WO2021195555A1 - Palmitoylethanolamide compounds - Google Patents

Abstract

The present application provides palmitoylethanolamide compounds useful for treating a disease or disorder in a subject in need thereof. Pharmaceutical compositions comprising the compounds and methods of treating diseases or disorders are also provided.

Description

Palmitoylethanolamide Compounds

CROSS-REFERENCE TO RELATED APPLICATIONS

This application claims benefit of U.S. Provisional Application Serial No. 63/000,782, filed on March 27, 2020, the disclosure of which is incorporated herein by reference in its entirety.

TECHNICAL FIELD

The present application provides palmitoylethanolamide compounds useful for treating a disease or disorder in a subject in need thereof.

BACKGROUND

Palmitoylethanolamide (PEA) is an endogenous fatty acid amide of antineuroinflammation and neuroprotective profile, primarily via its agonistic activity against Peroxisome Proliferator- Activated Receptor alpha (PPAR-α) with therapeutic potential for neurological disorders (e.g. see Br J Pharmacol. 2016; 173(12): 1899).

SUMMARY

The present invention relates to, inter alia , compounds of Formula I:

I or pharmaceutically acceptable salts, wherein constituent members are defined herein.

The present invention further provides pharmaceutical compositions comprising a compound of Formula I, or a pharmaceutically acceptable salt thereof, and a pharmaceutically acceptable carrier.

The present invention further provides methods of treating a disease or disorder in a subject, comprising administering to a subject in need thereof a therapeutically effective amount of a compound of Formula I, or a pharmaceutically acceptable salt thereof.

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 this invention belongs. Methods and materials are described herein for use in the present invention; other, suitable methods and materials known in the art can also be used. The materials, methods, and examples are illustrative only and not intended to be limiting. All publications, patent applications, patents, sequences, database entries, and other references mentioned herein are incorporated by reference in their entirety. In case of conflict, the present specification, including definitions, will control.

DESCRIPTION OF DRAWINGS

FIGs. 1A-3B show representative thin layer chromatography (TLC) images from the iodine (FIGs. 1 A, 2A, and 3 A) and cupric acetate (FIGs. IB, 2B, and 3B) stain analysis described in Example 4.

FIGs. 4-6 shows the concentration of Compounds 1 -3 measured in HIP samples from WT cells, cells transfected with D97A mutant Mfsd2a, and empty vector.

FIGs. 7A-7B show that Compound 1 exhibits cytoprotective properties in hiPSC-RPE cells.

DETAILED DESCRIPTION

The present application presents palmitoylethanolamide (PEA) derivatives that may improve the Blood Brain Barrier (BBB) penetrating profile of PEA, resulting in increased central nervous system (CNS) exposure. The improvement in the BBB penetrating profile of the compounds provided herein may facilitate delivery to the targeted site of action and/or neuronal cells. In addition, the compounds provided herein may be useful in combination therapies, partnered with other endocannabinoids (e.g. , anandamide, 2-arachidonoyl glycerol, and oleoylethanolamide) of synergistic potential.

Compounds

Accordingly, the present application provides a compound of Formula I:

I or a pharmaceutically acceptable salt thereof, wherein:

L¹ is C(O) or PO₂-;

X¹ is selected from the group consisting of C_1-4 alkylene, C_1-4 alkylene- OC(O)O-C_1-4 alkylene, C_1-4 alkylene-OC(O)NH-C_1-4 alkylene, C_1-4 alkylene- OC(O)C_1-4 alkylene, C_1-4 alkylene-O-C_1-4 alkylene, C_1-4 alkylene-O-C_1-4 alkylene-O- C_1-4 alkylene, and C_1-4 alkylene-O-C_1-4 alkylene-O-C(O)C_1-4 alkylene, wherein each C_1-4 alkylene is optionally substituted by OH or CO₂H;

X² is C_1-6 alkylene, which is optionally substituted by OH or CO₂H;

Y¹ is selected from the group consisting of O, S, and NR⁵;

R² is selected from the group consisting of H and C_1-6 alkyl, wherein the C_1-6 alkyl is optionally substituted by OH or CO₂H;

R³ is selected from the group consisting of H and C_1-6 alkyl;

R^3a is selected from the group consisting of H and C_1-6 alkyl;

R^3b is selected from the group consisting of H and C_1-6 alkyl;

R⁴ is selected from the group consisting of H and C_1-6 alkyl, wherein the C_1-6 alkyl is optionally substituted by OH or CO₂H; and

R⁵ is selected from the group consisting of H and C_1-6 alkyl, wherein the C_1-6 alkyl is optionally substituted by OH or CO₂H.

In some embodiments:

L¹ is C(O) or PO₂-;

X¹ is selected from the group consisting of C_1-4 alkylene, C_1-4 alkylene-OC(O)O-C_1-4 alkylene, C_1-4 alkylene-OC(O)NH-C_1-4 alkylene, C_1-4 alkylene- OC(O)C_1-4 alkylene, C_1-4 alkylene-O-C_1-4 alkylene, C_1-4 alkylene-O-C_1-4 alkylene-O- C_1-4 alkylene, and C_1-4 alkylene-O-C_1-4 alkylene-O-C(O)C_1-4 alkylene, wherein each C_1-4 alkylene is optionally substituted by CO₂H;

X² is C_1-6 alkylene, which is optionally substituted by OH or CO₂H;

Y¹ is selected from the group consisting of O, S, and NH;

R² is selected from the group consisting of H and C_1-6 alkyl, wherein the C_1-6 alkyl is optionally substituted by OH or CO₂H;

R³ is selected from the group consisting of H and C_1-6 alkyl;

R^3a is selected from the group consisting of H and C_1-6 alkyl; R^3b is selected from the group consisting of H and C_1-6 alkyl; and R⁴ is selected from the group consisting of H and C_1-6 alkyl, wherein the C_1-6 alkyl is optionally substituted by OH or CO₂H; provided that when L¹ is PO₂-, X¹ is an unsubstituted C_1-2 alkylene, X² is CH₂, R² is H, and R⁴ is H or methyl, then at least one of R³, R^3a, and R^3b is not methyl.

In some embodiments:

L¹ is C(O) or PO₂-;

X¹ is selected from the group consisting of C_1-4 alkylene, C_1-4 alkylene- OC(O)O-C_1-4 alkylene, C_1-4 alkylene-OC(O)C 1 -4 alkylene, C_1-4 alkylene-O-C_1-4 alkylene, C_1-4 alkylene-O-C_1-4 alkyl ene-O-C 1 -4 alkylene, and C_1-4 alkylene-O-C 1 -4 alkyl ene-O-C(O)C 1 -4 alkylene, wherein each C_1-4 alkylene is optionally substituted by OH or CO₂H;

X² is C_1-6 alkylene, which is optionally substituted by OH or CO₂H;

Y¹ is selected from the group consisting of O, S, and NR⁵;

R² is selected from the group consisting of H and C_1-6 alkyl, wherein the C_1-6 alkyl is optionally substituted by OH or CO₂H;

R³ is selected from the group consisting of H and C_1-6 alkyl;

R^3a is selected from the group consisting of H and C_1-6 alkyl;

R^3b is selected from the group consisting of H and C_1-6 alkyl;

R⁴ is selected from the group consisting of H and C_1-6 alkyl, wherein the C_1-6 alkyl is optionally substituted by OH or CO₂H; and

R⁵ is selected from the group consisting of H and C_1-6 alkyl, wherein the C_1-6 alkyl is optionally substituted by OH or CO₂H.

In some embodiments, L¹ is C(O). In some embodiments, L¹ is PO₂-.

In some embodiments, X¹ is selected from the group consisting of C_1-4 alkylene, C_1-4 alkylene-OC(O)O-C_1-4 alkylene, C_1-4 alkylene-OC(O)NH-C 1 -4 alkylene, C_1-4 alkylene-OC(O)C_1-4 alkylene, C_1-4 alkylene-O-C_1-4 alkylene, C_1-4 alkylene-O-C 1-4 alkylene-O-C 1-4 alkylene, and C_1-4 alkylene-O-C_1-4 alkylene-O-C(O)C_1-4 alkylene, wherein each C_1-4 alkylene is optionally substituted by OH or CO₂H. In some embodiments, X¹ is selected from the group consisting of C_1-4 alkylene, C_1-4 alkylene- OC(O)O-C_1-4 alkylene, C_1-4 alkylene-OC(O)C_1-4 alkylene, C_1-4 alkylene-O-C 1 -4 alkylene, C_1-4 alkylene-O-C_1-4 alkyl ene-O-C 1-4 alkylene, and C_1-4 alkyl ene-O-C 1-4 alkylene-O-C(O)C_1-4 alkylene, wherein each C_1-4 alkylene is optionally substituted by OH or CO₂H.

In some embodiments, X¹ is selected from the group consisting of C_1-4 alkylene, C_1-4 alkylene-OC(O)O-C_1-4 alkylene, C_1-4 alkylene-OC(O)NH-C_1-4 alkylene, C_1-4 alkylene-OC(O)C_1-4 alkylene, C_1-4 alkylene-O-C_1-4 alkylene, C_1-4 alkylene-O-C_1-4 alkylene-O-C_1-4 alkylene, and C_1-4 alkylene-O-C_1-4 alkylene-O-C(O)C_1-4 alkylene, wherein each C_1-4 alkylene is optionally substituted by CO₂H. In some embodiments, X¹ is selected from the group consisting of C_1-4 alkylene, C_1-4 alkylene-OC(O)O-C 1 -4 alkylene, C_1-4 alkylene-OC(O)C_1-4 alkylene, C_1-4 alkylene-O-C_1-4 alkylene, C_1-4 alkylene-O-C_1-4 alkylene-O-C_1-4 alkylene, and C_1-4 alkylene-O-C_1-4 alkylene-O- C(O)C_1-4 alkylene, wherein each C_1-4 alkylene is optionally substituted by CO₂H.

In some embodiments, X¹ is selected from the group consisting of CH₂, CH₂OC(O)OCH₂CH₂, CH₂OC(O)NHCH₂CH₂, CH₂OC(O)OCH₂CH₂CH₂,CH₂OC(O)CH₂CH₂CH₂, and CH₂OCH(CH₃)OCH₂CH₂. In some embodiments, X¹ is selected from the group consisting of CH₂, CH₂OC(O)OCH₂CH₂, CH₂OC(O)OCH₂CH₂CH₂, CH₂OC(O)CH₂CH₂CH₂, and CH₂OCH(CH₃)OCH₂CH₂. In some embodiments, X¹ is CH₂. In some embodiments, X¹ is unsubstituted C_1-2 alkylene and at least one of R³, R^3a, and R^3b is not methyl.

In some embodiments, X² is C_1-3 alkylene which is optionally substituted with OH or CO₂H. In some embodiments, X² is selected from CH₂ and CHCO₂H. In some embodiments, X² is CH₂. In some embodiments, X² is CHCO₂H.

In some embodiments, Y¹ is O. In some embodiments, Y¹ is S. In some embodiments, Y¹ is NR⁵. In some embodiments, Y¹ is NH.

In some embodiments:

Y¹ is NH;

X¹ is unsubstituted C_1-2 alkylene; and at least one of R³, R^3a, and R^3b is not methyl.

In some embodiments:

L¹ is PO₂-;

X¹ is an unsubstituted C_1-2 alkylene;

X² is CH₂;

R² is H; R⁴ is H or methyl; and and at least one of R³, R^3a, and R^3b is not methyl.

In some embodiments, R² is selected from the group consisting of H and C_1-3 alkyl which is optionally substituted by OH or CO₂H. In some embodiments, R² is C_{1- 3} alkyl, which is optionally substituted by OH or CO₂H. In some embodiments, R² is

H.

In some embodiments, R³ is selected from the group consisting of H and C_1-3 alkyl. In some embodiments, R³ is H. In some embodiments, R³ is C_1-3 alkyl. In some embodiments, R³ is methyl.

In some embodiments, R^3a is selected from the group consisting of H and C_1-3 alkyl. In some embodiments, R^3a is H. In some embodiments, R^3a is C 1-₃ alkyl. In some embodiments, R^3a is methyl.

In some embodiments, R^3b is selected from the group consisting of H and C_1-3 alkyl. In some embodiments, R^3b is H. In some embodiments, R^3b is C_1-3 alkyl. In some embodiments, R^3b is methyl.

In some embodiments, two or more of R³, R^3a, and R^3b are the same. In some embodiments, R³, R^3a, and R^3b are the same. In some embodiments, R³, R^3a, and R^3b are each H. In some embodiments, R³, R^3a, and R^3b are each an independently selected C_1-3 alkyl. In some embodiments, R³, R^3a, and R^3b are each methyl.

In some embodiments, R⁴ is selected from the group consisting of H and C1-3 alkyl which is optionally substituted by OH or CO₂H. In some embodiments, R⁴ is C_{1- 3} alkyl, which is optionally substituted by OH or CO₂H. In some embodiments, R⁴ is

H.

In some embodiments:

L¹ is PO₂-;

X¹ is Ci-3 alkylene, which is optionally substituted by OH or CO₂H; X² is C_1-3 alkylene, which is optionally substituted by OH or CO₂H; Y¹ is NH;

R² is selected from the group consisting of H and C_1-3 alkyl which is optionally substituted by OH or CO₂H;

R³ is selected from the group consisting of H and C_1-3 alkyl;

R^3a is selected from the group consisting of H and C_1-3 alkyl; R^3b is selected from the group consisting of H and C_1-3 alkyl; and R⁴ is selected from the group consisting of H and C_1-3 alkyl which is optionally substituted by OH or CO₂H.

In some embodiments:

L¹ is PO₂-;

X¹ is C_1-3 alkylene, which is optionally substituted by CO₂H;

X² is C_1-3 alkylene, which is optionally substituted by OH or CO₂H;

Y¹ is NH;

R² is selected from the group consisting of H and C_1-3 alkyl which is optionally substituted by OH or CO₂H;

R³ is selected from the group consisting of H and C_1-3 alkyl;

R^3a is selected from the group consisting of H and C_1-3 alkyl;

R^3b is selected from the group consisting of H and C_1-3 alkyl; and R⁴ is selected from the group consisting of H and C_1-3 alkyl which is optionally substituted by OH or CO₂H.

In some embodiments:

L¹ is PO₂-;

X¹ is X¹ is selected from the group consisting of CH₂, CH₂OC(O)OCH₂CH₂, CH₂OC(O)NHCH₂CH₂, CH₂OC(O)OCH₂CH₂CH₂, CH₂OC(O)CH₂CH₂CH₂, and CH₂OCH(CH₃)OCH₂CH₂;

X² is C_1-3 alkylene, which is optionally substituted by OH or CO₂H;

Y¹ is NH;

R² is selected from the group consisting of H and C_1-3 alkyl which is optionally substituted by OH or CO₂H;

R³ is selected from the group consisting of H and C_1-3 alkyl;

R^3a is selected from the group consisting of H and C_1-3 alkyl;

R^3b is selected from the group consisting of H and C_1-3 alkyl; and R⁴ is selected from the group consisting of H and C_1-3 alkyl which is optionally substituted by OH or CO₂H.

In some embodiments:

L¹ is PO₂-;

X¹ is C_1-3 alkylene, which is optionally substituted by OH or CO₂H; X² is Ci-3 alkylene, which is optionally substituted by OH or CO₂H;

R² is selected from the group consisting of H and C1-3 alkyl which is optionally substituted by OH or CO₂H;

R³ is selected from the group consisting of H and C_1-3 alkyl;

R^3a is selected from the group consisting of H and C1-3 alkyl; and R^3b is selected from the group consisting of H and C1-3 alkyl.

In some embodiments:

L¹ is PO₂- ;

X¹ is C_1-4 alkylene;

X² is C_1-3 alkylene, which is optionally substituted by CO₂H;

R² is selected from the group consisting of H and C_1-3 alkyl;

R³ is selected from the group consisting of H and C_1-3 alkyl;

R^3a is selected from the group consisting of H and C_1-3 alkyl; and R^3b is selected from the group consisting of H and C_1-3 alkyl.

In some embodiments:

L¹ is PO₂-;

X¹ is C_1-4 alkylene;

X² is Ci-3 alkylene, which is optionally substituted by CO₂H;

Y¹ is NH;

R² is selected from the group consisting of H and C1-3 alkyl;

R³ is selected from the group consisting of H and C_1-3 alkyl;

R^3a is selected from the group consisting of H and C_1-3 alkyl; and R^3b is selected from the group consisting of H and C_1-3 alkyl.

In some embodiments:

L¹ is PO₂-;

X¹ is selected from the group consisting of CH₂, CH₂OC(O)OCH₂CH₂, CH₂OC(O)NHCH₂CH₂, CH₂OC(O)OCH₂CH₂CH₂, CH₂OC(O)CH₂CH₂CH₂, and CH₂OCH(CH3)OCH₂CH₂;

X² is C_1-3 alkylene, which is optionally substituted by CO₂H;

Y¹ is NH;

R² is selected from the group consisting of H and C_1-3 alkyl;

R³ is selected from the group consisting of H and C_1-3 alkyl; R^3a is selected from the group consisting of H and C1-3 alkyl; and R^3b is selected from the group consisting of H and C1-3 alkyl.

In some embodiments:

L¹ is PO₂-;

X¹ is selected from the group consisting of CH₂, CH₂OC(O)OCH₂CH₂, CH₂OC(O)NHCH₂CH₂, CH₂OC(O)OCH₂CH₂CH₂, CH₂OC(O)CH₂CH₂CH₂, and CH₂OCH(CH₃)OCH₂CH₂ ;

X² is C_1-3 alkylene, which is optionally substituted by CO₂H;

Y¹ is NH;

R² is selected from the group consisting of H and C1-3 alkyl;

R³ is selected from the group consisting of H and C1 -3 alkyl;

R^3a is selected from the group consisting of H and C1-3 alkyl; and R^3b is selected from the group consisting of H and C1-3 alkyl; when X¹ is CH₂ then at least one of R³, R^3a, and R^3b is not methyl.

In some embodiments, the compound of Formula I is a compound of Formula

II:

II or a pharmaceutically acceptable salt thereof, wherein variables L¹, X², R³, R^3a, and R^3b are defined according to the definitions provided herein for compounds of Formula I.

In some embodiments, the compound of Formula I is a compound of Formula

III:

III or a pharmaceutically acceptable salt thereof, wherein variables X², R³, R^3a, and R^3b are defined according to the definitions provided herein for compounds of Formula I.

In some embodiments, the compound provided herein is selected from the group consisting of:

or a pharmaceutically acceptable salt thereof.

In some embodiments, the compound provided herein is selected from the group consisting of:

or a pharmaceutically acceptable salt thereof.

In some embodiments, the compound of Formula I is not:

Synthesis

As will be appreciated, the compounds provided herein, including salts thereof, can be prepared using known organic synthesis techniques and can be synthesized according to any of numerous possible synthetic routes.

Accordingly, the present application provides process of preparing a compound of Formula la:

la or a pharmaceutically acceptable salt thereof, comprising reacting a compound of Formula IV:

IV with a compound of Formula V :

V in the presence of a first base and an amine coupling agent, wherein:

R⁶ is C_1-6 alkylene;

R⁷ is C_1-6 alkylene; and each R^a, R^b, R^C is independently selected from the group consisting of H and C_1-6 alkyl.

In some embodiments, the first base is a tri(C_1-6 alkyl) amine base. In some embodiments, the first base is N,N-diisopropylethylamine (DIPEA). In some embodiments, the amine coupling agent is 1 - [bis(dimethylamino)methylene]-lH-l,2,3-triazolo[4,5-b]pyridinium 3 -oxide hexafluorophosphate (HATU).

In some embodiments, the reacting comprises mixing the amine coupling agent with the compound of Formula V to afford a first mixture; and adding the first base and the compound of Formula IV, sequentially, to the first mixture.

In some embodiments, the reacting comprises mixing the amine coupling agent with the compound of Formula V to afford a first mixture; and adding the compound of Formula IV and then the first base, sequentially, to the first mixture.

In some embodiments, the reacting comprises mixing the amine coupling agent with the compound of Formula V to afford a first mixture; and adding the first base and the compound of Formula IV to the first mixture concurrently.

In some embodiments, about 1 to about 5 equivalents (e.g., about 1 equivalent, about 1.5 equivalents, about 2 equivalents, about 2.5 equivalents, about 3 equivalents, about 4 equivalents, or about 5 equivalents) of the compound of Formula IV is used based on 1 equivalent of the compound of Formula V.

In some embodiments, the reacting is performed at a temperature of from about 20°C to about 30°C (i. e. , about room temperature, for example about 20°C, about 21°C, about 22°C, about 23°C, about 24°C, about 25°C, about 26°C, about 27°C, about 28°C, about 29°C, or about 30°C).

In some embodiments, the reacting is performed for about 4 hours to about 40 hours, e.g., about 4 hours, about 6 hours, about 8 hours, about 10 hours, about 12 hours, about 14 hours, about 16 hours, about 18 hours, about 20 hours, about 24 hours, about 28 hours, about 30 hours, about 32 hours, about 36 hours, or about 40 hours.

In some embodiments, the reacting is performed in the presence of a first solvent component. In some embodiments, the first solvent component comprises dimethylformamide (DMF).

In some embodiments, the process further comprises separating and/or isolating the by-products and/or unreacted compounds (i.e., unreacted compound of Formula IV and unreacted compound of Formula V) from the desired product (i.e., compound of Formula la, or a pharmaceutically acceptable salt thereof). In some embodiments, the process further comprises isolating the compound of Formula la, or a pharmaceutically acceptable salt thereof.

In some embodiments, the compound of Formula la is a compound of Formula lb

lb or a pharmaceutically acceptable salt thereof.

The present application further provides a process of preparing a compound of

Formula lb:

lb or a pharmaceutically acceptable salt thereof, comprising reacting a compound of Formula IVa:

IVa with a compound of Formula V:

V in the presence of Ν,Ν-diisopropylethylamine (DIPEA) and 1- [bis(dimethylamino)methylene]-lH-1,2,3-triazolo[4,5-b]pyridinium 3-oxide hexafluorophosphate (HATU).

The present application provides a process of preparing a compound of

Formula Ic:

Ic or a pharmaceutically acceptable salt thereof, comprising reacting a compound of Formula VI:

VI with a compound of Formula VII:

VII in the presence of a second base, wherein:

Pg¹ is an amine protecting group;

R⁶ is C_1-6 alkylene;

R⁷ is C_1-6 alkylene, wherein the C_1-6 alkylene is optionally substituted OH or CO₂H; and

R⁸ is a carboxylic acid activating group.

In some embodiments, the second base is an amine base. In some embodiments, the second base is tri(C_1-6 alkyl) amine base. In some embodiments, the second base is triethylamine.

In some embodiments, R⁸ is 2,5 -dioxopyrrolidin-1-yl.

In some embodiments, the reacting comprises adding the second base and the compound of Formula VI, sequentially, to the compound of Formula VII.

In some embodiments, the reacting comprises adding the compound of Formula VI and then the second base, sequentially, to the compound of Formula VII.

In some embodiments, the reacting comprises adding the second base and the compound of Formula VI to the compound of Formula VII concurrently.

In some embodiments, the compound of Formula VII may be dissolved in a first solvent component that will be described below. It is therefore to be understood that the compound of Formula VII is partially or fully dissolved in said first solvent component. In some embodiments, about 1 to about 5 equivalents (e.g. , about 1 equivalent, about 1.5 equivalents, about 2 equivalents, about 2.5 equivalents, about 3 equivalents, about 4 equivalents, or about 5 equivalents) of the compound of Formula VI is used based on 1 equivalent of the compound of Formula VII.

In some embodiments, the reacting is performed at a temperature of from about 20°C to about 30°C (i.e., about room temperature, for example about 20°C, about 21°C, about 22°C, about 23°C, about 24°C, about 25°C, about 26°C, about 27°C, about 28°C, about 29°C, or about 30°C).

In some embodiments, the reacting is performed for about 4 hours to about 40 hours, e.g., about 4 hours, about 6 hours, about 8 hours, about 10 hours, about 12 hours, about 14 hours, about 16 hours, about 18 hours, about 20 hours, about 24 hours, about 28 hours, about 30 hours, about 32 hours, about 36 hours, or about 40 hours.

In some embodiments, the reacting is performed in the presence of a first solvent component. In some embodiments, the first solvent component comprises dimethylformamide (DMF).

In some embodiments, the process further comprises separating and/or isolating the by-products and/or unreacted compounds (i.e., unreacted compound of Formula IV and unreacted compound of Formula V) from the desired product (i.e., compound of Formula la, or a pharmaceutically acceptable salt thereof). In some embodiments, the process further comprises isolating the compound of Formula la, or a pharmaceutically acceptable salt thereof.

In some embodiments, the compound of Formula Ic is a compound of Formula

Id or le:

Id

Ie or a pharmaceutically acceptable salt thereof The present application further provides a process of preparing a compound of

Formula Id:

Id or a pharmaceutically acceptable salt thereof, comprising reacting a compound of Formula Via:

VIa with a compound of Formula Vila:

VIIa in the presence of triethylamine.

The present application further provides a process of preparing a compound of

Formula Ie:

Ie or a pharmaceutically acceptable salt thereof, comprising reacting a compound of Formula VIb:

VIb with a compound of Formula VIIa:

Vila in the presence of triethylamine.

In some embodiments, the process for preparing compounds of Formula Ic, Id and Ie, further comprising a deprotection step in the presence of a deprotecting agent. In some embodiments, the deprotecting agent comprises an acid. In some embodiments, the acid is trifluoroacetic acid (TFA).

In some embodiments, the deprotection step is performed in the presence of a second solvent component. In some embodiments, the second solvent component comprises dichloromethane (DCM).

In some embodiments, the deprotection step is performed at a temperature of from about 20°C to about 30°C (i.e., about room temperature, for example about 20°C, about 21°C, about 22°C, about 23°C, about 24°C, about 25°C, about 26°C, about 27°C, about 28°C, about 29°C, or about 30°C).

In some embodiments, the deprotection step is performed for about 1 hour to about 40 hours, e.g., about 1 hour, about 2 hours, about 4 hours, about 6 hours, about 8 hours, about 10 hours, about 12 hours, about 14 hours, about 16 hours, about 18 hours, about 20 hours, about 24 hours, about 28 hours, about 30 hours, about 32 hours, about 36 hours, or about 40 hours.

In some embodiments, the process further comprises isolating the compounds of Formula Ic, Id and Ie, or a pharmaceutically acceptable salt thereof.

The present application further provides a compound of Formula Vila:

Vila or salt thereof.

In some embodiments, the compounds provided herein can be prepared according to the general procedures shown in Schemes 1-4 (where Pg¹ is an amine protecting group such as Boc), using appropriately substituted starting materials. Scheme 1.

Compound 1

Scheme 2.

Compound 2

Scheme 3.

Compound 3

Scheme 4.

Compound 4

It will be appreciated by one skilled in the art that the processes described are not the exclusive means by which compounds provided herein may be synthesized and that a broad repertoire of synthetic organic reactions is available to be potentially employed in synthesizing compounds provided herein. The person skilled in the art knows how to select and implement appropriate synthetic routes. Suitable synthetic methods of starting materials, intermediates and products may be identified by reference to the literature, including reference sources such as: Advances in Heterocyclic Chemistry, Vols. 1-107 (Elsevier, 1963-2012); Journal of Heterocyclic Chemistry Vols. 1-49 (Journal of Heterocyclic Chemistry, 1964-2012); Carreira, et al. (Ed.) Science of Synthesis, Vols. 1-48 (2001-2010) and Knowledge Updates KU2010/1-4; 2011/1-4; 2012/1-2 (Thieme, 2001-2012); Katritzky, et al. (Ed.) Comprehensive Organic Functional Group Transformations, (Pergamon Press, 1996); Katritzky et al. (Ed.); Comprehensive Organic Functional Group Transformations II (Elsevier, 2^nd Edition, 2004); Katritzky et al. (Ed.), Comprehensive Heterocyclic Chemistry (Pergamon Press, 1984); Katritzky et al., Comprehensive Heterocyclic Chemistry II, (Pergamon Press, 1996); Smith et al., March's Advanced Organic Chemistry: Reactions, Mechanisms, and Structure, 6^th Ed. (Wiley, 2007); Trost et al. (Ed.), Comprehensive Organic Synthesis (Pergamon Press, 1991).

Preparation of compounds described herein can involve the protection and deprotection of various chemical groups. The need for protection and deprotection, and the selection of appropriate protecting groups can be readily determined by one skilled in the art. The chemistry of protecting groups can be found, for example, in T. W. Greene and P. G. M. Wuts, Protective Groups in Organic Synthesis, 3^rd Ed., Wiley & Sons, Inc., New York (1999).

Reactions can be monitored according to any suitable method known in the art. For example, product formation can be monitored by spectroscopic means, such as nuclear magnetic resonance spectroscopy (e.g., ¹H or ¹³C), infrared spectroscopy, spectrophotometry (e.g., UV-visible), mass spectrometry, or by chromatographic methods such as high performance liquid chromatography (HPLC), liquid chromatography-mass spectroscopy (LCMS), or thin layer chromatography (TLC). Compounds can be purified by those skilled in the art by a variety of methods, including high performance liquid chromatography (HPLC) and normal phase silica chromatography.

As used herein, the term “reacting” is used as known in the art and generally refers to the bringing together of chemical reagents in such a manner so as to allow their interaction at the molecular level to achieve a chemical or physical transformation. In some embodiments, the reacting involves two reagents, wherein one or more equivalents of second reagent are used with respect to the first reagent. The reacting steps of the processes described herein can be conducted for a time and under conditions suitable for preparing the identified product.

As used herein, the term “amine base” refers to a mono-substituted amine group (i.e., primary amine base), di-substituted amine group (i.e., secondary amine base), or a tri-substituted amine group (i.e., tertiary amine base). Exemplary amine bases include, bt are not limited to, methyl amine, ethyl amine, propyl amine, butyl amine, dimethylamine, diethylamine, dipropylamine, dibutylamine, trimethylamine, triethylamine, tripropylamine, triisopropylamine, tributylamine, tri-tert-butylamine, Ν,Ν-dimethylethanamine, N,N-diisopropylethylamine (DIPEA), pyridine, and the like.

As used herein, the term “Pg” refers to a protecting group (e.g., an amine protecting groups). Exemplary amine protecting groups include, but are not limited to, benzyloxycarbonyl (Cbz), 2,2,2-trichloroethoxycarbonyl (Troc), 2- (trimethylsilyl)ethoxycarbonyl (Teoc), 2-(4- trifluoromethylphenylsulfonyl)ethoxycarbonyl (Tsc), t-butoxycarbonyl (BOC), 1- adamantyloxycarbonyl (Adoc), 2-adamantylcarbonyl (2-Adoc), 2,4-dimethylpent-3 - yloxycarbonyl (Doc), cyclohexyloxycarbonyl (Hoc), 1,1 -dimethyl-2, 2,2- trichloroethoxycarbonyl (TcBOC), vinyl, 2-chloroethyl, 2-phenylsulfonylethyl, allyl, benzyl, 2-nitrobenzyl, 4-nitrobenzyl, diphenyl-4-pyridylmethyl, N’,N’- dimethylhydrazinyl, methoxymethyl, t-butoxymethyl (Bum), benzyloxymethyl (BOM), or 2-tetrahydropyranyl (THP), tri(C_1-4 alkyl)silyl (e.g., tri(isopropyl)silyl),

1 , 1 -diethoxymethyl, or N -pivaloyloxymethyl (POM). In some embodiments, the protecting group is benzyloxycarbonyl (Cbz).

As used herein, the terms “deprotecting” or “deprotection conditions” refer to conditions suitable to cleave a protecting group (e.g. , an amine protecting group). In some embodiments, deprotection conditions may include cleavage of a protecting group in the presence of a strong acid, in the presence of a strong base, in the presence of a reducing agent, or in the presence of an oxidizing agent. Deprotection of an amine protecting group can be accomplished by methods known in the art for the removal of particular protecting groups for amines, such as those in Wuts and Greene, Protective Groups in Organic Synthesis, 4th ed., John Wiley & Sons: New Jersey, pages 696-887 (and, in particular, pages 872-887) (2007), the disclosure of which is incorporated herein by reference in its entirety.

The reactions of the processes described herein can be carried out in suitable solvents which can be readily selected by one of skill in the art of organic synthesis. Suitable solvents can be substantially nonreactive with the starting materials (reactants), the intermediates, or products at the temperatures at which the reactions are carried out, e.g., temperatures which can range from the solvent's freezing temperature to the solvent's boiling temperature. A given reaction can be carried out in one solvent or a mixture of more than one solvent. Depending on the particular reaction step, suitable solvents for a particular reaction step can be selected. In some embodiments, reactions can be carried out in the absence of solvent, such as when at least one of the reagents is a liquid or gas.

As used herein, a “solvent component” refers to one solvent or a mixture of two or more solvents.

As used herein, “second”, “third, " " fourth”, etc. as a prefix to the phrase “solvent component” or “amine base” is used to differentiate the solvent component or amine base from other solvent components or amine bases used in earlier or later steps of the process and does not indicate that multiple solvents or bases must be present.

Exemplary halogenated solvents include, but are not limited to, carbon tetrachloride, chloroform, dichloromethane, and the like.

Exemplary ether solvents include, but are not limited to, tetrahydrofuran, 1,3- dioxane, 1 ,4-dioxane, furan, diethyl ether, ethylene glycol dimethyl ether, ethylene glycol diethyl ether, diethylene glycol dimethyl ether (diglyme), t-butyl methyl ether, and the like.

Exemplary protic solvents include, but are not limited to, water, methanol (MeOH), ethanol (EtOH), ethylene glycol, 1 -propanol, 2-propanol, 1 -butanol, 2- butanol, i-butyl alcohol, t-butyl alcohol, and the like.

Exemplary aprotic solvents include, but are not limited to, tetrahydrofuran (THF), N,N -dimethylformamide (DMF), N,N -dimethylacetamide (DMA), 1,3- dimethyl-3 ,4,5,6-tetrahydro-2( lH)-pyrimidinone (DMPU), 1 ,3 -dimethyl-2 - imidazolidinone (DMI), N-methylpyrrolidinone (NMP), formamide, N- methylacetamide, N-methylformamide, acetonitrile (ACN), dimethyl sulfoxide (DMSO), acetone, ethyl methyl ketone, ethyl acetate (EtOAc), and the like.

Exemplary hydrocarbon solvents include, but are not limited to, benzene, cyclohexane, pentane, hexane, toluene, and the like.

The reactions of the processes described herein can be carried out in air or under an inert atmosphere. Typically, reactions containing reagents or products that are substantially reactive with air can be carried out using air-sensitive synthetic techniques that are well known to the skilled artisan.

In some embodiments, preparation of compounds can involve the addition of acids or bases to affect, for example, catalysis of a desired reaction or formation of salt forms such as acid addition salts.

Exemplary inorganic acids include, but are not limited to, hydrochloric acid, hydrobromic acid, sulfuric acid, phosphoric acid, nitric acid, and the like. Exemplary organic acids include, but are not limited to, formic acid, acetic acid, propionic acid, butanoic acid, benzoic acid, methanesulfonic acid, p-toluenesulfonic acid, benzenesulfonic acid, trifluoroacetic acid (TFA), and the like. Exemplary bases include, but are not limited to, lithium hydroxide, sodium hydroxide, potassium hydroxide, lithium carbonate, sodium carbonate, potassium carbonate, and sodium bicarbonate. Exemplary strong bases include, but are not limited to, hydroxide, alkoxides, metal amides, metal hydrides, metal dialkylamides, and arylamines, wherein; alkoxides include lithium, sodium and potassium salts of methyl, ethyl and t-butyl oxides; metal amides include sodium amide, potassium amide and lithium amide; metal hydrides include sodium hydride, potassium hydride and lithium hydride; and metal dialkylamides include lithium, sodium, and potassium salts of methyl, ethyl, n-propyl, i-propyl, n-butyl, t-butyl, trimethylsilyl and cyclohexyl substituted amides.

At various places in the present specification, divalent linking substituents are described. It is specifically intended that each divalent linking substituent include both the forward and backward forms of the linking substituent. For example, - NR(CR’R”)n- includes both -NR(CR’R”)_n- and -(CR’R”)_nNR-. Where the structure clearly requires a linking group, the Markush variables listed for that group are understood to be linking groups.

At various places in the present specification, substituents of compounds of the invention are disclosed in groups or in ranges. It is specifically intended that the invention include each and every individual subcombination of the members of such groups and ranges. For example, the term “C_1-6 alkyl” is specifically intended to individually disclose methyl, ethyl, C₃ alkyl, C₄ alkyl, C₅ alkyl, and C₆ alkyl.

As used herein, the phrase “optionally substituted” means unsubstituted or substituted. As used herein, the term “substituted” means that one or more hydrogen atoms are removed and replaced by one or more substituents. It is to be understood that substitution at a given atom is limited by valency.

Throughout the definitions, the term “C_n-m” indicates a range which includes the endpoints, wherein n and m are integers and indicate the number of carbons. Examples include C_1-4, _C1-6, and the like.

As used herein, the term “C_n-m alkyl ene”, employed alone or in combination with other terms, refers to a divalent alkyl linking group having n to m carbons. Examples of alkyl ene groups include, but are not limited to, methylene, ethan-1,2- diyl, propan- 1,3 -diyl, propan- 1 ,2-diyl, and the like. In some embodiments, the alkylene moiety contains 1 to 6, 1 to 3, or 1 to 2 carbon atoms.

As used herein, the term “C_n-_m alkyl”, employed alone or in combination with other terms, refers to a saturated hydrocarbon group that may be straight-chain or branched, having n to m carbons. Examples of alkyl moieties include, but are not limited to, chemical groups such as methyl, ethyl, n-propyl, isopropyl, n-butyl, tert- butyl, isobutyl, sec-butyl; higher homologs such as 2 -methyl- 1 -butyl, n-pentyl, 3- pentyl, n-hexyl, 1 ,2,2-trimethylpropyl, and the like. In some embodiments, the alkyl group contains from 1 to 6 carbon atoms, from 1 to 4 carbon atoms, from 1 to 3 carbon atoms, or 1 to 2 carbon atoms.

The term “compound” as used herein is meant to include all stereoisomers, geometric isomers, tautomers, and isotopes of the structures depicted. Compounds herein identified by name or structure as one particular tautomeric form are intended to include other tautomeric forms unless otherwise specified.

Compounds provided herein also include tautomeric forms. Tautomeric forms result from the swapping of a single bond with an adjacent double bond together with the concomitant migration of a proton. Tautomeric forms include prototropic tautomers which are isomeric protonation states having the same empirical formula and total charge. Example prototropic tautomers include ketone — enol pairs, amide - imidic acid pairs, lactam - lactim pairs, enamine - imine pairs, and annular forms where a proton can occupy two or more positions of a heterocyclic system, for example, 1H- and 3H-imidazole, 1H-, 2H- and 4H- 1 ,2,4-triazole, 1H- and 2H- isoindole, and 1H- and 2H-pyrazole. Tautomeric forms can be in equilibrium or sterically locked into one form by appropriate substitution.

All compounds, and pharmaceutically acceptable salts thereof, can be found together with other substances such as water and solvents (e.g. hydrates and solvates) or can be isolated.

In some embodiments, preparation of compounds can involve the addition of acids or bases to affect, for example, catalysis of a desired reaction or formation of salt forms such as acid addition salts.

Example acids can be inorganic or organic acids and include, but are not limited to, strong and weak acids. Some example acids include hydrochloric acid, hydrobromic acid, sulfuric acid, phosphoric acid, p-toluenesulfonic acid, 4- nitrobenzoic acid, methanesulfonic acid, benzenesulfonic acid, trifluoroacetic acid, and nitric acid. Some weak acids include, but are not limited to acetic acid, propionic acid, butanoic acid, benzoic acid, tartaric acid, pentanoic acid, hexanoic acid, heptanoic acid, octanoic acid, nonanoic acid, and decanoic acid.

Example bases include lithium hydroxide, sodium hydroxide, potassium hydroxide, lithium carbonate, sodium carbonate, potassium carbonate, and sodium bicarbonate. Some example strong bases include, but are not limited to, hydroxide, alkoxides, metal amides, metal hydrides, metal dialkylamides and arylamines, wherein; alkoxides include lithium, sodium and potassium salts of methyl, ethyl and t-butyl oxides; metal amides include sodium amide, potassium amide and lithium amide; metal hydrides include sodium hydride, potassium hydride and lithium hydride; and metal dialkylamides include lithium, sodium, and potassium salts of methyl, ethyl, n-propyl, iso-propyl, n-butyl, tert- butyl, trimethylsilyl and cyclohexyl substituted amides.

In some embodiments, the compounds and salts provided herein are substantially isolated. By “substantially isolated” is meant that the compound is at least partially or substantially separated from the environment in which it was formed or detected. Partial separation can include, for example, a composition enriched in the compounds provided herein. Substantial separation can include compositions containing at least about 50%, at least about 60%, at least about 70%, at least about 80%, at least about 90%, at least about 95%, at least about 97%, or at least about 99% by weight of the compounds provided herein, or salt thereof. Methods for isolating compounds and their salts are routine in the art.

The term, “room temperature” or “RT” as used herein, are understood in the art, and refer generally to a temperature (e.g., a reaction temperature) that is about the temperature of the room in which the reaction is carried out, for example, a temperature from about 20°C to about 30°C.

The compounds described herein can be asymmetric (e.g., having one or more stereocenters). All stereoisomers, such as enantiomers and diastereomers, are intended unless otherwise indicated. Compounds of the present invention that contain asymmetrically substituted carbon atoms can be isolated in optically active or racemic forms. Methods on how to prepare optically active forms from optically inactive starting materials are known in the art, such as by resolution of racemic mixtures or by stereoselective synthesis. Many geometric isomers of olefins, C=N double bonds, and the like can also be present in the compounds described herein, and all such stable isomers are contemplated in the present invention. Cis and trans geometric isomers of the compounds of the present invention are described and may be isolated as a mixture of isomers or as separated isomeric forms.

Resolution of racemic mixtures of compounds can be carried out by any of numerous methods known in the art. An example method includes fractional recrystallizaion using a chiral resolving acid which is an optically active, salt-forming organic acid. Suitable resolving agents for fractional recrystallization methods are, for example, optically active acids, such as the D and L forms of tartaric acid, diacetyltartaric acid, dibenzoyltartaric acid, mandelic acid, malic acid, lactic acid or the various optically active camphorsulfonic acids such as β-camphorsulfonic acid. Other resolving agents suitable for fractional crystallization methods include stereoisomerically pure forms of a-methylbenzylamine (e.g., S and R forms, or diastereomerically pure forms), 2-phenylglycinol, norephedrine, ephedrine, N- methylephedrine, cyclohexyl ethylamine, 1 ,2-diaminocyclohexane, and the like.

Resolution of racemic mixtures can also be carried out by elution on a column packed with an optically active resolving agent (e.g., dinitrobenzoylphenylglycine). Suitable elution solvent composition can be determined by one skilled in the art.

The phrase “pharmaceutically acceptable” is employed herein to refer to those compounds, materials, compositions, and/or dosage forms which are, within the scope of sound medical judgment, suitable for use in contact with the tissues of human beings and animals without excessive toxicity, irritation, allergic response, or other problem or complication, commensurate with a reasonable benefit/risk ratio.

The present application also includes pharmaceutically acceptable salts of the compounds described herein. As used herein, “pharmaceutically acceptable salts” refers to derivatives of the disclosed compounds wherein the parent compound is modified by converting an existing acid or base moiety to its salt form. Examples of pharmaceutically acceptable salts include, but are not limited to, mineral or organic acid salts of basic residues such as amines; alkali or organic salts of acidic residues such as carboxylic acids; and the like. The pharmaceutically acceptable salts of the present application include the conventional non-toxic salts of the parent compound formed, for example, from non-toxic inorganic or organic acids. The pharmaceutically acceptable salts of the present application can be synthesized from the parent compound which contains a basic or acidic moiety by conventional chemical methods. Generally, such salts can be prepared by reacting the free acid or base forms of these compounds with a stoichiometric amount of the appropriate base or acid in water or in an organic solvent, or in a mixture of the two; generally, non-aqueous media like ether, ethyl acetate, alcohols (e.g. , methanol, ethanol, iso-propanol, or butanol) or acetonitrile (MeCN) are preferred. Lists of suitable salts are found in Remington’s Pharmaceutical Sciences, 17th ed., Mack Publishing Company, Easton, Pa., 1985, p. 1418 and Journal of Pharmaceutical Science, 66, 2 (1977). Conventional methods for preparing salt forms are described, for example, in Handbook of Pharmaceutical Salts: Properties, Selection, and Use, Wiley- VCH, 2002.

Methods of Use

The present application further provides methods of treating a disease or disorder in a subject. As used herein, the term “subject,” refers to any animal, including mammals. Exemplary subjects include, but are not limited to, mice, rats, rabbits, dogs, cats, swine, cattle, sheep, horses, primates, and humans. In some embodiments, the subject is a human. In some embodiments, the method comprises administering to the subject (e.g. , a subject in need thereof) a therapeutically effective amount of a compound provided herein (e.g. , a compound of Formula I), or a pharmaceutically acceptable salt thereof.

In some embodiments, the disease or disorder is selected from the group consisting of pain, a pain-related disease or disorder, a mood disease or disorder, a disease or disorder of the central nervous system, an optical disease or disorder, cancer, a gastrointestinal disease or disorder, a renal disease or disorder, a renal- related disease or disorder, a cardiovascular disease or disorder, and a skin disease or disorder. In some embodiments, the disease or disorder is pain or a pain-related disease or disorder. In some embodiments, the pain or a pain-related disease or disorder is selected from the group consisting of acute pain, chronic pain, neuropathic pain, nociceptive pain, inflammatory pain, cancer pain, fibromyalgia, rheumatoid arthritis, osteoarthritis, surgery-related pain, and osteoporosis. In some embodiments, the disease or disorder is selected from the group consisting of rheumatoid arthritis and osteoarthritis. In some embodiments, the disease or disorder is pain related to rheumatoid arthritis or pain related to osteoarthritis.

In some embodiments, the disease or disorder is a mood disease or disorder. In some embodiments, the mood disease or disorder is selected from the group consisting of anxiety, depression, a sleeping disorder, an eating disorder, post- traumatic stress disorder, symptoms of drug or alcohol withdrawal or abuse, schizophrenia, obsessive-compulsive disorder, bipolar disorder, sexual dysfunction, attention deficit disorder (ADD), and attention deficit hyperactivity disorder (ADHD).

In some embodiments, the disease or disorder is a disease or disorder of the central nervous system, a disease or disorder of the peripheral nervous system, or an optical disease or disorder. In some embodiments, the disease or disorder is a disease or disorder of the central nervous system. In some embodiments, the disease or disorder is a disease or disorder of the peripheral nervous system. In some embodiments, the disease or disorder is an optical disease or disorder. In some embodiments, the disease or disorder of the central nervous system, peripheral nervous system, or optical disease or disorder is selected from the group consisting of a demyelinating disease, glaucoma, age-related macular degeneration (AMD), amyotrophic lateral sclerosis (ALS), a cognitive disorder, Alzheimer’s disease, a movement disorder, Huntington’s chorea, Tourette’s syndrome, Niemann-Pick disease, Parkinson's disease, epilepsy, a cerebrovascular disorder, ischemic stroke, and brain injury.

In some embodiments, the demyelinating disease is selected from the group consisting of multiple sclerosis (MS), neuromyelitis optica (NMO), Devic’s disease, central nervous system neuropathy, central pontine myelinolysis, syphilitic myelopathy, leukoencephalopathies, leukodystrophies, Guillain-Barre syndrome, chronic inflammatory demyelinating polyneuropathy, anti -myelin-associated glycoprotein (MAG) peripheral neuropathy, Charcot-Marie-Tooth disease, peripheral neuropathy, myelopathy, optic neuropathy, progressive inflammatory neuropathy, optic neuritis, and transverse myelitis.

In some embodiments, the disease or disorder is cancer. In some embodiments, the cancer is selected from the group consisting of leukemia, mantle cell lymphoma, Hodgkin lymphoma, Non-Hodgkin lymphoma, hepatocellular carcinoma, ovarian cancer, colorectal cancer, pancreatic cancer, prostate cancer, breast cancer, glioma, skin cancer, renal carcinoma, and lung cancer.

In some embodiments, the disease or disorder is a gastrointestinal disease or disorder. In some embodiments, the gastrointestinal disease or disorder is selected from the group consisting of inflammatory bowel disease, gastroesophageal reflux disease, paralytic ileus, secretory diarrhoea, gastric ulcer, nausea, emesis, celiac disease, irritable bowel syndrome, and a liver disorder.

In some embodiments, the liver disease is selected from the group consisting of acute liver failure, Alagille syndrome, hepatitis, enlarged liver, Gilbert’s syndrome, liver cyst, liver haemangioma, fatty liver disease, steatohepatitis, primary sclerosing cholangitis, fascioliasis, primary bilary cirrhosis, Budd-Chiari syndrome, hemochromatosis, Wilson’s disease, and transthyretin-related hereditary amyloidosis.

In some embodiments, the disease or disorder is a renal disease or disorder or a renal-related disease or disorder. In some embodiments, the renal disease or disorder or a renal-related disease or disorder is selected from the group consisting of diabetes, diabetic nephropathy, acute inflammatory kidney injury, renal ischemia urinary incontinence, and overactive bladder.

In some embodiments, the disease or disorder is a skin disease or disorder. In some embodiments, the skin disease or disorder is selected from the group consisting of atopic dermatitis, psoriasis, and lupus.

In some embodiments, the disease or disorder is a cardiovascular disease or disorder. In some embodiments, the cardiovascular disease or disorder is selected from the group consisting of cardiovascular disease, vascular inflammation, idiopathic pulmonary fibrosis, cough, and hypertension. The present application further provides a compound described herein, or a pharmaceutically acceptable salt thereof, for use in any of the methods described herein.

The present application further provides use of a compound described herein, or a pharmaceutically acceptable salt thereof, for the preparation of a medicament for use in any of the methods described herein.

As used herein, the phrase “therapeutically effective amount” refers to the amount of active compound or pharmaceutical agent that elicits the biological or medicinal response that is being sought in a tissue, system, animal, individual or human by a researcher, veterinarian, medical doctor or other clinician.

As used herein, the term “treating” or “treatment” refers to one or more of: (1) inhibiting the disease; for example, inhibiting a disease, condition or disorder in an individual who is experiencing or displaying the pathology or symptomatology of the disease, condition or disorder (i.e., arresting further development of the pathology and/or symptomatology); and (2) ameliorating the disease; for example, ameliorating a disease, condition or disorder in an individual who is experiencing or displaying the pathology or symptomatology of the disease, condition or disorder (i.e., reversing the pathology and/or symptomatology) such as decreasing the severity of disease or reducing or alleviating one or more symptoms of the disease.

Labeled Compounds

The present application further provides isotopically-labeled compounds provided herein. An “isotopically” or “radio-labeled” compound is a compound wherein one or more atoms are replaced or substituted by an atom having an atomic mass or mass number different from the atomic mass or mass number typically found in nature (i.e., naturally occurring). Suitable radionuclides that may be incorporated in compounds of the present disclosure include but are not limited to ²H (also written as D for deuterium), ³H (also written as T for tritium), ¹¹C, ¹³C, ¹⁴C, ¹³N, ¹⁵N, ¹⁵O, ¹⁷O, and ¹⁸O. For example, one or more hydrogen atoms in a compound of the present disclosure can be replaced by deuterium atoms (e.g., one or more hydrogen atoms of a C_1-6 alkyl group as described herein can be optionally substituted with deuterium atoms, such as -CD₃ being substituted for -CH₃). In some embodiments, alkyl groups of the componuds provided herein can be perdeuterated.

One or more constituent atoms of the compounds presented herein can be replaced or substituted with isotopes of the atoms in natural or non-natural abundance. In some embodiments, the compound provided herein comprises at least one deuterium atom. In some embodiments, the compound provided herein comprises two or more deuterium atoms. In some embodiments, the compound provided herein comprises 1-2, 1-3, 1-4, 1-5, 1-6, 1-8, 1-10, 1-12, 1-14, 1-16, 1-18, or 1-20 deuterium atoms. In some embodiments, all of the hydrogen atoms in a compound provided herein are replaced or substituted by deuterium atoms.

Synthetic methods for including isotopes into organic compounds are known in the art (Deuterium Labeling in Organic Chemistry by Alan F. Thomas (New York, N.Y., Appleton-Century-Crofts, 1971; The Renaissance of H/D Exchange by Jens Atzrodt, Volker Derdau, Thorsten Fey and Jochen Zimmermann, Angew. Chem. Int. Ed. 2007, 7744-7765; The Organic Chemistry of Isotopic Labelling by James R. Hanson, Royal Society of Chemistry, 2011). Isotopically labeled compounds can be used in various studies such as NMR spectroscopy, metabolism experiments, and/or assays. In some embodiments, the present application further provides synthetic methods for incorporating radio-isotopes into the compounds described herein. Synthetic methods for incorporating radio-isotopes into organic compounds are well known in the art, and an ordinary skill in the art will readily recognize the methods applicable for the compounds described herein.

Substitution with heavier isotopes, such as deuterium, may afford certain therapeutic advantages resulting from greater metabolic stability, for example, increased in vivo half-life or reduced dosage requirements, and hence may be preferred in some circumstances, (see e.g., A. Kerekes et. al. J. Med. Chem. 2011, 54, 201-210; R. Xu et. al. J. Label Compd. Radiopharm. 2015, 58, 308-312). In particular, substitution at one or more metabolism sites may afford one or more of the therapeutic advantages. Combination Therapies

One or more additional therapeutic agents such as, for example, chemotherapeutic agents, anesthetics (e.g., for use in combination with a surgical procedure), or other agents useful for treating the diseases or disorders provided herein can be used in combination with the compounds and salts provided herein.

Exemplary compounds that may be useful in combination therapies include, but are not limited to, compounds disclosed in International Application No.: PCT/US2020/013150, the disclosure of which is incorporated herein by reference its entirety. In some embodiments, the compounds provided herein are administered in combination with a compound selected from the group consisting of:

or a pharmaceutically acceptable salt thereof.

In some embodiments, the compounds provided herein are administered in combination with a compound selected from the group consisting of anandamide, 2- arachidonoyl glycerol, and oleoylethanolamide, or a pharmaceutically acceptable salt thereof. In some embodiments, the compounds provided herein are administered in combination with anandamide, or a pharmaceutically acceptable salt thereof. In some embodiments, the compounds provided herein are administered in combination with 2-arachidonoyl glycerol, or a pharmaceutically acceptable salt thereof. In some embodiments, the compounds provided herein are administered in combination with oleoylethanolamide, or a pharmaceutically acceptable salt thereof. Exemplary anesthetics include, but are not limited to, local anesthetics (e.g. , lidocaine, procain, ropivacaine) and general anesthetics (e.g., desflurane, enflurane, halothane, isoflurane, methoxyflurane, nitrous oxide, sevoflurane, amobarbital, methohexital, thiamylal, thiopental, diazepam, lorazepam, midazolam, etomidate, ketamine, propofol, alfentanil, fentanyl, remifentanil, buprenorphine, butorphanol, hydromorphone levorphanol, meperidine, methadone, morphine, nalbuphine, oxymorphone, and pentazocine).

In some embodiments, the additional therapeutic agent is administered simultaneously with the compound or salt provided herein. In some embodiments, the additional therapeutic agent is administered after administration of the compound or salt provided herein. In some embodiments, the additional therapeutic agent and compound or salt provided herein are administered concurrently. In some embodiments, the additional therapeutic agent is administered prior to administration of the compound or salt provided herein. In some embodiments, the additional therapeutic agent and compound or salt provided herein are administered sequentially.

In some embodiments, the compound or salt provided herein is administered during a surgical procedure. In some embodiments, the compound or salt provided herein is administered in combination with an additional therapeutic agent during a surgical procedure.

In some embodiments, there is also provided methods of using a combination therapy, comprising a compound or salt provided herein and one or more additional therapeutic agents such as anandamide, 2-arachidonoyl glycerol, and/or oleoylethanolamide, or a pharmaceutically acceptable salt thereof, to treat a disease or disorder as described herein, in a subject.

Pharmaceutical Formulations

When employed as pharmaceuticals, the compounds and salts provided herein can be administered in the form of pharmaceutical compositions. These compositions can be prepared as described herein or elsewhere, and can be administered by a variety of routes, depending upon whether local or systemic treatment is desired and upon the area to be treated. Administration may be topical (e.g. , transdermal, epidermal, ophthalmic and to mucous membranes including intranasal, vaginal and rectal delivery), pulmonary (e.g. , by inhalation or insufflation of powders or aerosols, including by nebulizer; intratracheal or intranasal), oral, or parenteral. Parenteral administration includes intravenous, intraarterial, subcutaneous, intraperitoneal intramuscular or injection or infusion; or intracranial (e.g., intrathecal or intraventricular, administration).

Parenteral administration can be in the form of a single bolus dose, or may be, for example, by a continuous perfusion pump.

In some embodiments, the compounds provided herein (e.g., compounds of Formula I) are suitable for parenteral administration. In some embodiments, the compounds provided herein (e.g., the compounds of Formula I) are suitable for intravenous administration. Pharmaceutical compositions and formulations for topical administration may include transdermal patches, ointments, lotions, creams, gels, drops, suppositories, sprays, liquids and powders. Conventional pharmaceutical carriers, aqueous, powder or oily bases, thickeners and the like may be necessary or desirable. In some embodiments, the pharmaceutical compositions provided herein are suitable for parenteral administration. In some embodiments, the compositions provided herein are suitable for intravenous administration.

Also provided are pharmaceutical compositions which contain, as the active ingredient, a compound provided herein, or a pharmaceutically acceptable salt thereof, in combination with one or more pharmaceutically acceptable carriers (excipients). In making the compositions provided herein, the active ingredient is typically mixed with an excipient, diluted by an excipient or enclosed within such a carrier in the form of, for example, a capsule, sachet, paper, or other container. When the excipient serves as a diluent, it can be a solid, semi-solid, or liquid material, which acts as a vehicle, carrier or medium for the active ingredient. Thus, the compositions can be in the form of tablets, pills, powders, lozenges, sachets, cachets, elixirs, suspensions, emulsions, solutions, syrups, aerosols (as a solid or in a liquid medium), ointments, soft and hard gelatin capsules, suppositories, sterile injectable solutions, and sterile packaged powders.

Some examples of suitable excipients include, without limitation, lactose, dextrose, sucrose, sorbitol, mannitol, starches, gum acacia, calcium phosphate, alginates, tragacanth, gelatin, calcium silicate, microcrystalline cellulose, polyvinylpyrrolidone, cellulose, water, syrup, and methyl cellulose. The formulations can additionally include, without limitation, lubricating agents such as talc, magnesium stearate, and mineral oil; wetting agents; emulsifying and suspending agents; preserving agents such as methyl- and propylhydroxy-b enzoates ; sweetening agents; flavoring agents, or combinations thereof.

EXAMPLES

The invention will be described in greater detail by way of specific examples. The following examples are offered for illustrative purposes and are not intended to limit the invention in any manner. Those of skill in the art will readily recognize a variety of noncritical parameters which can be changed or modified to yield essentially the same results. Analytical methods described throughout the Examples were performed according to the following procedures:

LC-MS - Method A

LC-MS - Method B

LC-MS - Method C μL

LC-MS - Method D μL

PREP-HPLC Method A

PREP-HPLC Method B

PREP-HPLC Method C

Intermediate 1. 2,5-Dioxopyrrolidin-l-yl palmitate

To a stirred solution of palmitic acid (1 g, 3.91 mmol, 1.0 eq.) in ethyl acetate (20 mL) was added NHS (885 mg, 4.30 mmol, 1.1 eq.) at 25 °C for 10 min. After 10 min, DCC (494 mg, 4.30 mmol, 1.1 eq.) was added at 25°C. The reaction mixture was stirred at room temperature for 18h. The reaction mixture was filtered through Celite pad and washed with ethyl acetate (2 X 15 mL). The filtrate was concentrated under reduced pressure to obtain crude solid (1.2 g), which was purified from silica gel chromatography (60-120 mesh) and eluted with 10-20% of ethyl Acetate/hexane. Collected pure fractions were concentrated under reduced pressure to afford 2,5- dioxopyrrolidin-1-yl palmitate (800 mg) as a white solid. ¹H-NMR (400 MHz,

CDCl₃): δ 2.83 (bs, 4H), 2.59 (t, J=7.6 Hz, 2H), 1.78-1.70 (m, 2H), 1.44-1.35 (m, 2H), 1.34-1.22 (m, 22H), 0.88 (t, J= 6.8 Hz, 3H).

Example 1. 2-Palmitamidoethyl (2-(trimethylammonio)ethyl) phosphate (Compound 1)

Step 1. Benzyl (2 -hydroxy ethyl) carbamate

To a stirred solution of 2-aminoethan- 1 -ol (50 g, 0.819 mol, 1.0 eq.) in DCM (1.5 L) at 0°C was added Et₃N (137 mL, 0.983 mol, 1.2 eq.) dropwise. The reaction mixture was stirred for 10 min at 0 °C and after 10 min, benzyl carbonochloridate (Cbz-Cl; 50 %; 302 mL, 1.064 mol, 1.2 eq) was added to the reaction mixture, dropwise at 0°C. Then the reaction was stirred for 3 h at 0 °C and monitored by thin layer chromatography (TLC). The reaction mixture was then quenched with water (500 mL)and extracted with DCM (3 X 500 mL). The total organic layer was then dried over anhydrous Na₂SO₄, filtered, and concentrated to obtain crude (200 g) compound, which was purified by silica gel column chromatography using gradient elution with 3% MeOH/DCM to afford benzyl (2-hydroxyethyl)carbamate (95 g) as a white solid. Mass [m/z]: 196.09 [M+H]⁺. Yield: 95 g (59.7%).

Step 2. 2-(( (Benzyloxy)carbonyl)amino)ethyl(2-(trimethylammonio)ethyl)phosphate

To a stirred solution of benzyl (2-hydroxyethyl)carbamate (5 g, 25.641 mmol,

1 eq.) in chloroform (100 mL) was added Et₃N (5.5 mL, 38.461 mmol, 1.5 eq.) followed by POCl₃ (2.65 mL, 28.205 mmol, 1.1 eq.) at -10 °C. Then the reaction mixture was stirred for 1 h at 25 °C, and monitored by TLC. After 1 h, pyridine (17.5 mL, 220.512 mmol, 8.6 eq.) and choline tosylate (10.55 g, 38.461 mmol, 1.5 eq) were added at -10 °C and the resulting mixture was stirred at 25 °C for 18 h. The reaction mass was then cooled to 0 °C, quenched with water (20 mL), and extracted with DCM (3 X 100 mL). The aqueous layer was purified by flash chromatography [Column: Biotage-C18, 60 g Duo-100 A 30 μm; Mobile phase: [ACN: Water + 0.1% TFA];

B%: B%: 0-8%, 0-20min / 8% 20-45 min.] to afford 2-

(((benzyloxy)carbonyl)amino)ethyl(2-(trimethylammonio)ethyl)phosphate (2.4 g) as a colorless liquid. Mass [m/z]: 361.01 [M+H]⁺. Yield: 2.4 g (26%).

Step 3. 2-Aminoethyl(2-(trimethylammonio)ethyl)phosphate

To a stirred solution of 2-(((benzyloxy)carbonyl)amino)ethyl(2- (trimethylammonio)ethyl)phosphate (2.3 g, 0.638 mmol, 1.0 eq.) in isopropyl alcohol (IP A; 20 mL) was added 10% Pd/C (50% wet; 500 mg) at 25 °C. The reaction mixture was stirred under hydrogen pressure at 25 °C for 18 h. The resulting mixture was filtered through a celite pad and washed with IP A. The filtrate was concentrated and dried under vacuum to yield 2-aminoethyl(2-(trimethylammonio)ethyl)phosphate (1.6 g) as a colorless liquid. The product was used in the following Examples without further purification. Mass [m/z]: 227.5 [M+H]⁺. Yield: 1.6 g (93%).

Step 4. 2-Palmitamidoethyl (2-(trimethylammonio)ethyl) phosphate (Compound 1)

To a stirred solution of palmitic acid (100 mg, 0.39 mmol, 1.0 eq.) in DMF (5 mL) was added HATU (178 mg, 0.46 mmol, 1.2 eq.) and stirred at 25 °C for 30 min. After 30 min, DIPEA (0.2 mL, 1.17 mmol, 3.0 eq.) and 2-aminoethyl(2- (trimethylammonio)ethyl)phosphate (131 mg, 0.855 mmol, 1.5 eq.) were added to the reaction mixture at 25 ^°C. The mixture was stirred at room temperature for 18 h. The reaction mixture was diluted with IPE (20 mL) and the solvent was decanted to obtain crude solid (300g), which was purified according to PREP-HPLC Method- A: Kinetex EVO Cl 8, 250 X 21.2 mm, 5 μm; Mobile phase: [0.1% of FA in Water: ACN]; time/ B%: 0/10, 20/95) to afford the title compound as a pale brown semisolid (75 mg). Mass [m/z]: 465.2 [M+H]⁺.(LCMS Method A) ¹H-NMR (400 MHz, DMSO-d₆): δ 8.28 (t, J=4.8 Hz, 1H), 4.06-3.99 (m, 2H), 3.67-3.60 (m, 2H), 3.52-3.47 (m, 2H), 3.19-3.13 (m, 2H), 3.12 (s, 9H), 2.02 (t, J=7.2 Hz, 2H), 1.46 (t, J=6.8 Hz, 2H), 1.26- 1.17 (m, 24H), 0.85 (t, . J=6.8 Hz, 3H).³¹P-NMR (400 MHz, DMSO-d₆): δ -0.23 Example 2. 2-Ammonioethyl (2-palmitamidoethyl) phosphate (Compound 2)

Step 1. 2-(((Benzyloxy)carbonyl)amino)ethyl (2-((tert-butoxycarbonyl)amino)ethyl) phosphate

To a stirred solution of benzyl (2-hydroxyethyl)carbamate (1 g, 5.128 mmol, 1 eq.) in chloroform (20 mL) was added Et₃N (1.1 mL, 7.692 mmol, 1.5 eq.) followed by POCI₃ (0.52 mL, 5.641 mmol, 1.1 eq.) at -10 °C. The reaction mixture was then stirred for 1 h at 25 °C, followed by the addition of pyridine (3.6 mL, 44.102 mmol, 8.6 eq.) and N-Boc ethanol amine (1.23 g, 7.692 mmol, 1.5 eq) at -10°C. The reaction mixture was stirred for 18 h at 25 °C. The mixture was then cooled to 0°C, water was added (20 mL), and the mixture was extracted with dichloromethane (DCM; 3 x 30 mL). The organic layer was concentrated and purified by flash chromatography Column: Biotage-C18, 30g Duo-100 Å 30 μm; Mobile phase: ACN: Water + 0.015% NH₄HCO₃; min/B%: 0/0, 5/5, 30/5, 35/14, 55/14, 58/100. The pure fractions were lyophilized to afford the title compound (200 mg) as off-white solid. [M-H]⁺ (m/z): 417.2; ¹H-NMR (400 MHz, DMSO-d₆): δ 7.69 (br s, 1H), 7.38-7.27 (m, 5H), 7.09 (br s, 1H), 4.99 (s, 2H), 3.72-3.57 (m, 4H), 3.14-3.10 (m, 2H), 3.05-3.01 (m, 2H), 1.36 (9H). ³¹P NMR (400 MHz, DMSO-d₆): 0.14.

Step 2. 2-Aminoethyl (2-((tert-butoxycarbonyl)amino)ethyl) phosphate

To a stirred solution of 2-(((benzyloxy)carbonyl)amino)ethyl (2-((tert- butoxycarbonyl)amino)ethyl) phosphate (50 mg, 0.119 mmol, 1.0 eq.) in isopropanol (IPA; 2 mL) was added 10% Pd/C (50% wet) (15 mg) at 25°C. The reaction mixture was stirred under hydrogen atmosphere at 25°C for 2 h and was filtered through celite pad, and washed with IP A (5 mL). The filtrate was concentrated and dried under vacuum to afford the title product (40 mg, crude) as off-white solid, which was used for the next step without purification. Mass [m/z]: 285.2 [M+H]⁺. ¹H-NMR (400 MHz, DMSO-d₆): δ 8.36 (hr s, 2H), 6.96 (hr s, 1H), 3.87-3.79 (m, 2H), 3.68-3.60 (m, 2H), 3.10-3.02 (m, 2H), 2.97-2.92 (m, 2H), 1.37 (s, 9H). ³¹P NMR (400 MHz, DMSO-d₆): 0.82.

Step 3. 2-((tert-Butoxycarbonyl)amino)ethyl (2-palmitamidoethyl) phosphate

To a stirred solution of 2,5-dioxopyrrolidin-l-yl palmitate (Intermediate 1, 248 mg, 0.706 mmol, 1.0 eq.) in DMF (8 mL) was added Et₃N (0.4 mL, 2.826 mmol, 4.0 eq.) and 2-aminoethyl (2-((tert-butoxycarbonyl)amino)ethyl) phosphate (200 mg, 0.706 mmol, 1.0 eq.) and stirred at 25°C for 18 hours. The reaction mass was evaporated under reduced pressure to afford the crude semisolid, which was used in the next step without any further purification. Mass [m/z]: 523.4 [M+H]⁺. (LCMS Method B)

Step 4. 2-Ammonioethyl (2-palmitamidoethyl) phosphate (Compound 2)

To a stirred solution of 2-((tert-butoxycarbonyl)amino)ethyl (2- palmitamidoethyl) phosphate (20 mg, 0.038 mmol, 1.0 eq.) in DCM (2 ml) was added trifluoroacetic acid (TFA, 0.05 ml) at 0°C, then stirred at 25°C for 2 h. The reaction mixture was then concentrated under reduced pressure and the crude solid (20 mg) was purified from PREP-HPLC (Method-B: Kinetex EVO Cl 8, 250 X 21.2 mm, 5 μm; Mobile phase: [0.1% of FA in Water: ACN]; time/ B%: 0/20, 20/90). The resulting fractions were lyophilized to afforded title compound as an off-white solid (8.5 mg). Mass [m/z]: 423.0 [M+H]⁺. (LCMS Method B) ¹H-NMR (400 MHz, CD₃OD): δ 4.06-3.99 (m, 2H), 3.93-3.86 (m, 2H), 3.39 (t, .J=5.6 Hz, 2H), 3.14 (t, .J=5.2 Hz, 2H), 2.19 (t, .J=7.6 Hz, 2H), 1.59 (t, J=6.8Hz, 2H), 1.37-1.22 (m, 24H),

0.89 (t, .J=6.8 Hz, 3H). ³¹P NMR (400 MHz, CD₃OD): δ 0.53 Example 3. 2-Ammonio-3-((oxido(2- palmitamidoethoxy)phosphoryl)oxy)propanoate (Compound 3)

Step 1. 2-(((Benzyloxy)carbonyl)amino)ethyl (3-(tert-butoxy)-2-((tert- butoxycarbonyl)amino)-3-oxopropyl) phosphate

To a stirred solution of benzyl (2-hydroxyethyl)carbamate (1 g, 5.128 mmol, 1.0 eq.) in chloroform (20 mL) was added Et₃N (1.1 mL, 7.692 mmol, 1.5 eq.), followed by POCl₃ (0.52 mL, 5.641 mmol, 1.1 eq.) at -10°C. The reaction mixture was then stirred for 1 h at 25°C. Pyridine (3.6 mL, 44.102 mmol, 8.6 eq.) and Boc-L- Serine tert-butyl ester (2 g, 7.692 mmol, 1.5 eq) were added at -10°C and the mixture was stirred at 25°C for 18 h. The reaction mixture was then cooled to 0°C, and water was added (20 mL). The mixture was extracted with DCM (3 x 30 mL) and the organic layer was concentrated and purified by flash chromatography (Column: Biotage-C18, 30g Duo-100 A 30 μm; Mobile phase: ACN: Water + 0.015% NH4HCO3; min.ZB%: 0/0, 30/5, 35/20, 50/20, 55/100). Pure fractions were lyophilized to afford the title product (220 mg) as an off-white solid. [M-H]⁺ (m/z): 517.2; ¹H- NMR (400 MHz, DMSO-d₆): δ 7.94-7.89 (m, 1H), 7.64-7.58 (m, 1H), 7.39-7.27 (m, 5H), 7.09 (bs, 3H), 4.99 (s, 2H), 3.88-3.78 (m, 3H), 3.67-3.58 (m, 2H), 3.16-3.08 (m, 2H), 1.42-1.32 (m, 18H). ³¹P NMR (400 MHz, DMSO-d₆): 0.66.

Step 2. 2-Aminoethyl (3-(tert-butoxy)-2-((tertbutoxycarbonyl)amino)-3-oxopropyl) phosphate

To a stirred solution of 2-(((benzyloxy)carbonyl)amino)ethyl (3 - (tert-butoxy)- 2-((tert-butoxycarbonyl)amino)-3-oxopropyl) phosphate (50 mg, 0.096 mmol, 1.0 eq.) in IP A (2 mL) was added 10% Pd/C (50% wet) (15 mg) at 25 °C. The reaction mixture was stirred under the atmosphere of hydrogen gas at 25°C for 2 h. The reaction mixture was filtered through celite pad, washed with IP A (5 mL), and the filtrate was concentrated and dried under vacuum to afford the title compound (22 mg) as an off- white solid. [M+H]⁺ (m/z): 385.3; ¹H-NMR (400 MHz, DMSO-d₆): δ 8.23 (bs, 2H), 3.95-3.81 (m, 5H), 2.97-2.93 (m, 2H), 1.44-1.35 (m, 18H). ³¹P NMR (400 MHz, DMSO-d₆): 1.09.

Step 3. 3-(tert-Butoxy)-2-((tert-butoxycarbonyl)amino)-3-oxopropyl (2- palmitamidoethyl) phosphate o

O^'

N NHBoc

H

COOtBu

To a stirred solution of 2,5-dioxopyrrolidin-l-yl palmitate (41 mg, 0.117 mmol, 1.0 eq.) in DMF (2 mL) was added Et₃N (0.067 mL, 0.468 mmol, 4.0 eq.) and 2-aminoethyl (3-(tert-butoxy)-2-((tertbutoxycarbonyl)amino)-3-oxopropyl) phosphate (45 mg, 0.117 mmol, 1.0 eq.) and stirred at 25°C for 18 hours. The reaction mass was evaporated under reduced pressure to afford the crude semisolid, which was used in the next step without any further purification. Mass [m/z]: 623.5 [M+H]⁺. (LCMS Method B)

Step 4. 2-Ammonio-3-((oxido(2-palmitamidoethoxy)phosphoryl)oxy)propanoate (Compound 3)

To a stirred solution of 3-(tert-butoxy)-2-((tert-butoxycarbonyl)amino)-3- oxopropyl (2 -palmitamidoethyl) phosphate (70 mg, 0.112 mmol, 1.0 eq.) in DCM (5 mL) was added TFA (0.03 ml) at 0°C, then stirred at 25°C for 36 hours. The reaction mixture was then concentrated under reduced pressure and the crude solid (60 mg) was purified from PREP-HPLC (Method-C: Gemini Cl 8, 250 x 21.2 x 5 pm, 1.6 μm; Mobile phase: [ACN/0.1% of formic acid in water]; min./B%: 0/10, 15/70, 30/95). The resulting fractions were lyophilized to afforded title compound as an off-white semi solid (15.0 mg). Mass [m/z]: 467.4 [M+H]⁺.(LCMS Method C) ¹H-NMR (400 MHz, DMSO-d₆): δ 8.03 (t, J= 5.2 Hz, 1H), 4.07-4.00 (m, 3H), 3.68-3.60 (m, 2H), 3.21-3.13 (m, 2H), 2.03 (t, J=7.2 Hz, 2H), 1.46 (t, J=7.2 Hz, 2H), 1.31-1.14 (m, 22H), 0.85 (t, .J=6.4 Hz, 3H). ³¹P NMR (400 MHz, DMSO-d₆): 0.66.

Example 4. In vitro MfsdZa Transport

Methodology

The assay was conducted in a low throughput 6-well format with HEK293 cells prepared and then transfected in duplicate wells with plasmids containing either the wildtype (WT) version of hMfsd2a, the D97A mutant version, or an empty vector as control. Uptake into the cells was assessed by both thin-layer chromatography (TLC) and ultrahigh performance liquid chromatography hyphenated mass spectrometry.

Cell Transfection

HEK293 cells were seeded at 6.25 x 10⁵ per 6-well in 2mL of DMEM with 10% FBS and 1% penicillin-streptomycin (P/S) media (Sigma) and incubated overnight at 37°C in 5% CO₂. Cells were checked for confluency the next morning.

On a per well basis the following lipid mix was generated; 6 μL of Lipofectamine 2000 was added dropwise to 200 μL of OptiMEM, this was left to stand for 5 minutes at room temperature (RT). 1 pg of hMfsd2a WT, D97A or empty plasmid was prepared in 200 μL of OptiMEM as appropriate for each well; the Lipofectamine 2000 in OptiMEM solution was then added dropwise to a total volume of 400 μL (this can be scaled to support the number of wells/plates to be assayed). This transfection preparation was then incubated at RT for 20 minutes. DMEM with 10% FBS and no P/S media was warmed to 37°C, the HEK293 plate media was changed and the cells washed carefully with 1 mL of the warmed DMEM with 10% FBS no P/S media, 1.6 mL of the warmed DMEM with 10% FBS no P/S media was then added to each well. Four hundred μL of the transfection preparation was then added dropwise to each well as appropriate and the plate was gently swirled in a circular motion. The plate was then incubated overnight at 37°C in 5% CO₂.

Compound Incubation and Preparation of Analysis Samples

Compound stock solutions were prepared in a 12% BSA in PBS solution such that a 40 μL spike into 2 mL of plain DMEM would yield a concentration of 50 μM of test compound (the compound treated media). Remaining compound stock solution in 12% BSA in PBS was frozen at -20°C to allow for media stability testing. The HEK293 6-well plate was removed from the incubator and the wells gently rinsed with 1 mL of plain DMEM that had been prewarmed to 37°C. Two mL of the compound treated media was then added to each well. A 100 μL sample of the compound treated media was sampled to represent a T = 0h sample; 5 μL of this sample (the remainder was reserved and frozen in case of re-analysis being required) was diluted with 45 μL of DMEM and crashed with 50 μL MeCN in a 96-well plate which was sealed and kept on ice. The HEK293 6-well plate was then incubated at 37°C in 5% CO₂ for 1 hour, the plate was then removed from the incubator and a 100 pL sample of media taken from each well to represent a T = 1h sample; 5 μL of this sample (the remainder was reserved and frozen in case of re-analysis being required) was diluted with 45 μL of DMEM and crashed with 50 μL MeCN into the 96-well plate which was re-sealed and stored at -20°C. The remaining media was then removed from the HEK293 6-well plate, the wells were gently rinsed twice with 1 mL of 0.5% BSA in DMEM, the media was then removed and the 6-well plate allowed to dry completely at RT. 1 mL of 3:2 Hexane:lsopropanol (H IP) was added to each well in a fume cupboard and the plate allowed to stand for 30 minutes at RT without shaking. The HIP solution was then transferred to 2 mL Eppendorf tubes, and the process was repeated with a second 1 mL aliquot of HIP and the two aliquots combined. The HIP samples were then dried down under a nitrogen stream.

Thin Layer Chromatography (TLC) Analysis

Silica gel plates were prepared in a fume cupboard by initially drawing a line 1.5 cm from the bottom edge of the plate and then drawing sample lanes with a width of 1 cm and a separation of 0.5 cm between lanes. TLC buffer for phospholipids was prepared as a 31:62:7 solvent mix of Methanol:Chloroform: Ammonium Hydroxide. Plates were pre-run in a humid chamber containing 200 mL of the TLC buffer until the solvent front was 1.5 cm from the plate edge, the plate was then allowed to dry. HIP samples prepared as described above were reconstituted in 50 μL of chloroform, briefly vortexed 3 times and then kept on ice.

Samples were loaded onto the plate (along with reference compound) by streaking gently with a pipette tip, samples were allowed to dry between streaks. On completion of the sample loading the plate was run in a sealed humid chamber containing the TLC buffer as described above for approximately 1.5 hours or until the solvent front had nearly reached the top of the plate. The plate was removed from the chamber and dried. An initial image was taken using Bio-Rad Image lab 6.0. Iodine crystals were added to a new chamber which was sealed to allow the iodine vapor to saturate the container, the plate was then exposed to the iodine vapor in the chamber to allow visualization of bands of unsaturated fatty acids, once the plate was developed a second image was taken using Bio-Rad Image lab 6.0. The plate was then air-dried to remove the iodine. The plate was then saturated using a spray bottle with cupric acetate solution consisting of 3% cupric acetate by weight, 8% phosphoric acid by volume made up in an aqueous solution. The plate was allowed to dry for 5 minutes at RT and then heated in a fume cupboard using a hot air gun to make the bands more visible. A final image was acquired using the Bio-Rad Image lab 6.0. The difference in intensity between the bands generated from hMfsd2a (WT) or D97A transfected HEK293 cells were compared to the empty vector (EV) transfected cells, allowed for uptake into the cells driven by hMfsd2a to be identified against the reference (REF). Results of the TLC analysis are shown in FIGs. 1A-3B. FIGs. 1A-1B show representative TLC images from the iodine and cupric acetate stain respectively as described above with Compound 1 — lane from left: reference compound, HIP sample from WT cells, cells transfected with D97A mutant Mfsd2a and empty vector, with bands corresponding to the Compound 1 not showing significant differentiation of intensity in WT cells compared with cells transfected with D97A mutant Mfsd2a and/or empty vector. Hence, it cannot be confirmed if the compound is transported via Mfsd2a by this method.

FIGs. 2A-2B show the TLC images from the iodine and cupric acetate stain respectively as described above with Compound 2 - lane from left: reference compound, HIP sample from WT cells, cells transfected with D97A mutant Mfsd2a and empty vector, with bands corresponding to the Compound 2 not showing significant differentiation of intensity in WT cells compared with cells transfected with D97A mutant Mfsd2a and/or empty vector. Hence, it cannot be confirmed if the compound is transported via Mfsd2a by this method.

FIGs. 3A-3B show the TLC images from the iodine and cupric acetate stain respectively as described above with Compound 3 - lane from left: reference compound, HIP sample from WT cells, cells transfected with D97A mutant Mfsd2a and empty vector, with bands corresponding to the Compound 3 not showing significant differentiation of intensity in WT cells compared with cells transfected with D97A mutant Mfsd2a and/or empty vector. Hence, it cannot be confirmed if the compound is transported via Mfsd2a by this method.

UPLC-MS-MS Analysis

HIP samples prepared as described above were reconstituted in 100 μL of MeCN, vortex mixed and inverted multiple times to ensure all surfaces of the Eppendorf tube were rinsed with the MeCN and finally pulse centrifuged. A 50 μL aliquot of the MeCN reconstitution solution was then taken as a non-diluted HIP extract sample and added to the 96-well plate, alongside this a 1:10 dilution sample was prepared by taking a 5 μL aliquot and diluting with 45 μL of MeCN; 50 μL of Millipore water was added to each sample. A bioanalytical calibration line was prepared to cover a range of concentration from 0.0001 to 10 μM by spiking 2 μL of a 0.5 mM DMSO stock of the test compound into 98 μL of MeCN to generate a 10 μM top standard that was then serial diluted with MeCN to produce 6 calibration standard stocks. Fifty μL of each calibration standard stock was added to the 96 -well plate and diluted with 50 μL of Millipore water. Fifty μL of an appropriate internal standard in MeCN was then added to each of the wells in the 96-well plate that contained either a sample or calibration standard, the plate was sealed and transferred to the UPLC- MS-MS system for analysis. Uptake of the test compound into the HEK293 cells determined from the HIP sample analysis with the impact of hMfsd2a assessed by comparing the ratio of the concentration of test compound in the hMfsd2a and D97A transfected cells to the empty vector transfected cells, as shown in FIGs. 4-6.

FIG. 4 shows the concentration of Compound 1 measured in HIP samples from WT cells, cells transfected with D97A mutant Mfsd2a and empty vector, with more Compound 1 detected in WT cells compared to cells transfected with D97A mutant Mfsd2a and/or empty vector, thus confirming the compound is transported via Mfsd2a.

FIG. 5 shows the concentration of Compound 2 measured in HIP samples from WT cells, cells transfected with D97A mutant Mfsd2a and empty vector, with more Compound 2 detected in WT cells compared to cells transfected with D97A mutant Mfsd2a and/or empty vector, thus confirming the compound is transported via Mfsd2a.

FIG. 6 shows the concentration of Compound 3 measured in HIP samples from WT cells, cells transfected with D97A mutant Mfsd2a and empty vector, with more Compound 3 detected in cells transfected with empty vector compared to cells transfected with D97A mutant Mfsd2a or WT cells. Hence, it cannot be definitively confirmed if the compound is transported via Mfsd2a by this method.

Bioanalytical Analysis

Bioanalytical samples were prepared according to the procedures described above for LC -MS-MS analysis. The samples were analyzed by LC-MSMS utilizing the AB Sciex QTRAP 5500. The instrument was set to operate in positive ion mode for all analyses and the parameters are shown below in Table below. - MS/MS TUNE PARAMETERS

ADDITIONAL MS/MS PARAMETERS

Example 5. In vivo AD ME

Protocol 1 : PO terminal rat PK study at 20 mg/kg

Compound 1 was orally dosed at 20 mg/kg to a group of 15 individually housed male Sprague Dawley rats that were fasted overnight and fed 4 hours post- dose. Dosing was performed with 5 mL/kg dosing volumes with 5% DMSO 95% water used as a dosing vehicle. Terminal blood samples were taken from groups of 3 animals at each of 5 time-points post dose (1 hour, 2 hours, 4 hours, 8 hours and 24 hours )by cardiac puncture inder CO2 into lithium heparin coated tubes. Immediately following collection of blood samples, whole body perfusion was performed using phosphate buffered saline (PBS) and the brain excised, rinsed in ice-cold PBS, blotted dry, weighed, and snap frozen in liquid nitrogen. Collected tissue samples were stored in the freezer at -70°C until bioanalysis.

Protocol 2: Bioanalytical samples preparation for blood samples

Blood samples were defrosted and acetonitrile (containing internal standard or IS) was used to precipitate the proteins. All the samples were mixed, centrifuged and the supernatants analysed by LC -MS/MS according to the following procedures:

1. 50 μL blood was transferred to a clean 1.5 mL tube.

2. 150 μL acetonitrile (ACN) containing IS was added to each tube.

3. The tubes were mixed using a vortex for 1 minute.

4. The samples were centrifuged for 5 minutes at 3000 rpm.

5. 150 μL supernatant was transferred to a clean v-bottom 96-well plate containing 150 μL MilliQ water with 0.1% formic acid.

6. Plates were placed into the LC -MS/MS and analysed for Compound 1.

The resulting PK parameters of Compound 1 in blood are shown below in Table 1.

Table 1.

Protocol 3: Bioanalytical samples preparation for brain samples

Brain homogenates were prepared from whole frozen brains harvested from Protocol 1 using ACN and methanol (MeOH). Harvested whole brains were weighed and 2x weight equivalent volume of ice-cold 1 : 1 ACN :MeOH added before homogenization. Analysis of the brain samples was conducted according to the following procedures and the data are shown in Table 2.

1. 200 μL of brain homogenate was transferred to a clean 10 mL tube.

2. 4 mL of ice-cold 1 : 1 ACN:MeOH and 2 μL IS in MeOH was added into each tube.

3. The tubes were mixed on a vortex briefly, sonicated for 15 minutes in a cold water bath and vortex vigorously for 30 seconds. 4. The samples were centrifuged for 5 minutes at 3000 rpm.

5. The supernatant was transferred into a fresh 10 mL tube.

6. Samples were concentrated using nitrogen then washed down with 1 mL MeOH.

7. Samples were further concentrated using nitrogen and the residue reconstituted in 100 μL of 90:10 ACN:MilliQ water and mixed vigorously on a vortex for 5 minutes.

8. Samples were then analyzed by LC-MS/MS.

The resulting PK parameters of Compound 1 in brain are shown below in Table 2.

Table 2.

Protocol 4: Bioanalytical Analysis

Bioanalytical samples were prepared according to the procedures described above for LC-MS/MS analysis. The samples were analyzed by LC-MS/MS utilizing the AB Sciex QTRAP 6500+ coupled to a HPLC system. The LC-MS/MS conditions used are shown below in Table 3.

Table 3.

Example 6. Cytoprotective Effect

This Example demonstrate the cytoprotective effects of Compound 1 in human induced pluripotent stem cell (iPSC) derived retinal pigment epithelium (RPE) cells. Human iPSC-RPE cells were differentiated on 96 -well plates for 16 days. Cells were exposed to Compound 1 at 5 μΜ in quadruplicates for 48 hours before induction of oxidative stress (day 19) with tert- butyl hydroperoxide solution (tBHP) (0-10 mM) at 37°C 5% CO₂. Lactate dehydrogenase (LDH) was measured on samples collected after 24 hours tBHP-incubation as a readout of the cytoprotective effect of Compound

1.

Materials and Methods hiPSC derived Retinal Pigment Epithelium

Human iPSC-RPE cells (PCi-RPE, p2, Phenocell SAS, France) were cultured according to manufacturer’s instructions. Cells (passage 2) were expanded for 14 days in culture medium containing 70% DMEM, high glucose (Gibco Thermo Fischer Scientific, USA), 30% Ham’s F12 Nutrient Mix (Gibco Thermo Fischer Scientific, USA), 2% B-27® Supplement (Gibco Thermo Fischer Scientific, USA), 1% Antibiotic-Antimycotic (Gibco Thermo Fischer Scientific, USA) in Matrigel^® coated cell culture dishes (Coming, USA, final density of Matrigel 8-10 pg/cm²) at 37°C 5% CO₂. iPSC-RPE cells acquired their characteristic polygonal morphology and were pigmented at passage 3 in Matrigel^® coated cell culture dishes. Cells were passaged (passage 4) onto Matrigel^® coated 96-well plates at a density of 100,000 cells/cm², and cultured until fully differentiated into RPE cells for 16 days.

Compounds, Delivery, Storage, and Administration

Compound 1 was dissolved in 100% DMSO (Sigma- Aldrich, USA) at concentration 10 mM. Thereafter, dilutions of study compounds were prepared in RPE medium with B-27 AO neg. (Gibco Thermo Fischer Scientific, USA, 100 mM) and sterile filtered through 0.22 pm. Sterile aliquots were stored at -20°C. Final working dilutions of study compounds (5 μM, 0.1% DMSO) were prepared fresh in RPE medium with B-27 AO neg. Vehicle was 0.1% DMSO in RPE medium with B- 27 AO neg. RPE cells were pre-treated with each of the study compounds or vehicle 48 hours before induction of oxidative stress. LDH Assay Protocol

Human iPSC-RPE cells were pre-incubated with Compound 1 at 5 μΜ in quadruplicates for 48 hours before exposure to whole concentration range of tBHP at 37°C for 24 hours (co-incubation period during which iPSC-RPE cells will be continually exposed to Compound 1). LDH quantification was performed on samples collected after 6 hours tBHP incubation, and again after the 24 hours tBHP- incubation, according to the previously published protocol (Kaja et al. 2015). Samples (50 μL ) of cell culture supernatants were collected (quadruplicates) and incubated in 1 mM INT (Iodonitrotetrazolium Chloride, Sigma- Aldrich, USA), 1.6 mM NAD (Beta- Nicotinotinamide Adenine Dinucleotide Sodium Salt, Sigma-Aldrich, USA), 80 mM lithium L-lactate (Sigma-Aldrich, USA), 7.5 μΜ MPMS ( 1 -Methoxyphenazine methosulfate, Sigma-Aldrich, USA) in 0.2 M Tris-HCl (Sigma-Aldrich, USA), pH 8.2 for 30 minutes. The reaction was stopped using 1 M acetic acid (Fisher, USA). Absorbance at 490 nm was measured using Cytation 3 multi-mode reader (BioTek, Winooski, VT, USA) .

Kaja et al. An Optimized Lactate Dehydrogenase Release Assay for Screening of Drug Candidates in Neuroscience. J Pharmacol Toxicol Methods. May-Jun 2015;73:1-6. doi: 10.1016/j.vascn.2015.02.001.

Data Analysis

All the assay data was normalized to cell viability of vehicle treated control cells (not induced with tBHP). All values are presented as mean ± standard deviation (SD). Parametric data were analyzed using One-way- AN O V A followed by Dunnett’s multiple comparisons posthoc test to compare each group to the vehicle group. Differences were considered to be statistically significant at P<0.05.

Cytoprotection of human iPSC-RPE cells readout

As can be seen from Figs. 7A-7B, Compound 1 displays cytoprotective effects. Compound 1 shows lower LDH release at 0.6 mM tBHP compared to the vehicle, indicating that this compound has cytoprotective properties in hiPSC-RPE cells.

It is noteworthy that Compound 1 demonstrated a trend towards better cell viability (lower LDH release at 6 hours ) than vehicle at high tBHP concentrations (1- 10 mM) indicating acute (0-6 hours) protective effects against oxidative stress (refer to Figs. 7A-7B).

Example 7. 2-(((2-palmitamidoethoxy)carbonyl)amino)ethyl (2- (trimethylammonio)ethyl) phosphate

Mass [m/z] : 552.5 [M+H]^{+ 1}H-NMR (400 MHz, DMSO-d₆): δ 8.00 (t, .J=4.8 Hz, 1H), 7.46 (t, .J=4.8 Hz, 1H), 4.06-3.98 (m, 2H), 3.91 (t, .J=6.0 Hz, 2H), 3.69-3.63 (m, 2H), 3.51-3.48 (m, 2H), 3.21 (q, .J=6.0 Hz, 2H), 3.12 (s, 9H), 3.11-3.08 (m, 2H), 2.04 (t, J=7.6 Hz, 2H), 1.45 (t, J=6.8 Hz, 2H), 1.29-1.17 (m, 24H), 0.85 (t, J=6.8 Hz, 3H). ³¹P-NMR (162 MHz, DMSO-d₆): δ -0.25.

It will be appreciated that all other compounds disclosed in the present application can be prepared according to the procedures described in Examples 1 , 2 and 3 and/or Schemes 1, 2, 3 and 4, using appropriately substituted starting materials.

OTHER EMBODIMENTS

It is to be understood that while the invention has been described in conjunction with the detailed description thereof, the foregoing description is intended to illustrate and not limit the scope of the invention, which is defined by the scope of the appended claims. Other aspects, advantages, and modifications are within the scope of the following claims. It should be appreciated by those persons having ordinary skill in the art(s) to which the present invention relates that any of the features described herein in respect of any particular aspect and/or embodiment of the present invention can be combined with one or more of any of the other features of any other aspects and/or embodiments of the present invention described herein, with modifications as appropriate to ensure compatibility of the combinations. Such combinations are considered to be part of the present invention contemplated by this disclosure.

Claims

WHAT IS CLAIMED IS:

1. A compound of Formula I:

I or a pharmaceutically acceptable salt thereof, wherein:

L¹ is C(O) or PO₂ ^";

X¹ is selected from the group consisting of C₁-₄ alkylene, C₁-₄ alkylene- OC(O)O-C_1-4 alkylene, C_1-4 alkylene-OC(O)NH-C 1 -4 alkylene, C_1-4 alkylene- OC(O)C_1-4 alkylene, C_1-4 alkylene-O-C 1 -4 alkylene, C_1-4 alkylene-O-C 1 -4 alkylene-O- C_1-4 alkylene, and C_1-4 alkylene-O-C_1-4 alkylene-O-C(O)C 1 -4 alkylene, wherein each C_1-4 alkylene is optionally substituted by CO₂H;

X² is C_1-6 alkylene, which is optionally substituted by OH or CO₂H;

Y¹ is selected from the group consisting of O, S, and NH;

R² is selected from the group consisting of H and C_1-6 alkyl, wherein the C_1-6 alkyl is optionally substituted by OH or CO₂H;

R³ is selected from the group consisting of H and C_1-6 alkyl;

R^3a is selected from the group consisting of H and C_1-6 alkyl;

R^3b is selected from the group consisting of H and C_1-6 alkyl; and

R⁴ is selected from the group consisting of H and C_1-6 alkyl, wherein the C_1-6 alkyl is optionally substituted by OH or CO₂H; provided that when L¹ is PO₂ ^", X¹ is an unsubstituted C₁-₂ alkylene, X² is CH₂, R² is H, and R⁴ is H or methyl, then at least one of R³, R^3a, and R^3b is not methyl.

2. The compound of claim 1, or a pharmaceutically acceptable salt thereof, wherein L¹ is C(O).

3. The compound of claim 1, or a pharmaceutically acceptable salt thereof, wherein L¹ is PO₂ ^".

4. The compound of any one of claims 1 to 3, or a pharmaceutically acceptable salt thereof, wherein X¹ is selected from the group consisting of C_1-4 alkylene, C_1-4 alkylene-OC(O)O-C_1-4 alkylene, C_1-4 alkylene-OC(O)NH-C_1-4 alkylene, C_1-4 alkylene- OC(O)C_1-4 alkylene, C_1-4 alkylene-O-C_1-4 alkylene, C_1-4 alkylene-O-C_1-4 alkylene-O- C_1-4 alkylene, and C_1-4 alkylene-O-C_1-4 alkylene-O-C(O)C i -4 alkylene, wherein each C_1-4 alkylene is optionally substituted by CO₂H.

5. The compound of any one of claims 1 to 4, or a pharmaceutically acceptable salt thereof, wherein X¹ is CH₂.

6. The compound of any one of claims 1 to 5, or a pharmaceutically acceptable salt thereof, wherein X² is C ₁-₃ alkylene which is optionally substituted with OH or CO₂H.

7. The compound of any one of claims 1 to 6, or a pharmaceutically acceptable salt thereof, wherein R² is selected from the group consisting of H and C_1-3 alkyl which is optionally substituted by OH or CO₂H.

8. The compound of any one of claims 1 to 7, or a pharmaceutically acceptable salt thereof, wherein R² is H.

9. The compound of any one of claims 1 to 8, or a pharmaceutically acceptable salt thereof, wherein R³ is selected from the group consisting of H and C_1-3 alkyl.

10. The compound of any one of claims 1 to 9, or a pharmaceutically acceptable salt thereof, wherein R³ is H.

11. The compound of any one of claims 1 to 9, or a pharmaceutically acceptable salt thereof, wherein R³ is methyl .

12. The compound of any one of claims 1 to 11 , or a pharmaceutically acceptable salt thereof, wherein R^3a is selected from the group consisting of H and C_1-3 alkyl.

13. The compound of any one of claims 1 to 12, or a pharmaceutically acceptable salt thereof, wherein R^3a is H.

14. The compound of any one of claims 1 to 12, or a pharmaceutically acceptable salt thereof, wherein R^3a is methyl.

15. The compound of any one of claims 1 to 14, or a pharmaceutically acceptable salt thereof, wherein R^3b is selected from the group consisting of H and C_1-3 alkyl.

16. The compound of any one of claims 1 to 15, or a pharmaceutically acceptable salt thereof, wherein R^3b is H.

17. The compound of any one of claims 1 to 15, or a pharmaceutically acceptable salt thereof, wherein R^3b is methyl.

18. The compound of any one of claims 1 to 8, or a pharmaceutically acceptable salt thereof, wherein R³, R^3a, and R^3b are each H.

19. The compound of any one of claims 1 to 8, or a pharmaceutically acceptable salt thereof, wherein R³, R^3a, and R^3b are each methyl.

20. The compound of claim 1 , or a pharmaceutically acceptable salt thereof, wherein:

L¹ is PO₂-;

X¹ is C_1-3 alkylene, which is optionally substituted by CO₂H;

X² is C_1-3 alkylene, which is optionally substituted by OH or CO₂H; R² is selected from the group consisting of H and C_1-3 alkyl which is optionally substituted by OH or CO₂H;

R³ is selected from the group consisting of H and C_1-3 alkyl;

R^3a is selected from the group consisting of H and C_1-3 alkyl; and R^3b is selected from the group consisting of H and C_1-3 alkyl.

21. The compound of claim 1, or a pharmaceutically acceptable salt thereof, wherein:

L¹ is POT;

X¹ is C_1-4 alkylene;

X² is Ci-3 alkylene, which is optionally substituted by CO₂H;

R² is selected from the group consisting of H and C_1-3 alkyl;

R³ is selected from the group consisting of H and C_1-3 alkyl;

R^3a is selected from the group consisting of H and C1-3 alkyl; and R^3b is selected from the group consisting of H and C1-3 alkyl.

22. The compound of claim 1, wherein the compound of Formula I is a compound of Formula II:

II or a pharmaceutically acceptable salt thereof.

23. The compound of claim 1, wherein the compound is selected from the group consisting of:

or a pharmaceutically acceptable salt thereof.

24. A pharmaceutical composition comprising a compound of any one of claims 1 to 23, or a pharmaceutically acceptable salt thereof, and one or more pharmaceutically

25. A method of treating a disease or disorder selected from the group consisting of pain, a pain-related disease or disorder, a mood disease or disorder, a disease or disorder of the central nervous system, a disease or disorder of the peripheral nervous system, an optical disease or disorder, cancer, a gastrointestinal disease or disorder, a renal disease or disorder, a renal-related disease or disorder, a cardiovascular disease or disorder, and a skin disease or disorder in a subject in need thereof, comprising administering to the subject a therapeutically effective amount of a compound of any one of claims 1 to 23, or a pharmaceutically acceptable salt thereof, or a pharmaceutical composition of claim 24.

26. A method of treating a disease or disorder selected from the group consisting of pain, a pain-related disease or disorder, a mood disease or disorder, a disease or disorder of the central nervous system, a disease or disorder of the peripheral nervous system, an optical disease or disorder, cancer, a gastrointestinal disease or disorder, a renal disease or disorder, a renal-related disease or disorder, a cardiovascular disease or disorder, and a skin disease or disorder in a subject in need thereof, comprising administering to the subject a therapeutically effective amount of a compound of Formula I:

I or a pharmaceutically acceptable salt thereof, wherein:

L¹ is C(O) or PO₂-;

X¹ is selected from the group consisting of C_1-4 alkylene, C_1-4 alkylene- OC(O)O-C_1-4 alkylene, C_1-4 alkylene-OC(O)NH-C_1-4 alkylene, C_1-4 alkylene- OC(O)C_1-4 alkylene, C_1-4 alkylene-O- C_1-4 alkylene, C_1-4 alkylene-O-C_1-4 alkylene-O- C_1-4 alkylene, and C_1-4 alkylene-O- C_1-4 alkylene-O-C(O)C_1-4 alkylene, wherein each C_1-4 alkylene is optionally substituted by OH or CO₂H;

X² is C_1-6 alkylene, which is optionally substituted by OH or CO₂H;

Y¹ is selected from the group consisting of O, S, and NR⁵; R² is selected from the group consisting of H and C_1-6 alkyl, wherein the C_1-6 alkyl is optionally substituted by OH or CO₂H;

R³ is selected from the group consisting of H and C_1-6 alkyl;

R^3a is selected from the group consisting of H and C_1-6 alkyl;

R^3b is selected from the group consisting of H and C_1-6 alkyl;

R⁴ is selected from the group consisting of H and C_1-6 alkyl, wherein the C_1-6 alkyl is optionally substituted by OH or CO₂H; and

R⁵ is selected from the group consisting of H and C_1-6 alkyl, wherein the C_1-6 alkyl is optionally substituted by OH or CO₂H.

27. The method of claim 26, wherein the compound is selected from the group consisting of:

or a pharmaceutically acceptable salt thereof.

28. The method of any one of claims 25 to 27, wherein the disease or disorder is pain or a pain-related disease or disorder.

29. The method of claim 28, wherein the pain or a pain-related disease or disorder is selected from the group consisting of acute pain, chronic pain, neuropathic pain, nociceptive pain, inflammatory pain, cancer pain, fibromyalgia, rheumatoid arthritis, osteoarthritis, surgery-related pain, and osteoporosis.

30. The method of any one of claims 25 to 27, wherein the disease or disorder is a mood disease or disorder.

31. The method of claim 30, wherein the mood disease or disorder is selected from the group consisting of anxiety, depression, a sleeping disorder, an eating disorder, post-traumatic stress disorder, symptoms of drug or alcohol withdrawal or abuse, schizophrenia, obsessive-compulsive disorder, bipolar disorder, sexual dysfunction, attention deficit disorder (ADD), and attention deficit hyperactivity disorder (ADHD).

32. The method of any one of claims 25 to 27, wherein the disease or disorder is a disease or disorder of the central nervous system, a disease or disorder of the peripheral nervous system, or an optical disease or disorder.

33. The method of claim 32, wherein the disease or disorder of the central nervous system, a disease or disorder of the peripheral nervous system, or an optical disease or disorder is selected from the group consisting of a demyelinating disease, glaucoma, age-related macular degeneration (AMD), amyotrophic lateral sclerosis (ALS), a cognitive disorder, Alzheimer’s disease, a movement disorder, Huntington’s chorea, Tourette’s syndrome, Niemann-Pick disease, Parkinson's disease, epilepsy, a cerebrovascular disorder, ischemic stroke, and brain injury.

34. The method of claim 33, wherein the demyelinating disease is selected from the group consisting of multiple sclerosis (MS), neuromyelitis optica (NMO), Devic’s disease, central nervous system neuropathy, central pontine myelinolysis, syphilitic myelopathy, leukoencephalopathies, leukodystrophies, Guillain-Barre syndrome, chronic inflammatory demyelinating polyneuropathy, anti -myelin-associated glycoprotein (MAG) peripheral neuropathy, Charcot-Marie-Tooth disease, peripheral neuropathy, myelopathy, optic neuropathy, progressive inflammatory neuropathy, optic neuritis, and transverse myelitis.

35. The method of any one of claims 25 to 27, wherein the disease or disorder is cancer.

36. The method of claim 35, wherein the cancer is selected from the group consisting of leukemia, mantle cell lymphoma, Hodgkin lymphoma, Non-Hodgkin lymphoma, hepatocellular carcinoma, ovarian cancer, colorectal cancer, pancreatic cancer, prostate cancer, breast cancer, glioma, skin cancer, renal carcinoma and lung cancer

37. The method of any one of claims 25 to 27, wherein the disease or disorder is a gastrointestinal disease or disorder.

38. The method of claim 37, wherein the gastrointestinal disease or disorder is selected from the group consisting of inflammatory bowel disease, gastroesophageal reflux disease, paralytic ileus, secretory diarrhoea, gastric ulcer, nausea, emesis, celiac disease, irritable bowel syndrome, and a liver disorder.

39. The method of claim 38, wherein the liver disorder is selected from the group consisting of acute liver failure, Alagille syndrome, hepatitis, enlarged liver, Gilbert’s syndrome, liver cyst, liver haemangioma, fatty liver disease, steatohepatitis, primary sclerosing cholangitis, fascioliasis, primary bilary cirrhosis, Budd-Chiari syndrome, hemochromatosis, Wilson’s disease, and transthyretin-related hereditary amyloidosis.

40. The method of any one of claims 25 to 27, wherein the disease or disorder is a renal disease or disorder or a renal-related disease or disorder.

41. The method of claim 40, wherein the renal disease or disorder or a renal- related disease or disorder is selected from the group consisting of diabetes, diabetic nephropathy, acute inflammatory kidney injury, renal ischemia urinary incontinence, and overactive bladder.

42. The method of any one of claims 25 to 27, wherein the disease or disorder is a skin disease or disorder.

43. The method of claim 42, wherein the skin disease or disorder is atopic dermatitis, psoriasis or lupus.

44. The method of any one of claims 25 to 27, wherein the disease or disorder is a cardiovascular disease or disorder.

45. The method of claim 44, wherein the cardiovascular disease or disorder is selected from the group consisting of cardiovascular disease, vascular inflammation, idiopathic pulmonary fibrosis, cough, and hypertension.

46. The method of any one of claims 25 to 45, further comprising administering to the subject one or more additional therapeutic agents.

47. A combination therapy comprising a compound of any one of claims 1 to 23, or a pharmaceutically acceptable salt thereof, and at least one compound selected from the group consisting of anandamide, 2 -arachidonoyl glycerol, and oleoylethanolamide, or a pharmaceutically acceptable salt thereof.

48. A method of treating a disease or disorder selected from the group consisting of pain, a pain-related disease or disorder, a mood disease or disorder, a disease or disorder of the central nervous system, a disease or disorder of the peripheral nervous system, an optical disease or disorder, cancer, a gastrointestinal disease or disorder, a renal disease or disorder, a renal-related disease or disorder, a cardiovascular disease or disorder, and a skin disease or disorder in a subject, comprising administering to the subject a combination therapy of claim 47.

49. A process of preparing a compound of Formula la:

la or a pharmaceutically acceptable salt thereof, comprising reacting a compound of

Formula IV:

IV with a compound of Formula V:

V in the presence of a first base and an amine coupling agent, wherein:

R⁶ is C_1-6 alkylene;

R⁷ is C_1-6 alkylene; and each R^A, R^B, R^C is independently selected from the group consisting of H and C_1-6 alkyl.

50. The process of claim 49, wherein the first base is a tri(C_1-6 alkyl) amine base.

51. The process of claim 49, wherein the first base is N,N-diisopropylethylamine

(DIPEA).

52. The process of any one of claims 49 to 51 , wherein the amine coupling agent is l-[bis(dimethylamino)methylene]-1H-1,2,3-triazolo[4,5-b]pyridinium 3-oxide hexafluorophosphate (HATU).

53. The process of claim 49, wherein the compound of Formula la is a compound

lb or a pharmaceutically acceptable salt thereof.

54. A process of preparing a compound of Formula lb

lb or a pharmaceutically acceptable salt thereof, comprising reacting a compound of Formula IVa

IVa with a compound of Formula V

V in the presence of Ν,Ν-diisopropylethylamine (DIPEA) and 1- [bis(dimethylamino)methylene]-1H-1,2,3-triazolo[4,5-b]pyridinium 3-oxide hexafluorophosphate (HATU).

55. A process of preparing a compound of Formula Ic:

Ic or a pharmaceutically acceptable salt thereof, comprising reacting a compound of Formula VI:

VI with a compound of Formula VII:

VII in the presence of a first base, wherein:

Pg¹ is an amine protecting group;

R⁶ is C_1-6 alkylene;

R⁷ is C_1-6 alkylene, wherein the C_1-6 alkylene is optionally substituted OH or CO₂H; and

R⁸ is a carboxylic acid activating group.

56. The process of claim 55, wherein the first base is an amine base.

57. The process of claim 55, wherein the first base is tri(C_1-6 alkyl) amine base.

58. The process of claim 55, wherein the first base is triethylamine.

59. The process of any one of claims 55 to 58, wherein R⁸ is 2,5 -dioxopyrrolidin-

1-yl.

60. A process of preparing a compound of Formula Id:

Id or a pharmaceutically acceptable salt thereof, comprising reacting a compound of Formula Via:

Via with a compound of Formula Vila:

O

Vila in the presence of triethylamine.

61. A process of preparing a compound of Formula Ie:

Ie or a pharmaceutically acceptable salt thereof, comprising reacting a compound of Formula VIb:

VIb with a compound of Formula Vila:

Vila in the presence of triethylamine.

62. A compound of Formula Vila:

Vila or salt thereof.

63. The process of any one of claims 55 to 61, further comprising a deprotection step in the presence of a deprotecting agent.

64. The process of claim 63, wherein the deprotecting agent comprises an acid.

65. The process of claim 64, wherein the acid is trifluoroacetic acid (TFA).