Thiadiazolone Derivatives Useful as AMPK Activators

Abstract

The present invention relates to a compound of formula I, or a pharma- ceutically acceptable salt or solvate thereof, 1 , R 2 , R 3 and R 4 are as defined herein, and these compounds are useful, particularly as prodrugs, in the treatment of disorders or conditions ameliorated by activation of AMPK.

Description

Detailed Description of the Invention

[0001] [Field of the Invention] The present invention relates to novel compounds and their use in medicine. In particular, the present invention relates to compounds useful in the treatment of disorders or conditions ameliorated by activation of AMP-activated protein kinase.

[0002] [Background of the invention] AMP-activated protein kinase (AMPK) is a protein kinase enzyme consisting of three protein subunits that is activated by hormones, cytokines, exercise, and stresses that decrease cellular energy status (e.g., glucose deprivation). Activation of AMPK increases processes that generate adenosine 5'-triphosphate (ATP) (e.g., fatty acid oxidation) and suppresses others that consume ATP but are not rapidly required for survival, such as fatty acid, glycerolipid, and protein synthesis. Conversely, when cells are presented with a sustained excess of glucose, AMPK activity is decreased and fatty acid, glycerolipid, and protein synthesis are enhanced. Thus, AMPK is a protein kinase enzyme that plays a key role in cellular energy homeostasis. Thus, activation of AMPK is coupled with glucose-lowering effects and induces several other biological effects, including cholesterol synthesis, lipogenesis, inhibition of triglyceride synthesis, and reduction of hyperinsulinemia.

[0003] Considering the above, AMPK is the preferred target for the treatment of metabolic syndrome, especially type 2 diabetes. AMPK also participates in many pathways that are important for many different diseases (for example, AMPK also participates in many pathways that are important in CNS disorders, fibrosis, osteoporosis, heart failure, and sexual dysfunction).

[0004] AMPK is also involved in many pathways important in cancer. Several tumor suppressors are part of the AMPK pathway. AMPK acts as a negative regulator of the mammalian TOR (mTOR) and EF2 pathways, which are important regulators of cell growth and proliferation. Thus, deregulation may be associated with diseases such as cancer (and diabetes). Thus, AMPK activators may be useful as anticancer agents.

[0005] AMPK activators (e.g., metformin and 4-chloro-N-[2-[(4-chlorophenyl)methyl]-3-oxo-1,2,4-thiadiazol-5-yl]benzamide (i.e., the compound of formula II below)) have been shown to be effective in treating pain. Das and coworkers report that post-injury treatment in mice with an AMPK activator reduces mechanical hypersensitivity after lumbar disc puncture (Das V, et al. Reg Anesth Pain Med 2019;0:1-5. doi:10.1136 / rapm-2019-100839). Similarly, Das and coworkers have reported that early treatment with AMPK activators reduces mechanical hypersensitivity in a postoperative pain model in mice (Das V, et al. Reg Anesth Pain Med 2019;0:1-6. doi:10.1136 / rapm-2019-100651). These drugs also normalize the AMPK pathway in the dorsal root ganglion. Thus, AMPK activators can be used to treat pain, especially postoperative pain.

[0006] It has also been shown that hepatic steatosis can be regulated by AMPK (Zhao et al. J. Biol. Chem. 2020 295: 12279-12289). Activation of AMPK promotes fatty acid oxidation (β-oxidation) in the liver while inhibiting de novo lipogenesis. AMPK activation also reduces free fatty acid release from adipose tissue and prevents hepatic steatosis. Pharmacological activation of AMPK in the liver was reported to promote beneficial effects on multiple aspects of nonalcoholic fatty liver disease (NAFLD). For example, activation of AMPK was found to ameliorate nonalcoholic steatohepatitis (NASH) in both mouse and monkey animal models. Thus, AMPK activators may be useful for the treatment of NAFLD and NASH.

[0007] An example of an AMPK activator is 4-chloro-N-[2-[(4-chlorophenyl)methyl]-3-oxo-1,2,4-thiadiazol-5-yl]benzamide (i.e., the compound of formula II), which was first disclosed in WO2011 / 004162. [ka]

[0008] As AMPK agonists (i.e., AMPK activators), the compounds of formula II are useful in treating disorders or conditions that are ameliorated by activation of AMPK. The compounds may be useful in treating cardiovascular disease (such as heart failure), diabetic kidney disease, type 2 diabetes, insulin resistance, nonalcoholic fatty liver disease, nonalcoholic steatohepatitis, pain, opioid addiction, obesity, cancer, inflammation (including chronic inflammatory disease), autoimmune disease, osteoporosis, and intestinal disease.

[0009] Although many AMPK activators are known, there remains a need for the development of new compounds for the treatment of disorders or conditions that are improved by the activation of AMPK.The present inventors have now found new compounds that are metabolized in vivo to form known AMPK activators, and surprisingly improve the bioavailability of the AMPK activators.The compounds have also been found to activate AMPK.

[0010] The listing or discussion of an apparently prior-published document in this specification should not necessarily be taken as an acknowledgement that the document is part of the prior art or is common general knowledge.

[0011] [Detailed Description of the Invention] In a first aspect of the present invention, there is provided a compound of formula I, or a pharma- ceutically acceptable salt or solvate thereof: [ka] R 1 and R 2 are independently selected from the group consisting of hydrogen, the side chains of proteinogenic amino acids, and C optionally substituted by one or more substituents selected from the group consisting of -OH, -NH2, and -C(O)NH2. 1-6 selected from the group consisting of alkyl; R 3 and R 4 are independently hydrogen, C optionally substituted with a phenyl group 1-4 Alkyl, and A 1 or R 1 (or R 2 ) and R 3 (or R 4 ), together with the carbon and nitrogen atoms to which they are attached, form a 4- to 6-membered heterocycloalkyl group; A 1 teeth, [ka] or [ka] represents A 2 is hydrogen, C optionally substituted with phenyl groups 1-4 Alkyl, [ka] or [ka] represents Each A 3 are independently hydrogen or C optionally substituted with a phenyl group. 1-4 represents alkyl; and R 5a and R 5b is independently a C optionally substituted by one or more substituents selected from the group consisting of hydrogen, the side chain of a proteinogenic amino acid, and -OH, -NH2, and -C(O)NH2. 1-6 alkyl.

[0012] These compounds, including their pharma- ceutically acceptable salts and solvates, may be referred to herein as "compounds of the invention."

[0013] In a preferred embodiment of the first aspect of the present invention, A 2 is hydrogen, [ka] or [ka] represents Each A 3 represents hydrogen.

[0014] The compounds of the present invention have been found to be metabolized in vivo to form 4-chloro-N-[2-[(4-chlorophenyl)methyl]-3-oxo-1,2,4-thiadiazol-5-yl]benzamide (referred to herein as the compound of formula II), which is known to be an AMPK activator. In this regard, the compounds of the present invention may be considered to be prodrugs of 4-chloro-N-[2-[(4-chlorophenyl)methyl]-3-oxo-1,2,4-thiadiazol-5-yl]benzamide.

[0015] Pharmaceutically acceptable salts of potential utility include those discussed in Berge et al., J. Pharmaceutical Sciences, 66: 1-19 (1977). Pharmaceutically acceptable salts of the compounds of formula I may be prepared according to techniques well known to those skilled in the art.

[0016] Examples of pharma- ceutically acceptable addition salts include those derived from organic acids, such as citric acid, tartaric acid, acetic acid (including halogenated forms of acetic acid, such as trifluoroacetic acid), malic acid, lactic acid, fumaric acid, benzoic acid, glycolic acid, gluconic acid, succinic acid, and arylsulfonic acids; and those derived from mineral acids, such as hydrochloric acid, hydrobromic acid, phosphoric acid, metaphosphoric acid, nitric acid, and sulfuric acid. In certain embodiments, the pharma-ceutically acceptable salt is a citrate or trifluoroacetate salt of the compound of formula I.

[0017] Unless otherwise specified, alkyl groups as defined herein may be straight-chained or, when there is a sufficient number (i.e., at least two or three, as appropriate) of carbon atoms, branched-chained and / or cyclic (thus forming a cycloalkyl group). When there is a sufficient number (i.e., at least four) of carbon atoms, the group may also be partially cyclic (thus forming a partial cycloalkyl group). For example, cycloalkyl groups that may be listed include cyclopropyl, cyclobutyl, cyclopentyl, and cyclohexyl. Similarly, partial cyclic alkyl groups (which may also be referred to as "partial cycloalkyl" groups) that may be listed include cyclopropylmethyl.

[0018] Heterocycloalkyl groups that may be mentioned include non-aromatic monocyclic heterocycloalkyl groups in which at least one (e.g., 1 to 4) of the atoms in the ring system is an atom other than carbon (i.e., a heteroatom such as sulfur, oxygen or especially nitrogen) and the total number of atoms in the ring system is 4 to 6. The carbon atoms of the heterocycloalkyl groups referred to herein may be substituted by one or more =O substituents.

[0019] For the avoidance of doubt, even if the identities of two or more substituents in a compound of formula I are the same, the actual identities of the respective substituents are not in any way interdependent.

[0020] Where a group is referred to herein as being optionally substituted, it is specifically intended that the optional substituent may be absent (i.e., the reference to the optional substituent may be omitted), in which case the optionally substituted group may, in certain embodiments, be referred to as being unsubstituted.

[0021] Pharmaceutically acceptable salts of compounds of formula I may be prepared according to techniques well known to those skilled in the art. For example, compounds of formula I may be reacted with a suitable organic or mineral acid. Salt switching techniques may be used to convert one salt into another.

[0022] The compounds disclosed herein may exist in unsolvated as well as solvated forms with pharma- ceutically acceptable solvents such as water and ethanol, and the present invention is intended to encompass both solvated and unsolvated forms of the compounds of the present invention.

[0023] The term "solvate" refers to a complex formed by a solute and a solvent with variable stoichiometry. For the purposes of the present invention, the solvent may not interfere with the biological activity of the solute. Examples of suitable solvents include, but are not limited to, water, methanol, ethanol, and acetic acid. Solvates in which water is the solvent molecule are typically referred to as hydrates. Hydrates include compositions that contain stoichiometric amounts of water and compositions that contain variable amounts of water.

[0024] The compounds of formula I contain double bonds and therefore may exist as E (entgegen) and Z (zusammen) geometric isomers about each individual double bond, and all such isomers and mixtures thereof are included within the scope of the present invention.

[0025] Compounds of formula I may exist as positional isomers and may exhibit tautomerism, all tautomers and mixtures thereof are included within the scope of the invention.

[0026] The present invention also encompasses isotopically labeled compounds of formula I, which are identical to those described herein, but which have one or more atoms replaced with atoms having atomic masses or mass numbers different from the atomic masses or mass numbers normally found in nature (or the atomic masses or mass numbers most abundantly found in nature).All isotopes of any particular atom or element as defined herein are contemplated to be within the scope of the present invention.Therefore, compounds of formula I also include deuterated compounds, i.e., compounds of formula I in which one or more hydrogen atoms are replaced with hydrogen isotopes deuterium.

[0027] Those of skill in the art will appreciate that the compounds of the invention that are the subject of the present invention include stable compounds, i.e., compounds of the invention include compounds that are sufficiently robust to be isolated to a useful degree of purity, for example, from a reaction mixture.

[0028] Throughout this specification, structures may or may not be shown by chemical names. In case of doubt about nomenclature, the structures shall prevail. If a compound can exist as tautomers (e.g., alternative resonance forms), the structure shown represents one of the possible tautomers, and the tautomer actually observed may vary depending on environmental factors such as solvent, temperature, or pH. All tautomeric (and resonance) forms and mixtures thereof are within the scope of the present invention.

[0029] Unless otherwise defined, all technical and scientific terms used herein have the common meaning as understood by one of ordinary skill in the art to which this invention belongs.

[0030] For the avoidance of doubt, those skilled in the art will understand that reference herein to a particular aspect of the invention (such as the first aspect of the invention) includes reference to all embodiments and specific features thereof, and that these embodiments and specific features may be used in combination to form further embodiments and features of the invention.

[0031] Amino acids are organic compounds containing an amino functional group (-NH2) and a carboxyl functional group (-CO2H) and (most often) have a side chain. α-amino acids are those in which the amino and carboxyl functional groups are attached to the same carbon atom (i.e., the α-carbon atom). Among the α-amino acids that may be mentioned are the proteinogenic amino acids. Proteinogenic amino acids are those amino acids that are biosynthetically incorporated into proteins during translation. The proteinogenic amino acids are glycine (Gly), alanine (Ala), arginine (Arg), asparagine (Asn), aspartic acid (Asp), cysteine ​​(Cys), glutamic acid (Glu), glutamine (Gln), histidine (His), isoleucine (Ile), leucine (Leu), lysine (Lys), methionine (Met), phenylalanine (Phe), proline (Pro), pyrrolysine (Pyl), selenocysteine ​​(Sec), serine (Ser), threonine (Thr), tryptophan (Trp), tyrosine (Tyr), and valine (Val).

[0032] Thus, the term "side chain of a proteinogenic amino acid" refers to a group other than hydrogen attached to the α-carbon atom of a proteinogenic amino acid. In the context of the present invention, the side chain of one or more proteinogenic amino acids is R 1 , R 2 , R 5a or R 5b The amino acid residues may be located at any of the positions. For the avoidance of doubt, the compounds of the invention may contain multiple different side chains at these positions, i.e., the prodrugs of compound II may contain chains with two or three different amino acids linked to the thiadiazole ring. Particular examples of proteinogenic amino acid side chains include methyl substituted with imidazolyl or indolyl groups (for histidine and tryptophan), propyl substituted with guanidino groups (for arginine) and butyl substituted with amino groups (for lysine).

[0033] Particular compounds of the invention that may be enumerated are 1 and R2 For example, R 1 and R 2 may both be hydrogen.

[0034] Other compounds of the invention that may be mentioned are R 1 and R 2 For example, R 1 is the side chain of a proteinogenic amino acid, and R 2 may be hydrogen.

[0035] Thus, the compounds of the first aspect of the invention may contain one or more asymmetric carbon atoms and may therefore exhibit optical isomerism and / or diastereoisomerism. 1 and R 2 are different (e.g., R 1 is methyl and R 2 is hydrogen), compounds of formula I can be prepared by those skilled in the art in which R 1 and R 2 The compound may be one in which the carbon substituted with R is in the L-configuration or the D-configuration. 5a and / or R 5b groups, those R 5a and R 5b Any chiral centers to which the groups are attached may also independently be of either the L- or D-configuration.

[0036] In certain embodiments, R 1 and R 2 If different, R 1 and R 2 The carbon substituted with is in the L-configuration. 1 and R 2 If different, R 1 and R 2 The carbon substituted with may be in the D-configuration. For example, a compound of formula I may be a compound of formula IA or a compound of formula IB. [ka] R 1 , R 2 , R 3 and R 4 is as described herein (i.e. as described in the first aspect of the invention, including all embodiments and specific features and combinations thereof). Particular examples of compounds of the invention include 1 and R 2 are both hydrogen or R 1 and R 2 In the compound, the carbon substituted with is in the L-configuration, e.g., R 1 and R 2 When one of these is hydrogen, the other is a side chain of a proteinogenic amino acid.

[0037] Similarly, R 3 or R 4 A 1 If you want to represent 1 teeth [ka] and A 2 is hydrogen, C optionally substituted with phenyl groups 1-4 Alkyl, [ka] may represent A 3 , R 5a and R 5b A is as described herein (i.e. as described in the first aspect of the invention, including all embodiments and specific features and combinations thereof). Particular examples of compounds of the invention include A 1 is a compound that represents a proteinogenic amino acid or two linked proteinogenic amino acids.

[0038] Preferred examples of the compounds of the invention that may be mentioned include 2 is hydrogen, [ka] or [ka] Each A 3 represents hydrogen; and R 5b is as described herein.

[0039] Preferably, R 1 is optionally substituted by one or more substituents selected from the group consisting of hydrogen, the side chain of a proteinogenic amino acid, and -OH, -NH2, and -C(O)NH2. 1-6 alkyl, or R 1 and R 3 are linked to form a pyrrolidine ring (as found in proline); and R 2 is hydrogen.

[0040] In certain embodiments, R 2 is hydrogen, and R 1 is hydrogen or the side chain of a proteinogenic amino acid selected from the group consisting of Glu, particularly Arg, His, Lys, Ser, Thr, Asn, Gln, Cys, Sec, Ala, Ile, Leu, Met, Phe, Trp, Tyr, Asp and Val; or R 2 is hydrogen, and R 1 and R 3 form a pyrrolidine ring together with the carbon and nitrogen atoms to which they are attached.

[0041] In certain embodiments, R 2 is hydrogen, and R 1 is hydrogen or the side chain of an amino acid selected from the group consisting of Arg, His, Lys, and Trp (e.g., Arg, His, and Lys); or R 2 is hydrogen, and R 1 and R 3form a pyrrolidine ring together with the carbon and nitrogen atoms to which they are attached.

[0042] In a further embodiment, R 2 is hydrogen, and R 1 is hydrogen or the side chain of an amino acid selected from the group consisting of Lys, Ala, Ile, Phe, Leu, Ser and Asp; or R 2 is hydrogen, and R 1 and R 3 form a pyrrolidine ring together with the carbon and nitrogen atoms to which they are attached.

[0043] Preferably, R 2 is hydrogen.

[0044] Certain embodiments of the present invention include compounds of formula I, wherein R 1 is hydrogen or the side chain of an amino acid selected from the group consisting of Arg, His, Lys, and Trp (e.g., Arg, His, and Lys); or R 1 and R 3 form a pyrrolidine ring together with the carbon and nitrogen atoms to which they are attached; and R 2 is hydrogen.

[0045] In other embodiments, R 3 and R 4 are independently hydrogen, C optionally substituted with one or more phenyl groups (e.g., methyl or benzyl groups). 1-3 Alkyl, [ka] and [ka] and R 5a is hydrogen or the side chain of a proteinogenic amino acid. In a preferred embodiment, R 3 and R4 are independently hydrogen and C optionally substituted with one or more phenyl groups. 1-3 alkyl (e.g., R 3 and R 4 are independently hydrogen, methyl or benzyl. 3 and R 4 are independently selected from the group consisting of hydrogen and methyl (e.g., R 3 and R 4 are both methyl).

[0046] In another preferred embodiment, R 3 and R 4 are independently hydrogen, C optionally substituted with one or more phenyl groups (e.g., methyl or benzyl groups). 1-3 Alkyl, [ka] and [ka] or R 1 and R 3 form a pyrrolidine ring together with the carbon and nitrogen atoms to which they are attached, and R 4 is hydrogen, C optionally substituted with one or more phenyl groups 1-3 Alkyl, [ka] and [ka] and R 5a is the side chain of a proteinogenic amino acid (e.g., Phe, Val, Lys, Asp, or Arg).

[0047] Preferred compounds of the invention that may be mentioned include those having R 1 is hydrogen or a side chain of a proteinogenic amino acid selected from the group consisting of Glu, in particular Arg, His, Lys, Ser, Thr, Asn, Gln, Cys, Sec, Ala, Ile, Leu, Met, Phe, Trp, Tyr, Asp and Val (e.g., Lys, Ala, Ile, Phe, Leu, Ser and Asp; or Arg, His, Lys and Trp); R 2 represents hydrogen; R 3 and R 4 are independently selected from hydrogen and C optionally substituted with one or more phenyl groups (e.g., methyl). 1-3 alkyl; or R 1 and R 3 together with the carbon and nitrogen atoms to which they are attached form a pyrrolidine ring.

[0048] Other preferred compounds of the invention that may be mentioned include those having R 2 represents hydrogen; R 1 is hydrogen or a side chain of a proteinogenic amino acid selected from the group consisting of Glu, Arg, His, Lys, Ser, Thr, Asn, Gln, Cys, Sec, Ala, Ile, Leu, Met, Phe, Trp, Tyr, Asp, and Val (e.g., Lys, Ala, Ile, Phe, Leu, Ser, and Asp); R 3 and R 4 are independently hydrogen, C optionally substituted with one or more phenyl groups (e.g., methyl or benzyl groups), 1-3 Alkyl, [ka] and [ka] or R 1 and R 3form a pyrrolidine ring together with the carbon and nitrogen atoms to which they are attached, and R 4 is optionally substituted by hydrogen, one or more phenyl groups (e.g., methyl or benzyl groups); 1-3 Alkyl, [ka] and [ka] R 5a is hydrogen or a side chain of a proteinogenic amino acid selected from the group consisting of Glu, Arg, His, Lys, Ser, Thr, Asn, Gln, Cys, Sec, Ala, Ile, Leu, Met, Phe, Trp, Tyr, Asp, and Val (e.g., Phe, Val, Lys, Asp, and Arg); and A 2 and A 3 are independently as defined herein.

[0049] In an embodiment of the present invention, A 2 teeth [ka] R 5b is preferably hydrogen or the side chain of a proteinogenic amino acid selected from the group consisting of Glu, Arg, His, Lys, Ser, Thr, Asn, Gln, Cys, Sec, Ala, Ile, Leu, Met, Phe, Trp, Tyr, Asp and Val (e.g., Phe, Val, Lys, Asp and Arg); and 3 are independently hydrogen and C 1-3 It is selected from the group consisting of alkyl (eg, methyl).

[0050] As mentioned above, it has been found that the compounds of the present invention are metabolized in vivo to form 4-chloro-N-[2-[(4-chlorophenyl)methyl]-3-oxo-1,2,4-thiadiazol-5-yl]benzamide (referred to herein as compound of formula II). Nevertheless, the exact mechanism by which the compounds of the present invention are converted to compounds of formula II may vary depending on the compound of the present invention. Without wishing to be bound by theory, it is believed that the compounds of the present invention are degraded to compounds of formula II in two steps. First, the ester is hydrolyzed (chemically or enzymatically) to form an N-hydroxymethyl intermediate (referred to elsewhere in the present specification as compound of formula III), which is then converted to a compound of formula II.

[0051] The applicant has found that some compounds are hydrolyzed more rapidly under acidic conditions (e.g., conditions expected to be present in the human stomach), whereas other compounds are hydrolyzed more rapidly under more alkaline conditions (e.g., conditions similar to those in the small intestine).

[0052] Compounds of the present invention that are hydrolyzed more quickly under alkaline conditions (e.g., under conditions similar to those in the small intestine) than under acidic conditions are advantageous in that they may provide additional clinical benefits, and may be useful, for example, for patients who cannot achieve sufficient conversion in the stomach (e.g., cannot achieve conversion at the required rate or to the required extent). Such patients may suffer, for example, from esterase enzyme dysfunction. Furthermore, the intermediate formed as a result of hydrolysis (i.e., the compound of formula III) may be more easily taken up in the intestine compared to the compound of formula II, and therefore the formation of the compound of formula III in the intestine may be advantageous to achieve optimal systemic exposure for patients.

[0053] R 1 / R 2 Compounds of the invention containing a lipophilic (alkyl) group in R have been found to have relatively high stability at pH 7.4. 1 / R 2Groups that are H or -NH2-containing amino acid side chains (e.g., glycine- or lysine-based) are less stable at this pH. 1 / R 2 When one of the groups is an electron-withdrawing amino acid side chain (eg, an aspartic acid-, arginine- or glutamic acid-based side chain), the stability at this pH is much lower.

[0054] Particular compounds that may be mentioned in this regard are those of formula A, or a pharma- ceutically acceptable salt or solvate thereof: [ka] (i)R 1 represents hydrogen, R 2 represents the side chain of aspartic acid, the side chain of arginine, or the side chain of glutamic acid, and R 3 is hydrogen or methyl, R 4 is hydrogen or methyl; or (ii)R 1 , R 2 and R 3 represents hydrogen, R 4 A 1 A stands for 1 teeth [ka] A stands for 2 is hydrogen or methyl, and A 3 is hydrogen or methyl, R 5a represents the side chain of aspartic acid, the side chain of arginine, or the side chain of glutamic acid.

[0055] Further compounds that may be mentioned in this regard are compounds of formula B or a pharma- ceutically acceptable salt or solvate thereof: [ka] (i)R 1 represents hydrogen, R 2represents the side chain of aspartic acid or the side chain of arginine, and R 3 is hydrogen or methyl, R 4 is hydrogen or methyl; or (ii)R 1 , R 2 and R 3 represents hydrogen, R 4 A 1 A stands for 1 teeth [ka] A stands for 2 is hydrogen or methyl, and A 3 is hydrogen or methyl, R 5a represents the side chain of aspartic acid or the side chain of arginine.

[0056] Further particular compounds that may be listed in this regard are compounds of formula C, or a pharma- ceutically acceptable salt or solvate thereof: [ka] (i)R 1 , R 3 and R 4 represents hydrogen, R 2 represents the side chain of aspartic acid, the side chain of arginine or the side chain of glutamic acid; or (ii)R 1 , R 2 and R 3 represents hydrogen, R 4 A 1 A stands for 1 teeth [ka] R 5a represents the side chain of aspartic acid, the side chain of arginine, or the side chain of glutamic acid.

[0057] In another embodiment, the compound of the present invention is 2is hydrogen, and R 1 is the side chain of aspartic acid, and optionally, R 3 and R 4 is independently hydrogen or methyl.

[0058] Further compounds which may be mentioned in this respect are the following compounds and their pharma- ceutically acceptable salts or solvates: [ka]

[0059] In a further embodiment, the present invention also relates to compounds of formula I as defined herein (including compounds of formula I as defined according to any of the preferred embodiments defined herein), provided that: (i) the compound is not a compound of formula A; or (ii) the compound is not a compound of formula B; or (iii) the compound is not a compound of formula C; or (iv) The compound is not selected from the group consisting of: [ka] or (v) The compound is R 2 is hydrogen and R 1 is the side chain of aspartic acid (optionally, R 3 and R 4 is not a compound of formula I, wherein (vi) The compound [ka] isn't it.

[0060] Particularly preferred compounds of the present invention are the following compounds or pharma- ceutically acceptable salts or solvates thereof: [ka] [ka] [ka]

[0061] The compounds of the present invention can be considered to be prodrugs of 4-chloro-N-[2-[(4-chlorophenyl)methyl]-3-oxo-1,2,4-thiadiazol-5-yl]benzamide since they have been found to degrade in vivo to form the above compound, which can be represented as the compound of formula II. [ka]

[0062] The term "prodrug" refers to a compound that, after oral or parenteral administration, is metabolized in vivo to yield a compound that is pharmacologic active in an experimentally detectable amount within a given time (e.g., within a 6-24 hour dosing interval (i.e., 1-4 times per day)). For the avoidance of doubt, the term "parenteral" administration includes all forms of administration other than oral administration. General information regarding prodrugs can be found, for example, in Bundegaard, H. "Design of Prodrugs" p. l-92, Elsevier, New York-Oxford (1985).

[0063] It has been surprisingly found that administration of the compounds of the present invention significantly increases the bioavailability and systemic exposure of 4-chloro-N-[2-[(4-chlorophenyl)methyl]-3-oxo-1,2,4-thiadiazol-5-yl]benzamide compared to the bioavailability and systemic exposure observed following administration of 4-chloro-N-[2-[(4-chlorophenyl)methyl]-3-oxo-1,2,4-thiadiazol-5-yl]benzamide.

[0064] The bioavailability of a drug is the amount of the administered dose that reaches the systemic circulation in the form of that drug. Sufficient bioavailability is important to achieve a therapeutically effective concentration at the site of action. An improvement (i.e., an increase) in bioavailability is the increase in the C concentration in the blood of a subject after administration of a compound (or a pharmaceutical formulation thereof) to the subject. max Or it may be proved by measuring the area under the curve (AUC). The compounds and formulations of the present invention are useful for the therapy described herein in the subject who needs the therapy. The subject that may be mentioned includes animals such as mammals. The specific mammals that may be mentioned include, for example, primates (e.g., human, male or female), cows, horses, dogs and cats. Preferably, the subject is a human.

[0065] The term “C max It will be appreciated by those of skill in the art that " and "AUC" in this context refer to the peak plasma concentration of 4-chloro-N-[2-[(4-chlorophenyl)methyl]-3-oxo-1,2,4-thiadiazol-5-yl]benzamide following administration (e.g., to a human subject) and the integral of the concentration / time curve for that substance following administration of a compound of the invention (or a formulation thereof), respectively.

[0066] It has been found that administration of the compounds of the present invention in particular increases the bioavailability of 4-chloro-N-[2-[(4-chlorophenyl)methyl]-3-oxo-1,2,4-thiadiazol-5-yl]benzamide in vivo, as evidenced by the data in the Examples.These data show that administration of the compounds of the present invention to mammalian subjects increases the plasma exposure of 4-chloro-N-[2-[(4-chlorophenyl)methyl]-3-oxo-1,2,4-thiadiazol-5-yl]benzamide compared to the plasma exposure observed after administration of 4-chloro-N-[2-[(4-chlorophenyl)methyl]-3-oxo-1,2,4-thiadiazol-5-yl]benzamide.

[0067] Thus, administration of the compound of the present invention can increase the bioavailability of 4-chloro-N-[2-[(4-chlorophenyl)methyl]-3-oxo-1,2,4-thiadiazol-5-yl]benzamide compared to administration of 4-chloro-N-[2-[(4-chlorophenyl)methyl]-3-oxo-1,2,4-thiadiazol-5-yl]benzamide. In this context, the expression "increase bioavailability" means that administration of the compound of the present invention results in a greater systemically available fraction of 4-chloro-N-[2-[(4-chlorophenyl)methyl]-3-oxo-1,2,4-thiadiazol-5-yl]benzamide in vivo compared to administration of 4-chloro-N-[2-[(4-chlorophenyl)methyl]-3-oxo-1,2,4-thiadiazol-5-yl]benzamide. The increase in the systemically available fraction of 4-chloro-N-[2-[(4-chlorophenyl)methyl]-3-oxo-1,2,4-thiadiazol-5-yl]benzamide may be at least about 10%, (at least) about 20%, about 30%, about 40%, about 50%, about 60%, about 70%, about 80%, about 90%, about 100% (i.e., 2-fold), about 200% (i.e., 3-fold), about 300% (i.e., 4-fold), about 400% (i.e., 5-fold), about 500% (i.e., 6-fold), about 600% (i.e., 7-fold), about 700% (i.e., 8-fold), about 800% (i.e., 9-fold) or about 900% (i.e., 10-fold).

[0068] The improved bioavailability provided by the compound of the present invention may be demonstrated by using suitable methods known in the art.For example, the improved bioavailability may be demonstrated by comparing the pharmacokinetic data (e.g., AUC data) of the subject administered with the compound of the present invention with the corresponding data of the subject administered with 4-chloro-N-[2-[(4-chlorophenyl)methyl]-3-oxo-1,2,4-thiadiazol-5-yl]benzamide.

[0069] As shown in the examples, the compounds of the present invention have improved stability under acidic conditions (e.g., pH 1.2), and are therefore suitable for the preparation of pharmaceutical formulations for oral administration to patients who require gastric passage.

[0070] The compound of the present invention is also found to be surprisingly effective in activating AMPK, as evidenced by the data in the examples.As an AMPK activator (i.e., an AMPK agonist), the compound of the present invention may be useful in treating disorders or conditions that are improved by the activation of AMPK.Therefore, the compound may be useful in treating certain diseases as described herein.

[0071] The compounds of the present invention may be prepared from 4-chloro-N-[2-[(4-chlorophenyl)methyl]-3-oxo-1,2,4-thiadiazol-5-yl]benzamide. 4-chloro-N-[2-[(4-chlorophenyl)methyl]-3-oxo-1,2,4-thiadiazol-5-yl]benzamide may in turn be prepared according to techniques well known to those skilled in the art. For example, 4-chloro-N-[2-[(4-chlorophenyl)methyl]-3-oxo-1,2,4-thiadiazol-5-yl]benzamide may be prepared according to the techniques described in International Patent Application WO2011 / 004162, the contents of which are incorporated herein by reference in their entirety.

[0072] The compounds of the invention described herein may be prepared according to techniques well known to those skilled in the art, such as those described in the Examples provided below.

[0073] Compounds of formula I may be obtained from available starting materials using appropriate reagents and reaction conditions according to standard techniques, by analogy with methods known in the literature or by conventional synthetic procedures. In this regard, the skilled person may refer in particular to "Comprehensive Organic Synthesis" by BM Trost and I. Fleming, Pergamon Press, 1991.

[0074] For example, there is provided a process for the preparation of the compounds of the invention defined herein, (i) a compound of formula III; [ka] Reaction with compounds of formula IV [ka] or (ii) a compound of formula V, [ka] with a compound of formula IV [ka] R 1 and R 2 is as defined herein, and R 3 and R 4 is as defined herein or independently represents a suitable protecting group (e.g., Boc), in the presence of a suitable base (e.g., 4-dimethylaminopyridine), a suitable amide coupling agent (e.g., N,N'-dicyclohexylcarbodiimide) and a suitable solvent (e.g., tetrahydrofuran) according to procedures known to those skilled in the art.

[0075] For the avoidance of doubt, the processes described herein for preparing the compounds of the invention include processes which comprise the sequential reaction of a compound of formula III with up to three (e.g., two) compounds of formula IV, for example comprising the steps of: (i) reacting a compound of formula III with a compound of formula IV according to the methods described herein; (ii) optionally performing one or more deprotection steps; (iii) reacting the product of step (i) or step (ii) with a compound of formula IV according to the methods described herein; (iv) optionally performing one or more deprotection steps; (v) optionally reacting the product of step (iii) or step (iv) with a compound of formula IV according to the methods described herein; and (vi) optionally carrying out one or more deprotection steps.

[0076] In the above embodiment, the compound of formula IV in each of steps (i), (iii) and (v) may be the same or different.

[0077] Those skilled in the art will appreciate that in the processes described above and below, the functional groups of intermediate compounds may need to be protected with protecting groups. For example, in certain embodiments, in compounds of formula IV, R 3 and R 4 One of the groups may represent hydrogen and the other a suitable protecting group (e.g. Boc). Protection and deprotection of functional groups may be carried out before or after the above reactions.

[0078] Protective groups are well known to those skilled in the art and may be removed according to techniques such as those described below. For example, the protected compounds / intermediates described herein may be chemically converted to unprotected compounds using standard deprotection techniques. The use of protecting groups is fully described in "Protective Groups in Organic Synthesis", 3rd edition, TW Greene & PGM Wutz, Wiley-Interscience (1999).

[0079] Thus, specific transformation steps that may be employed to form compounds of formula I include deprotection steps, for example deprotection of an N-Boc protecting group by reaction in the presence of acid, or a hydroxy group protected as a silyl ether (e.g. a tert-butyl-dimethylsilyl protecting group) may be deprotected by reaction with an acid or a fluoride ion source, for example using the reagent tetrabutylammonium fluoride (TBAF).

[0080] The compound of formula III may be obtained by conventional synthetic procedures from available starting materials using suitable reagents and reaction conditions according to standard techniques. For example, the compound of formula III (referred to as compound 2 in the examples) may be prepared by reacting the compound of formula II with formaldehyde in the presence of a suitable base (e.g., triethylamine) and a suitable solvent (e.g., N,N-dimethylformamide) according to procedures known to those skilled in the art.

[0081] Similarly, the compound of formula II (i.e., 4-chloro-N-[2-[(4-chlorophenyl)methyl]-3-oxo-1,2,4-thiadiazol-5-yl]benzamide; referred to in the Examples as Compound 1) may be prepared according to techniques well known to those skilled in the art, for example, the techniques described in International Patent Application No. WO2011 / 004162.

[0082] Those skilled in the art will recognize that compounds of formula V may be obtained by the synthetic procedures exemplified herein from available starting materials using appropriate reagents and reaction conditions.

[0083] The compounds of the invention may be isolated from their reaction mixtures and, if necessary, purified using conventional techniques known to those skilled in the art. Thus, the processes for the preparation of the compounds of the invention described herein may comprise, as a final step, the isolation and optional purification of the compounds of the invention.

[0084] [Pharmaceutical preparations] As provided herein, the compounds of the present invention are useful as therapeutic agents for treating a variety of medical disorders or conditions. Typically, the compounds of the present invention are administered to a subject in need thereof in the form of a pharmaceutical formulation.

[0085] According to a second aspect of the invention, there is provided a pharmaceutical formulation comprising a compound of formula I (or a pharma- ceutically acceptable salt or solvate thereof). Such a formulation is referred to herein as the formulation of the invention. All of the embodiments and particular features thereof described herein in relation to the first aspect of the invention are disclosed herein in relation to the second aspect of the invention.

[0086] Pharmaceutical formulations according to the second aspect of the invention may be prepared in accordance with standard and / or accepted pharmaceutical practice.

[0087] In an embodiment of the second aspect of the invention, the compound of the invention (or a pharma- ceutically acceptable salt or solvate thereof) is the only pharma- ceutical active ingredient present in the formulation. In a further embodiment of the second aspect of the invention, the compound of the invention (or a pharma- ceutical acceptable salt or solvate thereof) may be present in a formulation with one or more other pharma- ceutical active ingredients or may be administered as part of a combination therapy with one or more other pharma- ceutical active ingredients.

[0088] The formulation of the second aspect of the present invention is generally provided as a mixture containing the compound of the present invention (or a pharma- ceutically acceptable salt or solvate thereof) and one or more pharma- ceutically acceptable excipients. The one or more pharma- ceutically acceptable excipients may be selected according to standard pharmaceutical practice with due consideration given to the intended route of administration. The pharma- ceutical acceptable excipients are preferably chemically inert to the active compounds and preferably have no adverse side effects or toxicity under the conditions of use. Suitable pharmaceutical formulations may be found, for example, in Remington The Science and Practice of Pharmacy, 19th ed., Mack Printing Company, Easton, Pennsylvania (1995). A brief review of methods of drug delivery may be found, for example, in Langer, Science 249, 1527 (1990).

[0089] For example, the formulations of the present invention may include lubricants, binders, fillers, surfactants, diluents, antiadherents, coatings, flavoring agents, coloring agents, glidants, preservatives, sweetening agents, disintegrants, adsorbents, buffers, antioxidants, chelating agents, solubility enhancers, solubility retarders and / or wetting agents.

[0090] Specific pharma- ceutically acceptable excipients that may be mentioned include microcrystalline cellulose, lactose monohydrate, crospovidone, magnesium stearate, colloidal silicon dioxide, anhydrous lactose, dicalcium phosphate, mannitol, pregelatinized starch, hydroxypropylcellulose, povidone, low-substituted hydroxypropylcellulose, croscarmellose sodium, sodium starch glycolate, sodium stearyl fumarate, talc, hydroxypropylmethylcellulose, polysorbate 80, sodium lauryl sulfate, poloxamer 188, poloxamer 407, propylene glycol, titanium dioxide, hypromellose phthalate, hypromellose acetate succinate, methacrylic acid, methyl methacrylate copolymer, Opadry® I, Opadry® II.

[0091] In preparing pharmaceutical formulations of the compounds of the invention for oral administration, the compounds of the invention may be mixed, together or separately, with one or more of the above-mentioned pharmaceutical excipients.

[0092] The mixture of the compound of the present invention and one or more pharma- ceutically acceptable excipients may be processed into pellets or granules or compressed into tablets.Thus, the pharmaceutical formulation of the present invention includes a formulation provided in the form of tablets, mini-tablets, blocks, pellets, particles, granules or powders for oral administration.The mixture of the compound of the present invention and one or more pharma-ceutically acceptable excipients may also be provided in a form suitable for subcutaneous or intramuscular administration, such as a form of lyophilized powder suitable for reconstitution in an injection solution or suitable fluid before administration.

[0093] Those skilled in the art will understand that the formulations described herein may act systemically and therefore may be administered using suitable techniques known to those skilled in the art.The formulations described herein are usually administered orally, subcutaneously or intramuscularly in suitable pharmaceutically acceptable dosage forms.The pharmaceutical formulation of the second aspect of the present invention is preferably an oral pharmaceutical formulation, especially in view of the improved stability under acidic conditions observed for the compounds of the present invention.

[0094] The formulation of the present invention may be prepared for oral administration in the form of capsule.For example, capsules such as soft gelatin capsules may be prepared to contain the compound of the present invention (or its pharmaceutically acceptable salt or solvate) alone, optionally with suitable vehicle, such as vegetable oil, fat, etc.Similarly, hard gelatin capsules may contain the compound of the present invention (or its pharmaceutically acceptable salt or solvate) alone or in combination with solid powder components such as disaccharides (e.g., lactose or saccharose), alcohol sugars (e.g., sorbitol or mannitol), vegetable starches (e.g., potato starch or corn starch), polysaccharides (e.g., amylopectin or cellulose derivatives) or gelling agents (e.g., gelatin).

[0095] Particular pharmaceutical formulations of the present invention include those that may be recited, for example, as formulations provided in the form of a capsule or tablet for oral administration.

[0096] Pharmaceutical formulations that may be mentioned include those in which the compound of the invention (or a pharma- ceutically acceptable salt or solvate thereof) is present in a total amount that is at least 1% by weight (or at least 10% by weight, at least 30% by weight or at least 50% by weight) and up to 99% by weight of the formulation. The weight ratio of the compound of the invention (or a pharma- ceutically acceptable salt or solvate thereof) to the total of the components of the pharmaceutical formulation (i.e., the compound of the invention and all pharmaceutical excipients, such as adjuvants, diluents and carriers) is at least 1:99 (or at least 10:90, at least 30:70 or at least 50:50) and up to 99:1.

[0097] As used herein, "therapeutically effective amount", "effective amount" and "dosage" refer to an amount of the compound of the present invention (or a pharma- ceutically acceptable salt or solvate thereof) sufficient to produce a desired clinical effect, which may be a therapeutic effect and / or a beneficial effect. The effective amount or dosage will vary depending on the age or general health of the subject (e.g., human), the severity of the condition being treated, the particular agent being administered, the duration of treatment, the nature of any co-treatment, the pharma- ceutically acceptable excipients used, and similar factors within the knowledge and expertise of the skilled artisan. Suitably, a "therapeutically effective amount", "effective amount" or "dosage" in each individual case may be determined by the skilled artisan by reference to the relevant texts and literature and / or by using routine experimentation. The skilled artisan will understand that the therapeutic effect need not be complete or curative, so long as some benefit is provided to the subject.

[0098] Those skilled in the art will appreciate that the compounds of the present invention and their formulations may be administered in a variety of doses (e.g., by one or more of the preparations described herein), and suitable doses are readily determined by those skilled in the art. The total dose of the compounds of the present invention (or a pharma- ceutically acceptable salt or solvate thereof) administered to a subject in need thereof may range from about 1 milligram per day (mg / day) to about 3000 mg / day, about 2 mg / day to about 2000 mg / day, or about 5 mg / day to about 1000 mg / day (e.g., about 10 mg / day to about 500 mg / day). Such doses may be, for example, oral doses of the formulations of the second aspect of the present invention. When the compounds of the present invention (or a pharma- ceutically acceptable salt or solvate thereof) are administered intramuscularly or subcutaneously, those skilled in the art will appreciate that the dose should be adjusted accordingly.

[0099] When administered orally, treatment with the above formulations may include administration of a unit dose formulation containing about 1 mg to about 3000 mg, for example about 2 mg to about 2000 mg, or about 5 mg to about 1000 mg (for example about 10 mg to about 500 mg) of the compound of the present invention. Advantageously, treatment may include administration of the compound of the present invention (for example in the form of one or more capsules containing the above formulations) using a single dose per day. Alternatively, the total daily dose of the compound of the present invention may be administered in divided doses two, three or four times per day (for example twice a day with reference to the doses described herein, for example 100 mg, 250 mg, 500 mg or 1000 mg doses per day). The specialist will recognize that the dosage will vary from subject to subject.

[0100] In certain embodiments, the daily dose of the compound of the present invention administered to a subject ranges from about 1 mg to about 3000 mg, preferably from about 1 mg to about 1000 mg.

[0101] The term "about" as used herein, when referring to a measurable value such as the amount of a compound, dosage, time, temperature, etc., refers to a variation of 20%, 10%, 5%, 1%, 0.5%, or even 0.1% of the particular amount. In each case, it is contemplated that the term may be replaced with a notation such as "±10%" (or by indicating the variance of the particular amount calculated based on the relevant value). It is also contemplated that in each case, the term may be omitted.

[0102] For the avoidance of doubt, in the context of the present invention, the dose administered to a subject, particularly a human subject, should be sufficient to achieve a therapeutic response in the subject over a reasonable time frame.Those skilled in the art will recognize that the selection of the exact dose and composition and the most appropriate delivery regimen will also be influenced by, among other things, the pharmacological properties of the formulation, the nature and severity of the condition being treated, and the physical condition and mental acuity of the recipient, as well as the potency of the specific compound, the age, condition, weight, sex and response of the subject being treated, and the stage / severity of the disease.

[0103] In any event, a physician or other skilled artisan will be able to routinely determine the actual dosage that will be most suitable for an individual subject. The above dosage amounts are exemplary of the average case; there can, of course, be individual instances in which higher or lower dosage ranges are merited, and such are within the scope of this invention.

[0104] [Medical Use] As demonstrated herein, the compounds of the present invention are useful as pharmaceuticals. The compounds of the present invention are useful because they have pharmacological activity and / or are metabolized in the body after oral or parenteral administration to form compounds that have pharmacological activity.

[0105] Thus, according to a third aspect of the invention, there is provided a compound of the invention as previously defined herein (i.e. a compound defined in the first aspect of the invention), or a pharmaceutical formulation as defined in relation to the second aspect of the invention, for use in medicine. For the avoidance of doubt, references to compounds defined in the first aspect of the invention include references to compounds of formula I (including all embodiments thereof) and pharma- ceutically acceptable salts and solvates thereof.

[0106] The compounds of the invention (i.e. compounds as defined in the first aspect of the invention), and formulations comprising same, may be particularly useful in the treatment of disorders or conditions ameliorated by activation of AMP-activated protein kinase (AMPK).Accordingly, in a fourth aspect of the invention, there is provided a compound of the invention, or a formulation comprising said compound, for use in the treatment of disorders or conditions ameliorated by activation of AMPK.

[0107] Similarly, there is provided the use of a compound of the invention, or a formulation comprising said compound, in the manufacture of a medicament for the treatment of a disorder or condition ameliorated by activation of AMPK.In a further alternative fourth aspect of the invention, there is provided a method of treating a disorder or condition ameliorated by activation of AMPK comprising administering a compound of the invention (or a formulation comprising said compound) to a subject (e.g., a human) in need thereof.

[0108] By "activating AMPK" is meant that the steady state level of phosphorylation of the Thr-172 moiety of the AMPK-α (AMPK-alpha) subunit is increased compared to the steady state level of phosphorylation in the absence of a compound of Formula I or an active metabolite thereof (e.g., 4-chloro-N-[2-[(4-chlorophenyl)methyl]-3-oxo-1,2,4-thiadiazol-5-yl]benzamide). Alternatively or additionally, it means that there is a higher steady state level of phosphorylation of any other protein downstream of AMPK, such as acetyl-CoA carboxylase (ACC).

[0109] Those skilled in the art will understand that the term "disorders or conditions improved by the activation of AMPK" includes cardiovascular disease (e.g., heart failure), diabetic kidney disease, diabetes (e.g., type 2 diabetes), insulin resistance, nonalcoholic fatty liver disease, nonalcoholic steatohepatitis, pain, opioid addiction, obesity, cancer, inflammation (including chronic inflammatory disease), autoimmune disease, osteoporosis, intestinal disease, and hyperinsulinemia associated with obesity or cardiovascular disease. Other diseases or conditions that may be improved by the activation of AMPK include hyperinsulinemia and related conditions, conditions / disorders in which fibrosis plays a role, sexual dysfunction, and neurodegenerative diseases.

[0110] The term "cancer" will be understood by those skilled in the art to include one or more diseases in the class of disorders characterized by the uncontrolled division of cells and the ability of these cells to invade other tissues, either by direct growth into adjacent tissues through invasion, proliferation, or by transplantation to distant sites through metastasis. "Proliferation" includes an increase in the number and / or size of cancer cells. "Metastasis" refers to the movement or migration (e.g., invasion) of cancer cells from a primary tumor site in a subject's body to one or more other areas in the subject's body (cells can then form secondary tumors).

[0111] Thus, the compounds of the present invention may be suitable for use in the treatment of any type of cancer, including all tumors (non-solid and preferably solid tumors, e.g. carcinoma, adenoma, adenocarcinoma, regardless of organ, blood cancer).For example, the cancer cells may be selected from the group consisting of breast, bile duct, brain, colon, stomach, reproductive organs, thyroid, hematopoietic system, lung and airway, skin, gallbladder, liver, nasopharynx, neuronal cells, kidney, prostate, lymphatic gland and gastrointestinal tract cancer cells.Preferably, the cancer is selected from the group consisting of colon cancer (including colorectal adenoma), breast cancer (e.g. postmenopausal breast cancer), endometrial cancer, hematopoietic system cancer (e.g. leukemia, lymphoma, etc.), thyroid cancer, kidney cancer, esophageal adenocarcinoma, ovarian cancer, prostate cancer, pancreatic cancer, gallbladder cancer, liver cancer and cervical cancer.More preferably, the cancer is selected from the group consisting of colon, prostate, and especially breast cancer. When the cancer is a non-solid tumor, preferably the cancer is a hematopoietic tumor such as leukemia (e.g., acute myeloid leukemia (AML), chronic myeloid leukemia (CML), acute lymphocytic leukemia (ALL), or chronic lymphocytic leukemia (CLL)). Preferably, the cancer cells are breast cancer cells.

[0112] The term "diabetes" (i.e., diabetes mellitus) is understood by those skilled in the art to refer to both type 1 (insulin-dependent) diabetes and type 2 (non-insulin-dependent) diabetes, both of which involve dysfunction of glucose homeostasis. The compounds of the present invention and their formulations may be particularly suitable for use in treating type 1 diabetes and / or type 2 diabetes. The compounds of the present invention are particularly suitable for treating type 2 diabetes.

[0113] In addition to being useful for treating diabetes, the compounds of the present invention are also suitable for treating diabetic kidney disease (i.e., diabetic nephropathy). "Diabetic kidney disease" refers to kidney damage caused by diabetes, and is a serious complication of type 1 diabetes and type 2 diabetes. Diabetic kidney disease affects the kidney's ability to remove waste products from the blood and excrete them in urine, which can lead to kidney failure.

[0114] The compounds of the present invention are also suitable for treating chronic kidney disease, including chronic kidney disease in the absence of type 2 diabetes. "Chronic kidney disease" is a health condition characterized by the gradual loss of kidney function over time.Chronic kidney disease usually occurs as a result of one or more other diseases or health conditions that affect the kidney, such as high blood pressure, diabetes, high cholesterol, kidney infection, glomerulonephritis, polycystic kidney disease, urinary tract obstruction in urine flow, and long-term drug use.

[0115] The term "hyperinsulinemia or related conditions" will be understood by those skilled in the art to include hyperinsulinemia, type 2 diabetes, glucose intolerance, insulin resistance, metabolic syndrome, dyslipidemia, childhood hyperinsulinemia, hypercholesterolemia, hypertension, obesity, fatty liver conditions, diabetic nephropathy, diabetic neuropathy, diabetic retinopathy, cardiovascular disease, atherosclerosis, cerebrovascular conditions such as stroke, systemic lupus erythematosus, neurodegenerative diseases such as Alzheimer's disease, and polycystic ovarian syndrome. Other disease conditions include progressive renal diseases such as chronic renal failure.

[0116] In particular, the compounds of the present invention and formulations thereof may be suitable for use in the treatment of obesity associated with hyperinsulinemia and / or cardiovascular disease associated with hyperinsulinemia.

[0117] The compounds of the present invention and their formulations can also be suitable for use in the treatment of cardiovascular disease (e.g., heart failure), which is not associated with hyperinsulinemia.Similarly, the compounds of the present invention and their formulations can also be suitable for use in the treatment of obesity, which is not associated with hyperinsulinemia.For the avoidance of doubt, the treatment of obesity and / or cardiovascular disease (such as heart failure) in which AMPK activation can be beneficial is included within the scope of the present invention.

[0118] Health conditions / disorders in which fibrosis plays a role include, but are not limited to, scar healing, keloids, scleroderma, pulmonary fibrosis (including idiopathic pulmonary fibrosis), nephrogenic systemic fibrosis, and cardiovascular fibrosis (including endomyocardial fibrosis), systemic sclerosis, liver cirrhosis, ocular macular degeneration, retinal and vitreoretinopathy, Crohn's disease / inflammatory bowel disease, surgical scar tissue formation, radiation and chemotherapy drug induced fibrosis, and cardiovascular fibrosis.

[0119] The compounds of the invention may also be useful in the treatment of sexual dysfunction (eg, the treatment of erectile dysfunction).The compounds of the invention may also be useful in the treatment of inflammation.

[0120] Neurodegenerative diseases that may be mentioned include Alzheimer's disease, Parkinson's disease and Huntington's disease, amyotrophic lateral sclerosis, polyglutamine disorders such as spinal and bulbar muscular atrophy (SBMA), dentate and pallidoluysian atrophy (DRPLA), and some spinocerebellar ataxias (SCAs).

[0121] The compounds of the invention may be useful in the treatment of non-alcoholic fatty liver disease (NAFLD).

[0122] Nonalcoholic fatty liver disease (NAFLD) is defined by excess fat accumulation in the form of triglycerides (steatosis) in the liver (histologically referred to as accumulation in more than 5% of liver cells). It is the most common liver disorder in developed countries (e.g., affecting about 30% of US adults), and most patients are asymptomatic. If left untreated, the condition can progressively worsen and eventually lead to cirrhosis. NAFLD is particularly common in obese patients, with about 80% believed to have the disease.

[0123] NAFLD may be diagnosed when a patient's alcohol intake is not considered to be a major causative factor. Typical thresholds for diagnosing fatty liver disease as "not alcohol-related" are daily intakes of less than 20 g in women and less than 30 g in men.

[0124] Specific diseases or health conditions associated with NAFLD include metabolic conditions such as diabetes, hypertension, obesity, dyslipidemia, dyslipidemia, glycogen storage disease, Weber-Christian disease, acute fatty liver of pregnancy, and lipodystrophy. Other non-alcohol related factors associated with fatty liver disease include malnutrition, total parenteral nutrition, severe weight loss, refeeding syndrome, jejunal oil film bypass, gastric bypass, polycystic ovary syndrome, and diverticulosis.

[0125] Nonalcoholic steatohepatitis (NASH) is the most aggressive form of NAFLD, a condition in which excess fat accumulation (steatosis) is accompanied by inflammation of the liver. In advanced cases, NASH can lead to the development of scar tissue in the liver (fibrosis) and ultimately to cirrhosis. As mentioned above, the compounds of the present invention have been found to be useful in treating NAFLD and inflammation. Thus, the compounds of the present invention are also useful in treating NASH. Thus, in a further embodiment, the treatment is the treatment of nonalcoholic steatohepatitis (NASH).

[0126] It has been shown that AMPK activator compounds (e.g., 4-chloro-N-[2-[(4-chlorophenyl)methyl]-3-oxo-1,2,4-thiadiazol-5-yl]benzamide (i.e., the compound of formula II)) can treat pain (Das V, et al. Reg Anesth Pain Med 2019;0:1-5. doi:10.1136 / rapm-2019-100839 and Das V, et al. Reg Anesth Pain Med 2019;0:1-6. doi:10.1136 / rapm-2019-100651), and such compounds can be considered analgesics. Thus, since the compounds of the present invention can activate AMPK or are metabolized in vivo to form known AMPK-activating compounds, it follows that the compounds of the present invention can be useful for treating pain. In particular, the compounds of the invention may be useful in treating patients with severe pain, chronic pain, or in managing post-operative pain.

[0127] Opioid-based therapy, such as opioid analgesics, is used to treat severe chronic cancer pain, acute pain (e.g., surgery recovery and sudden pain), and their use is increasing in the management of chronic non-malignant pain.However, the increase in the use of opioid-based therapy to treat pain has led to an increase in opioid dependence (e.g., opioid addiction).As an AMPK activator, the compound of the present invention can be used to treat pain instead of opioid-based therapy, as known to those skilled in the art.Therefore, the compound of the present invention can be useful for treating opioid addiction.

[0128] Specific autoimmune diseases known to those skilled in the art include Crohn's disease / inflammatory bowel disease, systemic lupus erythematosus and type 1 diabetes.

[0129] Specific bowel diseases to mention include Crohn's disease / inflammatory bowel disease and cancer of the gastrointestinal tract.

[0130] Those skilled in the art will understand that references to "treatment" of a particular health condition (or, equivalently, "treatment" of that health condition) take their ordinary meaning in the medical arts. In particular, the term may refer to achieving a reduction in the severity and / or frequency of occurrence of one or more clinical symptoms associated with the health condition, as determined by the attending physician of a subject having or susceptible to such a condition.

[0131] Those skilled in the art will understand that the above treatment or prevention is carried out on a subject in need thereof. The need of a subject for the above treatment or prevention may be assessed by those skilled in the art using routine techniques. In the context of the present invention, a "subject in need" of the compound of the present invention includes a subject suffering from a disorder or condition that is improved by the activation of AMPK. As used herein, the terms "disease" and "disorder" (as well as the terms "condition", "disease", "medical problem", etc.) may be used interchangeably.

[0132] Without wishing to be bound by theory, it is believed that administration of the compounds of the present invention increases the bioavailability of 4-chloro-N-[2-[(4-chlorophenyl)methyl]-3-oxo-1,2,4-thiadiazol-5-yl]benzamide in the systemic circulation. Administration of a formulation containing the compounds of the present invention has been shown to increase the plasma concentration of 4-chloro-N-[2-[(4-chlorophenyl)methyl]-3-oxo-1,2,4-thiadiazol-5-yl]benzamide by approximately 10-fold under certain circumstances compared to administration of a formulation containing 4-chloro-N-[2-[(4-chlorophenyl)methyl]-3-oxo-1,2,4-thiadiazol-5-yl]benzamide.

[0133] The compounds of the invention (and their formulations), whether for use in the above indications or otherwise, may have the advantage of being more effective, less toxic, longer acting, more potent, with fewer side effects, more easily absorbed, and / or having a better pharmacokinetic profile (e.g., higher oral bioavailability and / or lower clearance), and / or having other useful pharmacological, physical, or chemical properties, than other treatments known in the prior art. In particular, the compounds of the invention may have the advantage of being more effective and / or exhibiting advantageous properties in vivo.

[0134] [figure] The following drawings are provided to illustrate various aspects of the inventive concepts and are not intended to limit the scope of the invention unless otherwise specified herein.

[0135] FIG. 1 is a Western blot image showing that Compound 3 increases phosphorylation of AMPK in a dose-dependent manner.

[0136] FIG. 2 shows the comparative results of oral pharmacokinetic studies using Compound 1 and Compound 3.

[0137] FIG. 3 shows comparative results of oral pharmacokinetic studies using compounds 5, 6, 7, 9, 10, 13, 14, 16 and 17.

[0138] [Example] The invention will now be described in more detail in the following non-limiting examples.

[0139] The reaction schemes set forth below are intended to provide a general description of the procedures employed in the preparation of the compounds of the invention. The examples provided herein are offered to illustrate, but not limit, the compounds of the invention and the preparation of such compounds and intermediates.

[0140] All starting materials, building blocks, reagents, acids, bases, dehydrating agents, solvents and catalysts used to synthesize the compounds of the invention are either commercially available or can be routinely prepared by procedures described in the literature, e.g., Houben-Weyl "Science of Synthesis" volumes 1-48, Georg Thieme Verlag and subsequent versions.

[0141] The reaction may be carried out in the presence of a suitable solvent or diluent or mixtures thereof in a manner known to those skilled in the art of organic synthesis. The reaction may also be carried out, if necessary, in the presence of an acid or base, with cooling or heating, for example, in a temperature range of about -30°C to about 150°C. In some embodiments, the reaction is carried out in a temperature range of about 0°C to about 100°C, more particularly, in a temperature range of room temperature to about 80°C, in an open or closed reaction vessel and / or in an atmosphere of an inert gas, for example, nitrogen.

[0142] [Abbreviation] The abbreviations used herein will be known to those of skill in the art. In particular, the following abbreviations may be used herein: AUC: Area under the concentration-time curve aq: water-based bw: weight C max Peak plasma concentration d: doublet DCC: N,N'-dicyclohexylcarbodiimide DCM: dichloromethane DMAP: 4-dimethylaminopyridine DMSO: Dimethyl sulfoxide eq: equivalent ESI: electrospray ionization Et3N: Triethylamine EtOH: Ethanol g: grams h: time HPLC: High-performance liquid chromatography LC: Liquid chromatography LCMS: Liquid chromatography-mass spectrometry LC-MS / MS: Liquid chromatography-(tandem) mass spectrometry LLOQ: Lower limit of quantification m: multiplet MeOH: Methanol MeOD: methanol-d4 Min: Minutes mL: milliliter MRT: Average residence time ND: Not detected NMR: nuclear magnetic resonance RT: room temperature s: singlet T 1 / 2 :Half-life TBDMS-Cl: tert-butyldimethylsilyl chloride T max :Time to peak plasma concentration THF: tetrahydrofuran

[0143] [Table 1]

[0144] The present invention is further described with reference to the following examples, which are not intended to limit the scope of the invention.

[0145] Example 1: Preparation of dimethylaminoacetic acid 5-[(Z)-4-chloro-benzoylimino]-2-(4-chloro-benzyl)-3-oxo-[1,2,4]thiadiazolidin-4-ylmethyl ester (Compound 3) [ka]

[0146] Compound 1: 4-chloro-N-[2-[(4-chlorophenyl)methyl]-3-oxo-1,2,4-thiadiazol-5-yl]benzamide Compound 1 was prepared according to the procedure described in WO2011 / 004162.

[0147] Compound 2: 4-chloro-N-{2-[(4-chlorophenyl)methyl]-4-(hydroxymethyl)-3-oxo-1,2,4-thiadiazolidin-5-ylidene}benzamide To a stirred solution of compound 1 (100 g, 0.26 mol) in THF (2.0 L) was added EtN (150 mL, 1.05 mol) followed by 37% formaldehyde (85 mL, 1.05 mol). The reaction mixture was stirred at RT for 12-14 h.

[0148] The reaction mixture was then concentrated to remove THF. The resulting material was treated with toluene (2×100 mL) and dried under reduced pressure at 35-40° C. to give compound 2 as a white solid (103 g, 95.5% yield).

[0149] Compound 3: Dimethylaminoacetic acid 5-[(Z)-4-chloro-benzoylimino]-2-(4-chloro-benzyl)-3-oxo-[1,2,4]thiadiazolidin-4-ylmethyl ester To a stirred solution of N,N-dimethylglycine (50.3 g, 0.48 mol) in THF (2.0 L) was added DMAP (8.93 g, 0.07 mol) and DCC (100.6 g, 0.48 mol). After 10 min, compound 2 (200 g, 0.49 mol) was added to the reaction mixture in four portions over 30 min. The reaction mixture was stirred at 40-50 °C for 16-18 h. The reaction mixture was cooled to 20-30 °C, filtered through Celite®, washed with THF (200 mL), and the filtrate was concentrated to remove THF. The resulting material was slurried at RT using MeOH (300 mL) for 30 min, filtered, and washed with MeOH (50 mL). The collected solid was dried under reduced pressure at 35-40 °C to give compound 3 as a white solid (102 g, 42% yield). 1 H NMR (400 MHz, DMSO) δ (ppm): 2.20 (s, 6H), 3.21 (s, 2H), 4.84 (s, 2H), 6.03 (s, 2H), 7.40 (dd, 4H), 7.63 (d, 2H), 8.15 (d, 2H) LCMS: 495.2 [M+H] HPLC: 96.3% purity at 8.57 min.

[0150] Example 2 Preparation of dimethylaminoacetic acid 5-[(Z)-4-chloro-benzoylimino]-2-(4-chloro-benzyl)-3-oxo-[1,2,4]thiadiazolidin-4-ylmethyl ester citrate (Compound 4) [ka]

[0151] To a stirred solution of dimethylaminoacetic acid 5-[(Z)-4-chloro-benzoylimino]-2-(4-chloro-benzyl)-3-oxo-[1,2,4]thiadiazolidin-4-ylmethyl ester (compound 3; 550 mg, 1.1 mmol) in acetone (30.0 v) was added citric acid (0.9 eq.). The reaction was stirred at RT for 2.3 h. The precipitated compound was then collected by filtration and dried to give compound 4 (450 mg). 1 H NMR (400 MHz, DMSO) δ (ppm): 2.20 (s, 6H), 3.21 (s, 2H), 4.84 (s, 2H), 6.03 (s, 2H), 7.40 (dd, 4H), 7.63 (d, 2H), 8.15 (d, 2H) LCMS: 495.2 [M+H] HPLC: 98.3% purity at 9.95 min

[0152] [Example 3] Preparation of 2-({[5-(4-chlorobenzamido)-2-[(4-chlorophenyl)methyl]-3-oxo-1,2,4-thiadiazolidin-4-yl]methoxy}carbonyl)pyrrolidin-1-ium trifluoroacetate (Compound 5) [ka]

[0153] (Step 1) 1-[(tert-butoxy)carbonyl]pyrrolidine-2-carboxylic acid (Compound A2) To a suspension of L-proline (A1) (1.0 g, 8.68 mmol) in DCM (10 mL) was added triethylamine (1.34 mL, 9.55 mmol) at 0° C., followed by di-tert-butyl dicarbonate (2.18 mL, 9.55 mmol), and the resulting reaction mixture was stirred at RT for 16 h. The reaction mixture was carefully acidified to pH 3 with 5% citric acid solution. The product was extracted with ethyl acetate, and the extract was dried and evaporated in vacuum to give compound A2 as a white solid (1.55 g, 83% yield).

[0154] (Step 2) 1-tert-butyl 2-[5-(4-chlorobenzamido)-2-[(4-chlorophenyl)methyl]-3-oxo-1,2,4-thiadiazolidin-4-yl]methylpyrrolidine-1,2-dicarboxylate (Compound A3) To a stirred solution of compound A2 (0.55 g, 2.56 mmol) in THF (11 mL, 20 V), DMAP (0.06 g, 0.49 mmol) was added followed by DCC (0.52 g, 2.56 mmol) at 0-5 °C and stirred for 20 min. Compound 2 (0.7 g, 1.7 mmol) was added to the reaction mixture at 0-5 °C and the reaction mixture was stirred at ambient temperature (25-30 °C) for 16 h. The reaction mixture was filtered, and the filtrate was concentrated and purified by silica gel column chromatography to give compound A3 as a white solid (0.5 g, 50% yield).

[0155] (Step 3) 2-({[5-(4-chlorobenzamido)-2-[(4-chlorophenyl)methyl]-3-oxo-1,2,4-thiadiazolidin-4-yl]methoxy}carbonyl)pyrrolidin-1-ium trifluoroacetate (Compound 5) To a stirred solution of compound A3 (0.5 g, 0.82 mmol) in dichloromethane (5 mL) was added trifluoroacetic acid (1 mL, 2.0 vol) at 0-5 °C. The reaction mixture was stirred at ambient temperature (25-30 °C) for 16 h. The reaction mixture was concentrated to remove dichloromethane. The resulting residue was reprecipitated with methanol / ether and filtered under vacuum to give compound 5 as a white solid (0.2 g, 39% yield). 1 H NMR (400 MHz, DMSO) δ (ppm): 1.90 (m, 2H), 1,98 (m, 1H), 2.22 (m, 1H), 3,18 (m, 2H), 4.48 (m, 1H), 4.84 (s, 2H), 6.13 (s, 2H), 7.42 (dd, 4H), 7.63 (d, 2H), 8.16 (d, 2H) LCMS: 507.3 [M+H] HPLC: 95.3% purity at 10.07 min.

[0156] In step 1 of this example, L-proline, the L-enantiomer of the naturally occurring amino acid, was used. In all subsequent examples, the L-enantiomer of each naturally occurring amino acid (or its alkylated and / or protected derivatives) was used.

[0157] [Example 4] Preparation of 6-{[5-(4-chlorobenzamido)-2-[(4-chlorophenyl)methyl]-3-oxo-1,2,4-thiadiazolidin-4-yl]methoxy}-6-oxohexane-1,5-bis(aminium) di-trifluoroacetate (Compound 6) [ka]

[0158] (Step 1) [5-(4-chlorobenzamido)-2-[(4-chlorophenyl)methyl]-3-oxo-1,2,4-thiadiazolidin-4-yl]methyl 2,6-bis({[(tert-butoxy)carbonyl]amino})hexanoate (Compound B2) To a stirred solution of 2,6-bis({[(tert-butoxy)carbonyl]amino})hexanoic acid (compound B1; 0.84 g, 2.43 mmol) in THF (17 mL, 20 V), DMAP (0.074 g, 0.60 mmol) was added followed by DCC (0.50 g, 2.43 mmol) at 0-5 °C and stirred for 20 min. Compound 2 (0.5 g, 1.21 mmol) was added to the reaction mixture at 0-5 °C and the reaction mixture was stirred at ambient temperature (25-30 °C) for 16 h. The reaction mixture was filtered, and the filtrate was concentrated and purified by silica gel column chromatography to give compound B2 as a white solid (0.5 g, 55% yield).

[0159] (Step 2) 6-{[5-(4-chlorobenzamido)-2-[(4-chlorophenyl)methyl]-3-oxo-1,2,4-thiadiazolidin-4-yl]methoxy}-6-oxohexane-1,5-bis(aminium) di-trifluoroacetate (6) To a stirred solution of compound B2 (0.5 g, 0.67 mmol) in dichloromethane (5 mL) was added trifluoroacetic acid (1 mL, 2.0 vol) at 0-5 °C. The reaction mixture was stirred at ambient temperature (25-30 °C) for 16 h. The reaction mixture was concentrated to remove dichloromethane. The resulting residue was reprecipitated with acetone / hexane and filtered under vacuum to give compound 6 as a white solid (0.28 g, 63% yield). 1 H NMR (400 MHz, DMSO) δ (ppm): 1.40 (m, 4H), 1,75 (m, 2H), 2.62 (m, 2H), 4,05 (m, 1H), 4.85 (s, 2H), 6.13 (s, 2H), 7.42 (dd, 4H), 7.63 (d, 2H), 8.16 (d, 2H) LCMS: 538.3 [M+H] HPLC: 95.0% purity at 8.28 min.

[0160] [Example 5] Preparation of (6-azaniumyl-1-{[5-(4-chlorobenzamido)-2-[(4-chlorophenyl)methyl]-3-oxo-1,2,4-thiadiazolidin-4-yl]methoxy}-1-oxohexan-2-yl)dimethylazanium di-trifluoroacetate (Compound 7) [ka]

[0161] (Step 1) 6-{[(tert-butoxy)carbonyl]amino}-2-(dimethylamino)hexanoic acid (Compound C2) To a solution of Boc-Lys-OH (C1) (2.0 g, 8.1 mmol) in methanol (20 mL), 37% formaldehyde (2.6 mL, 32.4 mmol) was added followed by Pd / C (0.2 g, 10% w / w), and the resulting mixture was hydrogenated in a Parr apparatus (5 kg, H2 pressure) for 16 h. After completion of the reaction, the reaction mixture was filtered through Celite® and concentrated under reduced pressure. The residue was triturated with ether to give compound C2 as a white solid (1.5 g, 67% yield).

[0162] (Step 2) [5-(4-chlorobenzamido)-2-[(4-chlorophenyl)methyl]-3-oxo-1,2,4-thiadiazolidin-4-yl]methyl 6-{[(tert-butoxy)carbonyl]amino}-2-(dimethylamino)hexanoate (Compound C3) To a stirred solution of compound C2 (0.66 g, 2.43 mmol) in THF (14 mL, 20 V), DMAP (0.035 g, 0.29 mmol) was added followed by DCC (0.50 g, 2.43 mmol) at 0-5 °C and stirred for 20 min. Compound 2 (0.4 g, 0.97 mmol) was added to the reaction mixture at 0-5 °C and the reaction mixture was stirred at ambient temperature (25-30 °C) for 16 h. The reaction mixture was filtered and the filtrate was concentrated and purified by reverse phase column chromatography (8:2 acetonitrile and water) to give compound C3 as a white solid (0.18 g, 64.9% yield).

[0163] (Step 3) (6-azanyl-1-{[5-(4-chlorobenzamido)-2-[(4-chlorophenyl)methyl]-3-oxo-1,2,4-thiadiazolidin-4-yl]methoxy}-1-oxohexan-2-yl)dimethylazanium di-trifluoroacetate (Compound 7) To a stirred solution of compound C3 (0.18 g, 0.027 mmol) in dichloromethane (3.6 mL) was added trifluoroacetic acid (0.36 mL, 2.0 vol) at 0-5 °C. The reaction mixture was stirred at ambient temperature (25-30 °C) for 3 h. The reaction mixture was concentrated to remove dichloromethane. The resulting residue was reprecipitated with acetone / hexane and filtered under vacuum to give compound 7 as an off-white solid (0.15 g, 81% yield). 1 H NMR (400 MHz, D2O) δ (ppm): 1.24 (m, 2H), 1,39 (m, 2H), 1.80 (m, 2H), 2.73 (s, 6H), 4.77 (s, 2H), 6.16 (m, 2H), 7.35 (dd, 4H), 7.58 (d, 2H), 8.12 (d, 2H) LCMS: 566.4 [M+H] HPLC: 97.5% purity at 8.09 min.

[0164] [Example 6] Preparation of (2-{[5-(4-chlorobenzamido)-2-[(4-chlorophenyl)methyl]-3-oxo-1,2,4-thiadiazolidin-4-yl]methoxy}-2-oxoethyl)(methyl)azanium trifluoroacetate (Compound 8) [ka]

[0165] (Step 1) [5-(4-chlorobenzamido)-2-[(4-chlorophenyl)methyl]-3-oxo-1,2,4-thiadiazolidin-4-yl]methyl 2-{[(tert-butoxy)carbonyl](methyl)amino}acetate (D2) To a stirred solution of N-Boc sarcosine (compound D1; 0.57 g, 3.04 mmol) in THF (12 mL, 20 V), DMAP (0.044 g, 0.36 mmol) was added followed by DCC (0.62 g, 3.04 mmol) at 0-5 °C and stirred for 20 min. Compound 2 (0.5 g, 1.21 mmol) was added to the reaction mixture at 0-5 °C and the reaction mixture was stirred at ambient temperature (25-30 °C) for 16 h. The reaction mixture was filtered and the filtrate was concentrated and purified by reverse phase column chromatography to give compound D2 as a white solid (0.22 g, 31% yield).

[0166] (Step 2) (2-{[5-(4-chlorobenzamido)-2-[(4-chlorophenyl)methyl]-3-oxo-1,2,4-thiadiazolidin-4-yl]methoxy}-2-oxoethyl)(methyl)azanium trifluoroacetate (Compound 8) To a stirred solution of compound D2 (0.2 g, 0.34 mmol) in dichloromethane (4.0 mL) was added trifluoroacetic acid (0.4 mL, 2.0 vol) at 0-5° C. The reaction mixture was stirred at ambient temperature (25-30° C.) for 16 h. The reaction mixture was concentrated to remove dichloromethane. The resulting residue was triturated with diethyl ether to give compound 8 as a white solid (0.17 g, 84% yield). 1 H NMR (400 MHz, DMSO) δ (ppm): 2.57 (s, 3H), 4.09 (s, 2H), 4.85 (s, 2H), 6.13 (m, 2H), 7.42 (dd, 4H), 7.64 (d, 2H), 8.17 (d, 2H) LCMS: 481.3 [M+H] HPLC: 98.6% purity at 8.91 min.

[0167] [Example 7] Preparation of 2-{[5-(4-chlorobenzamido)-2-[(4-chlorophenyl)methyl]-3-oxo-1,2,4-thiadiazolidin-4-yl]methoxy}-2-oxoethan-1-aminium trifluoroacetate (Compound 9) [ka]

[0168] (Step 1) [5-(4-chlorobenzamido)-2-[(4-chlorophenyl)methyl]-3-oxo-1,2,4-thiadiazolidin-4-yl]methyl 2-{[(tert-butoxy)carbonyl]amino}acetate (Compound E2) To a stirred solution of N-Boc glycine (compound E1; 0.53 g, 3.04 mmol) in THF (11 mL, 20 V), DMAP (0.044 g, 0.36 mmol) was added followed by DCC (0.62 g, 3.04 mmol) at 0-5 °C and stirred for 20 min. To the reaction mixture, compound 2 (0.5 g, 1.21 mmol) was added at 0-5 °C and the reaction mixture was stirred at ambient temperature (25-30 °C) for 16 h. The reaction mixture was filtered and the filtrate was concentrated and purified by reverse phase column chromatography to give compound E2 as a white solid (0.32 g, 46% yield).

[0169] (Step 2) 2-{[5-(4-chlorobenzamido)-2-[(4-chlorophenyl)methyl]-3-oxo-1,2,4-thiadiazolidin-4-yl]methoxy}-2-oxoethan-1-aminium trifluoroacetate (Compound 9) To a stirred solution of compound E2 (0.3 g, 0.52 mmol) in dichloromethane (3.0 mL) was added trifluoroacetic acid (0.6 mL, 2.0 vol) at 0-5 °C. The reaction mixture was stirred at ambient temperature (25-30 °C) for 2 h. The reaction mixture was concentrated to remove dichloromethane. The resulting residue was triturated with diethyl ether to give compound 9 as a white solid (0.23 g, 76% yield). 1 H NMR (400 MHz, DMSO) δ (ppm): 3.91 (s, 2H), 4.85 (s, 2H), 6.13 (m, 2H), 7.42 (dd, 4H), 7.64 (d, 2H), 8.17 (d, 2H) LCMS: 467.2 [M+H] HPLC: 98.2% purity at 8.55 min.

[0170] [Example 8] Preparation of 2-({[5-(4-chlorobenzamido)-2-[(4-chlorophenyl)methyl]-3-oxo-1,2,4-thiadiazolidin-4-yl]methoxy}carbonyl)-1-methylpyrrolidine (Compound 10) [ka]

[0171] (Step 1) 1-Methylpyrrolidine-2-carboxylic acid (Compound F1) To a solution of L-proline (compound A1; 2.0 g, 17.37 mmol) in methanol (20 mL), 37% formaldehyde (1.54 mL, 19.1 mmol) was added, followed by Pd / C (0.5 g, 25% w / w), and the resulting mixture was hydrogenated in a Parr apparatus (1 kg H2 pressure) for 16 h. After completion of the reaction, the reaction mixture was filtered through Celite® and concentrated under reduced pressure. The residue was triturated with ether to give compound F1 as a white solid (1.9 g, 84% yield).

[0172] (Step 2) 2-({[5-(4-chlorobenzamido)-2-[(4-chlorophenyl)methyl]-3-oxo-1,2,4-thiadiazolidin-4-yl]methoxy}carbonyl)-1-methylpyrrolidin-1-ium trifluoroacetate (Compound 10) To a stirred solution of compound F1 (0.39 g, 3.0 mmol) in THF (10 mL, 20 V), DMAP (0.074 g, 0.60 mmol) was added followed by DCC (0.62 g, 3.04 mmol) at 0-5 °C and stirred for 20 min. Compound 2 (0.5 g, 0.1.21 mmol) was added to the reaction mixture at 0-5 °C and the reaction mixture was stirred at ambient temperature (25-30 °C) for 16 h. The reaction mixture was filtered, and the filtrate was concentrated and purified by reverse phase column chromatography, then reprecipitated using acetone / diethyl ether to give compound 10 as a white solid (0.25 g, 39% yield). 1H NMR (400 MHz, DMSO) δ (ppm): 1.83 (br, 1H), 2,03 (m, 2h), 2.40 (br, 1H), 2,87 (br, 1H), 3,13 (br, 1H), 3.57 (br, 1H), 4.85 (s, 2H), 6.13 (m, 2H), 7.41 (dd, 4H), 7.64 (d, 2H), 8.15(d, 2H) LCMS: 521.3 [M+H] HPLC: 96.9% purity at 9.16 min.

[0173] [Example 9] Preparation of 1-{[5-(4-chlorobenzamido)-2-[(4-chlorophenyl)methyl]-3-oxo-1,2,4-thiadiazolidin-4-yl]methoxy}-1-oxopropan-2-aminium trifluoroacetate (Compound 11) [ka]

[0174] (Step 1) [5-(4-chlorobenzamido)-2-[(4-chlorophenyl)methyl]-3-oxo-1,2,4-thiadiazolidin-4-yl]methyl 2-{[(tert-butoxy)carbonyl]amino}propanoate (Compound G2) To a stirred solution of N-Boc alanine (compound G1) (0.57 g, 3.04 mmol) in THF (10 mL, 20 V), DMAP (0.044 g, 0.36 mmol) was added followed by DCC (0.62 g, 3.04 mmol) at 0-5 °C and stirred for 20 min. Compound 2 (0.5 g, 1.21 mmol) was added to the reaction mixture at 0-5 °C and the reaction mixture was stirred at ambient temperature (25-30 °C) for 16 h. The reaction mixture was filtered and the filtrate was concentrated and purified by reverse phase column chromatography to give compound G2 as a white solid (0.31 g, 43% yield).

[0175] (Step 2) 1-{[5-(4-chlorobenzamido)-2-[(4-chlorophenyl)methyl]-3-oxo-1,2,4-thiadiazolidin-4-yl]methoxy}-1-oxopropane-2-aminium trifluoroacetate (Compound 11) To a stirred solution of compound G2 (0.3 g, 0.51 mmol) in dichloromethane (3.0 mL) was added trifluoroacetic acid (0.9 mL, 3.0 vol) at 0-5° C. The reaction mixture was stirred at ambient temperature (25-30° C.) for 2 h. The reaction mixture was concentrated to remove dichloromethane. The resulting residue was triturated with diethyl ether to give compound 11 as a white solid (0.14 g, 45% yield). 1 H NMR (400 MHz, DMSO) δ (ppm): 1.33 (d, 3H), 4.19 (t, 1h), 4.85 (s, 2H), 6.12 (m, 2H), 7.41 (dd, 4H), 7.64 (m, 1H), 8.15(d, 2H) LCMS: 481.3 [M+H] HPLC: 95.7% purity at 9.14 min.

[0176] [Example 10] Preparation of 1-{[5-(4-chlorobenzamido)-2-[(4-chlorophenyl)methyl]-3-oxo-1,2,4-thiadiazolidin-4-yl]methoxy}-3-methyl-1-oxopentan-2-aminium trifluoroacetate (Compound 12) [ka]

[0177] (Step 1) [5-(4-chlorobenzamido)-2-[(4-chlorophenyl)methyl]-3-oxo-1,2,4-thiadiazolidin-4-yl]methyl 2-{[(tert-butoxy)carbonyl]amino}-3-methylpentanoate (Compound H2) To a stirred solution of N-Boc isoleucine (compound H1; 0.56 g, 3.04 mmol) in THF (10 mL, 20 V), DMAP (0.044 g, 0.36 mmol) was added followed by DCC (0.62 g, 3.04 mmol) at 0-5 °C and stirred for 20 min. Compound 2 (0.5 g, 1.21 mmol) was added to the reaction mixture at 0-5 °C and the reaction mixture was stirred at ambient temperature (25-30 °C) for 16 h. The reaction mixture was filtered and the filtrate was concentrated and purified by reverse phase column chromatography to give compound H2 as a white solid (0.4 g, 52% yield).

[0178] (Step 2) 1-{[5-(4-chlorobenzamido)-2-[(4-chlorophenyl)methyl]-3-oxo-1,2,4-thiadiazolidin-4-yl]methoxy}-3-methyl-1-oxopentan-2-aminium trifluoroacetate (Compound 12) To a stirred solution of compound H2 (0.4 g, 0.64 mmol) in dichloromethane (3.0 mL) was added trifluoroacetic acid (1.2 mL, 3.0 vol) at 0-5 °C. The reaction mixture was stirred at ambient temperature (25-30 °C) for 2 h. The reaction mixture was concentrated to remove dichloromethane. The resulting residue was triturated with diethyl ether to give compound 12 as a white solid (0.22 g, 53% yield). 1 H NMR (400 MHz, DMSO) δ (ppm): 0.67 (t, 3H), 0.84 (d, 2H), 1.15 (m, 1H), 1.33 (m, 1H), 1.81 (br, 1H), 4.10 (s, 1h), 4.86 (s, 2H), 6.03 (d, 1H), 6.20 (d, 1H), 7.45 (dd, 4H), 7.64 (d, 1H), 8.17(d, 2H) LCMS: 523.3 [M+H] HPLC: 98.0% purity at 10.38 min.

[0179] Example 11 Preparation of 5-(4-chlorobenzamido)-2-[(4-chlorophenyl)methyl]-3-oxo-1,2,4-thiadiazolidin-4-yl]methyl 2-(dimethylamino)-3-phenylpropanoate (Compound 13) [ka]

[0180] (Step 1) 2-(Dimethylamino)-3-phenylpropanoic acid (Compound I2) To a solution of Phe-Ala-OH (compound I1; 1.0 g, 6.05 mmol) in methanol (20 mL), 37% formaldehyde (1.96 mL, 24.21 mmol) was added, followed by Pd / C (0.1 g, 10% w / w), and the resulting mixture was hydrogenated in a Parr apparatus (5 kg H2 pressure) for 16 h. After completion of the reaction, the reaction mixture was filtered through Celite® and concentrated under reduced pressure. The residue was triturated with ether to give compound I2 as a white solid (1.0 g, 86% yield).

[0181] (Step 2) [5-(4-chlorobenzamido)-2-[(4-chlorophenyl)methyl]-3-oxo-1,2,4-thiadiazolidin-4-yl]methyl 2-(dimethylamino)-3-phenylpropanoate (Compound 13) To a stirred solution of compound I2 (0.47 g, 2.43 mmol) in THF (10 mL, 20 V), DMAP (0.044 g, 0.36 mmol) was added followed by DCC (0.62 g, 3.04 mmol) at 0-5 °C and stirred for 20 min. Compound 2 (0.5 g, 1.21 mmol) was added to the reaction mixture at 0-5 °C and the reaction mixture was stirred at ambient temperature (25-30 °C) for 16 h. The reaction mixture was filtered and the filtrate was concentrated and purified by reverse phase column chromatography to give compound 13 as a white solid (0.22 g, 30% yield). 1H NMR (400 MHz, DMSO) δ (ppm): 2.49 (s, 6H), 2.92 (m, 2h), 3.51 (t, 1H), 4.89 (s, 2h), 5.97 (m, 2H), 7.07 (m, 5H), 7.40 (d, 2H), 7.45 (d, 2H), 7.64 (d, 2H), 8.15 (d, 2H) LCMS: 586.0 [M+H] HPLC: 97.5% purity at 11.54 min.

[0182] [Example 12] Preparation of 1-{[5-(4-chlorobenzamido)-2-[(4-chlorophenyl)methyl]-3-oxo-1,2,4-thiadiazolidin-4-yl]methoxy}-1-oxo-3-phenylpropan-2-aminium trifluoroacetate (Compound 14) [ka]

[0183] (Step 1) 2-{[(tert-butoxy)carbonyl]amino}-3-phenylpropanoic acid (Compound J1) To a solution of Phe-Ala-OH (compound I1; 1.0 g, 6.05 mmol) in DCM (10 mL) was added triethylamine (1.70 mL, 12.17 mmol) at 0° C., followed by di-tert-butyl pyrocarbonate (2.18 mL, 9.08 mmol), and the resulting reaction mixture was stirred at room temperature for 16 h. The reaction mixture was carefully acidified to pH 4 with 5% citric acid solution. The product was extracted with DCM, the extract was dried, evaporated in vacuum, and purified by reverse phase column chromatography to give compound J1 as a colorless wax (1.3 g, 81% yield).

[0184] (Step 2) [5-(4-chlorobenzamido)-2-[(4-chlorophenyl)methyl]-3-oxo-1,2,4-thiadiazolidin-4-yl]methyl 2-{[(tert-butoxy)carbonyl]amino}-3-phenylpropanoate (Compound J2) To a stirred solution of compound J1 (0.64 g, 2.4 mmol) in THF (10 mL, 20 V), DMAP (0.044 g, 0.36 mmol) was added followed by DCC (0.62 g, 3.04 mmol) at 0-5 °C and stirred for 20 min. Compound 2 (0.5 g, 1.21 mmol) was added to the reaction mixture at 0-5 °C and the reaction mixture was stirred at ambient temperature (25-30 °C) for 16 h. The reaction mixture was filtered and the filtrate was concentrated and purified by reverse phase column chromatography to give compound J2 as a white solid (0.3 g, 37% yield).

[0185] (Step 3) 1-{[5-(4-chlorobenzamido)-2-[(4-chlorophenyl)methyl]-3-oxo-1,2,4-thiadiazolidin-4-yl]methoxy}-1-oxo-3-phenylpropan-2-aminium trifluoroacetate (Compound 14) To a stirred solution of compound J2 (0.3 g, 0.45 mmol) in dichloromethane (3.0 mL) was added trifluoroacetic acid (0.9 mL, 3.0 vol) at 0-5° C. The reaction mixture was stirred at ambient temperature (25-30° C.) for 2 h. The reaction mixture was concentrated to remove dichloromethane. The resulting residue was triturated with diethyl ether to give compound 14 as a white solid (0.2 g, 66% yield). 1 H NMR (400 MHz, DMSO) δ (ppm): 2.98 (m, 1H), 3.14 (m, 1H), 4.44 (m, 1H), 4.85 (s, 2h), 5.877 (s, 1H), 6.20 (d, 1H), 7.07 (m, 5H), 7.41 (d, 2H), 7.47 (d, 2H), 7.64 (d, 2H), 8.16 (d, 2H) LCMS: 558.0 [M+H] HPLC: 98.6% purity at 10.47 min.

[0186] [Example 13] Preparation of 1-{[5-(4-chlorobenzamido)-2-[(4-chlorophenyl)methyl]-3-oxo-1,2,4-thiadiazolidin-4-yl]methoxy}-4-methyl-1-oxopentan-2-aminium trifluoroacetate (Compound 15) [ka]

[0187] (Step 1) [5-(4-chlorobenzamido)-2-[(4-chlorophenyl)methyl]-3-oxo-1,2,4-thiadiazolidin-4-yl]methyl 2-{[(tert-butoxy)carbonyl]amino}-4-methylpentanoate (Compound K2) To a stirred solution of 2-{[(tert-butoxy)carbonyl]amino}-4-methylpentanoic acid (compound K1; 0.56 g, 2.43 mmol) in THF (10 mL, 20 V), DMAP (0.044 g, 0.36 mmol) was added followed by DCC (0.62 g, 3.04 mmol) at 0-5 °C and stirred for 20 min. Compound 2 (0.5 g, 1.21 mmol) was added to the reaction mixture at 0-5 °C and the reaction mixture was stirred at ambient temperature (25-30 °C) for 16 h. The reaction mixture was filtered and the filtrate was concentrated and purified by reverse phase column chromatography to give compound K2 as a white solid (0.4 g, 75% yield).

[0188] (Step 2) 1-{[5-(4-chlorobenzamido)-2-[(4-chlorophenyl)methyl]-3-oxo-1,2,4-thiadiazolidin-4-yl]methoxy}-4-methyl-1-oxopentan-2-aminium trifluoroacetate (Compound 15) To a stirred solution of compound K2 (0.35 g, 0.56 mmol) in dichloromethane (3.0 mL) was added trifluoroacetic acid (1.05 mL, 3.0 vol) at 0-5° C. The reaction mixture was stirred at ambient temperature (25-30° C.) for 2 h. The reaction mixture was concentrated to remove dichloromethane. The resulting residue was triturated with diethyl ether to give compound 15 as a white solid (0.27 g, 93% yield). 1 H NMR (400 MHz, DMSO) δ (ppm): 2.98 (m, 1H), 3.14 (m, 1H), 4.44 (m, 1H), 4.85 (s, 2h), 5.877 (s, 1H), 6.20 (d, 1H), 7.07 (m, 5H), 7.41 (d, 2H), 7.47 (d, 2H), 7.64 (d, 2H), 8.16 (d, 2H) LCMS: 523.4 [M+H] HPLC: 98.0% purity at 10.56 min.

[0189] [Example 14] Preparation of 1-{[5-(4-chlorobenzamido)-2-[(4-chlorophenyl)methyl]-3-oxo-1,2,4-thiadiazolidin-4-yl]methoxy}-3-hydroxy-1-oxopropan-2-aminium trifluoroacetate (Compound 16) [ka]

[0190] (Step 1) 2-{[(tert-butoxy)carbonyl]amino}-3-[(tert-butyldimethylsilyl)oxy]propanoic acid (compound L2) To a solution of Boc-Ser-OH (compound L1; 1.0 g, 4.87 mmol) in DCM (10 mL) was added imidazole (0.53 mg, 7.79 mmol) at 0° C., followed by TBDMS-Cl (1.10 g, 7.30 mmol), and the resulting reaction mixture was stirred at room temperature for 16 h. The product was extracted with DCM, the extract was dried, evaporated in vacuum, and purified by reverse-phase column chromatography to give compound L2 as a colorless wax (0.7 g, 46% yield).

[0191] (Step 2) [5-(4-chlorobenzamido)-2-[(4-chlorophenyl)methyl]-3-oxo-1,2,4-thiadiazolidin-4-yl]methyl 2-{[(tert-butoxy)carbonyl]amino}-3-[(tert-butyldimethylsilyl)oxy]propanoate (Compound L3) To a stirred solution of compound L2 (0.77 g, 2.43 mmol) in THF (10 mL, 20 V), DMAP (0.044 g, 0.36 mmol) was added followed by DCC (0.62 g, 3.04 mmol) at 0-5 °C and stirred for 20 min. Compound 2 (0.5 g, 1.21 mmol) was added to the reaction mixture at 0-5 °C and the reaction mixture was stirred at ambient temperature (25-30 °C) for 16 h. The reaction mixture was filtered and the filtrate was concentrated and purified by reverse phase column chromatography to give compound L3 as a white solid (0.4 g, 46% yield).

[0192] (Step 3) 1-{[5-(4-chlorobenzamido)-2-[(4-chlorophenyl)methyl]-3-oxo-1,2,4-thiadiazolidin-4-yl]methoxy}-3-hydroxy-1-oxopropane-2-aminium trifluoroacetate (Compound 16) To a stirred solution of compound L3 (0.4 g, 0.56 mmol) in dichloromethane (4.0 mL) was added trifluoroacetic acid (1.2 mL, 3.0 vol) at 0-5 °C. The reaction mixture was stirred at ambient temperature (25-30 °C) for 2 h. The reaction mixture was concentrated to remove dichloromethane. The resulting residue was purified by reverse phase column chromatography to give compound 16 as a white solid (0.1 g, 29% yield). 1 H NMR (400 MHz, DMSO) δ (ppm): 3.80 (m, 2H), 4.27 (m, 1H), 4.85 (s, 2h), 5.60 (t, 1H), 6.12 (q, 2H), 7.07 (m, 5H), 7.44 (dd, 4H), 7.63 (d, 2H), 8.18 (d, 2H) LCMS: 497.3 [M+H] HPLC: 91.6% purity at 8.52 min.

[0193] [Example 15] Preparation of 3-carboxy-1-{[5-(4-chlorobenzamido)-2-[(4-chlorophenyl)methyl]-3-oxo-1,2,4-thiadiazolidin-4-yl]methoxy}-1-oxopropan-2-aminium trifluoroacetate (Compound 17) [ka]

[0194] (Step 1) 4-(tert-butoxy)-2-{[(tert-butoxy)carbonyl]amino}-4-oxobutanoic acid (Compound M2) To a solution of L-aspartic acid 4-tert-butyl ester (compound M1; 2.0 g, 10.5 mmol) in dioxane:water (7:3, 20 mL), NaOH (0.84 g, 21.1 mmol) was added, followed by di-tert-butyl pyrocarbonate (3.64 mL, 15.85 mmol), and the resulting reaction mixture was stirred at room temperature for 16 h. The reaction mixture was carefully acidified to pH 3 with 1.5 N HCl acid solution. The product was extracted with ethyl acetate, and the extract was dried and evaporated in vacuum to give compound M2 as a white solid (2.5 g, 81% yield).

[0195] (Step 2) tert-Butyl 1-[5-(4-chlorobenzamido)-2-[(4-chlorophenyl)methyl]-3-oxo-1,2,4-thiadiazolidin-4-yl]methyl 2-{[(tert-butoxy)carbonyl]amino}butanedioate (Compound M3) To a stirred solution of compound M2 (0.88 g, 3.04 mmol) in THF (18 mL, 20 V), DMAP (0.044 g, 0.36 mmol) was added followed by DCC (0.62 g, 3.04 mmol) at 0-5 °C and stirred for 20 min. Compound 2 (0.5 g, 1.21 mmol) was added to the reaction mixture at 0-5 °C and the reaction mixture was stirred at ambient temperature (25-30 °C) for 16 h. The reaction mixture was filtered and the filtrate was concentrated and purified by reverse phase column chromatography to give compound M3 as a white solid (0.45 g, 54% yield).

[0196] (Step 3) 3-Carboxy-1-{[5-(4-chlorobenzamido)-2-[(4-chlorophenyl)methyl]-3-oxo-1,2,4-thiadiazolidin-4-yl]methoxy}-1-oxopropan-2-aminium trifluoroacetate (Compound 17) To a stirred solution of compound M3 (0.45 g, 0.66 mmol) in dichloromethane (4.5 mL) was added trifluoroacetic acid (0.9 mL, 2.0 vol) at 0-5 °C. The reaction mixture was stirred at ambient temperature (25-30 °C) for 6 h. The reaction mixture was concentrated to remove dichloromethane. The resulting residue was purified by reverse phase column chromatography to give compound 17 as a white solid (0.12 g, 28% yield). 1 H NMR (400 MHz, DMSO) δ (ppm): 2.85 (m, 2H), 4.41 (m, 1H), 4.84 (s, 2h), 6.12 (dd, 2H), 6.12 (q, 2H), 7.45 (dd, 4H), 7.63 (d, 2H), 8.18 (d, 2H) LCMS: 525.3 [M+H] HPLC: 94.2% purity at 8.29 min.

[0197] Example 16 Preparation of 5-(4-chlorobenzamido)-2-[(4-chlorophenyl)methyl]-3-oxo-1,2,4-thiadiazolidin-4-yl]methyl 2-[(2S)-2-azanyl-3-phenylpropanamide]acetate chloride (Compound 18) [ka]

[0198] (Step 2) Preparation of methyl 2-[(2S)-2-{[(tert-butoxy)carbonyl]amino}-3-phenylpropanamido]acetate (N2) To a stirred solution of (2S)-2-{[(tert-butoxy)carbonyl]amino}-3-phenylpropanoic acid (J1 synthesized as above; 2.6 g, 9.8 mmol) in DCM (52 mL, 20 V), DCC (2.42 g, 11.76 mmol), DMAP (119 mg, 0.98 mmol) and triethylamine (2.71 mL, 19.6 mmol) were added at 0-5 °C and stirred for 20 min. To the above reaction mixture, 2-aminoacetate methyl hydrochloride (compound N1) (1.6 g, 12.74 mmol) was added at 0-5 °C and the reaction mixture was stirred at ambient temperature (25-30 °C) for 16 h. The reaction product was filtered to remove the by-product (urea), and the filtrate was concentrated and purified by silica gel (60-120 mesh) column chromatography to give compound N2 as an off-white solid (2.73 g, 83% yield).

[0199] (Step 3) Preparation of 2-[(2S)-2-{[(tert-butoxy)carbonyl]amino}-3-phenylpropanamido]acetic acid (Compound N3) To a stirred solution of compound N2 (3.0 g, 8.91 mmol) in methanol (30 mL, 10 V) was added 2 M NaOH solution (12 mL, 4 V) at 0-5 °C, and the reaction mixture was stirred at ambient temperature (25-30 °C) for 16 h. The reaction product was concentrated to remove volatiles, diluted with water (50 mL), and washed with MTBE (100 mL). The aqueous layer was carefully acidified to pH 4-5 with 1.5 N HCl solution and extracted with ethyl acetate (200 mL × 2). The organic layers were combined, dried over Na2SO4, and concentrated under reduced pressure. The resulting residue was washed with hexane (50 mL × 2) to give compound N3 as a white solid (1.6 g, 56% yield).

[0200] (Step 4) Preparation of [5-(4-chlorobenzamido)-2-[(4-chlorophenyl)methyl]-3-oxo-1,2,4-thiadiazolidin-4-yl]methyl 2-[(2S)-2-{[(tert-butoxy)carbonyl]amino}-3-phenylpropanamide]acetate (Compound N4) To a stirred solution of compound N3 (1.6 g, 4.96 mmol) in THF (32 mL, 20 V), DCC (1.53 g, 7.45 mmol) was added followed by DMAP (0.3 g, 2.48 mmol) at 0-5 °C and stirred for 20 min. 4-Chloro-N-{2-[(4-chlorophenyl)methyl]-4-(hydroxymethyl)-3-oxo-1,2,4-thiadiazolidin-5-ylidene}benzamide 6 (1.42 g, 3.47 mmol) was added to the resulting reaction mixture at 0-5 °C and stirred at ambient temperature (25-30 °C) for 16 h. The reaction product was filtered, and the filtrate was concentrated and purified by reverse phase column chromatography to give compound N4 as a white solid (354 mg, 10% yield).

[0201] (Step 5) Preparation of [5-(4-chlorobenzamido)-2-[(4-chlorophenyl)methyl]-3-oxo-1,2,4-thiadiazolidin-4-yl]methyl 2-[(2S)-2-azanyl-3-phenylpropanamide]acetate chloride (Compound 18) To a stirred solution of compound N4 (0.2 g, 0.279 mmol) in THF (1.0 mL, 5 vol) was added hydrochloric acid in diethyl ether (4.0 mL, 20 vol) at 0-5°C, and the reaction mixture was stirred at the same temperature for 20 min. The reaction was concentrated to remove volatiles, and the resulting residue was triturated with diethyl ether to give compound 18 as a white solid (150 mg, 83% yield). 1 H NMR (300 MHz, DMSO) δ (ppm): 2.94 (m, 1H), 3.15 (m, 1H), 4.05 (m, 3h), 4.84 (s, 2H), 6.07 (s, 2H), 7.27 (m, 5H), 7.42 (q, 4H), 7.62 (d, 2H), 8.17 (m, 2H), 8,23 (br, 2h, NH), ), 9.14 (t, 1H, NH) LCMS: 614.31 [M+H] HPLC: 97.35% purity at 11.82 min.

[0202] Example 17 Preparation of 5-(4-chlorobenzamido)-2-[(4-chlorophenyl)methyl]-3-oxo-1,2,4-thiadiazolidin-4-yl]methyl (2S)-2-[(2S)-2-azanyl-3-methylbutanamido]-3-methylbutanoate chloride (Compound 19) [ka]

[0203] (Step 1) Preparation of [5-(4-chlorobenzamido)-2-[(4-chlorophenyl)methyl]-3-oxo-1,2,4-thiadiazolidin-4-yl]methyl (2S)-2-{[(tert-butoxy)carbonyl]amino}-3-methylbutanoate (Compound O2) To a stirred solution of (2S)-2-{[(tert-butoxy)carbonyl]amino}-3-methylbutanoic acid (O1; 3.0 g, 13.8 mmol) in THF (60 mL, 20 V), DMAP (0.84 g, 6.91 mmol) was added followed by DCC (4.27 g, 20.73 mmol) at 0-5 °C and stirred for 20 min. Compound 2 (2.83 g, 6.91 mmol) was added to the above reaction mixture at 0-5 °C and the resulting reaction mixture was stirred at ambient temperature (25-30 °C) for 16 h. The reaction product was diluted with dichloromethane (200 mL), washed with water (2 × 100 mL), and concentrated under reduced pressure to give compound O2 as a white solid (6.0 g, 77%).

[0204] Note: The crude compound O2 was carried on to the next step without purification.

[0205] (Step 2) Preparation of (2S)-1-{[5-(4-chlorobenzamido)-2-[(4-chlorophenyl)methyl]-3-oxo-1,2,4-thiadiazolidin-4-yl]methoxy}-3-methyl-1-oxobutan-2-aminium trifluoroacetate (Compound O3) To a stirred solution of [5-(4-chlorobenzamido)-2-[(4-chlorophenyl)methyl]-3-oxo-1,2,4-thiadiazolidin-4-yl]methyl (2S)-2-{[(tert-butoxy)carbonyl]amino}-3-methylbutanoate (compound O2; 6.0 g, 9.85 mmol) in dichloromethane (25 mL) was added trifluoroacetic acid (24 mL, 4.0 vol) at 0-5° C. and stirred at ambient temperature (25-30° C.) for 4 h. The reaction mixture was concentrated to remove volatiles and the resulting residue was triturated with ether and filtered under vacuum to give compound O3 as a white solid (1.5 g, 24% yield).

[0206] (Step 3) Preparation of [5-(4-chlorobenzamido)-2-[(4-chlorophenyl)methyl]-3-oxo-1,2,4-thiadiazolidin-4-yl]methyl (2S)-2-(2-{[(tert-butoxy)carbonyl]amino}-3-methylbutanamido)-3-methylbutanoate (Compound O4) To a stirred solution of (2S)-2-{[(tert-butoxy)carbonyl]amino}-3-methylbutanoic acid (compound O1; 1.30 g, 6.02 mmol) in THF (30 mL, 20 V), DMAP (0.088 g, 0.72 mmol) was added followed by DCC (1.39 g, 6.75 mmol) at 0-5° C. and stirred for 20 min. To the above reaction mixture, (2S)-1-{[5-(4-chlorobenzamido)-2-[(4-chlorophenyl)methyl]-3-oxo-1,2,4-thiadiazolidin-4-yl]methoxy}-3-methyl-1-oxobutan-2-aminium trifluoroacetate (compound O3; 1.5 g, 2.41 mmol) was added at 0-5° C. and stirred at ambient temperature (25-30° C.) for 16 h. The reaction mixture was filtered, and the filtrate was concentrated and purified by reverse phase column chromatography to give compound O4 as a white solid (0.65 g, 37% yield).

[0207] (Step 4) Preparation of [5-(4-chlorobenzamido)-2-[(4-chlorophenyl)methyl]-3-oxo-1,2,4-thiadiazolidin-4-yl]methyl (2S)-2-[(2S)-2-azanyl-3-methylbutanamido]-3-methylbutanoate trifluoroacetate (Compound O5) To a stirred solution of [5-(4-chlorobenzamido)-2-[(4-chlorophenyl)methyl]-3-oxo-1,2,4-thiadiazolidin-4-yl]methyl (2S)-2-(2-{[(tert-butoxy)carbonyl]amino}-3-methylbutanamido)-3-methylbutanoate (compound O4; 0.66 g, 0.93 mmol) in dichloromethane (25 mL) was added trifluoroacetic acid (1.98 mL, 3.0 vol) at 0-5 °C and the reaction mixture was stirred at ambient temperature (25-30 °C) for 4 h. The reaction mixture was concentrated to remove volatiles and the resulting residue was triturated with ether and filtered under vacuum to give compound O5 as a white solid (0.45 g, 67% yield).

[0208] (Step 5) Preparation of [5-(4-chlorobenzamido)-2-[(4-chlorophenyl)methyl]-3-oxo-1,2,4-thiadiazolidin-4-yl]methyl (2S)-2-[(2S)-2-azanyl-3-methylbutanamido]-3-methylbutanoate chloride (Compound 19) To a stirred solution of [5-(4-chlorobenzamido)-2-[(4-chlorophenyl)methyl]-3-oxo-1,2,4-thiadiazolidin-4-yl]methyl (2S)-2-[(2S)-2-azaniumyl-3-methylbutanamido]-3-methylbutanoate trifluoroacetate (compound O5; 0.45 g, 0.62 mmol) in THF (2.25 mL, 5.0 vol), HCl in diethyl ether (9.0 mL, 20 vol) was added at 0-5°C and the reaction mixture was stirred at the same temperature for 20 min. The reaction was concentrated to remove volatiles and the resulting residue was triturated with diethyl ether to give compound 19 as a white solid (240 mg, 60% yield). 1H NMR (300 MHz, DMSO) δ (ppm): 0.8-0.9 (bm, 12H, -CH3), 2,07 (m, 2H), 3.7 (dd, 1H), 4.26 (m, 1H), 4.84 (s, 2h), 5.97 (dd, 1H), 6.11 (q, 1H), 7.43 (dd, 4H), 7.63 (d, 2H), 8.15 (m, 2K), 8.19 (m, 3H, NH3+), 8.6 (m, 1H, NH) LCMS: 608.53 [M+H] HPLC: 79.5% purity at 11.125 min and 17.3% purity (rotamers) at 11.97 min

[0209] Example 18 Preparation of 5-(4-chlorobenzamido)-2-[(4-chlorophenyl)methyl]-3-oxo-1,2,4-thiadiazolidin-4-yl]methyl (2R)-2-[(2R)-2-azanyl-3-methylbutanamido]-3-methylbutanoate chloride (Compound 20) [ka]

[0210] The method used was similar to that described for the preparation of compound 19 in Example 17, starting from (2R)-2-{[(tert-butoxy)carbonyl]amino}-3-methylbutanoic acid, to give compound 20 as a white solid (300 mg, 88% yield). 1 H NMR (300 MHz, DMSO) δ (ppm): 0.8-0.9 (bm, 12H, -CH3), 2,07 (m, 2H), 3.69 (dd, 1H), 4.26 (m, 1H), 4.84 (s, 2h), 5.97 (dd, 1H), 6.11 (q, 1H), 7.43 (dd, 4H), 7.63 (d, 2H), 8.15 (m, 2K), 8.19 (m, 3H, NH3+), 8.6 (m, 1H, NH) LCMS: 608.53 [M+H] HPLC: 75.93% purity at 13.43 min and 22.45% purity (rotamers) at 14.5 min

[0211] Example 19 Preparation of 5-(4-chlorobenzamido)-2-[(4-chlorophenyl)methyl]-3-oxo-1,2,4-thiadiazolidin-4-yl]methyl 2-[(2S)-2,6-diazanimylhexanamide]acetate dichloride (Compound 21) [ka]

[0212] (Step 1) Preparation of methyl 2-[(2S)-2,6-bis({[(tert-butoxy)carbonyl]amino})hexanamide]acetate (Compound P2) To a stirred solution of (2S)-2,6-bis({[(tert-butoxy)carbonyl]amino})hexanoic acid (compound B1; 2.6 g, 7.51 mmol) in DCM (52 mL, 20 V), DCC (3.86 g, 18.78 mmol), DMAP (275 mg, 2.25 mmol) and triethylamine (2.07 mL, 15.02 mmol) were added at 0-5 °C and stirred for 20 min. Methyl 2-aminoacetate hydrochloride P1 (939 mg, 7.51 mmol) was added to the above reaction mixture at 0-5 °C and further stirred at ambient temperature (25-30 °C) for 16 h. The reaction product was filtered to remove the by-product (urea) and the filtrate was concentrated under reduced pressure. The resulting residue was purified by silica gel column chromatography to obtain compound P2 as a colorless liquid (1.81 g, 58% yield).

[0213] (Step 2) Preparation of 2-[(2S)-2,6-bis({[(tert-butoxy)carbonyl]amino})hexanamide]acetic acid (Compound P3) To a stirred solution of methyl 2-[(2S)-2,6-bis({[(tert-butoxy)carbonyl]amino})hexanamide]acetate (compound P2; 3.0 g, 7.19 mmol) in methanol (30 mL, 10 V) was added 2 M NaOH solution (12 mL, 4 V) at 0-5 °C. The resulting reaction mixture was stirred at ambient temperature (25-30 °C) for 16 h. The reaction product was concentrated to remove methanol, diluted with water (50 mL), and washed with MTBE (50 mL × 2). The aqueous layer was carefully acidified to pH 4-5 with 1.5 N aqueous HCl solution and extracted with ethyl acetate (100 mL × 2). The organic layers were combined, dried over Na2SO4, and concentrated under reduced pressure. The residue was washed with hexane to give compound P3 as a colorless liquid (2.75 g, 95% yield).

[0214] (Step 3) Preparation of [5-(4-chlorobenzamido)-2-[(4-chlorophenyl)methyl]-3-oxo-1,2,4-thiadiazolidin-4-yl]methyl 2-[(2S)-2,6-bis({[(tert-butoxy)carbonyl]amino})hexanamide]acetate (Compound P4) To a stirred solution of (2-[(2S)-2,6-bis({[(tert-butoxy)carbonyl]amino})hexanamide]acetic acid (compound P3; 1.75 g, 4.34 mmol) in THF (35 mL, 20 V), DMAP (0.26 g, 2.17 mmol) was added followed by EDC.HCl (1.24 g, 6.51 mmol) at 0-5 °C and stirred for 20 min. Compound 2 (0.89 g, 2.17 mmol) was added to the above reaction at 0-5 °C and stirred at ambient temperature (25-30 °C) for 16 h. The reaction product was diluted with dichloromethane (200 mL), washed with water (2 × 100 mL), and concentrated under reduced pressure. The residue obtained after evaporation of the solvent was purified by reverse phase column chromatography to give compound P4 as a white solid (0.3 g, 12% yield).

[0215] (Step 4) Preparation of [5-(4-chlorobenzamido)-2-[(4-chlorophenyl)methyl]-3-oxo-1,2,4-thiadiazolidin-4-yl]methyl 2-[(2S)-2,6-diazaniumylhexanamide]acetate di-trifluoroacetate (Compound P5) To a stirred solution of [5-(4-chlorobenzamido)-2-[(4-chlorophenyl)methyl]-3-oxo-1,2,4-thiadiazolidin-4-yl]methyl 2-[(2S)-2,6-bis({[(tert-butoxy)carbonyl]amino})hexanamide]acetate (compound P4; 0.3 g, 0.37 mmol) in dichloromethane (4.5 mL, 15 V) was added trifluoroacetic acid (1.2 mL, 4 vol) at 0-5° C. and stirred at ambient temperature (25-30° C.) for 4 h. The reaction mixture was concentrated to remove volatiles and the resulting residue was washed with ether, filtered and dried under vacuum to give compound P5 as a white solid (0.25 g, 96% yield).

[0216] (Step 5) Preparation of [5-(4-chlorobenzamido)-2-[(4-chlorophenyl)methyl]-3-oxo-1,2,4-thiadiazolidin-4-yl]methyl 2-[(2S)-2,6-diazaniumylhexanamide]acetate dichloride (Compound 21) To a stirred solution of [5-(4-chlorobenzamido)-2-[(4-chlorophenyl)methyl]-3-oxo-1,2,4-thiadiazolidin-4-yl]methyl 2-[(2S)-2,6-diazaniumylhexanamide]acetate di-trifluoroacetate (compound P5; 0.11 g, 0.15 mmol) in THF (0.5 mL, 5.0 vol), HCl in diethyl ether (2.2 mL, 20 vol) was added at 0-5°C and the reaction mixture was stirred at the same temperature for 20 min. The reaction was concentrated to remove volatiles and the resulting residue was triturated with diethyl ether to give compound 21 as a white solid (90 mg, 90% yield). 1H NMR (300 MHz, DMSO) δ (ppm): 1,44 (br, 2H), 1.60 (br, 2H), 1,75 (br, 2H), 2.78 (br, 2H), 3.87 (br, 1H), 4.04 (m, 2h), 4.86 (s, 2H), 6.05 (s, 2H), 7.45 (m, 4H), 7.65 (d, 2H), 8.00 (br, 3H, NH3+), 8.18 (d, 2H). 8.35 (br, 3H, NH3+), 9.14 (br, 1H, NH) LCMS: 595.49 [M+H] HPLC: 94.09% purity at 8.68 min.

[0217] [Example 20] Preparation of (1S)-2-carboxy-1-[(2-{[5-(4-chlorobenzamido)-2-[(4-chlorophenyl)methyl]-3-oxo-1,2,4-thiadiazolidin-4-yl]methoxy}-2-oxoethyl)carbamoyl]ethane-1-aminium chloride (Compound 22) [ka]

[0218] (Step 1) Preparation of benzyl 2-{[(tert-butoxy)carbonyl]amino}acetate (Compound Q2) To a stirred solution of 2-{[(tert-butoxy)carbonyl]amino}acetic acid (compound E1; 2.0 g, 11.42 mmol) in DCM (40 mL, 20 V) was added DMAP (130 mg, 1.14 mmol) followed by EDC.HCl (2.65 g, 17.12 mmol) at 0-5 °C and the resulting mixture was stirred for 5 min. Phenylmethanol (Q1; 1.35 g, 12.55 mmol) was added to the above reaction mixture at 0-5 °C and stirred at ambient temperature (25-30 °C) for 16 h.

[0219] The reaction mixture was diluted with dichloromethane (200 mL), washed with water (2×100 mL), and concentrated under reduced pressure. The residue obtained after evaporation of the solvent was purified by silica gel flash column chromatography to give compound Q2 (2 g, 66% yield) as a white solid.

[0220] (Step 2) Preparation of 2-aminoacetate benzyl (compound Q3) To a stirred solution of 2-{[(tert-butoxy)carbonyl]amino}benzyl acetate (compound Q2; 2 g, 7.54 mmol) in dichloromethane (30 mL, 15 V) was added trifluoroacetic acid (4 mL, 2 V) at 0-5 °C. The reaction mixture was stirred at ambient temperature (25-30 °C) for 3 h. The reaction was concentrated to remove volatiles and the resulting residue was triturated with diethyl ether to give compound Q3 as a white solid (1.8 g, 85% yield).

[0221] (Step 3) Preparation of tert-butyl (3S)-3-{[2-(benzyloxy)-2-oxoethyl]carbamoyl}-3-{[(tert-butoxy)carbonyl]amino}propanoate (Compound Q5) To a stirred solution of (2S)-4-(tert-butoxy)-2-{[(tert-butoxy)carbonyl]amino}-4-oxobutanoic acid (compound Q4; 4.37 g, 15.1 mmol) in DCM (20 mL, 20 V) was added DMAP (140 mg, 1.21 mmol) followed by EDC.HCl (3.23 g, 16.95 mmol) at 0-5 °C and stirred for 5 min. To the above reaction mixture was added 2-aminoacetate benzyl (compound Q3; 1.0 g, 6.05 mmol) and DIPEA (2.96 mL, 18.16 mmol) at 0-5 °C and the reaction mixture was stirred at ambient temperature (25-30 °C) for 16 h. The reaction product was diluted with dichloromethane (400 mL), washed with water (2 x 200 mL) and concentrated under reduced pressure. The residue obtained after evaporation of the solvent was purified by silica gel flash column chromatography to give compound Q5 as a pale yellow wax (1 g, 64% yield).

[0222] (Step 4) Preparation of 2-[(2S)-4-(tert-butoxy)-2-{[(tert-butoxy)carbonyl]amino}-4-oxobutanamido]acetic acid (Compound Q6) To a solution of (3S)-tert-butyl 3-{[2-(benzyloxy)-2-oxoethyl]carbamoyl}-3-{[(tert-butoxy)carbonyl]amino}propanoate (compound Q5; 1.0 g, 2.29 mmol) in ethyl acetate (30 mL, 30 V), Pd / C (0.1 g, 10% w / w) was added and the resulting mass was hydrogenated in a Parr apparatus (5 kg H2 pressure) for 3 h. The reaction product was filtered through a celite pad and concentrated under reduced pressure. The resulting residue was triturated with diethyl ether to give compound Q6 as a white solid (0.67 g, 84% yield).

[0223] (Step 5) Preparation of tert-butyl (3S)-3-{[(tert-butoxy)carbonyl]amino}-3-[(2-{[5-(4-chlorobenzamido)-2-[(4-chlorophenyl)methyl]-3-oxo-1,2,4-thiadiazolidin-4-yl]methoxy}-2-oxoethyl)carbamoyl]propanoate (Compound Q7) To a solution of 2-[(2S)-4-(tert-butoxy)-2-{[(tert-butoxy)carbonyl]amino}-4-oxobutanamido]acetic acid (compound Q6; 0.64 g, 1.87 mmol) in THF (7 mL, 20 V), DMAP (0.052 g, 0.42 mmol) was added followed by DCC (0.43 g, 2.13 mmol) at 0-5 °C and stirred for 20 min. Compound 2 (0.35 g, 0.85 mmol) was added to the above reaction mixture at 0-5 °C and the reaction mixture was stirred at ambient temperature (25-30 °C) for 16 h. The reaction mixture was filtered, and the filtrate was concentrated and purified by reverse phase column chromatography to give compound Q7 as a white solid (0.5 g, 79% yield).

[0224] (Step 6) Preparation of (1S)-2-carboxy-1-[(2-{[5-(4-chlorobenzamido)-2-[(4-chlorophenyl)methyl]-3-oxo-1,2,4-thiadiazolidin-4-yl]methoxy}-2-oxoethyl)carbamoyl]ethane-1-aminium trifluoroacetate (Compound Q8) To a stirred solution of (3S)-3-{[(tert-butoxy)carbonyl]amino}-3-[(2-{[5-(4-chlorobenzamido)-2-[(4-chlorophenyl)methyl]-3-oxo-1,2,4-thiadiazolidin-4-yl]methoxy}-2-oxoethyl)carbamoyl]tert-butyl propanoate (compound Q7; 0.5 g, 0.67 mmol) in dichloromethane (7.5 mL) was added trifluoroacetic acid (2 mL, 4.0 vol) at 0-5° C. and stirred at ambient temperature (25-30° C.) for 4 h. The reaction product was concentrated to remove volatiles and the resulting residue was triturated with diethyl ether to give compound Q8 as a white solid (0.25 g, 53% yield).

[0225] (Step 7) Preparation of (1S)-2-carboxy-1-[(2-{[5-(4-chlorobenzamido)-2-[(4-chlorophenyl)methyl]-3-oxo-1,2,4-thiadiazolidin-4-yl]methoxy}-2-oxoethyl)carbamoyl]ethane-1-aminium chloride (Compound 22) To a stirred solution of (1S)-2-carboxy-1-[(2-{[5-(4-chlorobenzamido)-2-[(4-chlorophenyl)methyl]-3-oxo-1,2,4-thiadiazolidin-4-yl]methoxy}-2-oxoethyl)carbamoyl]ethane-1-aminium trifluoroacetate (compound Q8; 0.25 g, 0.35 mmol) in THF (1.25 mL, 5.0 vol), HCl in diethyl ether (5.0 mL, 20 vol) was added at 0-5°C and the reaction mixture was stirred at the same temperature for 20 min. The reaction was concentrated to remove volatiles and the resulting residue was triturated with diethyl ether to give compound 22 as a white solid (200 mg, 90% yield). 1H NMR (400 MHz, DMSO) δ (ppm): 2.85 (m, 2H), 4.07 (m, 3H), 4.83 (s, 2h), 6.04 (s, 2H), 7.44 (m, 4H), 7.65 (d, 2H), 7.63 (d, 2H), 8.14 (d, 2H), 8.25 (br, 3H, NH3+), 9.01 (s, 1H), 12-14 (br, 1H, -COOH) LCMS: 582.3 M+H] HPLC: 95.26% purity at 11.43 min.

[0226] Example 21 Preparation of azaniumyl({[(4S)-4-azaniumyl-4-[(2-{[5-(4-chlorobenzamido)-2-[(4-chlorophenyl)methyl]-3-oxo-1,2,4-thiadiazolidin-4-yl]methoxy}-2-oxoethyl)carbamoyl]butyl]amino})methylidene]azanium trichloride (Compound 23) [ka]

[0227] (Step 1) Preparation of (2S)-2-{[(tert-butoxy)carbonyl]amino}-5-{[(1E)-{[(tert-butoxy)carbonyl]amino}({[(tert-butoxy)carbonyl]imino})methyl]amino}pentanoic acid (Compound R2) To a solution of (2S)-2-amino-5-carbamimidamidopentanoic acid (compound R1; 8.8 g, 50.51 mmol) in t-butanol (158 mL, 18 V) and water (158 mL, 18 V) was added NaOH (7.07 g, 176.80 mmol) and Boc2O (46.42 mL, 202.06 mmol) at 0 °C. The reaction was stirred at ambient temperature (25-30 °C) for 48 h. The reaction product was evaporated of volatiles and carefully acidified to pH 3-4 with saturated citric acid solution. The product was extracted with ethyl acetate, and the extract was dried over Na2SO4, filtered, evaporated in vacuo, and purified by reverse phase column chromatography to give compound R2 as a white solid (3.7 g, 15% yield).

[0228] (Step 2) Preparation of 2-[(2S)-2-{[(tert-butoxy)carbonyl]amino}-5-{[(1E)-{[(tert-butoxy)carbonyl]amino}({[(tert-butoxy)carbonyl]imino})methyl]amino}pentanamido]benzyl acetate (Compound R3) To a stirred solution of (2S)-2-{[(tert-butoxy)carbonyl]amino}-5-{[(1E)-{[(tert-butoxy)carbonyl]amino}({[(tert-butoxy)carbonyl]imino})methyl]amino}pentanoic acid (compound R2; 1.33 g, 2.15 mmol) in THF (8 mL, 20 V) was added DMAP (0.052 g, 0.43 mmol) followed by EDC.HCl (0.54 g, 2.86 mmol) at 0-5 °C and stirred for 20 min. To the above reaction mixture was added benzyl aminoacetate (compound Q3; 0.4 g, 1.43 mmol) in TEA (0.6 mL, 4.30 mmol) at 0-5 °C and the reaction mixture was stirred at ambient temperature (25-30 °C) for 16 h. The reaction product was diluted with water (100 mL), the product was extracted with dichloromethane (200 mL), the extract was dried and evaporated in vacuum, and the residue was purified by silica gel column chromatography to give compound R3 as a white solid (0.6 g, 67% yield).

[0229] (Step 3) Preparation of 2-[(2S)-2-{[(tert-butoxy)carbonyl]amino}-5-{[(1E)-{[(tert-butoxy)carbonyl]amino}({[(tert-butoxy)carbonyl]imino})methyl]amino}pentanamido]acetic acid (Compound R4) To a solution of 2-[(2S)-2-{[(tert-butoxy)carbonyl]amino}-5-{[(1E)-{[(tert-butoxy)carbonyl]amino}({[(tert-butoxy)carbonyl]imino})methyl]amino}pentanamido]benzyl acetate (compound R3; 0.6 g, 0.96 mmol) in ethyl acetate (6 mL, 10 V), Pd / C (0.06 g, 10% w / w) was added and hydrogenated in a Parr apparatus (5 kg H2 pressure) for 3 h. The reaction product was filtered through a celite bed and the filtrate was concentrated under reduced pressure. The residue was washed with ether to give compound R4 as an off-white solid (0.43 g, 84% yield).

[0230] (Step 4) Preparation of [5-(4-chlorobenzamido)-2-[(4-chlorophenyl)methyl]-3-oxo-1,2,4-thiadiazolidin-4-yl]methyl 2-[(2S)-2-{[(tert-butoxy)carbonyl]amino}-5-{[(1E)-{[(tert-butoxy)carbonyl]amino}({[(tert-butoxy)carbonyl]imino})methyl]amino}pentanamide]acetate (Compound R5) To a stirred solution of 2-[(2S)-2-{[(tert-butoxy)carbonyl]amino}-5-{[(1E)-{[(tert-butoxy)carbonyl]amino}({[(tert-butoxy)carbonyl]imino})methyl]amino}pentanamido]acetic acid (compound R4; 0.41 g, 0.77 mmol) in THF (3.2 mL, 20 V), DMAP (0.014 g, 0.11 mmol) was added followed by DCC (0.18 g, 0.89 mmol) at 0-5 °C and stirred for 20 min. Compound 2 (0.16 g, 0.38 mmol) was added to the above reaction mixture at 0-5 °C and further stirred at ambient temperature (25-30 °C) for 16 h. The reaction mixture was filtered, and the filtrate was concentrated and purified by reverse phase column chromatography to give compound R5 as a white solid (0.25 g, 69% yield).

[0231] (Step 5) Preparation of [azaniumyl({[(4S)-4-azaniumyl-4-[(2-{[5-(4-chlorobenzamido)-2-[(4-chlorophenyl)methyl]-3-oxo-1,2,4-thiadiazolidin-4-yl]methoxy}-2-oxoethyl)carbamoyl]butyl]amino})methylidene]azanium trichloride (Compound 23) To a stirred solution of [5-(4-chlorobenzamido)-2-[(4-chlorophenyl)methyl]-3-oxo-1,2,4-thiadiazolidin-4-yl]methyl 2-[(2S)-2-{[(tert-butoxy)carbonyl]amino}-5-{[(1E)-{[(tert-butoxy)carbonyl]amino}({[(tert-butoxy)carbonyl]imino})methyl]amino}pentanamide]acetate (compound R5; 0.2 g, 0.21 mmol) in THF (1.0 mL, 5.0 vol), HCl in diethyl ether (4.0 mL, 20 vol) was added at 0-5°C and the reaction mixture was stirred at the same temperature for 4 h. The reaction was concentrated to remove volatiles and the resulting residue was triturated with diethyl ether to give compound 23 as a white solid (90 mg, 60% yield). 1H NMR (400 MHz, DMSO) δ (ppm):1.45 (br, 3H), 1.78 (br, 4H), 3.14 (m, 2H), 3.88 (m, 1H), 4.05 (m, 2H), 4.84 (s, 2H), 6.04 (s, 2H), 7.45 (q, 5H), 7.65 (dd, 2H), 7.83 (t, 1H), 8.18 (d, 2H), 8.35 (br, 4H), 9.20 (t, 1H) LCMS: 623.3 [M+H] HPLC: 92.07% purity at 8.52 min.

[0232] [Example 22] Compound stability 5 μM solutions of compound 1 and compound 4 (i.e., the citrate salt of compound 3) in HCl buffer (pH: 1.20) were prepared using 5 mM DMSO stock solutions, and the solutions were incubated at 37° C. for 120 minutes while shaking at 400 rpm using a thermomixer.

[0233] At 0, 15, 30, 60, and 120 minute intervals, sufficient aliquots of compound solution were removed, diluted, and analyzed to determine the stability of compounds 1 and 3 in acidic solution.

[0234] (result) The results of the stability studies are shown in Table 3 below.

[0235] As a result, it was found that compound 3 has higher stability than compound 1 in an acidic solution (ie, pH=1.2).

[0236] [Table 2]

[0237] [Example 23] Activation of AMPK (Culture of INS-1E insulinoma cells and compound treatment) INS-1E cells were cultured as described in Steneberg et al., JCI Insight. 2018;3(12):e99114. https: / / doi.org / 10.1172 / jci.insight.99114, substituting 5% fetal bovine serum for 10% fetal bovine serum when plating for treatment.

[0238] Compound 3 was dissolved in DMSO at 10 mM and frozen at -20°C.

[0239] INS-1E cells were treated in serum-free medium with increasing doses of compound 3 for 4 hours according to the method described in Steneberg et al., JCI Insight. 2018;3(12):e99114. https: / / doi.org / 10.1172 / jci.insight.99114.

[0240] [Western blot analysis] Western blot analysis of INS-1E cells was performed as described in Steneberg et. al., JCI Insight. 2018;3(12):e99114. https: / / doi.org / 10.1172 / jci.insight.99114. Cell lysates were passed through a 30-gauge needle and centrifuged approximately eight times at 14000 rpm for 10 min at +4°C. Quantitative values ​​of AMPKα and phosphorylated-T172 AMPKα were normalized to quantitative values ​​of β-actin.

[0241] (result) The results of the Western blot analysis are presented below in Table 4 and illustrated graphically in FIG.

[0242] The results showed that compound 3 increased phosphorylated-T172 AMPK in a dose-dependent manner in cultured INS-1E cells, and thus compound 3 is an agonist of AMPK.

[0243] [Table 3]

[0244] [Example 24] Single oral administration pharmacokinetic study of compounds 1 and 3 in Sprague Dawley rats Formulation of Compound 1 10.0 mL of a 2% w / v solution of methylcellulose in phosphate buffer (pH 7.5) was added to a 100 mL conical flask along with 1 g of 2 mm glass beads. The solution was vigorously stirred with a magnetic stirrer. 100 mg of compound 1 was slowly added to the solution, and the solution was vigorously stirred for approximately 1 hour. The pH of the formulation was measured to be 7.49. Each formulation was freshly prepared before administration to the animals. The final concentration of compound 1 in the formulation was 10 mg / mL. The formulations were administered at 5 mL / kg body weight.

[0245] Formulation of Compound 3 6 mL of a 2% w / v solution of methylcellulose in phosphate buffer (pH 7.5) was added to a 25 mL conical flask along with 1 g of 2 mm glass beads. The solution was vigorously stirred with a magnetic stirrer. 78 mg of compound 3 was slowly added to the solution, and the solution was vigorously stirred for approximately 1 hour. The pH of the formulation was measured to be 7.41. Each formulation was freshly prepared before administration to the animals. The final concentration of compound 3 in the formulation was 13 mg / mL, which corresponds to 10 mg / mL of compound 1. The formulations were administered at 5 mL / kg body weight.

[0246] (Dose selection and justification for selection) For equimolar dose comparison, doses of 50 mg / kg and 65 mg / kg b.w. were chosen for Compound 1 and Compound 3, respectively.

[0247] (Administration) Healthy adult male Sprague Dawley rats aged 8-10 weeks were used for the experiments after a minimum of 3 days of acclimation. Fed animals were orally administered the compound 1 or compound 3 formulations by gavage at doses of 50 mg / kg or 65 mg / kg body weight, respectively.

[0248] (Blood collection) Under mild isoflurane anesthesia, blood samples were collected into pre-labeled tubes containing anticoagulant (K2EDTA: 2 mg / mL blood) by retro-orbital puncture using a capillary tube during the 72-h period after dosing, details of which are shown in Table 6. When blood was collected at multiple time points, the right and left eyes were used alternately. Collected blood samples were centrifuged at 6000 rpm at 4 °C for 10 min, and plasma samples were separated and stored at -80 °C until analysis.

[0249] (result) The results of the single oral dose pharmacokinetic study in rats are shown in Tables 5 and 6 below and are graphically depicted in Figure 2. In Table 5, C max , T max , AUC last , AUC inf、 AUC extrap , T 1 / 2 and MRT last The values ​​are for 4-chloro-N-[2-[(4-chlorophenyl)methyl]-3-oxo-1,2,4-thiadiazol-5-yl]benzamide (Compound 1) detected after administration of both Compound 1 and Compound 3.

[0250] As a result, the systemic exposure of 4-chloro-N-[2-[(4-chlorophenyl)methyl]-3-oxo-1,2,4-thiadiazol-5-yl]benzamide was surprisingly increased 10-fold when compound 3 was administered compared to compound 1 (Cmax after compound 1 administration was 19.60 μg compound 1 per mL, whereas Cmax after compound 3 administration was 207.00 μg compound 1 per mL). Thus, the systemic exposure of 4-chloro-N-[2-[(4-chlorophenyl)methyl]-3-oxo-1,2,4-thiadiazol-5-yl]benzamide is increased by administering the compound in the form of a compound of the present invention.

[0251] [Table 4]

[0252] [Table 5]

[0253] [Example 25] Single oral administration pharmacokinetic study of compounds 5, 6, 7, 9, 10, 13, 14, and 16 in Sprague Dawley rats (Formulation of Compound) The formulations were prepared according to the method described in Example 19.

[0254] (Selection of dosage and justification for selection) For equimolar dose comparison, a dose equivalent to 50 mg / kg of Compound 1 was chosen.

[0255] (Administration and Blood Collection) Dosing and blood sampling were performed as described in Example 19.

[0256] (result) The results of a single oral dose pharmacokinetic study in rats are shown in Table 7 and below, and are graphically depicted in Figure 3. max , T max , AUC last , AUC inf、 AUC extrap and T 1 / 2 The value of is the value of 4-chloro-N-[2-[(4-chlorophenyl)methyl]-3-oxo-1,2,4-thiadiazol-5-yl]benzamide (Compound 1) detected after administration of the compound.

[0257] The results showed that administration of compounds 5, 6, 7, 9, 10, 13, 14, 16 and 17 resulted in mean maximum plasma concentrations (Cmax) of 4-chloro-N-[2-[(4-chlorophenyl)methyl]-3-oxo-1,2,4-thiadiazol-5-yl]benzamide ranging from 71 to 137 μg / mL.

[0258] [Table 6]

[0259] [Table 7]

[0260] [Example 26] Stability of Compound 17 The stability of compound 17 was evaluated in buffers of pH 1.20, 7.40, and 9.20. The pH 1.20 study was performed using HCl buffer. Phosphate buffer was used for the pH 7.40 medium, and alkaline borate buffer was used for the pH 9.20 medium.

[0261] (HCl buffer) Potassium chloride (1.491 g) was dissolved in 100 mL of ultrapure water type-1 to obtain a 0.2 M KCl solution. Approximately 50 mL of 0.2 M KCl was placed in a beaker and the pH was adjusted to 1.20 using 0.2 M HCl solution. The final volume was made up to 200 mL using ultrapure water type-1 and stored at room temperature (22 ± 3 °C).

[0262] (Phosphate buffer solution) Potassium phosphate monobasic (2.722 g) was dissolved in 100 mL of ultrapure water type-1 to obtain a 0.2 M solution. Approximately 50 mL of the 0.2 M solution was placed in a beaker and the pH was adjusted to 7.40 using 0.2 M NaOH solution. The final volume was made up to 200 mL using ultrapure water type-1 and stored at room temperature (22 ± 3 °C).

[0263] (Alkaline borate buffer) Boric acid (1.237 g) and potassium chloride (1.491 g) were dissolved in 100 mL of ultrapure water type-1 to obtain a 0.2 M solution. Approximately 50 mL of the 0.2 M solution was taken in a beaker and the pH was adjusted to 9.20 using 0.2 M NaOH solution. The final volume was made up to 200 mL using ultrapure water type-1 and stored at room temperature (22 ± 3 °C).

[0264] A 5 μM solution of compound 17 in each buffer was prepared using a 5 mM DMSO stock solution and incubated at 22±3° C. for 120 minutes with shaking at 400 rpm in a thermomixer.

[0265] At 0, 15, 30, 60, and 120 minute intervals, sufficient aliquots of the compound solutions were removed, diluted, and analyzed to determine the stability of compound 17 in each solution and the amount of compound 1 present. The amount of compound 1 present was compared to the amount expected assuming complete conversion of compound 17 to compound 1.

[0266] (result) The results of the stability studies are shown in Table 9 below.

[0267] [Table 8]

[0268] The results showed that compound 17 had the greatest stability in a low pH environment (pH = 1.2), but was rapidly decomposed above pH 7.40.

[0269] Example 27: Additional Stability Results The stability of compounds 3, 6, 9, 20 and 23 was tested in the same manner as described above for compound 17. The amount of each compound remaining at 15 and 30 minutes is shown in Table 10.

[0270] [Table 9]

[0271] Compounds 17 and 23 were found to degrade rapidly at pH 7.40 and relatively stable at pH 1.2. These data suggest that when the amino acid side chain is lipophilic (e.g., alkyl, see compound 20), the stability is rather high at pH 7.4. When the side chain contains H or -NH2 (glycine or lysine base), the stability is low. When aspartic acid is used (compound 17), the stability is very low at higher pH. The compounds will remain intact in the stomach, but may be hydrolyzed in the intestine. [Brief description of the drawings]

[0272] [Figure 1] 1 is a Western blot image showing that Compound 3 increases phosphorylation of AMPK in a dose-dependent manner. [Diagram 2] 1 shows comparative results of oral pharmacokinetic studies using Compound 1 and Compound 3. [Diagram 3] Comparative results of oral pharmacokinetic studies using compounds 5, 6, 7, 9, 10, 13, 14, 16 and 17.

Claims

1. A compound of formula I or a pharmaceutically acceptable salt or solvate thereof: 【Chemical 1】 R １ and R ２ are independently hydrogen, the side chain of a proteinogenic amino acid, and —OH, —NH ２ and —C(O)NH ２ C optionally substituted with one or more substituents selected from the group consisting of １－６ selected from the group consisting of alkyl; R ３ and R ４ are independently hydrogen, C optionally substituted with a phenyl group １－４ Alkyl, and A １ or R １ (or R ２ ) and R ３ (or R ４ ) together with the carbon and nitrogen atoms to which they are attached form a 4- to 6-membered heterocycloalkyl group; A １ teeth, 【Chemistry 2】 or 【Chemistry 3】 represents A ２ is hydrogen, C optionally substituted with phenyl groups １－４ Alkyl, 【Chemistry 4】 or 【Chemistry 5】 represents Each A ３ are independently hydrogen or C optionally substituted with a phenyl group. １－４ represents alkyl; and R ５ａ and R ５ｂ are independently hydrogen, the side chains of proteinogenic amino acids, and -OH, -NH ２ and —C(O)NH ２ C optionally substituted with one or more substituents selected from the group consisting of １－６ alkyl.

2. R ２ is hydrogen, and further, R １ is hydrogen or the side chain of a proteinogenic amino acid selected from the group consisting of Glu, Arg, His, Lys, Ser, Thr, Asn, Gln, Cys, Sec, Ala, Ile, Leu, Met, Phe, Trp, Tyr, Asp and Val; or R ２ is hydrogen, and further, R １ and R ３ 10. The compound of claim 1, wherein: together with the carbon atom and nitrogen atom to which they are attached, form a pyrrolidine ring.

3. R ２ is hydrogen, and further, R １ is hydrogen or the side chain of a proteinogenic amino acid selected from the group consisting of Arg, His, Lys, and Trp, or R ２ is hydrogen, and further, R １ and R ３ 10. The compound of claim 1, wherein: together with the carbon atom and nitrogen atom to which they are attached, form a pyrrolidine ring.

4. R ２ is hydrogen, and further, R １ The compound of claim 1 , wherein is the side chain of aspartic acid.

5. R ３ and R ４ are independently hydrogen, C optionally substituted with one or more phenyl groups １－３ Alkyl, 【Chemistry 6】 and 【Chemistry 7】 and further selected from the group consisting of: R ５ａ is hydrogen or a side chain of a proteinogenic amino acid.

6. R ３ and R ４ are independently hydrogen, C optionally substituted with one or more phenyl groups １－３ Alkyl, 【Chemistry 8】 and 【Chemistry 9】 or selected from the group consisting of R １ and R ３ form a pyrrolidine ring together with the carbon atom and nitrogen atom to which they are attached, and further, R ４ C optionally substituted with hydrogen, one or more phenyl groups １－３ Alkyl, 【Chemistry 10】 and 【Chemistry 11】 and R ５ａ 2. The compound of claim 1, wherein is a side chain of a proteinogenic amino acid selected from the group consisting of Phe, Val, Lys, Asp, and Arg.

7. R ３ and R ４ are independently hydrogen and C optionally substituted with one or more phenyl groups; １－３ 2. The compound of claim 1, wherein the compound is selected from the group consisting of alkyl.

8. R ３ and R ４ 10. The compound of claim 1, wherein is independently selected from the group consisting of hydrogen and methyl.

9. R ２ is hydrogen, and R １ is hydrogen or a side chain of a proteinogenic amino acid selected from the group consisting of Glu, Arg, His, Lys, Ser, Thr, Asn, Gln, Cys, Sec, Ala, Ile, Leu, Met, Phe, Trp, Tyr, Asp, and Val; R ３ and R ４ are independently hydrogen, C optionally substituted with one or more phenyl groups １－３ Alkyl, 【Chemistry 12】 and 【Chemistry 13】 or selected from the group consisting of R ２ is hydrogen, and R １ and R ３ form a pyrrolidine ring together with the carbon atom and nitrogen atom to which they are attached, and further, R ４ C optionally substituted with hydrogen, one or more phenyl groups １－３ Alkyl, 【Chemistry 14】 and 【Chemistry 15】 and R ５ａ is hydrogen or a side chain of a proteinogenic amino acid selected from the group consisting of Glu, Arg, His, Lys, Ser, Thr, Asn, Gln, Cys, Sec, Ala, Ile, Leu, Met, Phe, Trp, Tyr, Asp, and Val.

10. The compound is Dimethylaminoacetic acid 5-[(Z)-4-chloro-benzoylimino]-2-(4-chloro-benzyl)-3-oxo-[1,2,4]thiadiazolidin-4-ylmethyl ester; Dimethylaminoacetic acid 5-[(Z)-4-chloro-benzoylimino]-2-(4-chloro-benzyl)-3-oxo-[1,2,4]thiadiazolidin-4-ylmethyl ester citrate salt; 2-({[5-(4-chlorobenzamido)-2-[(4-chlorophenyl)methyl]-3-oxo-1,2,4-thiadiazolidin-4-yl]methoxy}carbonyl)pyrrolidin-1-ium trifluoroacetate; 6-{[5-(4-chlorobenzamido)-2-[(4-chlorophenyl)methyl]-3-oxo-1,2,4-thiadiazolidin-4-yl]methoxy}-6-oxohexane-1,5-bis(aminium) di-trifluoroacetate; (6-azaniumyl-1-{[5-(4-chlorobenzamido)-2-[(4-chlorophenyl)methyl]-3-oxo-1,2,4-thiadiazolidin-4-yl]methoxy}-1-oxohexan-2-yl)dimethylazanium di-trifluoroacetate; (2-{[5-(4-chlorobenzamido)-2-[(4-chlorophenyl)methyl]-3-oxo-1,2,4-thiadiazolidin-4-yl]methoxy}-2-oxoethyl)(methyl)azanium trifluoroacetate; 2-{[5-(4-chlorobenzamido)-2-[(4-chlorophenyl)methyl]-3-oxo-1,2,4-thiadiazolidin-4-yl]methoxy}-2-oxoethane-1-aminium trifluoroacetate; 2-({[5-(4-chlorobenzamido)-2-[(4-chlorophenyl)methyl]-3-oxo-1,2,4-thiadiazolidin-4-yl]methoxy}carbonyl)-1-methylpyrrolidin-1-ium trifluoroacetate; 1-{[5-(4-chlorobenzamido)-2-[(4-chlorophenyl)methyl]-3-oxo-1,2,4-thiadiazolidin-4-yl]methoxy}-1-oxopropan-2-aminium trifluoroacetate; 1-{[5-(4-chlorobenzamido)-2-[(4-chlorophenyl)methyl]-3-oxo-1,2,4-thiadiazolidin-4-yl]methoxy}-3-methyl-1-oxopentan-2-aminium trifluoroacetate; [5-(4-chlorobenzamido)-2-[(4-chlorophenyl)methyl]-3-oxo-1,2,4-thiadiazolidin-4-yl]methyl 2-(dimethylamino)-3-phenylpropanoate; 1-{[5-(4-chlorobenzamido)-2-[(4-chlorophenyl)methyl]-3-oxo-1,2,4-thiadiazolidin-4-yl]methoxy}-1-oxo-3-phenylpropan-2-aminium trifluoroacetate; 1-{[5-(4-chlorobenzamido)-2-[(4-chlorophenyl)methyl]-3-oxo-1,2,4-thiadiazolidin-4-yl]methoxy}-4-methyl-1-oxopentan-2-aminium trifluoroacetate; 1-{[5-(4-chlorobenzamido)-2-[(4-chlorophenyl)methyl]-3-oxo-1,2,4-thiadiazolidin-4-yl]methoxy}-3-hydroxy-1-oxopropan-2-aminium trifluoroacetate; 3-carboxy-1-{[5-(4-chlorobenzamido)-2-[(4-chlorophenyl)methyl]-3-oxo-1,2,4-thiadiazolidin-4-yl]methoxy}-1-oxopropan-2-aminium trifluoroacetate; [5-(4-chlorobenzamido)-2-[(4-chlorophenyl)methyl]-3-oxo-1,2,4-thiadiazolidin-4-yl]methyl 2-[(2S)-2-azaniumyl-3-phenylpropanamide]acetate chloride; [5-(4-chlorobenzamido)-2-[(4-chlorophenyl)methyl]-3-oxo-1,2,4-thiadiazolidin-4-yl]methyl (2S)-2-[(2S)-2-azaniumyl-3-methylbutanamido]-3-methylbutanoate chloride; [5-(4-chlorobenzamido)-2-[(4-chlorophenyl)methyl]-3-oxo-1,2,4-thiadiazolidin-4-yl]methyl (2R)-2-[(2R)-2-azaniumyl-3-methylbutanamido]-3-methylbutanoate chloride; [5-(4-chlorobenzamido)-2-[(4-chlorophenyl)methyl]-3-oxo-1,2,4-thiadiazolidin-4-yl]methyl 2-[(2S)-2,6-diazaniumylhexanamide]acetate dichloride; (1S)-2-carboxy-1-[(2-{[5-(4-chlorobenzamido)-2-[(4-chlorophenyl)methyl]-3-oxo-1,2,4-thiadiazolidin-4-yl]methoxy}-2-oxoethyl)carbamoyl]ethane-1-aminium chloride; and [azaniumyl({[(4S)-4-azaniumyl-4-[(2-{[5-(4-chlorobenzamido)-2-[(4-chlorophenyl)methyl]-3-oxo-1,2,4-thiadiazolidin-4-yl]methoxy}-2-oxoethyl)carbamoyl]butyl]amino})methylidene]azanium trichloride 2. The compound of claim 1 selected from the group consisting of:

11. A pharmaceutical formulation comprising a compound as defined in any one of claims 1 to 10, or a pharmaceutically acceptable salt or solvate thereof, and a pharmaceutically acceptable excipient.

12. 12. A pharmaceutical formulation as defined in claim 11 for use in medicine.

13. 12. A pharmaceutical formulation as defined in claim 11 for use in the treatment of a disorder or condition ameliorated by activation of AMPK.

14. 11. Use of a compound of formula I, or a pharmaceutically acceptable salt or solvate thereof, as defined in any one of claims 1 to 10, in the manufacture of a medicament for the treatment of a disorder or condition ameliorated by activation of AMPK.

15. 11. A method of treating a disorder or condition ameliorated by activation of AMPK, comprising administering to a subject in need thereof a compound of formula I as defined in any one of claims 1 to 10, or a pharmaceutically acceptable salt or solvate thereof.

16. 14. The pharmaceutical formulation of claim 13, wherein the disorder or condition that is improved by activation of AMPK is selected from the group consisting of cardiovascular disease (such as heart failure), diabetic kidney disease, type 2 diabetes, insulin resistance, non-alcoholic fatty liver disease, non-alcoholic steatohepatitis, pain, opioid addiction, obesity, cancer, inflammation (including chronic inflammatory disease), autoimmune disease, osteoporosis, intestinal disease, and hyperinsulinemia associated with obesity or cardiovascular disease.

17. (i) a compound of formula III; and 【Chemistry 16】 Reaction with a compound of formula IV 【Chemistry 17】 or (ii) a compound of formula V; 【Chemistry 18】 with a compound of formula IV, 【Chemistry 19】 R １ and R ２ is as defined in any one of claims 1 to 10; and R ３ and R ４ is as defined in any one of claims 1 to 10 or independently represents a protecting group.

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