Treatment of a non-alcoholic fatty liver disease

Abstract

Described are methods of treatment of a non-alcoholic fatty liver disease via a reduction in HSD17B13 expression in humans in need of treatment.

Description

TREATMENT OF A NON-ALCOHOLIC FATTY LIVER DISEASECROSS-REFERENCE

[0001] This application claims the benefit of U.S. provisional application number 63 / 588,365 filed on October 6, 2023. which is hereby incorporated by reference in its entirety.FIELD OF THE INVENTION

[0002] Disclosed herein are methods of treatment of a non-alcoholic fatty liver disease (NAFLD) such as non-alcoholic steatohepatitis (NASH), which can reduce (e.g., inhibit or silence) hepatic lipid droplet protein 17P-hydroxysteroid dehydrogenase type 13 (also referred to as HSD17013, HSD17B13, 170-HSD13, 17beta-HSD13, 17beta-HSD type 13, or 17B-HSD13) gene expression and its protein expression.BRIEF SUMMARY

[0003] In some aspects, the present disclosure provides a method for treating a nonalcoholic fatty liver disease in a human subject, comprising administering to the human subject a compound or a pharmaceutically acceptable salt thereof, at a dose of about 300 to about 800 mg, calculated based on a free acid form of the compound, wherein the compound comprises a double-stranded oligonucleotide linked to a ligand, wherein the double-stranded oligonucleotide reduces expression of HSD17P13 in the human subject, and wherein the ligand comprises a chemical structure of:stereoisomer thereof, or a pharmaceutically acceptable salt thereof.BRIEF DESCRIPTION OF THE DRAWINGS

[0004] FIG. 1 shows predicted change from baseline in hepatic HSD 17(313 protein for 1000 subjects to biopsy on Day 30, 60, 90, 120 and 150 for a single subcutaneous dose of 100, 200, 300, 400, 600, or 800 mg of a test compound.

[0005] FIG. 2 shows an overlay of predicted mean changes from baseline in hepatic HSD17013 protein for 1000 subjects to biopsy sampling over a year from a single dose of 100, 300, 400, 600, or 800 mg of a test compound.

[0006] FIG. 3A shows a target engagement model predicting the percentage of subj ects for example at over 85% hepatic HSD 17( 13 protein knockdown for 1000 subjects on Day 60. This compares the doses of 100mgQ4W (dose on Day 1, 28. and 56) and 200mgQ12W (dose on Day 1) with the doses of 400mgQ12W (dose on Day 1) and 800mgQ12W (dose on Day 1). On Day 60 there are approximately 71% and 75% more subjects with >85% knock down with 400 mg and 800 mg compared to 200 mg dose, respectively. FIG. 3B shows a target engagement model predicting the percentage of subjects for example at over 85% hepatic HSD17J313 protein knockdown for 1000 subjects on Day 84. This compares the 100 mg Q4W (dose on Day 1 , 28, and 56) and 200 mg QI 2W (dose on Day 1) with the doses of 400mg Q12W (dose on Day 1) and 800mg Q12W (dose on Day 1). On Day 84 there are approximately 67% and 87% more subjects with >85% HSD17(313 knock down with 400 mg and 800 mg compared to 200 mg. respectively.

[0007] FIG. 4 shows the details of the population pharmacokinetic (PK) target engagement model. PK and Keo: individual estimates fixed from a population PK- pharmacodynamic model of PK and ALT; Kout: fixed based on estimate of fractional synthesis rate from patients with fibrosis; baseline (BSL): semi-quantitative western blot measurements; Kin: estimated as BSL x Kout; ImaX: fixed to 0.99; ICso: model estimated to percent change in western blot measurements.

[0008] FIG. 5A to 5D shows a chemical structure of the test compound described in Example 2, in a free acid form. The circled numerical numbers indicate the connecting points of parts of the structure split into four pages due to length.

[0009] FIG. 6A to 6D shows a chemical structure of the test compound described in Example 2, in a sodium salt form. The circled numerical numbers indicate the connecting points of parts of the structure split into four pages due to length.

[0010] FIG. 7 shows study enrollment schema and associated data reviews.

[0011] FIG. 8 shows the clinically observed data compared with the accurately predicted data from the population PK target engagement model (FIG. 4) in the plasma concentration of a test compound after a single subcutaneous dose of 200 and 400 mg. Black circles represent individual plasma concentrations of the test compound in the Phase 2a clinical study. The black solid lines and black dotted lines are the median and 95% quantile of the model predictions.

[0012] FIG. 9 shows the strong correlation between the systemic exposure [area under the concentration-time curve (AUCO-24)] of the test compound after a single subcutaneous dose of 200 and 400 mg and the body weight of the Phase 2a clinical study participants.

[0013] FIG. 10 shows the clinically observed data compared with the accurately predicted data from the population PK target engagement model (FIG. 4) in the mean change from baseline in hepatic HSD17|313 protein after a single subcutaneous dose of 200 and 400 mg of a test compound. Black circles represent individual change from baseline in hepatic HSD17[313 protein at approximately weeks 8 and 12 in the Phase 2a clinical study after a single subcutaneous dose of 200 and 400 mg. The thick black solid lines and shaded region are the median and 95% quantile of the model predictions. The clinically observed changes in hepatic HSD17P13 protein are adequately captured within the model predictions confirming the accuracy over time and across different doses.

[0014] FIG. 11A and FIG. 11B show the clinical efficacy of the test compound on the mean changes from baseline of liver enzymes alanine aminotransferase (ALT) and aspartate aminotransferase (AST) after a single subcutaneous dose of 400 mg of a test compound in patients with NASH or NAFLD.DETAILED DESCRIPTION

[0015] In some cases, treatment disclosed herein can reduce hepatocyte production of the HSD17P13 and provide a therapeutic and / or protective effect in human subjects having or suspected of having NAFLD and potentially other chronic liver diseases. In some instances of human patients with NASH or suspected NASH (e g., NAFLD with high ALT), the treatment can lead to significant reduction in HSD17P13 mRNA or protein expression at a dose of more than 200 mg, for example from about 300 mg to about 800 mg. In some instances, the treatment inhibits HSD17P13 in subjects carrying the PNPLA3 rs738409 CG or GG genotypes that are associated with increased risk of NASH cirrhosis, and such inhibition effect can be substantially the same compared to subjects lacking the PNPLA3mutation. In some instances, reduced HSD17[313 expression can lead to or be accompanied by reductions (e.g., at least 10%) in ALT and AST, which are surrogate markers of hepatic inflammation in NASH. In some instances, the treatment disclosed herein results in an effective and / or sustained reduction in HSD17P13 mRNA or protein expression over the course of 150 days or more in human subjects having or suspected of having NAFLD. for example a decrease of about 90% to about 99% on Day 60, or about 60% to about 90% on Day 84, 90, or 120, or about 40% to about 60% on Day 150, in the protein expression level after the initial administration of a compound described herein. In some instances, the human subjects for the treatments described herein may have a body weight within a range of 50 to 150 kg, e.g., 50 to 100 kg, 60 to 120 kg, 80 to 120 kg, or 80 to 140 kg.

[0016] Unless indicated otherwise, the term “about” used herein means plus or minus 10% of a referred number.

[0017] Unless indicated otherwise, the term “a”, “an”, and “the” includes singular and plural references.Compounds for Use

[0018] Compounds used in treating human subjects disclosed herein can comprise, for example an RNA interference (RNAi) agent that reduces expression of HSD17B13, referred herein as “HSD17B13 RNAi agent.” In some instances, the compound comprises HSD17B13-targeting double-stranded oligonucleotide linked to galactose-containing ligands that can be preferentially taken up by hepatocytes to promote liver specificity, following which the compound will engage the cell’s RNAi machinery to target HSD17B13 mRNA for degradation, thereby decreasing the amount of free HSD17B13 mRNA for translation. These factors may minimize off-target effects or extrahepatic inhibition of the compound and contribute to the compound’s acceptable safety profile. In some instances, the compound herein targets engagement with a knockdown of HSD17B13, mimicking carriers of the enzy matic LOF (loss of function) mutation in HSD17B13 and confer the protective benefits against NASH and potentially other chronic liver diseases.

[0019] As used herein, the terms “reduce.” “silence,” “inhibit.” “down-regulate,” or “knockdown,” when referring to expression of HSD17B13, mean that the expression of the gene, as measured by the level of mRNA (messenger RNA) transcribed from the gene, or the level of protein (or protein subunit) translated from the mRNA, in a cell, group of cells, tissue, organ, or subject in which the gene is transcribed, is reduced when the cell, group of cells, tissue, organ, or subject is treated with the RNAi agent as compared to a second cell.group of cells, tissue, organ, or subject that has not been so treated. HSD17B13 reduction may be measured by any appropriate assay or method known in the art. The non-limiting exemplary7assays include microarray, Affymetrix gene chips, and qRT-PCR (quantitative Reverse Transcriptase Polymerase Chain Reaction) for HSD17B13 gene quantitation, and gel electrophoresis such as Western Blot and mass spectrometry7for HSD17B13 protein measurements, as well as those assays set forth in International Patent Application Publication No. WO 2020 / 061177 (Patent Application No. PCT / US2019 / 051707), which is incorporated by reference herein in its entirety. A reference HSD17B13 mRNA gene transcript for normal humans can be transcript variant A, GenBank NM_178135.4. In some instances, the reduction in gene expression is measured by comparing the baseline levels of HSD17B13 mRNA or HSD17B13 protein in a human subject prior to a treatment disclosed herein, with the HSD17B13 mRNA or HSD17B13 protein levels after administration of the compound.

[0020] Compounds disclosed herein may contain one or more asymmetric centers (also referred to as a chiral centers) and may. therefore, exist as individual enantiomers, diastereoisomers, or other forms of stereoisomers, or as mixtures thereof. Chiral centers, such as chiral carbon atoms, may also be present in a substituent such as an alkyl group. Where the stereochemistry of a chiral center in any chemical structure illustrated herein, is not specified, the structure is intended to encompass any stereoisomer and all mixtures thereof. In some instances, compounds disclosed herein containing one or more chiral centers may be used as racemic modifications including racemic mixtures and racemates, enantiomerically-enriched mixtures, or as enantiomerically-pure individual stereoisomers. In some instances, a compound herein is provided as a salt, mixed salt, or free acid.

[0021] Individual stereoisomers which contain one or more asymmetric centers may be resolved by methods known to those skilled in the art. For example, such resolution may be carried out (1) by formation of diastereoisomeric salts, complexes or other derivatives; (2) by selective reaction with a stereoisomer-specific reagent, for example by enzymatic oxidation or reduction; or (3) by gas-liquid or liquid chromatography in a chiral environment, for example, on a chiral support such as silica with a bound chiral ligand or in the presence of a chiral solvent. It will be appreciated that where the desired stereoisomer is converted into another chemical entity by one of the separation procedures described above, a further step is required to liberate the desired form. Alternatively, specific stereoisomers may be synthesized by asymmetric synthesis using optically active reagents, substrates, catalysts orsolvents, or by converting one enantiomer to the other by asymmetric transformation. In some cases, a compound for use herein comprises a sense strand (also referred to as a passenger strand) and an antisense strand (also referred to as a guide strand) that are annealed to form a duplex.

[0022] In some instances, the compound comprises a double-stranded oligonucleotide, for example comprising an RNA, chemically modified RNA, DNA. or chemically modified DNA oligonucleotide molecule, any of which can have two strands of complementary sequence. In some instances, the double-stranded oligonucleotide is capable of degrading or inhibiting translation of messenger RNA (mRNA) transcripts of HSD17B13 mRNA in a sequence specific manner. In some instances, the compound disclosed herein may operate through the RNA interference mechanism (i.e., inducing RNA interference through interaction with the RNA interference pathway machinery (RNA-induced silencing complex or RISC) of mammalian cells), or by any alternative mechanism(s) or pathway(s). While it is believed that the HSD17B13 RNAi agents, as that term is used herein, operate primarily through the RNA interference mechanism, the disclosed RNAi agents are not bound by or limited to any particular pathway or mechanism of action. In some instances, the compound comprises a short or small interfering RNA (siRNA), double-strand RNA (dsRNA), micro RNA (miRNA), short hairpin RNA (shRNA), or dicer substrate. In some instances, the compounds for use herein include an antisense strand that has a region of complementarity to at least a portion of an HSD17B13 mRNA.

[0023] In some instances, a compound for use herein has a sense strand of 15 to 49 nucleotides in length, and an antisense strand of 18 to 49 nucleotides in length. In some instances, the sense and antisense strands are independently 18 to 26 nucleotides in length. In some instances, the sense and antisense strands are independently 21 to 26 nucleotides in length. In some instances, the sense and antisense strands are independently 21 to 24 nucleotides in length. In some instances, the sense strands are 15, 16, 17, 18, 19, 20, 21, 22, 23, 24, 25, 26, 27, 28, 29, or 30 nucleotides in length. In some instances, the antisense strands are 18, 19, 20, 21, 22, 23. 24. 25, 26, 27, 28, 29, or 30 nucleotides in length. In some instances, the sense strand and the antisense strand are both 21 nucleotides in length. The sense and antisense strands can be either the same length or different lengths. The sense and antisense strands can also form overhanging nucleotides on one or both ends of the nucleotide chain.

[0024] Compounds suitable for use herein can be covalently linked to a ligand that includesone or more N-acetyl-galactosamine moieties. The N-acetyl-galactosamine moieties can facilitate the targeting of the HSD17B13 RNAi agent to the asialoglycoprotein receptors (ASGPR) readily present on the surface of hepatocytes, which leads to internalization of the HSD17B13 RNAi agent by endocytosis or other means.Modified Nucleotides and Modified Internucleoside Linkages

[0025] Compounds for example HSD17B13 RNAi agents disclosed herein can comprise modified nucleotides, which can preserve activity of the RNAi agent while at the same time increasing the serum stability, as well as minimize the possibility of activating interferon activity in humans. As used herein, a '‘modified nucleotide” is a nucleotide other than a ribonucleotide (2'-hydroxyl nucleotide). In some instances, at least 50% (e.g., at least 60%, at least 70%, at least 80%, at least 90%, at least 95%, at least 97%, at least 98%, at least 99%, or 100%) of the nucleotides are modified nucleotides. As used herein, modified nucleotides include but are not limited to, deoxyribonucleotides, nucleotide mimics, 2'- modified nucleotides, inverted nucleotides, modified nucleobase-comprising nucleotides, bridged nucleotides, peptide nucleic acids (PNAs), 2',3'-seco nucleotide mimics (unlocked nucleobase analogues), locked nucleotides, 3’-O-methoxy (2' internucleoside linked) nucleotides. 2'-F-arabino nucleotides. 5'-Me, 2'-fluoro nucleotides, morpholino nucleotides, vinyl phosphonate-containing nucleotides, and cyclopropyl phosphonate-containing nucleotides. In some instances, the modified nucleotides of an HSD17B13 RNAi agent are 2'-modified nucleotides (i.e., a nucleotide with a group other than a hydroxyl group at the 2' position of the five-membered sugar ring). 2’-modified nucleotides include, but are not limited to, 2'Omethyl nucleotides, 2'-deoxy-2'-fluoro nucleotides (commonly referred to simply as 2’ -Fluoro nucleotides), 2'-deoxy nucleotides, 2'-methoxy ethyl (2'-O-2- methoxyethyl) nucleotides, 2'-amino nucleotides, and 2'-alkyl nucleotides. It may not be necessary for all nucleotides in a given RNAi agent to be uniformly modified. In some instances, more than one modification can be incorporated in a single HSD17B13 RNAi agent or even in a single nucleotide thereof. The HSD17B13 RNAi agent sense strands and antisense strands can be synthesized and / or modified by methods known in the art. Modification at one nucleotide can be independent of modification at another nucleotide.

[0026] In some instances, a nucleobase (often referred to as simply the “base”) disclosed herein may be modified. Natural nucleobases can include the primary purine bases adenine and guanine, and the primary pyrimidine bases cytosine, thymine, and uracil. A nucleobase may be modified to include, without limitation, universal bases, hydrophobic bases,promiscuous bases, size-expanded bases, and fluorinated bases. The synthesis of such modified nucleobases (including phosphoramidite compounds that include modified nucleobases) is known in the art.

[0027] Modified nucleobases include, for example, 5-substituted pyrimidines, 6- azapyrimidines and N-2, N-6 and 0-6 substituted purines, (e.g.. 2-aminopropyladenine, 5- propynyluracil, or 5-propynylcytosine), 5-methylcytosine (5-me-C), Shydroxymethyl cytosine, inosine, xanthine, hypoxanthine, 2-aminoadenine, 6-alkyl (e.g., 6-methyl, 6-ethyl, 6-isopropyl, or 6-n-butyl) derivatives of adenine and guanine, 2-alkyl (e.g., 2-methyl, 2- ethyl, 2-isopropyl, or 2-n-butyl) and other alkyd derivatives of adenine and guanine, 2- thiouracil, 2-thiothymine, 2-thiocytosine, 5-halouracil, cytosine, 5-propynyl uracil. 5- propynyl cytosine, 6-azo uracil, 6-azo cytosine, 6-azo thymine, 5-uracil (pseudouracil), 4- thiouracil, 8-halo, 8-amino, 8-sulfhydryl, 8-thioalkyl, 8-hydroxyl and other 8-substituted adenines and guanines, 5-halo (e.g., 5-bromo), 5-trifluoromethyl, and other 5-substituted uracils and cytosines, 7-methylguanine and 7-methyladenine, 8-azaguanine and 8- azaadenine, 7-deazaguanine, 7-deazaadenine. 3-deazaguanine. and 3-deazaadenine.

[0028] In some instances, all or substantially all the nucleotides of a compound disclosed herein for example an HSD17B13 RNAi agent are modified nucleotides. As used herein, an RNAi agent wherein substantially all the nucleotides are modified nucleotides is an RNAi agent having four or fewer (i.e.. 0, 1, 2. 3, or 4) nucleotides in both the sense strand and the antisense strand being ribonucleotides (i.e., unmodified). As used herein, a sense strand wherein substantially all the nucleotides are modified nucleotides is a sense strand having two or fewer (i.e., 0, 1, or 2) nucleotides in the sense strand being ribonucleotides. As used herein, an antisense sense strand wherein substantially all the nucleotides are modified nucleotides is an antisense strand having two or fewer (i.e., 0, 1. or 2) nucleotides in the sense strand being ribonucleotides.

[0029] In some instances, one or more nucleotides of a compound disclosed herein for example an HSD17B13 RNAi agent are linked by non-standard linkages or backbones (i.e., modified intemucleoside linkages or modified backbones). Modified intemucleoside linkages or backbones include, but are not limited to, phosphorothioate groups, chiral phosphorothioates, thiophosphates, phosphorodithioates, phosphotriesters, aminoalkylphosphotriesters, alkyl phosphonates (e.g., methyl phosphonates or 3'-alkylene phosphonates), chiral phosphonates, phosphinates, phosphoramidates (e.g., 3'-amino phosphoramidate, aminoalkylphosphoramidates, or thionophosphoramidates), thionoalkyl-phosphonates, thionoalkylphosphotriesters. morpholino linkages, boranophosphates having normal 3 '-5 ' linkages. 2'-5' linked analogs of boranophosphates, or boranophosphates having inverted polarity wherein the adjacent pairs of nucleoside units are linked 3'-5' to 5'-3' or 2'- 5' to 5 '-2'. In some instances, a modified intemucleoside linkage or backbone lacks a phosphorus atom. Modified internucleoside linkages lacking a phosphorus atom include, but are not limited to, short chain alkyl or cycloalkyl inter-sugar linkages, mixed heteroatom and alkyl or cycloalkyl inter-sugar linkages, or one or more short chain heteroatomic or heterocyclic inter-sugar linkages. In some instances, modified intemucleoside backbones include, but are not limited to. siloxane backbones, sulfide backbones, sulfoxide backbones, sulfone backbones, formacetyl and thioformacetyl backbones, methylene formacetyl and thioformacetyl backbones, alkene-containing backbones, sulfamate backbones, methyleneimino and methyl enehydrazino backbones, sulfonate and sulfonamide backbones, amide backbones, and other backbones having mixed N, O, S, and CH2 components.

[0030] In some instances, a sense strand of a compound disclosed herein for example an HSD17B13 RNAi agent can contain 1, 2, 3, 4. 5, or 6 phosphorothioate linkages, an antisense strand of an HSD17B13 RNAi agent can contain 1 , 2, 3, 4, 5, or 6 phosphorothioate linkages, or both the sense strand and the antisense strand independently can contain 1, 2, 3, 4, 5, or 6 phosphorothioate linkages. In some instances, a sense strand of an HSD17B13 RNAi agent can contain 1. 2, 3, or 4 phosphorothioate linkages, an antisense strand of an HSD17B13 RNAi agent can contain 1, 2, 3, or 4 phosphorothioate linkages, or both the sense strand and the antisense strand independently can contain 1, 2, 3, or 4 phosphorothioate linkages.

[0031] In some instances, a sense strand in a compound disclosed herein for example an HSD17B13 RNAi agent contains at least two phosphorothioate intemucleoside linkages. In some instances, the at least two phosphorothioate intemucleoside linkages are between the nucleotides at positions 1-3 from the 3' end of the sense strand. In some instances, the at least two phosphorothioate intemucleoside linkages are between the nucleotides at positions 1-3, 2-4, 3-5, 4-6, 4-5, or 6-8 from the 5' end of the sense strand. In some instances, phosphorothioate intemucleoside linkages are used to link the terminal nucleotides in the sense strand to capping residues present at the 5’-end, the 3’-end, or both the 5’- and 3’-ends of the nucleotide sequence. In some instances, phosphorothioate intemucleoside linkages are used to link a ligand to the sense strand.

[0032] In some instances, an antisense strand of a compound disclosed herein for example an HSD17B13 RNAi agent contains three or four phosphorothioate intemucleoside linkages.In some instances, the antisense strand contains three phosphorothioate intemucleoside linkages. In some instances, the three phosphorothioate intemucleoside linkages are between the nucleotides at positions 1-3 from the 5' end of the antsense strand and between the nucleotides at positions 19-21, 20-22, 21-23, 22-24, 23-25, or 24-26 from the 5' end. In some instances, the compound contains at least two phosphorothioate intemucleoside linkages in the sense strand and three or four phosphorothioate intemucleoside linkages in the antisense strand.

[0033] In some instances, a compound disclosed herein for example an HSD17B13 RNAi agent contains one or more modified nucleotides and one or more modified intemucleoside linkages. In some instances, a 2'modified nucleoside is combined with modified intemucleoside linkage.Table 1A. Exemplary Antisense Strand SequenceTable IB. Exemplary Sense Strand Nucleotide Sequence (shown as modified version without inverted abasic residues or NAG ligand)

[0034] As used in Tables 1 A, IB, and 2 herein, the following notations are used to indicate modified nucleotides, ligand groups, and linking groups: A, C, G, and U represent adenosine, cytidine, guanosine, and uridine, respectively; a, c, g, and u represent 2'-O-methyl adenosine, cytidine, guanosine, and uridine, respectively; Af, Cf, Gf, and Uf represent 2'- fluoro adenosine, cytidine, guanosine, and uridine, respectively; s represents a phosphorothioate linkage; (invAb) represents an inverted abasic deoxyribose residue (see Table 2); and (NAG37)s represents the structure shown in Table 3, below.

[0035] Unless otherwise indicated by the sequence (such as, for example, by a phosphorothioate linkageCLs”), when present in a strand, the monomers can be mutually linked by 5 ’-3 ’-phosphodi ester bonds. In some instances, the inclusion of a phosphorothioate linkage as shown in the modified nucleotide sequences disclosed herein replaces the phosphodiester linkage ty pically present in oligonucleotides. In some instances, the terminal nucleotide at the 3’ end of a given oligonucleotide sequence would typicallyhave a hydroxyl (-OH) group at the respective 3‘ position of the given monomer instead of a phosphate moiety ex vivo. In some instances, when viewing the respective strand 5’ -> 3’. the inverted abasic residues are inserted such that the 3’ position of the deoxyribose is linked at the 3 ’ end of the preceding monomer on the respective strand. Moreover, as the person of ordinary skill would readily understand and appreciate, while the phosphorothioate chemical structures depicted herein typically show the anion on the sulfur atom, the present disclosure encompasses phosphorothioate tautomers (e.g., where the sulfur atom has a double-bond and the anion is on an oxygen atom). In some instances, each sense strand and / or antisense strand can have any linking groups disclosed herein, linked to the 5' and / or 3' end of the sequence. Capping Residues

[0036] In some instances, a sense strand disclosed herein may include one or more capping residues, sometimes referred to as a “cap,” a “terminal cap,” or a “capping residue.” As used herein, a “capping residue” is a non-nucleotide compound or other moiety that can be incorporated at one or more termini of a nucleotide sequence of an RNAi agent disclosed herein. A capping residue can provide the RNAi agent with certain beneficial properties, for example, protection against exonuclease degradation. In some instances, inverted abasic residues (invAb) (also referred to as “inverted abasic sites”) are added as capping residues (see Table 2). Capping residues include, for example, inverted abasic residues as well as carbon chains such as a terminal C3H7 (propyl), CeHis (hexyl), or C12H25 (dodecyl) groups. In some instances, a capping residue is present at either the 5' terminal end, the 3' terminal end, or both the 5' and 3' terminal ends of the sense strand. In some instances, the 5’ end and / or the 3' end of the sense strand may include more than one inverted abasic deoxyribose moiety as a capping residue.

[0037] In some instances, one or more inverted abasic residues (invAb) are added to the 3' end of a sense strand. In some instances, one or more inverted abasic residues (invAb) are added to the 5' end of the sense strand. In some instances, one or more inverted abasic residues or inverted abasic sites are inserted between the ligand and the nucleotide sequence of the sense strand of the RNAi agent. In some instances, the inclusion of one or more inverted abasic residues or inverted abasic sites at or near the terminal end or terminal ends of the sense strand of an RNAi agent allows for enhanced activity or other desired properties of an RNAi agent.

[0038] In some instances, one or more inverted abasic residues (invAb) are added to the 5' end of a sense strand. In some instances, one or more inverted abasic residues can be insertedbetween the ligand and the nucleotide sequence of the sense strand of the RNAi agent. The inverted abasic residues may be linked via phosphate, phosphorothioate (e.g., shown herein as (invAb)s)), or other intemucleoside linkages. The chemical structures for inverted abasic deoxyribose residues are show n in Table 2 below, as well as in the chemical structures shown in Figure 5A to 5D and Figure 6A to 6D.

[0039] In some cases, a compound for use herein can include a RNA molecule such as an HSD17B13 RNAi agent, linked to one or more non-nucleotide groups such as a ligand that can enhance targeting or delivery of the RNAi agent. Examples include three N-acetyl-galactosamine moieties, are described in the present disclosure. The ligand can be covalently linked to the 3' and / or 5' end of either the sense strand and / or the antisense strand. In some instances, the compound contains a ligand linked to the 3' and / or 5' end of the sense strand. In some instances, a ligand is linked to the 5' end of the sense strand. In some instances, the ligand comprises, consists essentially of, or consists of the structure (NAG37)s, and is linked to the 5' end of the sense strand. A ligand can be linked directly or indirectly to the RNAi agent via a linker / linking group. In some instances, a ligand is linked to the RNAi agent via a labile, cleavable, or reversible bond or linker. In some instances, a ligand is linked to an inverted abasic residue at the 5’ end of the sense strand.

[0040] Ligands disclosed herein can enhance the pharmacokinetic or biodistribution properties of a RNAi agent to which they are attached to improve cell-specific distribution and cell-specific uptake of the RNAi agent. In some instances, the ligand enhances endocytosis of the RNAi agent. In some instances, the ligand can be monovalent, divalent, trivalent, tetravalent, or have higher valency for the target to which it is directed. Representative ligands include, without limitation, compounds with affinity to cell surface molecules, cell receptor ligands, haptens, antibodies, monoclonal antibodies, antibody fragments, and antibody mimics with affinity' to cell surface molecules.

[0041] The preparation of ligands, such as galactose derivative clusters that include N- acetyl-galactosamine, is described in, for example, International Patent Application Publication No. WO 2018 / 044350 (Patent Application No. PCT / US2017 / 021147) and International Patent Application Publication No. WO 2017 / 156012 (Patent Application No. PCT / US2017 / 021175), both of which are incorporated by reference herein in their entirety'.

[0042] For example, a ligand linked to the HSD17B13 RNAi agent described in Tables 1 A and IB can have the chemical structure of (NAG37)s, as shown in Table 3.Pharmaceutical Compositions

[0043] The compounds for example HSD17B13 RNAi agents suitable for use herein can be formulated as pharmaceutical compositions for administration to human subjects. The pharmaceutical compositions can be used to treat or prevent a subject having a disease or disorder that would benefit from inhibition of expression of HSD17B13 mRNA or reduction in the level of HSD17B13 protein, such as human subjects having a non-alcoholic Patty liverdisease. In some instances, one or more pharmaceutically acceptable excipients (including vehicles, carriers, diluents, and / or delivery polymers) are added to the pharmaceutical compositions that include an HSD17B13 RNAi agent, thereby forming a pharmaceutical formulation suitable for in vivo delivery' to a human subject.

[0044] As used herein, “pharmacologically effective amount,'’ “therapeutically effective amount,” or simply “effective amount” refers to that amount of an active agent to produce a pharmacological, therapeutic or preventive result. In some instances, a therapeutically or prophylactically effective amount of one or more of pharmaceutical compositions is administered to a subject in need thereof, to decrease the number, severity, and / or frequency of symptoms of a disease in the subject.

[0045] As used herein, a pharmaceutical composition or medicament includes a pharmacologically effective amount of at least one compound disclosed herein and one or more pharmaceutically acceptable excipients. Excipients are substances other than the Active Pharmaceutical Ingredient (API, therapeutic product, e g., HSD17B13 RNAi agent) that are intentionally included in the drug delivery system. Excipients do not exert or are not intended to exert a therapeutic effect at the intended dosage. Excipients can act to a) aid in processing of the drug delivery system during manufacture, b) protect, support, or enhance stability, bioavailability or patient acceptability of the API, c) assist in product identification, and / or d) enhance any other attribute of the overall safety, effectiveness, of delivery of the API during storage or use.

[0046] Excipients may include, but are not limited to: absorption enhancers, antiadherents, anti-foaming agents, anti-oxidants, binders, buffering agents, carriers, coating agents, colors, delivery enhancers, delivery polymers, dextran, dextrose, diluents, disintegrants, emulsifiers, extenders, fillers, flavors, glidants. humectants, lubricants, oils, polymers, preservatives, saline, salts, solvents, sugars, suspending agents, sustained release matrices, sweeteners, thickening agents, tonicity agents, vehicles, water-repelling agents, and wetting agents.

[0047] Pharmaceutical compositions suitable for injection include sterile aqueous solutions (where water soluble). For subcutaneous or intravenous or intramuscular administration, suitable carriers may include physiological saline, bacteriostatic water, Cremophor® ELTM (BASF, Parsippany, NJ) or phosphate buffer. It should be stable under the conditions of manufacture and storage and should be preserved against the contaminating action of microorganisms such as bacteria and fungi. The carrier can be a solvent ordispersion medium containing, for example, water, ethanol, polyol (for example, glycerol, propylene glycol, and liquid polyethylene glycol), and suitable mixtures thereof.[004S] Sterile injectable solutions can be prepared by incorporating the active compound in the required amount in an appropriate solvent with one or a combination of ingredients enumerated above, as required, followed by filter sterilization. In some instances, dispersions are prepared by incorporating the active compound into a sterile vehicle, which contains a basic dispersion medium and the required other ingredients from those enumerated above.

[0049] A pharmaceutical composition can contain other additional components commonly found in pharmaceutical compositions. Such additional components be selected from active agents such as anti-pruritics, astringents, local anesthetics, anti-inflammatory agents, or antihistamines.

[0050] The described pharmaceutically acceptable formulations can be packaged into kits, containers, packs, or dispensers. The pharmaceutical compositions described herein can be packaged in pre-filled syringes or vials.Methods of Treatment

[0051] As used herein, the terms “treat,” “treatment,” “treating,” and the like, may include the prevention, management, prophylactic treatment, reduction, and / or inhibition of the number, severity, and / or frequency of a disease and / or one or more symptoms of a disease in a subject. In some cases, the terms “treat,” “treatment,” “treating,” and the like mean management, reduction, and / or inhibition of the number, severity, and / or frequency of a disease and / or one or more symptoms of a disease in a subject. Unless otherw ise expressly- noted, treatment also can include relief from or alleviation of underlying conditions and / or symptoms of the disease. A dose for the treatment herein can be more than 200 mg and up to about 800 mg, for example in a range of: about 300 to about 400 mg, about 300 to about 500 mg, about 300 to about 600 mg, about 300 about 700 mg, about 300 to about 800 mg, about 250 to about 350 mg, about 250 mg to about 500 mg, about 350 to about 400 mg, about 350 to about 450 mg, about 350 to about 500 mg, about 350 to about 600 mg. about 350 to about 700 mg, about 400 to about 500 mg, about 400 to about 600 mg, about 400 to about 700 mg, or about 400 to about 800 mg.

[0052] Embodiment 1 : A method for treating a non-alcoholic fatty liver disease in a human subject in need thereof, comprising administering to the human subject a compound or a pharmaceutically acceptable salt thereof, at a dose more than 200 mg and up to about 800mg, e.g.. in a range of about 300 to about 800 mg, wherein the dose is calculated based on a free acid form of the compound, wherein the compound comprises a double-stranded oligonucleotide linked to a ligand, wherein the double-stranded oligonucleotide reduces expression of HSD17013 in the human subject, and wherein the ligand comprises a chemical structure of:stereoisomer thereof, or a salt (e.g., pharmaceutically acceptable salt) thereof.

[0053] Embodiment 2: The method of Embodiment 1, wherein the administration is a single dose, or repeated about even' 1 , 2, or 3 months or longer, for example every 4, 8, or 12 weeks. In some instances, the administration is repeated every 4 weeks, e.g.. at the dose of about 300 or about 400 mg. In some instances, the administration is repeated every 8 weeks, e.g., at the dose of about 300 or about 400 mg. In some instances, the administration is repeated every 12 weeks, e.g., at the dose of about 300 or about 400 mg.

[0054] Embodiment 3 : The method of Embodiment 1 or 2, wherein the dose is about 600 mg to about 800 mg, for example 800 mg.

[0055] Embodiment 4: The method of Embodiment 1 or 2, wherein the dose is about 300 mg to about 400 mg, for example about 350 mg.

[0056] Embodiment 5: The method of Embodiment 4, wherein the dose is about 300 mg.

[0057] Embodiment 6: The method of Embodiment 4, wherein the dose is about 400 mg.

[0058] Embodiment 7: The method of any one of Embodiments 1-6, wherein the compound is in a sodium salt form.

[0059] Embodiment 8: The method of any one of Embodiments 1-7, wherein the compound is administered by an injection, for example at a concentration of 200 mg / ml.

[0060] Embodiment 9: The method of Embodiment 8, wherein the injection is a subcutaneous injection.

[0061] Embodiment 10: The method of any one of Embodiments 1-9, wherein the compound is present in a unit dosage form, for example including a single (unit) dosage form.

[0062] Embodiment 11: The method of any one of Embodiments 1-10, wherein the nonalcoholic fatty liver disease is non-alcoholic steatohepatitis (NASH), e.g., pre-cirrhotic nonalcoholic steatohepatitis, NASH-F3, NASH-F4, NASH with bridging fibrosis, or biopsy- proven NASH.

[0063] Embodiment 12 : The method of any one of Embodiments 1-11, wherein the human subject has a HSD17[313 rs72613567 mutation, PNPLA3 rs738409 (I148M) mutation, or a combination thereof. In some instances, the human subject has a HSD17|313 rs72613567 mutation. In some instances, the human subject has a PNPLA3 rs738409 (I148M) mutation.

[0064] Embodiment 13: The method of any one of Embodiments 1-12, wherein the double-stranded oligonucleotide comprises an antisense strand that comprises a modified nucleotide sequence that differs by 0 or 1 nucleotides from one of the following sequences (5' 3'): usCfsasUfcUfaucagAfcUfuCfuUfaCfsg (nucleotide sequence SEQ ID NO: 1); usCfsasUfcUfaUfcAfgAfcUfuCfuUfaCfsg (nucleotide sequence SEQ ID NO:5); usGfsasUfcCfaAfaAfaUfgUfcCfuAfgGfsc (nucleotide sequence SEQ IDNO:6); or usGfsasUfcCfaaaaaUfgUfcCfuAfgGfsc (nucleotide sequence SEQ ID NO:7); and wherein a. c, g, and u represent 2'-O-methyl adenosine, 2'-O-methyl cytidine, 2'-O-methyl guanosine, and 2'-O-methyl uridine, respectively; Af, Cf, Gf, and Uf represent 2'-fluoro adenosine, 2'-fluoro cytidine, 2'-fluoro guanosine, and 2'-fluoro uridine, respectively; and s represents a phosphorothioate linkage.

[0065] Embodiment 14: The method of any one of Embodiments 1-13. wherein the double-stranded oligonucleotide comprises a sense strand that comprises a modified nucleotide sequence that differs by 0 or 1 nucleotides from one of the following sequences (5' 3'): cguaagaaGfuCfuGfauagauga (nucleotide sequence SEQ ID NO:3); cguaagaaGfUfCfugauagauga (nucleotide sequence SEQ ID NO: 8);gccuaggaCfAfUfuuuugiauca (nucleotide sequence SEQ ID NO:9); or gccuaggaCfaUfuUfuugiauca (nucleotide sequence SEQ ID NO: 10); and wherein a, c, g, i, and u represent 2'-O-methyl adenosine, 2'-O-methyl cytidine, 2'-O-methyl guanosine, 2'-O-methyl inosine, and 2'-O-methyl uridine, respectively: Af. Cf. Gf. and Uf represent 2’-fluoro adenosine, 2'-fluoro cytidine, 2'- fluoro guanosine, and 2'-fluoro uridine, respectively; and s represents a phosphorothioate linkage.

[0066] Embodiment 15: The method of any one of Embodiments 1-14, wherein the double-stranded oligonucleotide comprises an antisense strand that comprises a sequence: usCfsasUfcUfaucagAfcUfuCfuUfaCfsg (nucleotide sequence SEQ ID NO: 1), and a sense strand that comprises a sequence: (NAG37)s(invAb)scguaagaaGfuCfuGfauagaugas(invAb) (nucleotide sequence SEQ ID NO: 11), and wherein a, c, g, and u represent 2'-O-methyl adenosine, 2'-O-methyl cytidine, 2'-O-methyl guanosine, and 2'-O-methyl uridine, respectively; Af, Cf, Gf, and Uf represent 2'-fluoro adenosine, 2'-fluoro cytidine, 2'-fluoro guanosine, and 2'-fluoro uridine, respectively; s represents a phosphorothioate linkage; and (invAb) represents an inverted abasic deoxyribose residue.

[0067] Embodiment 16: The method of any one of Embodiments 1-15, wherein the method reduces the expression level of HSD17P13 mRNA in the human subject by at least about 50% (e.g., at least about 62%. at least about 79%, at least about 84%, at least about 90%, or at least about 96%) compared with that prior to the administration, as measured by quantitative Reverse Transcriptase Polymerase Chain Reaction (qRT-PCR) with a liver biopsy sample from the human subject, for example about day 71 or 113 from the first administration.

[0068] Embodiment 17 : The method of any one of Embodiments 1-16, wherein the method reduces the expression of HSD17P13 protein in the human subject by at least about 30% (e.g., at least about 40%, at least about 50%, at least about 60%, at least about 83%, at least about 92%, or at least about 97%) compared with that prior to the administration, as measured by Western Blot with a liver biopsy sample from the human subject, for example about day 71 or 113 from the first administration.

[0069] Embodiment 18: The method of any one of Embodiments 1-17, wherein the method reduces ALT (alanine aminotransferase) level in the human subject by at least about 7.7% (e.g., at least about 14%, at least about 26%, at least about 36%, or at least about 39%), or atleast about 40% (e.g., at least about 51%, at least about 53%, or at least about 60%) compared with that prior to the administration, as measured in the human subject's serum, for example about day 71 or 113 from the first administration.

[0070] Embodiment 19: The method of any one of Embodiments 1-18, wherein the method reduces AST (aspartate aminotransferase) level in the human subject by at least about 7.5% (e.g., at least about 15%). or at least about 20% (e.g., at least about 24%, or at least about 28%) compared with that prior to the administration, as measured in the human subject’s serum, for example about day 71 or 113 from the first administration.

[0071] Embodiment 20: The method of any one of Embodiments 1-19, wherein the method reduces or slows an increase of the human subject’s liver fat fraction (e.g., a reduction of at least 4-41% or at least 7-8%) compared with that prior to the administration, as measured by magnetic resonance imaging, for example about day 71 or 113 from the first administration.

[0072] Embodiment 21 : The method of any one of Embodiments 1-20, wherein the method reduces or slows an increase of the human subject’s liver stiffness (e.g., a reduction of at least 4-37%) compared with that prior to the administration, as measured by transient elastography, for example about day 71 or 113 from the first administration.

[0073] Embodiment 22: The method of any one of Embodiments 1-21, wherein the chemical structure of the ligand comprises:

[0074] Embodiment 23: The method of Embodiment 22, wherein the compound is in a free acid form having a chemical structure as illustrated in FIG. 5 A to 5D.

[0075] Embodiment 24: The method of any one of Embodiments 1-21, wherein the chemical structure of the ligand comprises:

[0076] Embodiment 25: The method of Embodiment 24, wherein the compound is in a sodium salt form having a chemical structure as illustrated in FIG. 6A to 6D.

[0077] Embodiment 26: The method of any one of Embodiments 1-25, wherein the method reduces GGT (gamma-glutamyl transferase) level in the human subject by at least about 1.4% or at least 8.3% or at least 9%, compared with that prior to the administration, as measured in the human subject's serum, for example about day 71 or 113 from the first use.

[0078] Embodiment 27: A compound or a pharmaceutically acceptable salt thereof for use in treating non-alcoholic fatty liver disease in a human subject, wherein the use comprises administering to the human subject the compound or pharmaceutically acceptable salt thereof, at a dose more than 200 mg and up to about 800 mg, e.g., in a range of about 300 mg to about 800 mg, wherein the dose is calculated based on a free acid form of the compound, wherein the compound comprises a double-stranded oligonucleotide linked to a ligand, wherein the double-stranded oligonucleotide reduces expression ofHSD17pi3 in the human subject, and wherein the ligand comprises a chemical structure of:stereoisomer thereof, or a salt (e.g., a pharmaceutically acceptable salt) thereof.

[0079] Embodiment 28: The compound or a pharmaceutically acceptable salt thereof for use according to Embodiment 27, wherein the administration is a single dose, or repeated at an interval of about: 1, 2, or 3 months or longer, for example every' 4, 8, or 12 weeks. Embodiment 28a: The compound or a pharmaceutically acceptable salt thereof for use according to Embodiment 27, wherein the administration is about 300 mg every' 4, 8, or 12 weeks. Embodiment 28b: The compound or a pharmaceutically7acceptable salt thereof for use according to Embodiment 27, wherein the administration is about 400 mg every 4, 8, or 12 weeks.

[0080] Embodiment 29: The compound or a pharmaceutically acceptable salt thereof for use according to Embodiment 27 or 28, wherein the dose is about 600 mg to about 800 mg, for example 800 mg.

[0081] Embodiment 30: The compound or a pharmaceutically acceptable salt thereof for use according to Embodiments 27 or 28, wherein the dose is about 300 mg to about 400 mg. for example about 350 mg.

[0082] Embodiment 31 : The compound or a pharmaceutically acceptable salt thereof for use according to Embodiment 30, wherein the dose is about 300 mg.

[0083] Embodiment 32: The compound or a pharmaceutically acceptable salt thereof for use according to Embodiment 30, wherein the dose is about 400 mg.

[0084] Embodiment 33: The compound or a pharmaceutically' acceptable salt thereof for use according to any one of Embodiments 27-32, wherein the compound is in a sodium salt form.

[0085] Embodiment 34: The compound or a pharmaceutically acceptable salt thereof for use according to any one of Embodiments 27-33. wherein the compound is administered by an injection, for example at a concentration of 200 mg / ml.

[0086] Embodiment 35: The compound or a pharmaceutically acceptable salt thereof for use according to Embodiment 34, wherein the injection is subcutaneous.

[0087] Embodiment 36: The compound or a pharmaceutically acceptable salt thereof for use according to any one of Embodiments 27-35, wherein the compound is administered in a unit dosage form, e.g., present in a single unit dosage form.

[0088] Embodiment 37: The compound or a pharmaceutically acceptable salt thereof for use according to any one of Embodiments 27-36, wherein the non-alcoholic fatty liver disease is non-alcoholic steatohepatitis (NASH), e.g., pre-cirrhotic non-alcoholic steatohepatitis, NASH-F3, NASH-F4, NASH with bridging fibrosis, or biopsy-proven NASH.

[0089] Embodiment 38: The compound or a pharmaceutically acceptable salt thereof for use according to any one of Embodiments 27-37, wherein the human subject has a HSD17|313 rs72613567 mutation, PNPLA3 rs738409 (1148M) mutation, or a combination thereof.

[0090] Embodiment 39: The compound or a pharmaceutically acceptable salt thereof for use according to any one of Embodiments 27-38, wherein the double-stranded oligonucleotide comprises an antisense strand that comprises a modified nucleotide sequence that differs by 0 or 1 nucleotides from one of the following sequences (5' 3'): usCfsasUfcUfaucagAfcUfuCfuUfaCfsg (nucleotide sequence SEQ ID NO: 1); usCfsasUfcUfaUfcAfgAfcUfuCfuUfaCfsg (nucleotide sequence SEQ ID NO:5); usGfsasUfcCfaAfaAfaUfgUfcCfuAfgGfsc (nucleotide sequence SEQ IDNO:6); or usGfsasUfcCfaaaaaUfgUfcCfuAfgGfsc (nucleotide sequence SEQ ID NO:7); and wherein a. c, g, and u represent 2'-O-methyl adenosine, 2'-O-methyl cytidine, 2'-O-methyl guanosine, and 2'-O-methyl uridine, respectively; Af, Cf, Gf, and Uf represent 2'-fluoro adenosine, 2'-fluoro cytidine, 2'-fluoro guanosine, and 2'-fluoro uridine, respectively; and s represents a phosphorothioate linkage.

[0091] Embodiment 40: The compound or a pharmaceutically acceptable salt thereof for use according to any one of Embodiments 27-39, wherein the double-strandedoligonucleotide comprises a sense strand that comprises a modified nucleotide sequence that differs by 0 or 1 nucleotides from one of the following sequences (5' - 3'): cguaagaaGfuCfuGfauagauga (nucleotide sequence SEQ ID NO:3); cguaagaaGfUfCfugauagauga (nucleotide sequence SEQ ID NO: 8); gccuaggaCfAfUfuuuugiauca (nucleotide sequence SEQ ID NO:9); or gccuaggaCfaUfuUfuugiauca (nucleotide sequence SEQ ID NO: 10); and wherein a, c, g, i, and u represent 2'-O-methyl adenosine, 2'-O-methyl cytidine, 2'-O-methyl guanosine, 2'-O-methyl inosine, and 2'-O-methyl uridine, respectively; Af, Cf, Gf, and Uf represent 2'-fluoro adenosine, 2'-fluoro cytidine, 2'- fluoro guanosine, and 2'-fluoro uridine, respectively; and s represents a phosphorothioate linkage.

[0092] Embodiment 41 : The compound or a pharmaceutically acceptable salt thereof for use according to any one of Embodiments 27-40, wherein the double-stranded oligonucleotide comprises an antisense strand that comprises a sequence: usCfsasUfcUfaucagAfcUfuCfuUfaCfsg (nucleotide sequence SEQ ID NO: 1), and a sense strand that comprises a sequence: (NAG37)s(invAb)scguaagaaGfuCfuGfauagaugas(invAb) (nucleotide sequence SEQ ID NO: 11), and wherein a, c, g, and u represent 2'-O-methyl adenosine, 2'-O-methyl cytidine. 2'-O-methyl guanosine, and 2'-O-methyl uridine, respectively; Af, Cf, Gf, and Uf represent 2'-fluoro adenosine, 2'-fluoro cytidine, 2'-fluoro guanosine, and 2'-fluoro uridine, respectively; s represents a phosphorothioate linkage; and (invAb) represents an inverted abasic deoxyribose residue.

[0093] Embodiment 42: The compound or a pharmaceutically acceptable salt thereof for use according to any one of Embodiments 27-41, wherein the use reduces the expression of HSD17[313 mRNA in the human subject by at least about 50% (e.g., at least about 62%, at least about 79%, at least about 84%, at least about 90%, or at least about 96%) compared with that prior to the administration, as measured by quantitative Reverse Transcriptase Polymerase Chain Reaction (qRT-PCR) with a liver biopsy sample from the human subject, for example about day 71 or 113 from the first administration.

[0094] Embodiment 43: The compound or a pharmaceutically acceptable salt thereof for use according to any one of Embodiments 27-42, wherein the use reduces the expression of HSD17P13 protein in the human subject by at least about 30% (e.g., at least about 40%, at least about 50%, at least about 60%, at least about 83%, at least about 92%, or at least about97%) compared with that prior to the administration, as measured by Western Blot with a liver biopsy sample from the human subject, for example about day 71 or 113 from the first administration.

[0095] Embodiment 44: The compound or a pharmaceutically acceptable salt thereof for use according to any one of Embodiments 27-43, wherein the use reduces ALT (alanine aminotransferase) level in the human subject by at least about 7.7% (e.g.. at least about 14%. at least about 26% at least about 36%, or at least about 39%), or at least about 40% (e.g., at least about 51%, at least about 53%, or at least about 60%) compared with that prior to the administration, as measured in the human subject’s serum, for example about day 71 or 113 from the first administration.

[0096] Embodiment 45: The compound or a pharmaceutically acceptable salt thereof for use according to any one of Embodiments 27-44, wherein the use reduces AST (aspartate aminotransferase) level in the human subject by at least about 7.5% (e.g., at least about 15%), or at least about 20% (e.g., at least about 24%, or at least about 28%) compared with that prior to the administration, as measured in the human subject’s serum, for example about day 71 or 1 13 from the first administration.

[0097] Embodiment 46: The compound or a pharmaceutically acceptable salt thereof for use according to any one of Embodiments 27-45, wherein the use reduces or slows an increase of the human subject’s liver fat fraction (e.g., a reduction of at least 4-41% or at least about 7-8%) compared with that prior to the administration, as measured by magnetic resonance imaging, for example about day 71 or 113 from the first administration.

[0098] Embodiment 47 : The compound or a pharmaceutically acceptable salt thereof for use according to any one of Embodiments 27-46, wherein the use reduces or slows an increase of the human subject’s liver stiffness (e.g.. a reduction of at least 4-37%) compared with that prior to the administration, as measured by transient elastography, for example about day 71 or 113 from the first administration.

[0099] Embodiment 48: The compound or a pharmaceutically acceptable salt thereof for use according to any one of Embodiments 27-47, wherein the chemical structure of the ligand comprises:

[0100] Embodiment 49: The compound or a pharmaceutically acceptable salt thereof for use according to Embodiment 48, wherein the compound is in a free acid form having a chemical structure as illustrated in FIG. 5A to 5D.

[0101] Embodiment 50: The compound or a pharmaceutically acceptable salt thereof for use according to any one of Embodiments 27-47, wherein the chemical structure of the ligand

[0102] Embodiment 51 : The compound or a pharmaceutically acceptable salt thereof for use according to Embodiment 50, wherein the compound is in a sodium salt form having a chemical structure as illustrated in FIG. 6A to 6D.

[0103] Embodiment 52: The compound or a pharmaceutically acceptable salt thereof for use according to any one of Embodiments 27-51, wherein the use reduces GGT (gammaglutamyl transferase) level in the human subject by at least about 1.4% or at least 8.3% or at least 9%, compared with that prior to the administration, as measured in the human subject’s serum, for example about day 71 or 113 from the first use.

[0104] Embodiment 53: Use of a compound or a pharmaceutically acceptable salt thereof for the manufacture of a medicament for the treatment of non-alcoholic fatty liver disease, wherein the compound or pharmaceutically acceptable salt thereof is used at a dose of more than 200 mg and up to about 800 mg, e g., in a range of about 300 mg to about 800 mg, wherein the dose is calculated based on a free acid form of the compound, wherein the compound comprises a double-stranded oligonucleotide linked to a ligand, wherein the double-stranded oligonucleotide reduces expression of I4SD17P13, and wherein the ligand comprises a chemical structure of:stereoisomer thereof, or a pharmaceutically acceptable salt thereof.

[0105] Embodiment 54: The use of Embodiment 53, wherein the dose is a single dose, or repeated at an interval of about: 1, 2, or 3 months, between doses, for example every 4, 8, or 12 weeks. Embodiment 55a: The use of Embodiment 53, wherein the administration is about 300 mg every 4, 8, or 12 weeks. Embodiment 55b: The use of Embodiment 53, wherein the administration is about 400 mg every 4, 8, or 12 weeks.

[0106] Embodiment 55: The use of Embodiment 53 or 54. wherein the dose is about 600 mg to about 800 mg, for example 800 mg.

[0107] Embodiment 56: The use of Embodiment 53 or 54. wherein the dose is about 300 mg to about 400 mg, for example about 350 mg.

[0108] Embodiment 57: The use of Embodiment 56, wherein the dose is about 300 mg.

[0109] Embodiment 58: The use of Embodiment 56, wherein the dose is about 400 mg.

[0110] Embodiment 59: The use of any one of Embodiments 53-58, wherein the compound is in a sodium salt form.

[0111] Embodiment 60: The use of any one of Embodiments 53-59, wherein the compound is used by an injection, for example at a concentration of 200 mg / ml.

[0112] Embodiment 61 : The use of Embodiment 60, wherein the injection is subcutaneous.

[0113] Embodiment 62: The use of any one of Embodiments 53-61, wherein the compound is in a unit dosage form.

[0114] Embodiment 63: The use of any one of Embodiments 53-62, wherein the nonalcoholic fatty7liver disease is non-alcoholic steatohepatitis (NASH), e.g., pre-cirrhotic nonalcoholic steatohepatitis, NASH-F3, NASH-F4, NASH with bridging fibrosis, or biopsy- proven NASH.

[0115] Embodiment 64: The use of any one of Embodiments 53-63, wherein the treatment is to a subject having a HSD17P13 rs72613567 mutation, PNPLA3 rs738409 (I148M) mutation, or a combination thereof.

[0116] Embodiment 65: The use of any one of Embodiments 53-64, wherein the doublestranded oligonucleotide comprises an antisense strand that comprises a modified nucleotide sequence that differs by70 or 1 nucleotides from one of the following sequences (5' 3'): usCfsasUfcUfaucagAfcUfuCfuUfaCfsg (nucleotide sequence SEQ ID NO: 1); usCfsasUfcUfaUfcAfgAfcUfuCfuUfaCfsg (nucleotide sequence SEQ ID NO:5); usGfsasUfcCfaAfaAfaUfgUfcCfuAfgGfsc (nucleotide sequence SEQ IDNO:6); or usGfsasUfcCfaaaaaUfgUfcCfuAfgGfsc (nucleotide sequence SEQ ID NO:7); and wherein a. c, g, and u represent 2'-O-methyl adenosine, 2'-O-methyl cytidine, 2'-O-methyl guanosine, and 2'-O-methyl uridine, respectively; Af, Cf, Gf, and Uf represent 2'-fluoro adenosine, 2'-fluoro cytidine, 2'-fluoro guanosine, and 2'-fluoro uridine, respectively; and s represents a phosphorothioate linkage.

[0117] Embodiment 66: The use of any one of Embodiments 53-65, wherein the doublestranded oligonucleotide comprises a sense strand that comprises a modified nucleotide sequence that differs by 0 or 1 nucleotides from one of the following sequences (5' -> 3'): cguaagaaGfuCfuGfauagauga (nucleotide sequence SEQ ID NO:3); cguaagaaGfUfCfugauagauga (nucleotide sequence SEQ ID NO: 8); gccuaggaCfAfUfuuuugiauca (nucleotide sequence SEQ ID NO:9); or gccuaggaCfaUfuUfuugiauca (nucleotide sequence SEQ ID NOTO); and wherein a, c, g, i, and u represent 2'-O-methyl adenosine, 2'-O-methyl cytidine, 2'-O-methyl guanosine, 2'-O-methyl inosine, and 2'-O-methyl uridine, respectively; Af. Cf. Gf. and Uf represent 2'-fluoro adenosine, 2'-fluoro cytidine, 2'- fluoro guanosine, and 2'-fluoro uridine, respectively; and s represents a phosphorothioate linkage.

[0118] Embodiment 67: The use of any one of Embodiments 53-66, wherein the doublestranded oligonucleotide comprises an antisense strand that comprises a sequence: usCfsasUfcUfaucagAfcUfuCfuUfaCfsg (nucleotide sequence SEQ ID NOT), and a sense strand that comprises a sequence: (NAG37)s(invAb)scguaagaaGfuCfuGfauagaugas(invAb) (nucleotide sequence SEQ ID NO: 11), and wherein a, c, g, and u represent 2'-O-methyl adenosine, 2'-O-methyl cytidine. 2'-O-methyl guanosine, and 2'-O-methyl uridine, respectively; Af, Cf, Gf, and Uf represent 2'-fluoro adenosine, 2'-fluoro cytidine, 2'-fluoro guanosine, and 2'-fluoro uridine, respectively; s represents a phosphorothioate linkage; and (invAb) represents an inverted abasic deoxyribose residue.

[0119] Embodiment 68: The use of any one of Embodiments 53-67, wherein the use reduces the expression of HSD17013 mRNA by at least about 50% (e.g., at least about 62%, at least about 79%, at least about 84%, at least about 90%, or at least about 96%) compared with that prior to the use, as measured by quantitative Reverse Transcriptase Polymerase Chain Reaction (qRT-PCR) with a liver biopsy sample, for example about day 71 or 113 from the first use.

[0120] Embodiment 69: The use of any one of Embodiments 53-68, wherein the use reduces the expression of HSD17 13 protein by at least about 30% (e.g.. at least about 40%. at least about 50%, at least about 60%, at least about 83%, at least about 92%, or at least about 97%) compared with that prior to the use, as measured by Western Blot with a liver biopsy sample, for example about day 71 or 113 from the first use.

[0121] Embodiment 70: The use of any one of Embodiments 53-69, wherein the use reduces ALT (alanine aminotransferase) level by at least about 7.7% (e.g., at least about 14%, at least about 26%, at least about 36%, or at least about 39%), or at least about 40% (e.g., at least about 51%, at least about 53%, or at least about 60%) compared with that prior to the use, as measured in serum, for example about day 71 or 113 from the first use.

[0122] Embodiment 71: The use of any one of Embodiments 53-70, wherein the use reduces AST (aspartate aminotransferase) level by at least about 7.5% (e.g., at least about 15%), or at least about 20% (e.g., at least about 24%, or at least about 28%) compared with that prior to the use, as measured in serum, for example about day 71 or 113 from the first use.

[0123] Embodiment 72: The use of any one of Embodiments 53-71, wherein the use reduces or slows an increase of liver fat fraction (e.g., a reduction of at least 4-41% or at least 7-8%) compared with that prior to the use, as measured by magnetic resonance imaging, for example about day 71 or 113 from the first use.

[0124] Embodiment 73: The use of any one of Embodiments 53-72. wherein the use reduces or slows an increase of liver stiffness (e.g., a reduction of at least 4-37%) compared with that prior to the use, as measured by transient elastography, for example about day 71 or 113 from the first use.

[0125] Embodiment 74: The use of any one of Embodiments 53-73, wherein the chemical structure of the ligand comprises:

[0126] Embodiment 75: The use of Embodiment 74, wherein the compound is in a free acid form having a chemical structure as illustrated in FIG. 5A to 5D.

[0127] Embodiment 76: The use of any one of Embodiments 53-73, wherein the chemical structure of the ligand comprises:

[0128] Embodiment 77: The use of Embodiment 76, wherein the compound is in a sodium salt form having a chemical structure as illustrated in FIG. 6A-6D.

[0129] Embodiment 78: The use of any one of Embodiments 53-77, wherein the use reduces GGT (gamma-glutamyl transferase) level by at least about 1.4% or at least 8.3% or at least 9%, compared with that prior to the use, as measured in serum, for example at about day 71 or 113 from the first use.

[0130] In some cases, one or more human subjects treated with a method described herein achieve => 1 stage (i.e., equal to or greater than 1 stage) improvement in histological fibrosis with no worsening of NASH, for example at week 52 from the start of the treatment. In some instances, improvement in histological fibrosis is assessed with Clinical Research Netw ork (CRN) Scoring. In some instances, no w orsening of NASH is defined as no increase in the NAFLD Activity' Score (NAS) for steatosis, ballooning, or inflammation.

[0131] In some cases, one or more human subjects treated with a method described herein achieve NASH resolution with no w orsening of fibrosis, for example at week 52 from the start of the treatment. In some instances, NASH resolution is defined as a ballooning scoreof 0 and an inflammation score of 0-1. In some instances, no worsening of fibrosis is defined as no increase in CRN fibrosis score.

[0132] In some cases, one or more human subjects treated with a method described herein achieve >30% relative reduction in liver fat from baseline at week 24 and week 52 from the start of the treatment, for example measured by MRI-PDFF.

[0133] In some cases, a treatment disclosed herein results in about 80% to about 99% (e.g.. >85% or >90%) reduction at Week 8 post-dose, about 60% to about 90% (e.g., >75%) at Week 12 post-dose, and / or about 40% to about 80% (e.g., >60%) at Week 16 post-dose, in HSD17B13 protein observed in a liver biopsy sample, compared to baseline (pre-dose), whereby the NAFLD is treated and / or the dose can be reduced. In some cases, a treatment disclosed herein results in about 80% to about 99% reduction at Week 8 post-dose, about 60% to about 90% at Week 12 post-dose, and / or about 40% to about 80% at Week 16 postdose, in HSD17B13 mRNA expression observed in a liver biopsy sample, compared to baseline (pre-dose), whereby the NAFLD is treated and / or the dose can be reduced. In some cases, a treatment disclosed herein results in Cmax of 1000-3000 ng / mL. e.g., 1100-1200. 1600-1800, or 2200-2400 ng / mL, and / or AUC0-24h of 10000-30000 h ng / mL, e.g., 12000- 13500, 18000-20000, or 24000-26000 h ng / mL. In some cases, a treatment disclosed herein results in significant reductions (e.g., at least 10%) in ALT, AST, cytokeratin-18, RBP-4, liver fat (using MRI-PDFF), LSM. ELF score, Pro-3 (at EoS) marker measurements compared to baseline (pre-dose).EXAMPLESExample 1. Synthesis and Formulation of HSD17B13 RNAi Agents

[0134] The compounds of RNAi agents suitable for use herein can be synthesized using standard phosphoramidite technology on solid phase oligonucleotide synthesis as is known in the art. Commercially available oligonucleotide synthesizers (e.g., MerMade96E® (Bioautomation) or MerMadel2® (Bioautomation)) may be used. Syntheses can be performed on a solid support made of controlled pore glass (CPG, 500 A or 600 A, obtained from Prime Synthesis, Aston, PA, USA). The monomer positioned at the 3’ end of the respective strand may be attached to the solid support as a starting point for synthesis. All 2'-modified RNA phosphoramidites, and inverted abasic phosphorami dites can be purchased commercially. Ligand-containing phosphoramidites can be synthesized that are suitable for addition to the 5' end of the sense strand. Standard cleavage, deprotection, purification, and annealing steps can be utilized as is known in the art. Further description related to thesynthesis of HSD17B13 RNAi agents may be found, for example, in International Patent Application Publication No. WO 2020 / 061177 (Patent Application No. PCT / US2019 / 051707) and WO 2018 / 044350 (PCT / US2017 / 021147), each of which is incorporated by reference herein in its entirety7. HSD17B13 RNAi agents can then be formulated by dissolving in suitable pharmaceutically acceptable excipients.Example 2. Phase 2a, Single Dose, Open-Label, Dose-Exploration Study to Assess The PK-PD Activity, Safety, And Tolerability of HSD17B13 RNAi Agent In Adult Participants with NASH And Suspected NASH

[0135] The test compound exemplified herein reduces expression of HSD 17(313 messenger RNA (mRNA) in hepatocytes. Its sequences are shown in Table 4, and physical properties are listed in Table 5.Table 4. Test CompoundTable 5. Properties of the Test Compound

[0136] Full chemical structure representations are shown in FIG. 5 A to 5D (free acid form) and FIG. 6A to 6D (sodium salt form).

[0137] This is a Phase 2a, single dose, open-label, multi-center, dose exploration study designed to assess the PK and PD activity, safety, and tolerability of the test compound in adult participants with confirmed NASH or suspected NASH. The study includes 4 treatment arms and all participants receives a single dose via SC injection. Following a screening period of 6 weeks, eligible participants are assigned into 1 of the 4 treatment arms: 25 mg, 200 mg, 400 mg or 800 mg. There are 12 participants allocated to each treatment arm. Eachtreatment arm are split into 2 cohorts. The total study duration, including screening procedures, dosing and post-dose follow-up, is approximately 24 weeks for each participant. All participants have a liver biopsy performed prior to dosing on Day 1 to collect liver tissue for quantification of baseline hepatic HSD17B13 mRNA and protein. Participants are assigned for a second liver biopsy either at 8, 12 or 16 weeks post-dose. Tissue from the second liver biopsy is used to assess extent of knockdown of hepatic HSD17B13 mRNA and HSD17B13 protein compared to baseline.Objectives, Endpoints, and Estimands:Table 6. Objectives and EndpointsPrimary Objective Estimands

[0138] The primary clinical questions of interest for the primary objective are:1) What is the predicted mean percent change from baseline of HSD17B13 protein levels, derived from the PK-PD target engagement model at each treatment arm in adult participants with proven or suspected NASH regardless of prohibited medication, increase in self-reported alcohol consumption, or delays in post-dose biopsy times?2) What is the predicted mean percent change from baseline of HSD17B13 mRNA levels, derived from the PK-PD target engagement model at each treatment arm in adult participants with proven or suspected NASH regardless of prohibited medication, increase in self-reported alcohol consumption, or delays in post-dose biopsy times?The estimands are described by the following attributes in Table 7:

[0139] Rationale for primary estimand: It is of interest to estimate the mean and 95% CI of percent change from baseline based on the PK-PD target engagement model regardless of the use of prohibited medication or increased self-reported alcohol consumption to reflect the real-world setting.

[0140] Rationale for additional estimand: It is of interest to estimate the mean and 95% CI of percent change from baseline based on the PK-PD target engagement model up until the use of prohibited medication and / or increased alcohol consumption which may influence the PK-PD target engagement model parameter estimates.Secondary Objective Estimand s

[0141] The secondary and safety study objectives are assessed using the endpoints described above. They share the same estimand elements of intervention, population, intercurrent events and strategies as the primary estimands.Table 8. Summary MeasuresOverall Study Design

[0142] This is a Phase 2a, single dose, open-label, multi-center, dose-exploration study designed to assess the PK and PD activity, safety, and tolerability of the test compound in adult participants with NASH and suspected NASH. The study include 4 treatment arms (with the possibility of 5 total treatment arms), all receiving a single dose via SC injection.Following a screening period of 6 weeks, it is planned that eligible participants are assigned into 1 of the 4 treatment arms: 25 mg, 200 mg, 400 mg or 800 mg. There are approximately 12 participants allocated to each treatment arm, for a total of 48 participants planned across 4 treatment arms. Each treatment arm are split into 2 dose cohorts. Approximately 6 participants are enrolled in each dose cohort. Participants are randomized to dose cohorts, where feasible. The total study duration, including screening procedures, dosing and postdose follow-up, is approximately 24 weeks for each participant. Part A of Figure 7 outlines enrollment into the dose cohorts. Once a schedule has been enrolled, the next commences. If an additional treatment arm (e.g., 600 mg as intermediate prior to 800 mg) is recommended, this take at Schedule 8 as Cohort 4. Part B of Figure 7 outlines the coordinated data review and the number of participants from a given dose cohort included in the data review upon completion of the associated schedules outlined in the part A.

[0143] All participants have a liver biopsy performed prior to dosing on Day 1 to collect liver tissue for quantification of baseline hepatic HSD17B13 mRNA and protein. In Cohorts la, 2a, 3a, and 4a (25 mg, 200 mg, 400 mg, and 800 mg respectively), participants are assigned for a second liver biopsy 8 weeks post-dose, while participants in Cohorts lb. 2b. and 3b (25 mg, 200 mg and 400 mg respectively) have a liver biopsy performed 12 weeks and Cohort 4b (800 mg) have a liver biopsy performed at 16 weeks post-dose. Tissue from the second liver biopsy is used to assess extent of knockdow n of hepatic HSD17B13 mRNA and HSD17B13 protein compared to baseline. A fifth treatment arm may be added to the study, with the highest dose in this study not to exceed 800 mg. If a fifth treatment arm is added, the potential total number of participants for this study is approximately 60 (n= up to 12 parti cipants / treatment arm).For doses 400 mg and above:

[0144] Sentinel dosing is employed for the first participant of each treatment arm, starting from 400 mg and above. The sentinel participant is dosed in advance of the rest of the participants in their respective treatment arm. The sentinel participant is observed for approximately 3 days post-dose, with the first 24 hours confined at the clinical site. The participant may be discharged on Day 2 and return to the clinical site for outpatient visits.

[0145] The test compound is rapidly cleared in plasma with a half-life of approximately 4 hours. An observation period of 3 days is considered sufficient duration for clearance of higher dosing in this study. If it is deemed necessary, following review of the data fromsentinel dosing of a participant, sentinel dosing of an additional participant may be required.

[0146] Safety and PK data (where available) are reviewed when 6 participants in a treatment arm reach Day 75 and assessed prior to progression to the next dose escalation up to 800 mg (treatment arm 4 [or treatment arm 5, if applicable]), if recommended.Scientific Rationale for Study Design

[0147] The test compound was well tolerated at doses up to 200 mg and demonstrated effective pharmacological response via reduced hepatic HSD17B13 mRNA and HSD17B13 protein levels. As a non-invasive biomarker for target engagement is not yet available to facilitate dose selection, in this study, liver biopsies at baseline and after dosing are required to measure the effect of the test compound on HSD17B13 gene and protein expression. This study measures baseline hepatic HSD17B13 mRNA and protein levels and post-dose levels at multiple time points after a single dose, ranging from 25 mg up to 800 mg. The outcome of this study enables characterization of the dose-exposure-target engagement relationship as it pertains to the extent and duration of hepatic HSD17B13 knockdown and investigate correlations with other non-invasive tests utilized in NASH trials (e.g., Pro-C3, ELF, ALT, FibroScan®, MRI-PDFF).Justification for Dose

[0148] The goal of this study is to characterize the target engagement of the test compound on hepatic HSD17B13 knockdown over time following single SC administration of ascending doses. The doses to be included in this study are based on the observed safety and PK / PD modeling of a previously conducted Phase l / 2a study data.

[0149] In the previous study, healthy participants received single escalating doses of 25 mg, 50 mg, 100 mg, or 200 mg SC on Day 1, and confirmed or suspected NASH participants received 25 mg, 100 mg, or 200 mg on Day 1 and Day 29. The test compound was well tolerated in both NHVs and participants with confirmed or suspected NASH, with no serious AEs, no AE-related study or drug discontinuations, no DLTs, or deaths. Hepatic HSD17B13 mRNA and protein levels. PK, and serum ALT data were obtained from the NHV and NASH participants who received SC doses of 25 mg to 200 mg. A population PK / PD model was developed and can successfully predict the reduction in ALT over 113 days post-dose and hepatic HSD17B13 protein up to 71 days post-dose following a single or two SC injections of up to 200 mg of the test compound.

[0150] A study with a broader dose range would provide a better understanding of the extent and duration of target engagement (hepatic HSD17B13 protein and mRNA knockdown). A wide range of doses (25 mg, 200 mg, 400 mg, and 800 mg) was selected based on the simulation results (FIG. 1 to FIG. 4) and the previously observed PK target engagement model of change in HSD17B13 protein to investigate maximum target engagement and duration of inhibition of hepatic HSD17B13 translation. A placebo group is not necessary as the objective of the study is to measure the change in HSD17B13 protein and mRNA after administration of study intervention in addition to reduce patient burden and feasibility concerns. The lowest dose of 25 mg was selected given the minimal effect observed in the previous study, which may help inform the lower portion of the exposureresponse curve.Anticipated Exposure for the Selected Dose Range

[0151] Predicted single dose plasma PK exposures (Cmax and AUC) of the test compound for the planned doses are provided in Table 9. These are planned doses with the assumption of linear PK at doses greater than the 200 mg evaluated previously, meaning if saturation is observed at the higher doses, the likelihood of achieving NOAEL (no-observed-adverse- effect level) exposures may also be reduced / modulated. The specific doses in this study are subject to change based upon emerging PK, PD, and safety data gathered in this study.Table 9. Predicted Single Dose PK for Proposed Doses Following SC AdministrationPopulation Inclusion Criteria

[0152] Participants are eligible to be included in the study only if all following criteria apply:1. Participant must be 18 to 75 years of age inclusive, at the time of screening.Participant must be capable of giving signed informed consent which includes compliance with the requirements and restrictions listed in the ICF and in this protocol. Participants who have proven diagnosis of NASH or suspected NASH demonstrated by the following: a. Proven diagnosis of NASH confirmed by historical liver biopsy (performed within 12 months prior to screening)OR b. Suspected NASH is defined as presence of: T2DM or pre-T2DM, OR at least 2 metabolic risk factors (e.g., obesity, dyslipidemia, hypertension), AND all of the criteria below: o Evidence of hepatic steatosis >8 % measured by centrally- assessed MRI-PDFF o ALT >ULN (Based on central laboratory measurement, 1 repeat test is allowed) o FibroScan® LSM >7.5 kPA (1 repeat test is allowed) Participants using any of the following medications can be included, but only if the dose has been stable for > 6 months prior to Screening 1 : glucagon-like peptide-1 agonists, PPAR agonists (e.g., pioglitazone, saroglitazar), sodium glucose cotransporter 2 inhibitors; thyroid hormone receptor agonists; famesoid X receptor agonists; or Vitamin E (at doses greater than or equal to 800 lU / day). Participant’s laboratory parameters must meet the criteria below:• ALT and AST <5xULN (or <250 U / L)• ALP <2xULN or <250 U / L• Total bilirubin < ULN (unless due to a diagnosis of Gilbert’s syndrome or hemolytic anemia) o For participants with Gilbert's syndrome or hemolytic anemia, elevated direct (conjugated) bilirubin <0.3 mg / dL• Platelets >LLN• INR <1.3 (unless due to anticoagulant)• Albumin >3.5g / dL• HbAl c <9.5%• Urinary7albumin creatinine ratio <500 mg / gBased on central laboratory measurement, 1 repeat test per laboratory parameter is allowed if clinically indicated.6. A female participant is eligible to participate, if she is not pregnant or breastfeeding and one of the following conditions applies:• Is a woman of childbearing potential WOCBP:• For permanently sterile individuals due to an alternate medical cause other than the above, (e.g., Mullerian agenesis, androgen insensitivity, gonadal dysgenesis), investigator discretion should be applied to determining study entry. If reproductive status is questionable, additional evaluation should be considered.• Is a WOCBP and using a contraceptive method that is highly effective, with a failure rate of <1%. Contraceptive measures must start from the time of negative serum pregnancy test at screening (Week -10 to Week-4) and at least 28 days before administration of study intervention. Contraceptive measures should then continue for at least 18 weeks after administration of study intervention. The investigator should evaluate the potential for contraceptive method failure (e.g., noncompliance, recently initiated in relationship to the first dose of study intervention. o A WOCBP must have a negative highly sensitive pregnancy test (urine as required by local regulations) within 24 h before administration of study intervention.Study InterventionTable 10. Study Intervention AdministeredTable 11. Study Arm(s)Target Engagement and Pharmacodynamic Assessments in LiverLiver-Biopsy-Based Assessments

[0153] Samples from biopsies collected from the liver at baseline and following a single dose undergo analysis to determine hepatic HSD17B13 protein and mRNA expression levels to determine target engagement.HSD17B13 Protein Expression

[0154] The effect of the test compound on hepatic HSD17B13 protein expression levels is assessed using the electrochemiluminescence immunoassay.HSD17B13 mRNA Expression

[0155] The effect of the test compound on hepatic HSD17B13 mRNA expression is assessed using reverse transcription-quantitative polymerase chain reaction.Physical Examinations

[0156] A complete physical examination includes, at a minimum, assessments of the cardiovascular, respiratory, gastrointestinal (including liver and spleen), dermatologic, and neurological systems. Height and weight (on an empty bladder and wearing light clothing) are measured and recorded.

[0157] A symptom-directed physical examination is performed only if the participant indicates during questioning that there might be some underlying pathology(ies) or if deemed necessary by the investigator or delegate. Significant weight gain or loss may require further physical examination at the discretion of the investigator.

[0158] If an exam is done by a HHS professional, a symptom-directed examination is performed.Vital Signs

[0159] Temperature, pulse rate, respiratory rate, and blood pressure should be assessed prior to blood collection for laboratory tests.

[0160] Blood pressure and pulse measurements are assessed with a completely automated device. Manual techniques are used only if an automated device is not available.

[0161] Blood pressure and pulse measurements should be taken sitting and preceded by at least 5 minutes of rest for the participant in a quiet setting without distractions (e.g., television, cell phones).

[0162] One pulse and 3 blood pressure measurements are taken. For blood pressure measurements, 3 consecutive blood pressure readings are recorded at intervals of at least 1 minute. The average of the 3 blood pressure readings is recorded.Electrocardiograms

[0163] Triplicate 12-lead ECGs are obtained using an ECG machine that automatically calculates the heart rate and measures PR, QRS, QT, and QTc intervals.

[0164] The QTc is the QT interval corrected for heart rate according Fridericia’s formula (QTcF). It is either machine-read or manually over-read.Fridericia’s fonnula:

[0165] ECG readings should be taken after 5 minutes rest in the supine position.

[0166] At each timepoint at which triplicate ECGs are required, 3 individual ECG tracings should be obtained as closely as possible in succession, but no more than 2 minutes apart.Genetic Analyses

[0167] A 6 mL blood sample for DNA isolation is collected from all participants who have consented to participate in the study. The DNA from the samples is genotyped for HSD17B13 rs72613567 to exclude participants who are homozygous for the TA variant to reduce noise-to-signal for the objectives / endpoints that relate to target engagement, PD and PK, as it is not fully understood whether knockdown of the homozygous variant transcript is significantly different than the wild- pe or heterozygous variants.

[0168] The assessment of the HSD17B13 genoty pe is performed by the CLIA approvedGenotyping Assay (Illumina SNP Array). SNP array is a type of DNA microarray containing designed probes harboring the SNP positions, which is hybridized with fragmented DNA to determine the specific alleles of all SNPs on the array for the hybridized DNA sample.Statistical Analyses

[0169] Final analyses are performed after the completion of the study and final dataset authorization. Where appropriate, baseline assessment is the last available assessment prior to the time of dosing.Primary Endpoints / Estimands Analysis

[0170] Details of estimand strategies for primary' endpoints can be found above. The primary analysis should fit the target engagement model of inhibition of HSD17B13 protein and mRNA expression. The following analyses are performed for the primary endpoints:Secondary Endpoints / Estimands Analyses

[0171] The following analyses are performed for the secondary endpoints:Safety AnalysisPharmacodynamics Analysis

[0172] This analysis validates the extrapolation of the test compound population PK target engagement model. As described above, the model was developed with data from a Phase l / 2a study of subcutaneous doses up to 200 mg. This population PK target engagement model simulated 1000 subjects after a single subcutaneous dose of the test compound at the respective dose levels and was compared to the Phase 2a study data (200 and 400 mg) presented herein. Overall, the population PK target engagement model was able to predict the clinically observed test compound plasma concentration data in the 11 participants enrolled in the Phase 2a study, as shown in FIG. 8 where 95% of the clinically observed data points were well within the model predicted range. The clinically observed data along with the model predictions supports that plasma concentrations increased substantially as the doses increase beyond 200 mg and suggests the greater exposure resulted in much greater target engagement.

[0173] Body weight can also be an important driver of the systemic concentrations, and the same dose can result in different exposures [area under the concentration-time curve (AUCO-24)] depending on the body weight of the individual. FIG. 9 clearly shows the impact of body weight on the systemic exposures. As the body weight increases, the corresponding exposures can decrease. Due to this weight effect, the exposures after a single subcutaneous dose of 200 and 400 mg in different individuals of drastically different body weights can result in some overlap.

[0174] The increased target engagement with increasing doses is shown in FIG. 10 in which the average change from baseline in hepatic HSD17P13 protein was marginally greater with a 400 mg dose of the test compound as compared to a 200 mg dose. The dose increase appears to provide a longer duration of hepatic HSD17P13 protein knockdown as predicted by the population PK target engagement model FIG. 2 and validated by the clinically observed data. Consistent with the model predictions, the dose increase resulted in a less variable, more complete knockdown of HSD 17(313 protein.

[0175] The reduced hepatic HSD17P13 protein led to reductions in serum biomarkers of liver injury and inflammation. The mean reduction of ALT levels were approximately 50% with 400 mg (FIG. 11A). Similarly, AST levels were also reduced by approximately 25% to 30% for the 400 mg dose (FIG. 11B).

Claims

CLAIMSWhat is claimed is:

1. A method for treating a non-alcoholic fatty liver disease in a human subject in need thereof, comprising administering to the human subject a compound or a pharmaceutically acceptable salt thereof, at a dose of about 300 to about 800 mg, wherein the dose is calculated based on a free acid form of the compound, wherein the compound comprises a double-stranded oligonucleotide linked to a ligand, wherein the double-stranded oligonucleotide reduces expression of HSD17P13 in the human subject, and wherein the ligand comprises a chemical structure of:((NAG37)s), a stereoisomer thereof, or a pharmaceutically acceptable salt thereof.

2. The method of claim 1, wherein the administration is repeated at an interval of about: 1, 2, or 3 months, or longer.

3. The method of claim 1 or 2, wherein the dose is about 800 mg.

4. The method of claim 1 or 2. wherein the dose is from about 300 to about 400 mg.

5. The method of claim 4, wherein the dose is about 300 mg.

6. The method of claim 4, wherein the dose is about 400 mg.

7. The method of any one of claims 1-6, wherein the compound is in a sodium salt form.

8. The method of any one of claims 1-7, wherein the compound is administered by an injection.

9. The method of claim 8, wherein the injection is a subcutaneous injection.

10. The method of any one of claims 1-9, wherein the compound is present in a single unit dosage form.

11. The method of any one of claims 1-10, wherein the non-alcoholic fatty liver disease is non-alcoholic steatohepatitis (NASH).

12. The method of any one of claims 1-11, wherein the human subj ect has a HSD 17 13 rs72613567 mutation, PNPLA3 rs738409 (I148M) mutation, or a combination thereof.

13. The method of any one of claims 1-12, wherein the double-stranded oligonucleotide comprises an antisense strand that comprises a modified nucleotide sequence that differs by 0 or 1 nucleotides from one of the following sequences (5 '3'): usCfsasUfcUfaucagAfcUfuCfuUfaCfsg (nucleotide sequence SEQ ID NO: 1); usCfsasUfcUfaUfcAfgAfcUfuCfuUfaCfsg (nucleotide sequence SEQ ID NO:5); usGfsasUfcCfaAfaAfaUfgUfcCfuAfgGfsc (nucleotide sequence SEQ IDNO:6); or usGfsasUfcCfaaaaaUfgUfcCfuAfgGfsc (nucleotide sequence SEQ ID NO:7); and wherein a. c, g, and u represent 2'-O-methyl adenosine, 2'-O-methyl cytidine, 2'-O-methyl guanosine, and 2'-O-methyl uridine, respectively; Af, Cf, Gf, and Uf represent 2'-fluoro adenosine, 2'-fluoro cytidine, 2'-fluoro guanosine, and 2'-fluoro uridine, respectively; and s represents a phosphorothioate linkage.

14. The method of any one of claims 1-13, wherein the double-stranded oligonucleotide comprises a sense strand that comprises a modified nucleotide sequence that differs by 0 or 1 nucleotides from one of the following sequences (5' -> 3'): cguaagaaGfuCfuGfauagauga (nucleotide sequence SEQ ID NO:3); cguaagaaGfUfCfugauagauga (nucleotide sequence SEQ ID NO: 8); gccuaggaCfAfUfuuuugiauca (nucleotide sequence SEQ ID NO:9); or gccuaggaCfaUfuUfuugiauca (nucleotide sequence SEQ ID NOTO);and wherein a. c, g, i, and u represent 2'-0-methyl adenosine, 2'-O-methyl cytidine. 2'-O-methyl guanosine, 2'-O-methyl inosine, and 2'-O-methyl uridine, respectively; Af, Cf, Gf, and Uf represent 2'-fluoro adenosine, 2'-fluoro cytidine, 2'- fluoro guanosine, and 2'-fluoro uridine, respectively; and s represents a phosphorothioate linkage.

15. The method of any one of claims 1-14, wherein the double-stranded oligonucleotide comprises an antisense strand that comprises a sequence (5' 3'): usCfsasUfcUfaucagAfcUfuCfuUfaCfsg (nucleotide sequence SEQ ID NO: 1), and a sense strand that comprises a sequence (5’ -> 3'):(NAG37)s(inv Ab)scguaagaaGfuC fuGfauagaugas(inv Ab) (nucleotide sequenceSEQ ID NO: 11), and wherein a, c, g, and u represent 2'-O-methyl adenosine, 2'-O-methyl cytidine. 2'-O-methyl guanosine, and 2'-O-methyl uridine, respectively; Af, Cf, Gf, and Uf represent 2'-fluoro adenosine. 2'-fluoro cytidine, 2'-fluoro guanosine, and 2'-fluoro uridine, respectively; s represents a phosphorothioate linkage; and (invAb) represents an inverted abasic deoxyribose residue.

16. The method of any one of claims 1-15, wherein the method reduces the expression of HSD17013 mRNA in the human subject by at least about 50% compared with that prior to the administration, as measured by quantitative Reverse Transcriptase Polymerase Chain Reaction (qRT-PCR) with a liver biopsy sample from the human subject.

17. The method of any one of claims 1-16, wherein the method reduces the expression of HSD17013 protein in the human subject by at least about 30% compared with that prior to the administration, as measured by Western Blot with a liver biopsy sample from the human subject.

18. The method of any one of claims 1-17, wherein the method reduces ALT (alanine aminotransferase) level in the human subject by at least about 40% compared with that prior to the administration, as measured in the human subject’s serum.

19. The method of any one of claims 1-18, wherein the method reduces AST (aspartate aminotransferase) level in the human subject by at least about 20% compared with that prior to the administration, as measured in the human subject’s serum.

20. The method of any one of claims 1-19, wherein the method reduces or slows an increase of the human subject's liver fat fraction compared with that prior to the administration, as measured by magnetic resonance imaging.

21. The method of any one of claims 1-20, wherein the method reduces or slows an increase of the human subject’s liver stiffness compared with that prior to the administration, as measured by transient elastography.

22. The method of any one of claims 1-21, wherein the chemical structure of the ligand comprises:

23. The method of claim 22, wherein the compound is in a free acid form having a chemical structure as illustrated in Figure 5 A to 5D.

24. The method of any one of claims 1-21, wherein the chemical structure of the ligand comprises:

25. The method of claim 24, wherein the compound is in a sodium salt form having a chemical structure as illustrated in Figure 6 A to 6D.

26. A compound or a pharmaceutically acceptable salt thereof for use in treating nonalcoholic fatty liver disease in a human subject, wherein the use comprises administering to the human subject the compound or pharmaceutically acceptable salt thereof, at a dose of about 300 to about 800 mg, wherein the dose is calculated based on a free acid form of the compound, wherein the compound comprises a double-stranded oligonucleotide linked to a ligand, wherein the double-stranded oligonucleotide reduces expression of HSD17P13 in the human subject, and wherein the ligand comprises a chemical structure of:stereoisomer thereof, or a pharmaceutically acceptable salt thereof.

27. The compound or pharmaceutically acceptable salt thereof for use according to claim 26, wherein the compound is in a free acid form having a chemical structure as illustrated in Figure 5A to 5D or in a sodium salt form having a chemical structure as illustrated in Figure 6A to 6D.

28. Use of a compound or a pharmaceutically acceptable salt thereof for the manufacture of a medicament for the treatment of non-alcoholic Patty liver disease, wherein the compound or a pharmaceutically acceptable salt thereof is used at a dose of about 300 to about 800 mg. wherein the dose is calculated based on a free acid form of the compound, wherein the compound comprises a double-stranded oligonucleotide linked to a ligand, wherein the double-stranded oligonucleotide reduces expression of HSD17P13 in the human subject, and wherein the ligand comprises a chemical structure of:stereoisomer thereof, or a pharmaceutically acceptable salt thereof.

29. The use of claim 28. wherein the compound is in a free acid form having a chemical structure as illustrated in Figure 5 A to 5D or in a sodium salt form having a chemical structure as illustrated in Figure 6A to 6D.

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