Compounds and their use for the treatment of α1-antitrypsin deficiency

Abstract

The present invention relates to carboxylic acid compounds designated by formula (1), and pharmaceutical compositions containing the compounds. The compounds may be inducers of alpha 1-antitrypsin (A1AT) and may be used in the treatment of diseases or disorders, such as alpha 1-antitrypsin deficiency (A1AD or AATD).

Description

Technical Field

[0001] The present invention relates to certain carboxylic acids and their medical uses.

Background Art

[0002] α1-Antitrypsin (A1AT) is a member of the serpin superfamily produced by the liver and secreted into the blood. It inhibits various serine proteases, particularly neutrophil elastase. When the blood level of A1AT is low, excessive neutrophil elastase activity degrades lung tissue, resulting in respiratory complications such as chronic obstructive pulmonary disease (COPD).

[0003] The reference range of A1AT in the blood is 0.9 - 2.3 g / L. Lower levels are typical of α1-antitrypsin deficiency (A1AD or AATD), a genetic disorder caused by mutations in the SERPINA1 gene encoding A1AT. The most common cause of AATD, the Z mutation, is a substitution of glutamic acid to lysine at position 366 of A1AT (UniProtKB - P01009 (A1AT_HUMAN)) corresponding to position 342 in the mature protein (Z A1AT). The Z mutation affects the folding of A1AT, resulting in only a small proportion acquiring the native / active state. The remainder is cleared as misfolded protein or accumulates in the liver as stable polymers. As a consequence of misfolding, homozygous carriers (ZZ) of the Z mutation have plasma levels of A1AT that are 10 - 15% of normal and are predisposed to being carriers of COPD. The accumulation of Z A1AT polymers in hepatocytes predisposes to being carriers of cirrhosis, liver cancer, and other liver pathologies.

[0004] Current treatment for the lung symptoms of AATD involves augmentation therapy using A1AT concentrates prepared from the plasma of blood donors. The US FDA has approved the use of four A1AT products: Prolastin, Zemaira, Glassia, and Aralast. The dosing is via once-weekly intravenous infusion. Augmentation therapy has been demonstrated to slow the progression of COPD. The liver symptoms of AATD (e.g., cirrhosis and cancer) are treated with steroids and liver transplantation. Research approaches towards improved treatment of liver symptoms include increased clearance of polymers through inhibition of Z A1AT polymerization and activation of autophagy. Research approaches towards improved treatment of both lung and liver symptoms are directed towards improving the folding and secretion of Z A1AT.

[0005] Elliott et al (Protein Science, 2000, 9, 1274-1281) have described the X-ray crystal structure of A1AT and identified five cavities that are potential targets for rational drug design to develop agents that affect Z A1AT polymerization.

[0006] Parfrey et al (J. Biol. Chem., 2003, 278, 35, 33060-33066) have further defined a single cavity that is a potential target for rational drug design to develop agents that affect Z A1AT polymerization.

[0007] Knaupp et al (J. Mol. Biol., 2010, 396, 375-383) have shown that bis-ANS (4,4’-dianilino-1,1’-binaphthyl-5,5’-disulfonate) can bind to Z A1AT in a 1:1 stoichiometry and with a K of 700 nM d but cannot bind to wild-type A1AT (M).

[0008] Chang et al (J. Cell. Mol. Med., 2009, 13, 8B, 2304-2316) reported a series of peptides, including Ac-TTAI-NH2, which inhibit Z A1AT polymerization.

[0009] Burrows et al (Proc. Nat. Acad. Sci., 2000, 97, 4, 1796-1801) showed that a series of non-selective chaperones, including 4-phenylbutyric acid, glycerol and trimethylamine oxide, can increase Z A1AT levels in cell supernatants and mouse models.

[0010] Bouchecareilh et al. (Journal of Biological Chemistry, 2012, 287, 45, 38265-38278) described the use of histone deacetylase inhibitors, particularly SAHA (suberoylanilide hydroxamic acid), to increase the secretion of both wild-type (M)A1AT and Z A1AT from cells.

[0011] Berthelier et al (PLOS ONE, May 11, 2015) demonstrated that S-(4-nitrobenzyl)-6-thioguanosine can prevent Z A1AT polymerization in vitro.

[0012] Mallya et al (J. Med. Chem., 2007, 50, 22, 5357-5363) described that a series of phenols, such as N-(4-hydroxy-3,5-dimethylphenyl)-2,5-dimethylthiophene-3-sulfonamide, can block the polymerization of Z A1AT in vitro.

[0013] Huntington (XIIIth International Symposium on Proteinases, Inhibitors and Biological Control, September 23, 2012, and 7 thThe International Symposium on Serpin Biology, Structure and Function (April 1, 2014) discussed the cavity from the X-ray crystal structure of Z A1AT, a potential target for rational drug design to develop agents that affect Z A1AT polymerization.

[0014] U.S. Patent No. 8,436,013 (B2) discloses various structures that can increase the secretion of Z A1AT from cells in the micromolar concentration range.

[0015] Angewandte Chemie International Edition vol 56, no 33, 2017, 9881-9885 discloses 1-((4-chlorophenyl)sulfonyl)piperidine-4-carboxylic acid as a herbicide.

[0016] Journal of Applicable Chemistry vol 2, no 6, 2013, 1501-1508 discloses the synthesis of 1-(4-(trifluoromethyl)phenylsulfonyl)piperidine-4-carboxylic acid as an antibacterial agent.

[0017] U.S. Patent Application Publication No. 2011 / 0065707 (A1) discloses the use of 1-(2-chlorobenzene-sulfonyl)-piperidine-4-carboxylic acid as a reagent.

[0018] European Patent Application Publication No. 0520336 (A2) discloses 1-(8-quinoyl-sulfonyl)-piperidine-4-carboxylic acid.

[0019] International Publication No. 2019 / 243841 (A1) discloses oxindoline-4-carboxamide compounds as modulators of alpha-1-antitrypsin and their use in the treatment of diseases associated with alpha-1-antitrypsin.

[0020] International Publication No. 2020 / 081257 (A1) discloses pyrrolo-indazolyl-propanoic acid compounds as modulators of alpha-1-antitrypsin.

[0021] U.S. Patent Application Publication No. 2020 / 0361939 (A1) discloses further pyrrolo-indazolyl-propanoic acid compounds as modulators of alpha-1-antitrypsin.

[0022] A prior art search based on the structure of 1-((2-(trifluoromethyl)phenyl)sulfonyl)piperidine-3-carboxylic acid was carried out after the present invention was made. The closest prior art molecule identified retrospectively by the search was the racemic compound, 1-((2-(trifluoromethyl)phenyl)sulfonyl)piperidine-3-carboxylic acid (CAS Registry Number 891392-68-0). This compound is listed as being commercially available from Aurora, ChemDiv and FCH Group, but no publications are recorded. Another close prior art molecule is 1-(1-tosyl-1,2,5,6-tetrahydropyridin-3-yl)ethan-1-one (Example 16 in U.S. Patent No. 9084782 (B2)). This compound is described as inhibiting angiogenesis and reducing cellular cholesterol levels (however, no biological data for this compound are provided in U.S. Patent No. 9084782 (B2)). SUMMARY OF THE INVENTION

[0023] According to one aspect of the present invention, a compound (carboxylic acid) of formula (1):

Chemical formula

[0024] wherein in formula (1), · R1 is an aryl or heteroaryl ring system which may be substituted or fused. ·m and n are independently 1, 2, or 3, provided that the combination where both n and m are 1 is excluded. ·When the combination of m and n results in a chiral center, both enantiomers and racemic mixtures may be included. and the compound is ·1-(quinolin-8-ylsulfonyl)piperidine-4-carboxylic acid or ·1-((2-chlorophenyl)sulfonyl)piperidine-4-carboxylic acid or ·1-((3-chlorophenyl)sulfonyl)piperidine-4-carboxylic acid or ·1-((2,3-dichlorophenyl)sulfonyl)piperidine-4-carboxylic acid or ·(S)-1-((3-fluorophenyl)sulfonyl)piperidine-3-carboxylic acid or ·(S)-1-((3-chlorophenyl)sulfonyl)piperidine-3-carboxylic acid or ·(R)-1-((3-fluorophenyl)sulfonyl)piperidine-3-carboxylic acid or ·(R)-1-((3-chlorophenyl)sulfonyl)piperidine-3-carboxylic acid or ·1-((4-chlorophenyl)sulfonyl)piperidine-4-carboxylic acid or ·1-((2-(trifluoromethyl)phenyl)sulfonyl)piperidine-4-carboxylic acid or ·1-((3-(trifluoromethyl)phenyl)sulfonyl)piperidine-4-carboxylic acid or ·1-((4-(trifluoromethyl)phenyl)sulfonyl)piperidine-4-carboxylic acid or ·1-((2,5-bis(trifluoromethyl)phenyl)sulfonyl)piperidine-4-carboxylic acid or ·1-((2-(trifluoromethoxy)phenyl)sulfonyl)piperidine-4-carboxylic acid or ·(S)-1-((2-(trifluoromethyl)phenyl)sulfonyl)pyrrolidine-3-carboxylic acid or · (R)-1-((2-(Trifluoromethyl)phenyl)sulfonyl)pyrrolidine-3-carboxylic acid or · (S)-1-((2-(Trifluoromethyl)phenyl)sulfonyl)piperidine-3-carboxylic acid or · (R)-1-((2-(Trifluoromethyl)phenyl)sulfonyl)piperidine-3-carboxylic acid There is provided a compound which is

[0025] Surprisingly, the compounds of the present invention have been found by us to be highly effective in increasing the level of correctly folded and thus active Z A1AT without affecting the secretion of wild-type (M) A1AT or the Siiyama variant of A1AT. BRIEF DESCRIPTION OF THE DRAWINGS

[0026]

Figure 1

BRIEF DESCRIPTION OF THE INVENTION

[0027] Also provided according to the invention is a mixture of two enantiomers of any compound according to claim 1 having a chiral center, which is a racemate or has one enantiomer in excess over the other enantiomer.

[0028] The compounds or mixtures of the invention may be in pharmaceutically acceptable salt or crystalline forms.

[0029] The term "pharmaceutically acceptable salt" refers to pharmaceutically acceptable organic or inorganic monosalts of the compounds of the invention. This can include those derived from bases such as sodium hydroxide, potassium hydroxide, lithium hydroxide, calcium hydroxide, 1-deoxy-2-(methylamino)-D-glucitol, magnesium hydroxide, zinc hydroxide, aluminum hydroxide, ferrous or ferric hydroxide, ammonium hydroxide or organic amines such as N-methylglucamine, choline, arginine and the like. For other examples of pharmaceutically acceptable salts, reference can be made to Gould (1986, Int J Pharm 33: 201-217).

[0030] According to a further aspect of the invention, there is provided a pharmaceutical composition comprising a compound or mixture of the invention described herein and a pharmaceutically or therapeutically acceptable excipient or carrier.

[0031] The term "pharmaceutically or therapeutically acceptable excipient or carrier" refers to a solid or liquid filler, diluent, or encapsulating substance that does not interfere with the effectiveness or biological activity of the active ingredient and is not toxic to the host, which may be either human or animal, to which it is administered. Depending on the particular route of administration, various pharmaceutically acceptable carriers, such as those well-known to those skilled in the art, may be used. Non-limiting examples include sugars, starches, cellulose and its derivatives, malt, gelatin, talc, calcium sulfate, vegetable oils, synthetic oils, polyols, alginic acid, phosphate buffered solutions, emulsifiers, isotonic saline, and pyrogen-free water.

[0032] All suitable modes of administration are contemplated in accordance with the present invention. For example, administration of the medicament may be via oral, subcutaneous, direct intravenous, slow intravenous infusion, continuous intravenous infusion, intravenous or epidural patient-controlled analgesia (PCA and PCEA), intramuscular, intrathecal, epidural, intracistemal, intraperitoneal, transdermal, topical, transmucosal, buccal, sublingual, transmucosal, inhalation, intranasal, intra-atricular, intranasal, rectal, or ocular routes. The medicament may be formulated in separate dosage units and may be prepared by any of the methods well-known in the pharmaceutical arts.

[0033] All suitable pharmaceutical dosage forms are contemplated. Administration of the medicament may be, for example, in the form of oral solutions and suspensions, tablets, capsules, lozenges, effervescent tablets, transmucosal films, suppositories, buccal products, oral mucosa-retentive products, topical creams, ointments, gels, films, and patches, transdermal patches, abuse-deterrent and abuse-resistant formulations, sterile solutions, suspensions, and depots for parenteral use, and the like, which may be administered as immediate release, sustained release, delayed release, controlled release, extended release, and the like.

[0034] Another aspect of the present invention is the use of the compounds or mixtures of the invention as defined herein in the manufacture of a medicament for the treatment of a disease or disorder.

[0035] A further aspect of the invention is the compound or mixture of the invention for use as an inducer of Z A1AT secretion.

[0036] There is further provided a compound or mixture of the invention as defined herein for use in the treatment of a disease or disorder.

[0037] The invention also encompasses a method of treating a disease or disorder comprising administering to a patient in need thereof a compound or mixture or pharmaceutical composition of the invention as defined herein.

[0038] The invention further encompasses the use of a compound or mixture of the invention as an inducer of Z A1AT secretion. The use may be in the treatment of a disease or disorder. Additionally or alternatively, the use may be in vitro, for example in an in vitro assay.

[0039] Diseases or disorders suitable for treatment according to related aspects of the invention are those characterized by low plasma levels of A1AT, such as AATD.

[0040] The invention also provides the use of a racemic compound of formula (1) in the manufacture of a medicament for the treatment of a disease or disorder, wherein the disease or disorder is AATD.

[0041] There is also provided the use of a racemic compound of formula (1) as an inducer of Z A1AT secretion.

[0042] The use of numerical ranges herein is clearly intended to include within the scope of the invention all individual integers within that range and all combinations of the numbers of the upper and lower limits within the broadest scope given for that range.

[0043] As used herein, the term "comprising" should be read as having both the meanings of "including" and "consisting of". As a result, when the present invention relates to a pharmaceutical composition "comprising a compound as an active ingredient", this expression is intended to cover both compositions in which other active ingredients may be present and compositions consisting of only one active ingredient as defined.

[0044] Unless otherwise defined, all scientific and technical terms used herein shall have the same meaning as commonly understood by one of ordinary skill in the art to which the present invention pertains. Similarly, all publications, patent applications, all patents, and all other references mentioned herein are hereby incorporated by reference in their entirety (where legally permissible).

[0045] Certain non-limiting specific examples of the present invention are described hereinafter.

Example

[0046] General Method The compound of formula 1 was prepared using the following synthetic procedure.

Chemical formula

[0047] Carboxylic acid (1 equivalent), potassium hydroxide (1 equivalent), and potassium carbonate (2 equivalents) were added to water and stirred. Sulfonyl chloride (1 equivalent) was added, and the reaction mixture was stirred at room temperature for 3 hours. The reaction mixture was cooled to 0 °C and acidified with 2M hydrochloric acid to obtain a white precipitate. This precipitate was dried and triturated with n-pentane to obtain the compound of formula (1).

[0048] Example 1: 1-(Quinolin-8-ylsulfonyl)piperidine-4-carboxylic acid The compound of Example 1 was prepared using the general method and 1-quinolin-8-ylsulfonyl chloride and piperidine-4-carboxylic acid.

[0049] 11H NMR (400 MHz, d6 DMSO) δ 12.26 (1H, s), 9.07 (1H, s), 8.54 (1H, d), 8.36 (1H, d), 8.30 (1H, d), 7.74 (1H, m), 7.70 (1H, m), 3.81 (2H, m), 2.82 (2H, m), 2.31 (1H, m), 1.82 (2H, m), 1.44 (1H, m).

[0050] Example 2: 1-((2-Chlorophenyl)sulfonyl)piperidine-4-carboxylic acid The compound of Example 2 was prepared using the general method and 1-(2-chlorophenyl)sulfonyl chloride and piperidine-4-carboxylic acid.

[0051] 1 1H NMR (400 MHz, d6 DMSO) δ 12.38 (1H, br s), 7.97 (1H, d), 7.88 (2H, m), 7.56 (1H, m), 3.60 (2H, m), 2.83 (2H, t), 2.40 (1H, m), 1.86 (2H, m), 1.48 (2H, m).

[0052] Example 3: 1-((3-Chlorophenyl)sulfonyl)piperidine-4-carboxylic acid The compound of Example 3 was prepared using the general method and 1-(3-chlorophenyl)sulfonyl chloride and piperidine-4-carboxylic acid.

[0053] 1 1H NMR (400 MHz, CDCl3) δ 7.77 (1H, s), 7.76 (1H, d), 7.63 (1H, m), 7.60 (1H, m), 3.68 (2H, m), 2.55 (2H, t), 2.37 (1H, m), 2.02 (2H, m), 1.86 (2H, m).

[0054] Example 4: 1-((2,3-Dichlorophenyl)sulfonyl)piperidine-4-carboxylic acid The compound of Example 4 was prepared using the general method and 1-(2,3-dichlorophenyl)sulfonyl chloride and piperidine-4-carboxylic acid.

[0055] 1 1H NMR (400 MHz, d6 DMSO) δ 12.36 (1H, br s), 7.97 (2H, d), 7.59 (1H, m), 3.64 (2H, m), 2.90 (2H, t), 2.43 (1H, m), 1.80 (2H, m), 1.54 (2H, m).

[0056] Example 5: (S)-1-((3-Fluorophenyl)sulfonyl)piperidine-3-carboxylic acid The compound of Example 5 was prepared using the general method and 1-(3-fluorophenyl)sulfonyl chloride and (S)-piperidine-3-carboxylic acid.

[0057] 1 1H NMR (400 MHz, CD3OD) δ 7.52 (2H, m), 7.48 (1H, m), 7.44 (1H, m), 3.73 (1H, d), 3.53 (1H, d), 2.60 (3H, m), 1.97 (1H, m), 1.81 (1H, m), 1.59 (1H, m), 1.50 (1H, m).

[0058] Example 6: (S)-1-((3-Chlorophenyl)sulfonyl)piperidine-3-carboxylic acid The compound of Example 6 was prepared using the general method and 1-(3-chlorophenyl)sulfonyl chloride and (S)-piperidine-3-carboxylic acid.

[0059] 1 1H NMR (400 MHz, CD3OD) δ 7.79 (1H, s), 7.68 (2H, m), 7.62 (1H, m), 3.72 (1H, d), 3.51 (1H, d), 2.60 (3H, m), 1.97 (1H, m), 1.81 (1H, m), 1.58 (1H, m), 1.51 (1H, m).

[0060] Example 7: (R)-1-((3-Fluorophenyl)sulfonyl)piperidine-3-carboxylic acid The compound of Example 7 was prepared using the general method and 1-(3-fluorophenyl)sulfonyl chloride and (R)-piperidine-3-carboxylic acid.

[0061] 1 1H NMR (400 MHz, CD3OD) δ 7.52 (2H, m), 7.48 (1H, m), 7.44 (1H, m), 3.73 (1H, d), 3.53 (1H, d), 2.60 (3H, m), 1.97 (1H, m), 1.81 (1H, m), 1.59 (1H, m), 1.50 (1H, m).

[0062] Example 8: (R)-1-((3-chlorophenyl)sulfonyl)piperidine-3-carboxylic acid The compound of Example 8 was prepared using the general method and 1-(3-chlorophenyl)sulfonyl chloride and (R)-piperidine-3-carboxylic acid.

[0063] 1 1H NMR (400 MHz, CD3OD) δ 7.79 (1H, s), 7.68 (2H, m), 7.62 (1H, m), 3.73 (1H, d), 3.53 (1H, d), 2.60 (3H, m), 1.97 (1H, m), 1.81 (1H, m), 1.59 (1H, m), 1.50 (1H, m).

[0064] Example 9: 1-((4-chlorophenyl)sulfonyl)piperidine-4-carboxylic acid The compound of Example 9 was prepared using the general method and 1-(4-chlorophenyl)sulfonyl chloride and piperidine-4-carboxylic acid.

[0065] 1 1H NMR (400 MHz, d6 DMSO) δ 12.32 (1H, s), 7.73 (4H, m), 3.47 (2H, m), 2.44 (2H, m), 2.28 (1H, m), 1.86 (2H, m), 1.57 (2H, m).

[0066] Example 10: 1-((2-(trifluoromethyl)phenyl)sulfonyl)piperidine-4-carboxylic acid The compound of Example 10 was prepared using the general method and 1-(2-(trifluoromethyl)phenyl)sulfonyl chloride and piperidine-4-carboxylic acid.

[0067] 1 H NMR(400MHz, d6 DMSO) δ 12.36(1H, s), 8.03(2H, m), 7.90(2H, m), 3.61(2H, m), 2.85(2H, m), 2.41(1H, m), 1.90(2H, m), 1.55(2H, m).

[0068] Example 11: 1-((3-(trifluoromethyl)phenyl)sulfonyl)piperidine-4-carboxylic acid The compound of Example 11 was prepared using the general method and 1-(3-(trifluoromethyl)phenyl)sulfonyl chloride and piperidine-4-carboxylic acid.

[0069] 1 H NMR(400MHz, d6 DMSO) δ 12.33(1H, s), 8.13(1H, m), 8.07(1H, m), 7.95(1H, m), 7.90(1H, m), 3.53(2H, m), 2.45(2H, m), 2.32(1H, m), 1.89(2H, m), 1.57(2H, m).

[0070] Example 12: 1-((4-(trifluoromethyl)phenyl)sulfonyl)piperidine-4-carboxylic acid The compound of Example 12 was prepared using the general method and 1-(4-(trifluoromethyl)phenyl)sulfonyl chloride and piperidine-4-carboxylic acid.

[0071] 1 H NMR(400MHz, d6 DMSO) δ 8.00(2H, m), 7.92(2H, m), 3.20(2H, m), 2.54(2H, m), 1.70(3H, m), 1.57(2H, m).

[0072] Example 13: 1-((2,5-bis(trifluoromethyl)phenyl)sulfonyl)piperidine-4-carboxylic acid The compound of Example 13 was prepared using a general method and 1-(2,5-bis(trifluoromethyl)phenyl)sulfonyl chloride and piperidine-4-carboxylic acid.

[0073] 1 H NMR (400 MHz, d6 DMSO) δ 12.36 (1H, s), 8.30 (2H, m), 8.24 (1H, s), 3.65 (2H, m), 2.87 (2H, m), 2.43 (1H, m), 1.89 (2H, m), 1.54 (2H, m).

[0074] Example 14: 1-((2-(Trifluoromethoxy)phenyl)sulfonyl)piperidine-4-carboxylic acid The compound of Example 14 was prepared using a general method and 1-(2-(trifluoromethoxy)phenyl)sulfonyl chloride and piperidine-4-carboxylic acid.

[0075] 1 H NMR (400 MHz, d6 DMSO) δ 12.33 (1H, s), 7.93 (1H, m), 7.83 (1H, m), 7.62 (2H, m), 3.56 (2H, m), 2.71 (2H, m), 2.36 (1H, m), 1.87 (2H, m), 1.52 (2H, m).

[0076] Example 15: (S)-1-((2-(Trifluoromethyl)phenyl)sulfonyl)pyrrolidine-3-carboxylic acid The compound of Example 15 was prepared using a general method and 1-(2-(trifluoromethyl)phenyl)sulfonyl chloride and (S)-pyrrolidine-3-carboxylic acid.

[0077] 1 H NMR (400 MHz, d6 DMSO) δ 12.62 (1H, br s), 8.05 (2H, m), 7.90 (2H, m), 3.48 (2H, m), 3.38 (2H, m), 3.15 (1H, m), 2.11 (2H, m).

[0078] Example 16: (R)-1-((2-(Trifluoromethyl)phenyl)sulfonyl)pyrrolidine-3-carboxylic acid The compound of Example 16 was prepared using the general method and 1-(2-(trifluoromethyl)phenyl)sulfonyl chloride and (R)-pyrrolidine-3-carboxylic acid.

[0079] 1 H NMR (400 MHz, d6 DMSO) δ 12.62 (1H, br s), 8.05 (2H, m), 7.90 (2H, m), 3.48 (2H, m), 3.38 (2H, m), 3.15 (1H, m), 2.11 (2H, m).

[0080] Example 17: (S)-1-((2-(Trifluoromethyl)phenyl)sulfonyl)piperidine-3-carboxylic acid (S)-1-((2-(Trifluoromethyl)phenyl)sulfonyl)piperidine-3-carboxylic acid was prepared using the following synthetic procedure.

Chemical formula

[0081] (S)-Piperidine-3-carboxylic acid (1 g, 7.7 mmol), potassium hydroxide (434 mg, 7.7 mmol) and potassium carbonate (2.14 g, 15.4 mmol) were added to water (20 ml) and stirred. 2-(Trifluoromethyl)benzenesulfonyl chloride (1.89 g, 7.7 mmol) was added and the reaction mixture was stirred at room temperature for 3 hours. The reaction mixture was cooled to 0 °C and acidified with 2 M hydrochloric acid to obtain a white precipitate. This precipitate was dried and triturated with n-pentane to obtain (S)-1-((2-(trifluoromethyl)phenyl)sulfonyl)piperidine-3-carboxylic acid.

[0082] Tlc Rf 0.3, 70% ethyl acetate in hexane.

[0083] m / z: 337.98 (calculated 338.03) 11H NMR (400 MHz, d6 DMSO) δ 12.33 (1H, s), 8.04 (2H, m), 7.90 (2H, m), 3.69 (1H, dd), 3.50 (1H, dd), 2.93 (1H, m), 2.81 (1H, m), 1.91 (1H, m), 1.72 (1H, m), 1.50 (2H, m).

[0084] Example 18: (R)-1-((2-(Trifluoromethyl)phenyl)sulfonyl)piperidine-3-carboxylic acid (R)-1-((2-(Trifluoromethyl)phenyl)sulfonyl)piperidine-3-carboxylic acid was prepared in the same manner as (S)-1-((2-(trifluoromethyl)phenyl)sulfonyl)piperidine-3-carboxylic acid but using (R)-piperidine-3-carboxylic acid.

[0085] Tlc Rf 0.3, 70% ethyl acetate in hexane.

[0086] m / z: 338.03 (calculated 338.03) 1H NMR (400 MHz, d6 DMSO) δ 12.53 (1H, s), 8.04 (2H, m), 7.90 (2H, m), 3.69 (1H, dd), 3.49 (1H, dd), 2.93 (1H, m), 2.81 (1H, m), 1.90 (1H, m), 1.72 (1H, m), 1.49 (2H, m).

[0087] Example 19: Activity of the compounds of Examples 1 - 18 in an A1AT cell secretion assay using HEK-Z cells Method Human embryonic kidney cell line HEK-Z cells stably transfected with the human Z A1AT gene were plated overnight at 37 °C in a humidified atmosphere containing 5% CO2 in 96-well plates (3.0×10 5cells / ml, 200 μl of medium / well). After incubation, the cells were washed three times with 200 μl of serum-free medium and replaced with serum-free medium containing vehicle, 10 μM suberoylanilide hydroxamic acid (SAHA), or the compounds of Examples 1-18 (at concentrations of 10, 33, 100, and 333 nM) in a final volume of 200 μl and incubated for 48 h in a 37 °C incubator in quadruplicate treatments. At the end of the incubation step, the supernatant was removed from the wells and centrifuged at 1000×g for 10 min at 4 °C, and the human A1AT level was assayed by ELISA (Human Serpin A1 / α1-antitrypsin duo set ELISA, R&D Systems, DY1268) as per the manufacturer's instructions.

[0088] Briefly, a 96-well plate was coated overnight at room temperature with human A1AT capture antibody (1:180 dilution from stock, final volume of 100 μl / well). The capture antibody was then removed, and the plate was washed three times well with 300 μl of wash buffer (0.05% Tween® 20 in PBS), and then 200 μl of reagent diluent (25% Tween® 20 in PBS) was incubated in each well at room temperature for 1 h. Diluted samples, standards (A1AT at 125, 250, 500, 1000, 2000, 4000, and 8000 pg / ml), or blanks were then added in duplicate to each well, the plate was covered with a plate sealer, and allowed to stand at room temperature for 2 h. At the end of the sample incubation step, the samples were removed, all wells were washed as before, 100 μl of detection antibody (1:180 dilution from stock) was added to each well, and incubated for an additional 2 h at room temperature. After incubation with the detection antibody, the supernatant was removed, the wells were washed as before, and 100 μl of streptavidin-HRP solution (1:200 dilution from stock) was added to each well in the dark for 20 min. Thereafter, 50 μl of stop solution (2M H2SO4) was added, and the optical density (OD) of each well was read at 450 nm with blank subtraction at 570 nm from each well using a microplate reader. A four-parameter logistic curve was constructed using GraphPad Prism 7, and the A1AT concentration in each sample was determined by interpolation from the standard curve and multiplication by the appropriate dilution factor.

[0089] Results The amount of human A1AT secreted into the medium from transfected HEK-EBNA cells was measured by ELISA. 10 μM SAHA was used as a positive control for all in vitro A1AT secretion experiments.

[0090] The data in Table 1 show that the compounds of Examples 1-18 increase the secretion of human Z A1AT from HEK-Z cells in a dose-dependent manner when measured by ELISA. [Table 1]

[0091] Example 20: Activity of the compounds of Examples 1 - 18 in an A1AT cell secretion assay using HEK-M cells Method The human embryonic kidney cell line HEK-M cells stably transfected with M A1AT were plated in 96-well plates (3.0×10 5 cells / ml, 200 μl of medium / well) in a humidified atmosphere containing 5% CO2 at 37 °C overnight. After incubation, the cells were washed three times with 200 μl of serum-free medium and then incubated in a 37 °C incubator for 48 h. The medium was replaced with serum-free medium containing vehicle, 10 μM suberoylanilide hydroxamic acid (SAHA), or the compounds of Examples 1 - 16 in six replicates at a final volume of 200 μl. At the end of the incubation step, the supernatant was removed from the wells and centrifuged at 1000×g for 10 min at 4 °C. Human A1AT levels were assayed by ELISA (Human Serpin A1 / α1-antitrypsin duo set ELISA, R & D Systems, DY1268) as per the manufacturer's instructions.

[0092] Briefly, 96-well plates were coated overnight at room temperature with human A1AT capture antibody (1:180 dilution from stock, final volume of 100 μl / well). The capture antibody was then removed, and the wells were washed three times thoroughly with 300 μl of wash buffer (0.05% Tween® 20 in PBS), and then 200 μl of reagent diluent (25% Tween® 20 in PBS) was incubated in each well at room temperature for 1 h. Diluted samples, standards (A1AT at 125, 250, 500, 1000, 2000, 4000, and 8000 pg / ml), or blanks were then added in duplicate to each well, the plate was covered with a plate sealer, and left to stand at room temperature for 2 h. At the end of the sample incubation step, the samples were removed, all wells were washed as before, 100 μl of detection antibody (1:180 dilution from stock) was added to each well, and incubated at room temperature for a further 2 h. After incubation with the detection antibody, the supernatant was removed, the wells were washed as before, and 100 μl of streptavidin-HRP solution (1:200 dilution from stock) was added to each well in the dark for 20 min. Subsequently, 50 μl of stop solution (2 M H2SO4) was added, and the optical density (OD) of each well was read at 450 nm with blank subtraction at 570 nm from each well using a microplate reader. A four-parameter logistic curve was constructed using GraphPad Prism 7, and the A1AT concentration in each sample was determined by interpolation from the standard curve and multiplication by the appropriate dilution factor.

[0093] Results The amount of human M A1AT secreted into the medium from transfected HEK-EBNA cells was measured by ELISA. 10 μM SAHA was used as a positive control for all in vitro A1AT secretion experiments. The compounds of Examples 1, 3, 4, 10, 17, and 18 did not lead to an increase in the secretion of human M A1AT from HEK-M cells at 10 μM.

[0094] Example 21: Activity of the compounds of Examples 1 and 17 in an A1AT cell secretion assay using HEK-Siiyama cells The rare Siiyama mutation (Ser 53 to Phe, numbering of mature A1AT) was identified in Japanese men with AATD (Seyama et al J Biol Chem (1991) 266:12627-32). Ser53 is a conserved serpin residue and is thought to be important for the organization of the internal core of the A1AT molecule. The change from an uncharged polar amino acid to a large nonpolar amino acid in the conserved backbone of the protein affects the folding and intracellular processing of Siiyama A1AT.

[0095] Method Human embryonic kidney cell line HEK-Siiyama cells stably transfected with the human Siiyama A1AT gene were plated overnight at 37 °C in a humidified atmosphere containing 5% CO2 in 96-well plates (3.0×10 5 cells / ml, 200 μl of medium / well). After incubation, the cells were washed three times with 200 μl of serum-free medium and replaced with serum-free medium containing vehicle, 10 μM suberoylanilide hydroxamic acid (SAHA) or the compound of Example 1 (1 and 10 μM) in a final volume of 200 μl in eight replicates in an incubator at 37 °C for 48 h. At the end of the incubation step, the supernatant was removed from the wells and centrifuged at 1000×g for 10 min at 4 °C, and the human A1AT level was assayed by ELISA (Human Serpin A1 / α1-antitrypsin duo set ELISA, R & D Systems, DY1268) as per the manufacturer's instructions.

[0096] Briefly, a 96-well plate was coated overnight at room temperature with human A1AT capture antibody (1:180 dilution from stock, 100 μl final volume / well). The capture antibody was then removed, and the plate was washed three times well with 300 μl of wash buffer (0.05% Tween® 20 in PBS), and then 200 μl of reagent diluent (25% Tween® 20 in PBS) was incubated in each well at room temperature for 1 h. Diluted samples, standards (A1AT at 125, 250, 500, 1000, 2000, 4000, and 8000 pg / ml), or blanks were then added in duplicate to each well, the plate was covered with a plate sealer, and left standing at room temperature for 2 h. At the end of the sample incubation step, the samples were removed, all wells were washed as before, 100 μl of detection antibody (1:180 dilution from stock) was added to each well, and incubated at room temperature for a further 2 h. After incubation with the detection antibody, the supernatant was removed, the wells were washed as before, and 100 μl of streptavidin-HRP solution (1:200 dilution from stock) was added to each well in the dark for 20 min. Subsequently, 50 μl of stop solution (2 M H2SO4) was added, and the optical density (OD) of each well was read at 450 nm with blank subtraction at 570 nm from each well using a microplate reader. A four-parameter logistic curve was constructed using GraphPad Prism 7, and the A1AT concentration in each sample was determined by interpolation from the standard curve and multiplication by the appropriate dilution factor.

[0097] Results The amount of human Siiyama A1AT secreted into the medium from transfected HEK-EBNA cells was measured by ELISA. 10 μM SAHA was used as a positive control for all in vitro A1AT human secretion experiments. The exemplary compounds of Examples 1 and 17 did not stimulate the secretion of Siiyama A1AT from HEK-Siiyama cells at 1 or 10 μM as measured by ELISA. In contrast, the positive control, 10 μM SAHA, stimulated an increase in Siiyama A1AT secretion.

[0098] Example 22: Activity of the Compounds of Examples 1 and 17 in Mice Expressing Human Z (huZ Mice) HuZ mice (also referred to as PiZZ mice) are transgenic mouse lines containing multiple copies of the Z variant of the human A1AT gene developed by two separate groups (Dycaico et al Science (1988) 242:1409-12) and Carlson et al J. Clin Invest (1989) 83:1183-90). HuZ mice are on a C57Bl / 6 background and express human Z A1AT protein in liver tissue. The mice used in this study were from the Carlson and colleagues' line (transgenic line Z11.03). HuZ mice were used as a tool to evaluate the effect of the exemplary compounds of the present invention on the increase in the circulating levels of Z A1AT in plasma or the effect of the compounds on the accumulation of Z A1AT polymers in the liver and the associated liver pathology.

[0099] HuZ mice (n = 4 / group; male or female) with a basal human Z A1AT plasma level of 200 - 600 μg / ml were treated twice daily for 14 consecutive days by oral gavage with either vehicle or one of the compounds of Example 1 or 17 at 5, 15 or 50 mg / kg. Mice had free access to food (standard mouse chow, SAFE diet) and water. On day 14 of the study, dosing of each mouse was performed 1 hour prior to the sacrifice procedure. Blood was collected from the tail vein of each mouse on days -12, -7 and -5 prior to dosing, and on days 12, 13 and 14 of dosing. Blood was collected into microvette containing EDTA and plasma was prepared by centrifugation at 2700×g for 10 minutes at 4°C. Plasma was aliquoted and stored at -80°C for bioanalysis. Plasma samples from days -12, -7 and -5 prior to dosing were used to determine the mean basal level of human Z A1AT for each mouse. Plasma samples collected on the last three dosing days (days 12, 13 and 14) of the study were used to measure human Z A1AT levels and to determine the effect of the compounds of Example 1 or 17 on human Z A1AT secretion by comparing with the basal level for each mouse. Human Z A1AT levels in mouse plasma samples were measured by ELISA (Human Serpin A1 / α1antitrypsin duo set ELISA, R & D Systems, DY1268) as per the manufacturer's instructions.

[0100] Briefly, a 96-well plate was coated overnight at room temperature with a human A1AT capture antibody (1:180 dilution from stock, 100 μl final volume / well). The capture antibody was then removed, and the plate was washed three times well with 300 μl of wash buffer (0.05% Tween® 20 in PBS), and then 200 μl of reagent diluent (25% Tween® 20 in PBS) was incubated in each well at room temperature for 1 h. Diluted samples, standards (A1AT at 125, 250, 500, 1000, 2000, 4000, and 8000 pg / ml), or blanks were then added in duplicate to each well, the plate was covered with a plate sealer, and left standing at room temperature for 2 h. At the end of the sample incubation step, the samples were removed, all wells were washed as before, 100 μl of detection antibody (1:180 dilution from stock) was added to each well, and incubated at room temperature for a further 2 h. After incubation with the detection antibody, the supernatant was removed, the wells were washed as before, 100 μl of streptavidin-HRP solution (1:200 dilution from stock) was added to each well in the dark for 20 min. Subsequently, 50 μl of stop solution (2M H2SO4) was added, and the optical density (OD) of each well was read at 450 nm with blank subtraction at 570 nm from each well using a microplate reader. A four-parameter logistic curve was constructed using GraphPad Prism 7, and the A1AT concentration in each sample was determined by interpolation from the standard curve and multiplication by the appropriate dilution factor.

[0101] Results The effect of the compound of Example 1 or 17 on the circulating levels of human Z A1AT was evaluated in the huZ mouse model.

[0102] The compounds of Example 1 and 17 stimulated the secretion of human Z A1AT in huZ mice compared to the baseline level. Figure 1 shows the data at each treatment dose for the compound of Example 17. In an embodiment of the present invention, for example, the following items are provided. (Item 1) Formula (1):

Chem.

Claims

Use of a compound selected from: - 1-(quinolin-8-ylsulfonyl)piperidine-4-carboxylic acid or - 1-((2-chlorophenyl)sulfonyl)piperidine-4-carboxylic acid or - 1-((3-chlorophenyl)sulfonyl)piperidine-4-carboxylic acid or - 1-((2,3-dichlorophenyl)sulfonyl)piperidine-4-carboxylic acid or - (S)-1-((3-fluorophenyl)sulfonyl)piperidine-3-carboxylic acid or or - (S)-1-((3-chlorophenyl)sulfonyl)piperidine-3-carboxylic acid or - (R)-1-((3-fluorophenyl)sulfonyl)piperidine-3-carboxylic acid or or - (R)-1-((3-chlorophenyl)sulfonyl)piperidine-3-carboxylic acid or - 1-((4-chlorophenyl)sulfonyl)piperidine-4-carboxylic acid or - 1-((2-(trifluoromethyl)phenyl)sulfonyl)piperidine-4-carboxylic acid or - 1-((3-(trifluoromethyl)phenyl)sulfonyl)piperidine-4-carboxylic acid or - 1-((4-(trifluoromethyl)phenyl)sulfonyl)piperidine-4-carboxylic acid or - 1-((2,5-bis(trifluoromethyl)phenyl)sulfonyl)piperidine-4-carboxylic acid or - 1-((2-(trifluoromethoxy)phenyl)sulfonyl)piperidine-4-carboxylic acid or - (S)-1-((2-(trifluoromethyl)phenyl)sulfonyl)pyrrolidine-3-carboxylic acid or - (R)-1-((2-(trifluoromethyl)phenyl)sulfonyl)pyrrolidine-3-carboxylic acid or - (S)-1-((2-(trifluoromethyl)phenyl)sulfonyl)piperidine-3-carboxylic acid or - (R)-1-((2-(trifluoromethyl)phenyl)sulfonyl)piperidine-3-carboxylic acid in the manufacture of a medicament for the treatment of α1-antitrypsin deficiency (AATD). Use of a mixture of the two enantiomers of any of the compounds as defined in claim 1 having a chiral center, said mixture being a racemate or having one enantiomer in excess over the other enantiomer, in the manufacture of a medicament for the treatment of AATD. Use according to claim 1 or 2, wherein each enantiomer of the compound or the mixture is in a pharmaceutically acceptable salt form. Use of a racemate of a compound as defined in claim 1 in the manufacture of a medicament for the treatment of AATD.

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