administration of the LPA RNAi construct, or compared to circulating Lp(a) levels / concentrations in a patient not receiving the LPA RNAi construct. Thus, in some embodiments, the present invention provides a method of reducing serum or plasma Lp(a) levels (or concentrations) in a patient in need thereof, comprising administering an LPA RNAi construct to the patient according to any of the dosing regimens 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 medicament for reducing serum or plasma Lp(a) levels (or concentrations) in a patient in need thereof, the medicament being administered or formulated for administration according to any of the dosing regimens described herein. 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 of reducing serum or plasma Lp(a) levels (or concentrations) in a patient in need thereof, the method comprising administering the LPA RNAi construct according to any of the dosing regimens described herein. In some embodiments, the patient in need of reducing serum or plasma Lp(a) levels (or concentrations) is a patient diagnosed with or at risk of cardiovascular disease, such as 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 in need of reducing serum or plasma Lp(a) levels (or concentrations) has a history of myocardial infarction or has a family history of myocardial infarction.

[0032] In certain embodiments, the patient who needs to reduce serum or plasma Lp(a) level (or concentration) is diagnosed with acute coronary syndrome. Acute coronary syndrome refers to the condition associated with the sudden reduction of blood flow to the heart, which is often caused by the rupture of atherosclerotic plaque and partial or complete thrombosis of coronary artery. Acute coronary syndrome includes acute myocardial ischemia or infarction, such as non-ST elevation myocardial infarction (NSTEMI) and ST elevation MI (STEMI), and unstable angina. Even if acute coronary syndrome does not initially cause infarction, it is a sign of high risk of infarction and must be diagnosed and treated promptly. Signs and symptoms of acute coronary syndrome typically begin suddenly and may include chest pain (angina) or discomfort, pain radiating from the chest to the shoulders, arms, upper abdomen, back, neck, or jaw, nausea or vomiting, indigestion, shortness of breath, sudden profuse sweating, dizziness, or fainting, unusual or inexplicable fatigue, and feelings of restlessness or anxiety.

[0033] In certain other embodiments, the patient in need of reducing serum or plasma Lp(a) levels (or concentrations) is diagnosed with chronic kidney disease. Chronic kidney disease generally refers to the gradual damage to the kidneys and loss of function. Chronic kidney disease can worsen over time and put the patient at higher risk for other cardiovascular diseases. In one embodiment, the patient to be treated according to the methods of the present invention has stage 3 chronic kidney disease. The stage of kidney disease is determined by the estimated glomerular filtration rate (eGFR), a value based on the amount of creatinine in the blood. Stage 3 chronic kidney disease is defined as a kidney filtration rate of approximately 30 mL / min / 1.73 m 2 ~ approx. 59mL / min / 1.73m 2 Stage 3 chronic kidney disease is characterized by an eGFR of about 15 mL / min / 1.73 m and may be more or less than normal, with some initial symptoms such as swelling in the hands and feet, back pain, and urination. Stage 3 chronic kidney disease patients may also have other health-related problems such as high blood pressure, 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. Stage 4 chronic kidney disease patients have an eGFR of about 15 mL / min / 1.73 m. 2~ approx. 29mL / min / 1.73m 2 and typically presents with more or less symptomatic swelling in the hands and feet, back pain, and urination than is normal.

[0034] In some embodiments, administration of an LPA RNAi construct to a patient according to the methods of the invention reduces the level (or concentration) of Lp(a) in serum or plasma in the patient 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 level (or concentration) of Lp(a) in serum or plasma in the patient before administration of the RNAi construct (e.g., a baseline Lp(a) level or concentration), or compared to the level (or concentration) of Lp(a) in serum or plasma in a patient who has not received the RNAi construct. In these and other embodiments, following administration of the LPA RNAi construct (e.g., administration of a single dose of the LPA RNAi construct), circulating Lp(a) levels or concentrations are reduced in a 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 the LPA RNAi construct (e.g., a single dose of the LPA RNAi construct) reduces serum or plasma Lp(a) levels (or concentrations) in the patient by more than 50% for at least 12 weeks, compared to the patient's baseline serum or plasma Lp(a) levels (or concentrations). Baseline serum or plasma Lp(a) levels (or concentrations) refer to the serum or plasma Lp(a) levels (or concentrations) in the patient before administration of the LPA RNAi construct (i.e., pretreatment levels or concentrations). The baseline level / concentration may be a single measurement taken before the patient receives the LPA RNAi construct, or the baseline level / concentration 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 serum or plasma Lp(a) levels (or concentrations) in a patient by more than 50% for at least 24 weeks, compared to the patient's baseline serum or plasma Lp(a) levels (or concentrations). 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 serum or plasma Lp(a) levels (or concentrations) in a patient by more than 80% for at least 12 weeks, compared to the patient's baseline serum or plasma Lp(a) levels (or concentrations). 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 serum or plasma Lp(a) levels (or concentrations) in a patient by more than 80% for at least 24 weeks, compared to the patient's baseline serum or plasma Lp(a) levels (or concentrations). In certain embodiments of the methods of the present invention, administration of an LPA RNAi construct (e.g., a single dose of an LPA RNAi construct) reduces serum or plasma Lp(a) levels (or concentrations) in a patient by more than 80% compared to the patient's baseline serum or plasma Lp(a) levels (or concentrations) for at least 32 weeks.In some embodiments of the methods of the invention, administration of an LPA RNAi construct (e.g., a single dose of an LPA RNAi construct) reduces serum or plasma Lp(a) levels (or concentrations) in a patient by more than 90% for at least 12 weeks, compared to the patient's baseline serum or plasma Lp(a) levels (or concentrations). In other embodiments of the methods of the invention, administration of an LPA RNAi construct (e.g., a single dose of an LPA RNAi construct) reduces serum or plasma Lp(a) levels (or concentrations) in a patient by more than 90% for at least 16 weeks, compared to the patient's baseline serum or plasma Lp(a) levels (or concentrations).

[0036] In certain embodiments, administration of an LPA RNAi construct to a patient according to the methods of the invention reduces the absolute Lp(a) level (or concentration) in serum or plasma in the patient 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 methods of the invention reduces the absolute Lp(a) level (or concentration) in serum or plasma in the patient to about 125 nmol / L or less. In another embodiment, administration of an LPA RNAi construct to a patient according to the methods of the invention reduces the absolute Lp(a) level (or concentration) in the serum or plasma of the patient to about 100 nmol / L or less. In another embodiment, administration of an LPA RNAi construct to a patient according to the methods of the invention reduces the absolute Lp(a) level (or concentration) in the serum or plasma of the patient to about 75 nmol / L or less. In yet another embodiment, administration of an LPA RNAi construct to a patient according to the methods of the invention reduces the absolute Lp(a) level (or concentration) in the serum or plasma of the patient to about 50 nmol / L or less.

[0037] Although there is a preference to measure 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 methods of the invention may reduce the Lp(a) level (or concentration) in serum or plasma in the patient 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 that is standardized to detect and quantify Lp(a) particles independent of apo(a) isoform size. In these and other embodiments, the assay used to measure Lp(a) levels is normalized to the IFCC reference material SRM2B in nmol / L (Marcovina et al., Clin. Chem., Vol. 46:1946-1967, 2000).

[0039] As described above, elevated levels of circulating Lp(a) are associated with increased risk of cardiovascular disease. Thus, the methods of the present invention are also useful for reducing the risk of cardiovascular events in patients with elevated serum or plasma levels of Lp(a). Thus, in certain embodiments, the present invention provides a method for reducing the risk of cardiovascular events in patients with atherosclerotic cardiovascular disease, comprising administering to the patient an effective amount of an LPA RNAi construct according to any of the dosing regimens 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 medicament for reducing the risk of cardiovascular events in patients with atherosclerotic cardiovascular disease, the medicament being administered or formulated for administration according to any of the dosing regimens described herein. 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 for reducing the risk of cardiovascular events in a patient with atherosclerotic cardiovascular disease, the method comprising administering the LPA RNAi construct according to any of the dosing regimens described herein.

[0040] In some embodiments, the cardiovascular event is one or more of the following: cardiovascular death, myocardial infarction, stroke (e.g., ischemic stroke), coronary revascularization, hospitalization for unstable angina, hospitalization for heart failure, peripheral revascularization, acute limb ischemia, transient ischemic attack, major limb amputation surgery for ischemia, cerebral revascularization (all resulting in death). In certain embodiments, the cardiovascular event is cardiovascular death, myocardial infarction, stroke (e.g., ischemic stroke), and / or coronary revascularization. In some such embodiments, the cardiovascular event is cardiovascular death, myocardial infarction, and / or coronary revascularization. In other such embodiments, the cardiovascular event is myocardial infarction and / or coronary revascularization. In other embodiments, the 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 surgery, and peripheral revascularization for 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 a coronary revascularization.

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

[0042] In certain other embodiments, a patient administered an LPA RNAi construct according to the methods of the invention has 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, a patient administered an LPA RNAi construct according to the methods of the invention has 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, a patient administered an LPA RNAi construct according to the methods of the invention has 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, a patient administered an LPA RNAi construct according to the methods of the invention has an absolute risk reduction of about 2.0% to about 3.0% for any one of cardiovascular death, myocardial infarction, and ischemic stroke.

[0043] In any of the above embodiments, the patient to whom the LPA RNAi construct is administered according to the methods of the present invention to reduce cardiovascular events may have a history of coronary 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 certain embodiments, the patient to whom the LPA RNAi construct is administered according to the methods of the present invention to reduce cardiovascular events has experienced a myocardial infarction. For example, in some such embodiments, the patient to whom the LPA RNAi construct is administered according to the methods of the present invention to reduce cardiovascular events has experienced a myocardial infarction within 1 year, 2 years, 3 years, 4 years, or 5 years of receiving the first administration of the LPA RNAi construct. In one such embodiment, the patient to whom the LPA RNAi construct is administered according to the methods of the present invention to reduce cardiovascular events has experienced a myocardial infarction within 1 year of receiving the first administration of the LPA RNAi construct. In certain other embodiments, the patient to whom the LPA RNAi construct is administered according to the methods of the present invention to reduce cardiovascular events is hospitalized or has recently been hospitalized for acute coronary syndrome or unstable angina.

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

[0045] In less preferred embodiments where circulating Lp(a) levels (or concentrations) are measured in mass concentration units, a patient to whom an LPA RNAi construct is to be administered according to the methods of the invention may have a circulating Lp(a) level (or concentration) of about 30 mg / dL or more, about 35 mg / dL or more, about 40 mg / dL or more, about 45 mg / dL or more, about 50 mg / dL or more, about 55 mg / dL or more, about 60 mg / dL or more, about 65 mg / dL or more, about 70 mg / dL or more, about 75 mg / dL or more, about 90 mg / dL or more, or about 100 mg / dL or more. In one embodiment, a patient is administered an RNAi construct of the invention if the serum or plasma Lp(a) level (or concentration) prior to the first administration of an LPA RNAi construct is about 50 mg / dL or more. In another embodiment, a patient is administered an RNAi construct of the present invention if the serum or plasma Lp(a) level (or concentration) before the first administration of an LPA RNAi construct is about 60 mg / dL or higher. In yet another embodiment, a patient is administered an RNAi construct of the present invention if the serum or plasma Lp(a) level (or concentration) before the first administration of an LPA RNAi construct is about 70 mg / dL or higher. In yet another embodiment, a patient is administered an RNAi construct of the present invention if the serum or plasma Lp(a) level (or concentration) before the first administration of an LPA RNAi construct is about 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 that is standardized to detect and quantify Lp(a) particles independent of apo(a) isoform size. In these and other embodiments, the assay used to measure Lp(a) levels is normalized to the IFCC reference material SRM2B in nmol / L (Marcovina et al., Clin. Chem., Vol. 46:1946-1967, 2000).

[0047] In some embodiments, the patient to whom the LPA RNAi construct is administered according to the method of the present invention may have a serum low-density lipoprotein cholesterol (LDL-C) level within the normal range or may be adjusted to within the normal range through treatment with one or more lipid-lowering therapies.For example, in one embodiment, the patient to whom the LPA RNAi construct is administered according to the method of the present invention has a serum LDL-C level of about 100 mg / dL or less before the first administration of the LPA RNAi construct.In another embodiment, the patient to whom the LPA RNAi construct is administered according to the method of the present invention has a serum LDL-C level of about 70 mg / dL or less before the first administration of the LPA RNAi construct.In a related embodiment, the patient to whom the LPA RNAi construct is administered according to the method of the present invention is undergoing one or more lipid-lowering therapies. Lipid-lowering therapies include, but are not limited to, PCSK9 inhibitors, such as PCSK9 antagonist monoclonal antibodies (e.g., evolocumab, alirocumab) and PCSK9-targeted siRNA (e.g., inclisiran), statins (e.g., atorvastatin, cerivastatin, fluvastatin, lovastatin, mevastatin, pitavastatin, pravastatin, rosuvastatin, simvastatin), cholesterol absorption inhibitors (e.g., ezetimibe), bempedoic acid, nicotinic acid (e.g., niacin), fibric acids (e.g., gemfibrozil, fenofibrate), bile acid adsorbents (e.g., cholestyramine, colestipol, colesevelam), LDL apheresis, or combinations thereof. In certain embodiments, the patient to whom the LPA RNAi construct is to be administered according to the methods of the present invention is receiving a lipid-lowering therapy selected from the group consisting of a PCSK9 antagonist monoclonal antibody, a statin, ezetimibe, bempedoic acid, or a combination thereof.

[0048] In certain embodiments, the patient to be administered the LPA RNAi construct according to the method of the present invention has a serum triglyceride level of less than about 500mg / dL before the first administration of the LPA RNAi construct.For example, the patient can have a serum triglyceride level at baseline (e.g., before the first administration of the LPA RNAi construct) of less than about 400mg / dL, less than about 375mg / dL, less than about 350mg / dL, less than about 325mg / dL, less than about 300mg / dL, less than about 275mg / dL, less than about 250mg / dL, less than about 225mg / dL, less than about 200mg / dL, less than about 175mg / dL, or less than about 150mg / dL.In one embodiment, the patient to be administered the LPA RNAi construct according to the method of the present invention has a serum triglyceride level of less than about 400mg / dL before the first administration of the LPA RNAi construct. In another embodiment, a patient to whom an LPA RNAi construct is to be administered according to the method of the present invention has a serum triglyceride level of about 50 mg / dL to about 400 mg / dL before the first administration of the LPA RNAi construct. In yet another embodiment, a patient to whom an LPA RNAi construct is to be administered according to the method of the present invention has a serum triglyceride level of about 150 mg / dL to about 375 mg / dL before 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 by a standard lipid panel using a blood sample from the patient. In some embodiments, the patient fasts for at least 9 hours, preferably 12 hours, before the blood sample is drawn. Thus, the levels / concentrations of the above-mentioned lipid biomarkers (e.g., LDL-C, triglycerides) can be fasting levels.

[0050] In some embodiments, the patient who will be administered LPA RNAi construct according to the method of the present invention has a glycohemoglobin A1C level that does not indicate untreated or insufficiently controlled type 2 diabetes.For example, the patient who will be administered LPA RNAi construct according to the method of the present invention has a glycohemoglobin A1C level at baseline (e.g., before the first administration of LPA RNAi construct) of less than about 10.0%, less than about 9.5%, less than about 9.0%, less than about 8.5%, less than about 8.0%, less than about 7.5%, less than about 7.0%, less than about 6.5%, less than about 6.0%, or less than about 5.5%.In one embodiment, the patient who will be administered LPA RNAi construct according to the method of the present invention has a glycohemoglobin A1C level of less than about 8.5% before the first administration of LPA RNAi construct. In another embodiment, a patient to whom an LPA RNAi construct is to be administered in accordance with the methods of the present invention has a glycated hemoglobin A1C level of less than about 7.0% prior to the first administration of the LPA RNAi construct.

[0051] In other embodiments, the patient to whom the LPA RNAi construct is administered according to the method of the present invention does not show uncontrolled hypertension in systolic and / or diastolic blood pressure.For example, the patient to whom the LPA RNAi construct is administered according to the method of the present invention has a mean resting systolic blood pressure at baseline (e.g., before the first administration of the LPA RNAi construct) of less than about 180mmHg, less than about 160mmHg, less than about 140mmHg, less than about 135mmHg, less than about 130mmHg, less than about 125mmHg, or less than about 120mmHg, and a mean resting diastolic blood pressure at baseline (e.g., before the first administration of the LPA RNAi construct) of less than about 120mmHg, less than about 110mmHg, less than about 100mmHg, less than about 85mmHg, less than about 90mmHg, or less than about 80mmHg. In one embodiment, the patient to whom the LPA RNAi construct is administered according to the method of the present invention has a resting mean systolic blood pressure of less than about 180 mmHg and a resting mean diastolic blood pressure of less than about 110 mmHg before the first administration of the LPA RNAi construct. In another embodiment, the patient to whom the LPA RNAi construct is administered according to the method of the present invention has a resting mean systolic blood pressure of less than about 160 mmHg and a resting mean diastolic blood pressure of less than about 100 mmHg before the first administration of the LPA RNAi construct. In yet another embodiment, the patient to whom the LPA RNAi construct is administered according to the method of the present invention has a resting mean systolic blood pressure of less than about 140 mmHg and a resting mean diastolic blood pressure of less than about 90 mmHg before the first administration of the LPA RNAi construct.

[0052] In some embodiments, the patient to whom the LPA RNAi construct is administered according to the method of the present invention has no signs of severe renal insufficiency.Thus, in certain embodiments, the patient to whom the LPA RNAi construct is administered according to the method of the present invention has an eGFR at baseline (e.g., before the first administration of the LPA RNAi construct) of at least about 30 mL / min / 1.73 m 2 , at least about 45 mL / min / 1.73 m 2 , at least about 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 In one particular embodiment, a patient to whom an LPA RNAi construct is to be administered according to the methods of the invention has an eGFR of about 30 mL / min / 1.73 m prior to the first administration of the LPA RNAi construct. 2 That's all.

[0053] In other embodiments, the patient to whom the LPA RNAi construct is 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 is administered according to the method of the present invention has an ALT level at baseline (e.g., before the first administration of the LPA RNAi construct) that is 3 times or less than the upper limit of normal (ULN). In a related embodiment, the patient to whom the LPA RNAi construct is administered according to the method of the present invention has an ALT level at baseline (e.g., before the first administration of the LPA RNAi construct) that is 3 times or less than the ULN. In these and other embodiments, patients to whom an LPA RNAi construct is to be administered according to the methods of the invention have a baseline total bilirubin level (e.g., before the first administration of an LPA RNAi construct) of less than or equal to 2x the ULN. In some embodiments, patients to whom an LPA RNAi construct is to be administered according to the methods of the invention have a baseline (e.g., before the first administration of an 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 aspect, the method of the present invention comprises administering to a patient an effective amount of LPA RNAi construct. "Effective amount" refers to an amount sufficient to treat, alleviate or ameliorate cardiovascular disease or one or more symptoms of cardiovascular disease, particularly conditions or symptoms associated with cardiovascular disease, or otherwise prevent, hinder, delay or stop the progression of cardiovascular disease or any other undesirable symptoms associated with cardiovascular disease in any way. Effective amount may also refer to an amount sufficient to reduce the occurrence or severity of sequelae resulting from cardiovascular disease. For example, in some embodiments, an effective amount of LPA RNAi construct is an amount sufficient to reduce the occurrence or severity of cardiovascular events, such as myocardial infarction, stroke, or revascularization of coronary, cerebral, or peripheral arteries, in patients with atherosclerosis or other cardiovascular disease.

[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 that is administered to all patients regardless of patient-specific factors, such as body weight. Thus, the fixed dose is not adjusted by the patient based on the patient's body weight. In some embodiments of the method of the present invention, the LPA RNAi construct can be administered to the patient in a fixed amount of about 9 mg to about 675 mg with an administration interval of at least 8 weeks. For example, a fixed dose of an LPA RNAi construct can be about 9 mg, about 10 mg, about 15 mg, about 30 mg, about 50 mg, about 60 mg, about 70 mg, about 75 mg, about 80 mg, about 90 mg, about 100 mg, about 125 mg, about 150 mg, about 175 mg, about 200 mg, about 225 mg, about 250 mg, about 300 mg, about 350 mg, about 400 mg, about 450 mg, about 500 mg, about 550 mg, about 600 mg, about 650 mg, or about 675 mg, with the doses administered at an interval of at least 8 weeks. Also contemplated are ranges between any and all of these endpoints, for example, a fixed dose of an LPA RNAi construct administered to a patient in the methods of the invention can 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, with the dose administered at an interval of at least 8 weeks.

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

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

[0058] In certain embodiments of the methods of the 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 methods of the invention comprise administering to the patient an LPA RNAi construct 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 methods of the invention comprise administering to the patient an LPA RNAi construct at a fixed dose of about 50 mg to about 100 mg once every 12 weeks or once every 3 months. In yet other embodiments, the methods of the invention comprise administering to the patient an LPA RNAi construct at a fixed dose of about 75 mg to about 225 mg once every 12 weeks or once every 3 months. In yet other embodiments, the methods of the invention comprise administering to the patient an LPA RNAi construct 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 comprises administering to the patient an LPA RNAi construct at a fixed dose of about 10 mg once every 12 weeks or once every 3 months. In another embodiment, the method of the present invention comprises administering to the patient an LPA RNAi construct at a fixed dose of about 30 mg once every 12 weeks or once every 3 months. In another embodiment, the method of the present invention comprises administering to the patient an LPA RNAi construct at a fixed dose of about 75 mg once every 12 weeks or once every 3 months. In another embodiment, the method of the present invention comprises administering to the patient an LPA RNAi construct at a fixed dose of about 100 mg once every 12 weeks or once every 3 months. In another embodiment, the method of the present invention comprises administering to the patient an LPA RNAi construct at a fixed dose of about 125 mg once every 12 weeks or once every 3 months. In yet another embodiment, the method of the present invention comprises administering to the patient an LPA RNAi construct at a fixed dose of about 150 mg once every 12 weeks or once every 3 months. In another embodiment, the method of the invention comprises administering to a patient an LPA RNAi construct at a fixed dose of about 175 mg once every 12 weeks or once every 3 months. In another embodiment, the method of the invention comprises administering to a patient an LPA RNAi construct at a fixed dose of about 200 mg once every 12 weeks or once every 3 months.In yet another embodiment, the methods of the invention comprise administering to a patient an LPA RNAi construct at a fixed dose of about 225 mg once every 12 weeks or once every three months.

[0059] In certain other embodiments of the methods of the 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 methods of the invention comprise administering to the patient an LPA RNAi construct 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 methods of the invention comprise administering to the patient an LPA RNAi construct at a fixed dose of about 225 mg to about 450 mg once every 24 weeks or once every 6 months. In yet other embodiments, the methods of the invention comprise administering to the patient an LPA RNAi construct 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 methods of the invention comprise administering to the patient an LPA RNAi construct at a fixed dose of about 225 mg once every 24 weeks or once every 6 months. In another embodiment, the method of the invention comprises administering to a patient an LPA RNAi construct at a fixed dose of about 300 mg once every 24 weeks or once every 6 months. In yet another embodiment, the method of the invention comprises administering to a patient an LPA RNAi construct at a fixed dose of about 450 mg once every 24 weeks or once every 6 months. In yet another embodiment, the method of the invention comprises administering to a patient an LPA RNAi construct at a fixed dose of about 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 treatment period. The "treatment period" begins with administration of the first dose of the LPA RNAi construct and ends with administration of the final dose of the LPA RNAi construct. The treatment period may be from about 12 weeks to about 240 weeks, about 24 weeks to about 144 weeks, about 3 months to about 60 months, about 6 months to about 48 months, for example, about 12 weeks, about 24 weeks, about 36 weeks, about 48 weeks, about 60 weeks, about 72 weeks, about 84 weeks, about 96 weeks, about 108 weeks, about 120 weeks, about 132 weeks, about 144 weeks, about 156 weeks, about 168 weeks, about 180 weeks, about 192 weeks, about 20 weeks, about 30 weeks, about 40 weeks, about 50 weeks, about 60 weeks, about 70 weeks, about 80 weeks, about 90 weeks, about 100 weeks, about 120 weeks, about 140 weeks, about 150 weeks, about 160 weeks, about 180 weeks, about 192 weeks, about 20 ...20 weeks, about 20 weeks, about 20 weeks, about 20 weeks, about 20 weeks, about 20 weeks, about 20 weeks, about 20 weeks, about 20 weeks, about 20 weeks, about 20 weeks, about 20 weeks, about 2 The treatment period may be about 4 weeks, about 216 weeks, about 228 weeks, about 240 weeks, about 3 months, about 6 months, about 9 months, about 12 months, about 15 months, about 18 months, about 21 months, about 24 months, about 27 months, about 30 months, about 33 months, about 36 months, about 39 months, about 42 months, about 45 months, about 48 months, about 51 months, about 54 months, about 57 months, or about 60 months. In some embodiments, the treatment period is about 48 weeks. In other embodiments, the treatment period is about 192 weeks. In yet other embodiments, the treatment period is about 12 months. In yet other embodiments, the treatment period is about 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 particular embodiment, the LPA RNAi construct is administered for a treatment period of at least about 36 weeks to provide a statistically significant percent reduction from baseline in serum or plasma Lp(a) levels compared to subjects not receiving the LPA RNAi construct. In another particular embodiment, the LPA RNAi construct is administered for a treatment period of at least about 48 weeks to provide a statistically significant percent reduction from baseline in serum or plasma Lp(a) levels compared to subjects not receiving the LPA RNAi construct.

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

[0062] Double-stranded RNA molecules may contain chemical modifications to ribonucleotides, including modifications to the ribose sugar, base, or backbone components of the ribonucleotides, such as those described herein or known in the art. Any modification used in double-stranded RNA molecules (e.g., siRNA, shRNA, etc.) is encompassed by the term "double-stranded RNA" for 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 comprising the first sequence can hybridize to a polynucleotide comprising the second sequence to form a duplex region. Other such conditions may include moderate or stringent hybridization conditions known to those skilled in the art. A first sequence is considered to be fully complementary (100% complementary) to a second sequence if the polynucleotide comprising the first sequence base pairs with a polynucleotide comprising 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. Percent complementarity can be calculated by dividing the number of bases in a first sequence that are complementary to the bases at corresponding positions in a second sequence or target sequence by the total length of the first sequence. Also, if there are no more than 5, 4, 3, or 2 mismatches throughout the entire 30 base pair duplex region when the two sequences hybridize, one sequence can be said to be substantially complementary to the other sequence. In general, if any nucleotide overhangs are present as defined herein, the sequence of such overhangs is not taken into account when determining the degree of complementarity between two sequences. As an example, a 21 nucleotide long sense strand and a 21 nucleotide long antisense strand that hybridize to form a 19 base pair duplex region with a 2 nucleotide overhang at the 3' end of each strand would be considered to be fully complementary as the term is used herein.

[0064] In some embodiments, a region of the antisense strand comprises a sequence that is substantially or completely complementary to a region of the target LPA RNA sequence (e.g., LPA mRNA). In such embodiments, the sense strand may comprise a sequence that is completely complementary to the sequence of the antisense strand. In other such embodiments, the sense strand may comprise a sequence that is substantially complementary to the sequence of the antisense strand, such as a sequence that has 1, 2, 3, 4, or 5 mismatches in the duplex region formed by the sense strand and the antisense strand. In certain embodiments, it is preferred that any mismatches occur in the terminal regions (e.g., within 6, 5, 4, 3, or 2 nucleotides of the 5'-end and / or 3'-end of the strand). In one embodiment, any mismatches in the duplex region formed by the sense strand and the antisense strand occur within 6, 5, 4, 3, or 2 nucleotides of the 5'-end of the antisense strand.

[0065] In certain embodiments, the sense strand and antisense strand of double-stranded RNA can be two separate molecules that hybridize to form a duplex region, but are otherwise separate.This type of double-stranded RNA molecule that is formed from two separate strands is called "small interfering RNA" or "short interfering RNA" (siRNA).Therefore, in some embodiments, the LPA RNAi construct used in the method of the present invention comprises siRNA.

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

[0067] The LPA RNAi construct used in the method of the present invention comprises a sense strand and an antisense strand, and the antisense strand comprises a region having a sequence that is substantially or completely complementary to the LPA messenger RNA (mRNA) sequence. As used herein, "LPA mRNA sequence" refers to the messenger RNA sequence, including allelic variants and splice variants, that encode apo(a) protein, including variants or isoforms of apo(a) protein from any species (e.g., non-human primates, humans). The LPA gene (also known as AK38, APOA, and LP) encodes apo(a) protein, which is the major 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 is highly polymorphic due to differences between individuals in the copy number of the kringle IV type 2 (KIV-2) domain, which can range from 2 to over 40 copies of the gene alleles (see, e.g., Kronenberg and Utermann, J. Intern. Med., Vol. 273:6-30, 2013).

[0068] LPA mRNA sequence also includes the transcript sequence expressed as its complementary DNA (cDNA) sequence.cDNA sequence refers to the sequence of the mRNA transcript expressed as DNA bases (e.g., guanine, adenine, thymine, and cytosine) rather than RNA bases (e.g., guanine, adenine, uracil, and cytosine).Thus, the antisense strand of the LPA RNAi construct used in the method of the present invention can comprise a region having a substantially or completely complementary sequence to the target LPA mRNA sequence or LPA cDNA sequence. The LPA mRNA or cDNA sequence may include, but is not limited to, 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). In certain embodiments, the LPA mRNA sequence is the human transcript described in the NCBI database as reference sequence NM_005577.4.

[0069] The sense strand of the LPA RNAi construct typically contains a sequence sufficiently complementary to that of the antisense strand such that the two strands hybridize to form a duplex region under physiological conditions. "Duplex region" refers to regions within two complementary or substantially complementary polynucleotides that base pair with each other by either Watson-Crick base pairing or other hydrogen bonding interactions to create a duplex between the two polynucleotides. The duplex region of the LPA RNAi construct should be of sufficient length to allow the LPA RNAi construct to enter the RNA interference pathway, for example, by binding the Dicer enzyme and / or the RISC complex. For example, in some embodiments, the duplex region is about 15 to about 30 base pairs in length. Other lengths of the duplex region within this range are also suitable, such as 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 certain embodiments, the duplex region is about 17 to about 26 base pairs in length. In other embodiments, the duplex region is about 19 to about 21 base pairs in length. In one embodiment, the duplex region is about 19 base pairs in length. In another embodiment, the duplex region is about 21 base pairs in length.

[0070] For embodiments in which the sense and antisense strands are two separate molecules (e.g., the LPA RNAi construct comprises an siRNA), the sense and antisense strands need not be the same length as the length of the duplex region. For example, one or both strands may be longer than the duplex region and have one or more unpaired nucleotides or mismatches flanking the duplex region. Thus, in some embodiments, the RNAi construct comprises at least one nucleotide overhang. As used herein, a "nucleotide overhang" refers to an unpaired nucleotide or nucleotides at the end of a strand that extend beyond the duplex region. A nucleotide overhang is 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. The length of a nucleotide overhang is 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 comprises 1, 2, 3, 4, 5, or 6 nucleotides. In one particular embodiment, the nucleotide overhang comprises 1-4 nucleotides. In certain embodiments, the nucleotide overhang comprises 2 nucleotides. In certain other embodiments, the nucleotide overhang comprises a single nucleotide. When the nucleotide overhang is present in the antisense strand, the nucleotides in the overhang may be complementary to the target gene sequence, may form a mismatch with the target gene sequence, or may comprise some other sequence (e.g., a polypyrimidine or polypurine sequence, e.g., UU, TT, AA, GG, etc.).

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

[0072] The RNAi construct may comprise a nucleotide overhang at one end of the double-stranded RNA molecule and a blunt end at the other end. "Blunt end" means that the sense strand and the antisense strand are perfectly base-paired at the ends of the molecule and there are no unpaired nucleotides extending beyond the duplex region. In some embodiments, the LPA RNAi construct comprises a nucleotide overhang at the 3' end of the sense strand and a blunt end at the 5' end of the sense strand and the 3' end of the antisense strand. In other embodiments, the LPA RNAi construct comprises a nucleotide overhang at the 3' end of the antisense strand and a blunt end at the 5' end of the antisense strand and the 3' end of the sense strand. In certain embodiments, the LPA RNAi construct comprises a blunt end at both ends of the double-stranded RNA molecule. In such embodiments, the sense strand and the antisense strand have the same length and the duplex region is the same length as the sense strand and the antisense strand (i.e., the molecule is double-stranded over its entire length).

[0073] The sense strand and the antisense strand in the LPA RNAi construct used in the method of the present invention can 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 the 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 the antisense strand form a duplex region having the same length but shorter than the strands, such that the LPA RNAi construct has a two nucleotide overhang. For example, in one embodiment, the LPA RNAi construct comprises (i) a sense strand and an antisense strand each 21 nucleotides long, (ii) a duplex region 19 base pairs long, and (iii) a nucleotide overhang 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 comprises (i) a sense strand and an antisense strand each 23 nucleotides long, (ii) a duplex region 21 base pairs long, and (iii) a nucleotide overhang 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 sense strand and the antisense strand have the same length and form a duplex region over their entire length such that there are no nucleotide overhangs at either end of the double-stranded molecule. In a particular 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 21 nucleotides long, and (ii) a duplex region 21 base pairs long. In another specific embodiment, the LPA RNAi construct used in the methods of the present invention is blunt-ended and comprises (i) sense and antisense strands, each 19 nucleotides in length, and (ii) a duplex region 19 base pairs in length.

[0074] In another embodiment, the sense strand or antisense strand is longer than the other strand, and the two strands form a duplex region with a length equal to the length of the shorter strand, so that the LPA RNAi construct comprises at least one nucleotide overhang.For example, in one embodiment, the LPA RNAi construct used in the method of the present invention comprises (i) a sense strand with a length of 19 nucleotides, (ii) an antisense strand with a length of 21 nucleotides, (iii) a duplex region with a length of 19 base pairs, 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 comprises (i) a sense strand with a length of 21 nucleotides, (ii) an antisense strand with a length of 23 nucleotides, (iii) a duplex region with a length of 21 base pairs, 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 that has one or more chemical modifications to a nucleoside, nucleobase, pentose ring, or phosphate group. Modified nucleotides as used herein do not include ribonucleotides that contain adenosine monophosphate, guanosine monophosphate, uridine monophosphate, and cytidine monophosphate. However, the LPA RNAi construct may contain a combination of modified nucleotides and ribonucleotides. The 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 susceptibility of the molecule to nucleases and other degradative processes. Also, the incorporation of modified nucleotides can enhance the efficacy of the LPA RNAi construct to reduce the expression of the LPA gene.

[0076] In certain embodiments, modified nucleotides have modifications of the ribose sugar. 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 at the 2' position other than OH. Such 2' modifications include 2'H (e.g., deoxyribonucleotides), 2'-O-alkyl (e.g., O-C1-C2-alkyl ... 10 Or O-C1~C 10 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 a modification of a pentose ring in which two atoms of the ring are linked by a bridge to form a second ring, resulting in a bicyclic sugar structure. In some embodiments, a bicyclic sugar modification comprises a bridge between the 4' and 2' carbons of the pentose ring. Nucleotides containing a sugar moiety having a bicyclic sugar modification are referred to herein as bicyclic nucleic acids or BNAs. Exemplary bicyclic sugar modifications include α-L-methyleneoxy (4'-CH2-O-2') bicyclic nucleic acids (BNAs); β-D-methyleneoxy (4'-CH2-O-2') BNAs (also referred to as locked nucleic acids or LNAs); ethyleneoxy (4'-(CH2)2-O-2') BNAs; aminooxy (4'-CH2-ON(R)-2') BNAs; oxyamino (4'-CH2-N(R)-O-2') BNAs; methyl(methyleneoxy) (4'-CH(CH3)-O-2') BNAs (constrained Examples of sugar-modified nucleotides include, but are not limited to, 4'-CH2-S-2' BNAs, methylene-amino (4'-CH2-N(R)-2') BNAs, methyl carbocyclic (4'-CH2-CH(CH3)-2') BNAs, propylene carbocyclic (4'-(CH2)3-2') BNAs, and methoxy(ethyleneoxy) (4'-CH(CH2OMe)-O-2') BNAs (also referred to as constrained MOE or cMOE). These and other sugar-modified nucleotides that can be incorporated into the LPA RNAi constructs used in the methods of the present invention are described in U.S. Pat. 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 deoxyribonucleotides may be terminal nucleotides at the 3' end and / or 5' end of the sense strand or antisense strand. In embodiments where a deoxyribonucleotide is a terminal nucleotide, the terminal nucleotide may be an inverted nucleotide, i.e., linked to the adjacent nucleotide via a 3'-3' internucleotide bond (if on the 3' end of the strand) or via a 5'-5' internucleotide bond (if on the 5' end of the strand) rather than the natural 3'-5' internucleotide bond.

[0079] Both the sense strand and the antisense strand of the LPA RNAi construct may contain one or more modified nucleotides. For example, in some embodiments, the sense strand contains 1, 2, 3, 4, 5, 6, 7, 8, 9, 10 or more modified nucleotides. In certain embodiments, all nucleotides in the sense strand are modified nucleotides. In some embodiments, the antisense strand contains 1, 2, 3, 4, 5, 6, 7, 8, 9, 10 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'-fluoro modified nucleotides, 2'-O-methyl modified nucleotides, or a combination thereof.

[0080] In certain embodiments, modified nucleotides incorporated into one or both strands of the LPA RNAi construct used in the methods of the present invention have a modification of the nucleobase (also referred to herein as "base"). "Modified nucleobase" or "modified base" refers to a base other than the naturally occurring purine bases adenine (A) and guanine (G), and the pyrimidine bases thymine (T), cytosine (C), and uracil (U). Modified nucleobases may be synthetic or naturally occurring modifications, including but not limited to the 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, 6-azouracil, 6-amino ... These 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, particularly 5-bromo, 5-trifluoromethyl, and other 5-substituted uracils and cytosines, 7-methylguanine and adenine, 8-azaguanine and adenine, 7-deazaguanine and adenine, and 3-deazaguanine and adenine.

[0081] In some embodiments, the modified base is a universal base. "Universal base" refers to a base analogue that indiscriminately base pairs with all natural bases in RNA and DNA without changing 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 the LPA RNAi construct include 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 will appreciate that guanine, cytosine, adenine, thymine, and uracil can be replaced by other nucleobases, such as the modified nucleobases described above, without substantially altering the base pairing properties of the polynucleotide that contains the nucleotide with such a substituted nucleobase.

[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 abasic nucleotides. An "abasic nucleotide" or an "abasic nucleoside" is a nucleotide or nucleoside that lacks a nucleobase at the 1' position of the ribose sugar. In certain embodiments, an abasic nucleotide is incorporated at the end of the sense and / or antisense strand of the RNAi construct. In one embodiment, the sense strand contains an abasic nucleotide as a terminal nucleotide at its 3' end, its 5' end, or both its 3' end and 5' end. In another embodiment, the antisense strand contains an abasic nucleotide as a terminal nucleotide at its 3' end, its 5' end, or both its 3' end and 5' end. In those embodiments where the abasic nucleotide is a terminal nucleotide, the terminal nucleotide may be an inverted nucleotide, i.e., linked to the adjacent nucleotide via a 3'-3' internucleotide linkage (if on the 3' end of the strand) or via a 5'-5' internucleotide linkage (if on the 5' end of the strand) rather than the natural 3'-5' internucleotide linkage. The abasic nucleotide may also include a sugar modification, such as any of the sugar modifications described above. In certain embodiments, the abasic nucleotide includes a 2' modification, such as a 2'-fluoro modification, a 2'-O-methyl modification, or a 2'-H (deoxy) modification. In one embodiment, the abasic nucleotide includes a 2'-O-methyl modification. In another embodiment, the abasic nucleotide includes a 2'-H modification (i.e., a deoxy abasic nucleotide).

[0084] The LPA RNAi construct used in the method of the present invention may also include one or more modified internucleotide linkages. As used herein, the term "modified internucleotide linkage" refers to an internucleotide linkage other than the natural 3'-5' phosphodiester linkage. In some embodiments, the modified internucleotide linkage is a phosphorus-containing internucleotide linkage, such as a phosphotriester, an aminoalkyl phosphotriester, an alkyl phosphonate (e.g., methyl phosphonate, 3'-alkylene phosphonate), a phosphinate, a phosphoramidate (e.g., 3'-amino phosphoramidate and aminoalkyl phosphoramidate), a phosphorothioate (P=S), a chiral phosphorothioate, a phosphorodithioate, a thionophosphoramidate, a thionoalkyl phosphonate, a thionoalkyl phosphotriester, and a boranophosphate. In one embodiment, the modified internucleotide linkage is a 2'-5' phosphodiester linkage. In other embodiments, the modified internucleotide linkage is a non-phosphorus-containing internucleotide linkage, and may therefore be referred to as a modified internucleoside linkage. Such non-phosphorus containing linkages include, but are not limited to, morpholino linkages (formed in part from the sugar portion of the nucleoside); siloxane linkages (-O-Si(H)2-O-); sulfide, sulfoxide, and sulfone linkages; formacetyl and thioformacetyl linkages; alkene-containing backbones; sulfamate backbones; methylenemethylimino (-CH2-N(CH3)-O-CH2-) and methylenehydrazino linkages; sulfonate and sulfonamide linkages; amide linkages; and others with mixed N, O, S, and CH2 component moieties. In one embodiment, the modified internucleoside linkage is a peptide-based linkage (e.g., aminoethylglycine) to create peptide nucleic acids or PNAs, such as those described in U.S. Pat. Nos. 5,539,082, 5,714,331, and 5,719,262.Other suitable modified internucleotide and internucleoside linkages that can be used in the LPA RNAi constructs are described in U.S. Pat. No. 6,693,187, U.S. Pat. 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 comprises one or more phosphorothioate internucleotide linkages. The phosphorothioate internucleotide linkages may be present in the sense strand, the antisense strand, or both strands of the LPA RNAi construct. For example, in some embodiments, the sense strand comprises 1, 2, 3, 4, 5, 6, 7, 8 or more phosphorothioate internucleotide linkages. In other embodiments, the antisense strand comprises 1, 2, 3, 4, 5, 6, 7, 8 or more phosphorothioate internucleotide linkages. In yet other embodiments, both strands comprise 1, 2, 3, 4, 5, 6, 7, 8 or more phosphorothioate internucleotide linkages. The LPA RNAi construct may comprise one or more phosphorothioate internucleotide linkages at the 3' end, the 5' end, or both the 3' end and the 5' end of the sense strand, the antisense strand, or both strands. For example, in certain embodiments, the LPA RNAi construct comprises about 1 to about 6 or more (e.g., about 1, 2, 3, 4, 5, 6 or more) consecutive phosphorothioate internucleotide linkages at the 3'-end of the sense strand, the antisense strand, or both strands. In other embodiments, the LPA RNAi construct comprises about 1 to about 6 or more (e.g., about 1, 2, 3, 4, 5, 6 or more) consecutive phosphorothioate internucleotide linkages at the 5'-end of the sense strand, the antisense strand, or both strands. In any of the embodiments in which one or both strands comprise one or more phosphorothioate internucleotide linkages, the remaining internucleotide linkages in the strands can be natural 3' to 5' phosphodiester linkages. For example, in some embodiments, each internucleotide linkage in the sense strand and the antisense strand is selected from phosphodiester and phosphorothioate, and at least one internucleotide linkage is phosphorothioate.

[0086] In some embodiments of the RNAi construct of the present invention, the 5' end of the sense strand, the antisense strand, or both the antisense strand and the sense strand comprises a phosphate moiety. The term "phosphate moiety" as used herein refers to a terminal phosphate group, including unmodified phosphate (-OP=O)(OH)OH) and modified phosphate. Modified phosphate includes phosphate in which one or more of O and OH groups are replaced with H, O, S, N(R), or alkyl (wherein R is H, an amino protecting group, or unsubstituted or substituted alkyl). Exemplary phosphate moieties include, but are not limited to, 5'-monophosphate; 5'-diphosphate; 5'-triphosphate; 5'-guanosine cap (7-methylated or unmethylated); 5'-adenosine cap, or any other modified or unmodified nucleotide cap structure; 5'-monothiophosphate (phosphorothioate); 5'-monodithiophosphate (phosphorodithioate); 5'-alpha-thiotriphosphate; 5'-gamma-thiotriphosphate; 5'-phosphoramidate; 5'-vinyl phosphate; 5'-alkyl phosphonates (e.g., alkyl = methyl, ethyl, isopropyl, propyl, etc.); 5'-cyclopropyl phosphonate, and 5'-alkyl ether phosphonates (e.g., alkyl ether = methoxymethyl, ethoxymethyl, etc.).

[0087] Modified nucleotides that may be incorporated into LPA RNAi constructs suitable for use in the methods of the present invention may have multiple chemical modifications as described herein. For example, modified nucleotides may have modifications to the ribose sugar and modifications to the nucleobase. By way of example, modified nucleotides may include 2' sugar modifications (e.g., 2'-fluoro or 2'-O-methyl) and may include modified bases (e.g., 5-methylcytosine or pseudouracil). In other embodiments, modified nucleotides may include sugar modifications combined with modifications to the 5' phosphate, resulting in modified internucleotide or internucleoside linkages when the modified nucleotide is incorporated into a polynucleotide. For example, in some embodiments, modified nucleotides may include sugar modifications such as 2'-fluoro, 2'-O-methyl, or bicyclic sugar modifications, and 5' phosphorothioate groups. Thus, in some embodiments, one or both strands of the RNAi constructs of the present invention include a combination of 2' modified nucleotides or BNAs and phosphorothioate internucleotide linkages. In certain embodiments, both the sense and antisense strands of an RNAi construct of the invention comprise a combination of 2'-fluoro modified nucleotides, 2'-O-methyl modified nucleotides, and phosphorothioate internucleotide linkages.

[0088] The LPA gene is expressed primarily in the liver. Thus, in certain embodiments, the LPA It is desirable to specifically deliver RNAi constructs to liver cells.Thus, in some embodiments, the LPA RNAi constructs used in the methods of the present invention can comprise a targeting moiety to specifically target LPA RNAi constructs to liver cells (e.g., hepatocytes) using various approaches as described in more detail below.In certain embodiments, the LPA RNAi constructs comprise a targeting moiety that comprises a ligand that binds to 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 a ligand that binds to or interacts with a protein expressed on the surface of liver cells. For example, in certain embodiments, the ligand can comprise an antigen-binding protein (e.g., an antibody or a binding fragment thereof (e.g., Fab, scFv)) that specifically binds to a receptor expressed on liver cells, such as the asialoglycoprotein receptor and the LDL receptor. In a particular embodiment, the ligand comprises an antibody or a binding fragment thereof that specifically binds to ASGR1 and / or ASGR2. In another embodiment, the ligand comprises a Fab fragment of an antibody that specifically binds to ASGR1 and / or ASGR2. A "Fab fragment" is composed of one immunoglobulin light chain (i.e., the light chain variable region (VL) and constant region (CL)) and one immunoglobulin heavy chain CH1 region and variable region (VH). In another embodiment, the ligand comprises a single chain variable antibody fragment (scFv fragment) of an antibody that specifically binds to ASGR1 and / or ASGR2. An "scFv fragment" comprises the VH and VL regions of an antibody, which are present in a single polypeptide chain, and optionally includes a peptide linker between the VH and VL regions that allows the Fv to form the desired structure for antigen binding. Exemplary antibodies and binding fragments thereof that specifically bind to ASGR1 that can be used as asialoglycoprotein receptors in the targeting moiety of the LPA RNAi construct used in the methods of the present invention are described in WO 2017 / 058944 (incorporated herein by reference in its entirety). Other antibodies or binding fragments thereof that specifically bind to ASGR1, LDL receptor, or other liver surface expressed proteins suitable for use as targeting moieties in LPA RNAi constructs are commercially available.

[0090] In certain embodiments, the targeting moiety comprises a carbohydrate. "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 with an oxygen, nitrogen, or sulfur atom bonded to each carbon atom. 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 carbohydrate incorporated in the targeting moiety is a monosaccharide selected from pentose, hexose, or heptose, as well as disaccharides and trisaccharides that include such monosaccharide units. In other embodiments, the carbohydrate incorporated in the targeting moiety is an amino sugar, such as galactosamine, glucosamine, N-acetylgalactosamine, and N-acetylglucosamine.

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

[0092] In certain embodiments, the targeting moiety in the LPA RNAi construct comprises a multivalent carbohydrate moiety. As used herein, a "multivalent carbohydrate moiety" refers to a moiety that comprises two or more carbohydrate units that can bind or interact independently with other molecules. For example, a multivalent carbohydrate moiety comprises two or more binding domains composed of carbohydrates that can bind to two or more different molecules, or to two or more different sites on the same molecule. The valency of the carbohydrate moiety indicates the number of individual binding domains within the carbohydrate moiety. For example, the terms "monovalent", "bivalent", "trivalent", and "tetravalent" in reference to a carbohydrate moiety refer to a carbohydrate moiety that has one, two, three, and four binding domains, respectively. A multivalent carbohydrate moiety may comprise a multivalent lactose moiety, a multivalent galactose moiety, a multivalent glucose moiety, a multivalent N-acetyl-galactosamine moiety, a multivalent N-acetyl-glucosamine moiety, a multivalent mannose moiety, or a multivalent fucose moiety. In some embodiments, the targeting moiety comprises a multivalent galactose moiety. In other embodiments, the targeting moiety comprises a multivalent N-acetyl-galactosamine moiety. In these and other embodiments, the polyvalent carbohydrate moiety can be divalent, trivalent, or tetravalent. In such embodiments, the polyvalent carbohydrate moiety can be biantennary or triantennary. In a particular embodiment, the polyvalent N-acetyl-galactosamine moiety is trivalent or tetravalent. In another particular embodiment, the polyvalent galactose moiety is trivalent or tetravalent.

[0093] The targeting moiety can be directly or indirectly linked or conjugated to the RNA molecule of the LPA RNAi construct. For example, in some embodiments, the targeting moiety is directly covalently linked to the sense or antisense strand of the LPA RNAi construct. In other embodiments, the targeting moiety is covalently linked to the sense or antisense strand of the LPA RNAi construct via a linker. The targeting moiety can be linked to the nucleobase, 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 the purine nucleobase or its derivative can occur at any position, including endocyclic and exocyclic atoms. In certain embodiments, the 2-, 6-, 7-, or 8-position of the purine nucleobase is linked to the targeting moiety. Also, conjugation or binding to the pyrimidine nucleobase or its derivative can occur at any position. In some embodiments, the 2-, 5-, and 6-position of the pyrimidine nucleobase can be linked to the targeting moiety. Conjugation or attachment to the sugar moiety of a nucleotide can occur at any carbon atom. Exemplary carbon atoms of the sugar moiety that can be attached to a targeting moiety include the 2', 3', and 5' carbon atoms. Also, for example, in an abasic nucleotide, the 1' position can be attached to a targeting moiety. Internucleotide linkages can also assist in the attachment of targeting moieties. For phosphorus-containing linkages (e.g., phosphodiester, phosphorothioate, phosphorodithioate, phosphoroamidate, etc.), the targeting moiety can be attached directly to the phosphorus atom or to an O atom, N atom, or S atom that is attached to the phosphorus atom. For amine- or amide-containing internucleoside linkages (e.g., PNA), the targeting moiety can be attached to the nitrogen atom of the amine or amide or to an adjacent carbon atom.

[0094] In some embodiments, the targeting moiety may be attached to the 3' or 5' end of either the sense strand or the antisense strand. In certain preferred embodiments, the targeting moiety is covalently attached to the 5' end of the sense strand. In such embodiments, the targeting moiety is attached to the 5' terminal nucleotide of the sense strand. In these and other embodiments, the targeting moiety is attached at the 5' position of the 5' terminal nucleotide of the sense strand. In embodiments where the inverted abasic nucleotide or inverted deoxyribonucleotide is the 5' terminal nucleotide of the sense strand and is linked to the adjacent nucleotide via a 5'-5' internucleotide bond, the targeting moiety may be attached at the 3' position of the inverted abasic nucleotide or inverted deoxyribonucleotide. In other embodiments, the targeting moiety is covalently attached to the 3' end of the sense strand. For example, in some embodiments, the targeting moiety is attached to the 3' terminal nucleotide of the sense strand. In certain such embodiments, the targeting moiety is attached at the 3' position of the 3' terminal nucleotide of the sense strand. In embodiments where the inverted abasic nucleotide or inverted deoxyribonucleotide is the 3'-terminal nucleotide of the sense strand and is linked to the adjacent nucleotide via a 3'-3' internucleotide bond, the targeting moiety can be attached at the 5'-position of the inverted abasic 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 1, 2, 3, or 4 terminal nucleotides). 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 attached to the sense or antisense strand via a linker. A "linker" is an atom or group of atoms that covalently attaches a ligand to a polynucleotide component of an LPA RNAi construct. The linker can be about 1 to about 30 atoms in length, about 2 to about 28 atoms in length, about 3 to about 26 atoms in length, about 4 to about 24 atoms in length, about 6 to about 20 atoms in length, about 7 to about 20 atoms in length, about 8 to about 20 atoms in length, about 8 to about 18 atoms in length, about 10 to about 18 atoms in length, and about 12 to about 18 atoms in length. In some embodiments, the linker can 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 a compound of interest (e.g., the sense or antisense strand of an RNAi construct), and the other is selected to essentially bind to any selected group, such as a targeting moiety or component thereof, as described herein. In certain embodiments, the linker comprises a chain structure or oligomer of repeating units such as ethylene glycol units or amino acid units. Examples of functional groups typically used in bifunctional linking moieties include, but are not limited to, electrophiles for reacting with nucleophilic groups, and nucleophiles for reacting with electrophilic groups. In some embodiments, bifunctional linking moieties include amino, hydroxyl, carboxylic acid, thiol, unsaturation (e.g., double or triple bonds), and the like.

[0096] Linkers that can be used to attach a targeting moiety to the sense or antisense strand in the LPA RNAi constructs used in the methods of the invention include pyrrolidine, 8-amino-3,6-dioxaoctanoic acid, succinimidyl 4-(N-maleimidomethyl)cyclohexane-1-carboxylate, 6-aminohexanoic acid, substituted C1-C 10 Alkyl, substituted or unsubstituted C2-C 10 Alkenyl, or substituted or unsubstituted C2-C 10The 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.Other types of linkers suitable for connecting targeting moiety to sense or antisense strand in the LPA RNAi construct used in the method of the present invention are known in the art, and can 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 attached to the sense or antisense strand of the LPA RNAi construct used in the method of the present invention comprises a GalNAc moiety, such as a multivalent GalNAc moiety. In some embodiments, the multivalent GalNAc moiety is a trivalent GalNAc moiety and is attached to the 3' end of the sense strand. In other embodiments, the multivalent GalNAc moiety is a trivalent GalNAc moiety and is attached to the 5' end of the sense strand. In yet other embodiments, the multivalent GalNAc moiety is a tetravalent GalNAc moiety and is attached to the 3' end of the sense strand. In yet other embodiments, the multivalent GalNAc moiety is a tetravalent GalNAc moiety and is attached to the 5' end of the sense strand.

[0098] In certain embodiments, the LPA RNAi construct used in the methods of the invention comprises a targeting moiety having the following structure [Structure 1]: [ka] In a preferred embodiment, a targeting moiety having this structure is covalently attached to the 5' end of the sense strand via a phosphorothioate or phosphodiester bond.

[0099] In certain embodiments, an LPA RNAi construct suitable for use in the methods of the invention comprises: a sense strand and an antisense strand, each about 19 to about 23 nucleotides in length, wherein the antisense strand comprises a sequence complementary to the LPA mRNA sequence, and the sense strand comprises a sequence complementary to that of the antisense strand; a targeting moiety comprising an asialoglycoprotein receptor ligand, said targeting moiety being covalently attached to the 5' end of the sense strand; 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 to each other to form a duplex region of 21 base pairs long. In other such embodiments, the sense strand and the antisense strand are each 19 nucleotides long and hybridize to each other to form a duplex region of 19 base pairs long. In other embodiments, the LPA RNAi construct has two nucleotide overhangs. In one such embodiment, the LPA RNAi construct comprises (i) a sense strand and an antisense strand each 21 nucleotides long, (ii) a duplex region of 19 base pairs long, and (iii) a nucleotide overhang 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 targeting moiety comprises a trivalent GalNAc moiety, such as any of the trivalent GalNAc moieties described in U.S. Patent No. 10,246,709, which is incorporated by reference in its entirety. In a preferred embodiment, the targeting moiety has the structure of Structure 1 described above.

[0101] In certain embodiments, the antisense strand of the LPA RNAi construct comprises a sequence substantially complementary or fully complementary to nucleotides 2706-2726 of the human LPA mRNA transcript set forth in NCBI Reference Sequence NM_005577.4, nucleotides 2697-2726 of the human LPA mRNA transcript set forth in NCBI Reference Sequence NM_005577.4, or nucleotides 2708-2725 of the human LPA mRNA transcript set forth in NCBI Reference Sequence NM_005577.4. In such embodiments, the LPA RNAi construct may comprise a sense strand that is substantially complementary or fully complementary to the antisense strand that targets this region. Thus, in these embodiments, the sense strand may comprise a sequence identical to nucleotides 2706-2726, nucleotides 2697-2726, or nucleotides 2725-2708 of the human LPA mRNA transcript set forth in NCBI Reference Sequence NM_005577.4.

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

[0103] Examples of LPA RNAi constructs suitable for use in the methods of the invention are described in WO 2017 / 059223, which is incorporated herein by reference in its entirety. The duplexes AD03851, AD03853, and AD03536 described in WO 2017 / 059223 are particularly useful in the methods of the invention. In certain preferred embodiments, the LPA RNAi constructs used in the methods of the invention comprise a sense strand that comprises or consists of the sequence 5'-CAGCCCCUUAUUGUUAUACGA-3' (SEQ ID NO: 3) and an antisense strand that comprises or consists of the sequence 5'-UCGUAUAACAAUAAGGGGCUG-3' (SEQ ID NO: 4). In related embodiments, the LPA RNAi constructs used in the methods of the invention comprise a sense strand that comprises or consists of the sequence 5'-CAGCCCCUUAUUGUUAUACGA-3' (SEQ ID NO: 4). The RNAi construct comprises a sense strand that comprises or consists of a sequence of modified nucleotides according to the sequence 5'-csagccccuUfAfUfuguuauacgs(invdA)-3' (SEQ ID NO:5), and an antisense strand that comprises or consists of a sequence of modified nucleotides according to the sequence 5'-usCfsgUfaUfaacaaUfaAfgGfgGfcsUfsg-3' (SEQ ID NO:6), where a, g, c, and u are each 2'-O- Af, Gf, Cf, and Uf are 2'-deoxy-2'-fluoro ("2'-fluoro") adenosine, 2'-fluoro guanosine, 2'-fluoro cytidine, and 2'-fluoro uridine, respectively; invdA is inverted deoxyadenosine (3'-3' linked nucleotide), and s is a phosphorothioate bond. In some such embodiments, a targeting moiety having the structure of Structure 1 described herein is covalently attached 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 invention comprises a sense strand comprising the sequence 5'-GCCCCUUAUUGUUAUACGAUU-3' (SEQ ID NO:7) and an antisense strand comprising the sequence 5'-UCGUAUAACAAUAAGGGGCUU-3' (SEQ ID NO:8). In a related embodiment, the LPA RNAi construct used in the method of the invention comprises a sense strand comprising or consisting of a sequence of modified nucleotides according to the sequence 5'-gsccccuUfAfUfuguuauacgauus(invAb)-3' (SEQ ID NO:9) and an antisense strand comprising or consisting of a sequence of modified nucleotides according to the sequence 5'-usCfsgUfaUfaacaaUfaAfgGfgGfcsusu-3' (SEQ ID NO:10), where a, g, c, and u are each 2'- Af, Gf, Cf, and Uf are 2'-deoxy-2'-fluoro ("2'-fluoro") adenosine, 2'-fluoro guanosine, 2'-fluoro cytidine, and 2'-fluoro uridine, respectively; invAb is an inverted abasic nucleotide (a 3'-3' linked nucleotide), and s is a phosphorothioate bond. In some such embodiments, a targeting moiety having the structure of Structure 1 described herein is covalently attached to the 5' end of the sense strand via a phosphorothioate bond.In another related embodiment, the LPA RNAi construct used in the method of the invention comprises a sense strand that comprises or consists of a sequence of modified nucleotides according to the sequence 5'-(invAb)GfcCfcCfuUfAfUfuGfuUfaUfaCfgausu(invAb)-3' (SEQ ID NO:11), and an antisense strand that comprises or consists of a sequence of modified nucleotides according to the sequence 5'-usCfsgsUfaUfaAfCfAfauaAfgGfgGfcusu-3' (SEQ ID NO:12), where a, g, c, and u are 2'-O-methyladenosine, 2'-aminobutyric acid ... Af, Gf, Cf, and Uf are 2'-deoxy-2'-fluoro ("2'-fluoro") adenosine, 2'-fluoro guanosine, 2'-fluoro cytidine, and 2'-fluoro uridine, respectively; invAb is an inverted abasic nucleotide (5'-5' linked nucleotide if on the 5' end of the strand, and 3'-3' linked nucleotide if on the 3' end of the strand), and s is a phosphorothioate linkage. In some of these embodiments, a targeting moiety having the structure of Structure 1 described herein is covalently attached to the 5' end of the sense strand 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 olpaciran. The structure of olpaciran is also shown diagrammatically in FIG. 1 and described in WO 2017 / 059223, where olpaciran is designated duplex no. AD03851. Olpaciran is a double-stranded siRNA molecule comprising two separate strands, a sense strand and an antisense strand, each 21 nucleotides long. The nucleobase sequences of the sense strand and the antisense strand are fully complementary to each other and hybridize to form a 21 base pair long duplex. The nucleotide sequences for the sense strand and the antisense strand of olpaciran are shown in SEQ ID NO: 3 and SEQ ID NO: 4, respectively. Both the sense strand and the antisense strand of olpaciran are composed of modified nucleotides, and the modified sequences for each strand are shown in SEQ ID NO: 5 (sense strand) and SEQ ID NO: 6 (antisense strand). A trivalent GalNAc moiety having the structure of Structure 1 (and represented as R1 in Figure 1) is covalently attached to the 5' end of the sense strand of olpaciran by a phosphorothioate linkage. The term olpaciran refers to the free acid of the compound shown in Figure 1, as well as pharma- ceutically acceptable salts thereof, such as the sodium salt.

[0106] The LPA RNAi construct used in the method of the present invention can be easily produced using techniques known in the art, for example, using conventional solid-phase nucleic acid synthesis. The polynucleotide of the RNAi construct can be assembled on a suitable nucleic acid synthesizer using standard nucleotide or nucleoside precursors (e.g., phosphoramidites). 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 of synthesizing LPA RNAi constructs and selecting targeting moieties are described in the examples of WO2017 / 059223 and U.S. Patent No. 10,246,709, both of which are incorporated herein by reference in their entirety.

[0107] 2' silyl protecting groups can be used in conjunction with acid labile dimethoxytrityl (DMT) at the 5' position of ribonucleosides to synthesize oligonucleotides via phosphoramidite chemistry. Final deprotection conditions are known not to significantly degrade the RNA product. All syntheses can be performed on large, medium, or small scale with any automated or manual synthesizer. Syntheses can also be performed in multiple well plates, columns, or glass slides.

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

[0109] The choice of protecting groups used for the phosphite triesters and phosphotriesters can modify the stability of the triesters towards fluoride: methyl protection of the phosphotriester or phosphite triester can stabilize the bond towards fluoride ion and improve process yields.

[0110] Because ribonucleosides have a reactive 2' hydroxyl substituent, it may be desirable to protect the reactive 2' position in the RNA with a protecting group that is orthogonal to the 5'-O-dimethoxytrityl protecting group (e.g., a protecting group that is stable to acid treatment). Silyl protecting groups meet this requirement and can be easily removed in a 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 of synthesizing the LPA RNAi constructs described herein will be apparent to those skilled in the art. In addition, various synthetic steps can be carried out in alternative sequences or orders to obtain desired compounds. Other synthetic chemical transformations, protective groups (e.g., for hydroxyl, amino, etc. present on bases) and protective group methodologies (protection and deprotection) useful in the synthesis of RNAi constructs are known in the art, including, for example, those described in R.Larock, Comprehensive Organic Transformations, VCH Publishers (1989); TWGreene and PGMWuts, Protective Groups in Organic Synthesis, 2d.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 their subsequent editions. Custom synthesis of RNAi agents is also available from several commercial 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] The LPA RNAi construct is usually administered to a patient in a pharmaceutical composition that may include a pharma- ceutically acceptable carrier, excipient, or diluent. Thus, the present invention also includes pharmaceutical compositions and formulations that include the LPA RNAi construct and a pharma- ceutically acceptable carrier, excipient, or diluent used in the methods of the present invention described herein. For clinical use, pharmaceutical compositions and formulations will be prepared in a form appropriate for the intended use. In general, this will involve preparing compositions that are essentially free of pyrogens as well as other impurities that may be harmful to humans or animals.

[0114] The phrase "pharmacologically acceptable" or "pharmacologically acceptable" refers to molecular entities and compositions that do not produce adverse, allergic, or other untoward reactions when administered to animals or humans. As used herein, "pharmacologically acceptable carriers, excipients, or diluents" include solvents, buffers, solutions, dispersion media, coatings, antibacterial and antifungal agents, isotonicity agents, and absorption delaying agents that are acceptable for use in formulating pharmaceuticals, such as pharmaceuticals suitable for administration to humans. The use of such media and agents for pharmaceutically active substances is well known in the art. Any conventional media or agent is contemplated for use in therapeutic compositions, except insofar as it is incompatible with the LPA RNAi constructs described herein. Supplementary active ingredients may also be incorporated into the compositions, provided that they do not inactivate the LPA RNAi constructs of the compositions.

[0115] Compositions and methods for the formulation of pharmaceutical compositions depend on several criteria, including but not limited to the route of administration, the type and extent 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 delivery route. For example, in certain embodiments, the pharmaceutical composition is formulated for parenteral delivery. Parenteral delivery forms include intravenous, intraarterial, 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 comprises an effective amount of the LPA RNAi construct. The effective amount of the LPA RNAi construct, particularly olpaciran, may be any of the doses described herein.

[0116] The administration of the pharmaceutical composition comprising the LPA RNAi construct according to the method of the present invention can be via any common route, as long as the target tissue is available via 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 the pharmaceutical composition comprising the LPA RNAi construct is administered parenterally to the patient. For example, in certain embodiments, the LPA RNAi construct or the pharmaceutical composition comprising the LPA RNAi construct is administered intravenously. In other embodiments, the LPA RNAi construct or the pharmaceutical composition comprising the LPA RNAi pharmaceutical composition is administered subcutaneously, for example by subcutaneous injection. In such an embodiment, the subcutaneous injection volume is about 2 mL or less, e.g., 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 the embodiment in which the pharmaceutical composition is administered by parenteral injection, the pharmaceutical composition can be administered to the patient by syringe.In some embodiments, the syringe is pre-filled with the pharmaceutical composition.In other embodiments in which the pharmaceutical composition is administered to the patient by parenteral injection, for example subcutaneous injection, the pharmaceutical composition is administered by injection device, including device for self-administration.Such devices are commercially available and include, but are not limited to, autoinjector, administration pen, microinjection pump, on-body injector, and pre-filled syringe. Exemplary devices for administering a pharmaceutical composition comprising an effective amount of an LPA RNAi construct (e.g., olpaciran) according to the methods of the 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 prefilled syringes (BD Sterifill™, BD Hypak™, Baxter pre-filled syringes). In some embodiments, the pharmaceutical composition comprising an effective amount of the LPA RNAi construct (e.g., olpaciran) is administered to the patient by a pre-filled syringe. In other embodiments, the pharmaceutical composition comprising an effective amount of the LPA RNAi construct (e.g., olpaciran) is administered to the patient by an auto-injector. In certain such embodiments, the injection volume of the syringe, auto-injector, 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, auto-injector, or other injection device is about 1 mL or less. In another embodiment, the injection volume of the syringe, autoinjector, or other injection device is about 1 mL. In yet another embodiment, the injection volume of the syringe, autoinjector, or other injection device is about 1.5 mL.

[0118] Colloidal dispersion systems, such as macromolecular 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. A preferred colloidal system used as an in vivo delivery vehicle is a liposome (i.e., an artificial membrane vesicle). The LPA RNAi constructs can be encapsulated in liposomes or complexed to liposomes, particularly cationic liposomes. Alternatively, the LPA RNAi constructs can be complexed to lipids, particularly cationic lipids. Suitable lipids and liposomes include neutral (e.g., dioleoylphosphatidylethanolamine (DOPE), dimyristoylphosphatidylcholine (DMPC), and dipalmitoylphosphatidylcholine (DPPC)), distearoylphosphatidylcholine), anionic (e.g., dimyristoylphosphatidylglycerol (DMPG)), and cationic (e.g., dioleoyltetramethylaminopropyl (DOTAP) and dioleoylphosphatidylethanolamine (DOTMA)). The preparation and use of such colloidal dispersion systems is well known in the art. Also, exemplary formulations are disclosed in U.S. Pat. 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 WO 03 / 093449.

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

[0120] Pharmaceutical compositions containing LPA RNAi constructs suitable for injection use include, for example, sterile aqueous solutions or dispersions, and sterile powders for extemporaneous preparation of sterile solutions or dispersions for injection. Generally, these preparations are sterile and fluid to the extent that they can be easily injected. Preparations must be stable under the conditions of manufacture and storage, and must be protected against the contaminating action of microorganisms such as bacteria and fungi. Suitable solvents or dispersion media may contain, for example, water, ethanol, polyol (for example, glycerol, propylene glycol, liquid polyethylene glycol, etc.), suitable mixtures thereof, and vegetable oils. Proper fluidity can be maintained, for example, by the use of a coating such as lecithin, by maintaining the required particle size in the case of dispersion, and by the use of surfactants. Prevention of the action of microorganisms can be brought about by various antibacterial and antifungal agents, for example, parabens, chlorobutanol, phenol, sorbic acid, thimerosal, etc. In many cases, it is preferable to include an isotonic agent, for example, sugar or sodium chloride. Prolonged absorption of the injectable compositions can be brought about by the use in the compositions of agents delaying absorption, for example, aluminum monostearate and gelatin.

[0121] Sterile injection solution can be prepared by incorporating the active compound in the appropriate amount in a solvent with any other ingredients as required (for example, as listed above), and then sterilizing by filtration.Generally, dispersion is prepared by incorporating various sterilized active ingredients into a sterile vehicle containing a basic dispersion medium and other desired ingredients, for example, the ingredients listed above.In the case of sterile powder for preparing sterile injection solution, the preferred preparation method includes vacuum drying and freeze-drying techniques, which obtain a powder of active ingredient plus any additional desired ingredients from its previously sterile-filtered solution.

[0122] The compositions used in the methods of the present invention can generally be formulated in neutral or salt form. Pharmaceutically acceptable salts include, for example, acid addition salts (formed with free amino groups) 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 free carboxyl groups can also be derived from inorganic bases (e.g., sodium, potassium, ammonium, calcium, or ferric hydroxide) or from organic bases (e.g., isopropylamine, trimethylamine, histidine, procaine, etc.). Sodium salts of LPA RNAi constructs are particularly useful for therapeutic administration to human subjects. Thus, in certain preferred embodiments, LPA The RNAi construct, particularly olpaciran, is in the form of a sodium salt.In another embodiment, the LPA RNAi construct (e.g., olpaciran) is in the form of a potassium salt.

[0123] For parenteral administration by aqueous solution, for example, the solution is usually suitably buffered and the liquid diluent is first rendered isotonic, for example, with sufficient saline or glucose. Such aqueous solutions can be used, for example, for intravenous, intramuscular, subcutaneous, and intraperitoneal administration. It is preferable to use a sterile aqueous medium as known to those skilled in the art, especially in light of the present disclosure. By way of illustration, a single dose may be dissolved in 1 ml of isotonic NaCl solution and added to 1000 ml of subcutaneous infusion or injected at the proposed site of infusion or injection (see, for example, "Remington's Pharmaceutical Sciences" 15th Edition, pages 1035-1038 and 1570-1580). For human administration, preparations 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 the LPA RNAi construct (e.g., olpaciran) described herein. In other embodiments, the pharmaceutical composition used in the methods of the invention comprises or consists of an LPA RNAi construct described herein (e.g., olpaciran) and sterile water (e.g., water for injection, WFI). In yet other embodiments, the pharmaceutical composition used in the methods of the invention comprises or consists of an LPA RNAi construct described herein (e.g., olpaciran) and phosphate buffered saline (PBS).

[0124] In certain embodiments, pharmaceutical compositions useful for treating, ameliorating, preventing, or reducing the risk of cardiovascular disease according to the methods of the present invention comprise an effective amount of an LPA RNAi construct (e.g., olpaciran), 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., olpaciran), 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., olpaciran), about 15 mM to about 25 mM potassium phosphate, and about 70 mM to about 90 mM sodium chloride. In yet other embodiments, the pharmaceutical composition comprises about 140 mg / ml to about 160 mg / ml of an LPA RNAi construct (e.g., olpaciran), 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 can be within the range of about 6.4 to about 7.2 (e.g., a pH of about 6.4, about 6.6, about 6.8, about 7.0, or about 7.2).

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

[0126] Any of the LPA RNAi constructs described herein can be incorporated into any of the pharmaceutical compositions described above and administered to a patient according to the methods of the present invention. In a specific embodiment, the LPA RNAi construct incorporated into any of the pharmaceutical compositions described above and administered to a patient according to the methods of the present invention is olpaciran.

[0127] In some embodiments, the pharmaceutical composition of the present invention is packaged with or stored in a device for administration, such as any of the above-mentioned injection devices (e.g., pre-filled syringes, auto-injectors, injection pumps, on-body injectors, and injection pens). Devices for aerosolized formulations or powder formulations include, but are not limited to, inhalers, insufflators, inhalers, etc. Thus, the present invention includes an administration device that comprises 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 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. A Phase 1, Randomized, Double-Blind, Placebo-Controlled, Single Ascending Dose Study to Evaluate the Safety, Tolerability, Pharmacokinetics, and Pharmacodynamics of Olpaciran in Subjects With Elevated Plasma Lipoprotein(a) Mendelian and epidemiologic randomized studies have recently established lipoprotein(a) (Lp(a)) as a strong causative risk factor for myocardial infarction and other atherosclerotic complications. There are currently no approved drugs that selectively target Lp(a) and have demonstrated reduction in cardiovascular events. Olpaciran (also known as AMG 890) is an siRNA designed to reduce Lp(a) production by targeting the mRNA transcribed from the LPA gene. The structure of olpaciran is shown in Figure 1.

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

[0131] Eligible subjects were non-fertile women and men, both aged 18-60 years (inclusive) for cohorts 1-5; 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; for cohorts 6-9, at least 6 subjects in each cohort had a stable statin dose for at least 6 weeks at the time of enrollment. Subjects were free of any clinically significant abnormalities in their medical history at the time of randomization.

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

[0133] [Table 1]

[0134] For cohorts 1-5 and cohort 9, the first 2 enrolled subjects in each cohort were randomized to receive olpaciran or placebo in a 1:1 ratio (sentinel pair) administered in a blinded fashion at the same study site on the same day. The remaining cohort subjects were administered the same dose as deemed safe by the investigator and 24 hours or more after the sentinel pair administration. Enrollment into cohorts 1-5 was staggered. After the dosing regimen in the preceding cohort was found to be safe and reasonably well tolerated by the Dose Level Review Team (DLRT), all subjects were dosed into the subsequent cohort based on available safety data through study day 15. Enrollment into cohorts 5-7 began after the dosing regimen in cohort 4 was found to be safe and reasonably well tolerated by the DLRT based on available safety data through study day 15. Subjects returned to the site for post-treatment evaluation on Day 113 for Cohorts 1 and 2, and on Day 225 for Cohorts 3-7. Subjects returned for follow-up (approximately every 2 weeks for Cohorts 1 and 2, and monthly for Cohorts 3-7) until Lp(a) concentrations were at least 80% of baseline. Blood and urine samples were collected throughout the study for evaluation of olpaciran pharmacokinetics (PK) and pharmacodynamics (PD). Safety variables were also assessed periodically.

[0135] The primary endpoints were safety and tolerability as measured by treatment-emergent adverse events (TEAEs), safety laboratory analytes, vital signs, and electrocardiograms (ECGs). Secondary endpoints were maximum observed concentration (C max ), the time point of maximum observed concentration (t maxolpaciran PK parameters, including but not limited to, PK, concentration, and area under the concentration-time curve (AUC); and PD parameters, including change and percentage change in plasma Lp(a) levels at each scheduled visit. The baseline value for Lp(a) was defined as the average of screening and 1 day prior to dosing. If for any reason only one value was available, that value was used as the baseline. Exploratory endpoints included 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 received olpaciran or placebo (Cohorts 1-5: olpaciran, n=30, doses: 3 mg, 9 mg, 30 mg, 75 mg, 225 mg; placebo, n=10; Cohorts 6-7: olpaciran, n=18, doses: 9 mg and 75 mg; placebo, n=6). For subjects who received olpaciran in Cohorts 1-5 (n=30), subjects had a mean (SD) age of 43.9 (13.5) years, 30.0% were female, 63.3% were of Hispanic or Latino ethnicity, 30.0% were black or African American, and 70.0% were white. For subjects receiving placebo in cohorts 1-5 (n=10), subjects had a mean (SD) age of 46.3 (8.5) years, 30.0% were female, 50.0% were of Hispanic or Latino ethnicity, 30.0% were black or African American, and 70.0% were white. For subjects receiving olpaciran in cohorts 6 and 7 (n=18), subjects had a mean (SD) age of 52.7 (9.4) years, 33.3% were female, 27.8% were of Hispanic or Latino ethnicity, and 88.9% were white. For subjects who received placebo in cohorts 6 and 7 (n=6), subjects had a mean (SD) age of 57.8 (5.8) years, 66.7% were female, 33.3% were of Hispanic or Latino ethnicity, 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. There was no use of lipid-modifying medications in cohorts 1-5, but a sizable 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) for subjects who received placebo in cohorts 1-5 and 122 nmol / L (97, 146) for subjects who received olpaciran in cohorts 1-5.The median (Q1, Q3) baseline Lp(a) concentration was 272 nmol / L (233, 307) in subjects receiving placebo in cohorts 6 and 7 and 253 nmol / L (224, 334) for subjects receiving olpaciran in cohorts 6 and 7.

[0137] Olpaciran appeared to be well tolerated. There were no treatment-related serious adverse events. One placebo subject had a serious non-cardiac chest pain adverse event, which was considered unrelated to treatment. In cohorts 1-5, the most common TEAE was upper respiratory tract infection (10% placebo, 13% olpaciran). See Table 2 below. In cohorts 6-7, the most common TEAEs were headache (50% placebo, 28% olpaciran) and upper respiratory tract infection (17% placebo, 17% olpaciran) (Table 2). Only one subject in the study experienced an injection site reaction. There was no apparent dose relationship with the frequency of adverse events. No clinically relevant changes in liver tests, 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 olpaciran effectively lowered mean Lp(a) levels from baseline by 71-96% (based on dose) at day 43 and by 80-94% at day 113 (cohorts 2-5). In cohorts 6 and 7, a single dose of olpaciran effectively lowered mean Lp(a) levels from baseline by 75% and 89%, respectively, at day 43 and by 61% and 80%, respectively, at day 113 (Figure 2). A steep decline in Lp(a) was observed beginning on day 15, with maximum Lp(a) suppression observed from days 43-71. Lp(a) concentrations gradually recovered but remained well below placebo levels at day 225. A single dose of 9 mg or greater caused a reduction in Lp(a) that persisted for 3-6 months.

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

[0141] [Table 3]

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

[0143] The depth and duration of suppression of Lp(a) levels observed with this low single dose of olpaciran was significantly better than expected from the proposed human dose based on the study of olpaciran in cynomolgus monkeys. Based on the efficacy data for olpaciran in cynomolgus monkeys (see, for example, Example 18 in WO 2017 / 059223), the proposed human dose of 75 mg was predicted to reduce Lp(a) levels by about 80% for at least one month. Of note, as mentioned above, a single dose of olpaciran as low as 9 mg reduced Lp(a) levels in human subjects by more than 80% for more than three months. Single olpaciran doses of 75 mg and 225 mg suppressed Lp(a) levels by more than 80% for more than six months. Thus, olpaciran can be administered at lower doses and at longer dosing intervals (including up to once every six months) to human patients in need of Lp(a) reduction. Such a dosing regimen has several different benefits, including increased patient loyalty, reduced drug costs, and reduced injection volume and frequency.

[0144] Example 2. Olpaciran PK / PD model for design of dosing regimen for optimal Lp(a) reduction Based on the Phase 1 data described in Example 1, a mathematical model was developed to characterize olpaciran pharmacokinetics and Lp(a) suppression in healthy volunteers with elevated plasma Lp(a) (>70 nmol / L to <200 nmol / L for cohorts 1-5, >200 nmol / L for cohorts 6-7). The pharmacokinetics (PK) of olpaciran was described using a PK model with first order absorption from the subcutaneous administration site into the circulation, distribution to the liver via asialoglycoprotein receptor (ASGPR) absorption, recycling of olpaciran from the liver back to the circulation, and elimination via degradation from the systemic circulation and liver. Olpaciran serum exposure was found to correlate with baseline Lp(a). Thus, the ability to modulate olpaciran bioavailability by baseline Lp(a) was also included in the model. By describing the suppression of Lp(a) from baseline using the PK / PD model, the model-predicted olpaciran liver concentrations promoted the degradation of LPA mRNA, leading to a decrease in Lp(a) production and suppression during periods of sufficient olpaciran concentrations in the liver. The relationship between olpaciran liver concentrations and LPA mRNA degradation was evaluated using E max The model was used to model changes in LPA mRNA concentrations. Changes in LPA mRNA concentrations were predicted based on the degree of Lp(a) suppression. Baseline Lp(a) levels informed the rates of Lp(a) synthesis and degradation. Higher baseline values ​​were associated with greater rates of production.

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

[0146] The model was able to adequately predict the significant observed reduction in olpaciran exposure in subjects with higher baseline Lp(a) (≧150 nmol / L), suggesting that higher doses of olpaciran may be necessary to achieve target Lp(a) suppression in this patient population. Simulations of the model were performed to explore Q3M and Q6M dosing regimens and to extrapolate predicted Lp(a) suppression after multiple doses of olpaciran. For each candidate dosing regimen, the proportion of subjects achieving target Lp(a) suppression (≧80% reduction from baseline) and the proportion of subjects achieving absolute Lp(a) values ​​≦50 nmol / L at the end of the dosing interval were calculated.

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

[0148] [Table 4]

[0149] [Table 5]

[0150] Model simulations based on Phase 1 data predict that a dose of 10 mg olpaciran administered quarterly (Q3M) will result in approximately 42% of subjects with baseline Lp(a) levels of ≥ 150 nmol / L achieving at least an 80% reduction in Lp(a) from baseline by Month 12. Doses of ≥ 75 mg olpaciran administered quarterly are predicted to achieve ≥ 80% Lp(a) suppression in at least 90% of subjects with baseline Lp(a) levels of ≥ 150 nmol / L as early as 3 months after receiving a single dose of olpaciran. A similar percentage of subjects are predicted to achieve absolute Lp(a) levels of ≤ 50 nmol / L with these same dosing regimens.

[0151] Simulations were also performed on a model for a twice-yearly (Q6M) dosing schedule of olpaciran. Figures 4A-4F show predicted Lp(a) levels as a percentage of baseline for Q6M dosing of olpaciran at doses of 10 mg, 75 mg, 150 mg, 225 mg, 450 mg, and 675 mg for subjects with baseline Lp(a) levels of ≥ 150 nmol / L. The model predicts that doses of at least 75 mg will suppress Lp(a) levels by ≥ 80% throughout the entire six-month dosing interval. Table 6 below shows the predicted proportion of subjects achieving at least an 80% reduction in Lp(a) levels from baseline with different doses of olpaciran administered once every six months (Q6M dosing) at each dosing interval, while Table 7 shows the predicted proportion of subjects achieving absolute Lp(a) levels of ≤ 50 nmol / L with the same olpaciran dosing schedule.

[0152] [Table 6]

[0153] [Table 7]

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

[0155] Based on recent Mendelian randomization studies, a reduction in Lp(a) levels of 80% or more from baseline is expected to provide 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). Thus, olpaciran PK / PD modeling was focused on identifying a dosing regimen of olpaciran that could reduce Lp(a) levels greater than this threshold. The results of olpaciran PK / PD modeling and the simulations described in this example show the following: The 10 mg dose administered once every 3 months or once every 12 weeks achieved an Lp(a) reduction of ≥ 80% in approximately half (42%) of subjects with baseline Lp(a) levels of ≥ 150 nmol / L by month 12, and a median Lp(a) reduction from baseline of approximately 77% at months 6 and 12; · A dose of 75 mg administered once every 3 months or once every 12 weeks is expected to achieve an Lp(a) reduction of ≥ 80% from baseline within 2-3 doses in the majority of subjects (94%), and approximately 90% of subjects are expected to achieve absolute Lp(a) concentrations of ≤ 50 nmol / L with this dosing regimen; A dose of 225 mg administered once every 3 months or once every 12 weeks is expected to achieve an Lp(a) reduction of ≥ 80% in 98% of subjects and to reduce Lp(a) levels to absolute concentrations below 50 nmol / L in 96% of subjects; For doses of 10 mg or greater, a dosing frequency of once every 3 months or once every 12 weeks results in suppression of Lp(a) levels to less than 20% of baseline throughout the 3-month dosing interval in the majority (≥ 90%) of subjects with baseline Lp(a) levels of ≥ 150 nmol / L; and A 225 mg dose administered once every 6 months or once every 24 weeks resulted in a median Lp(a) reduction from baseline of 88%, with approximately 74% of subjects achieving absolute Lp(a) concentrations below 50 nmol / L.

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

[0157] Approximately 240 subjects will be randomized in a 1:1:1:1:1 ratio. Four arms will be treated with olpaciran 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 dosing at days 1, 12, 24, and 36. After week 48, safety follow-up will continue for ≧40 weeks without further dosing with olpaciran or placebo until Lp(a) returns to 80% of baseline (whichever occurs later). Subjects will continue standard of care (including stable lipid altering therapy) according to local guidelines during the treatment period and long-term safety follow-up period.

[0158] After signing informed consent, subjects enter a screening phase (up to 4 weeks) during which subject eligibility criteria are assessed. Eligible subjects include adults aged 18-80 years with atherosclerotic cardiovascular disease who have an Lp(a) >150 nmol / L during screening. Specifically, subjects are enrolled in the study if they meet all of the following key inclusion criteria: Age: 18-80 years During central laboratory screening, Lp(a) >150nmol / L Atherosclerotic cardiovascular disease based on one of the following: History of coronary revascularization by percutaneous coronary intervention (PCI) or coronary artery bypass graft (CABG); · Diagnosis of coronary artery disease with or without previous myocardial infarction; A diagnosis of atherosclerotic cerebrovascular disease; or Diagnosis of peripheral arterial disease For subjects not on lipid-altering therapy (not required for eligibility criteria), lipid-altering therapy, including statin dose, must have remained stable according to local guidelines for ≥4 weeks prior to screening and during screening Subjects will be excluded from the study if they meet any of the following important exclusion criteria: Estimated glomerular filtration rate (eGFR) < 30mL / min / 1.73m during screening 2 Severe renal insufficiency defined as History or clinical evidence of liver 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 Day 1 (excluding non-melanoma skin cancer, cervical intraepithelial neoplasia, ductal intraepithelial neoplasia, or stage 1 prostate cancer) Uncontrolled hypertension on Day 1 (defined as mean systolic blood pressure ≥ 160mmHg or mean diastolic blood pressure ≥ 100mmHg at rest) Fasting triglycerides of ≥ 400 mg / dL (4.5 mmol / L) during screening Type 1 diabetes or uncontrolled type 2 diabetes as determined by glycated hemoglobin (HbA1c) of ≥ 8.5% as determined by a core laboratory at screening

[0159] Eligible subjects for the study had a baseline Lp(a) of >150 nmol / L. This threshold was 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, based on the degree of absolute Lp(a) reduction required to demonstrate a corresponding effect on cardiovascular events, the enrolled population must have a high baseline Lp(a). Thus, the entry criterion of Lp(a)>150 nmol / L yields a study population with a median Lp(a) of approximately 200 nmol / L and allows for evaluation of the efficacy and safety of olpaciran in subjects with very high Lp(a).

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

[0161] Screening for Lp(a) is performed in a central laboratory using approved or investigational turbidimetric immunoassays that are standardized to detect and quantify Lp(a) particles independent of apo(a) isoform size, such as the Tina-quant® Lipoprotein(a) Gen.2 assay available from Roche Diagnostics. The assay is validated for the measurement of Lp(a) in nmol / L in serum samples with a detection limit of 7 nmol / L and is standardized in 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 by the central laboratory using standard methods.

[0162] The primary analysis will be done when all randomized subjects have had the opportunity to complete the Week 36 assessment or have terminated early. The treatment-duration analysis will be done when all subjects have had the opportunity to complete the Week 48 assessment or have terminated early. The final analysis will be done after the last subject has completed the long-term safety follow-up and has either terminated or terminated early from the study. The primary endpoint (percent change from baseline in Lp(a) at Week 36) will be compared between groups using a repeated measures linear effects model that includes treatment group duration, stratification factors, scheduled visits, and interactions of treatment with scheduled visits. The Hochberg procedure will be used to control for type I error for multiple comparisons between the active and placebo arms. Secondary endpoints percent change from baseline in Lp(a) at Week 48, ApoB and LDL-C at Weeks 36 and 48 will be analyzed similarly to the primary endpoint. Safety endpoints (e.g., treatment-emergent adverse events) will be summarized narratively. Baseline Lp(a) is defined as the average of the two most recent nonmissing Lp(a) values ​​measured by a central laboratory prior to or on Study Day 1. If for any reason only one value is available, that value will be used as the baseline.

[0163] A reduction in Lp(a) of 80% or more from baseline was observed with a single dose of olpaciran lasting for more than 3 months (see Example 1). It is expected that this sustained reduction in Lp(a) may provide clinically meaningful cardiovascular benefit by reducing the risk of cardiovascular events in patients with atherosclerotic cardiovascular disease. Recent Mendelian randomization studies suggest that in individuals with very high baseline Lp(a) concentrations, a reduction in Lp(a) of 80% to 90% would be 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., JAMA Cardiol., Vol. 3: 619-627, 2018). al., JAMA Cardiol., Vol. 4:575-579, 2019; and Madsen et al., Arterioscler. Thromb. Vasc. Biol., 40:255-266, 2020). Thus, the results of this study are expected to show that olpaciran will provide a significant percent reduction in Lp(a) from baseline in subjects with elevated Lp(a) and atherosclerotic cardiovascular disease at all doses tested compared to placebo. In particular, it is expected that olpaciran will effectively reduce Lp(a) levels to less than 50 nmol / L in the majority of subjects at doses as low as 10 mg administered once every 12 weeks. This is expected to result in a reduction in the risk of cardiovascular events in such subjects. Olpaciran 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.It is understood that the disclosed invention is not limited to the specific methodology, protocols, and materials described, which may vary.It is also understood that the terminology used herein is merely for the purpose of describing specific embodiments, and is not intended to limit the scope of the appended claims.

[0165] Those skilled in the art will recognize, or be able to ascertain using no more than routine experimentation, many equivalents to the specific embodiments of the invention described herein which equivalents are intended to be encompassed by the following claims. The present invention provides, for example, the following items. (Item 1) 1. A method of treating, attenuating, or preventing atherosclerosis in a patient in need thereof, comprising administering to said patient an LPA RNAi construct at a dose of about 9 mg to about 675 mg with an administration interval of at least 8 weeks, said LPA RNAi construct comprising 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, said targeting moiety being covalently attached to the 5' end of the sense strand. (Item 2) 1. A method of reducing serum or plasma Lp(a) levels in a patient in need thereof, comprising administering to said patient an LPA RNAi construct at a dose of about 9 mg to about 675 mg with an administration interval of at least 8 weeks, said LPA RNAi construct comprising 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, said targeting moiety being covalently attached to the 5' end of the sense strand. (Item 3) 3. The method of claim 2, wherein the patient is diagnosed with or at risk for cardiovascular disease. (Item 4) 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) 3. The method of claim 2, wherein the patient is diagnosed with coronary artery disease. (Item 6) 3. The method of claim 2, wherein the patient has a history of myocardial infarction. (Item 7) 3. The method of claim 2, wherein the patient is diagnosed with acute coronary syndrome. (Item 8) 1. A method of treating, attenuating, or preventing cardiovascular disease in a patient in need thereof, comprising administering to said patient an LPA RNAi construct at a dose of about 9 mg to about 675 mg with an administration interval of at least 8 weeks, said LPA RNAi construct comprising 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, said targeting moiety being covalently attached to the 5' end of the sense strand. (Item 9) 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) 1. A method for reducing the risk of cardiovascular events in a patient with atherosclerotic cardiovascular disease, comprising administering to said patient an LPA RNAi construct at a dose of about 9 mg to about 675 mg with an administration interval of at least 8 weeks, said LPA RNAi construct comprising 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, said targeting moiety being covalently attached to the 5' end of the sense strand. (Item 11) 11. The method of item 10, wherein the cardiovascular event is cardiovascular death, myocardial infarction, stroke, and / or coronary revascularization. (Item 12) 12. The method of claim 10 or 11, wherein the patient has a history of coronary 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 13) 13. The method of any one of items 10 to 12, wherein the patient has experienced a myocardial infarction within one year prior to the first administration of the LPA RNAi construct. (Item 14) 13. The method according to any one of items 10 to 12, wherein the patient is hospitalized due to acute coronary syndrome or unstable angina. (Item 15) 15. The method of any one of items 1 to 14, wherein the patient has a serum or plasma Lp(a) level of about 70 nmol / L or greater prior to the first administration of the LPA RNAi construct. (Item 16) 15. The method of any one of items 1 to 14, wherein the patient has a serum or plasma Lp(a) level of about 100 nmol / L or greater prior to the first administration of the LPA RNAi construct. (Item 17) 15. The method of any one of items 1 to 14, wherein the patient has a serum or plasma Lp(a) level of about 125 nmol / L or greater prior to the first administration of the LPA RNAi construct. (Item 18) 15. The method of any one of items 1 to 14, wherein the patient has a serum or plasma Lp(a) level of about 150 nmol / L or greater prior to the first administration of the LPA RNAi construct. (Item 19) 15. The method of any one of items 1 to 14, wherein the patient has a serum or plasma Lp(a) level of about 175 nmol / L or greater prior to the first administration of the LPA RNAi construct. (Item 20) 15. The method of any one of items 1 to 14, wherein the patient has a serum or plasma Lp(a) level of about 200 nmol / L or greater prior to the first administration of the LPA RNAi construct. (Item 21) 15. The method of any one of items 1 to 14, wherein the patient has a serum or plasma Lp(a) level of about 225 nmol / L or greater prior to the first administration of the LPA RNAi construct. (Item 22) 22. The method according to any one of items 1 to 21, wherein the administration interval is about 12 weeks. (Item 23) 22. The method according to any one of items 1 to 21, wherein the administration interval is about 24 weeks. (Item 24) 22. The method according to any one of items 1 to 21, wherein the LPA RNAi construct is administered to the patient at a dose of about 10 mg to about 225 mg once every 12 weeks. (Item 25) 25. The method according to item 24, wherein the LPA RNAi construct is administered to the patient at a dose of about 50 mg to about 100 mg once every 12 weeks. (Item 26) 25. The method of claim 24, wherein the LPA RNAi construct is administered to the patient at a dose of about 150 mg to about 225 mg once every 12 weeks. (Item 27) 25. The method of claim 24, wherein the LPA RNAi construct is administered to the patient at a dose of about 75 mg once every 12 weeks. (Item 28) 25. The method of claim 24, wherein the LPA RNAi construct is administered to the patient at a dose of about 150 mg once every 12 weeks. (Item 29) 25. The method of claim 24, wherein the LPA RNAi construct is administered to the patient at a dose of about 225 mg once every 12 weeks. (Item 30) 22. The method according to any one of items 1 to 21, wherein the LPA RNAi construct is administered to the patient at a dose of about 225 mg to about 675 mg once every 24 weeks. (Item 31) 31. The method of claim 30, wherein the LPA RNAi construct is administered to the patient at a dose of about 225 mg once every 24 weeks. (Item 32) 32. The method according to any one of items 1 to 31, wherein the patient is undergoing lipid-lowering therapy. (Item 33) 33. The method of item 32, wherein the lipid-lowering therapy is a statin, ezetimibe, a PCSK9 inhibitor, bempedoic acid, or a combination thereof. (Item 34) 34. The method of any one of items 1 to 33, wherein the patient has a serum low-density lipoprotein cholesterol (LDL-C) level of about 100 mg / dL or less prior to the first administration of the LPA RNAi construct. (Item 35) 34. The method of any one of items 1 to 33, wherein the patient has a serum low-density lipoprotein cholesterol (LDL-C) level of about 70 mg / dL or less prior to the first administration of the LPA RNAi construct. (Item 36) The patient has an estimated glomerular filtration rate of about 30 mL / min / 1.73 m before the first administration of the LPA RNAi construct. 2 The method according to any one of Items 1 to 35, wherein (Item 37) 37. The method of any one of items 1 to 36, wherein the patient has a mean systolic blood pressure of less than about 160 mmHg and a mean diastolic blood pressure of less than about 100 mmHg at rest prior to the first administration of the LPA RNAi construct. (Item 38) 38. The method of any one of items 1 to 37, wherein the patient has a glycated hemoglobin A1c level of less than about 8.5% prior to the first administration of the LPA RNAi construct. (Item 39) 39. The method of any one of items 1 to 38, wherein the patient has a serum triglyceride level of less than about 400 mg / dL prior to the first administration of the LPA RNAi construct. (Item 40) 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 a sequence of SEQ ID NO: 3, and the antisense strand of the LPA RNAi construct comprises or consists of a sequence of SEQ ID NO: 4. (Item 41) 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 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. (Item 42) The targeting portion of the LPA RNAi construct: [ka] 42. The method according to any one of items 1 to 41, having the structure: (Item 43) 43. The method of any one of items 1 to 42, wherein the LPA RNAi construct is olpaciran. (Item 44) 44. The method of any one of items 1 to 43, wherein administration of the LPA RNAi construct reduces serum or plasma Lp(a) levels in the patient by more than 50% for at least 12 weeks, compared to the patient's baseline serum or plasma Lp(a) levels. (Item 45) 44. The method of any one of items 1 to 43, wherein administration of the LPA RNAi construct reduces serum or plasma Lp(a) levels in the patient by more than 80% compared to the patient's baseline serum or plasma Lp(a) levels for at least 12 weeks. (Item 46) 44. The method of any one of items 1 to 43, wherein administration of the LPA RNAi construct reduces serum or plasma Lp(a) levels in the patient by more than 90% compared to the patient's baseline serum or plasma Lp(a) levels for at least 12 weeks. (Item 47) 44. The method of any one of items 1 to 43, wherein administration of the LPA RNAi construct reduces serum or plasma Lp(a) levels in the patient to about 100 nmol / L or less. (Item 48) 44. The method of any one of items 1 to 43, wherein administration of the LPA RNAi construct reduces serum or plasma Lp(a) levels in the patient to about 75 nmol / L or less. (Item 49) 44. The method of any one of items 1 to 43, wherein administration of the LPA RNAi construct reduces serum or plasma Lp(a) levels in the patient to about 50 nmol / L or less. (Item 50) 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 comprising potassium phosphate and sodium chloride. (Item 51) 51. The method of any one of items 1 to 50, wherein the LPA RNAi construct is administered to the patient by subcutaneous injection. (Item 52) 52. The method of claim 51, wherein the volume of the injection is about 1 mL or less. (Item 53) 1. An LPA RNAi construct for use in a method of treating, attenuating, or preventing atherosclerosis in a patient in need thereof, the method comprising administering to the patient the LPA RNAi construct at a dose of about 9 mg to about 675 mg with an administration interval of at least 8 weeks, the LPA RNAi construct comprising 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, the targeting moiety being covalently attached to the 5' end of the sense strand. RNAi constructs. (Item 54) 1. An LPA RNAi construct for use in a method of reducing serum or plasma Lp(a) levels in a patient in need thereof, the method comprising administering to the patient the LPA RNAi construct at a dose of from about 9 mg to about 675 mg with an administration interval of at least 8 weeks, the LPA RNAi construct comprising 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, the targeting moiety being covalently linked to the 5' end of the sense strand. (Item 55) 55. The LPA RNAi construct for use according to item 54, wherein the patient is diagnosed with or at risk of cardiovascular disease. (Item 56) 56. The LPA RNAi construct for use according to item 55, 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 57) 55. The LPA RNAi construct for use according to item 54, wherein the patient is diagnosed with chronic kidney disease. (Item 58) 55. The LPA RNAi construct for use according to item 54, wherein the patient has a history of myocardial infarction. (Item 59) The LPA for use according to item 54, wherein the patient is diagnosed with acute coronary syndrome. RNAi constructs. (Item 60) 1. An LPA RNAi construct for use in a method of treating, attenuating, or preventing cardiovascular disease in a patient in need thereof, the method comprising administering to the patient the LPA RNAi construct at a dose of about 9 mg to about 675 mg with an administration interval of at least 8 weeks, the LPA RNAi construct comprising 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, the targeting moiety being covalently linked to the 5' end of the sense strand. (Item 61) 61. The LPA RNAi construct for use according to item 60, 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 62) 1. An LPA RNAi construct for use in a method for reducing the risk of a cardiovascular event in a patient with atherosclerotic cardiovascular disease, the method comprising administering to the patient an LPA RNAi construct. 13. The method of claim 12, further comprising administering an LPA RNAi construct at a dose of about 9 mg to about 675 mg with an administration interval of at least 8 weeks, wherein 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 linked to the 5' end of the sense strand. (Item 63) 63. The LPA RNAi construct for use according to item 62, wherein the cardiovascular event is cardiovascular death, myocardial infarction, stroke, and / or coronary revascularization. (Item 64) 64. The LPA RNAi construct for use according to item 62 or 63, wherein the patient has a history of coronary 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 65) 65. The LPA RNAi construct for use according to any one of items 62 to 64, wherein the patient has experienced a myocardial infarction within one year prior to the first administration of the LPA RNAi construct. (Item 66) 65. The LPA RNAi construct for use according to any one of items 62 to 64, wherein the patient is hospitalized due to acute coronary syndrome or unstable angina. (Item 67) 67. The LPA RNAi construct for use according to any one of items 53 to 66, wherein the patient has a serum or plasma Lp(a) level of about 70 nmol / L or greater before the first administration of the LPA RNAi construct. (Item 68) 67. The LPA RNAi construct for use according to any one of items 53 to 66, wherein the patient has a serum or plasma Lp(a) level of about 100 nmol / L or more before the first administration of the LPA RNAi construct. (Item 69) 67. The LPA RNAi construct for use according to any one of items 53 to 66, wherein the patient has a serum or plasma Lp(a) level of about 125 nmol / L or greater before the first administration of the LPA RNAi construct. (Item 70) 67. The LPA RNAi construct for use according to any one of items 53 to 66, wherein the patient has a serum or plasma Lp(a) level of about 150 nmol / L or greater before the first administration of the LPA RNAi construct. (Item 71) 67. The LPA RNAi construct for use according to any one of items 53 to 66, wherein the patient has a serum or plasma Lp(a) level of about 175 nmol / L or greater before the first administration of the LPA RNAi construct. (Item 72) 67. The LPA RNAi construct for use according to any one of items 53 to 66, wherein the patient has a serum or plasma Lp(a) level of about 200 nmol / L or more before the first administration of the LPA RNAi construct. (Item 73) 67. The LPA RNAi construct for use according to any one of items 53 to 66, wherein the patient has a serum or plasma Lp(a) level of about 225 nmol / L or greater before the first administration of the LPA RNAi construct. (Item 74) 74. The LPA RNAi construct for use according to any one of items 53 to 73, wherein the administration interval is about 12 weeks. (Item 75) 74. The LPA RNAi construct for use according to any one of items 53 to 73, wherein the administration interval is about 24 weeks. (Item 76) 74. The LPA RNAi construct for use according to any one of items 53 to 73, wherein the LPA RNAi construct is administered to the patient at a dose of about 10 mg to about 225 mg once every 12 weeks. (Item 77) 77. The LPA RNAi construct for use according to item 76, wherein the LPA RNAi construct is administered to the patient at a dose of about 50 mg to about 100 mg once every 12 weeks. (Item 78) 77. The LPA RNAi construct for use according to item 76, wherein the LPA RNAi construct is administered to the patient at a dose of about 150 mg to about 225 mg once every 12 weeks. (Item 79) 77. The LPA RNAi construct for use according to item 76, wherein the LPA RNAi construct is administered to the patient at a dose of about 75 mg once every 12 weeks. (Item 80) 77. The LPA RNAi construct for use according to item 76, wherein the LPA RNAi construct is administered to the patient at a dose of about 150 mg once every 12 weeks. (Item 81) 77. The LPA RNAi construct for use according to item 76, wherein the LPA RNAi construct is administered to the patient at a dose of about 225 mg once every 12 weeks. (Item 82) 74. The LPA RNAi construct for use according to any one of items 53 to 73, wherein the LPA RNAi construct is administered to the patient at a dose of about 225 mg to about 675 mg once every 24 weeks. (Item 83) 83. The LPA RNAi construct for use according to item 82, wherein the LPA RNAi construct is administered to the patient at a dose of about 225 mg once every 24 weeks. (Item 84) 84. The LPA RNAi construct for use according to any one of items 53 to 83, wherein the patient is undergoing lipid-lowering therapy. (Item 85) 85. The LPA RNAi construct for use according to item 84, wherein the lipid-lowering therapy is a statin, ezetimibe, a PCSK9 inhibitor, bempedoic acid, or a combination thereof. (Item 86) 86. The LPA RNAi construct for use according to any one of items 53 to 85, wherein the patient has a serum low-density lipoprotein cholesterol (LDL-C) level of about 100 mg / dL or less prior to the first administration of the LPA RNAi construct. (Item 87) 86. The LPA RNAi construct for use according to any one of items 53 to 85, wherein the patient has a serum low-density lipoprotein cholesterol (LDL-C) level of about 70 mg / dL or less prior to the first administration of the LPA RNAi construct. (Item 88) The patient has an estimated glomerular filtration rate of about 30 mL / min / 1.73 m before the first administration of the LPA RNAi construct. 2 The LPA RNAi construct for use according to any one of Items 53 to 87. (Item 89) 89. The LPA RNAi construct for use according to any one of items 53 to 88, wherein the patient has a mean systolic blood pressure at rest of less than about 160 mmHg and a mean diastolic blood pressure of less than about 100 mmHg before the first administration of the LPA RNAi construct. (Item 90) 90. The LPA RNAi construct for use according to any one of items 53 to 89, wherein the patient has a glycated hemoglobin A1c level of less than about 8.5% prior to the first administration of the LPA RNAi construct. (Item 91) The LPA RNAi construct for use according to any one of items 53 to 90, wherein the patient has a serum triglyceride level of less than about 400 mg / dL prior to the first administration of the LPA RNAi construct. (Item 92) 92. The LPA RNAi construct for use according to 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) 93. The LPA RNAi construct for use according to 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 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. (Item 94) The targeting portion of the LPA RNAi construct: [ka] 94. The LPA RNAi construct for use according to any one of items 53 to 93, having the structure: (Item 95) 95. The LPA RNAi construct for use according to any one of items 53 to 94, wherein the LPA RNAi construct is olpaciran. (Item 96) 96. The LPA RNAi construct for use according to any one of items 53 to 95, wherein administration of the LPA RNAi construct reduces serum or plasma Lp(a) levels in the patient by more than 50% for at least 12 weeks, compared to the patient's baseline serum or plasma Lp(a) levels. (Item 97) 96. The LPA RNAi construct for use according to any one of items 53 to 95, wherein administration of the LPA RNAi construct reduces serum or plasma Lp(a) levels in the patient by more than 80% for at least 12 weeks, compared to the patient's baseline serum or plasma Lp(a) levels. (Item 98) 96. The LPA RNAi construct for use according to any one of items 53 to 95, wherein administration of the LPA RNAi construct reduces serum or plasma Lp(a) levels in the patient by more than 90% for at least 12 weeks, compared to the patient's baseline serum or plasma Lp(a) levels. (Item 99) 96. The LPA RNAi construct for use according to any one of items 53 to 95, wherein administration of the LPA RNAi construct reduces serum or plasma Lp(a) levels in the patient to about 100 nmol / L or less. (Item 100) 96. The LPA RNAi construct for use according to any one of items 53 to 95, wherein administration of the LPA RNAi construct reduces serum or plasma Lp(a) levels in the patient to about 75 nmol / L or less. (Item 101) 96. The LPA RNAi construct for use according to any one of items 53 to 95, wherein administration of the LPA RNAi construct reduces serum or plasma Lp(a) levels in the patient to about 50 nmol / L or less. (Item 102) 102. The LPA RNAi construct for use according to any one of items 53 to 101, wherein the LPA RNAi construct is administered to the patient in a pharmaceutical composition comprising potassium phosphate and sodium chloride. (Item 103) 103. The LPA RNAi construct for use according to any one of items 53 to 102, wherein the LPA RNAi construct is administered to the patient by subcutaneous injection. (Item 104) 104. The LPA RNAi construct for use according to item 103, wherein the volume of the injection is about 1 mL or less. (Item 105) 13. Use of an LPA RNAi construct for the preparation of a medicament for treating, attenuating or preventing atherosclerosis in a patient in need thereof, wherein the medicament is administered or formulated for administration at a dose of about 9 mg to about 675 mg with an administration interval of at least 8 weeks, the LPA RNAi construct comprising 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, the targeting moiety being covalently attached to the 5' end of the sense strand. (Item 106) 1. Use of an LPA RNAi construct for the preparation of a medicament for reducing serum or plasma Lp(a) levels in a patient in need thereof, wherein the medicament is administered or formulated for administration at a dose of from about 9 mg to about 675 mg with a dosing interval of at least 8 weeks, the LPA RNAi construct comprising 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, the targeting moiety being covalently attached to the 5' end of the sense strand. (Item 107) 107. The use according to item 106, wherein the patient is diagnosed with or at risk for cardiovascular disease. (Item 108) 108. The use according to item 107, 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 109) The use of item 106, wherein the patient is diagnosed with chronic kidney disease. (Item 110) The use according to item 106, wherein the patient has a history of myocardial infarction. (Item 111) Item 107. The use according to item 106, wherein the patient is diagnosed with acute coronary syndrome. (Item 112) 13. Use of an LPA RNAi construct for the preparation of a medicament for treating, attenuating or preventing cardiovascular disease in a patient in need thereof, wherein the medicament is administered or formulated for administration at a dose of about 9 mg to about 675 mg with an administration interval of at least 8 weeks, the LPA RNAi construct comprising 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, the targeting moiety being covalently attached to the 5' end of the sense strand. (Item 113) 13. 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 stenosis, stable or unstable angina, atrial fibrillation, heart failure, hyperlipidemia, heterozygous familial hypercholesterolemia, or homozygous familial hypercholesterolemia. (Item 114) 1. Use of an LPA RNAi construct for the preparation of a medicament for reducing the risk of cardiovascular events in a patient with atherosclerotic cardiovascular disease, wherein the medicament is administered or formulated for administration at a dose of about 9 mg to about 675 mg with an administration interval of at least 8 weeks, the LPA RNAi construct comprising 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, the targeting moiety being covalently attached to the 5' end of the sense strand. (Item 115) 15. The use according to item 114, wherein the cardiovascular event is cardiovascular death, myocardial infarction, stroke, and / or coronary revascularization. (Item 116) 116. The use according to item 114 or 115, wherein the patient has a history of coronary 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 a myocardial infarction within one year prior to the first administration of the agent. (Item 118) 117. 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) 119. The use according to any one of items 105 to 118, wherein the patient has a serum or plasma Lp(a) level of about 70 nmol / L or greater before the first administration of the agent. (Item 120) 119. The use according to any one of items 105 to 118, wherein the patient has a serum or plasma Lp(a) level of about 100 nmol / L or more before the first administration of the agent. (Item 121) 119. The use according to any one of items 105 to 118, wherein the patient has a serum or plasma Lp(a) level of about 125 nmol / L or greater before the first administration of the agent. (Item 122) 119. The use according to any one of items 105 to 118, wherein the patient has a serum or plasma Lp(a) level of about 150 nmol / L or greater before the first administration of the agent. (Item 123) 119. The use according to any one of items 105 to 118, wherein the patient has a serum or plasma Lp(a) level of about 175 nmol / L or greater before the first administration of the agent. (Item 124) 119. The use according to any one of items 105 to 118, wherein the patient has a serum or plasma Lp(a) level of about 200 nmol / L or greater before the first administration of the agent. (Item 125) 119. The use according to any one of items 105 to 118, wherein the patient has a serum or plasma Lp(a) level of about 225 nmol / L or greater before the first administration of the agent. (Item 126) 126. The use according to any one of items 105 to 125, wherein the administration interval is about 12 weeks. (Item 127) 126. The use according to any one of items 105 to 125, wherein the administration interval is about 24 weeks. (Item 128) 126. The use according to any one of items 105 to 125, wherein the medicament is administered to the patient at a dose of about 10 mg to about 225 mg once every 12 weeks, or is formulated for administration at that dose. (Item 129) 129. The use of item 128, wherein the medicament is administered to the patient at a dose of about 50 mg to about 100 mg once every 12 weeks, or is formulated for administration at that dose. (Item 130) 129. The use of item 128, wherein the medicament is administered to the patient at a dose of about 150 mg to about 225 mg once every 12 weeks, or is formulated for administration at that dose. (Item 131) 129. The use of item 128, wherein the medicament is administered to the patient in a dose of about 75 mg once every 12 weeks or is formulated for administration in that dose. (Item 132) 129. The use of item 128, wherein the medicament is administered to the patient in a dose of about 150 mg once every 12 weeks or is formulated for administration in that dose. (Item 133) 129. The use of item 128, wherein the medicament is administered to the patient in a dose of about 225 mg once every 12 weeks or is formulated for administration in that dose. (Item 134) 126. The use according to any one of items 105 to 125, wherein the medicament is administered to the patient at a dose of about 225 mg to about 675 mg once every 24 weeks, or is formulated for administration at that dose. (Item 135) 135. The use of item 134, wherein the medicament is administered to the patient in a dose of about 225 mg once every 24 weeks or is formulated for administration in that dose. (Item 136) 136. The use according to any one of items 105 to 135, wherein the patient is undergoing lipid-lowering therapy. (Item 137) 137. The use of item 136, wherein the lipid-lowering therapy is a statin, ezetimibe, a PCSK9 inhibitor, bempedoic acid, or a combination thereof. (Item 138) 138. The use according to any one of items 105 to 137, wherein the patient has a serum low-density lipoprotein cholesterol (LDL-C) level of about 100 mg / dL or less before the first administration of the agent. (Item 139) The use according to any one of items 105 to 137, wherein the patient has a serum low-density lipoprotein cholesterol (LDL-C) level of about 70 mg / dL or less before the first administration of the agent. (Item 140) The patient has an estimated glomerular filtration rate of about 30 mL / min / 1.73 m before the first dose of the drug. 2 The use according to any one of Items 105 to 139, wherein (Item 141) 141. The use according to any one of items 105 to 140, wherein the patient has a mean systolic blood pressure of less than about 160 mmHg and a mean diastolic blood pressure of less than about 100 mmHg at rest before the first administration of the agent. (Item 142) 142. The use according to any one of items 105 to 141, wherein the patient has a glycated hemoglobin A1c level of less than about 8.5% prior to the first administration of the agent. (Item 143) 143. The use according to any one of items 105 to 142, wherein the patient has a serum triglyceride level of less than about 400 mg / dL before the first administration of the agent. (Item 144) 144. The use according to any one of items 105 to 143, 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 145) 145. The use according to any one of items 105 to 144, wherein 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. (Item 146) The targeting portion of the LPA RNAi construct: [ka] 146. The use according to any one of items 105 to 145, having the structure: (Item 147) 147. The use according to any one of items 105 to 146, wherein the LPA RNAi construct is olpaciran. (Item 148) 148. The use of any one of items 105 to 147, wherein administration of the agent reduces serum or plasma Lp(a) levels in the patient by more than 50% compared to the patient's baseline serum or plasma Lp(a) levels for at least 12 weeks. (Item 149) 148. The use of any one of items 105 to 147, wherein administration of the agent reduces serum or plasma Lp(a) levels in the patient by more than 80% compared to the patient's baseline serum or plasma Lp(a) levels for at least 12 weeks. (Item 150) 148. The use according to any one of items 105 to 147, wherein administration of the agent reduces serum or plasma Lp(a) levels in the patient by more than 90% compared to the patient's baseline serum or plasma Lp(a) levels for at least 12 weeks. (Item 151) 148. The use according to any one of items 105 to 147, wherein administration of the agent reduces serum or plasma Lp(a) levels in the patient to below about 100 nmol / L. (Item 152) 148. The use according to any one of items 105 to 147, wherein administration of the agent reduces serum or plasma Lp(a) levels in the patient to below about 75 nmol / L. (Item 153) 148. The use according to any one of items 105 to 147, wherein administration of the agent reduces serum or plasma Lp(a) levels in the patient to below about 50 nmol / L. (Item 154) 154. The use according to any one of items 105 to 153, wherein the medicament is administered to the patient in a pharmaceutical composition comprising potassium phosphate and sodium chloride or is formulated for administration to the patient in such a pharmaceutical composition. (Item 155) 155. The use according to any one of items 105 to 154, wherein the medicament is administered to the patient by subcutaneous injection or is formulated for administration to the patient by subcutaneous injection. (Item 156) Item 156. The use according to item 155, wherein the volume of the injection is about 1 mL or less.

Claims

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

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