Prodrugs of itaconate and methyl itaconate

Abstract

The invention relates to prodrugs of itaconate and methyl itaconate. Disclosed herein are prodrugs of itaconic acid and 1-methyl itaconic acid and 4-methyl itaconic acid and their use for the treatment of diseases, disorders or conditions associated with inflammation. The present application further discloses a method for treating a disease, disorder or condition associated with inflammation, the method comprising administering to a subject in need thereof the prodrug as described above.

Description

[0001] This application is a divisional application of the application with the filing date of October 29, 2020, application number 202080088327.2, and invention title "Prodrugs of Itaconate and Methyl Itaconate". Background

[0003] Inflammatory macrophages show a large accumulation of itaconate, which has been shown to exert significant anti-inflammatory activity by inhibiting succinate dehydrogenase and inducing electrophilic stress that activates the NRF2-dependent antioxidant response. However, considering the charged nature of itaconate, itaconate itself does not exhibit good pharmacokinetic properties or cell permeability properties. Therefore, the therapeutic potential of administering exogenous itaconate has not been clarified. Overview

[0005] The subject matter disclosed by the present invention provides prodrugs of itaconic acid and 1-methylitaconic acid and 4-methylitaconic acid and their use for treating inflammation-related diseases, disorders or conditions.

[0006] More particularly, in some aspects, the subject matter disclosed by the present invention provides compounds of formula (I):

[0007]

[0008] Wherein:

[0009] R1 and R2 may be the same or different and each independently selected from one or more of the following and combinations thereof:

[0010] (a) -OR3, where R3 is H or a C1-C6 straight-chain or branched unsubstituted or substituted hydrocarbon group;

[0011] (b) Where n is an integer selected from 1, 2, 3, and 4; R4 is a C1-C6 straight-chain or branched unsubstituted or substituted hydrocarbon group or -OR5, where R5 is a C1-C6 straight-chain or branched unsubstituted or substituted hydrocarbon group;

[0012] (c) Where m is an integer selected from 1, 2, 3, and 4; p is an integer selected from 1, 2, 3, 4, 5, 6, 7, 8, 9, 10, 11, 12, 13, 14, 15, 16, 17, 18, 19, and 20; and R6 is a C1-C6 straight-chain or branched unsubstituted or substituted hydrocarbon group;

[0013] (d) Where R7 is selected from:

[0014] (i) -C(=O)-O-R8, where R8 is a C1-C6 straight-chain or branched unsubstituted or substituted hydrocarbon group;

[0015] (ii) wherein R9 is H or an unsubstituted or substituted C1-C4 straight-chain or branched-chain hydrocarbon group; R 10 is an unsubstituted or substituted C1-C6 straight-chain or branched-chain hydrocarbon group; R 11 and R 12 are each independently H or a protecting group; and R 13 is an unsubstituted or substituted C1-C6 straight-chain or branched-chain hydrocarbon group;

[0016] (iii) wherein q is an integer selected from 1, 2, 3, 4, 5, 6, 7, 8,

[0017] 9 and 10; R 11 and R 12 are each independently H or a protecting group; R 14 is H or an unsubstituted or substituted C1-C4 straight-chain or branched-chain hydrocarbon group; and R 15 is an unsubstituted or substituted C1-C6 straight-chain or branched-chain hydrocarbon group; and

[0018] (iv) wherein R 16 and R 17 are each independently selected from H, an unsubstituted or substituted C1-C4 straight-chain or branched-chain hydrocarbon group, and a protecting group; R 18 is an aryl group;

[0019] (e) wherein R 19 is an unsubstituted or substituted C1-C4 straight-chain or branched-chain hydrocarbon group;

[0020] (f) wherein u is an integer selected from 1, 2, 3, and 4; R 20 is H or an unsubstituted or substituted C1-C4 straight-chain or branched-chain hydrocarbon group; and R 21 is -OR 22 wherein R 22 is an unsubstituted or substituted C1-C6 straight-chain or branched-chain hydrocarbon group or -NR 23 R 24 wherein R 23 and R 24 are each independently H or an unsubstituted or substituted C1-C4 straight-chain or branched-chain hydrocarbon group;

[0021] provided that R1 and R2 cannot both be -OH or both be -OR3 at the same time;

[0022] and pharmaceutically acceptable salts thereof.

[0023] In some embodiments:

[0024] (a) R1 is -OR3 and R2 is selected from -OR3, wherein R7 is selected from -C(=O)-O-R8,

[0025]

[0026] (b) R1 is and R2 is selected from -OR3, wherein R7 is selected from -C(=O)-O-R8,

[0027] (c) R1 is and R2 is selected from -OR3, wherein R7 is selected from -C(=O)-O-R8,

[0028] (d) R1 is and R2 is selected from -OR3, wherein R7 is selected from -C(=O)-O-R8,

[0029] (e) R1 is and R2 is selected from -OR3, wherein R7 is selected from -C(=O)-O-R8,

[0030]

[0031] (f) R1 is wherein R7 is -C(=O)-O-R8 and R2 is selected from -OR3,

[0032]

[0033] wherein R7 is selected from -C(=O)-O-R8,

[0034] (g) R1 is wherein R7 is and R2 is selected from -OR3, wherein R7 is selected from -C(=O)-O-R8,

[0035] (h) R1 is wherein R7 is and R2 is selected from -OR3,

[0036]

[0037] wherein R7 is selected from -C(=O)-O-R8, and (i) R1 is wherein R7 is and R2 is selected from -OR3,

[0038]

[0039] wherein R7 is selected from -C(=O)-O-R8,

[0040] provided that R1 and R2 cannot both be -OH or both be -OR3 at the same time;

[0041] and its pharmaceutically acceptable salts.

[0042] In some embodiments:

[0043] (a-i) R1 is -OH and R2 is selected from:

[0044] wherein R7 is -C(=O)-O-R8;

[0045] (a-ii) R1 is -OR3 and R2 is selected from:

[0046] wherein R7 is selected from

[0047] -C(=O)-O-R8,

[0048] (b) R1 is and R2 is selected from:

[0049] -OR3,

[0050] (c) R1 is and R2 is -OR3;

[0051] (d) R1 is and R2 is selected from: -OR3 and

[0052] (e) R1 is wherein R7 is -C(=O)-O-R8 and R2 is -OR3;

[0053] (f) R1 is wherein R7 is and R2 is -OR3;

[0054] (g) R1 is wherein R7 is and R2 is -OR3;

[0055] (h) R1 is wherein R7 is and R2 is -OR3; and

[0056] (i) R1 is and R2 is -OR3.

[0057] In some embodiments, R3, R4, R5, R6, R8, R9, R 10 , R 13 , R 15 and R 22 are each independently selected from methyl, ethyl, n-propyl, isopropyl, n-butyl, sec-butyl, isobutyl, tert-butyl, pentyl, isopentyl, neopentyl, n-hexyl, 2-methylpentane, 3-methylpentane, 2,2-dimethylbutane, and 2,3-dimethylbutane.

[0058] In some embodiments, R9, R 14 , R 16 , R 17 , R 19 , R 20 , R 23 and R 24 are each independently selected from methyl, ethyl, n-propyl, isopropyl, n-butyl, sec-butyl, isobutyl, and tert-butyl.

[0059] In some embodiments, the protecting group is selected from tert-butoxycarbonyl (boc), benzyloxycarbonyl (Cbz), p-methoxybenzylcarbonyl (Moz or MeOZ), 9-fluorenylmethoxycarbonyl (Fmoc) group, benzyl (Bn), p-methoxybenzyl (PMB), 3,4-dimethoxybenzyl (DMPM), p-toluenesulfonyl (Ts), Troc (trichloroethyl chloroformate), (4-nitrophenyl)sulfonyl (Nosyl), and nitrophenylthio (Nps).

[0060] In some embodiments, R 18 is selected from phenyl.

[0061] In some embodiments:

[0062] (a-i) R1 is -OH and R2 is selected from:

[0063]

[0064] (a-ii) R1 is selected from -OCH3, -OCH(CH3)2 and -OC(CH3)3 and R2 is selected from:

[0065]

[0066] (b) R1 is and R2 is -OCH3(5, 6) or

[0067] (c) R1 is and R2 is -OCH3(12, 13);

[0068] (d) R1 is and R2 is selected from: -OCH3(11) and

[0069] (e) R1 is and R2 is -OCH3(14);

[0070] (f) R1 is and R2 is -OCH3(17, 19);

[0071] (g) R1 is and R2 is -OCH3(15, 16);

[0072] (h) R1 is and R2 is -OCH3(18, 20); and

[0073] (i) R1 is and R2 is -OCH3(3, 4).

[0074] In some embodiments, the compound of formula (I) is selected from:

[0075]

[0076]

[0077]

[0078] and its pharmaceutically acceptable salts.

[0079] The present application also provides a pharmaceutical composition, which comprises the compound as described above and a pharmaceutically acceptable excipient.

[0080] The present application further provides a method for treating a disease, disorder or condition associated with inflammation, the method comprising administering to a subject in need of such treatment the compound or composition as described above.

[0081] In other aspects, the subject matter disclosed by the present invention provides a pharmaceutical composition comprising a compound of formula (I) or a pharmaceutically acceptable salt thereof and a pharmaceutically acceptable excipient.

[0082] In still other aspects, the subject matter disclosed by the present invention provides a method for treating a disease, disorder or condition associated with inflammation, the method comprising administering to a subject in need of such treatment a compound of formula (I) or a pharmaceutical composition thereof.

[0083] Certain aspects of the subject matter disclosed by the present invention have been set forth above, all or part of which are presented by the subject matter disclosed by the present invention, and when considered in conjunction with the following appended examples and drawings which are described in detail below, other aspects will become apparent as the description proceeds. Brief Description of the Drawings

[0085] The subject matter disclosed by the present invention has been described so generally, and now reference will be made to the drawings, which are not necessarily drawn to scale, and in which:

[0086] Figure 1 Showing the plasma stability of a representative prodrug;

[0087] Figure 2 Showing the release of active monomethyl itaconate (in plasma) from a representative prodrug; and

[0088] Figure 3 Showing the release of active itaconic acid (in plasma) from a representative prodrug. Detailed Description

[0090] The subject matter disclosed by the present invention will now be described more fully hereinafter with reference to the drawings, in which certain but not all embodiments of the present invention are shown. The same numbers refer to the same elements throughout. The subject matter disclosed by the present invention may be embodied in many different forms and should not be construed as limited to the embodiments set forth herein; rather, these embodiments are provided so that this disclosure will satisfy applicable legal requirements. In fact, many modifications and other embodiments of the subject matter disclosed by the present invention described herein will come to mind to those skilled in the art having the benefit of the teachings presented in the foregoing description and the related drawings. Accordingly, it should be understood that the subject matter disclosed by the present invention is not limited to the specific embodiments disclosed and that modifications and other embodiments are intended to be included within the scope of the appended claims.

[0091] I. Novel prodrugs of itaconates and methyl itaconates

[0092] Inflammatory macrophages show a large accumulation of itaconates, which have been shown to exert significant anti-inflammatory activity by inhibiting succinate dehydrogenase and inducing electrophilic stress that activates the NRF2-dependent antioxidant response. However, considering the charged nature of itaconates, itaconates themselves do not exhibit good pharmacokinetic properties or cell permeability properties. Given the significant therapeutic potential of administering exogenous itaconates, the subject matter disclosed herein provides prodrugs of itaconic acid and 1-methylitaconic acid and 4-methylitaconic acid, which have the ability to achieve enhanced cell permeability and release active itaconic acid and the corresponding methyl itaconates after oral, systemic, or topical administration. The prodrugs disclosed herein also exert an anti-inflammatory effect on keratinocytes.

[0093] A. Representative compounds of formula (I)

[0094] In some embodiments, the subject matter disclosed herein provides compounds of formula (I):

[0095]

[0096] Wherein:

[0097] R1 and R2 may be the same or different and each independently selected from one or more of the following and combinations thereof:

[0098] (a) -OR3, wherein R3 is H or a C1-C6 straight-chain or branched, unsubstituted or substituted hydrocarbon group;

[0099] (b) Where n is an integer selected from 1, 2, 3, and 4; R4 is a C1-C6 straight-chain or branched, unsubstituted or substituted hydrocarbon group or -OR5, wherein R5 is a C1-C6 straight-chain or branched, unsubstituted or substituted hydrocarbon group;

[0100] (c) Where m is an integer selected from 1, 2, 3, and 4; p is an integer selected from 1, 2, 3, 4, 5, 6, 7, 8, 9, 10, 11, 12, 13, 14, 15, 16, 17, 18, 19, and 20; and R6 is a C1-C6 straight-chain or branched, unsubstituted or substituted hydrocarbon group;

[0101] (d) Wherein R7 is selected from:

[0102] (i) -C(=O)-O-R8, wherein R8 is a C1-C6 straight-chain or branched, unsubstituted or substituted hydrocarbon group;

[0103] (ii) wherein R9 is H or an unsubstituted or substituted C1-C4 straight-chain or branched-chain hydrocarbon group; R 10 is an unsubstituted or substituted C1-C6 straight-chain or branched-chain hydrocarbon group; R 11 and R 12 are each independently H or a protecting group; and R 13 is an unsubstituted or substituted C1-C6 straight-chain or branched-chain hydrocarbon group;

[0104] (iii) wherein q is an integer selected from 1, 2, 3, 4, 5, 6, 7, 8,

[0105] 9 and 10; R 11 and R 12 are each independently H or a protecting group; R 14 is H or an unsubstituted or substituted C1-C4 straight-chain or branched-chain hydrocarbon group; and R 15 is an unsubstituted or substituted C1-C6 straight-chain or branched-chain hydrocarbon group; and

[0106] (iv) wherein R 16 and R 17 are each independently selected from H, an unsubstituted or substituted C1-C4 straight-chain or branched-chain hydrocarbon group, and a protecting group; R 18 is an aryl group;

[0107] (e) wherein R 19 is an unsubstituted or substituted C1-C4 straight-chain or branched-chain hydrocarbon group;

[0108] (f) wherein u is an integer selected from 1, 2, 3, and 4; R 20 is H or an unsubstituted or substituted C1-C4 straight-chain or branched-chain hydrocarbon group; and R 21 is -OR 22 wherein R 22 is an unsubstituted or substituted C1-C6 straight-chain or branched-chain hydrocarbon group or -NR 23 R 24 wherein R 23 and R 24 are each independently H or an unsubstituted or substituted C1-C4 straight-chain or branched-chain hydrocarbon group;

[0109] provided that R1 and R2 cannot both be -OH or both be -OR3 at the same time;

[0110] and pharmaceutically acceptable salts thereof.

[0111] In certain embodiments of the compounds of formula (I):

[0112] (a) R1 is -OR3 and R2 is selected from -OR3, where R7 is selected from -C(=O)-O-R8,

[0113]

[0114] (b) R1 is and R2 is selected from -OR3, where R7 is selected from -C(=O)-O-R8,

[0115] (c) R1 is and R2 is selected from -OR3, where R7 is selected from -C(=O)-O-R8,

[0116] (d) R1 is and R2 is selected from -OR3, where R7 is selected from -C(=O)-O-R8,

[0117] (e) R1 is and R2 is selected from -OR3, where R7 is selected from -C(=O)-O-R8,

[0118]

[0119] (f) R1 is where R7 is -C(=O)-O-R8 and R2 is selected from -OR3,

[0120]

[0121] where R7 is selected from -C(=O)-O-R8,

[0122] (g) R1 is where R7 is and R2 is selected from -OR3, where R7 is selected from -C(=O)-O-R8,

[0123] (h) R1 is wherein R7 is and R2 is selected from -OR3,

[0124]

[0125] wherein R7 is selected from -C(=O)-O-R8, and (i) R1 is wherein R7 is and R2 is selected from -OR3,

[0126] wherein R7 is selected from -C(=O)-O-R8,

[0127] provided that R1 and R2 cannot both be -OH or both be -OR3 at the same time;

[0128] and its pharmaceutically acceptable salts.

[0129] In certain further embodiments of the compounds of formula (I):

[0130] (a-i) R1 is -OH and R2 is selected from:

[0131] wherein R7 is -C(=O)-O-R8;

[0132] (a-ii) R1 is -OR3 and R2 is selected from:

[0133] wherein R7 is selected from

[0134] -C(=O)-O-R8,

[0135] (b) R1 is and R2 is selected from:

[0136] -OR3,

[0137] (c) R1 is and R2 is -OR3;

[0138] (d) R1 is and R2 is selected from: -OR3 and

[0139] (e) R1 is wherein R7 is -C(=O)-O-R8 and R2 is -OR3;

[0140] (f) R1 is wherein R7 is and R2 is -OR3;

[0141] (g) R1 is wherein R7 is and R2 is -OR3;

[0142] (h) R1 is wherein R7 is and R2 is -OR3; and

[0143] (i) R1 is and R2 is -OR3.

[0144] In some embodiments of the compounds of formula (I), R3, R4, R5, R6, R8, R9, R 10 , R 13 , R 15 and R 22 can each independently be a C1, C2, C3, C4, C5 or C6 straight-chain or branched unsubstituted or substituted hydrocarbyl group selected from methyl, ethyl, n-propyl, isopropyl, n-butyl, sec-butyl, isobutyl, tert-butyl, pentyl, isopentyl, neopentyl, n-hexyl, 2-methylpentane, 3-methylpentane, 2,2-dimethylbutane, and 2,3-dimethylbutane.

[0145] In some embodiments of the compounds of formula (I), R9, R 14 , R 16 , R 17 , R 19 , R 20 , R 23 and R 24 can each independently be a C1, C2, C3 or C4 straight-chain or branched unsubstituted or substituted hydrocarbyl group selected from methyl, ethyl, n-propyl, isopropyl, n-butyl, sec-butyl, isobutyl, and tert-butyl.

[0146] Representative substituent groups include, but are not limited to, hydrocarbyl, substituted hydrocarbyl, halogen, aryl, substituted aryl, hydrocarbyloxy, hydroxy, nitro, amino, hydrocarbylamino, dihydrocarbylamino, sulfate, cyano, mercapto, and hydrocarbylthio.

[0147] In certain embodiments of the compounds of formula (I), the protecting group is selected from tert-butoxycarbonyl (boc), benzyloxycarbonyl (Cbz), p-methoxybenzylcarbonyl (Moz or MeOZ), 9-fluorenylmethoxycarbonyl (Fmoc) group, benzyl (Bn), p-methoxybenzyl (PMB), 3,4-dimethoxybenzyl (DMPM), p-toluenesulfonyl (Ts), Troc (trichloroethyl chloroformate), (4-nitrophenyl)sulfonyl (Nosyl), and nitrophenylthio (Nps). In a particular embodiment, the protecting group is tert-butoxycarbonyl (boc).

[0148] In some embodiments of the compounds of formula (I), R 18 is selected from phenyl.

[0149] In certain embodiments of the compounds of formula (I):

[0150] (a-i) R1 is -OH and R2 is selected from:

[0151]

[0152] (a-ii) R1 is selected from -OCH3, -OCH(CH3)2, and -OC(CH3)3 and R2 is selected from:

[0153]

[0154] (b) R1 is and R2 is -OCH3 (5, 6) or

[0155]

[0156] (c) R1 is and R2 is -OCH3 (12, 13);

[0157] (d) R1 is and R2 is selected from: -OCH3 (11) and

[0158] (e) R1 is and R2 is -OCH3 (14);

[0159] (f) R1 is and R2 is -OCH3 (17, 19);

[0160] (g) R1 is and R2 is -OCH3 (15, 16);

[0161] (h) R1 is and R2 is -OCH3 (18, 20); and

[0162] (i) R1 is and R2 is -OCH3 (3, 4).

[0163] In certain embodiments of the compounds of formula (I), the compounds are selected from:

[0164]

[0165]

[0166]

[0167] In some embodiments, the compounds of formula (I) include pharmaceutically acceptable salts. In some embodiments, the pharmaceutically acceptable salts are acidic salts. In certain embodiments, the pharmaceutically acceptable salt is trifluoroacetate (TFA).

[0168] Table 1 provides representative compounds of formula (I) and the active agents released by each corresponding prodrug.

[0169]

[0170]

[0171]

[0172]

[0173]

[0174] refers to 4-methyl itaconate; refers to 1-methyl itaconate

[0175] In still other embodiments, the subject matter disclosed herein provides a method for treating a disease, disorder or condition associated with inflammation, the method comprising administering to a subject in need of such treatment a compound of formula (I) or a pharmaceutical composition thereof.

[0176] As used herein, the term "treatment" can include reversing, alleviating, inhibiting the progression of the disease, disorder or condition to which such terms apply or one or more symptoms or manifestations of such disease, disorder or condition, and preventing or reducing the likelihood thereof. Prevention refers to preventing the disease, disorder, condition or its symptoms or manifestations from occurring, or preventing them from worsening in severity. Thus, the compounds disclosed herein can be administered prophylactically to prevent or reduce the occurrence or recurrence of a disease, disorder or condition.

[0177] The "subject" treated by the methods disclosed by the present invention in many of its embodiments is desirably a human subject. However, it should be understood that the methods described herein are effective with respect to all vertebrate species, and all vertebrate species are intended to be included within the term "subject". Thus, a "subject" can include a human subject for medical purposes, such as for the treatment of an existing condition or disease or for prophylactic treatment to prevent the onset of a condition or disease, or an animal subject for medical, veterinary, or developmental purposes. Suitable animal subjects include mammals, including but not limited to, primates, such as humans, monkeys, apes, etc.; bovines, such as domestic cattle, bulls, etc.; ovines, such as sheep, etc.; caprines, such as goats, etc.; porcines, such as pigs, hogs, etc.; equines, such as horses, donkeys, zebras, etc.; felines, including wild and domestic cats; canines, including dogs; lagomorphs, including rabbits, hares, etc.; and rodents, including mice, rats, etc. The animal can be a transgenic animal. In certain embodiments, the subject is a human, including but not limited to, fetal, neonatal, infant, juvenile, and adult subjects. Additionally, a "subject" can include a patient suffering from or suspected of suffering from a condition or disease. Thus, the terms "subject" and "patient" are used interchangeably herein. The term "subject" also refers to an organism, tissue, cell, or collection of cells from a subject.

[0178] Generally, an "effective amount" of an active agent or a drug delivery device refers to the amount necessary to elicit a desired biological response. As would be understood by one of ordinary skill in the art, the effective amount of an agent or device can vary depending on factors such as the desired biological endpoint, the agent to be delivered, the composition of the pharmaceutical composition, the target tissue, etc.

[0179] The term "combination" is used in its broadest sense and means that a subject is administered at least two agents, and more particularly, a compound of formula (I) and at least one β-lactam antibiotic, and optionally one or more antibacterial agents. More particularly, the term "combination" refers to the simultaneous administration of two (or more) active agents for the treatment of, for example, a single disease state. As used herein, the active agents can be combined and administered in a single dosage form, can be administered as separate dosage forms at the same time, or can be administered as separate dosage forms administered alternately or sequentially on the same or different days. In one embodiment of the subject matter disclosed by the present invention, the active agents are combined and administered in a single dosage form. In another embodiment, the active agents are administered as separate dosage forms (e.g., where it is desirable to vary the amount of one active agent while the amount of the other active agent remains unchanged). The single dosage form can include additional active agents for the treatment of the disease state.

[0180] In addition, the compounds of formula (I) described herein can be administered alone or in combination with an adjuvant that enhances the stability of the compounds of formula (I), alone or in combination with one or more antibacterial agents, and in certain embodiments facilitate the administration of pharmaceutical compositions containing them (which contain other active ingredients), provide increased dissolution or dispersion, increase inhibitory activity, provide adjuvant therapy, etc. Advantageously, such combination therapies utilize lower doses of conventional therapeutic agents, thereby avoiding the possible toxicities and adverse side effects that occur when those agents are used as monotherapies.

[0181] The timing of administration of the compounds of formula (I) and at least one additional therapeutic agent can be varied, provided that the beneficial effects of the combination of these agents are achieved. Thus, the phrase "in combination with" refers to the administration of the compounds of formula (I) and at least one additional therapeutic agent simultaneously, sequentially, or a combination thereof. Thus, a subject to whom a combination of a compound of formula (I) and at least one additional therapeutic agent is administered can receive the compound of formula (I) and at least one additional therapeutic agent at the same time (i.e., simultaneously) or at different times (i.e., on the same day or on different days, sequentially in any order), provided that the effect of the combination of the two agents is achieved in the subject.

[0182] When administered sequentially, the agents can be administered within 1 minute, 5 minutes, 10 minutes, 30 minutes, 60 minutes, 120 minutes, 180 minutes, 240 minutes, or more of each other. In other embodiments, the agents administered sequentially can be administered within 1 day, 5 days, 10 days, 15 days, 20 days, or more of each other. When the compounds of formula (I) and at least one additional therapeutic agent are administered simultaneously, they can be administered to the subject as separate pharmaceutical compositions (each pharmaceutical composition containing the compound of formula (I) or at least one additional therapeutic agent), or they can be administered to the subject as a single pharmaceutical composition containing both agents.

[0183] When administered in combination, the effective concentration of each of the agents that elicits a particular biological response can be less than the effective concentration of each agent when administered alone, thereby allowing a reduction in the dose of one or more of the agents relative to the dose that would be required if the agent were administered as a single agent. The effects of multiple agents can, but need not be, additive or synergistic. The agents can be administered multiple times.

[0184] In some embodiments, when administered in combination, two or more agents can have a synergistic effect. As used herein, the terms “synergy,” “synergistic,” “synergistically,” and their derivatives, such as in “synergistic effect” or “synergistic combination” or “synergistic composition,” refer to the situation where the biological activity of a combination of a compound of formula (I) and at least one additional therapeutic agent is greater than the sum of the biological activities of the respective agents when administered alone.

[0185] Synergy can be expressed in terms of a “synergy index (SI),” which can generally be determined from the ratios determined by the method described by F.C. Kull et al., Applied Microbiology 9, 538 (1961):

[0186] Q a / Q A +Q b / Q B = synergy index (SI)

[0187] Wherein:

[0188] Q A is the concentration of component A acting alone that produces an endpoint related to component A;

[0189] Q a is the concentration of component A in the mixture that produces the endpoint;

[0190] Q B is the concentration of component B acting alone that produces an endpoint related to component B;

[0191] And

[0192] Q b is the concentration of component B in the mixture that produces the endpoint.

[0193] Generally, when the sum of Q a / Q A and Q b / Q B is greater than 1, it indicates antagonism. When the sum is equal to 1, it indicates additivity. When the sum is less than 1, it indicates a synergistic effect. The lower the SI, the greater the synergy shown by that particular mixture. Thus, a “synergistic combination” has an activity higher than can be expected based on the activities observed for the individual components when used alone. Additionally, a “synergistically effective amount” of a component refers to the amount of the component necessary to elicit a synergistic effect with, for example, another therapeutic agent present in the composition.

[0194] In another embodiment, the subject matter disclosed herein provides a pharmaceutical composition comprising a compound of formula (I) alone or a combination of a compound of formula (I) with one or more additional therapeutic agents, admixed with a pharmaceutically acceptable excipient. Those skilled in the art will recognize that pharmaceutical compositions include pharmaceutically acceptable salts of the compounds described above. Pharmaceutically acceptable salts are generally well known to those of ordinary skill in the art and include salts of the active compounds prepared from relatively non-toxic acids or bases, depending on the particular substituent moieties present on the compounds described herein. When the compounds of the present disclosure contain relatively acidic functional groups, base addition salts can be obtained by contacting the neutral form of such compounds with a sufficient amount of the desired base, either pure or in a suitable inert solvent, or by ion exchange, whereby one basic counterion (base) in the ion complex is replaced by another. Examples of pharmaceutically acceptable base addition salts include sodium, potassium, calcium, ammonium, organic amino or magnesium salts, or similar salts.

[0195] When the compounds of the present disclosure contain relatively basic functional groups, acid addition salts can be obtained by contacting the neutral form of such compounds with a sufficient amount of the desired acid, either pure or in a suitable inert solvent, or by ion exchange, whereby one acidic counterion (acid) in the ion complex is replaced by another. Examples of pharmaceutically acceptable acid addition salts include those derived from inorganic acids such as hydrochloric acid, hydrobromic acid, nitric acid, carbonic acid, monohydrogencarbonic acid, phosphoric acid, monohydrogenphosphoric acid, dihydrogenphosphoric acid, sulfuric acid, monohydrogensulfuric acid, hydroiodic acid or phosphorous acid, etc., and salts derived from relatively non-toxic organic acids such as acetic acid, propionic acid, isobutyric acid, maleic acid, malonic acid, benzoic acid, succinic acid, suberic acid, fumaric acid, lactic acid, mandelic acid, phthalic acid, benzenesulfonic acid, p-toluenesulfonic acid, citric acid, tartaric acid, methanesulfonic acid, etc. Also included are salts of amino acids such as arginine salts, etc., and salts of organic acids such as glucuronic acid or galacturonic acid, etc. (see, e.g., Berge et al., “Pharmaceutical Salts”, Journal of Pharmaceutical Science, 1977, 66, 1-19). Certain specific compounds of the present disclosure contain both basic and acidic functional groups, which allows the compounds to be converted into base addition salts or acid addition salts.

[0196] Accordingly, pharmaceutically acceptable salts suitable for use with the subject matter disclosed herein include, by way of example and not limitation, acetates, benzenesulfonates, benzoates, bicarbonates, bitartrates, bromides, calcium edetate, camphorsulfonates, carbonates, citrates, edetates, edisylates, estolates, esylates, fumarates, gluceptates, gluconates, glutamates, glycollylarsanilates, hexylresorcinate, hydrabamine, hydrobromides, hydrochlorides, hydroxynaphthoates, iodides, isethionates, lactates, lactobionates, malates, maleates, mandelates, mesylates, mucates, naphthalenesulfonates, nitrates, napsylates, pantothenates, phosphates / diphosphates, pectates, salicylates, stearates, subacetates, succinates, sulfates, tannates, tartrates, or teoclate. Other pharmaceutically acceptable salts can be found, for example, in Remington: The Science and Practice of Pharmacy (20th Edition) Lippincott, Williams & Wilkins (2000).

[0197] In therapeutic and / or diagnostic applications, the compounds of the present disclosure can be formulated for a variety of modes of administration, including systemic administration as well as topical or local administration. Techniques and formulations can generally be found in Remington: The Science and Practice of Pharmacy (20th Edition) Lippincott, Williams & Wilkins (2000).

[0198] Depending on the particular condition being treated, such agents can be formulated as liquid or solid dosage forms and administered systemically or locally. The agents can be delivered, for example, in a timed slow-release or sustained slow-release form known to those of skill in the art. Techniques for formulation and administration can be found in Remington: The Science and Practice of Pharmacy (20th Edition) Lippincott, Williams & Wilkins (2000). Suitable routes can include oral, buccal, by inhalation spray, sublingual, rectal, transdermal, vaginal, transmucosal, nasal, or intestinal administration; parenteral delivery, including intramuscular, subcutaneous, intramedullary injection, and intrathecal, direct intraventricular, intravenous, intra-articular, intrasternal, intrasynovial, intrahepatic, intralesional, intracranial, intraperitoneal, intranasal, or intraocular injection or other delivery modes.

[0199] For injection, the agents of the present disclosure can be formulated and diluted in an aqueous solution, such as in a physiologically compatible buffer such as Hank's solution, Ringer's solution, or saline buffer. For such transmucosal administration, a penetrant suitable for the barrier to be penetrated is used in the formulation. Such penetrants are generally known in the art.

[0200] It is within the scope of the present disclosure to use pharmaceutically acceptable inert carriers to formulate the compounds disclosed herein for practicing the present disclosure into a dosage suitable for systemic administration. With appropriate selection of carriers and suitable manufacturing practices, the compositions of the present disclosure, particularly those formulated as solutions, can be administered parenterally, such as by intravenous injection. The compounds can be readily formulated into a dosage suitable for oral administration using pharmaceutically acceptable carriers well known in the art. Such carriers enable the compounds of the present disclosure to be formulated into tablets, pills, capsules, liquids, gels, syrups, slurries, suspensions, etc. for oral ingestion by the subject to be treated (e.g., a patient).

[0201] For nasal or inhalation delivery, the agents of the present disclosure can also be formulated by methods known to those skilled in the art and can include, for example, but not limited to, examples of solubilizing, diluting, or dispersing substances such as saline; preservatives such as benzyl alcohol; absorption promoters; and fluorocarbons.

[0202] Drug compositions suitable for use in the present disclosure include those in which the active ingredient is included in an effective amount to achieve its intended purpose. Determination of the effective amount is entirely within the ability of those skilled in the art, particularly in light of the detailed disclosure provided herein. Generally, the compounds according to the present disclosure are effective over a wide dosage range. For example, in the treatment of an adult, dosages of from 0.01 mg to 1000 mg per day, from 0.5 mg to 100 mg per day, from 1 mg to 50 mg per day, and from 5 mg to 40 mg per day are examples of dosages that can be used. A non-limiting dosage is from 10 mg to 30 mg per day. The exact dosage will depend on the route of administration, the form in which the compound is administered, the subject to be treated, the weight of the subject to be treated, the bioavailability of the compound, the adsorption, distribution, metabolism, and excretion (ADME) toxicity of the compound, and the preference and experience of the attending physician.

[0203] In addition to the active ingredient, these drug compositions can contain suitable pharmaceutically acceptable carriers that include excipients and adjuvants that aid in processing the active compound into a pharmaceutically useful article. Articles formulated for oral administration can be in the form of tablets, dragees, capsules, or solutions.

[0204] Pharmaceutical preparations for oral use can be obtained by combining the active compound with a solid excipient, optionally grinding the resulting mixture, and processing the granule mixture, if necessary, after adding suitable auxiliaries to obtain tablets or dragee cores. Suitable excipients are, in particular, fillers such as sugars, including lactose, sucrose, mannitol or sorbitol; cellulose products, for example, corn starch, wheat starch, rice starch, potato starch, gelatin, tragacanth, methylcellulose, hydroxypropylmethylcellulose, sodium carboxymethylcellulose (CMC) and / or polyvinylpyrrolidone (PVP: povidone). If necessary, disintegrants such as cross-linked polyvinylpyrrolidone, agar or alginic acid or its salts such as sodium alginate can be added.

[0205] The dragee cores are provided with a suitable coating. For this purpose, concentrated sugar solutions can be used which may optionally contain gum arabic, talc, polyvinylpyrrolidone, carbomer gel, polyethylene glycol (PEG) and / or titanium dioxide, lacquer solutions and suitable organic solvents or solvent mixtures. Dyes or pigments can be added to the tablet or dragee coating for identifying or characterizing different combinations of the active compound dose.

[0206] Pharmaceutical preparations which can be used orally include push-fit capsules made of gelatin, as well as soft, sealed capsules made of gelatin and a plasticizer such as glycerol or sorbitol. The push-fit capsules can contain the active ingredient admixed with a filler such as lactose, a binder such as starch and / or a lubricant such as talc or magnesium stearate and, optionally, a stabilizer. In the soft capsules, the active compound can be dissolved or suspended in a suitable liquid, such as a fatty oil, a liquid paraffin or a liquid polyethylene glycol (PEG). Additionally, stabilizers can be added.

[0207] In still other embodiments, the compounds of formula (I) disclosed herein can be formulated as viscous lotions, creams, ointments, suspensions, pastes, gels, oils, sprays or aerosols and applied topically. Such viscous lotions, creams or ointments can be water-based and can also contain oils (water-in-oil or oil-in-water), such as liquid paraffin or vegetable oils, such as peanut oil or castor oil, or solvents, and can contain one or more other components, including but not limited to penetration enhancers, such as ethanol and propylene glycol, humectants, including but not limited to glycerin and / or glycerol, thickeners and / or gelling agents, including but not limited to soft paraffin, aluminum stearate, cetearyl alcohol, polyethylene glycol, lanolin, beeswax, carboxypolymethylene and cellulose derivatives and / or glyceryl monostearate and / or nonionic emulsifiers, stabilizers, dispersants and suspending agents. By way of example only, common liquid formulations can contain from about 10% to about 60% water, from about 10% to about 70% ethanol, from about 5% to about 10% propylene glycol and from about 2% to about 5% humectant.

[0208] II. Definitions

[0209] Although specific terms are employed herein, they are used in a general and descriptive sense only and not for purposes of limitation. Unless otherwise defined, all technical and scientific terms used herein have the same meaning as commonly understood by one of ordinary skill in the art to which the subject matter described by this invention pertains.

[0210] Although the following terms relating to the compounds of formula (I) are believed to be well understood by one of ordinary skill in the art, the following definitions are presented to facilitate the explanation of the subject matter disclosed herein. These definitions are intended to supplement and illustrate rather than exclude definitions that will be apparent to one of ordinary skill in the art upon review of the present disclosure.

[0211] As used herein, the term substituted (whether or not preceded by the term optionally), and substituents, refer to the ability to change one functional group on a molecule to another functional group, as understood by one of ordinary skill in the art, provided that the valence of all atoms is maintained. When more than one position in any given structure can be substituted with more than one substituent selected from a designated group, the substituents can be the same or different at each position. Substituents can also be further substituted (e.g., an aryl group substituent can have additional substituents distal to it, such as another aryl group, which another aryl group is further substituted at one or more positions).

[0212] In cases where substituent groups or linking groups are designated by their conventional chemical formulas written from left to right, they also encompass chemically identical substituents that would result from writing the structure from right to left. For example, -CH2O- is equivalent to -OCH2-; -C(=O)O- is equivalent to -OC(=O)-; -OC(=O)NR- is equivalent to -NRC(=O)O-, and so on.

[0213] When the term "independently selected from" is used, the substituents being referred to (e.g., R groups such as groups R1, R2, and the like, or variables such as "m" and "n") can be the same or different. For example, both R1 and R2 can be substituted hydrocarbon groups, or R1 can be hydrogen and R2 can be a substituted hydrocarbon group, and so on.

[0214] When the term "a", "an", or "a(n)" is used with respect to groups of substituents herein, it means at least one. For example, in cases where a compound is substituted by "a(n)" hydrocarbon group or aryl group, the compound is optionally substituted by at least one hydrocarbon group and / or at least one aryl group. Additionally, in cases where a moiety is substituted by R substituents, the moiety can be referred to as "R-substituted". In cases where a moiety is R-substituted, the moiety is substituted by at least one R substituent and each R substituent is optionally different.

[0215] The "R" or group named will generally have a structure recognized in the art as corresponding to the group having that name, unless otherwise specified herein. For illustrative purposes, certain representative "R" groups as presented above are defined below.

[0216] The description of the compounds of the present disclosure is limited by the principles of chemical bonding known to those skilled in the art. Thus, in cases where a group can be substituted by one or more of a number of substituents, such substitution is chosen so as to comply with the principles of chemical bonding and to give a compound that is not inherently unstable and / or would be considered likely to be unstable under environmental conditions such as aqueous, neutral, and certain known physiological conditions. For example, a heterocyclic hydrocarbon group or heteroaryl group is attached to the remainder of the molecule via a ring heteroatom, in accordance with the principles of chemical bonding known to those skilled in the art, so as to avoid inherently unstable compounds.

[0217] Unless otherwise explicitly defined, "substituent group" as used herein includes functional groups selected from one or more of the following moieties defined herein:

[0218] As used herein, the term "hydrocarbon" refers to any chemical group containing hydrogen and carbon. The hydrocarbon can be substituted or unsubstituted. As will be known to those skilled in the art, all valences must be satisfied when making any substitution. The hydrocarbon can be unsaturated, saturated, branched, unbranched, cyclic, polycyclic or heterocyclic. Illustrative hydrocarbons are further defined hereinafter and include, for example, methyl, ethyl, n-propyl, isopropyl, cyclopropyl, allyl, vinyl, n-butyl, tert-butyl, ethynyl, cyclohexyl and similar hydrocarbons.

[0219] Unless otherwise specified, the term "alkyl", alone or as part of another substituent, means a straight-chain (i.e., unbranched) or branched, acyclic or cyclic hydrocarbon group or a combination thereof, which can be fully saturated, monounsaturated or polyunsaturated and can include divalent and polyvalent groups, having a specified number of carbon atoms (i.e., C 1-10 means from one to ten carbons, including 1, 2, 3, 4, 5, 6, 7, 8, 9 and 10 carbons). In certain embodiments, the term "alkyl" refers to a C 1-20 (containing 1 and 20) including linear (i.e., "straight-chain"), branched or cyclic, saturated or at least partially unsaturated and in some cases fully unsaturated (i.e., alkenyl and alkynyl) hydrocarbon radicals containing 1, 2, 3, 4, 5, 6, 7, 8, 9, 10, 11, 12, 13, 14, 15, 16, 17, 18, 19 and 20 carbon atoms.

[0220] Representative saturated hydrocarbon groups include, but are not limited to, methyl, ethyl, n-propyl, isopropyl, n-butyl, isobutyl, sec-butyl, tert-butyl, n-pentyl, sec-pentyl, isopentyl, neopentyl, n-hexyl, sec-hexyl, n-heptyl, n-octyl, n-decyl, n-undecyl, dodecyl, cyclohexyl, (cyclohexyl)methyl, cyclopropylmethyl, and homologs and isomers thereof.

[0221] "Branched" refers to a hydrocarbon group in which a lower hydrocarbon group such as methyl, ethyl or propyl is attached to a linear hydrocarbon chain. "Lower hydrocarbon" refers to a hydrocarbon group having from 1 to about 8 carbon atoms such as 1, 2, 3, 4, 5, 6, 7 or 8 carbon atoms (i.e., C 1-8hydrocarbyl). "Higher hydrocarbyl" refers to a hydrocarbyl group having from about 10 to about 20 carbon atoms such as 10, 11, 12, 13, 14, 15, 16, 17, 18, 19 or 20 carbon atoms. In certain embodiments, "hydrocarbyl" particularly refers to C 1-8 linear hydrocarbyl. In other embodiments, "hydrocarbyl" particularly refers to C 1-8 branched hydrocarbyl.

[0222] The hydrocarbyl group may optionally be substituted with one or more hydrocarbyl group substituents ("substituted hydrocarbyl"), and the one or more hydrocarbyl group substituents may be the same or different. The term "hydrocarbyl group substituent" includes, but is not limited to, hydrocarbyl, substituted hydrocarbyl, halogen, arylamino, acyl, hydroxy, aryloxy, hydrocarbyloxy, hydrocarbylthio, arylthio, arylalkyloxy, arylalkylthio, carboxyl, hydrocarbyloxycarbonyl, oxo, and cycloalkyl. One or more oxygen, sulfur, or substituted or unsubstituted nitrogen atoms may optionally be inserted along the hydrocarbyl chain, where the nitrogen substituent is hydrogen, lower hydrocarbyl (also referred to herein as "hydrocarbylamino hydrocarbyl"), or aryl.

[0223] Thus, as used herein, the term "substituted hydrocarbyl" includes a hydrocarbyl group as defined herein, wherein one or more atoms or functional groups in the hydrocarbyl group are replaced by additional atoms or functional groups, and the additional atoms or functional groups include, for example, hydrocarbyl, substituted hydrocarbyl, halogen, aryl, substituted aryl, hydrocarbyloxy, hydroxy, nitro, amino, hydrocarbylamino, dihydrocarbylamino, sulfate, cyano, and mercapto.

[0224] Unless otherwise specified, the term "heterohydrocarbyl", alone or in combination with another term, means a stable straight-chain or branched-chain hydrocarbyl group having from 1 to 20 carbon atoms or heteroatoms, or a cyclic hydrocarbyl group having from 3 to 10 carbon atoms or heteroatoms, or a combination thereof, consisting of at least one carbon atom and at least one heteroatom selected from the group consisting of O, N, P, Si, and S, wherein the nitrogen, phosphorus, and sulfur atoms may optionally be oxidized and the nitrogen heteroatoms may optionally be quaternized. The heteroatoms O, N, P, S, and Si may be located at any internal position of the heterohydrocarbyl group or at the position where the hydrocarbyl group is attached to the remainder of the molecule. Examples include, but are not limited to, -CH2-CH2-O-CH3, -CH2-CH2-NH-CH3, -CH2-CH2-N(CH3)-CH3, -CH2-S-CH2-CH3, -CH2-CH2-S(O)-CH3, -CH2-CH2-S(O)2-CH3, -CH=CH-O-CH3, -Si(CH3)3, -CH2-CH=N-OCH3, -CH=CH-N(CH3)-CH3, O-CH3, -O-CH2-CH3, and -CN. Up to two or three heteroatoms may be consecutive, such as, for example, -CH2-NH-OCH3 and -CH2-O-Si(CH3)3.

[0225] As described above, heterohydrocarbyl groups as used herein include those groups attached to the remainder of the molecule through a heteroatom, such as -C(O)NR', -NR'R", -OR', -SR, -S(O)R, and / or -S(O2)R'. In instances where the term "heterohydrocarbyl" is followed by a recitation of a specific heterohydrocarbyl group (e.g., -NR'R" or a similar group), it will be understood that the terms heterohydrocarbyl and -NR'R" are not redundant or mutually exclusive. Rather, the specific heterohydrocarbyl group is recited for increased clarity. Thus, the term "heterohydrocarbyl" should not be construed herein as excluding specific heterohydrocarbyl groups such as -NR'R" or similar groups.

[0226] "Cyclic" and "cycloalkyl" refer to non-aromatic monocyclic or polycyclic ring systems having from about 3 to about 10 carbon atoms (e.g., 3, 4, 5, 6, 7, 8, 9, or 10 carbon atoms). The cycloalkyl group may optionally be partially unsaturated. The cycloalkyl group may also optionally be substituted with hydrocarbyl group substituents, oxo, and / or alkylene as defined herein. One or more oxygen, sulfur, or substituted or unsubstituted nitrogen atoms may optionally be inserted along the cycloalkyl chain, wherein the nitrogen substituent is hydrogen, unsubstituted hydrocarbyl, substituted hydrocarbyl, aryl, or substituted aryl, thus providing a heterocyclic group. Representative monocyclic cycloalkyl rings include cyclopentyl, cyclohexyl, and cycloheptyl. Polycyclic cycloalkyl rings include adamantyl, octahydronaphthyl, decalin, camphor, bornane, and noradamantyl, and fused ring systems such as dihydronaphthalene and tetrahydronaphthalene and the like.

[0227] Unless otherwise specified, the terms "cycloalkyl" and "heterocycloalkyl" alone or in combination with other terms represent the cyclic forms of "alkyl" and "heteroalkyl", respectively. In addition, for heterocycloalkyl, the heteroatom may occupy the position where the heterocycle is attached to the remainder of the molecule. Examples of cycloalkyl include, but are not limited to, cyclopentyl, cyclohexyl, 1-cyclohexenyl, 3-cyclohexenyl, cycloheptyl, and like groups. Examples of heterocycloalkyl include, but are not limited to, 1-(1,2,5,6-tetrahydropyridyl), 1-piperidyl, 2-piperidyl, 3-piperidyl, 4-morpholinyl, 3-morpholinyl, tetrahydrofuran-2-yl, tetrahydrofuran-3-yl, tetrahydrothiophen-2-yl, tetrahydrothiophen-3-yl, 1-piperazinyl, 2-piperazinyl, and like groups. The terms "cycloalkylene" and "heterocycloalkylene" refer to the divalent derivatives of cycloalkyl and heterocycloalkyl, respectively.

[0228] Unsaturated hydrocarbons have one or more double bonds or triple bonds. Examples of unsaturated hydrocarbyl groups include, but are not limited to, vinyl, 2-propenyl, crotyl, 2-isopentenyl, 2-(butadienyl), 2,4-pentadienyl, 3-(1,4-pentadienyl), ethynyl, 1-propynyl, and 3-propynyl, 3-butynyl, and higher homologs and isomers. An alkyl group that is restricted to a hydrocarbon group is referred to as "homoalkyl".

[0229] More particularly, as used herein, the term "alkenyl" refers to a C having at least one carbon-carbon double bond 2-20A monovalent group derived by removing a single hydrogen molecule from a straight-chain or branched hydrocarbon moiety (containing from 2 to 20 carbon atoms). Alkenyl groups include, for example, ethenyl (i.e., vinyl), propenyl, butenyl, 1-methyl-2-buten-1-yl, pentenyl, hexenyl, octenyl, allene, and butadiene groups.

[0230] As used herein, the term "cycloalkenyl" refers to a cyclic hydrocarbon containing at least one carbon-carbon double bond. Examples of cycloalkenyl groups include cyclopropenyl, cyclobutenyl, cyclopentenyl, cyclopentadiene, cyclohexenyl, 1,3-cyclohexadiene, cycloheptenyl, cycloheptatriene, and cyclooctenyl.

[0231] As used herein, the term "alkynyl" refers to a monovalent group derived from a straight-chain or branched C 2-20 hydrocarbon containing at least one carbon-carbon triple bond and having a designed number of carbon atoms. Examples of "alkynyl" include ethynyl, 2-propynyl (propargyl), 1-propynyl, pentynyl, hexynyl, and heptynyl groups and the like.

[0232] The term "alkylene", alone or as part of another substituent, refers to a straight-chain or branched divalent aliphatic hydrocarbon group derived from a hydrocarbon group having from 1 to about 20 carbon atoms (e.g., 1, 2, 3, 4, 5, 6, 7, 8, 9, 10, 11, 12, 13, 14, 15, 16, 17, 18, 19, or 20 carbon atoms). The alkylene group can be straight-chain, branched, or cyclic. The alkylene group can also optionally be unsaturated and / or substituted with one or more "hydrocarbon group substituents". One or more oxygen, sulfur, or substituted or unsubstituted nitrogen atoms (also referred to herein as "hydrocarbon aminoalkyl") can be optionally inserted along the alkylene group, where the nitrogen substituent is a hydrocarbon as previously described. Exemplary alkylene groups include methylene (-CH2-); ethylene (-CH2-CH2-); propylene (-(CH2)3-); cyclohexylene (-C6H 10 -); -CH=CH-CH=CH-; -CH=CH-CH2-; -CH2CH2CH2CH2-, -CH2CH=CHCH2-, -CH2CsCCH2-, -CH2CH2CH(CH2CH2CH3)CH2-, -(CH2) q -N(R)-(CH2) r-, where q and r are each independently an integer from 0 to about 20, for example, 0, 1, 2, 3, 4, 5, 6, 7, 8, 9, 10, 11, 12, 13, 14, 15, 16, 17, 18, 19 or 20, and R is hydrogen or a lower hydrocarbyl group; methylenedioxy (-O-CH2-O-); and ethylenedioxy (-O-(CH2)2-O-). The hydrocarbylene group can have from about 2 to about 3 carbon atoms and can also have 6 - 20 carbons. Typically, the hydrocarbyl (or hydrocarbylene) group will have from 1 to 24 carbon atoms, and those groups having 10 or fewer carbon atoms are some embodiments of the present disclosure. "Lower hydrocarbyl" or "lower hydrocarbylene" is a shorter-chain hydrocarbyl or hydrocarbylene group that typically has eight or fewer carbon atoms.

[0233] The term "heterohydrocarbylene", alone or as part of another substituent, means a divalent group derived from a heterohydrocarbyl (such as but not limited to -CH2-CH2-S-CH2-CH2- and -CH2-S-CH2-CH2-NH-CH2-). For the heterohydrocarbylene group, the heteroatom can also occupy one or both of the chain termini (such as, for example, hydrocarbyleneoxo, hydrocarbylenedioxy, hydrocarbyleneamino, hydrocarbylenediamino and the like). Still additionally, for the hydrocarbylene and heterohydrocarbylene linking groups, the direction in which the formula of the linking group is written does not imply the orientation of the linking group. For example, the formula -C(O)OR'- represents both -C(O)OR'- and -R'OC(O)-.

[0234] Unless otherwise indicated, the term "aryl" means an aromatic hydrocarbon substituent that can be monocyclic or polycyclic (such as from 1 to 3 rings) fused together or covalently linked. The term "heteroaryl" refers to an aryl group (or ring) containing from one to four heteroatoms selected from N, O, and S (in the case of polycyclic rings in each individual ring), where the nitrogen and sulfur atoms are optionally oxidized and the nitrogen atoms are optionally quaternized. The heteroaryl group can be attached to the remainder of the molecule through a carbon or heteroatom. Non-limiting examples of aryl and heteroaryl groups include phenyl, 1-naphthyl, 2-naphthyl, 4-biphenyl, 1-pyrrolyl, 2-pyrrolyl, 3-pyrrolyl, 3-pyrazolyl, 2-imidazolyl, 4-imidazolyl, pyrazinyl, 2-oxazolyl, 4-oxazolyl, 2-phenyl-4-oxazolyl, 5-oxazolyl, 3-isoxazolyl, 4-isoxazolyl, 5-isoxazolyl, 2-thiazolyl, 4-thiazolyl, 5-thiazolyl, 2-furyl, 3-furyl, 2-thienyl, 3-thienyl, 2-pyridyl, 3-pyridyl, 4-pyridyl, 2-pyrimidinyl, 4-pyrimidinyl, 5-benzothiazolyl, purinyl, 2-benzimidazolyl, 5-indolyl, 1-isoquinolyl, 5-isoquinolyl, 2-quinoxalinyl, 5-quinoxalinyl, 3-quinolyl, and 6-quinolyl. The substituents for each of the above aryl and heteroaryl ring systems are selected from the group of acceptable substituents described below. The terms "arylene" and "heteroarylene" refer to the divalent forms of aryl and heteroaryl, respectively.

[0235] For the sake of brevity, the term "aryl", when used in combination with other terms (e.g., aryloxy, arylthio, arylhydrocarbyl), includes both aryl and heteroaryl rings as defined above. Thus, the terms "arylhydrocarbyl" and "heteroarylhydrocarbyl" are intended to include those groups in which an aryl or heteroaryl group is attached to a hydrocarbyl group (e.g., benzyl, phenethyl, pyridylmethyl, furylmethyl, and the like), and the hydrocarbyl group includes those hydrocarbyl groups in which a carbon atom (e.g., a methylene group) has been replaced, for example, by an oxygen atom (e.g., phenoxymethyl, 2-pyridyloxymethyl, 3-(1-naphthyloxy)propyl, and the like). However, the term "haloaryl" as used herein is intended to include only aryl groups substituted with one or more halogens.

[0236] In the case where a heterohydrocarbyl, heterocyclohydrocarbyl, or heteroaryl includes a specific number of members (e.g., "3-membered to 7-membered"), the term "member" refers to a carbon or heteroatom.

[0237] In addition, as used herein, a structure generally represented by the following formula:

[0238]

[0239] Refers to a ring structure containing a substituent R group, such as, but not limited to, 3-carbon, 4-carbon, 5-carbon, 6-carbon, 7-carbon, etc., aliphatic and / or aromatic cyclic compounds, including saturated ring structures, partially saturated ring structures, and unsaturated ring structures, where the R group may or may not be present, and when present, one or more R groups may each be substituted on one or more available carbon atoms of the ring structure. The presence or absence of the R group and the number of R groups are determined by the value of the variable "n", which is an integer typically having a value in the range from 0 to the number of carbon atoms on the ring available for substitution. Each R group, if more than one, is substituted on the available carbon of the ring structure rather than on another R group. For example, the above structure where n is from 0 to 2 will include a group of compounds that includes, but is not limited to:

[0240]

[0241] and the like.

[0242] The dashed line representing a bond in the cyclic ring structure indicates that the bond may or may not be present in the ring. That is, the dashed line representing a bond in the cyclic ring structure indicates that the ring structure is selected from the group consisting of a saturated ring structure, a partially saturated ring structure, and an unsaturated ring structure.

[0243] The symbol represents the attachment point of the moiety to the remainder of the molecule.

[0244] When a named atom of an aromatic ring or a heteroaromatic ring is defined as "absent", the named atom is replaced by a direct bond.

[0245] As used herein, the term "acyl" refers to an organic acid group in which the -OH of the carboxyl group has been replaced by another substituent and has the general formula RC(=O)-, where R is an alkyl, alkenyl, alkynyl, aryl, carbocyclic, heterocyclic, or heteroaromatic group as defined herein. Thus, the term "acyl" specifically includes arylacyl groups, such as 2-(furan-2-yl)acetyl group and 2-phenylacetyl group. Specific examples of acyl groups include acetyl and benzoyl. The acyl group is also intended to include amides, -RC(=O)NR', esters, -RC(=O)OR', ketones, -RC(=O)R', and aldehydes, -RC(=O)H.

[0246] The terms "alkoxyl" or "alkoxy" are used interchangeably herein and refer to saturated (i.e., alkyl-O-) or unsaturated (i.e., alkenyl-O- and alkynyl-O-) groups attached to the parent molecular moiety through an oxygen atom, where the terms "alkyl", "alkenyl", and "alkynyl" are as previously described and may include C 1-20 (containing 1 and 20) linear, branched or cyclic, saturated or unsaturated oxyhydrocarbon chains, including, for example, methoxy, ethoxy, propoxy, isopropoxy, n-butoxy, sec-butoxy, tert-butoxy and n-pentyloxy, neopentyloxy, n-hexyloxy and similar groups.

[0247] As used herein, the term "alkoxyhydrocarbyl" refers to a hydrocarbyl-O-hydrocarbyl ether, for example, a methoxyethyl or ethoxymethyl group.

[0248] "Aryloxy" refers to an aryl-O- group, where the aryl group is as previously described, including substituted aryl. As used herein, the term "aryloxy" may refer to phenoxy or hexyloxy, and phenoxy or hexyloxy substituted with a hydrocarbyl, substituted hydrocarbyl, halogen, or alkoxy group.

[0249] "Arylhydrocarbyl" refers to an aryl-hydrocarbyl- group, where the aryl and hydrocarbyl are as previously described, and includes substituted aryl and substituted hydrocarbyl. Exemplary arylhydrocarbyl groups include benzyl, phenylethyl, and naphthylmethyl.

[0250] "Arylhydrocarbyloxy" refers to an arylhydrocarbyl-O- group, where the arylhydrocarbyl group is as previously described. An exemplary arylhydrocarbyloxy group is benzyloxy, i.e., C6H5-CH2-O-. The arylhydrocarbyloxy group may be optionally substituted.

[0251] "Alkoxycarbonyl" refers to an alkyl-O-C(=O)- group. Exemplary alkoxycarbonyl groups include methoxycarbonyl, ethoxycarbonyl, butoxycarbonyl, and tert-butoxycarbonyl.

[0252] "Aryloxycarbonyl" refers to an aryl-O-C(=O)- group. Exemplary aryloxycarbonyl groups include phenoxy-carbonyl and naphthyloxy-carbonyl.

[0253] "Arylhydrocarbyloxycarbonyl" refers to an arylhydrocarbyl-O-C(=O)- group. An exemplary arylhydrocarbyloxycarbonyl group is benzyloxycarbonyl.

[0254] "Carbamoyl" refers to an amide group of the formula -C(=O)NH2. "Hydrocarbylcarbamoyl" refers to an R'RN-C(=O)- group, where one of R and R' is hydrogen and the other of R and R' is a hydrocarbyl group and / or a substituted hydrocarbyl group as previously described. "Di(hydrocarbyl)carbamoyl" refers to an R'RN-C(=O)- group, where each of R and R' is independently a hydrocarbyl group and / or a substituted hydrocarbyl group as previously described.

[0255] As used herein, the term carbonyldioxy refers to a carbonate group of the formula -O-C(=O)-OR.

[0256] "Acyl-oxy" refers to an acyl-O- group, where the acyl group is as previously described.

[0257] The term "amino" refers to the -NH2 group and also refers to a nitrogen-containing group known in the art that is derived from ammonia by replacing one or more hydrogen groups with organic groups. For example, the terms "acylamino" and "hydrocarbylamino" refer to specific N-substituted organic groups having acyl and hydrocarbyl substituents, respectively.

[0258] As used herein, "aminohydrocarbyl" refers to an amino group covalently attached to a hydrocarbylene linking group. More particularly, as used herein, the terms hydrocarbylamino, di(hydrocarbyl)amino, and tri(hydrocarbyl)amino refer to one, two, or three hydrocarbyl groups as previously defined that are attached to the parent molecular moiety via a nitrogen atom, respectively. The term hydrocarbylamino refers to a group having the structure -NHR', where R' is a hydrocarbyl group as previously defined; and the term di(hydrocarbyl)amino refers to a group having the structure -NR'R", where R' and R" are each independently selected from the group consisting of hydrocarbyl groups. The term tri(hydrocarbyl)amino refers to a group having the structure -NR'R"R"', where R', R", and R"' are each independently selected from the group consisting of hydrocarbyl groups. In addition, R', R", and / or R"' taken together may optionally be -(CH2) k -, where k is an integer from 2 to 6. Examples include, but are not limited to, methylamino, dimethylamino, ethylamino, diethylamino, diethylaminocarbonyl, methylethylamino, isopropylamino, piperidino, trimethylamino, and propylamino.

[0259] The amino group is -NR'R", where R' and R" are typically selected from hydrogen, a substituted or unsubstituted hydrocarbyl group, a substituted or unsubstituted heterohydrocarbyl group, a substituted or unsubstituted cyclo-hydrocarbyl group, a substituted or unsubstituted heterocyclo-hydrocarbyl group, a substituted or unsubstituted aryl group, or a substituted or unsubstituted heteroaryl group.

[0260] The terms hydrocarbyl thioether and thiohydrocarbyloxy refer to saturated (i.e., alkyl-S-) or unsaturated (i.e., alkenyl-S- and alkynyl-S-) groups attached to the parent molecular moiety through a sulfur atom. Examples of thiohydrocarbyloxy moieties include, but are not limited to, methylthio, ethylthio, propylthio, isopropylthio, n-butylthio and like groups.

[0261] "Acylamino" refers to an acyl-NH- group, where acyl is as previously described. "Aroylamino" refers to an aroyl-NH- group, where aroyl is as previously described.

[0262] The term "carbonyl" refers to a -C(=O)- group and may include aldehyde groups represented by the general formula R-C(=O)H.

[0263] The term "carboxyl" refers to a -COOH group. Such a group is also referred to herein as a "carboxylic acid" moiety.

[0264] The term "cyano" refers to a -C≡N group.

[0265] As used herein, the terms "halo", "halide" or "halogen" refer to fluorine, chlorine, bromine and iodine groups. Further, terms such as "halohydrocarbyl" are intended to include both monohalohydrocarbyl and polyhalohydrocarbyl. For example, the term "halo(C 1-4 )hydrocarbyl" is intended to include, but is not limited to, trifluoromethyl, 2,2,2-trifluoroethyl, 4-chlorobutyl, 3-bromopropyl and like groups.

[0266] The term "hydroxy" refers to an -OH group.

[0267] The term "hydroxyhydrocarbyl" refers to a hydrocarbyl group substituted with an -OH group.

[0268] The term "mercapto" refers to a -SH group.

[0269] As used herein, the term "oxo" means an oxygen atom double-bonded to a carbon atom or another element.

[0270] The term "nitro" refers to a -NO2 group.

[0271] The term "thio" refers to compounds previously described herein where a carbon atom or an oxygen atom is replaced by a sulfur atom.

[0272] The term "sulfate" refers to a -SO4 group.

[0273] As used herein, the term thiohydroxy or thiol, refers to a group of the formula -SH.

[0274] More particularly, the term "sulfide" refers to a compound having a group of the formula -SR.

[0275] The term "sulfone" refers to a compound having a sulfonyl group -S(O2)R.

[0276] The term "sulfoxide" refers to a compound having a sulfinyl group -S(O)R.

[0277] The term ureido refers to a ureido group of the formula -NH—CO—NH2.

[0278] Throughout the specification and claims, a given chemical formula or name will encompass all tautomers, homologs, and optical and stereoisomers, as well as racemic mixtures (when such isomers and mixtures exist).

[0279] Certain compounds of the present disclosure may have asymmetric carbon atoms (optical or chiral centers) or double bonds; enantiomers, racemates, diastereomers, tautomers, geometric isomers, stereoisomeric forms, and individual isomers that can be defined as (R)- or (S)- or, for amino acids, as D- or L- according to absolute stereochemistry are included within the scope of the present disclosure. The present disclosure is intended to include compounds in racemic, scalemic, and optically pure forms. The optically active (R)- and (S)-isomers or D- and L-isomers can be prepared using chiral synthons or chiral reagents or resolved using conventional techniques. When the compounds described herein contain double bonds or other centers of geometric asymmetry, and unless otherwise specified, it is intended that the compounds include both E geometric isomers and Z geometric isomers.

[0280] Unless otherwise stated, the structures depicted herein also intend to include all stereochemical forms of the structure; that is, the R and S configurations for each asymmetric center. Accordingly, the individual stereoisomers of the compounds of the present invention, as well as enantiomeric and diastereomeric mixtures, are within the scope of the present disclosure.

[0281] It will be apparent to those skilled in the art that certain compounds of the present disclosure may exist in tautomeric forms, and all such tautomeric forms of the compounds are within the scope of the present disclosure. As used herein, the term "tautomer" refers to one of two or more structural isomers that exist in equilibrium and are readily convertible from one isomeric form to another.

[0282] Unless otherwise stated, the structures depicted herein also intend to include compounds that differ only in the presence of one or more isotopically enriched atoms. For example, compounds having the structure of the present invention with deuterium or tritium replacing hydrogen or carbon replaced by 13 C or 14 C-enriched carbon are within the scope of the present disclosure.

[0283] The compounds of the present disclosure may also contain unnatural proportions of atomic isotopes at one or more of the atoms that make up such compounds. For example, the compounds may be radiolabeled with radioactive isotopes such as, for example, tritium ( 3 H), iodine-125 ( 125 I), or carbon-14 ( 14 C). All isotopic variants of the compounds of the present disclosure, whether radioactive or not, are included within the scope of the present disclosure.

[0284] Certain compounds of the present disclosure may exist in unsolvated forms as well as solvated forms (including hydrated forms). In general, the solvated forms are equivalent to the unsolvated forms and are included within the scope of the present disclosure. Certain compounds of the present disclosure may exist in multiple crystalline or amorphous forms. In general, all physical forms are equivalent for the intended uses of the present disclosure and are intended to be within the scope of the present disclosure.

[0285] The term "protecting group" refers to a chemical moiety that blocks some or all of the reactive moieties of a compound and prevents such moieties from participating in chemical reactions until the protecting group is removed, for example, those moieties listed and described in: T.W. Greene, P.G.M. Wuts, Protective Groups in Organic Synthesis, 3rd Edition John Wiley & Sons (1999). In cases where different protecting groups are used, it may be advantageous for each (different) protecting group to be removable in a different manner. Protecting groups that cleave under completely different reaction conditions allow for the differential removal of such protecting groups. For example, protecting groups can be removed by acids, bases, and hydrogenolysis. Groups such as trityl, dimethoxytrityl, acetals, and tert-butyldimethylsilyl are acid-labile and can be used to protect carboxyl and hydroxyl reactive moieties in the presence of amino groups protected with Cbz groups (which are removable by hydrogenolysis) and Fmoc groups (which are base-labile). Carboxylic acid and hydroxyl reactive moieties can be blocked with base-labile groups such as, but not limited to, methyl, ethyl, and acetyl in the presence of acid-labile groups such as tert-butyl carbamate or amines blocked with carbamates that are stable to both acids and bases but removable by hydrolysis.

[0286] The carboxylic acid and hydroxy-reactive moieties may also be blocked with a hydrolyzable protecting group such as a benzyl group, while amine groups capable of hydrogen bonding to an acid may be blocked with a base-labile group such as Fmoc. The carboxylic acid-reactive moiety may be blocked with an oxidizable protecting group such as 2,4-dimethoxybenzyl, while co-existing amino groups may be blocked with a fluoride-labile silyl carbamate.

[0287] Allyl blocking groups are useful in the presence of acid and base protecting groups, as the former is stable and can subsequently be removed by a metal or pi-acid catalyst. For example, allyl-blocked carboxylic acids can be deprotected with a palladium(0)-catalyzed reaction in the presence of an acid-labile tert-butyl carbamate or a base-labile acetate amine protecting group. Another form of a protecting group is a resin to which a compound or intermediate can be attached. So long as the residue is attached to the resin, the functional group is blocked and cannot react. After release from the resin, the functional group is available for reaction.

[0288] Typical blocking / protecting groups include, but are not limited to, the following moieties: p-methoxybenzylcarbonyl (Moz or MeOZ), 3,4-dimethoxybenzyl (DMPM), Troc (trichloroethyl chloroformate), (4-nitrophenyl)sulfonyl (Nosyl), and nitrophenylthio (Nps), and

[0289]

[0290] Following long-standing patent law convention, the terms "a", "an", and "the" when used in this application (including the claims) refer to "one or more". Thus, for example, reference to "a subject" includes more than one subject unless the context clearly dictates otherwise (e.g., more than one subject), and so forth.

[0291] Throughout this specification and the claims, the terms "comprise", "comprises", and "comprising" are used in a non-exclusive sense unless the context requires otherwise. Similarly, the term "include" and its grammatical variants are intended to be non-limiting, such that the recitation of items in a list does not exclude other similar items that may be substituted or added to the listed items.

[0292] For the purposes of this specification and the appended claims, unless otherwise indicated, all numbers expressing quantities, dimensions, measurements, ratios, shapes, formulations, parameters, percentages, amounts, properties and other numerical values used in the specification and claims shall be understood to be modified in all instances by the term "about", even if the term "about" may not expressly appear with the value, amount or range. Accordingly, unless indicated to the contrary, the numerical parameters set forth in the following specification and appended claims are not and need not be exact, but may be approximate and / or larger or smaller as desired, reflecting tolerances, conversion factors, rounding, measurement error, and the like, as well as other factors known to those of skill in the art, depending upon the desired properties sought to be obtained by the subject matter disclosed herein. For example, when referring to a value, the term "about" can be intended to include differences of, in some embodiments, ±100%, in some embodiments ±50%, in some embodiments ±20%, in some embodiments ±10%, in some embodiments ±5%, in some embodiments ±1%, in some embodiments ±0.5%, and in some embodiments ±0.1% from the specified amount, as such differences are appropriate for carrying out the disclosed methods or using the disclosed compositions.

[0293] In addition, when used in conjunction with one or more numerical values or numerical ranges, the term "about" shall be understood to refer to all such numerical values, including all values within the range and modifying the range by extending the boundaries above and below the listed values. A numerical range stated by endpoints includes all values subsumed within that range, e.g., all integers, including their fractions (e.g., a statement of 1 to 5 includes 1, 2, 3, 4, and 5, as well as their fractions, e.g., 1.5, 2.25, 3.75, 4.1, etc.) and any range within that range. Examples

[0294] The following examples have been included to provide guidance to those of ordinary skill in the art for practicing representative embodiments of the subject matter disclosed herein. Based on the present disclosure and the general level of skill in the art, one of ordinary skill in the art will understand that the following examples are intended to be merely exemplary, and that many variations, modifications, and alterations can be employed without departing from the scope of the subject matter disclosed herein. The following descriptions of synthesis and specific examples are intended for illustrative purposes only and are not to be construed as limiting the preparation of the compounds of the present disclosure by any other method.

[0295] Example 1

[0296] Stability of prodrugs in mouse skin homogenate and plasma

[0297] Mouse skin homogenate stability was performed using washed mouse skin or CD1 mouse plasma diluted 10-fold in 0.1 M potassium phosphate buffer and homogenized using a tissue homogenizer. To evaluate the stability of the intact prodrug, 1 mL aliquots of skin homogenate or plasma were spiked with the prodrug to a final assay concentration of 20 μM. The spiked skin homogenate and plasma samples were incubated at 37 °C in an orbital shaker for 1 h and then quenched in triplicate with three volumes of acetonitrile containing an internal standard (IS; losartan: 0.5 μM). The samples were vortexed for 30 s and centrifuged at 10,000 × g for 10 min at 4 °C. 50 μL of the supernatant was diluted with 50 μL of water and transferred to a 250 μL polypropylene vial sealed with a Teflon cap. Release of itaconic acid or methyl itaconate was measured by liquid chromatography tandem mass spectrometry (LC-MS / MS).

[0298] Example 2

[0299] Pharmacokinetics in mice

[0300] Male CD1 mice (25 g - 30 g) were obtained from Harlan and maintained on a 12 h light-dark cycle with ad libitum access to food and water. IS-100-142, MK939, and MK941 were administered via oral gavage at a molar equivalent dose of 10 mg / kg of methyl itaconate. Blood samples (n = 3 per time point) were collected at 0.25 h and 1 h after dosing. At the designated time points after drug administration, the mice were euthanized with carbon dioxide and blood samples (~0.8 mL) were collected by cardiac puncture into heparinized microtubes. The blood samples were centrifuged at 3000 × g for 10 min at a temperature of 4 °C. Plasma samples (~300 μL) were collected in polypropylene tubes and stored at -80 °C until bioanalysis. Calibration standards were prepared using plasma from the first-in-experiment ( ) mice spiked with methyl itaconate. Methyl itaconate standards and samples were extracted from plasma by one-step protein precipitation using methanol (100% v / v) containing the internal standard dimethyl succinate (5 μM). Aliquots of the supernatant (100 μL) were diluted with water (100 μL) and transferred to a 250 μL polypropylene vial sealed with a Teflon cap and analyzed via LC-MS / MS.

[0301] Example 3

[0302] Treatment of human keratinocytes with itaconate prodrugs

[0303] Neonatal human epidermal keratinocytes (NHEK) isolated from neonatal foreskins were seeded at a density of 100,000 cells per well and maintained in KGM supplemented with growth factors (KGM-GOLD Bullet Kit, #192060). The prodrug was reconstituted in DMSO. NHEK were pretreated with vehicle (0.1% DMSO) or the prodrug. After 2 days, NHEK were treated with 50 μg / μl poly(I:C) for 24 h.

[0304] Example 4

[0305] RNA isolation and quantitative real-time PCR

[0306] Total RNA was isolated and purified from cultured NHEK using the RNeasy Mini Kit (Qiagen, Valencia, CA, #74106). After assessing RNA purity and concentration using a NanoDrop 2000 UV-Vis spectrophotometer, RNA was converted to cDNA using a reverse transcription kit and random hexamer primers (Applied Biosystems, #4368814). mRNA expression was determined by qRT-PCR using gene-specific, fluorophore-based TaqMan probes and a Universal Master Mix (Applied Biosystems, #4366072). The qRT-PCR reactions were multiplexed using the target probe and a reference gene (RPLP0) probe. Relative fold changes in mRNA were then quantified using the ΔΔCt method.

[0307] Example 5

[0308] Representative compounds

[0309] (S)-4-((1-Ethoxy-1-oxo-3-phenylpropan-2-yl)amino)-2-methylidene-4-oxobutanoic acid (IS-100-127)

[0310]

[0311] Itaconic anhydride (50 mg, 0.45 mmol) was dissolved in anhydrous THF (5 mL), and solid potassium carbonate (0.19 g, 1.34 mmol) was added followed by L-phenylalanine ethyl ester hydrochloride (0.1 g, 0.45 mmol). The reaction mixture was stirred at room temperature for 16 h. The volatiles were then evaporated, the residue was redissolved in DCM (30 mL), and extracted with 1 M aqueous HCl (10 mL) and brine (10 mL). The organic phase was dried over Na2SO4, the volatiles were evaporated, and the residue was subjected to final purification on reverse-phase HPLC to afford 100 mg (73%) of the desired compound as a colorless semi-solid.

[0312] 1 H NMR (401 MHz, DMSO-d6): δ H 1.25 (t, J = 7.1 Hz, 3H), 3.13 (dd, J = 13.9, 5.9 Hz, 2H), 3.28 (s, 2H), 4.18 (q, J = 7.2 Hz, 2H), 4.86 (dt, J = 7.8, 6.0 Hz, 1H), 5.90 (s, 1H), 6.46 (s, 1H), 6.65 (d, J = 8.0 Hz, 1H), 7.06–7.38 (m, 5H), 9.98 (s, 1H).

[0313] ESIMS: 304.1 ([M-H] + )。

[0314] HRMS (ESI): For C 16 H 18 O5N calcd 304.11905. Found: 304.11910.

[0315] (S)-Methyl 3-((1-ethoxy-1-oxo-3-phenylpropan-2-yl)carbamoyl)but-3-enoate (IS-100-128)

[0316]

[0317] β-Methyl itaconate (50 mg, 0.35 mmol) and HATU (0.13 g, 0.34 mmol) were dissolved in anhydrous DMF (3 mL), N,N-Diisopropylethylamine (0.12 mL, 0.69 mmol) was added and the mixture was stirred at room temperature for 5 minutes. L-Phenylalanine ethyl ester hydrochloride (80 mg, 0.35 mmol) was added and the reaction mixture was stirred at room temperature for 3 hours. Then the volatiles were evaporated, the residue was redissolved in DCM (30 mL) and extracted with 1 M aqueous HCl (10 mL) and brine (10 mL). The organic phase was dried over Na2SO4, the volatiles were evaporated and the residue was subjected to flash column chromatography (silica gel 60 mesh 70 - 230, solvent: cyclohexane / ethyl acetate 2:1) to afford 100 mg (90%) of compound IS-100-128 as a colorless semi-solid.

[0318] 1 H NMR (401 MHz, DMSO-d6): δ H 1.27 (t, J = 7.1 Hz, 3H), 3.19 (dd, J = 5.7, 4.6 Hz, 2H), 3.37 (d, J = 1.1 Hz, 2H), 3.69 (s, 3H), 4.20 (q, J = 7.1 Hz, 2H), 4.90 (dt, J = 7.6, 5.7 Hz, 1H), 5.50 (t, J = 1.2 Hz, 1H), 5.74 (s, 1H), 6.60 (d, J = 7.6 Hz, 1H), 7.09–7.19 (m, 2H), 7.21–7.36 (m, 3H).

[0319] 13 C NMR (101 MHz, CDCl3): δ C 12.76, 36.46, 36.53, 50.77, 52.04, 60.20, 120.63, 125.72, 127.13, 128.06, 134.53, 136.59, 165.52, 169.86, 170.01.

[0320] ESIMS: 342.1 ([M+Na] + )。

[0321] HRMS (ESI): For C 17 H 22 O5N calcd 320.14925. Found: 320.14892.

[0322] (S)-Methyl 3-((1-amino-1-oxo-3-phenylpropan-2-yl)carbamoyl)but-3-enoate (IS-100-129)

[0323]

[0324] β-Methyl itaconate (70 mg, 0.49 mmol) and HATU (0.18 g, 0.49 mmol) were dissolved in anhydrous DMF (5 mL), N,N-Diisopropylethylamine (0.17 mL, 0.97 mmol) was added and the mixture was stirred at room temperature for 5 minutes. L-Phenylalanine amide hydrochloride (97 mg, 0.49 mmol) was added and the reaction mixture was stirred at room temperature for 3 hours. Then the volatiles were evaporated, the residue was redissolved in DCM (30 mL), and extracted with 1 M aqueous HCl (10 mL) and brine (10 mL). The organic phase was dried over Na2SO4, the volatiles were evaporated, and the residue was subjected to flash column chromatography (silica gel 60 mesh 70 - 230, solvent: dichloromethane / methanol 20:1) to afford 120 mg (85%) of compound IS-100-129 as a colorless semi-solid.

[0325] 1 H NMR (401 MHz, DMSO-d6): δ H 3.14–3.31 (m, 3H), 3.46–3.56 (m, 1H), 3.68 (s, 3H), 4.77 (dt, J = 8.0, 6.8 Hz, 1H), 5.43 (s, 1H), 5.57–5.63 (m, 1H), 5.67 (s, 1H), 6.38 (s, 1H), 6.67 (d, J = 8.0 Hz, 1H), 7.21–7.38 (m, 5H).

[0326] 13 C NMR (101 MHz, CDCl3): δ C 37.45, 38.05, 52.24, 54.01, 121.42, 127.11, 128.76, 129.32, 136.60, 138.22, 167.87, 171.74, 173.19.

[0327] ESIMS: 313.1 ([M+Na] + )

[0328] HRMS (ESI): Calcd for C 15 H 18 O4N2Na 313.11588. Found: 313.11545.

[0329] 1-(((Isopropoxycarbonyl)oxy)methyl) 4-methyl 2-methylenesuccinate (IS-100-142)

[0330]

[0331] β-Methyl itaconate (0.2 g, 1.39 mmol), isopropyl(chloromethyl) carbonate (0.22 mL, 1.66 mmol), and potassium carbonate (0.29 g, 2.08 mmol) were dissolved in anhydrous MeCN (5 mL), and the mixture was stirred at room temperature for 16 h. EtOAc (60 mL) was added and the mixture was washed with brine (20 mL). The organic phase was dried over Na2SO4, the volatiles were evaporated, and the residue was subjected to flash column chromatography (silica gel 60 mesh 70 - 230, solvent: cyclohexane / ethyl acetate 5:1) to afford 90 mg (25%) of compound IS-100-142 as a colorless oil.

[0332] 1 H NMR (401 MHz, DMSO-d6): δ H 1.34 (d, J = 6.3 Hz, 6H), 3.38 (s, 2H), 3.72 (s, 3H), 4.86–5.02 (m, 1H), 5.82–5.88 (m, 3H), 6.46 (s, 1H).

[0333] 13 C NMR (101 MHz, CDCl3): δ C 21.64, 37.20, 52.12, 73.10, 82.20, 130.50, 132.76, 153.29, 164.52, 170.77.

[0334] ESIMS: 283.1 ([M+Na] + )。

[0335] HRMS (ESI): Calcd for C 11 H 16 O7Na 283.07882. Found: 283.07855.

[0336] 4-Methyl 1-((pivaloyloxy)methyl) 2-methylenesuccinate (IS-100-143)

[0337]

[0338] β-Methyl itaconate (0.2 g, 1.39 mmol), chloro(pivaloyloxy)methane (0.26 mL, 1.8 mmol), sodium iodide (50 mg, 0.33 mmol), and potassium carbonate (0.29 g, 2.08 mmol) were dissolved in anhydrous MeCN (5 mL), and the mixture was stirred at 40 °C for 16 h. EtOAc (60 mL) was added and the mixture was washed with brine (20 mL). The organic phase was dried over Na2SO4, the volatiles were evaporated, and the residue was subjected to flash column chromatography (silica gel 60 mesh 70 - 230, solvent: cyclohexane / ethyl acetate 5:1) to afford 0.33 g (92%) of compound IS-100-143 as a colorless oil.

[0339] 1 H NMR (401 MHz, DMSO-d6): δ H 1.20 (s, 9H), 3.34 (s, 2H), 3.68 (s, 3H), 5.79 (d, J = 1.1 Hz, 1H), 5.82 (s, 2H), 6.39 (s, 1H).

[0340] 13 C NMR (101 MHz, CDCl3): δ C 26.95, 37.38, 38.90, 52.21, 79.91, 130.30, 133.06, 164.82, 170.90, 177.19.

[0341] ESIMS: 281.1 ([M+Na] + )

[0342] HRMS (ESI): Calcd for C 12 H 18 O6Na 281.09956. Found: 281.09921.

[0343] 1-(tert-Butyl) 4-((pivaloyloxy)methyl) 2-methylenesuccinate (IS-100-146)

[0344]

[0345] 3-(tert-Butoxycarbonyl)but-3-enoic acid (0.2 g, 1.07 mmol), chloromethyl pivalate (0.2 mL, 1.4 mmol), sodium iodide (30 mg, 0.21 mmol), and potassium carbonate (0.22 g, 1.61 mmol) were dissolved in anhydrous MeCN (4 mL), and the mixture was stirred at 45 °C for 16 h. EtOAc (60 mL) was added and the mixture was washed with brine (20 mL). The organic phase was dried over Na2SO4, the volatiles were evaporated, and the residue was subjected to flash column chromatography (silica gel 60 mesh 70 - 230, solvent: cyclohexane / ethyl acetate 6:1) to afford 0.23 g (71%) of compound IS-100-143 as a colorless oil.

[0346] 1 H NMR (401 MHz, DMSO-d6): δ H 1.21 (s, 9H), 1.48 (s, 9H), 3.33 (d, J = 1.2 Hz, 2H), 5.63 (d, J = 1.2 Hz, 1H), 5.76 (s, 2H), 6.25 (d, J = 1.0 Hz, 1H).

[0347] 13 C NMR (101 MHz, CDCl3): δ C 27.00, 28.10, 37.72, 38.90, 79.88, 81.44, 127.92, 134.86, 165.13, 169.79, 177.23.

[0348] ESIMS: 323.2 ([M+Na] + )

[0349] HR ESIMS: Calcd for C 15 H 24 O6Na 323.14651. Found: 323.14622.

[0350] 2-Methylene-4-oxo-4-((pivaloyloxy)methoxy)butanoic acid (IS-100-147)

[0351]

[0352] 1-(tert-Butyl) 4-((pivaloyloxy)methyl) 2-methylenesuccinate (0.14 g, 0.47 mmol) was dissolved in anhydrous DCM (0.5 mL) and trifluoroacetic acid (4 mL), and the mixture was stirred at room temperature for 2 hours. The volatiles were evaporated, and the residue was dissolved in DCM (3 × 15 mL) and evaporated three times to afford 0.11 g (97%) of compound IS-100-147 as a colorless oil.

[0353] 1 H NMR (401 MHz, DMSO-d6): δ H 1.21 (s, 9H), 3.37 (s, 2H), 5.77 (s, 2H), 5.87 (s, 1H), 6.49 (s, 1H), 11.12 (s, 1H).

[0354] 13 C NMR (101 MHz, CDCl3): δ C 26.81, 36.97, 38.76, 79.66, 131.36, 132.54, 169.22, 171.11, 177.13.

[0355] ESIMS: 267.1 ([M+Na] + )

[0356] HRMS (ESI): Calculated for C 11 H 16 O6Na 267.08391. Found: 267.08375.

[0357] Bis((pivaloyloxy)methyl) 2-methylenesuccinate (IS-100-148)

[0358]

[0359] Itaconic acid (0.2 g, 1.53 mmol), pivaloyl chloride methyl ester (0.55 mL, 3.84 mmol), sodium iodide (50 mg, 0.3 mmol), and potassium carbonate (0.64 g, 4.60 mmol) were dissolved in anhydrous MeCN (5 mL), and the mixture was stirred at 45 °C for 16 hours. EtOAc (60 mL) was added and the mixture was washed with brine (20 mL). The organic phase was dried over Na2SO4, the volatiles were evaporated, and the residue was subjected to flash column chromatography (silica gel 60 mesh 70 - 230, solvent: cyclohexane / ethyl acetate 5:1) to afford 0.20 g (36%) of compound IS-100-148 as a colorless oil.

[0360] 11H NMR (401 MHz, DMSO-d6): δ H 1.20 (s, 18H), 3.38 (d, J = 1.0 Hz, 2H), 5.74 (s, 2H), 5.82 (s, 3H), 6.41 (s, 1H).

[0361] 13 13C NMR (101 MHz, CDCl3): δ C 26.83, 26.84, 37.12, 38.75, 38.78, 79.70, 79.82, 130.52, 132.33, 164.45, 169.12, 177.07.

[0362] ESIMS: 381.2 ([M+Na] + )

[0363] HRMS (ESI): For C 17 H 26 O8Na calcd 381.15199. Found: 381.15158.

[0364] Bis(((isopropoxycarbonyl)oxy)methyl) 2-methylenesuccinate (IS-100-149)

[0365]

[0366] Itaconic acid (0.2 g, 1.53 mmol), chloromethyl isopropyl carbonate (0.51 mL, 3.84 mmol), sodium iodide (50 mg, 0.3 mmol) and potassium carbonate (0.64 g, 4.60 mmol) were dissolved in anhydrous MeCN (5 mL), and the mixture was stirred at 45 °C for 16 h. EtOAc (60 mL) was added and the mixture was washed with brine (20 mL). The organic phase was dried over Na2SO4, the volatiles were evaporated, and the residue was subjected to flash column chromatography (silica gel 60 mesh 70–230, solvent: cyclohexane / ethyl acetate 5:1) to afford 80 mg (14%) of compound IS-100-149 as a colorless oil.

[0367] 1 1H NMR (401 MHz, DMSO-d6): δ H 1.31 (d, J = 6.3 Hz, 12H), 3.41 (s, 2H), 4.82–4.98 (m, 2H), 5.75 (s, 2H), 5.82 (s, 2H), 5.85 (s, 1H), 6.46 (s, 1H).

[0368] 1313C NMR (101 MHz, CDCl3): δ C 21.63, 36.95, 73.13, 82.00, 82.22, 131.05, 131.99, 153.26, 153.29, 164.28, 168.95。

[0369] ESIMS: 385.1 ([M+Na] + )。

[0370] HRMS (ESI): For C 15 H 22 O 10 Na calcd for 385.11052. Found: 385.11069.

[0371] 4-Methyl-1-((5-methyl-2-oxo-1,3-dioxolan-4-yl)methyl)-2-methylene-succinate (LTP1025)

[0372]

[0373] β-Methyl itaconate (200 mg, 1.39 mmol) was dissolved in anhydrous MeCN (5 mL), K2CO3 (384 mg, 2.78 mmol, 2 equiv) and NaI (416 mg, 2.78 mmol, 2 equiv) were added and the resulting mixture was heated at 40 °C for 10 min under an inert atmosphere. (4-Chloromethyl)-5-methyl-1,3-dioxolane-2-one (412 mg, 303 μL, 2.78 mmol, 2 equiv) was finally added and the mixture was heated at 40 °C for 48 h. The MeCN was evaporated, EtOAc (50 mL) was added and the organic phase was washed with 10% Na2S2O5 (50 mL), distilled H2O (50 mL) and saturated NaCl (2 × 50 mL). The organic phase was dried over MgSO4, the volatiles were evaporated and the residue was subjected to flash column chromatography (silica gel 60 mesh 70–230, solvent: cyclohexane / EtOAc, 2:1) to afford 320 mg (90%) of compound LTP1025 as a pale yellow oil.

[0374] 1 1H NMR (401 MHz, CDCl3): δ H 2.15 (s, 3H), 3.31 (s, 2H), 3.65 (s, 3H), 4.89 (s, 2H), 5.75 (s, 1H), 6.33 (s, 1H).

[0375] 13 13C NMR (101 MHz, CDCl3): δ C9.36, 37.36, 52.12, 54.28, 129.98, 132.99, 133.36, 140.30, 152.11, 165.51, 170.91。

[0376] ESIMS: 279.1 ([M+Na] + )。

[0377] HRMS(ESI): For C 11 H 12 O7Na, calculated value 279.04752. Found: 279.04757.

[0378] Hydrocarbyl β-methylitaconate

[0379]

[0380] R = hydrocarbyl, aryl, hydrocarbylsalicyl, Boc-tyrosylhydrocarbylamide, Boc-tyrosylester, (di)peptide with terminal tyrosylester, (di)peptide with terminal tyrosylhydrocarbylamide

[0381]

[0382] Examples of R-OH of hydrocarbyl β-methylitaconate include, but are not limited to:

[0383] The general procedure was carried out according to the modified method described in the following: Boschert, D.; Schneider-Chaabane, A.; Himmelsbach, A.; Eickenscheidt, A.; Lienkamp, K. Synthesis and Bioactivity of Polymer-Based Synthetic Mimics of Antimicrobial Peptides (SMAMPs) Made from Asymmetrically Disubstituted Itaconates. Chem. Eur. J. 2018, 24, 8217 - 8227.

[0384] Synthesis of L - tyrosylalkylamides according to the following: McKenna, C.E.; Kashemirov, B.A.; Krylov, I.S.; Zakharova, V.M. Method to improve antiviral activity of nucleotide analogue drugs. US9550803 B2, January 24, 2017; and Hidaka, K.; Gohda, K.; Teno, N.; Wanaka, K.; Tsuda, Y. Active site - directed plasmin inhibitors: Extension on the P2 residue. Bioorg. Med. Chem. 2016, 24, 545 - 553.

[0385] Dissolve β - methyl itaconate (1 mmol) in dry dichloromethane (3 mL). Add the appropriate hydroxy - derivative (1.2 mmol) and DMAP (1.5 mmol), and cool the solution to 0 °C. Dropwise add a solution of DCC (1.5 mmol) in dichloromethane (2 mL) under nitrogen, and stir the whole mixture at 0 °C for 1 h, and then stir overnight at room temperature. Filter the mixture, and extract the solution with 10% aqueous KHSO4 (3×5 mL) and saturated NaHCO3 (5 mL). Dry the organic phase over sodium sulfate, evaporate, and chromatograph the residue on a silica gel column in the system described below.

[0386] 1 - (2 - (hexadecyloxy)ethyl) 4 - methyl 2 - methylenesuccinate (MK - 933)

[0387]

[0388] Chromatography in a cyclohexane - ethyl acetate (80:15) system. Yield: 253 mg (61%) of a colorless liquid.

[0389] 1 H NMR (CDCl3, ppm) δ: 0.88 (t, 3H, J CH3,CH2 = 7.1, CH3(18′)), 1.38 - 1.20 (m, 26H, 13xCH2(5′ - 17′)), 1.57 (m, 2H, CH2(4′)), 3.35 (d, 2H, J 2,＝C H = 1.1, CH2(2)), 3.46 (t, 2H, J 3′,4′= 6.7, OCH2(3′)), 3.65(m, 2H, OCH2(2′)), 3.69(s, 3H, OCH3), 4.30(m, 2H, OCH2(1′)), 5.73(q, 1H, J gem = J H,2 = 1.1, =C H a ), 6.37(m, 1H, =C H b )。

[0390] 13 C NMR(CDCl3, ppm) δ: 14.11(C-18′), 22.68(C-17′), 26.05(C-5′), 29.35, 29.46, 29.61 and 29.68(C-4′, C-6′-C-15′), 31.91(C-16′), 37.48(C-2), 52.03(OCH3), 64.29(C-1′), 68.39(C-2′), 71.46(C-3′), 128.74(= C H2), 133.64(C-3), 166.08(C-4), 171.11(C-1)。

[0391] CI MS: 413.3(MH) + (60)。

[0392] HRMS(CI): For C 24 H 45 O5(MH) + the calculated value: 413.3267; found: 413.3264。

[0393] 4-Methyl 1-(4-(tetradecyloxy)butyl) 2-methylenesuccinate (MK-937)

[0394]

[0395] Chromatography in cyclohexane-ethyl acetate (7:1) system. Yield: 259 mg (63%) of a colorless liquid。

[0396] 1 H NMR(CDCl3, ppm) δ: 0.87(t, 3H, J CH3,CH2 = 7.1, CH3(18′)), 1.37 - 1.20(m, 22H, 11xCH2(7′-17′)), 1.55(m, 2H, CH2(6′)), 1.64(m, 2H, CH2(3′)), 1.74(m, 2H, CH2(2′)), 3.33(d, 2H, J 2,＝C H = 1.1, CH2(2)), 3.38 (t, 2H, J 5′,6′ = 6.7, OCH2(5′)), 3.42 (t, 2H, J 4′,3′ = 6.4, OCH2(4′)), 3.69 (s, 3H, OCH3), 4.18 (t, 2H, J 1′,2′ = 6.6, OCH2(1′)), 5.70 (q, 1H, J gem = J Ha,2 = 1.1, =C H a ), 6.32 (m, 1H, =C H b )。

[0397] 13 C NMR (CDCl3, ppm) δ: 14.10 (C-18′), 22.67 (C-17′), 25.46 (C-2′), 26.17 and 26.21 (C-3′, C-7′), 29.34, 29.49, 29.60, 29.65 and 29.73 (C-6′ and C-8′-C-15′), 31.90 (C-16′), 37.53 (C-2), 52.01 (OCH3), 64.93 (C-1′), 70.10 (C-4′), 71.05 (C-5′), 128.36 (= C H2), 133.89 (C-3), 166.11 (C-4), 171.15 (C-1)。

[0398] CI MS: 412.3 (M) + (6)。

[0399] HRMS (CI): For C 24 H 44 O5 (M) + calculated: 412.3189; found: 412.3187。

[0400] 1-(2-(Butoxycarbonyl)phenyl)-4-methyl-2-methylenesuccinate (MK-939)

[0401]

[0402] Chromatography in a cyclohexane - ethyl acetate (6:1) system. Yield: 230 mg (72%) of a colorless liquid.

[0403] 1 H NMR (CDCl3, ppm) δ: 0.94 (t, 3H, CH3(4′)), 1.42 (m, 2H, CH2(3′)), 1.67 (m, 2H, CH2(2′)), 3.48 (d, 2H, JCH2,H = 1.1, C H2 -C=O), 3.73 (s, 3H, OCH3), 4.25 (t, 2H, J 1′,2′ = 6.7, OCH2(1')), 5.94 (q, 1H, J gem = J Ha,CH2 = 1.1, =C H a ), 6.62 (m, 1H, =C H b ), 7.14 (ddd, 1H, J 3,4 = 8.1, J 3,5 = 1.2, J 3,6 = 0.6, H-3), 7.32 (ddd, 1H, J 5,6 = 7.9, J 5,4 = 7.4, J 5,3 = 1.2, H-5), 7.56 (ddd, 1H, J 4,3 = 8.1, J 4,5 = 7.4, J 4,6 = 1.7, H-4), 8.02 (ddd, 1H, J 6,5 = 7.9, J 6,4 = 1.7, J 6,3 = 0.3, H-6).

[0404] 13 C NMR (CDCl3, ppm) δ: 13.70 (C-4'), 19.16 (C-3'), 30.60 (C-2'), 37.43 C H2C=O), 52.09 (OCH3), 65.05 (C-1'), 123.70 (C-1), 123.80 (C-3), 126.06 (C-5), 130.41 (= C H2), 131.74 (C-6), 133.18 ( C =CH2), 133.64 (C-4), 150.43 (C-2), 164.52 ( C OOBu), 164.70 ( C OOPh), 170.95 ( C OOMe).

[0405] ESIMS: 343.1 (M+Na) + (100).

[0406] HRMS (ESI): For C 17 H 20 O6Na (M+Na) + Calculated value: 343.11521; Measured value: 343.11487.

[0407] (S)-1-(4-(2-(tert-Butoxycarbonyl)amino)-3-(octylamino)-3-oxopropyl)phenyl)4-methyl 2-methylenesuccinate (MK-940)

[0408]

[0409] Chromatography in the cyclohexane-ethyl acetate (2:1) system, followed by additional chromatography in the CHCl3 system containing 1.5% MeOH. Yield: 325 mg (63%) of a white amorphous solid.

[0410] 1 H NMR (CDCl3, ppm) δ: 0.86 (t, 3H, J CH3,CH2 = 7.1, CH3(11′)), 1.14 - 1.31 (m, 10H, 5xCH2(6′ - 10′)), 1.36 (m, 2H, CH2(5′)), 1.41 (s, 9H, CH3(t-Bu)), 3.02 and 3.06 (m, 2H, CH2(tyrosine)), 3.14 (m, 2H, CH2(4′)), 3.44 (s, 2H, CH2 -C=O), 3.71 (s, 3H, OCH3), 4.25 (m, 1H, CH -NH(2′)), 5.09 (bs, 1H, 2′- NH ), 5.83 (bs, 1H, 3′- NH ), 5.88 (m, 1H, =CH a ), 6.53 (s, 1H, =CH b ), 7.04 (d, 2H, J 2,3 = 8.4, H-2(aromatic)), 7.21 (d, 2H, J 3,2 = 8.4, H-3(aromatic)).

[0411] 13 C NMR (CDCl3, ppm) δ: 14.04 (C-11′), 22.58 (C-10′), 26.75 (C-6′), 28.24( C H3(t-Bu)), 29.10, 29.13 and 29.32 (C-5′, C-7′, C-8′), 31.74 (C-9′), 37.48( C H2C=O), 37.91 (C-1′), 39.49 (C-4′), 52.11 (O C H3), 55.90 (C-2′), 80.16( C(CH3)3), 121.59 (C-2), 130.08 (= C H2), 130.29 (C-3), 133.36 ( C =CH2), 134.51 (C-4), 149.61 (C-1), 155.38 (NH- C OO), 164.60 ( C OOPh), 170.75 and 170.91 (C-3′, CH3O- C =O).

[0412] ESIMS: 1059.6 (2M+Na) + (10), 541.3 (M+Na) + (100).

[0413] HRMS(ESI): For C 28 H 42 O7N2Na (M+Na) + Calculated value: 541.28842; Found value: 541.28735.

[0414] 1-(4-((S)-2-((S)-2-((tert-Butoxycarbonyl)amino)-3-methylbutanamido)-3-isopropoxy-3-oxopropyl)phenyl)-4-methyl-2-methylenesuccinate (MK-942)

[0415]

[0416] Chromatography in cyclohexane-acetone (4:1) system. Yield: 420 mg (77%) of white solid.

[0417] 1 H NMR (DMSO-d6, ppm) δ: 0.77 - 0.80 (m, 6H, CH( CH3 )2), 1.05 and 1.13 (2x d, 6H, J CH3,CH = 6.3, OCH( CH3 )2), 1.37 (s, 9H, C( CH3 )3), 1.85 (m, 1H, CH (CH3)2), 2.94 (dd, 1H, J 3′a,2′ = 8.5, J gem = 13.9, H-3′a), 3.00 (dd, 1H, J 3′b,2′ = 6.4, J gem = 13.9, H-3′b), 3.50 (s, 2H, CH2-COOMe), 3.62 (s, 3H, OCH3), 3.80 (dd, 1H, J 2″,NH = 9.2, J 2″,3″ = 7.1, H-2″), 4.42 (m, 1H, H-2′), 4.81 (septet, 1H, J CH,CH3 = 6.3, O CH (CH3)2), 6.02 and 6.42 (2x d, 2H, J gem = 1.1, =CH2), 6.60 (d, 1H, J NH,2″ = 9.2, NH -2″), 7.00 (m, 2H, H-aromatic(2)), 7.28 (m, 2H, H-aromatic(3)), 8.29 (d, J NH,2′ = 7.5, NH-2′).

[0418] 13 C NMR (DMSO-d6, ppm) δ: 18.39 and 19.35 (C-4″), 21.54 and 21.69 (O-CH( C H3)2), 28.38 (C( C H3)3), 30.73 (C-3″), 36.24 (C-3′), 37.16 ( C H2-COOMe), 52.01 (OCH3), 53.70 (C-2′), 59.61 (C-2″), 68.23 (O- CH (CH3)2), 78.16 ( C (CH3)3), 121.43 (C-2), 130.47 (C-3), 130.88 (=CH2), 133.59 ( C =CH2), 135.00 (C-4), 149.24 (C-1), 155.50 (NH- C OO), 164.61 ( C OO-Ph), 170.96 and 171.02 (C-1′, C OOMe), 171.63 (C-1″).

[0419] ESIMS: 571.3 (M+Na) + (100), 549.3 (MH) + (2).

[0420] HRMS (ESI): For C 28 H 40 O9N2Na (M+Na) + calculated value: 571.26260; found value: 571.26247.

[0421] (S)-1-(4-(3-Benzyloxy)-2-((tert-butoxycarbonyl)amino)-3-oxopropyl)phenyl)4-methyl 2-methylenesuccinate (MK-943)

[0422]

[0423] Chromatography in cyclohexane-acetone (4:1) system. Yield: 439 mg (88%) of colorless paste.

[0424] 1 H NMR (DMSO-d6, ppm) δ: 1.32 (s, 9H, C( CH3 )3), 2.90 (dd, 1H, J 3′a,2′ = 10.0, J gem = 13.8, H-3′a), 3.02 (dd, 1H, J 3′b,2′ = 5.4, J gem = 13.8, H-3′b), 3.51 (s, 2H, CH2 -COOMe), 3.63 (s, 3H, OCH3), 4.21 (ddd, 1H, J 2′,3′ = 10.0 and 5.4, J 2′,NH = 8.0, (H-2′), 5.10 (s, 2H, O- CH2 Ph), 6.04 and 6.43 (2x d, 2H, J gem = 1.1, =CH2), 7.02 (m, 2H, H-2 (aromatic)), 7.27 - 7.37 (m, 7H, H-3, H-2″, H-3″, H-4″ (aromatic)), 7.41 (bd, 1H, J NH,CH = 8.0, NH).

[0425] 13 C NMR (DMSO-d6, ppm) δ: 28.33 (C( C H3)3), 35.85 (C-3′), 37.18 ( C H2-COOMe), 52.04 (O C H3), 55.60 (C-2′), 66.17 (O- C H2Ph), 78.60 ( C (CH3)3), 121.51 (C-2), 128.05 (C-2″), 128.24 (C-4″), 128.59 (C-3″), 130.46 (C-3), 130.95 (=CH2), 133.59 ( C=CH2), 135.48 (C-4), 136.06 (C-1″), 149.21 (C-1), 155.68 (NH- C O), 164.68 ( C OO-C(1)), 171.04 ( C OO-Me), 172.19 (C-1′).

[0426] ESIMS: 1017.5 (2M + Na) + (15), 520.2 (M + Na) + (100).

[0427] HRMS(ESI): For C 27 H 31 O8NNa (M + Na) + calculated value: 520.19419; found value: 520.19394.

[0428] Deprotection of the tert-butoxycarbonyl (Boc) group. General procedure.

[0429] A mixture of dichloromethane and trifluoroacetic acid (1:1, 16 mL) was added to the appropriate Boc derivative (0.8 mmol). The solution was stirred at room temperature for 20 min and evaporated. The residue was crystallized (MK-944), or chromatographed on a silica gel column (100 mL) in the system described below, then purified on reverse-phase HPLC and lyophilized (compounds MK-941, MK-945).

[0430] The following compounds were synthesized:

[0431] (S)-1-(4-(2-Amino-3-(octylamino)-3-oxopropyl)phenyl)-4-methyl-2-methylenesuccinate trifluoroacetate (MK-941)

[0432]

[0433] Chromatography in the ethyl acetate - methanol (15:2) system gave 183 mg (43%) of a yellow foam, which was subjected to final purification on reverse-phase HPLC (gradient 20% to 60% CH3CN in 0.1% TFA / H2O, over 40 min, retention time 36.8 min). Yield: 124 mg (29%) of a white solid.

[0434] 1 H NMR (DMSO-d6, ppm) δ: 0.85 (t, 3H, J CH3,CH2= 7.0, CH3), 1.11 - 1.32(m, 12H, 6x CH2(2″ - 7″)), 2.94(m, 1H, NH - CH a ), 3.00(m, 2H, Ph - CH2 ), 3.11(m, 1H, NH - CH b ), 3.51(s, 2H, CH2 COOMe), 3.63(s, 3H, OCH3), 3.91(m, 1H, CH - NH3 + ), 6.05 and 6.42(2x d, J gem = 1.1, =CH2), 7.08(m, 2H, H - 2(aromatic)), 7.26(m, 2H, H - 3(aromatic)), 8.22(bs, 3H, NH3 + ), 8.31(bt, 1H, J NH,1″ = 5.6, NH)。

[0435] 13 C NMR(DMSO - d6, ppm) δ: 14.44(CH3(8″)), 22.57(CH2(7″)), 26.76(CH2(3″), 29.07, 29.13 and 29.15(CH2(2″, 4″, 5″)), 31.73(CH2(6″), 36.93(Ph - C H2), 37.37( C H2COOMe), 39.12(NH - C H2), 52.26(O C H3), 53.96( C H - NH3 + ), 122.04(C - 2(aromatic)), 131.05(C - 3(aromatic)), 131.21(= C H2), 133.12(C - 4(aromatic)), 133.80( C =CH2), 149.98(C - 1(aromatic)), 164.80(COO), 167.89(NH - C =O), 171.26( C OOMe)。

[0436] ESIMS: 441.2(M + Na) + (100), 419.3(MH) + (35)。

[0437] HRMS(ESI): For C 23 H 34 O5N2Na(M + Na)+ Calculated value: 441.23599; Measured value: 441.23569. For C 23 H 35 O5N2(MH) + Calculated value: 419.25405; Measured value: 419.25385.

[0438] 1-(4-((S)-2-((S)-2-Amino-3-methylbutanamido)-3-isopropoxy-3-oxopropyl)phenyl)-4-methyl-2-methylenesuccinate trifluoroacetate (MK-944)

[0439]

[0440] Crystallized from ethyl acetate with added diethyl ether. Yield: 324 mg (72%) of white crystals.

[0441] 1 H NMR (DMSO-d6, ppm) δ: 0.93 and 0.97 (2 x d, 6H, J CH3,CH = 6.9, CH( CH3 )2), 1.07 and 1.15 (2 x d, 6H, J CH3,CH = 6.2, O-CH( CH3 )2), 2.13 (m, 1H, CH (CH3)2), 2.99 (dd, 1H, J 3′a,2′ = 8.3, J gem = 14.1, H-3′a), 3.04 (dd, 1H, J 3′b,2′ = 6.3, J gem = 14.1, H-3′b), 3.51 (s, 2H, CH2 -COOMe), 3.63 (s, 3H, OCH3), 3.65 (d, 1H, J 2″,3″ = 5.1, H-2″), 4.50 (m, 1H, H-2′), 4.85 (septet, 1H, J CH,CH3 = 6.2, O- CH (CH3)2), 6.04 and 6.43 (2 x m, 2H, =CH2), 7.05 (m, 2H, H-2), 7.32 (m, 2H, H-3), 8.09 (bs, 3H, NH3 + ), 8.90 (d, 1H, J NH,2′ = 7.0, NH).

[0442] 13 C NMR (DMSO-d6, ppm) δ: 17.44 and 18.56 (C-4″), 21.52 and 21.69 (O-CH(C H3)2),30.10(C-3″),36.03(C-3′),37.16( C H2-COOMe),52.03(OCH3),54.27(C-2′),57.24(C-2″),68.66(O- CH (CH3)2),121.66(C-2),130.53(C-3),131.04(= C H2),133.54( C =CH2),134.68(C-4),149.41(C-1),164.71( C OO-Ph),168.41(NH- C O),170.56( C OO-iPr),171.07( C OOMe).

[0443] ESIMS:471.2(M+Na) + (58),449.2(MH) + (100).

[0444] HRMS(ESI): For C 23 H 32 O7N2Na(M+Na) + calculated value: 471.21017; found value: 471.20999. For C 23 H 33 O7N2(MH) + calculated value: 449.22823; found value: 449.22815.

[0445] (S)-1-(4-(2-Amino-3-(benzyloxy)-3-oxopropyl)phenyl)-4-methyl-2-methylenesuccinate trifluoroacetate (MK-945)

[0446]

[0447] Chromatography in the dichloromethane - methanol (50:2) system gave 270 mg (68%) of a yellow slurry, which was subjected to final purification on reverse-phase HPLC (gradient 15% to 50% CH3CN in 1% TFA / H2O, over 40 min, retention time 37.1 min). Yield: 85 mg (21%) of a viscous semi-solid.

[0448] 1 H NMR (DMSO-d6, ppm) δ: 3.08 (dd, 1H, J 3′a,2′ = 7.7, Jgem = 14.1, H-3′a), 3.16 (dd, 1H, J 3′b,2′ = 6.0, J gem = 14.1, H-3′b), 3.53 (s, 2H, CH2 -COOMe), 3.63 (s, 3H, OCH3), 4.40 (dd, 1H, J 2′,3′ = 7.6 and 6.1, H-2′), 5.13 and 5.17 (2x d, 2H, J gem = 12.3, O- CH2 Ph), 6.06 and 6.45 (2xd, 2H, J gem = 1.1, =CH2), 7.05 (m, 2H, H-2 (aromatic)), 7.24 (m, 2H, H-3 (aromatic)), 7.26 (m, 2H, H-2″), 7.34 - 7.38 (m, 3H, H-3″, H-4″ (aromatic)), 8.58 (bs, 3H, NH3 + ).

[0449] 13 C NMR (DMSO-d6, ppm) δ: 35.68 (C-3′), 37.21 ( C H2-COOMe), 52.07 (O C H3), 53.35 (C-2′), 67.44 (O- C H2Ph), 121.97 (C-2), 128.05 (C-2″), 128.67 - 128.69 (C-2″, C-3″, C-4″), 130.87 (C-3), 131.13 (= C H2), 132.48 (C-4), 133.56 ( C =CH2), 135.02 (C-1″), 149.86 (C-1), 164.62 (CH2- C OO), 169.20 ( C OO-Bn), 171.11 ( C OO-Me),

[0450] ESIMS: 420.1 (M+Na) + (38), 398.2 (MH) + (100).

[0451] HRMS (ESI): For C 22 H 23 O6NNa (M+Na) + the calculated value: 420.14176; the measured value: 420.14120. For C22 H 24 O6N(MH) + Calculated value: 398.15981; Measured value: 398.15939.

[0452] Hydrocarbyl α-methyl itaconate

[0453]

[0454] R = hydrocarbyl, aryl, hydrocarbyl salicyl, Boc-tyrosyl hydrocarbylamide, Boc-tyrosyl ester, (di)peptide having a terminal tyrosyl ester, (di)peptide having a terminal tyrosyl hydrocarbylamide

[0455] Examples of R-OH of hydrocarbyl α-methyl itaconate include but are not limited to:

[0456]

[0457] Dissolve α-methyl itaconate (1 mmol) in dry dichloromethane (3 mL). Add the appropriate hydroxy derivative (1.2 mmol) and DMAP (1.5 mmol), and cool the solution to 0 °C. Under nitrogen, add dropwise a solution of DCC (1.5 mmol) in dichloromethane (2 mL), and stir the whole mixture at 0 °C for 1 h, and then stir overnight at room temperature. Filter the mixture, and extract the solution with 10% aqueous KHSO4 (3 × 5 mL) and saturated NaHCO3 (5 mL). Dry the organic phase over sodium sulfate, evaporate, and subject the residue to chromatography on a silica gel column in the system described below.

[0458] 1-Methyl 4-(4-(tetradecyloxy)butyl) 2-methylenesuccinate (MK-956)

[0459]

[0460] Chromatography in a cyclohexane-ethyl acetate (10:1) system. Yield: 244 mg (59%) of a colorless liquid.

[0461] 1 H NMR (CDCl3, ppm) δ: 0.87 (t, 3H, J CH3,CH2 = 7.0, CH3(14)), 1.24 - 1.33 (m, 22H, 11xCH2(3 - 13)), 1.54 (m, 2H, CH2(2)), 1.62 (m, 2H, COO(CH2)2 CH2 CH2O), 1.69 (m, 2H, COO-CH2 CH2 ), 3.33 (d, 2H, J CH2,C＝CH2 = 1.2,CH2 -COO), 3.38 (t, 2H, J 1,2 = 6.7, CH2(1)), 3.41 (t, 2H, J CH2,CH2 = 6.4, COO-(CH2)3 CH2 O), 3.76 (s, 3H, OCH3), 4.12 (t, 2H, J CH2,CH2 = 6.5, COO-CH2), 5.70 (q, 1H, J Ha,CH2 = J gem 1.1, =CH a ), 6.32 (d, 1H, J gem 1.1, =CH b )。

[0462] 13 C NMR (CDCl3, ppm) δ: 14.11 (C-14), 22.67 (C-13), 25.42 (COO-CH2- C H2), 26.14 and 26.17 (C-3, COO(CH2)2 C H2CH2O), 29.34–29.73 (m, C-2, C-4–C-11), 31.91 (C-12), 37.75 ( C H2COO), 52.10 (OCH3), 64.81 (COO- C H2), 70.10 (COO-(CH2)3 C H2O), 71.05 (C-1), 128.44 (C= C H2), 133.75 ( C =CH2), 166.65 ( C OO-CH3), 170.72 (CH2- C OO)。

[0463] ESIMS: 847.6 (2M+Na) + (10), 435.3 (M+Na) + (100)。

[0464] HRMS (ESI): For C 24 H 44 O5Na (M+Na) + Calculated value for: 435.30810; Found: 435.30841.

[0465] 4-(3-(Hexadecyloxy)propyl)-1-methyl-2-methylenesuccinate (MK-957)

[0466]

[0467] Chromatography in the cyclohexane-ethyl acetate (80:15) system. Yield: 120 mg (28%) of a colorless liquid.

[0468] 1 1H NMR (CDCl3, ppm) δ: 0.87 (t, 3H, J CH3,CH2 = 7.0, CH3(16)), 1.23 - 1.33 (m, 26H, 13xCH2(3 - 15)), 1.54 (m, 2H, CH2(2)), 1.88 (p, 2H, OCH2 - CH2 -CH2O), 3.33 (d, 2H, J CH2,C＝CH2 = 1.2, CH2 -COO), 3.38 (t, 2H, J 1,2 = 6.7, OCH2(1)), 3.45 (t, 2H, J CH2,CH2 = 6.3, COO-(CH2)2 CH2 O), 3.76 (s, 3H, OCH3), 4.19 (t, 2H, J CH2,CH2 = 6.5, COO - CH2 ), 5.70 (q, 1H, J Ha,CH2 = J gem 1.1, =CH a ), 6.32 (d, 1H, J gem 1.1, =CH b )。

[0469] 13 13C NMR (CDCl3, ppm) δ: 14.10 (C - 16), 22.67 (C - 15), 26.14 (C - 3), 28.98 (OCH2 - C H2 - CH2O), 29.34 - 29.69 (m, C - 2, C - 4–C - 13), 31.91 (C - 14), 37.72 ( C H2COO), 52.10 (OCH3), 62.27 (COO - CH2), 66.99 (COO-(CH2)2 C H2O), 71.16 (C - 1), 128.44 (C = C H2), 133.73 ( C =CH2), 166.63 ( C OO - CH3), 170.64 (CH2 - C OO)。

[0470] ESIMS: 875.7 (2M + Na) + (5), 449.3 (M + Na)+ (100).

[0471] HRMS(ESI): For C 25 H 46 O5Na (M+Na) + Calculated value: 449.32375; Found value: 449.32382.

[0472] 4-(2-(Butoxycarbonyl)phenyl)-1-methyl-2-methylenesuccinate (MK-961)

[0473]

[0474] Chromatography in cyclohexane-ethyl acetate (6:1) system. Yield: 63 mg (20%) of colorless liquid.

[0475] 1 H NMR (CDCl3, ppm) δ: 0.97 (t, 3H, J CH3,CH2 = 7.4, CH3(4')), 1.45 (m, 2H, CH2(3')), 1.72 (m, 2H, CH2(2')), 3.67 (d, 2H, J CH2,C＝CH2 = 1.1, C H2 -C=O), 3.81 (s, 3H, OCH3), 4.27 (t, 2H, J 1′,2′ = 6.7, OCH2(1')), 5.89 (q, 1H, J gem = J Ha,CH2 = 1.1, =C H a ), 6.42 (d, 1H, J gem = 1.1, =C H b ), 7.11 (dddd, 1H, J 3,4 = 8.1, J 3,5 = 1.1, H-3), 7.30 (m, 1H, H-5), 7.54 (ddd, 1H, J 4,3 = 8.2, J 4,5 = 7.4, J 4,6 = 1.7, H-4), 8.00 (dd, 1H, J 6,5 = 7.9, J 6,4 = 1.7, H-6).

[0476] 13 C NMR (CDCl3, ppm) δ: 13.74 (C-4'), 19.20 (C-3'), 30.68 (C-2'), 37.50 CH2C=O), 52.20 (OCH3), 64.97 (C-1′), 123.43 (C-1), 123.79 (C-3), 126.04 (C-5), 129.30 (= C H2), 131.59 (C-6), 133.19 ( C =CH2), 133.67 (C-4), 150.56 (C-2), 164.38 ( C OOBu), 166.59 ( C OOCH3), 169.37 (CH2- C OO).

[0477] ESIMS: 343.1 (M+Na) + (100).

[0478] HRMS(ESI): For C 17 H 20 O6Na (M+Na) + Calculated value: 343.11521; Measured value: 343.11532.

[0479] 4-(4-((S)-2-((S)-2-((tert-Butoxycarbonyl)amino)-3-methylbutanamido)-3-isopropoxy-3-oxopropyl)phenyl)-1-methyl-2-methylenesuccinate (MK-962)

[0480]

[0481] Chromatography in a cyclohexane-acetone (4:1) system. Yield: 255 mg (47%) of white solid.

[0482] 1 H NMR (DMSO-d6, ppm) δ: 0.77 - 0.79 (m, 6H, CH( CH3 )2), 1.04 and 1.12 (2x d, 6H, J CH3,CH =6.2, OCH( CH3 )2), 1.37 (s, 9H, C( CH3 )3), 1.85 (m, 1H, CH (CH3)2), 2.93 (dd, 1H, J 3′a,2′ =8.5, J gem =14.0, H-3′a), 2.99 (dd, 1H, J 3′b,2′ =6.6, J gem =14.0, H-3′b), 3.62 (s, 2H, CH2-COOMe), 3.72 (s, 3H, OCH3), 3.80 (m, 1H, H-2″), 4.42 (q, 1H, J 2′,3′ = J 2′,NH = 7.4, H-2′), 4.81 (septet, 1H, J CH,CH3 = 6.2, O CH (CH3)2), 5.96 and 6.28 (2x s, 2H, =CH2), 6.57 (d, 1H, J NH,2″ = 9.2, NH -2″), 6.97 (m, 2H, H-aromatic(2)), 7.26 (m, 2H, H-aromatic(3)), 8.26 (d, J NH,2′ = 7.4, NH-2′).

[0483] 13 C NMR (DMSO-d6, ppm) δ: 18.35 and 19.31 (C-4″), 21.50 and 21.65 (O-CH( C H3)2), 28.36 (C( C H3)3), 30.71 (C-3″), 36.24 (C-3′), 37.38 ( C H2-COOMe), 52.29 (OCH3), 53.66 (C-2′), 59.61 (C-2″), 68.19 (O- CH (CH3)2), 78.16 ( C (CH3)3), 121.42 (C-2), 129.66 (=CH2), 130.41 (C-3), 133.62 ( C =CH2), 134.89 (C-4), 149.24 (C-1), 155.48 (NH- C OO), 166.27 ( C OO-Me), 169.38 ( C OO-Ph), 170.94 (C-1′), 171.59 (C-1″).

[0484] ESIMS: 571.3 (M+Na) + (100).

[0485] HRMS (ESI): For C 28 H 40 O9N2Na (M+Na) + calculated for: 571.26260; found: 571.26280.

[0486] (S)-4-(4-(3-Benzyloxy)-2-((tert-butoxycarbonyl)amino)-3-oxopropyl)phenyl) 1-methyl 2-methylenesuccinate (MK-963)

[0487]

[0488] Chromatography in cyclohexane-acetone (4:1) system. Yield: 223 mg (45%) of amorphous solid.

[0489] 1 H NMR (CDCl3, ppm) δ: 1.41 (s, 9H, C( CH3 )3), 3.04 (dd, 1H, J 3′a,2′ = 6.0, J gem = 14.0, H-3′a), 3.09 (dd, 1H, J 3′b,2′ = 6.0, J gem = 13.9, H-3′b), 3.56 (d, 2H, J CH2,C＝CH2 = 1.1, CH2 -COOMe), 3.80 (s, 3H, OCH3), 4.60 (m, 1H, CH -NH), 4.99 (d, J NH,CH = 8.3, NH), 5.09 and 5.17 (2x d, 2H, J gem = 12.2, O- CH2 Ph), 5.82 (q, 1H, J CH2,＝CHa = J gem = 1.1, =CH a ), 6.40 (d, 1H, J gem = 0.8, =CH b ), 6.95 (m, 2H, H-2 (aromatic)), 7.02 (m, 2H, H-3 (aromatic)), 7.28 - 7.38 (m, 5H, H-2″, H-3″, H-4″ (aromatic)).

[0490] 13 C NMR (CDCl3, ppm) δ: 28.25 (C( C H3)3), 37.51 (C-3′), 37.86 ( C H2-COOMe), 52.23 (O C H3), 54.31 (C-2′), 67.15 (O- C H2Ph), 79.97 ( C(CH3)3), 121.40 (C-2), 128.48 (C-4″), 128.58 and 128.60 (C-2″, C-3″), 129.00 (=CH2), 130.28 (C-3), 133.29 ( C =CH2), 133.50 (C-4), 135.06 (C-1″), 149.61 (C-1), 155.02 (NH- C O), 166.47 ( C OO-Me), 169.07 (CH2- C OO), 171.51 (C-1′).

[0491] ESIMS: 1017.7 (2M+Na) + (3), 520.3 (M+Na) + (100).

[0492] HRMS(ESI): For C 27 H 31 O8NNa (M+Na) + calculated value: 520.19419; found value: 520.19324. For C 27 H 32 O8N (MH) + calculated value: 498.21224; found value: 498.21161.

[0493] Deprotection of the tert-butoxycarbonyl (Boc) group. General procedure.

[0494] A mixture of dichloromethane and trifluoroacetic acid (1:1, 16 mL) was added to the appropriate Boc derivative (0.8 mmol). The solution was stirred at room temperature for 20 min and evaporated. The residue was treated as described below.

[0495] 4-(4-((S)-2-((S)-2-Amino-3-methylbutanamido)-3-isopropoxy-3-oxopropyl)phenyl)-1-methyl-2-methylenesuccinate trifluoroacetate (MK-964)

[0496]

[0497] The residue was crystallized from a mixture of ethyl acetate and diethyl ether. Yield: 340 mg (75%) of white crystals.

[0498] 1 H NMR (DMSO-d6, ppm) δ: 0.93 and 0.97 (2x d, 6H, CH( CH3), 1.06 and 1.14 (2x d, 6H, J CH3,CH = 6.2, O-CH( CH3 ), 2.12 (septet d, 1H, J 3″,4″ = 6.9, J 3″,2″ = 5.0, H-3″), 2.98 (dd, 1H, J 3′a,2′ = 8.2, J gem = 14.2, H-3′a), 3.03 (dd, 1H, J 3′b,2′ = 6.6, J gem = 14.2, H-3′b), 3.64 (m, 2H, CH2 -COOMe), 3.65 (d, 1H, J 2″,3″ = 5.1, H-2″), 3.73 (s, 3H, OCH3), 4.49 (m, 1H, H-2′), 4.84 (septet, 1H, J CH,CH3 = 6.2, O- CH (CH3)2), 5.96 (q, 1H, J CH2,＝CHa = J gem = 1.2, =CH a ), 6.28 (d, 1H, J gem = 1.2, =CH b ), 7.02 (m, 2H, H-2), 7.31 (m, 2H, H-3), 8.09 (bs, 3H, NH3 + ), 8.89 (d, 1H, J NH,2′ = 7.0, NH).

[0499] 13 C NMR (DMSO-d6, ppm) δ: 17.43 and 18.51 (C-4″), 21.47 and 21.65 (O-CH( C H3)2), 30.07 (C-3″), 36.03 (C-3′), 37.40 ( C H2-COOMe), 52.32 (OCH3), 54.23 (C-2′), 57.23 (C-2″), 68.60 (O- CH (CH3)2), 121.62 (C-2), 129.72 (= C H2), 130.47 (C-3), 133.60 ( C =CH2), 134.56 (C-4), 149.40 (C-1), 166.30 ( C OO-Me), 168.37 (NH- C O), 169.51 (CH2- COO),170.53( C OO-iPr).

[0500] ESIMS: 471.2 (M+Na) + (20), 449.2 (MH) + (100).

[0501] HRMS(ESI): For C 23 H 33 O7N2 (MH) + Calculated value: 449.22823; Found value: 449.22745.

[0502] (S)-4-(4-(2-Amino-3-(benzyloxy)-3-oxopropyl)phenyl)-1-methyl-2-methylenesuccinate trifluoroacetate (MK-965)

[0503]

[0504] The residue was chromatographed on a silica gel column (60 mL) with a gradient of cyclohexane - acetone (1:1 to 1:4), followed by a chloroform - methanol (1:1) system to give 380 mg (93%) of a white viscous foam.

[0505] 1 H NMR (DMSO-d6, ppm) δ: 3.07 (dd, 1H, J 3′a,2′ = 7.5, J gem = 14.1, H-3′a), 3.15 (dd, 1H, J 3′b,2′ = 6.1, J gem = 14.1, H-3′b), 3.66 (d, 2H, J CH2,C＝CH2 = 1.2, CH2 -COOMe), 3.73 (s, 3H, OCH3), 4.37 (dd, 1H, J 2′,3′ = 7.6 and 6.1, H-2′), 5.13 and 5.17 (2x d, 2H, J gem = 12.3, O- CH2 Ph), 5.98 (q, 1H, J gem = J CH2,＝CHa = 1.2, =CH a ), 6.30 (d, 1H, J gem = 1.2, =CH b), 7.02 (m, 2H, H-2 (aromatic)), 7.22 (m, 2H, H-3 (aromatic)), 7.26 (m, 2H, H-2″ (aromatic)), 7.34 - 7.37 (m, 3H, H-3″, H-4″ (aromatic)), 8.48 (bs, 3H, NH3 + ).

[0506] 13 C NMR (DMSO-d6, ppm) δ: 35.71 (C-3′), 34.73 ( C H2-COOMe), 52.33 (O C H3), 53.35 (C-2′), 67.35 (O- C H2Ph), 121.88 (C-2), 128.60 (C-2″, C-3″), 128.63 (C-4″), 129.72 ((= C H2), 130.80 (C-3′), 132.42 (C-4), 133.62 ( C =CH2), 135.01 (C-1″), 149.82 (C-1), 166.32 ( C OO-Me), 169.25 ( C OO-Bn), 169.40 (CH2- C OO).

[0507] ESIMS: 420.1 (M+Na) + (15), 398.2 (MH) + (100).

[0508] HRMS (ESI): For C 22 H 23 O6NNa (M+Na) + the calculated value: 420.14176; the measured value: 420.14105. For C 22 H 24 O6N (MH) + the calculated value: 398.15981; the measured value: 398.15930.

[0509] 4-(((Isopropoxycarbonyl)oxy)methyl)-1-methyl-2-methylenesuccinate (IS-101-088) (21)

[0510]

[0511] α-Methyl itaconate (0.2 g, 1.39 mmol), isopropyl chloromethyl carbonate (0.22 mL, 1.66 mmol), sodium iodide (50 mg, 0.33 mmol), and potassium carbonate (0.29 g, 2.08 mmol) were dissolved in anhydrous MeCN (5 mL), and the mixture was stirred at 50 °C for 16 h. EtOAc (60 mL) was added and the mixture was washed with brine (20 mL). The organic phase was dried over Na2SO4, the volatiles were evaporated, and the residue was subjected to flash column chromatography (silica gel 60 mesh 70–230, solvent: cyclohexane / ethyl acetate 5:1) to afford 0.34 g (93%) of compound IS-101-088 (21) as a colorless oil.

[0512] 1 H NMR (401 MHz, CDCl3): δ H 1.31 (d, J = 6.2 Hz, 6H), 3.37–3.43 (m, 2H), 3.76 (d, J = 0.6 Hz, 3H), 4.85–4.97 (m, 1H), 5.72–5.78 (m, 3H), 6.36 (d, J = 0.8 Hz, 1H).

[0513] 13 C NMR (101 MHz, CDCl3): δ C 21.6, 21.7, 37.3, 52.2, 73.1, 81.9, 129.2, 132.9, 153.3, 166.3, 169.2.

[0514] ESIMS: 283.1 ([M+Na] + )

[0515] HRMS (ESI): Calcd for C 11 H 16 O7Na 283.07882. Found: 283.07925.

[0516] 1-Methyl 4-((pivaloyloxy)methyl) 2-methylenesuccinate (IS-101-089) (23)

[0517]

[0518] α-Methyl itaconate (0.2 g, 1.39 mmol), chloroformate pivalate (0.26 mL, 1.8 mmol), sodium iodide (50 mg, 0.33 mmol), and potassium carbonate (0.29 g, 2.08 mmol) were dissolved in anhydrous MeCN (5 mL), and the mixture was stirred at 40 °C for 16 h. EtOAc (60 mL) was added and the mixture was washed with brine (20 mL). The organic phase was dried over Na2SO4, the volatiles were evaporated, and the residue was subjected to flash column chromatography (silica gel 60 mesh 70 - 230, solvent: cyclohexane / ethyl acetate 5:1) to afford 0.33 g (92%) of compound IS-101-089 (23) as a colorless oil.

[0519] 1 H NMR (401 MHz, CDCl3): δ H 1.21 (s, 9H), 3.38 (d, J = 1.2 Hz, 2H), 3.76 (s, 3H), 5.74 (d, J = 1.1 Hz, 1H), 5.76 (s, 2H), 6.35 (d, J = 0.8 Hz, 1H).

[0520] 13 C NMR (101 MHz, CDCl3): δ C 26.9, 37.4, 38.8, 52.2, 79.7, 129.0, 133.0, 166.3, 169.4, 177.1.

[0521] ESIMS: 281.1 ([M+Na] + )

[0522] HRMS (ESI): Calcd for C 12 H 18 O6Na 281.09956. Found: 281.09993.

[0523] 1-(((tert-Butoxycarbonyl)oxy)methyl) 4-methyl 2-methylenesuccinate (IS-102-081)

[0524]

[0525] β-Methyl itaconate (1.00 g, 6.94 mmol), tert-butyl (chloromethyl) carbonate (1.27 g, 7.60 mmol), sodium iodide (0.20 g, 1.33 mmol), and potassium carbonate (1.44 g, 10.4 mmol) were dissolved in anhydrous MeCN (20 mL), and the mixture was stirred at 50 °C for 16 h. The volatiles were then evaporated, the residue was redissolved in EtOAc (60 mL), and the mixture was washed with saturated sodium thiosulfate solution (20 mL) and brine (70 mL). The organic phase was dried over Na2SO4, the volatiles were evaporated, and the residue was subjected to flash column chromatography (silica gel 60 mesh 70 - 230, solvent: cyclohexane / ethyl acetate 5:1) to afford 1.65 g (87%) of compound IS-102-081 as a colorless oil.

[0526] 1 H NMR (401 MHz, CDCl3): δ H 1.49 (s, 9H), 3.34 (s, 2H), 3.68 (s, 3H), 5.78 (s, 2H), 5.81 (s, 1H), 6.42 (s, 1H).

[0527] 13 C NMR (101 MHz, CDCl3): δ C 27.7, 37.3, 52.2, 82.0, 83.7, 130.5, 133.0, 152.0, 164.7, 170.9.

[0528] ESIMS: 297.1 ([M+Na] + )

[0529] HRMS (ESI): Calcd for C 12 H 18 O7Na 297.09447. Found: 297.09465.

[0530] Scheme 1. Structure numbering for NMR assignment (alkyl β-methyl itaconate)

[0531]

[0532] Scheme 2. Structure numbering for NMR assignment (alkyl α-methyl itaconate)

[0533]

[0534] Example 6

[0535] In vitro data - mouse plasma stability

[0536] To evaluate the murine plasma stability of the intact prodrug over time, the prodrug was spiked into murine plasma to a final assay concentration of 10 μM. The spiked samples were incubated at 37 °C in an orbital shaker for 1 h and then the reaction was quenched with three volumes of acetonitrile containing an internal standard (IS; losartan: 0.5 μM). The samples were vortexed and centrifuged at 16,000 g for 5 min at 4 °C. 50 μL of the supernatant was diluted with 50 μL of water and transferred to a 250 μL polypropylene vial sealed with a Teflon cap. The disappearance of the prodrug over time was monitored using liquid chromatography mass spectrometry (LC-MS). Results for a representative prodrug disclosed herein are shown in Figure 1 are shown. Release of the active monomethyl itaconate (in plasma) from the representative prodrug is shown in Figure 2 are shown. Release of the active itaconic acid (in plasma) from the representative prodrug is shown in Figure 3 are shown.

[0537] References

[0538] All publications, patent applications, patents, and other references mentioned in the specification represent the level of those skilled in the art to which the disclosed subject matter of the invention pertains. All publications, patent applications, patents, and other references are hereby incorporated by reference to the extent as if each individual publication, patent application, patent, and other reference were specifically and individually indicated to be incorporated by reference. It will be understood that although many patent applications, patents, and other references are mentioned herein, such references do not constitute an admission that any of these documents forms part of the common general knowledge in the art.

[0539] International PCT Patent Application Publication No. WO2017142855, Artyomov et al., Immunomodulatory Agents and Methods of Use Thereof, published Aug. 24, 2017;

[0540] International PCT Patent Application Publication No. WO2019036509, Artyomov et al., Methods and Compositions for the Treatment of Diseases Associated with Cancer, Inflammation, or Immune Response, published Feb. 21, 2019;

[0541] O’Neill, L.A.J. and Artyomov, M.N., Itaconate: the poster child of metabolic reprogramming in macrophage function, Nature Reviews: Immunology, 19, 273 - 281 (2019).

[0542] Boschert, D.; Schneider - Chaabane, A.; Himmelsbach, A.; Eickenscheidt, A.; Lienkamp, K. Synthesis and Bioactivity of Polymer - Based Synthetic Mimics of Antimicrobial Peptides (SMAMPs) Made from Asymmetrically Disubstituted Itaconates. Chem. Eur. J. 2018, 24, 8217 - 8227.

[0543] McKenna, C.E.; Kashemirov, B.A; Krylov, I.S.; Zakharova, V.M. Method to improve antiviral activity of nucleotide analogue drugs. US9550803B2, January 24, 2017.

[0544] Hidaka, K.; Gohda, K.; Teno, N.; Wanaka, K.; Tsuda, Y. Active site - directed plasmin inhibitors: Extension on the P2 residue. Bioorg. Med. Chem. 2016, 24, 545 - 553.

[0545] Although the foregoing subject matter has been described in considerable detail for purposes of clarity of understanding by way of illustration and example, those skilled in the art will appreciate that certain changes and modifications may be made within the scope of the appended claims.

Claims

1. A compound of formula (I) or a pharmaceutically acceptable salt thereof: Wherein: R1 and R2 are selected from one or more of the following combinations: (a-i) R1 is -OR3, and R2 is selected from wherein R7 is selected from -C(=O)-O-R8, (a-ii) R1 is -OCH3 and R2 is (b) R1 is and R2 is selected from -OR3, wherein R7 is selected from -C(=O)-O-R8, (c) R1 is and R2 is selected from -OR3, wherein R7 is selected from -C(=O)-O-R8, (d) R1 is and R2 is selected from -OR3, wherein R7 is selected from -C(=O)-O-R8, (e) R1 is and R2 is selected from R7 is selected from -C(=O)-O-R8, (f) R1 is wherein R7 is -C(=O)-O-R8; and R2 is selected from -OR3, wherein R7 is selected from -C(=O)-O-R8, (g) R1 is wherein R7 is and R2 is selected from -OR3, wherein R7 is selected from -C(=O)-O-R8, (h) R1 is wherein R7 is and R2 is selected from -OR3, wherein R7 is selected from -C(=O)-O-R8, and (i) R1 is wherein R7 is and R2 is selected from -OR3, wherein R7 is selected from -C(=O)-O-R8, Where n is an integer selected from 1, 2, 3, and 4; Where m is an integer selected from 1, 2, 3, and 4; Where p is an integer selected from 1, 2, 3, 4, 5, 6, 7, 8, 9, 10, 11, 12, 13, 14, 15, 16, 17, 18, 19, and 20; Where u is an integer selected from 1, 2, 3, and 4; Where R3 is H or an unsubstituted or substituted C1-C6 straight-chain or branched-chain hydrocarbon group; Where R4 is an unsubstituted or substituted C1-C6 straight-chain or branched-chain hydrocarbon group or -OR5; Where R5 is an unsubstituted or substituted C1-C6 straight-chain or branched-chain hydrocarbon group; Where R6 is an unsubstituted or substituted C1-C6 straight-chain or branched-chain hydrocarbon group; Where R8 is an unsubstituted or substituted C1-C6 straight-chain or branched-chain hydrocarbon group; Where R9 is H or an unsubstituted or substituted C1-C4 straight-chain or branched-chain hydrocarbon group; wherein R 10 is an unsubstituted or substituted C1-C6 straight-chain or branched-chain hydrocarbon group; wherein R 11 and R 12 are each independently H or a protecting group; wherein R 13 is an unsubstituted or substituted C1-C6 straight-chain or branched-chain hydrocarbyl group; wherein R 14 is H or an unsubstituted or substituted C1-C4 linear or branched hydrocarbon group; wherein R 15 is an unsubstituted or substituted C1-C6 straight-chain or branched-chain hydrocarbon group; wherein R 16 and R 17 each independently selected from H, C1-C4 linear or branched unsubstituted or substituted hydrocarbon groups, and protecting groups; wherein R 18 is an aryl group; wherein R 19 is an unsubstituted or substituted C1-C4 straight-chain or branched-chain hydrocarbon group; wherein R 20 is H or an unsubstituted or substituted C1-C4 straight-chain or branched hydrocarbon group; wherein R 21 is -OR 22 wherein R 22 is an unsubstituted or substituted C1-C6 straight-chain or branched hydrocarbon group or -NR 23 R 24 ; wherein R 23 and R 24 each independently is H or an unsubstituted or substituted C1-C4 straight-chain or branched hydrocarbon group; Provided that R1 and R2 cannot both be -OH at the same time.

2. The compound or a pharmaceutically acceptable salt thereof according to claim 1, wherein: (a-i) R1 is -OH and R2 is selected from wherein R7 is selected from -C(=O)-O-R8; (a-i’) R1 is -OR3, and R2 is selected from wherein R7 is selected from -C(=O)-O-R8, (b) R1 is and R2 is selected from -OR3, (c) R1 is and R2 is -OR3; (d) R1 is and R2 is selected from -OR3 and (f) R1 is wherein R7 is -C(=O)-O-R8 and R2 is -OR3; (g) R1 is wherein R7 is and R2 is -OR3; (h) R1 is wherein R7 is and R2 is selected from -OR3; (i) R1 is wherein R7 is and R2 is -OR3.

3. The compound according to any one of claims 1-2 or a pharmaceutically acceptable salt thereof, wherein R3, R4, R5, R6, R8, R9, R 10 、R 13 、R 15 and R 22 are each independently selected from methyl, ethyl, n-propyl, isopropyl, n-butyl, sec-butyl, isobutyl, tert-butyl, pentyl, isopentyl, neopentyl, n-hexyl, 2-methylpentane, 3-methylpentane, 2,2-dimethylbutane and 2,3-dimethylbutane.

4. The compound according to any one of claims 1-3 or a pharmaceutically acceptable salt thereof, wherein R9, R 14 , R 16 , R 17 , R 19 , R 20 , R 23 and R 24 are each independently selected from methyl, ethyl, n-propyl, isopropyl, n-butyl, sec-butyl, isobutyl and tert-butyl.

5. The compound or a pharmaceutically acceptable salt thereof according to any one of claims 1-4, wherein the protecting group is selected from tert-butoxycarbonyl (boc), benzyloxycarbonyl (Cbz), p-methoxybenzylcarbonyl (Moz or MeOZ), 9-fluorenylmethoxycarbonyl (Fmoc) group, benzyl (Bn), p-methoxybenzyl (PMB), 3,4-dimethoxybenzyl (DMPM), p-toluenesulfonyl (Ts), Troc (trichloroethyl chloroformate), (4-nitrophenyl)sulfonyl (Nosyl), and nitrophenylthio (Nps).

6. The compound according to any one of claims 1-5 or a pharmaceutically acceptable salt thereof, wherein R 18 is selected from phenyl.

7. A compound of formula (I) or a pharmaceutically acceptable salt thereof: Wherein: (a-i) R1 is -OH and R2 is selected from: (a-ii) R1 is selected from -OCH3, -OCH(CH3)2, and -OC(CH3)3 and R2 is selected from: (b) R1 is and R2 is -OCH3 (5, 6) or (c) R1 is and R2 is -OCH3 (12, 13); (d) R1 is and R2 is selected from: -OCH3 (11) and (e) R1 is and R2 is -OCH3(14); (f) R1 is and R2 is -OCH3 (17, 19); (g) R1 is and R2 is -OCH3 (15, 16); or (h) R1 is and R2 is -OCH3 (18, 20).

8. A compound or a pharmaceutically acceptable salt thereof, wherein the compound is selected from:

9. A pharmaceutical composition comprising the compound or a pharmaceutically acceptable salt thereof according to any one of claims 1 to 8 and a pharmaceutically acceptable excipient.

10. Use of the compound or a pharmaceutically acceptable salt thereof according to any one of claims 1-8 or the pharmaceutical composition according to claim 9 in the preparation of a drug for treating a disease, disorder, or condition associated with inflammation in a subject.

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