Cyclic peptide antibiotics

Abstract

Antibacterial compounds are provided herein, which in some embodiments have broad-spectrum biological activity. In various embodiments, the compounds act by inhibiting lipoprotein signal peptidase II (LspA), a key protein in bacteria. Pharmaceutical compositions and methods of treatment using the compounds described herein are also provided.

Description

CROSS-REFERENCE TO RELATED APPLICATIONS

[0001] This application claims the benefit of priority application PCT / CN2019 / 078189, filed March 14, 2019, the disclosure of which is incorporated herein by reference. [Background technology]

[0002] Antibiotic resistance is a serious and growing phenomenon in modern medicine and has emerged as a major public health concern in the 21st century. Therefore, novel classes of broad-spectrum antibiotics, especially those targeting novel mechanisms of action, are needed to treat multidrug-resistant pathogens. The type II signal peptidase LspA plays a key role in the biosynthesis of Gram-negative bacterial lipoproteins. Blocking lipoprotein synthesis through inhibition of LspA function can not only deplete essential and virulence-associated lipoproteins but also cause the accumulation of mislocalized unprocessed lipoproteins, all of which lead to bacterial cell death. Therefore, LspA inhibitors offer a novel approach to combating Gram-negative bacterial infections. Summary of the Invention

[0003] Described herein are novel macrocyclic compounds for the treatment of microbial infections, such as bacterial infections. In various embodiments, the disclosure provides cyclic peptides, depsipeptides, and peptide-containing compounds for the treatment of bacterial infections. In various embodiments, the disclosure provides classes and subclasses of chemical compounds structurally related to globomycin for the treatment of bacterial infections. In various embodiments, the cyclic peptides, depsipeptides, and peptide-containing compounds act through the inhibition of lipoprotein signal peptidase II (LspA), a key enzyme involved in post-translational processing of lipoproteins in bacteria.

[0004] Formula (I): TIFF0007670618000001.tif81170 formula: R1a , R 2a , R 4a , R 5a and R 6a are each independently hydrogen or optionally substituted C1-C6 alkyl, R 3a is hydrogen, optionally substituted C1-C 20 Alkyl, optionally substituted C-C 20 Alkenyl, optionally substituted C-C 20 alkynyl, optionally substituted cycloalkyl, optionally substituted (C1-C6 alkyl)cycloalkyl, optionally substituted heterocycloalkyl, optionally substituted (C1-C6 alkyl)heterocycloalkyl, optionally substituted aryl, optionally substituted (C1-C6 alkyl)aryl, optionally substituted heteroaryl, or optionally substituted (C1-C6 alkyl)heteroaryl; R 20 is hydroxyl or -NR 1 R 2 and R 1 and R 2 are each independently hydrogen, optionally substituted C1-C6 alkyl, optionally substituted C2-C6 alkenyl, optionally substituted C2-C6 alkynyl, optionally substituted cycloalkyl, optionally substituted (C1-C6 alkyl)cycloalkyl, optionally substituted heterocycloalkyl, optionally substituted (C1-C6 alkyl)heterocycloalkyl, optionally substituted aryl, optionally substituted (C1-C6 alkyl)aryl, optionally substituted heteroaryl, optionally substituted (C1-C6 alkyl)heteroaryl, -S(=O)2R a , -S(=O)2NR b R c , -C(=O)R a , -C(=O)OR b , -C(=O)NR b R c Or -C(=NR b )NR b R c Or Or, R 1 and R2 together with the nitrogen atom to which they are attached form an optionally substituted heterocycloalkyl; R 3 and R 4 are each independently hydrogen, optionally substituted C1-C6 alkyl, optionally substituted C2-C6 alkenyl, optionally substituted C2-C6 alkynyl, optionally substituted cycloalkyl, optionally substituted heterocycloalkyl, optionally substituted aryl, or optionally substituted heteroaryl; Or, R 3 and R 4 together with the carbon atom to which they are attached form oxo, R 5 is hydrogen, halogen, optionally substituted C1-C6 alkyl, optionally substituted C2-C6 alkenyl, or optionally substituted C2-C6 alkynyl; R 6 and R 7 are each independently hydrogen, halogen, optionally substituted C1-C6 alkyl, optionally substituted C2-C6 alkenyl, optionally substituted C2-C6 alkynyl, optionally substituted cycloalkyl, optionally substituted (C1-C6 alkyl)cycloalkyl, optionally substituted heterocycloalkyl, optionally substituted (C1-C6 alkyl)heterocycloalkyl, optionally substituted aryl, optionally substituted (C1-C6 alkyl)aryl, optionally substituted heteroaryl, or optionally substituted (C1-C6 alkyl)heteroaryl; R 8 and R 9are each independently hydrogen, halogen, optionally substituted C1-C6 alkyl, optionally substituted C2-C6 alkenyl, optionally substituted C2-C6 alkynyl, optionally substituted cycloalkyl, optionally substituted (C1-C6 alkyl)cycloalkyl, optionally substituted heterocycloalkyl, optionally substituted (C1-C6 alkyl)heterocycloalkyl, optionally substituted aryl, optionally substituted (C1-C6 alkyl)aryl, optionally substituted heteroaryl, or optionally substituted (C1-C6 alkyl)heteroaryl; Or, R 3a and R 8 together with the atom to which they are attached form an optionally substituted heterocycloalkyl; R 10 and R 11 are each independently hydrogen, halogen, optionally substituted C1-C6 alkyl, optionally substituted C2-C 20 alkenyl, optionally substituted C2-C6 alkynyl, optionally substituted cycloalkyl, optionally substituted (C1-C6 alkyl)cycloalkyl, optionally substituted heterocycloalkyl, optionally substituted (C1-C6 alkyl)heterocycloalkyl, optionally substituted aryl, optionally substituted (C1-C6 alkyl)aryl, optionally substituted heteroaryl, or optionally substituted (C1-C6 alkyl)heteroaryl; R 12 and R 13 are each independently hydrogen, halogen, or an optionally substituted C1-C 20 alkyl, optionally substituted C2-C6 alkenyl, optionally substituted C2-C6 alkynyl, optionally substituted cycloalkyl, optionally substituted (C1-C6 alkyl)cycloalkyl, optionally substituted heterocycloalkyl, optionally substituted (C1-C6 alkyl)heterocycloalkyl, optionally substituted aryl, optionally substituted (C1-C6 alkyl)aryl, optionally substituted heteroaryl, or optionally substituted (C1-C6 alkyl)heteroaryl; R 14 and R15 are each independently hydrogen, halogen, optionally substituted C1-C6 alkyl, optionally substituted C2-C6 alkenyl, optionally substituted C2-C6 alkynyl, optionally substituted cycloalkyl, optionally substituted (C1-C6 alkyl)cycloalkyl, optionally substituted heterocycloalkyl, optionally substituted (C1-C6 alkyl)heterocycloalkyl, optionally substituted aryl, optionally substituted (C1-C6 alkyl)aryl, optionally substituted heteroaryl, or optionally substituted (C1-C6 alkyl)heteroaryl; R 16 and R 17 are each independently hydrogen, halogen, optionally substituted C1-C6 alkyl, optionally substituted C2-C6 alkenyl, optionally substituted C2-C6 alkynyl, optionally substituted cycloalkyl, optionally substituted (C1-C6 alkyl)cycloalkyl, optionally substituted heterocycloalkyl, optionally substituted (C1-C6 alkyl)heterocycloalkyl, optionally substituted aryl, optionally substituted (C1-C6 alkyl)aryl, optionally substituted heteroaryl, or optionally substituted (C1-C6 alkyl)heteroaryl; Each R a are independently optionally substituted C1-C6 alkyl, optionally substituted C2-C6 alkenyl, optionally substituted C2-C6 alkynyl, optionally substituted cycloalkyl, optionally substituted heterocycloalkyl, optionally substituted aryl, or optionally substituted heteroaryl; Each R b and R c are independently hydrogen, optionally substituted C1-C6 alkyl, optionally substituted C2-C6 alkenyl, optionally substituted C2-C6 alkynyl, optionally substituted cycloalkyl, optionally substituted heterocycloalkyl, optionally substituted aryl, or optionally substituted heteroaryl; Or, R b and R c together with the nitrogen atom to which they are attached form an optionally substituted heterocycloalkyl; or a pharmaceutically acceptable salt, solvate or stereoisomer thereof.

[0005] Formula (Ia): Also disclosed herein is the compound of TIFF0007670618000002.tif80170 or a pharmaceutically acceptable salt, solvate, or stereoisomer thereof.

[0006] Formula (Ib): Also disclosed herein is the compound of TIFF0007670618000003.tif81170 or a pharmaceutically acceptable salt, solvate, or stereoisomer thereof.

[0007] Formula (II): TIFF0007670618000004.tif81170 formula: R 1a , R 2a , R 3a , R 4a , R 5a and R 6a are each independently hydrogen or optionally substituted C1-C6 alkyl, R 1 and R 2 are each independently hydrogen, optionally substituted C1-C6 alkyl, optionally substituted C2-C6 alkenyl, optionally substituted C2-C6 alkynyl, optionally substituted cycloalkyl, optionally substituted (C1-C6 alkyl)cycloalkyl, optionally substituted heterocycloalkyl, optionally substituted (C1-C6 alkyl)heterocycloalkyl, optionally substituted aryl, optionally substituted (C1-C6 alkyl)aryl, optionally substituted heteroaryl, optionally substituted (C1-C6 alkyl)heteroaryl, -S(=O)2R a , -S(=O)2NR b R c , -C(=O)R a , -C(=O)OR b , -C(=O)NR b R c Or -C(=NR b)NR b R c Or Or, R 1 and R 2 together with the nitrogen atom to which they are attached form an optionally substituted heterocycloalkyl; R 3 and R 4 are each independently hydrogen, optionally substituted C1-C6 alkyl, optionally substituted C2-C6 alkenyl, optionally substituted C2-C6 alkynyl, optionally substituted cycloalkyl, optionally substituted heterocycloalkyl, optionally substituted aryl, or optionally substituted heteroaryl; Or, R 3 and R 4 together with the carbon atom to which they are attached form oxo, R 5 is hydrogen, halogen, optionally substituted C1-C6 alkyl, optionally substituted C2-C6 alkenyl, or optionally substituted C2-C6 alkynyl; R 6 and R 7 are each independently hydrogen, halogen, optionally substituted C1-C6 alkyl, optionally substituted C2-C6 alkenyl, optionally substituted C2-C6 alkynyl, optionally substituted cycloalkyl, optionally substituted (C1-C6 alkyl)cycloalkyl, optionally substituted heterocycloalkyl, optionally substituted (C1-C6 alkyl)heterocycloalkyl, optionally substituted aryl, optionally substituted (C1-C6 alkyl)aryl, optionally substituted heteroaryl, or optionally substituted (C1-C6 alkyl)heteroaryl; R 8 and R 9are each independently hydrogen, halogen, optionally substituted C1-C6 alkyl, optionally substituted C2-C6 alkenyl, optionally substituted C2-C6 alkynyl, optionally substituted cycloalkyl, optionally substituted (C1-C6 alkyl)cycloalkyl, optionally substituted heterocycloalkyl, optionally substituted (C1-C6 alkyl)heterocycloalkyl, optionally substituted aryl, optionally substituted (C1-C6 alkyl)aryl, optionally substituted heteroaryl, or optionally substituted (C1-C6 alkyl)heteroaryl; R 10 and R 11 are each independently hydrogen, halogen, or an optionally substituted C1-C 20 Alkyl, optionally substituted C-C 20 Alkenyl, optionally substituted C-C 20 alkynyl, optionally substituted cycloalkyl, optionally substituted (C1-C6 alkyl)cycloalkyl, optionally substituted heterocycloalkyl, optionally substituted (C1-C6 alkyl)heterocycloalkyl, optionally substituted aryl, optionally substituted (C1-C6 alkyl)aryl, optionally substituted heteroaryl, or optionally substituted (C1-C6 alkyl)heteroaryl; R 12 and R 13 are each independently hydrogen, halogen, optionally substituted C1-C6 alkyl, optionally substituted C2-C6 alkenyl, optionally substituted C2-C6 alkynyl, optionally substituted cycloalkyl, optionally substituted (C1-C6 alkyl)cycloalkyl, optionally substituted heterocycloalkyl, optionally substituted (C1-C6 alkyl)heterocycloalkyl, optionally substituted aryl, optionally substituted (C1-C6 alkyl)aryl, optionally substituted heteroaryl, or optionally substituted (C1-C6 alkyl)heteroaryl; Or, R 10 and R 12 taken together form an optionally substituted cycloalkyl or an optionally substituted cycloalkenyl; R14 and R 15 are each independently hydrogen, halogen, optionally substituted C1-C6 alkyl, optionally substituted C2-C6 alkenyl, optionally substituted C2-C6 alkynyl, optionally substituted cycloalkyl, optionally substituted (C1-C6 alkyl)cycloalkyl, optionally substituted heterocycloalkyl, optionally substituted (C1-C6 alkyl)heterocycloalkyl, optionally substituted aryl, optionally substituted (C1-C6 alkyl)aryl, optionally substituted heteroaryl, or optionally substituted (C1-C6 alkyl)heteroaryl; Or, R 14 and R 4a together with the atom to which they are attached form an optionally substituted heterocycloalkyl; R 16 and R 17 are each independently hydrogen, halogen, optionally substituted C1-C6 alkyl, optionally substituted C2-C6 alkenyl, optionally substituted C2-C6 alkynyl, optionally substituted cycloalkyl, optionally substituted (C1-C6 alkyl)cycloalkyl, optionally substituted heterocycloalkyl, optionally substituted (C1-C6 alkyl)heterocycloalkyl, optionally substituted aryl, optionally substituted (C1-C6 alkyl)aryl, optionally substituted heteroaryl, or optionally substituted (C1-C6 alkyl)heteroaryl; R 18 and R 19 are each independently hydrogen, halogen, optionally substituted C1-C6 alkyl, optionally substituted C2-C6 alkenyl, optionally substituted C2-C6 alkynyl, optionally substituted cycloalkyl, optionally substituted (C1-C6 alkyl)cycloalkyl, optionally substituted heterocycloalkyl, optionally substituted (C1-C6 alkyl)heterocycloalkyl, optionally substituted aryl, optionally substituted (C1-C6 alkyl)aryl, optionally substituted heteroaryl, or optionally substituted (C1-C6 alkyl)heteroaryl; Or, R 18and R 19 together with the carbon atom to which they are attached form oxo, Each R a are independently optionally substituted C1-C6 alkyl, optionally substituted C2-C6 alkenyl, optionally substituted C2-C6 alkynyl, optionally substituted cycloalkyl, optionally substituted heterocycloalkyl, optionally substituted aryl, or optionally substituted heteroaryl; Each R b and R c are independently hydrogen, optionally substituted C1-C6 alkyl, optionally substituted C2-C6 alkenyl, optionally substituted C2-C6 alkynyl, optionally substituted cycloalkyl, optionally substituted heterocycloalkyl, optionally substituted aryl, or optionally substituted heteroaryl; Or, R b and R c together with the nitrogen atom to which they are attached form an optionally substituted heterocycloalkyl; or a pharmaceutically acceptable salt, solvate or stereoisomer thereof, The compound is JPEG0007670618000005.jpg219170JPEG0007670618000006.jpg224170JPEG0007670618000007.jpg233170JPEG0007670618000008.jpg233170, or a pharmaceutically acceptable salt, solvate, or stereoisomer thereof.

[0008] Also disclosed herein are pharmaceutical compositions comprising a compound disclosed herein, or a pharmaceutically acceptable salt, solvate, or stereoisomer thereof, and a pharmaceutically acceptable excipient.

[0009] Also disclosed herein is the use of a compound disclosed herein, or a pharmaceutically acceptable salt, solvate, or stereoisomer thereof, to prepare a medicament for treating a bacterial infection in a patient.

[0010] Also disclosed herein is a method for treating a bacterial infection in a mammal, comprising administering to the mammal an effective amount of a compound disclosed herein, or a pharmaceutically acceptable salt, solvate, or stereoisomer thereof, at a frequency and for a duration sufficient to impart a beneficial effect to the mammal.

[0011] Incorporation by Reference All publications, patents, and patent applications mentioned in this specification are herein incorporated by reference to the same extent as if each individual publication, patent, or patent application was specifically and individually indicated to be incorporated by reference. DETAILED DESCRIPTION OF THE INVENTION

[0012] definition As used herein and in the appended claims, the singular forms "a," "and," and "the" include plural referents unless the context clearly dictates otherwise. Thus, for example, reference to "an agent" includes a plurality of such agents, and reference to "the cell" includes reference to one or more cells and equivalents thereof known to those of skill in the art, and so forth. When ranges are used herein for physical properties, such as molecular weight, or chemical properties, such as chemical formulas, all combinations and subcombinations of ranges and specific embodiments therein are intended to be included. The term "about" when referring to a number or numerical range means that the referenced number or numerical range is an approximation within experimental variability (or statistical experimental error); therefore, the number or numerical range will, in some instances, vary by 1% to 15% of the stated number or numerical range. The term "comprising" (and related terms such as "comprise" or "comprises" or "having" or "including") is not intended to exclude certain other embodiments, such as any compositions of matter, compositions, methods or processes described herein, from "consisting of" or "consist essentially of" the recited features.

[0013] As used in this specification and the appended claims, unless specified to the contrary, the following terms have the meanings indicated below.

[0014] "Alkyl" refers to an optionally substituted straight-chain or optionally substituted branched-chain saturated hydrocarbon monoradical having 1 to about 10 carbon atoms, or 1 to 6 carbon atoms, wherein an sp3 hybridized carbon of the alkyl residue is attached to the rest of the molecule by a single bond. Examples include, but are not limited to, methyl, ethyl, n-propyl, isopropyl, 2-methyl-1-propyl, 2-methyl-2-propyl, 2-methyl-1-butyl, 3-methyl-1-butyl, 2-methyl-3-butyl, 2,2-dimethyl-1-propyl, 2-methyl-1-pentyl, 3-methyl-1-pentyl, 4-methyl-1-pentyl, 2-methyl-2-pentyl, 3-methyl-2-pentyl, 4-methyl-2-pentyl, 2,2-dimethyl-1-butyl, 3,3-dimethyl-1-butyl, 2-ethyl-1-butyl, n-butyl, isobutyl, sec-butyl, t-butyl, n-pentyl, isopentyl, neopentyl, tert-amyl and hexyl, as well as longer alkyl groups such as heptyl, octyl, and the like. Whenever a numerical range appears herein, such as "C1-C6 alkyl," it means that the alkyl group consists of 1 carbon atom, 2 carbon atoms, 3 carbon atoms, 4 carbon atoms, 5 carbon atoms, or 6 carbon atoms; however, this definition also encompasses occurrences of the term "alkyl" where no numerical range is specified. In some embodiments, alkyl is any of C1-C6 alkyls. 20 Alkyl, C1-C 10The alkyl group may be alkyl, C1-C9 alkyl, C1-C8 alkyl, C1-C7 alkyl, C1-C6 alkyl, C1-C5 alkyl, C1-C4 alkyl, C1-C3 alkyl, C1-C2 alkyl, or C1 alkyl. Unless specifically stated otherwise in this specification, an alkyl group is optionally substituted as described below, for example, with oxo, halogen, amino, nitrile, nitro, hydroxyl, haloalkyl, alkoxy, aryl, cycloalkyl, heterocycloalkyl, heteroaryl, etc. In some embodiments, an alkyl is optionally substituted with oxo, halogen, —CN, —CF3, —OH, —OMe, —NH2, or —NO2. In some embodiments, an alkyl is optionally substituted with oxo, halogen, —CN, —CF3, —OH, or —OMe. In some embodiments, an alkyl is optionally substituted with halogen.

[0015] "Alkenyl" refers to an optionally substituted straight-chain or optionally substituted branched-chain hydrocarbon monoradical having one or more carbon-carbon double bonds and having from 2 to about 10 carbon atoms, more preferably from 2 to about 6 carbon atoms, where the sp2 hybridized carbon of the alkenyl residue is attached to the remainder of the molecule by a single bond. This group may be in either the cis or trans configuration about the double bond and should be understood to include both isomers. Examples include, but are not limited to, ethenyl (-CH=CH), 1-propenyl (-CHCH=CH), isopropenyl [-C(CH)=CH], butenyl, 1,3-butadienyl, and the like. Whenever a numerical range such as "C2-C6 alkenyl" appears herein, it means that the alkenyl group can consist of 2 carbon atoms, 3 carbon atoms, 4 carbon atoms, 5 carbon atoms, or 6 carbon atoms; however, this definition also encompasses occurrences of the term "alkenyl" where no numerical range is specified. In some embodiments, alkenyl is C-C 20 Alkenyl, C2-C 10Alkenyl is C2-C9 alkenyl, C2-C8 alkenyl, C2-C7 alkenyl, C2-C6 alkenyl, C2-C5 alkenyl, C2-C4 alkenyl, C2-C3 alkenyl, or C2 alkenyl. Unless specifically stated otherwise in this specification, alkenyl groups are optionally substituted as described below, for example, with oxo, halogen, amino, nitrile, nitro, hydroxyl, haloalkyl, alkoxy, aryl, cycloalkyl, heterocycloalkyl, heteroaryl, etc. In some embodiments, alkenyl is optionally substituted with oxo, halogen, -CN, -CF3, -OH, -OMe, -NH2, or -NO2. In some embodiments, alkenyl is optionally substituted with oxo, halogen, -CN, -CF3, -OH, or -OMe. In some embodiments, alkenyl is optionally substituted with halogen.

[0016] "Alkynyl" refers to an optionally substituted straight-chain or optionally substituted branched-chain hydrocarbon monoradical having one or more carbon-carbon triple bonds and having from 2 to about 10 carbon atoms, more preferably from 2 to about 6 carbon atoms. Examples include, but are not limited to, ethynyl, 2-propynyl, 2-butynyl, 1,3-butadiynyl, and the like. Whenever it appears herein, a numerical range such as "C2-C6 alkynyl" means that the alkynyl group can consist of 2 carbon atoms, 3 carbon atoms, 4 carbon atoms, 5 carbon atoms, or 6 carbon atoms, although this definition also encompasses occurrences of the term "alkynyl" where no numerical range is specified. In some embodiments, alkynyl refers to a C2-C6 alkynyl group. 20 Alkynyl, C2-C 10alkynyl, C2-C9 alkynyl, C2-C8 alkynyl, C2-C7 alkynyl, C2-C6 alkynyl, C2-C5 alkynyl, C2-C4 alkynyl, C2-C3 alkynyl, or C2 alkynyl. Unless specifically stated otherwise in this specification, alkynyl groups are optionally substituted as described below, for example, with oxo, halogen, amino, nitrile, nitro, hydroxyl, haloalkyl, alkoxy, aryl, cycloalkyl, heterocycloalkyl, heteroaryl, etc. In some embodiments, alkynyl is optionally substituted with oxo, halogen, -CN, -CF3, -OH, -OMe, -NH2, or -NO2. In some embodiments, alkynyl is optionally substituted with oxo, halogen, -CN, -CF3, -OH, or -OMe. In some embodiments, alkynyl is optionally substituted with halogen.

[0017] "Alkylene" refers to a straight or branched divalent hydrocarbon chain. Unless specifically stated otherwise in this specification, alkylene groups can be optionally substituted as described below, for example, with oxo, halogen, amino, nitrile, nitro, hydroxyl, haloalkyl, alkoxy, aryl, cycloalkyl, heterocycloalkyl, heteroaryl, etc. In some embodiments, alkylene is optionally substituted with oxo, halogen, -CN, -CF3, -OH, -OMe, -NH2, or -NO2. In some embodiments, alkylene is optionally substituted with oxo, halogen, -CN, -CF3, -OH, or -OMe. In some embodiments, alkylene is optionally substituted with halogen.

[0018] "Alkoxy" means a group of the formula -OR a where R ais an alkyl group as defined. Unless specifically stated otherwise in this specification, an alkoxy group can be optionally substituted as described below, for example, with oxo, halogen, amino, nitrile, nitro, hydroxyl, haloalkyl, alkoxy, aryl, cycloalkyl, heterocycloalkyl, heteroaryl, etc. In some embodiments, an alkoxy is optionally substituted with oxo, halogen, -CN, -CF3, -OH, -OMe, -NH2, or -NO2. In some embodiments, an alkoxy is optionally substituted with oxo, halogen, -CN, -CF3, -OH, or -OMe. In some embodiments, an alkoxy is optionally substituted with halogen.

[0019] "Aryl" refers to a group derived from a hydrocarbon ring system containing hydrogen, 6 to 30 carbon atoms, and at least one aromatic ring. Aryl groups can be monocyclic, bicyclic, tricyclic, or tetracyclic ring systems and can include fused ring systems (when fused to a cycloalkyl or heterocycloalkyl ring, the aryl is attached through an aromatic ring atom) or bridged ring systems. In some embodiments, an aryl is a 6- to 10-membered aryl. In some embodiments, an aryl is a 6-membered aryl. Aryl groups include, but are not limited to, aryl groups derived from the hydrocarbon ring systems of anthrylene, naphthylene, phenanthrylene, anthracene, azulene, benzene, chrysene, fluoranthene, fluorene, as-indacene, s-indacene, indane, indene, naphthalene, phenalene, phenanthrene, pleiadene, pyrene, and triphenylene. In some embodiments, an aryl is phenyl. Unless otherwise specifically stated herein, aryl can be optionally substituted as described below, for example, with halogen, amino, nitrile, nitro, hydroxyl, alkyl, alkenyl, alkynyl, haloalkyl, alkoxy, aryl, cycloalkyl, heterocycloalkyl, heteroaryl, etc. In some embodiments, aryl is optionally substituted with halogen, methyl, ethyl, -CN, -CF3, -OH, -OMe, -NH2, or -NO2. In some embodiments, aryl is optionally substituted with halogen, methyl, ethyl, -CN, -CF3, -OH, or -OMe. In some embodiments, aryl is optionally substituted with halogen or methyl. In some embodiments, aryl is optionally substituted with halogen.

[0020] "Cycloalkyl" refers to a stable, partially or fully saturated, monocyclic or polycyclic carbocyclic ring, which may include fused (when fused to an aryl or heteroaryl ring, the cycloalkyl is attached through a non-aromatic ring atom) or bridged ring systems. Representative cycloalkyls include those having 3 to 15 carbon atoms (C3 to C6). 15Cycloalkyl), 3 to 10 carbon atoms (C3 to C 10 Examples of cycloalkyl include, but are not limited to, cycloalkyls having 3 to 8 carbon atoms (C-C cycloalkyl), 3 to 6 carbon atoms (C-C cycloalkyl), 3 to 5 carbon atoms (C-C cycloalkyl), or 3 to 4 carbon atoms (C-C cycloalkyl). In some embodiments, cycloalkyls are 3- to 6-membered cycloalkyls. In some embodiments, cycloalkyls are 5- to 6-membered cycloalkyls. Monocyclic cycloalkyls include, for example, cyclopropyl, cyclobutyl, cyclopentyl, cyclohexyl, cycloheptyl, and cyclooctyl. Examples of polycyclic cycloalkyl or carbocycle include adamantyl, norbornyl, decalinyl, bicyclo[3.3.0]octane, bicyclo[4.3.0]nonane, cis-decalin, trans-decalin, bicyclo[2.1.1]hexane, bicyclo[2.2.1]heptane, bicyclo[2.2.2]octane, bicyclo[3.2.2]nonane, and bicyclo[3.3.2]decane, and 7,7-dimethyl-bicyclo[2.2.1]heptanyl. Examples of partially saturated cycloalkyl include cyclopentenyl, cyclohexenyl, cycloheptenyl, and cyclooctenyl. Unless otherwise specified in the specification, cycloalkyl is optionally substituted as described below, for example, with oxo, halogen, amino, nitrile, nitro, hydroxyl, alkyl, alkenyl, alkynyl, haloalkyl, alkoxy, aryl, cycloalkyl, heterocycloalkyl, heteroaryl, etc. In some embodiments, cycloalkyl is optionally substituted with oxo, halogen, methyl, ethyl, -CN, -CF3, -OH, -OMe, -NH2, or -NO2. In some embodiments, cycloalkyl is optionally substituted with oxo, halogen, methyl, ethyl, -CN, -CF3, -OH, or -OMe. In some embodiments, cycloalkyl is optionally substituted with halogen or methyl. In some embodiments, cycloalkyl is optionally substituted with halogen.

[0021] "Halo" or "halogen" refers to bromo, chloro, fluoro, or iodo. In some embodiments, halogen is fluoro or chloro. In some embodiments, halogen is fluoro.

[0022] "Haloalkyl" refers to an alkyl group, as defined above, that is substituted by one or more halo groups, as defined above, e.g., trifluoromethyl, difluoromethyl, fluoromethyl, trichloromethyl, 2,2,2-trifluoroethyl, 1,2-difluoroethyl, 3-bromo-2-fluoropropyl, 1,2-dibromoethyl, and the like.

[0023] "Heterocycloalkyl" refers to a stable 3- to 24-membered, partially or fully saturated ring radical containing 2 to 23 carbon atoms and 1 to 8 heteroatoms selected from the group consisting of nitrogen, oxygen, phosphorus, and sulfur. Unless specifically stated otherwise in the specification, a heterocycloalkyl group can be a monocyclic, bicyclic, tricyclic, or tetracyclic ring system, and can include fused (when fused to an aryl or heteroaryl ring, the heterocycloalkyl is attached through a non-aromatic ring atom) or bridged ring systems, in which the nitrogen, carbon, or sulfur atoms in the heterocycloalkyl group can be optionally oxidized, and the nitrogen atom can be optionally quaternized. In some embodiments, a heterocycloalkyl is a 3- to 6-membered heterocycloalkyl. In some embodiments, a heterocycloalkyl is a 5- to 6-membered heterocycloalkyl. Examples of such heterocycloalkyl groups include aziridinyl, azetidinyl, dioxolanyl, thienyl[1,3]dithianyl, decahydroisoquinolyl, imidazolinyl, imidazolidinyl, isothiazolidinyl, isoxazolidinyl, morpholinyl, octahydroindolyl, octahydroisoindolyl, 2-oxopiperazinyl, 2-oxopiperidinyl, 2-oxopyrrolidinyl, oxazolidinyl, piperidinyl, piperazinyl, 4-piperidonyl, pyrrolidinyl, Examples of heterocycloalkyl include, but are not limited to, pyrazolidinyl, quinuclidinyl, thiazolidinyl, tetrahydrofuryl, trithianyl, tetrahydropyranyl, thiomorpholinyl, thiamorpholinyl, 1-oxo-thiomorpholinyl, 1,1-dioxo-thiomorpholinyl, 1,3-dihydroisobenzofuran-1-yl, 3-oxo-1,3-dihydroisobenzofuran-1-yl, methyl-2-oxo-1,3-dioxol-4-yl, and 2-oxo-1,3-dioxol-4-yl. The term heterocycloalkyl also includes all ring forms of carbohydrates, including, but not limited to, monosaccharides, disaccharides, and oligosaccharides. Unless otherwise specified, heterocycloalkyls have 2 to 10 carbons in the ring.When referring to the number of carbon atoms in a heterocycloalkyl, it is understood that the number of carbon atoms in the heterocycloalkyl is not the same as the total number of atoms (including heteroatoms) comprising the heterocycloalkyl (i.e., skeletal atoms of the heterocycloalkyl ring). Unless specifically stated otherwise in this specification, a heterocycloalkyl is optionally substituted as described below, for example, with oxo, halogen, amino, nitrile, nitro, hydroxyl, alkyl, alkenyl, alkynyl, haloalkyl, alkoxy, aryl, cycloalkyl, heterocycloalkyl, heteroaryl, etc. In some embodiments, a heterocycloalkyl is optionally substituted with oxo, halogen, methyl, ethyl, -CN, -CF3, -OH, -OMe, -NH2, or -NO2. In some embodiments, a heterocycloalkyl is optionally substituted with oxo, halogen, methyl, ethyl, -CN, -CF3, -OH, or -OMe. In some embodiments, a heterocycloalkyl is optionally substituted with halogen or methyl. In some embodiments, a heterocycloalkyl is optionally substituted with halogen.

[0024] "Heteroalkyl" refers to an alkyl group in which one or more skeletal atoms of the alkyl are selected from atoms other than carbon, such as oxygen, nitrogen (e.g., -NH-, -N(alkyl)-), sulfur, or combinations thereof. The heteroalkyl is attached to the remainder of the molecule at a carbon atom of the heteroalkyl. In one aspect, the heteroalkyl is a C1-C6 heteroalkyl. Unless specifically stated otherwise in this specification, the heteroalkyl is optionally substituted as described below, for example, with oxo, halogen, amino, nitrile, nitro, hydroxyl, alkyl, alkenyl, alkynyl, haloalkyl, alkoxy, aryl, cycloalkyl, heterocycloalkyl, heteroaryl, etc. In some embodiments, the heteroalkyl is optionally substituted with oxo, halogen, methyl, ethyl, -CN, -CF3, -OH, -OMe, -NH2, or -NO2. In some embodiments, the heteroalkyl is optionally substituted with oxo, halogen, methyl, ethyl, -CN, -CF3, -OH, or -OMe. In some embodiments, the heteroalkyl is optionally substituted with halogen.

[0025] "Heteroaryl" refers to a 5- to 14-membered ring system containing a hydrogen atom, 1 to 13 carbon atoms, 1 to 6 heteroatoms selected from the group consisting of nitrogen, oxygen, phosphorus, and sulfur, and at least one aromatic ring. Heteroaryl groups can be monocyclic, bicyclic, tricyclic, or tetracyclic ring systems, and can include fused (when fused to a cycloalkyl or heterocycloalkyl ring, the heteroaryl is bonded through an aromatic ring atom) or bridged ring systems. The nitrogen, carbon, or sulfur atoms in the heteroaryl group can be optionally oxidized, and the nitrogen atom can be optionally quaternized. In some embodiments, the heteroaryl is a 5- to 10-membered heteroaryl. In some embodiments, the heteroaryl is a 5- to 6-membered heteroaryl.Examples include azepinyl, acridinyl, benzimidazolyl, benzothiazolyl, benzoindolyl, benzodioxolyl, benzofuranyl, benzoxazolyl, benzothiazolyl, benzothiadiazolyl, benzo[b][1,4]dioxepinyl, 1,4-benzodioxanyl, benzonaphthofuranyl, benzoxazolyl, benzodioxolyl, benzodioxinyl, benzopyranyl, benzopyranonyl, benzofuranyl, benzofuranonyl, benzothienyl (benzothiophenyl), benzotriazolyl, benzo[4,6]imidazo[1,2-a]pyridinyl, carbazolyl, cinnolinyl, dibenzofuranyl, dibenzothiophenyl, furanyl, furanonyl, isothiazolyl, imidazolyl, indazolyl, indolyl, indazolyl, isoindolyl, indophenyl, and indophenyl. Examples include, but are not limited to, dolinyl, isoindolinyl, isoquinolyl, indolizinyl, isoxazolyl, naphthyridinyl, oxadiazolyl, 2-oxoazepinyl, oxazolyl, oxiranyl, 1-oxidopyridinyl, 1-oxidopyrimidinyl, 1-oxidopyrazinyl, 1-oxidopyridazinyl, 1-phenyl-1H-pyrrolyl, phenazinyl, phenothiazinyl, phenoxazinyl, phthalazinyl, pteridinyl, purinyl, pyrrolyl, pyrazolyl, pyridinyl, pyrazinyl, pyrimidinyl, pyridazinyl, quinazolinyl, quinoxalinyl, quinolinyl, quinuclidinyl, isoquinolinyl, tetrahydroquinolinyl, thiazolyl, thiadiazolyl, triazolyl, tetrazolyl, triazinyl, and thiophenyl (i.e., thienyl). Unless specifically stated otherwise in this specification, heteroaryl is optionally substituted as described below, for example, with halogen, amino, nitrile, nitro, hydroxyl, alkyl, alkenyl, alkynyl, haloalkyl, alkoxy, aryl, cycloalkyl, heterocycloalkyl, heteroaryl, etc. In some embodiments, heteroaryl is optionally substituted with halogen, methyl, ethyl, -CN, -CF, -OH, -OMe, -NH, or -NO. In some embodiments, heteroaryl is optionally substituted with halogen, methyl, ethyl, -CN, -CF, -OH, or -OMe.In some embodiments, the heteroaryl is optionally substituted with halogen or methyl. In some embodiments, the heteroaryl is optionally substituted with halogen.

[0026] The term "oxo" means =O.

[0027] As used herein, the terms "treat," "prevent," "ameliorate," and "inhibit," as well as words derived therefrom, do not necessarily mean 100% or complete treatment, prevention, amelioration, or inhibition. Rather, there are various degrees of treatment, prevention, amelioration, and inhibition that those skilled in the art will recognize as having potential benefit or therapeutic effect. In this regard, the disclosed methods can provide any amount of treatment, prevention, amelioration, or inhibition of a disorder in a mammal at any level. For example, a disorder, including its symptoms or symptoms, may be reduced by, for example, about 100%, about 90%, about 80%, about 70%, about 60%, about 50%, about 40%, about 30%, about 20%, or about 10%. Furthermore, the treatment, prevention, amelioration, or inhibition provided by the methods disclosed herein can include treatment, prevention, amelioration, or inhibition of one or more symptoms or symptoms of a disorder, such as cancer or an inflammatory disease. Also, as used herein, "treatment," "prevention," "amelioration," or "inhibition" encompasses delaying the onset of a disorder, or its symptoms or symptoms.

[0028] As used herein, the term "effective amount" or "therapeutically effective amount" refers to a sufficient amount of a compound disclosed herein being administered to alleviate to some extent one or more symptoms of the disease or condition being treated, such as cancer or inflammatory disease. In some embodiments, the result is a reduction and / or alleviation of the signs, symptoms, or causes of a disease, or any other desired change in a biological system. For example, an "effective amount" for therapeutic use is the amount of a composition comprising a compound disclosed herein required to produce a clinically significant reduction in a disease symptom. In some embodiments, an appropriate "effective" amount in any individual case is determined using techniques such as a dose escalation study.

[0029] As used herein, "individual" (as in the subject of treatment) refers to both mammals and non-mammals. Mammals include, for example, humans, non-human primates such as apes and monkeys, and non-primates such as dogs, cats, cows, horses, sheep, and goats. Non-mammals include, for example, fish and birds.

[0030] The terms "disease" and "disorder" are used interchangeably to refer to a disease or condition in which a bacterial signal peptidase plays a role in the biochemical mechanisms involved in the disease or disorder, such that a therapeutically beneficial effect can be achieved by acting on the enzyme. "Affecting" a signal peptidase can include binding to and / or inhibiting the biological activity of the signal peptidase.

[0031] compound In one embodiment, a compound of formula (I): TIFF0007670618000009.tif80170 formula: R 1a , R 2a , R 4a , R 5a and R 6a are each independently hydrogen or optionally substituted C1-C6 alkyl, R3a is hydrogen, optionally substituted C1-C 20 Alkyl, optionally substituted C-C 20 Alkenyl, optionally substituted C-C 20 alkynyl, optionally substituted cycloalkyl, optionally substituted (C1-C6 alkyl)cycloalkyl, optionally substituted heterocycloalkyl, optionally substituted (C1-C6 alkyl)heterocycloalkyl, optionally substituted aryl, optionally substituted (C1-C6 alkyl)aryl, optionally substituted heteroaryl, or optionally substituted (C1-C6 alkyl)heteroaryl; R 20 is hydroxyl or -NR 1 R 2 and R 1 and R 2 are each independently hydrogen, optionally substituted C1-C6 alkyl, optionally substituted C2-C6 alkenyl, optionally substituted C2-C6 alkynyl, optionally substituted cycloalkyl, optionally substituted (C1-C6 alkyl)cycloalkyl, optionally substituted heterocycloalkyl, optionally substituted (C1-C6 alkyl)heterocycloalkyl, optionally substituted aryl, optionally substituted (C1-C6 alkyl)aryl, optionally substituted heteroaryl, optionally substituted (C1-C6 alkyl)heteroaryl, -S(=O)2R a , -S(=O)2NR b R c , -C(=O)R a , -C(=O)OR b , -C(=O)NR b R c Or -C(=NR b )NR b R c Or Or, R 1 and R 2 together with the nitrogen atom to which they are attached form an optionally substituted heterocycloalkyl; R 3 and R 4are each independently hydrogen, optionally substituted C1-C6 alkyl, optionally substituted C2-C6 alkenyl, optionally substituted C2-C6 alkynyl, optionally substituted cycloalkyl, optionally substituted heterocycloalkyl, optionally substituted aryl, or optionally substituted heteroaryl; Or, R 3 and R 4 together with the carbon atom to which they are attached form oxo, R 5 is hydrogen, halogen, optionally substituted C1-C6 alkyl, optionally substituted C2-C6 alkenyl, or optionally substituted C2-C6 alkynyl; R 6 and R 7 are each independently hydrogen, halogen, optionally substituted C1-C6 alkyl, optionally substituted C2-C6 alkenyl, optionally substituted C2-C6 alkynyl, optionally substituted cycloalkyl, optionally substituted (C1-C6 alkyl)cycloalkyl, optionally substituted heterocycloalkyl, optionally substituted (C1-C6 alkyl)heterocycloalkyl, optionally substituted aryl, optionally substituted (C1-C6 alkyl)aryl, optionally substituted heteroaryl, or optionally substituted (C1-C6 alkyl)heteroaryl; R 8 and R 9 are each independently hydrogen, halogen, optionally substituted C1-C6 alkyl, optionally substituted C2-C6 alkenyl, optionally substituted C2-C6 alkynyl, optionally substituted cycloalkyl, optionally substituted (C1-C6 alkyl)cycloalkyl, optionally substituted heterocycloalkyl, optionally substituted (C1-C6 alkyl)heterocycloalkyl, optionally substituted aryl, optionally substituted (C1-C6 alkyl)aryl, optionally substituted heteroaryl, or optionally substituted (C1-C6 alkyl)heteroaryl; Or, R 3a and R 8together with the atom to which they are attached form an optionally substituted heterocycloalkyl; R 10 and R 11 are each independently hydrogen, halogen, optionally substituted C1-C6 alkyl, optionally substituted C2-C 20 alkenyl, optionally substituted C2-C6 alkynyl, optionally substituted cycloalkyl, optionally substituted (C1-C6 alkyl)cycloalkyl, optionally substituted heterocycloalkyl, optionally substituted (C1-C6 alkyl)heterocycloalkyl, optionally substituted aryl, optionally substituted (C1-C6 alkyl)aryl, optionally substituted heteroaryl, or optionally substituted (C1-C6 alkyl)heteroaryl; R 12 and R 13 are each independently hydrogen, halogen, or an optionally substituted C1-C 20 alkyl, optionally substituted C2-C6 alkenyl, optionally substituted C2-C6 alkynyl, optionally substituted cycloalkyl, optionally substituted (C1-C6 alkyl)cycloalkyl, optionally substituted heterocycloalkyl, optionally substituted (C1-C6 alkyl)heterocycloalkyl, optionally substituted aryl, optionally substituted (C1-C6 alkyl)aryl, optionally substituted heteroaryl, or optionally substituted (C1-C6 alkyl)heteroaryl; R 14 and R 15 are each independently hydrogen, halogen, optionally substituted C1-C6 alkyl, optionally substituted C2-C6 alkenyl, optionally substituted C2-C6 alkynyl, optionally substituted cycloalkyl, optionally substituted (C1-C6 alkyl)cycloalkyl, optionally substituted heterocycloalkyl, optionally substituted (C1-C6 alkyl)heterocycloalkyl, optionally substituted aryl, optionally substituted (C1-C6 alkyl)aryl, optionally substituted heteroaryl, or optionally substituted (C1-C6 alkyl)heteroaryl; R 16 and R 17are each independently hydrogen, halogen, optionally substituted C1-C6 alkyl, optionally substituted C2-C6 alkenyl, optionally substituted C2-C6 alkynyl, optionally substituted cycloalkyl, optionally substituted (C1-C6 alkyl)cycloalkyl, optionally substituted heterocycloalkyl, optionally substituted (C1-C6 alkyl)heterocycloalkyl, optionally substituted aryl, optionally substituted (C1-C6 alkyl)aryl, optionally substituted heteroaryl, or optionally substituted (C1-C6 alkyl)heteroaryl; Each R a are independently optionally substituted C1-C6 alkyl, optionally substituted C2-C6 alkenyl, optionally substituted C2-C6 alkynyl, optionally substituted cycloalkyl, optionally substituted heterocycloalkyl, optionally substituted aryl, or optionally substituted heteroaryl; Each R b and R c are independently hydrogen, optionally substituted C1-C6 alkyl, optionally substituted C2-C6 alkenyl, optionally substituted C2-C6 alkynyl, optionally substituted cycloalkyl, optionally substituted heterocycloalkyl, optionally substituted aryl, or optionally substituted heteroaryl; Or, R b and R c together with the nitrogen atom to which they are attached form an optionally substituted heterocycloalkyl, or a pharmaceutically acceptable salt, solvate or stereoisomer thereof described herein.

[0032] In some embodiments of the compounds of Formula (I), R 20 is hydroxyl. In some embodiments of the compounds of Formula (I), R 20 Ha-NR 1 R 2 is.

[0033] In some embodiments of the compounds of Formula (I), R 5 is hydrogen.

[0034] In some embodiments of the compounds of Formula (I), R 7 is hydrogen.

[0035] In some embodiments of the compounds of Formula (I), R 13 is hydrogen.

[0036] In some embodiments of the compounds of Formula (I), R 15 is hydrogen.

[0037] In some embodiments of the compounds of Formula (I), R 17 is hydrogen.

[0038] In some embodiments of the compound of Formula (I), the compound has the formula (Ia): It has the structure TIFF0007670618000010.tif81170.

[0039] In some embodiments of the compound of Formula (I), the compound has the formula (Ib): It has the structure TIFF0007670618000011.tif84170.

[0040] In some embodiments of the compounds of Formula (I), R 1 is hydrogen.

[0041] In some embodiments of the compounds of Formula (I), R 2 is hydrogen.

[0042] In some embodiments of the compounds of Formula (I), R 3 and R 4 is hydrogen.

[0043] In some embodiments of the compounds of Formula (I), R 1a is hydrogen or C1-C6 alkyl. In some embodiments of the compound of Formula (I), R 1a is hydrogen.

[0044] In some embodiments of the compounds of Formula (I), R 2a is hydrogen or C1-C6 alkyl. In some embodiments of the compound of Formula (I), R 2a is hydrogen.

[0045] In some embodiments of the compounds of Formula (I), R 4a is hydrogen or C1-C6 alkyl. In some embodiments of the compound of Formula (I), R 4a is hydrogen.

[0046] In some embodiments of the compounds of Formula (I), R 5a is hydrogen or C1-C6 alkyl. In some embodiments of the compound of Formula (I), R 5a is hydrogen.

[0047] In some embodiments of the compounds of Formula (I), R 6a is hydrogen or C1-C6 alkyl. In some embodiments of the compound of Formula (I), R 6a is hydrogen.

[0048] In some embodiments of the compounds of Formula (I), R 9 is hydrogen.

[0049] In some embodiments of the compounds of Formula (I), R 10 is hydrogen.

[0050] In some embodiments of the compounds of Formula (I), R 11 is hydrogen.

[0051] In some embodiments of the compounds of Formula (I), R 8 is hydrogen or C1-C6 alkyl optionally substituted with 1, 2 or 3 halogens.

[0052] In some embodiments of the compounds of Formula (I), R 8is hydrogen or C1-C6 alkyl.

[0053] In some embodiments of the compounds of Formula (I), R 3a and R 8 together with the atom to which they are attached form an optionally substituted heterocycloalkyl.

[0054] In some embodiments of the compounds of Formula (I), R 3a and R 8 together with the atom to which they are attached form an azetidinyl, pyrrolidinyl, or piperidinyl.

[0055] In some embodiments of the compounds of Formula (I), R 6 is optionally substituted C1-C6 alkyl or optionally substituted heterocycloalkyl.

[0056] In some embodiments of the compounds of Formula (I), R 6 is an optionally substituted C1-C6 alkyl.

[0057] In some embodiments of the compounds of Formula (I), R 6 is C1-C6 alkyl optionally substituted with 1, 2 or 3 halogens, -OR b , -NR b R c , -NC(=NR b )NR b R c , -S(=O)2R a , -NR b S(=O)2R a , -S(=O)2NR b R c , -C(=O)R a , -OC(=O)R a , -C(=O)OR a , -OC(=O)OR b , -C(=O)NR b R c , -NR b C(=O)[C(R d )2] 1~4 NRb R c , -O[C(R d )2] 2~4 OR b , -O[C(R d )2] 2~4 NR b R c , -NR b [C(R d )2] 2~4 NR b R c , -O[C(R d )2] 2~4 O[C(R d )2] 2~4 NR b R c , -O[C(R d )2] 2~4 NC(=NR b )NR b R c , -OC(=O)NR b R c , -NR b C(=O)NR b R c , -NR b C(=O)R a , -NR b C(=O)OR b , -O-(cycloalkyl)NR b R c , —O-(heterocycloalkyl)NR b R c , -O-(aryl)NR b R c , —O-(heteroaryl)NR b R c , -O-(cycloalkyl)(C1-C6 alkyl)NR b R c , -O-(heterocycloalkyl)(C1-C6 alkyl)NR b R c , -O-(aryl)(C1-C6 alkyl)NR b R c , -O-(heteroaryl)(C1-C6 alkyl)NR b R c , aryl, or heteroaryl, and each R dare independently hydrogen, halogen, —OH, —OCH3, or C1-C6 alkyl.

[0058] In some embodiments of the compounds of Formula (I), R 6 can be 1, 2 or 3 -OR b , -NR b R c , -C(=O)NR b R c , -O[C(R d )2] 2~4 NR b R c , -NR b [C(R d )2] 2~4 NR b R c , -O-(cycloalkyl)NR b R c , -O-(cycloalkyl)(C1-C6 alkyl)NR b R c and each R is a C1-C6 alkyl optionally substituted with d , are independently hydrogen, halogen, —OH, —OCH3, or C1-C6 alkyl.

[0059] In some embodiments of the compounds of Formula (I), R 6 can be 1, 2 or 3 -OR b is C1-C6 alkyl optionally substituted with

[0060] In some embodiments of the compounds of Formula (I), R 6 teeth, JPEG0007670618000012.jpg242170JPEG0007670618000013.jpg186170. In some embodiments of the compound of Formula (I), R 6 teeth, JPEG0007670618000014.jpg71170. In some embodiments of the compound of Formula (I), R 6 teeth, JPEG0007670618000015.jpg17170.

[0061] In some embodiments of the compounds of Formula (I), R 12 is hydrogen or optionally substituted C1-C6 alkyl. In some embodiments of the compound of Formula (I), R 12 is C1-C6 alkyl optionally substituted with 1, 2, or 3 halogen or -OCH3. In some embodiments of the compound of Formula (I), R 12 is C1-C6 alkyl or C1-C6 haloalkyl. In some embodiments of the compound of Formula (I), R 12 teeth, JPEG0007670618000016.jpg33170.

[0062] In some embodiments of the compounds of Formula (I), R 14 is optionally substituted C1-C6 alkyl or optionally substituted (C1-C6 alkyl)cycloalkyl. In some embodiments of the compound of Formula (I), R 14 is a C1-C6 alkyl.

[0063] In some embodiments of the compounds of Formula (I), R 14 teeth JPEG0007670618000017.jpg30170. In some embodiments of the compound of Formula (I), R 14 teeth, JPEG0007670618000018.jpg23170.

[0064] In some embodiments of the compounds of Formula (I), R 16 is optionally substituted C1-C6 alkyl or optionally substituted cycloalkyl. In some embodiments of the compound of Formula (I), R 16 is C3-C7 cycloalkyl. In some embodiments of the compounds of Formula (I), R 16 is cyclohexyl.

[0065] In some embodiments of the compounds of Formula (I), R16 teeth, JPEG0007670618000019.jpg85170.

[0066] In some embodiments of the compounds of Formula (I), R 16 teeth, JPEG0007670618000020.jpg49170. In some embodiments of the compound of Formula (I), R 16 teeth, JPEG0007670618000021.jpg23170.

[0067] In some embodiments of the compounds of Formula (I), R 3a is an optionally substituted C1 to C 20 Alkyl, optionally substituted C-C 20 In some embodiments of the compound of Formula (I), R is alkynyl, optionally substituted (C1-C6 alkyl)cycloalkyl, or optionally substituted (C1-C6 alkyl)aryl. 3a is C1-C6 alkyl optionally substituted with 1, 2, or 3 halogens. In some embodiments of the compound of Formula (I), R 3a is (C1-C6 alkyl)aryl optionally substituted with 1, 2, or 3 C1-C6 alkyl. In some embodiments of compounds of Formula (I), R 3a is (C1-C6 alkyl)cycloalkyl. In some embodiments of the compound of Formula (I), R 3a teeth, JPEG0007670618000022.jpg147170. In some embodiments of the compound of Formula (I), R 3a teeth, JPEG0007670618000023.jpg67170.

[0068] In some embodiments of the compounds of Formula (I), each R a is independently C1 to C6 alkyl.

[0069] In some embodiments of the compounds of Formula (I), each R b and R c are independently hydrogen or C1-C6 alkyl.

[0070] In one embodiment, a compound of formula (II): TIFF0007670618000024.tif81170 formula: R 1a , R 2a , R 3a , R 4a , R 5a and R 6a are each independently hydrogen or optionally substituted C1-C6 alkyl, R 1 and R 2 are each independently hydrogen, optionally substituted C1-C6 alkyl, optionally substituted C2-C6 alkenyl, optionally substituted C2-C6 alkynyl, optionally substituted cycloalkyl, optionally substituted (C1-C6 alkyl)cycloalkyl, optionally substituted heterocycloalkyl, optionally substituted (C1-C6 alkyl)heterocycloalkyl, optionally substituted aryl, optionally substituted (C1-C6 alkyl)aryl, optionally substituted heteroaryl, optionally substituted (C1-C6 alkyl)heteroaryl, -S(=O)2R a , -S(=O)2NR b R c , -C(=O)R a , -C(=O)OR b , -C(=O)NR b R c Or -C(=NR b )NR b R c Or Or, R 1 and R 2 together with the nitrogen atom to which they are attached form an optionally substituted heterocycloalkyl; R 3 and R 4are each independently hydrogen, optionally substituted C1-C6 alkyl, optionally substituted C2-C6 alkenyl, optionally substituted C2-C6 alkynyl, optionally substituted cycloalkyl, optionally substituted heterocycloalkyl, optionally substituted aryl, or optionally substituted heteroaryl; Or, R 3 and R 4 together with the carbon atom to which they are attached form oxo, R 5 is hydrogen, halogen, optionally substituted C1-C6 alkyl, optionally substituted C2-C6 alkenyl, or optionally substituted C2-C6 alkynyl; R 6 and R 7 are each independently hydrogen, halogen, optionally substituted C1-C6 alkyl, optionally substituted C2-C6 alkenyl, optionally substituted C2-C6 alkynyl, optionally substituted cycloalkyl, optionally substituted (C1-C6 alkyl)cycloalkyl, optionally substituted heterocycloalkyl, optionally substituted (C1-C6 alkyl)heterocycloalkyl, optionally substituted aryl, optionally substituted (C1-C6 alkyl)aryl, optionally substituted heteroaryl, or optionally substituted (C1-C6 alkyl)heteroaryl; R 8 and R 9 are each independently hydrogen, halogen, optionally substituted C1-C6 alkyl, optionally substituted C2-C6 alkenyl, optionally substituted C2-C6 alkynyl, optionally substituted cycloalkyl, optionally substituted (C1-C6 alkyl)cycloalkyl, optionally substituted heterocycloalkyl, optionally substituted (C1-C6 alkyl)heterocycloalkyl, optionally substituted aryl, optionally substituted (C1-C6 alkyl)aryl, optionally substituted heteroaryl, or optionally substituted (C1-C6 alkyl)heteroaryl; R 10 and R 11 are each independently hydrogen, halogen, or an optionally substituted C1-C 20Alkyl, optionally substituted C-C 20 Alkenyl, optionally substituted C-C 20 alkynyl, optionally substituted cycloalkyl, optionally substituted (C1-C6 alkyl)cycloalkyl, optionally substituted heterocycloalkyl, optionally substituted (C1-C6 alkyl)heterocycloalkyl, optionally substituted aryl, optionally substituted (C1-C6 alkyl)aryl, optionally substituted heteroaryl, or optionally substituted (C1-C6 alkyl)heteroaryl; R 12 and R 13 are each independently hydrogen, halogen, optionally substituted C1-C6 alkyl, optionally substituted C2-C6 alkenyl, optionally substituted C2-C6 alkynyl, optionally substituted cycloalkyl, optionally substituted (C1-C6 alkyl)cycloalkyl, optionally substituted heterocycloalkyl, optionally substituted (C1-C6 alkyl)heterocycloalkyl, optionally substituted aryl, optionally substituted (C1-C6 alkyl)aryl, optionally substituted heteroaryl, or optionally substituted (C1-C6 alkyl)heteroaryl; Or, R 10 and R 12 taken together form an optionally substituted cycloalkyl or an optionally substituted cycloalkenyl; R 14 and R 15 are each independently hydrogen, halogen, optionally substituted C1-C6 alkyl, optionally substituted C2-C6 alkenyl, optionally substituted C2-C6 alkynyl, optionally substituted cycloalkyl, optionally substituted (C1-C6 alkyl)cycloalkyl, optionally substituted heterocycloalkyl, optionally substituted (C1-C6 alkyl)heterocycloalkyl, optionally substituted aryl, optionally substituted (C1-C6 alkyl)aryl, optionally substituted heteroaryl, or optionally substituted (C1-C6 alkyl)heteroaryl; Or, R 14 and R 4atogether with the atom to which they are attached form an optionally substituted heterocycloalkyl; R 16 and R 17 are each independently hydrogen, halogen, optionally substituted C1-C6 alkyl, optionally substituted C2-C6 alkenyl, optionally substituted C2-C6 alkynyl, optionally substituted cycloalkyl, optionally substituted (C1-C6 alkyl)cycloalkyl, optionally substituted heterocycloalkyl, optionally substituted (C1-C6 alkyl)heterocycloalkyl, optionally substituted aryl, optionally substituted (C1-C6 alkyl)aryl, optionally substituted heteroaryl, or optionally substituted (C1-C6 alkyl)heteroaryl; R 18 and R 19 are each independently hydrogen, halogen, optionally substituted C1-C6 alkyl, optionally substituted C2-C6 alkenyl, optionally substituted C2-C6 alkynyl, optionally substituted cycloalkyl, optionally substituted (C1-C6 alkyl)cycloalkyl, optionally substituted heterocycloalkyl, optionally substituted (C1-C6 alkyl)heterocycloalkyl, optionally substituted aryl, optionally substituted (C1-C6 alkyl)aryl, optionally substituted heteroaryl, or optionally substituted (C1-C6 alkyl)heteroaryl; Or, R 18 and R 19 together with the carbon atom to which they are attached form oxo, Each R a are independently optionally substituted C1-C6 alkyl, optionally substituted C2-C6 alkenyl, optionally substituted C2-C6 alkynyl, optionally substituted cycloalkyl, optionally substituted heterocycloalkyl, optionally substituted aryl, or optionally substituted heteroaryl; Each R b and R care independently hydrogen, optionally substituted C1-C6 alkyl, optionally substituted C2-C6 alkenyl, optionally substituted C2-C6 alkynyl, optionally substituted cycloalkyl, optionally substituted heterocycloalkyl, optionally substituted aryl, or optionally substituted heteroaryl; Or, R b and R c together with the nitrogen atom to which they are attached form an optionally substituted heterocycloalkyl; Compounds of or a pharmaceutically acceptable salt, solvate or stereoisomer thereof, as described herein, The compound is JPEG0007670618000025.jpg220170JPEG0007670618000026.jpg225170JPEG0007670618000027.jpg234170JPEG0007670618000028.jpg234170.

[0071] In some embodiments, the compound disclosed herein is selected from the compounds in Table 1 or a pharmaceutically acceptable salt, solvate, or stereoisomer thereof. JPEG0007670618000029.jpg252170JPEG0007670618000030.jpg255170JPEG0007670618000031.jpg248170 JPEG0007670618000032.jpg255170JPEG0007670618000033.jpg255170JPEG0007670618000034.jpg224170 JPEG0007670618000035.jpg246170JPEG0007670618000036.jpg251170JPEG0007670618000037.jpg240170 JPEG0007670618000038.jpg255170JPEG0007670618000039.jpg240170JPEG0007670618000040.jpg255170 JPEG0007670618000041.jpg255170JPEG0007670618000042.jpg255170JPEG0007670618000043.jpg255170 JPEG0007670618000044.jpg216170JPEG0007670618000045.jpg247170JPEG0007670618000046.jpg214170 JPEG0007670618000047.jpg255170JPEG0007670618000048.jpg246170JPEG0007670618000049.jpg214170 JPEG0007670618000050.jpg255170JPEG0007670618000051.jpg255170JPEG0007670618000052.jpg153170

[0072] Further forms of the compounds disclosed herein Isomers / stereoisomers In some embodiments, the compounds described herein exist as geometric isomers. In some embodiments, the compounds described herein have one or more double bonds. The compounds presented herein include all cis, trans, syn, anti, entgegen (E), and zusammen (Z) isomers, as well as their corresponding mixtures. In some circumstances, the compounds described herein have one or more chiral centers, with each center existing in the R or S configuration. The compounds described herein include all diastereomeric, enantiomeric, and epimeric forms, as well as their corresponding mixtures. In further embodiments of the compounds and methods provided herein, mixtures of enantiomers and / or diastereoisomers resulting from a single preparation step, combination, or interconversion are useful for the applications described herein. In some embodiments, the compounds described herein are prepared as individual stereoisomers by reacting a racemic mixture of the compound with an optically active resolving agent to form a pair of diastereoisomeric compounds, separating the diastereomers, and recovering the optically pure enantiomers. In some embodiments, dissociable complexes are preferred. In some embodiments, diastereomers have distinct physical properties (e.g., melting points, boiling points, solubilities, reactivity, etc.) and are separated by taking advantage of these dissimilarities. In some embodiments, diastereomers are separated by chiral chromatography, or preferably by separation / resolution techniques based upon differences in solubility. In some embodiments, the optically pure enantiomers are then recovered with the resolving agent.

[0073] Labeled Compounds In some embodiments, the compounds described herein exist in isotopically labeled form. In some embodiments, the methods disclosed herein include methods of treating disease by administering such isotopically labeled compounds. In some embodiments, the methods disclosed herein include methods of treating disease by administering such isotopically labeled compounds as pharmaceutical compositions. Thus, in some embodiments, the compounds disclosed herein include isotopically labeled compounds that are identical to the compounds cited herein, except for the fact that one or more atoms are replaced by atoms with atomic masses or mass numbers different from those usually found in nature. Examples of isotopes that can be incorporated into the compounds described herein, or their solvates or stereoisomers, include isotopes of hydrogen, carbon, nitrogen, oxygen, phosphorus, sulfur, fluorine, and chlorine, for example, 2 H, 3 H, 13 C. 14 C. 15 N, 18 O. 17 O. 31 P, 32 P, 35 S, 18 F and 36 Certain isotopically labeled compounds, such as 3 H, 14 Those into which radioactive isotopes such as C are incorporated are useful in drug and / or substrate tissue distribution assays. 3 H, and carbon-14, i.e., 14 C isotopes are particularly preferred for their ease of preparation and detectability. 2 Substitution with heavy isotopes, such as H, can result in greater metabolic stability, for example, a longer in vivo half-life or reduced dosage requirements, resulting in certain therapeutic advantages. In some embodiments, the isotopically labeled compound or a pharmaceutically acceptable salt, solvate, or stereoisomer thereof is prepared by any suitable method.

[0074] In some embodiments, the compounds described herein are labeled by other means, including but not limited to, the use of chromophores or fluorescent moieties, bioluminescent labels, or chemiluminescent labels.

[0075] Pharmaceutically acceptable salts In some embodiments, the compounds described herein are present as their pharmaceutically acceptable salts. In some embodiments, the methods disclosed herein include methods of treating diseases by administering such pharmaceutically acceptable salts. In some embodiments, the methods disclosed herein include methods of treating diseases by administering such pharmaceutically acceptable salts as pharmaceutical compositions. In some embodiments, the compounds described herein possess acidic or basic groups and therefore react with numerous inorganic or organic bases, and both inorganic and organic acids, to form pharmaceutically acceptable salts. In some embodiments, these salts are prepared in situ during the final isolation and purification of the compounds disclosed herein, or by separately reacting the purified compound in its free form with the appropriate acid or base and isolating the salt thus formed.

[0076] Examples of pharmaceutically acceptable salts include salts prepared by reaction of the compounds described herein with a mineral, organic acid, or inorganic base, such as acetate, acrylate, adipate, alginate, aspartate, benzoate, benzenesulfonate, bisulfate, bisulfite, bromide, butyrate, butyne-1,4-dioate, camphorate, camphorsulfonate, caproate, caprylate, chlorobenzoate, chloride, citrate, cyclopentanepropionate, decanoate, digluconate, dihydrogenphosphate, dinitrobenzoate, dodecyl sulfate, ethanesulfonate, formate, fumarate, glucoheptanoate, glycerophosphate, glycolate, hemisulfate, heptanoate, hexanoate, hexyne-1,6-dioate, hydroxybenzoate, carboxylate, gamma-hydroxybutyrate, hydrochloride, hydrobromide, Examples of suitable salts include hydroiodide, 2-hydroxyethanesulfonate, iodide, isobutyrate, lactate, maleate, malonate, methanesulfonate, mandelate metaphosphate, methanesulfonate, methoxybenzoate, methylbenzoate, monohydrogenphosphate, 1-naphthalenesulfonate, 2-naphthalenesulfonate, nicotinate, nitrate, palmoate, pectinate, persulfate, 3-phenylpropionate, phosphate, picrate, pivalate, propionate, pyrosulfate, pyrophosphate, propiolate, phthalate, phenylacetate, phenylbutyrate, propanesulfonate, salicylate, succinate, sulfate, sulfite, succinate, suberate, sebacate, sulfonate, tartrate, thiocyanate, tosylate, tosylateundeconate, and xylenesulfonate.

[0077] Additionally, the compounds described herein can be prepared by dissolving the free base form of the compounds in various forms with inorganic acids such as hydrochloric acid, hydrobromic acid, sulfuric acid, nitric acid, phosphoric acid, metaphosphoric acid, as well as acetic acid, propionic acid, hexanoic acid, cyclopentanepropionic acid, glycolic acid, pyruvic acid, lactic acid, malonic acid, succinic acid, malic acid, maleic acid, fumaric acid, p-toluenesulfonic acid, tartaric acid, trifluoroacetic acid, citric acid, benzoic acid, 3-(4-hydroxybenzoyl)benzoic acid, cinnamic acid, mandelic acid, arylsulfonic acid, methanesulfonic acid, ethanesulfonic acid, 1,2-ethanedisulfonic acid, 2-hydroxyethanesulfonic acid, They can be prepared as pharmaceutically acceptable salts formed by reaction with pharmaceutically acceptable inorganic or organic acids, including, but not limited to, benzenesulfonic acid, 2-naphthalenesulfonic acid, 4-methylbicyclo-[2.2.2]oct-2-ene-1-carboxylic acid, glucoheptonic acid, 4,4'-methylenebis-(3-hydroxy-2-ene-1-carboxylic acid), 3-phenylpropionic acid, trimethylacetic acid, tert-butylacetic acid, lauryl sulfuric acid, gluconic acid, glutamic acid, hydroxynaphthoic acid, salicylic acid, stearic acid, and muconic acid.

[0078] In some embodiments, compounds described herein containing free acid groups are reacted with a suitable base, such as hydroxides, carbonates, bicarbonates, sulfates of pharmaceutically acceptable metal cations, ammonia, or pharmaceutically acceptable organic primary, secondary, tertiary, or quaternary amines. Representative salts include alkali or alkaline earth salts, such as lithium, sodium, potassium, calcium, and magnesium, as well as aluminum salts. Specific examples of bases include sodium hydroxide, potassium hydroxide, choline hydroxide, sodium carbonate, N + (C 1~4 alkyl)4.

[0079] Representative organic amines useful for the formation of base addition salts include ethylamine, diethylamine, ethylenediamine, ethanolamine, diethanolamine, piperazine, and the like. It is understood that the compounds described herein also include the quaternization of any basic nitrogen-containing groups contained in the compounds. In some embodiments, water- or oil-soluble or dispersible products are obtained by such quaternization.

[0080] solvate In some embodiments, the compounds described herein exist as solvates. The present disclosure provides methods for treating diseases by administering such solvates. The present disclosure further provides methods for treating diseases by administering such solvates as pharmaceutical compositions. Solvates contain stoichiometric or non-stoichiometric amounts of solvent and, in some embodiments, are formed during the process of crystallization with a pharmaceutically acceptable solvent, such as water or ethanol. Hydrates are formed when the solvent is water, or alcoholates are formed when the solvent is alcohol. Solvates of the compounds described herein can be conveniently prepared or formed during the processes described herein. By way of example only, hydrates of the compounds described herein can be conveniently prepared by recrystallization from an aqueous / organic solvent mixture using organic solvents, including, but not limited to, dioxane, tetrahydrofuran, or methanol. Furthermore, the compounds provided herein can exist in unsolvated and solvated forms. In general, solvated forms are considered equivalent to unsolvated forms in the compounds and methods provided herein.

[0081] tautomers In some situations, compounds exist as tautomers.The compounds described herein include all possible tautomers within the formulas described herein.Tautomers are compounds that can be interconverted by the migration of a hydrogen atom, accompanied by the exchange of a single bond with an adjacent double bond.In bond configurations where tautomerization is possible, there is a chemical equilibrium of tautomers.All tautomeric forms of the compounds disclosed herein are contemplated.The exact ratio of tautomers depends on several factors, including temperature, solvent and pH.

[0082] Treatment method Also disclosed herein is a method of treating a mammal in need of such treatment, comprising administering to the mammal an antibacterial effective amount of any of the above compounds at a frequency and for a duration sufficient to provide a beneficial effect to the mammal. In one embodiment, the mammal has a bacterial-related infection that is resistant to treatment with one or more clinically used antibiotics. In a further embodiment, the bacterial species causing the bacterial infection is selected from the group consisting of Pseudomonas aeruginosa, Pseudomonas fluorescens, Pseudomonas acidovorans, Pseudomonas alcaligenes, Pseudomonas putida, Stenotrophomonas maltophilia, Burkholderia cepacia, Aeromonas hydrophilia, Escherichia coli, Citrobacter freundii, Salmonella typhimurium, typhimurium, Salmonella typhi, Salmonella paratyphi, Salmonella enteritidis, Shigella dysenteriae, Shigella flexneri, Shigella sonnei, Enterobacter cloacae, Enterobacter aerogenes, Klebsiella pneumoniae, Klebsiella oxytoca, Serratia marcescensmarcescens, Francisella tularensis, Morganella morganii, Proteus mirabilis, Proteus vulgaris, Providencia alcalifaciens, Providencia rettgeri, Providencia stuartii, Acinetobacter baumannii, Acinetobacter calcoaceticus, Acinetobacter haemolyticus, Yersinia enterocolitica enterocolitica, Yersinia pestis, Yersinia pseudotuberculosis, Yersinia intermedia, Bordetella pertussis, Bordetella parapertussis, Bordetella bronchiseptica, Haemophilus influenzae, Haemophilus parainfluenzae, Haemophilus haemolyticus, Haemophilus parahaemolyticus, Haemophilus ducreyi, Pasteurella multocida multocida, Pasteurella haemolytica, Branhamella catarrhaliscatarrhalis, Helicobacter pylori, Campylobacter fetus, Campylobacter jejuni, Campylobacter coli, Borrelia burgdorferi, Vibrio cholerae, Vibrio parahaemolyticus, Legionella pneumophila, Listeria monocytogenes, Neisseria gonorrhoeae, Neisseria meningitidis meningitidis, Kingella, Moraxella, Gardnerella vaginalis, Bacteroides fragilis, Bacteroides distasonis, Bacteroides 3452A homology group, Bacteroides vulgatus, Bacteroides ovalus, Bacteroides thetaiotaomicron, Bacteroides uniformis, Bacteroides eggerthii or Bacteroides plankunnicus In this embodiment, the bacterial infection is an infection involving Gram-negative bacteria.

[0083] In one embodiment, by way of example only, compounds described herein exhibit antibiotic activity useful in the treatment of bacterial infections such as various strains of E. coli, E. cloaceae, K. pneumoniae, A. baumannii, or P. aeruginosa.

[0084] Combination therapy Also disclosed herein are methods of treating a mammal in need of such treatment, comprising administering to the mammal a second therapeutic agent according to any of the aforementioned methods of treatment. In another embodiment, the second therapeutic agent is not an LspA inhibitor. In another embodiment, the second therapeutic agent is an aminoglycoside antibiotic, a fluoroquinolone antibiotic, a β-lactam antibiotic, a macrolide antibiotic, a glycopeptide antibiotic, rifampicin, chloramphenicol, fluoramphenicol, colistin, mupirocin, bacitracin, daptomycin, or linezolid.

[0085] In some embodiments, a method for treating a bacterial infection in a patient, preferably a human, is provided, wherein the treatment comprises administering a therapeutically or pharmacologically effective amount of a combination of 1) a β-lactam antibiotic, 2) a compound disclosed herein, or a pharmaceutically acceptable salt, solvate, or stereoisomer thereof, and 3) a pharmaceutically acceptable carrier. In some embodiments, the β-lactam antibiotic is a carbapenem, cephalosporin, cephamycin, monobactam, or penicillin. Exemplary carbapenem antibiotics include, but are not limited to, ertapenem, imipenem, biapenem, and meropenem. Exemplary cephalosporin antibiotics include, but are not limited to, ceftobiprole, ceftaroline, cefpirome, cefozopran, cefepime, cefotaxime, and ceftriazon. Exemplary penicillin antibiotics include, but are not limited to, ampicillin, amoxacillin, piperacillin, oxacillin, cloxacillin, methicillin, and nafcillin. In some embodiments, a β-lactam is administered with a β-lactamase inhibitor. In some embodiments, a carbapenem is administered with a DHP inhibitor, such as cilastatin.

[0086] In some embodiments, the β-lactam antibiotic and the compound disclosed herein are administered sequentially or simultaneously. In some embodiments, the β-lactam antibiotic and the compound disclosed herein are administered together. In some embodiments, the β-lactam antibiotic and the compound disclosed herein are administered in the same formulation or in separate formulations. In some embodiments, either the β-lactam or the compound disclosed herein is administered first. After administration of the first compound, the other compound is administered, for example, within 1 to 60 minutes, e.g., within 1, 2, 3, 4, 5, 10, 15, 30, or 60 minutes. In one aspect, when a β-lactamase inhibitor is used, the β-lactamase inhibitor is administered separately or in a formulation containing a compound disclosed herein and / or a β-lactam antibiotic. In one aspect, when a DHP inhibitor is used to improve the stability of a carbapenem, the DHP inhibitor is administered separately or in a formulation containing a compound disclosed herein and / or a carbapenem.

[0087] Further described herein are pharmaceutical compositions comprising a compound disclosed herein, a pharmaceutically acceptable carrier, and optionally a β-lactam antibiotic. In embodiments where a combination is used, the β-lactam antibiotic and the compound disclosed herein are present in amounts such that the combination constitutes a therapeutically effective amount. Due to the potentiating effect of the compound disclosed herein, the amount of the β-lactam antibiotic present in the combination may be less than the amount of the β-lactam antibiotic used alone. In certain embodiments, the composition further comprises a β-lactamase antibiotic.

[0088] In further embodiments, where the β-lactam antibiotic is a carbapenem, a pharmaceutical composition is provided comprising a carbapenem antibiotic, a DHP inhibitor, a compound disclosed herein, and a pharmaceutically acceptable carrier. In some embodiments, where the β-lactam antibiotic is a carbapenem, the carbapenem antibiotic is preferably selected from the group consisting of ertapenem, imipenem, and meropenem.

[0089] In some embodiments, a compound disclosed herein is for use in the treatment of a bacterial infection. In some embodiments, a compound disclosed herein is for use in the treatment of a bacterial infection in combination with one or more additional therapeutic agents, including a β-lactam antibiotic. In some embodiments, a compound disclosed herein is for use as a medicament for treating a bacterial infection. In some embodiments, a compound disclosed herein is for use in the preparation of a medicament for treating a bacterial infection. In some embodiments, a compound disclosed herein is for use in the preparation of a medicament for treating a bacterial infection in combination with one or more additional therapeutic agents, including a β-lactam antibiotic.

[0090] In some embodiments described herein, the compounds disclosed herein can enhance the activity of β-lactam antibacterial agents by inducing susceptibility to the antibacterial agent in drug-resistant strains, such as MRSA. In some embodiments, the compounds disclosed herein can enhance the activity of β-lactam antibacterial agents by reducing the dosage of the antibacterial agent required for therapeutic effect in drug-susceptible strains. For example, if a compound disclosed herein lowers the minimum inhibitory concentration (MIC) of the antibacterial agent in susceptible strains (the MIC is the lowest concentration of the antibacterial agent that completely inhibits growth), such treatment can be advantageous to allow for a reduction in the amount of the antibacterial agent administered (which can reduce antibiotic side effects) or to reduce the frequency of administration. In some embodiments, the compounds disclosed herein can enhance the activity of antibacterial agents, such as carbapenems, to prevent the emergence of resistant subpopulations in heterogeneous bacterial populations containing resistant subpopulations.

[0091] In some embodiments, potentiators are used to enhance the activity of antibacterial agents whose clinical effectiveness is limited by the increasing prevalence of resistant strains. In some embodiments described herein, the compounds disclosed herein are used as potentiators, and the compounds disclosed herein can be administered together (concurrently or sequentially) with a β-lactam antibiotic to enable effective treatment of infections involving resistant bacteria or to reduce the amount of antibacterial agent required to treat the infection.

[0092] Administration and Pharmaceutical Compositions The pharmaceutical compositions described herein comprise a therapeutically effective amount of a compound described herein (i.e., any compound disclosed herein) formulated with one or more pharmaceutically acceptable carriers. As used herein, the term "pharmaceutically acceptable carrier" refers to any type of non-toxic, inert solid, semi-solid, or liquid filler, diluent, encapsulating material, or formulation auxiliary. Some examples of materials which can serve as pharmaceutically acceptable carriers include sugars such as lactose, glucose, and sucrose; starches such as corn starch and potato starch; cellulose and its derivatives such as sodium carboxymethylcellulose, ethyl cellulose, and cellulose acetate; powdered tragacanth; malt; gelatin; talc; excipients such as cocoa butter and suppository wax; oils such as peanut oil, cottonseed oil, safflower oil, sesame oil, olive oil, corn oil, and soybean oil; glycols such as propylene glycol; esters such as ethyl oleate and ethyl laurate; agar; buffers such as magnesium hydroxide and aluminum hydroxide; alginic acid; pyrogen-free water; isotonic saline; Ringer's solution; ethyl alcohol; and phosphate buffers. In addition, other non-toxic, compatible lubricants such as sodium lauryl sulfate and magnesium stearate, as well as coloring agents, releasing agents, coating agents, sweetening, flavoring, and perfuming agents, preservatives, and antioxidants can also be present in the composition, according to the discretion of the formulator. The pharmaceutical compositions described herein can be administered to humans and other animals orally, rectally, parenterally, intracisternally, intravaginally, intraperitoneally, topically (such as by powders, ointments or drops), bucally, or as an oral or nasal spray, or as a liquid aerosol or dry powder formulation for inhalation.

[0093] Liquid dosage forms for oral administration include pharmaceutically acceptable emulsions, microemulsions, solutions, suspensions, syrups and elixirs.In addition to the active compound, liquid dosage forms optionally contain inert diluents commonly used in the art, such as water or other solvents, solubilizers and emulsifiers, and mixtures thereof, such as ethyl alcohol, isopropyl alcohol, ethyl carbonate, ethyl acetate, benzyl alcohol, benzyl benzoate, propylene glycol, 1,3-butylene glycol, dimethylformamide, oils (especially cottonseed oil, peanut oil, corn oil, germ oil, olive oil, castor oil and sesame oil), glycerol, tetrahydrofurfuryl alcohol, polyethylene glycol and fatty acid esters of sorbitan.In addition to inert diluents, oral compositions can also contain adjuvants such as wetting agents, emulsifiers and suspending agents, sweeteners, flavorings and fragrances.

[0094] Injectable preparations, for example, sterile injectable aqueous or oily suspensions, are optionally formulated using suitable dispersants or wetting agents and suspending agents according to known techniques.Sterile injectable preparations are optionally sterile injectable solutions, suspensions, or emulsions in non-toxic parenterally acceptable diluents or solvents, such as solutions in 1,3-butanediol.Optionally, acceptable vehicles and solvents are water, Ringer's solution, USP, and isotonic sodium chloride solution.In addition, sterile, fixed oils are conventionally used as solvents or suspending media.For this purpose, any non-irritating fixed oil can be used, including synthetic mono- or diglycerides.In addition, fatty acids such as oleic acid are used in the preparation of injectables.

[0095] Injectable preparations can be sterilized, for example, by filtration through a bacteria-retaining filter, or by incorporating sterilizing agents in the form of sterile solid compositions which can be dissolved or dispersed in sterile water or other sterile injectable medium prior to use.

[0096] To prolong the effect of a drug, it is often desirable to slow the absorption of the drug from subcutaneous or intramuscular injection. This is optionally accomplished by using a liquid suspension of crystalline or amorphous material with poor water solubility. The rate of drug absorption then depends on the dissolution rate of the drug, which in turn may depend on crystal size and crystalline form. Alternatively, delayed absorption of a parenterally administered drug form is optionally achieved by dissolving or suspending the drug in an oil vehicle. Injectable depot forms are produced by forming microencapsulated matrices of the drug in biodegradable polymers such as polylactide-polyglycolide. Depending on the ratio of drug to polymer and the nature of the specific polymer used, the rate of drug release can be controlled. Other examples of biodegradable polymers include poly(orthoesters) and poly(anhydrides). Depot injectable formulations are optionally prepared by entrapping the drug in liposomes or microemulsions that are compatible with body tissues.

[0097] Compositions for rectal or vaginal administration are preferably suppositories, which can be prepared by mixing a compound described herein (i.e., a compound disclosed herein) with a suitable non-irritating excipient or carrier, such as cocoa butter, polyethylene glycol or a suppository wax, which is solid at ambient temperature but liquid at body temperature and therefore will melt in the rectum or vaginal cavity and release the active compound.

[0098] Solid dosage forms for oral administration include capsules, tablets, pills, powders, and granules. In such solid dosage forms, the active compound is mixed with at least one inert, pharmaceutically acceptable excipient or carrier, such as sodium citrate or dicalcium phosphate, and / or a) fillers or extenders, such as starch, lactose, sucrose, glucose, mannitol, and silicic acid; b) binders, such as, for example, carboxymethylcellulose, alginate, gelatin, polyvinylpyrrolidinone, sucrose, and gum arabic; c) humectants, such as glycerol; d) agar-agar, calcium carbonate, potato or tapioca starch, alginic acid, certain silicates, and sodium carbonate; e) solution retarders, such as paraffin; f) absorption accelerators, such as quaternary ammonium compounds; g) humectants, such as, for example, acetyl alcohol and glycerol monostearate; (h) absorbents, such as kaolin and bentonite clay; and (i) lubricants, such as talc, calcium stearate, magnesium stearate, solid polyethylene glycols, sodium lauryl sulfate, and mixtures thereof. In the case of capsules, tablets and pills, the dosage forms optionally comprise buffering agents.

[0099] Solid compositions of a similar type are optionally employed as fillers in soft and hard-filled gelatin capsules using such excipients as lactose or milk sugar, as well as high molecular weight polyethylene glycols and the like.

[0100] The solid dosage forms of tablets, sugar-coated tablets, capsules, pills and granules can be prepared with coatings and shells, such as enteric coatings and other coatings known in the pharmaceutical formulation art.Tablets, sugar-coated tablets, capsules, pills and granules can optionally contain opacifying agents and can also be of a composition that releases active ingredient(s) only, or preferentially in a certain part of the intestinal tract, optionally in a delayed manner.Examples of embedding compositions that can be used include polymeric substances and waxes.

[0101] Solid compositions of a similar type are optionally employed as fillers in soft and hard-filled gelatin capsules using such excipients as lactose or milk sugar, as well as high molecular weight polyethylene glycols and the like.

[0102] The active compound may also be in microencapsulated form with one or more excipients, as described above. Solid dosage forms such as tablets, dragees, capsules, pills, and granules can be prepared with coatings and shells, such as enteric coatings, release-controlling coatings, and other coatings known in the pharmaceutical formulation art. In these solid dosage forms, the active compound is optionally mixed with at least one inert diluent, such as sucrose, lactose, or starch. These dosage forms may also optionally contain additional substances other than inert diluents, such as tableting lubricants and other tableting aids, such as magnesium stearate and microcrystalline cellulose, as is customary. In the case of capsules, tablets, and pills, the dosage form optionally contains a buffering agent. The dosage form may optionally contain an opacifying agent and may also be of a composition that releases the active ingredient(s) only, or preferentially in a certain part of the intestinal tract, optionally in a delayed manner. Examples of embedding compositions that can be used include polymeric substances and waxes.

[0103] The dosage form of the compound described herein for topical or transdermal administration includes ointment, paste, cream, lotion, gel, powder, liquid, spray, inhalant or patch.Active ingredient is mixed with pharmaceutically acceptable carrier under sterile conditions and optionally any necessary preservative or buffer.Ophthalmic preparation, ear drops, etc. are also contemplated.

[0104] The ointments, pastes, creams and gels may contain, in addition to the active compounds described herein, excipients such as animal and vegetable fats, oils, waxes, paraffin, starch, tragacanth, cellulose derivatives, polyethylene glycols, silicones, bentonite, silicic acid, talc and zinc oxide, or mixtures thereof.

[0105] The compositions described herein are optionally formulated to be delivered as liquid aerosol or inhalable dry powder.Liquid aerosol formulations are optionally atomized into particle sizes that can be delivered to the terminal bronchiole and respiratory bronchioles where bacteria are present in patients with bronchial infections such as chronic bronchitis and pneumonia.Pathogenic bacteria generally exist throughout the airways, particularly in the terminal bronchiole and respiratory bronchioles, up to bronchi, bronchioles and lung parenchyma.During the exacerbation of infection, bacteria can also exist in the alveoli.Liquid aerosol and inhalable dry powder formulations are preferably delivered to the terminal bronchiole and ultimately to the parenchymal tissue throughout the endobronchial tree.

[0106] The aerosolized formulations described herein are optionally delivered using an aerosol-forming device, such as a jet, vibrating porous plate, or ultrasonic nebulizer, selected to enable the formation of aerosol particles, preferably primarily having a mass medium average diameter of 1 μm to 5 μm. Furthermore, the formulations preferably have a balanced osmolality, ionic strength, and chloride concentration, as well as the smallest aerosolizable volume capable of delivering an effective dose of the compounds described herein (i.e., the compounds disclosed herein) to the site of infection. Furthermore, the aerosolized formulations preferably do not negatively impair airway functionality or cause undesirable side effects.

[0107] Aerosolization devices suitable for administering the aerosol formulations described herein include, for example, jet, vibrating porous plate, ultrasonic nebulizers, and energized dry powder inhalers capable of atomizing the formulation into aerosol particles primarily in the 1-5 μm size range. In this application, "primarily" means that at least 70%, but preferably greater than 90%, of all generated aerosol particles are in the 1-5 μm range. Jet nebulizers operate by using air pressure to break up liquid solutions into aerosol droplets. Vibrating porous plate nebulizers operate by using a sonic vacuum created by a rapidly vibrating porous plate to force solvent droplets through a porous plate. Ultrasonic nebulizers operate by using piezoelectric crystals to shear liquids into small aerosol droplets. A variety of suitable devices are available, such as the AeroNeb™ and AeroDose™ vibrating porous plate nebulizers (AeroGen, Inc., Sunnyvale, California), the Sidestream® nebulizer (Medic-Aid Ltd., West Sussex, England), the Pari LC® and Pari LC Star® jet nebulizers (Pari Respiratory Equipment, Inc., Richmond, Virginia), and the Aerosonic™ (DeVilbiss Medizinische Produkte (Deutschland) GmbH, Heiden, Germany) and UltraAire® (Omron Healthcare, Inc., Vernon Hills, Illinois) ultrasonic nebulizers.

[0108] In some embodiments, the compounds described herein (i.e., compounds disclosed herein) are formulated for use as topical powders and sprays containing, in addition to the compounds described herein, excipients such as lactose, talc, silicic acid, aluminum hydroxide, calcium silicate, and polyamide powder, or mixtures of these substances. The sprays optionally contain conventional propellants, such as chlorofluorohydrocarbons.

[0109] Transdermal patch has another advantage that compound can be delivered to the body in a controlled manner.This dosage form can be prepared by dissolving or distributing compound in suitable medium.Absorption enhancers can also be used to increase the flow of compound through the skin.This rate can be controlled by providing a rate-controlling membrane or dispersing compound in a polymer matrix or gel.

[0110] According to the methods of treatment described herein, bacterial infections are treated or prevented in a patient, such as a human or lower mammal, by administering to the patient a therapeutically effective amount of a compound described herein in an amount and for a time period necessary to achieve the desired result. A "therapeutically effective amount" of a compound described herein refers to an amount of compound sufficient to treat a bacterial infection at a reasonable benefit / risk ratio applicable to any medical treatment. However, it will be understood that the total daily usage of the compounds and compositions described herein will be determined by the attending physician within the scope of sound medical judgment. The specific therapeutically effective dose level for any particular patient will depend on a variety of factors, including the disorder being treated and the severity of the disorder; the activity of the specific compound used; the specific composition used; the patient's age, weight, overall health, sex, and diet; the administration time, route of administration, and excretion rate of the specific compound used; the duration of treatment; drugs used in combination with or concurrently with the specific compound used, and similar factors known in the medical field.

[0111] The total daily dose of the compounds described herein (i.e., compounds disclosed herein) administered to a human or other mammal in single or divided doses can be, for example, an amount of 0.01 mg / kg to 50 mg / kg body weight, or more usually 0.1 mg / kg to 25 mg / kg body weight. Single dose compositions can contain such amounts or multiples thereof to make up the daily dose. Generally, the treatment regimens described herein comprise administering to a patient in need of such treatment from about 10 mg to about 2000 mg of the compounds described herein per day, in single or multiple doses. [Example]

[0112] The compounds disclosed herein are made by the methods illustrated in the reaction schemes shown below. Provided herein, in combination with the knowledge of those skilled in the art of synthetic organic chemistry, are the procedures used in some embodiments to prepare the full range of compounds disclosed and claimed herein.

[0113] The starting materials and reagents used in the preparation of these compounds are available from commercial suppliers such as Aldrich Chemical Co., (Milwaukee, Wis.), Bachem (Torrance, Calif.), or Sigma (St. Louis, Mo.), or are prepared by methods known to those skilled in the art according to procedures described in references such as Fieser and Fieser's Reagents for Organic Synthesis, Volumes 1-17 (John Wiley and Sons, 1991); Rodd's Chemistry of Carbon Compounds, Volumes 1-5 and Supplementals (Elsevier Science Publishers, 1989); Organic Reactions, Volumes 1-40 (John Wiley and Sons, 1991), March's Advanced Organic Chemistry, (John Wiley and Sons, 4th Edition), and Larock's Comprehensive Organic Transformations (VCH Publishers Inc., 1989). These schemes are merely illustrative of some methods for synthesizing the compounds disclosed in some embodiments of the present invention; various modifications to these schemes will be apparent to those skilled in the art upon review of this disclosure. The starting materials and intermediates of the reactions, as well as the final products, can be isolated and purified, if desired, using conventional techniques, including but not limited to filtration, distillation, crystallization, chromatography, and the like. Such materials can be characterized using conventional means, including physical constants and spectral data. Compounds are typically isolated as trifluoroacetate salts by reverse-phase HPLC using CHCN / HO with trifluoroacetic acid as an additive. In some instances, purification is performed without trifluoroacetic acid, and the compounds are isolated as the free base. Abbreviation AcOH acetic acid DCE 1,2-dichloroethane DCM dichloromethane DIPEA N,N-Diisopropylethylamine DMA N,N-dimethylacetamide DMAP 4-dimethylaminopyridine DMF N,N-dimethylformamide DMSO dimethyl sulfoxide ESI electrospray ionization EtOAc ethyl acetate h time HATU 1-[bis(dimethylamino)methylene]-1H-1,2,3-triazolo[4,5-b]pyridinium 3-oxide hexafluorophosphate HBTU O-(benzotriazol-1-yl)-N,N,N',N'-tetramethyluronium hexafluorophosphate HFIP 1,1,1,3,3,3-hexafluoro-2-propanol HOBT Hydroxybenzotriazole HPLC High Pressure Liquid Chromatography IPAC Isopropyl Acetate LCMS Liquid Chromatography Mass Spectrometry LDA Lithium diisopropylamide M molar concentration min N specified concentration NMR nuclear magnetic resonance PE Petroleum Ether PyAOP (3-hydroxy-3H-1,2,3-triazolo[4,5-b]pyridinato-O)tri-1-pyrrolidinyl-phosphorus hexafluorophosphate R T retention time SFC Supercritical Fluid Chromatography TEA Triethylamine THF tetrahydrofuran TFA trifluoroacetic acid TFE 2,2,2-trifluoroethanol TfOH trifluoromethanesulfonic acid

[0114] The compounds disclosed herein were prepared as illustrated in general Schemes 1-6. Scheme 1. JPEG0007670618000053.jpg45170

[0115] As depicted in Scheme 1, β-hydroxy acid 4 can be prepared starting from Oppolzer's sultam 1, which can be acylated with an acid chloride to generate intermediate 2. Intermediate 2 can be subjected to Lewis acid-mediated aldol alkylation to generate the anti-aldol product 3. Compound 3 can be hydrolyzed with aqueous lithium hydroxide and hydrogen peroxide to give the desired β-hydroxy acid compound 4. Scheme 2. JPEG0007670618000054.jpg45170

[0116] As shown in Scheme 2, β-hydroxy acid compounds 4 can be alkylated with appropriately substituted tert-butyl 2,2-dioxo-oxathiazolidine-3-carboxylate to give esters 5. These esters can be saponified to give carboxylic acids 6. The amino protecting group can be converted from Boc to Fmoc by acidic deprotection, followed by reprotection with 9-fluorenylmethyl chloroformate to give intermediates 7. Scheme 3. JPEG0007670618000055.jpg87170

[0117] As shown in Scheme 3, chloro-(2-chlorotrityl) resin can be loaded with an Fmoc-protected amino acid and the Fmoc group removed with piperidine in DMF to give intermediate 8. Intermediate 8 can be elaborated through steps 2-4 using standard solid-phase chemistry techniques utilizing HOBt coupling with an Fmoc-protected amino acid and standard side-chain protecting groups to give the resin-bound peptide 9. Scheme 4. JPEG0007670618000056.jpg83170

[0118] As shown in Scheme 4, resin-bound peptide 9 can be coupled to carboxylic acid 6 or 7 using standard coupling reagents such as HATU or PyAOP, followed by Boc or Fmoc deprotection and resin cleavage to give protected peptide 10. The linear peptide can be cyclized using macrolactamization conditions such as HATU / DIPEA to give macrocycle 11. The side chain of macrocyclic depsipeptide 11 can be globally deprotected using palladium / hydrogenation and / or acidic conditions to give compound 12. Scheme 5. JPEG0007670618000057.jpg54170

[0119] As shown in Scheme 5, a benzyl or methyl ester protected amino acid can be subjected to sequential reductive amination to give the protected intermediate 13. Boc deprotection followed by Fmoc protection affords intermediate 14. Deprotection of the carboxylic acid using either saponification or hydrogenation conditions affords the Fmoc protected amino acid intermediate 15. Alternatively, intermediate 13 can be directly converted to the carboxylic acid to give the Boc protected amino acid intermediate 16. Scheme 6. JPEG0007670618000058.jpg84170

[0120] As shown in Scheme 6, resin-bound peptide 9 can be coupled to intermediates 15 or 16 under standard peptide coupling conditions. Deprotection of the Fmoc group with piperidine followed by cleavage from the resin or acidic cleavage from the resin to simultaneously remove the Boc protecting group provides linear peptide 17. The linear peptide can be cyclized using macrolactamization conditions such as HATU / DIPEA to give macrocycle 18. The side chain of macrocyclic depsipeptide 18 can be globally deprotected using palladium / hydrogenation and / or acidic conditions to give compound 19.

[0121] General experimental conditions 1H NMR spectra were recorded at ambient temperature using either a Varian Unity Inova (400 MHz) spectrometer, an Avance III (300 MHz) spectrometer, or a Bruker Ultrashield (400 MHz or 500 MHz) spectrometer equipped with a triple-resonance 5 mm probe. Chemical shifts are expressed in ppm relative to tetramethylsilane. The following abbreviations are used: br = broad signal, s = singlet, d = doublet, dd = double doublet, t = triplet, q = quartet, and m = multiplet.

[0122] Microwave experiments were performed using a CEM Discover, Smith Synthesiser, or Biotage Initiator 60™, all of which use a single-mode resonator and dynamic field tuning, giving reproducibility and control. Temperatures between 40 and 250°C can be achieved, and pressures up to 30 bar can be reached.

[0123] High-pressure liquid chromatography-mass spectrometry (LCMS) experiments to determine retention times (RT) and associated mass ions were performed using one of the following methods: The spectrometer had an electrospray source operating in positive and negative ion modes. UV absorbance was collected at 220 nm and 254 nm, and a full mass spectrometry scan was applied to all experiments. Additional detection was achieved using a Sedex 85 evaporative light scattering detector. A Thermo Scientific charged aerosol detector (model: Corona) was used to measure the purity of UV-insensitive analytes. CAD parameters were set as follows: Gas pressure: 35~40 psi ·Flow rate ratio: 0.51 Ion trap: 20.3v Charger voltage: 2.91kv ·Charger current: 1.01uA

[0124] LCMS Method A: Experiments were performed on an Agilent 1290 HPLC coupled to an Agilent MSD mass spectrometer using ESI as the ionization source. LC separation was performed using a Phenomenex XB-C18, 1.7 mm, 50 x 2.1 mm column at a flow rate of 0.4 mL / min. Solvent A was water with 0.1% formic acid, and solvent B was acetonitrile with 0.1% formic acid. The gradient consisted of 2 to 98% solvent B over 7 min, with a 1.5 min equilibration followed by a 1.5 min hold at 97% B. The LC column temperature was 40 °C.

[0125] LCMS Method B: Experiments were performed on an Agilent 1290 HPLC coupled to an Agilent MSD mass spectrometer using ESI as the ionization source. LC separation was performed on an Agilent Zorbax Eclipse XDB-C18, 3.5 mm, 100 x 3.0 mm column at a flow rate of 0.7 ml / min. MPA (mobile phase A) was water containing 0.1% FA, and MPB (mobile phase B) was acetonitrile containing 0.1% FA. The gradient started at 2% MPB over 25.5 min, ended at 98% MPB, and maintained at 98% B for 2.5 min after a 1.5 min equilibration. The LC column temperature was 40 °C. UV absorbance was collected at 220 nm and 254 nm, and a full mass scan was performed for all experiments.

[0126] LCMS Method C: Experiments were performed on a Waters Micromass ZQ2000 quadrupole mass spectrometer coupled to a Waters Acquity UPLC system equipped with a PDA UV detector. The spectrometer has an electrospray source operating in positive and negative ion modes. The system uses an Acquity BEH C18 1.7µm 100 x 2.1mm column maintained at 40°C or an Acquity BEH Shield RP18 1.7µm 100 x 2.1mm column maintained at 40°C and a flow rate of 0.4mL / min. The initial solvent system was 95% water containing 0.1% formic acid (solvent A) and 5% acetonitrile containing 0.1% formic acid (solvent B) for the first 0.4 minutes, followed by a gradient up to 5% solvent A and 95% solvent B over the next 5.6 minutes. This was maintained for 0.8 minutes before returning to 95% solvent A and 5% solvent B over the next 0.2 minutes.

[0127] Experiments were performed on a Waters Acquity UPLC equipped with a Waters LCT Premier XE mass spectrometer using LCMS Method D: ESI ionization. LC separation was performed on an Acquity UPLC BEH C18, 1.7 mm, 2.1 x 50 mm column at a flow rate of 0.6 ml / min. MPA (mobile phase A) was water containing 0.05% TFA, and MPB (mobile phase B) was acetonitrile. The gradient started at 2% MPB and ended at 98% MPB over 18.5 min, and was held at 98% MPB for 1.0 min after a 0.5 min equilibration. The LC column temperature was 40°C. UV data were collected at 220 nm and 254 nm, and mass spectrometry full scans were applied to all experiments.

[0128] LCMS Method E: Experiments were performed on a Dionex Ultimate 3000 coupled to a Thermo Scientific Q Exactive HRMS using ESI as the ionization source. LC separation was performed on an Agilent Zorbax Eclipse XDB-C18, 3.5 m, 100 x 3.0 mm column at a flow rate of 0.7 ml / min. MPA (mobile phase A) was water containing 0.1% TFA, and MPB (mobile phase B) was acetonitrile containing 0.1% TFA. The gradient started at 2% MPB over 25.5 min, ended at 98% MPB, and held at 98% B for 2.5 min after a 1.5 min equilibration. The LC column temperature was 40 °C. UV absorbance was collected by a DAD detector, and mass spectrometry full scans were applied to all experiments.

[0129] LCMS Method F: Experiments were performed on a Shimadzu UFLC coupled with a DAD detector, an ELSD detector, and a 2020EV MS. The spectrometer has an electrospray source operating in positive ion mode. The system uses a Shim-Pack XR-ODS C18 2.2 μm 50*3.0 mm column maintained at 40 °C and a flow rate of 1.2 mL / min. The initial solvent system was 95% water containing 0.05% trifluoroacetic acid (solvent A) and 5% acetonitrile containing 0.05% trifluoroacetic acid (solvent B) for the first 0.01 min, followed by a gradient up to 5% solvent A and 95% solvent B over the next 2.0 min. This was maintained for 0.7 min before returning to 95% solvent A and 5% solvent B over the next 0.3 min.

[0130] LCMS Method G: Experiments were performed on a Shimadzu UFLC coupled with a DAD detector, an ELSD detector, and a 2020EV MS. The spectrometer has an electrospray source operating in positive ion mode. The system uses a Shim-Pack XR-ODS C18 2.2 μm 50*3.0 mm column maintained at 40 °C and a flow rate of 1.2 mL / min. The initial solvent system was 95% water containing 0.05% trifluoroacetic acid (solvent A) and 5% acetonitrile containing 0.05% trifluoroacetic acid (solvent B) for the first 0.01 min, followed by a gradient up to 100% solvent B over the next 2.0 min. This was maintained for 0.7 min before returning to 95% solvent A and 5% solvent B over the next 0.3 min.

[0131] LCMS Method H: Experiments were performed on a Shimadzu UFLC coupled with a DAD detector, an ELSD detector, and a 2020EV MS. The spectrometer has an electrospray source operating in positive ion mode. The system uses a Shim-Pack XR-ODS C18 2.2 μm 50*3.0 mm column maintained at 40 °C and a flow rate of 1.2 mL / min. The initial solvent system was 70% water containing 0.05% trifluoroacetic acid (solvent A) and 30% acetonitrile containing 0.05% trifluoroacetic acid (solvent B) for the first 0.01 min, followed by a gradient up to 30% solvent A and 70% solvent B over the next 3.6 min. This was then ramped to 100% solvent B over the next 0.4 min. This was maintained for 0.5 min before returning to 95% solvent A and 5% solvent B over the next 0.3 min.

[0132] LCMS Method I: Experiments were performed on a Shimadzu UFLC coupled with a DAD detector, an ELSD detector, and a 2020EV MS. The spectrometer has an electrospray source operating in positive ion mode. The system uses a Shim-Pack XR-ODS C18 2.2 μm 50*3.0 mm column maintained at 40 °C and a flow rate of 1.2 mL / min. The initial solvent system was 95% water containing 0.05% trifluoroacetic acid (solvent A) and 5% acetonitrile containing 0.05% trifluoroacetic acid (solvent B) for the first 0.01 min, followed by a gradient up to 30% solvent A and 70% solvent B over the next 3.2 min. This was then ramped to 100% solvent B over the next 0.5 min. This was maintained for 0.8 min before returning to 95% solvent A and 5% solvent B over the next 0.3 min.

[0133] LCMS Method J: Experiments were performed on a Shimadzu UFLC coupled with a DAD detector, an ELSD detector, and a 2020EV MS. The spectrometer has an electrospray source operating in positive ion mode. The system uses a Shim-Pack XR-ODS C18 2.2 μm 50*3.0 mm column maintained at 40 °C and a flow rate of 1.2 mL / min. The initial solvent system was 95% water containing 0.05% trifluoroacetic acid (solvent A) and 5% acetonitrile containing 0.05% trifluoroacetic acid (solvent B) for the first 0.01 min, followed by a gradient up to 40% solvent A and 60% solvent B over the next 3.2 min. This was then ramped to 100% solvent B over the next 0.5 min. This was maintained for 0.8 min before returning to 95% solvent A and 5% solvent B over the next 0.3 min.

[0134] LCMS Method K: Experiments were performed on a Shimadzu UFLC coupled with a DAD detector, an ELSD detector, and a 2020EV MS. The spectrometer has an electrospray source operating in positive ion mode. The system uses a Shim-Pack XR-ODS C18 2.2 μm 50*3.0 mm column maintained at 40 °C and a flow rate of 1.2 mL / min. The initial solvent system was 80% water containing 0.05% trifluoroacetic acid (solvent A) and 20% acetonitrile containing 0.05% trifluoroacetic acid (solvent B) for the first 0.01 min, followed by a gradient up to 40% solvent A and 60% solvent B over the next 3.6 min. The gradient was then increased to 100% solvent B over the next 0.4 min. This was maintained for 0.5 min before returning to 95% solvent A and 5% solvent B over the next 0.3 min.

[0135] LCMS Method L: Experiments were performed on a Shimadzu UFLC coupled with a DAD detector, an ELSD detector, and a 2020EV MS. The spectrometer has an electrospray source operating in positive ion mode. The system uses a Shim-Pack XR-ODS C18 2.2 μm 50*3.0 mm column maintained at 40 °C and a flow rate of 1.2 mL / min. The initial solvent system was 95% water containing 0.05% trifluoroacetic acid (solvent A) and 5% acetonitrile containing 0.05% trifluoroacetic acid (solvent B) for the first 0.01 min, followed by a gradient up to 30% solvent A and 70% solvent B over the next 3.5 min. This was then ramped to 100% solvent B over the next 0.2 min. This was maintained for 1.0 min before returning to 95% solvent A and 5% solvent B over the next 0.3 min. Intermediate 1. (2R,3R)-3-Hydroxy-2-methylnonanoic acid TIFF0007670618000059.tif26170Step 1. 1-((3aR,6S,7aS)-8,8-dimethyl-2,2-dioxidetetrahydro-3H-3a,6-methanobenzo[c]isothiazol-1(4H)-yl)propan-1-one TIFF0007670618000060.tif41170

[0136] To a solution of (3aR,6S,7aS)-8,8-dimethylhexahydro-3H-3a,6-methanobenzo[c]isothiazole 2,2-dioxide (175 g, 813 mmol), DMAP (10.5 g, 81.3 mmol), and triethylamine (172 mL, 1220 mmol) in THF (1.4 L) at 0 °C was added propionyl chloride (844 g, 894 mmol). After 30 min, the reaction was warmed to room temperature and stirred for an additional 3 h. The reaction was evaporated under reduced pressure. The resulting residue was diluted with water and isopropyl acetate. The aqueous layer was extracted with isopropyl acetate (twice). The combined organic layers were washed with 1 M NaOH (twice), 1 M HCl (twice), brine, dried over magnesium sulfate, and evaporated in vacuo. The crude solid was recrystallized from DCM and heptane to give the title compound (213.3 g, 97% yield). Step 2. (2R,3R)-1-((3aR,6S,7aS)-8,8-dimethyl-2,2-dioxidetetrahydro-3H-3a,6-methanobenzo[c]isothiazol-1(4H)-yl)-3-hydroxy-2-methylnonan-1-one TIFF0007670618000061.tif41170

[0137] To a solution of triethylborane (91 mL, 1 mol / L in hexane) in DCM (50 mL) in a 500 mL three-necked round-bottom flask was added TfOH (13.825 g, 92.122 mmol) dropwise at 0° C. under nitrogen. The mixture was then stirred at 0° C. for 30 minutes and then cooled to −15° C. To this was added a solution of 1-((3aR,6S,7aS)-8,8-dimethyl-2,2-dioxidetetrahydro-3H-3a,6-methanobenzo[c]isothiazol-1(4H)-yl)propan-1-one (10.00 g, 36.8 mmol) and DIPEA (12.3 g) in DCM (50 mL) dropwise at −15° C. The resulting solution was stirred at −10° C. in a water / salt bath (mixture A) for 1 hour. Titanium tetrachloride (110 mL, 1 mol / L in DCM, 3.0 equiv.) was placed in a 250 mL three-necked round-bottom flask under nitrogen. To this solution was added heptaldehyde (6.3 g, 55.3 mmol, 1.5 equiv.) dropwise at −78° C., and the mixture was stirred at −50° C. for 30 minutes. The resulting solution was then transferred to an addition funnel (Mixture B).

[0138] Mixture B was added dropwise to mixture A at -78°C and stirred at -50°C for 4 hours. The reaction was quenched with 300 mL of saturated aqueous ammonium chloride solution and extracted with 2 x 500 mL of ethyl acetate. The organic layers were combined, washed with 1 x 300 mL of brine, dried over anhydrous sodium sulfate, filtered, and evaporated in vacuo. The residue was applied to a silica gel column eluted with ethyl acetate / petroleum ether (1:4). The appropriate fractions were combined and concentrated in vacuo to give the title compound (12 g) as a pale yellow oil (containing approximately 15% starting material) in 85% purity, which was carried on to the next step without purification. LCMS (ESI): [M+H] + =386. Step 3. (2R,3R)-3-Hydroxy-2-methylnonanoic acid

[0139] In a 250 mL round-bottom flask were placed THF (60 mL), 30% HO (6.3 mL), LiOH (1.96 g, 81.844 mmol), water (30 mL, 1.665 mol), and (2R,3R)-1-((3aR,6S,7aS)-8,8-dimethyl-2,2-dioxidetetrahydro-3H-3a,6-methanobenzo[c]isothiazol-1(4H)-yl)-3-hydroxy-2-methylnonan-1-one (6.30 g, 16.340 mmol). The resulting solution was stirred at 25 °C overnight. The reaction was then quenched by adding 100 mL of saturated aqueous NaSO solution. The pH of the solution was adjusted to 3 with 1 N HCl solution. The resulting solution was extracted with 3 x 200 mL of ethyl acetate, and the organic layers were combined, dried over anhydrous sodium sulfate, filtered, and concentrated under vacuum. The residue was applied to a silica gel column eluted with ethyl acetate / petroleum ether (1:2). The appropriate fractions were combined and concentrated under vacuum. This afforded 2.2 g (70% yield) of the title compound as a pale yellow oil. LCMS (ESI): [MH] - =187; 1 H NMR(400MHz,DMSO-d6)δ11.95(s,1H), 4.60(d,J=5.2Hz,1), 3.59(s,1H), 2.4 3-2.31(m,1H), 1.48-1.15(m,10H), 0.98(d,J=7.0Hz,3H), 0.92-0.82(m,3H). Intermediate 2. (2R,3R)-3-((R)-2-((tert-butoxycarbonyl)amino)propoxy)-2-methylnonanoic acid TIFF0007670618000062.tif35170Step 1. (R)-2-((tert-butoxycarbonyl)amino)propyl(2S,3R)-3-((R)-2-((tert-butoxycarbonyl)amino)propoxy)-2-methylnonanoate TIFF0007670618000063.tif51170

[0140] To a solution of (2R,3R)-3-hydroxy-2-methyl-nonanoic acid (10.13 g, 53.81 mmol, Intermediate 1) in anhydrous THF (100 mL) was added tert-butyl (4R)-4-methyl-2,2-dioxo-oxathiazolidine-3-carboxylate (38.31 g, 161.46 mmol) and tetrabutylammonium fluoride (1 mol / L THF solution, 80 mL, 53.81 mmol) at room temperature. Sodium hydride (60% in mineral oil, 8.61 g, 215.2 mmol) was added in portions at 0 °C. After the addition of sodium hydride was complete and hydrogen evolution ceased, the reaction mixture was warmed to room temperature and stirred at room temperature for 16 h. Aqueous HCl (1 mol / L) was carefully added with stirring until the pH reached approximately 6–7. The resulting solution was extracted with ethyl acetate (3 × 100 mL), and the organic layers were combined. The combined organic phases were dried over anhydrous sodium sulfate, filtered, and concentrated under reduced pressure. The residue was purified by flash chromatography on silica gel eluting with ethyl acetate / petroleum ether (1 / 9) to give the title compound (23.01 g, 85% yield) as a colorless oil. LCMS (ESI): [M+H] + =503.4. Step 3. (2R,3R)-3-((R)-2-((tert-butoxycarbonyl)amino)propoxy)-2-methylnonanoic acid

[0141] To a solution of (R)-2-((tert-butoxycarbonyl)amino)propyl(2S,3R)-3-((R)-2-((tert-butoxycarbonyl)amino)propoxy)-2-methylnonanoate (5.02 g, 9.98 mmol) in methanol (50 mL) was added a solution of lithium hydroxide (2.3 g, 99.87 mmol) in water (23 mL) at 25° C. The resulting solution was stirred at 30° C. for 16 hours. The resulting solution was concentrated under reduced pressure to approximately one-third of its volume (methanol was evaporated). Aqueous HCl (1 mol / L) was carefully added until the pH reached approximately 6. The resulting solution was extracted with ethyl acetate (3×100 mL). The combined organic layers were dried over anhydrous sodium sulfate and concentrated under reduced pressure. The residue was purified by flash chromatography on silica gel eluting with ethyl acetate / petroleum ether (1 / 6) to give the title compound (3.03 g, 87% yield) as a colorless oil. LCMS (ESI): [M+H] + =346.2; 1 H NMR(300MHz,DMSO-d6)δ12.11(s,1H), 6.56(d,J=8.1Hz,1H), 3.53-3.15(m,4H), 2.64-2.60(m,1H), 1 .37(s,9H), 1.35-1.25(m,10H), 0.99(d,J=6.9Hz,3H), 0.95(d,J=7.2Hz,3H), 0.86(t,J=6.6Hz,3H). Intermediate 3. (2R,3R)-3-((R)-2-((((9H-Fluoren-9-yl)methoxy)carbonyl)amino)propoxy)-2-methylnonanoic acid TIFF0007670618000064.tif52170

[0142] To a solution of (2S,3R)-3-[(2R)-2-(tert-butoxycarbonylamino)propoxy]-2-methyl-nonanoic acid (1.15 g, 3.33 mmol, Intermediate 2) in dioxane (10 mL) was added a solution of HCl / dioxane (40 mL, 4 mol / L). The mixture was stirred at 25° C. for 1 hour and concentrated under reduced pressure. The residue was dissolved in water (10 mL) and THF (30 mL), and a solution of lithium hydroxide (320.0 mg, 13.33 mmol) in water (5 mL) was added at 0° C., followed by a solution of 9-fluorenylmethyl chloroformate (1.03 g, 3.99 mmol) in THF (5 mL) at 0° C. The reaction mixture was stirred at 25° C. for 4 hours and concentrated under reduced pressure. The residue was diluted with water (10 mL), and aqueous HCl (1 M) was carefully added until the pH reached approximately 6. The resulting solution was concentrated under reduced pressure. The residue was purified by flash chromatography on silica gel eluting with PE / EtOAc (2 / 1) to give the title compound (1.12 g, 72% yield) as a colorless oil. LCMS (ESI): [M+H] + =468.2; 1 H NMR(400MHz,DMSO-d6)δ12.16(s,1H), 7.88(d,J=7.6Hz,2H), 7.69(d,J=7.6H z,2H), 7.43-7.39(m,2H), 7.34-7.30(m,2H), 7.14(d,J=8.4Hz,1H), 4.28-4.1 8(m,3H), 3.64-3.47(m,2H), 3.36-3.21(m,2H), 2.67-2.60(m,1H), 1.34-1.18 (m,10H), 1.04(d,J=6.4Hz,3H), 0.96(d,J=7.2Hz,3H), 0.80(t,J=6.4Hz,3H). Intermediate 4. (2R,3R)-3-((R)-2-((((9H-Fluoren-9-yl)methoxy)carbonyl)amino)propoxy)-2-methyl-4-(spiro[3.3]heptan-2-yl)butanoic acid TIFF0007670618000065.tif37170Step 1. 2-(spiro[3.3]heptan-2-yl)ethan-1-ol TIFF0007670618000066.tif21170

[0143] To a solution of triethylphosphonoacetate (11.47 g, 51.18 mmol) in THF (150 mL) was added NaH (2.23 g, 55.84 mmol, 60% in mineral oil) at 0 °C. The reaction mixture was stirred at room temperature for 1 hour and cooled to 0 °C. Spiro[3.3]heptan-2-one (5.13 g, 46.53 mmol) was added in portions at 0 °C. The reaction mixture was stirred at room temperature for 2 hours and then quenched by the addition of HCl (100 mL, 1 mol / L in water). The mixture was extracted with ethyl acetate (100 mL × 3), and the combined organic phases were dried over sodium sulfate and evaporated under reduced pressure. The residue was purified by flash chromatography on silica gel eluting with petroleum ether / ethyl acetate (5 / 1) to give ethyl 2-spiro[3.3]heptan-2-ylideneacetate (8.02 g, 95% yield) as a yellow oil.

[0144] To a solution of ethyl 2-spiro[3.3]heptan-2-ylideneacetate (8.02 g, 44.47 mmol) in ethyl acetate (150 mL) was added palladium (0.8 g, 10% loading on carbon) at 0 °C. The mixture was evacuated, refilled with hydrogen, and stirred at room temperature for 5 h under a hydrogen balloon. The mixture was filtered, and the filtered solution was evaporated under reduced pressure. The resulting residue was dissolved in THF (150 mL), and then LiAlH (2.48 g, 65.19 mmol) was added at 0 °C. The reaction mixture was stirred at room temperature for 3 h and quenched by carefully adding water (50 mL). The solid was filtered off, and the filtered solution was extracted with DCM (100 mL × 3). The combined organic phase was dried over sodium sulfate and concentrated under reduced pressure. The residue was purified by flash chromatography on silica gel eluting with PE / EtOAc (2 / 1) to give the title compound (5.52 g, 90% yield) as a colorless oil. TLC R f =0.4, PE / EtOAc=2 / 1. Step 2. 2-(spiro[3.3]heptan-2-yl)acetaldehyde TIFF0007670618000067.tif23170

[0145] To a solution of 2-spiro[3.3]heptan-2-ylethanol (5.52 g, 39.37 mmol) in DCM (300 mL) was added pyridinium chlorochromate (12.63 g, 58.74 mmol). The reaction mixture was stirred at room temperature for 3 hours. The solids were removed by filtration. The filtered solution was evaporated under reduced pressure and the crude product was used directly in the next step. TLC R f =0.5, PE / EA=4 / 1. Step 3. (2R,3R)-3-Hydroxy-2-methyl-4-(spiro[3.3]heptan-2-yl)butanoic acid TIFF0007670618000068.tif21170

[0146] The title compound was prepared following a procedure similar to that described for Intermediate 1, using 2-(spiro[3.3]heptan-2-yl)acetaldehyde instead of heptaldehyde. LCMS (ESI): [MH] - =211.1. Step 4. (2R,3R)-3-((R)-2-((((9H-fluoren-9-yl)methoxy)carbonyl)amino)propoxy)-2-methyl-4-(spiro[3.3]heptan-2-yl)butanoic acid

[0147] The title compound was prepared using (2R,3R)-3-hydroxy-2-methyl-4-(spiro[3.3]heptan-2-yl)butanoic acid following a procedure similar to that described for Intermediate 3. LCMS (ESI): [M+H] + =492.3. Intermediate 5. (2R,3R)-3-((R)-2-((((9H-Fluoren-9-yl)methoxy)carbonyl)amino)propoxy)-4-((1R,3s,5S)-bicyclo[3.2.1]octan-3-yl)-2-methylbutanoic acid TIFF0007670618000069.tif42170Step 1. 2-((1R,3s,5S)bicyclo[3.2.1]octan-3-yl)ethan-1-ol TIFF0007670618000070.tif19170

[0148] To a suspension of NaH (1.93 g, 48.3 mmol, 60% in mineral oil) in 1,2-dimethoxyethane (30 ml) was added triethylphosphonoacetate (11.7 g, 52.3 mmol) at 0° C. The resulting mixture was stirred at 0° C. for 15 minutes. Then, a solution of bicyclo[3.2.1]octan-3-one (4.06 g, 32.7 mmol) in 1,2-dimethoxyethane (20 ml) was added. The resulting solution was stirred at 0° C. for 15 minutes, stirred at room temperature for 3 days, and poured into an ice-cold solution of HCl (1 mol / L in water, 100 ml). The resulting solution was extracted with ethyl acetate (100 ml × 4), and the organic layers were combined. The organic layers were dried over anhydrous sodium sulfate and concentrated under vacuum. The residue was purified by flash chromatography on silica gel eluting with EtOAc / PE (1 / 4) to give ethyl 2-(3-bicyclo[3.2.1]octanylidene)acetate (2.51 g). This material was dissolved in ethyl acetate (100 mL). One drop of TFA was added, and then the flask was evacuated and flushed with nitrogen three times. Palladium (1.3 g, 10% loading on carbon) was added. The system was evacuated and filled with hydrogen. The mixture was stirred overnight at room temperature under a hydrogen balloon. The solids were filtered off, and the filtered solution was concentrated in vacuo to give crude ethyl 2-(3-bicyclo[3.2.1]octanyl)acetate (6.53 g). This material was dissolved in THF (120 mL) and treated portionwise with lithium aluminum hydride (2.31 g, 60.7 mmol) under nitrogen at 0 °C. The reaction mixture was allowed to warm to room temperature and stirred at this temperature for 2 h. It was cooled on an ice bath and quenched by the careful addition of 2.3 mL of water, 2.3 mL of aqueous sodium hydroxide (25%), and 2.3 mL of water, successively. The precipitate was removed by filtration, and the filtered solution was evaporated under vacuum. The residue was purified by flash chromatography on silica gel eluting with ethyl acetate / petroleum ether (11-15% ethyl acetate) to give the title compound (2.66 g, 84% yield) as a colorless oil. 1H NMR(400MHz,DMSO-d6)δ4.24(t,J=5.2Hz,1H), 3.39-3.34(m,2H), 2.13-2.12(m, 2H), 1.75-1.56(m,3H), 1.48-1.32(m,5H), 1.29-1.24(m,3H), 1.01-0.94(m,2H). 1 1 H NMR suggested a ratio of endo to exo isomers of approximately 6:1. Step 2. 2-((1R,3s,5S)-bicyclo[3.2.1]octan-3-yl)acetaldehyde TIFF0007670618000071.tif18170

[0149] To a solution of 2-3-bicyclo[3.2.1]octanyl)ethanol (4.21 g, 27.29 mmol) in DCM (150 mL) was added pyridinium chlorochromate (8.86 g, 41.21 mmol). The reaction mixture was stirred at room temperature for 3 hours. The solid was removed by filtration and the filtered liquid was evaporated under reduced pressure. The crude product was used directly in the next step without further purification. TLC R f =0.5, PE / EA=4 / 1. Step 3. (2R,3R)-4-((1R,3s,5S)-bicyclo[3.2.1]octan-3-yl)-3-hydroxy-2-methylbutanoic acid TIFF0007670618000072.tif34170

[0150] Trifluoromethanesulfonic acid (3.66 g, 24.4 mmol) was added dropwise to triethylborane (24.9 mL, 24.9 mmol, 1 M in THF) in DCM (30 mL) at −10° C., and the reaction mixture was stirred at room temperature for 30 minutes. Then, a solution of 1-((3aR,6S,7aS)-8,8-dimethyl-2,2-dioxidetetrahydro-3H-3a,6-methanobenzo[c]isothiazol-1(4H)-yl)propan-1-one (5.95 g, 21.92 mmol) and DIPEA (3.63 g, 28.14 mmol) in DCM (30 mL) was added at 0° C. The reaction mixture was stirred at 0° C. for 30 minutes. Then, the reaction mixture was stirred at −78° C. Titanium tetrachloride (28.4 mL, 1 M in DCM) was added. Then, a solution of 2-(3-bicyclo[3.2.1]octanyl)acetaldehyde (3.96 g, 26.01 mmol) in DCM (40 mL) was added. The reaction mixture was stirred at −78° C. for 2 hours. The reaction mixture was poured into saturated aqueous ammonium chloride solution (100 mL) and extracted with DCM (150 ml×3). The combined organic phase was dried over NaSO and evaporated under reduced pressure. The residue was purified by flash chromatography on silica gel eluting with petroleum ether / ethyl acetate (5 / 1) to give (2R,3R)-4-((1R,5S)-bicyclo[3.2.1]octan-3-yl)-1-((3aR,6S,7aS)-8,8-dimethyl-2,2-dioxidetetrahydro-3H-3a,6-methanobenzo[c]isothiazol-1(4H)-yl)-3-hydroxy-2-methylbutan-1-one (7.45 g, 17.6 mmol, 80.2% yield) as a yellow oil.

[0151] A solution of LiOH (1.67 g, 69.52 mmol) in HO (40 mL) was added to a solution of (2R,3R)-4-((1R,5S)-bicyclo[3.2.1]octan-3-yl)-1-((3aR,6S,7aS)-8,8-dimethyl-2,2-dioxidetetrahydro-3H-3a,6-methanobenzo[c]isothiazol-1(4H)-yl)-3-hydroxy-2-methylbutan-1-one (7.37 g, 17.39 mmol) in acetonitrile (120 mL) at 0 °C. The reaction mixture was stirred at room temperature for 16 h, then acidified to pH 6 by addition of HCl (1 mol / L in water) and extracted with ethyl acetate (100 mL × 3). The combined organic phases were dried over NaSO and evaporated under reduced pressure. The residue was purified by flash chromatography on silica gel eluting with petroleum ether / ethyl acetate (5 / 1) to give the title compound (2.96 g, 75% yield, endo / exo=6 / 1) as a pale red solid. The mixture (2.96 g) was dissolved in petroleum ether / ethyl acetate (5 / 1, 100 ml) at 60° C. and cooled to 0° C. The resulting crystalline solid was collected by filtration to give the title compound as the pure endo isomer (1.05 g). LCMS (ESI): [MH] - =225.2. Step 4. [(2R)-2-(tert-butoxycarbonylamino)propyl](2R,3R)-4-[(1S,5R)-3-bicyclo[3.2.1]octanyl]-3-[(2R)-2-(tert-butoxycarbonylamino)propoxy]-2-methyl-butanoate TIFF0007670618000073.tif53170

[0152] To a solution of the hydroxy acid from step 3 (3.6 g, 15.9 mmol) in anhydrous THF (36 mL) was added tert-butyl (4R)-4-methyl-2,2-dioxo-oxathiazolidine-3-carboxylate (11.33 g, 47.75 mmol) and TBAF (24 mL, 1 M in THF), followed by portionwise addition of NaH (2.55 g, 63.68 mmol, 60% in mineral oil) under nitrogen at 0 °C. The reaction mixture was stirred overnight at room temperature and quenched with HCl (1 M in water) to approximately pH 4-5. Ethyl acetate (200 mL) was added and the phases were separated. The aqueous phase was extracted with ethyl acetate (2x). The organic layers were combined, dried over anhydrous sodium sulfate, and concentrated in vacuo. The residue was purified by flash chromatography on silica gel eluting with ethyl acetate and petroleum ether (1:9) to give the title compound (3.412 g, 39.6% yield) as a colorless oil. LCMS (ESI): [M+H] + =541.4. Step 5. (2R,3R)-4-[(1S,5R)-3-bicyclo[3.2.1]octanyl]-3-[(2R)-2-(tert-butoxycarbonylamino)propoxy]-2-methyl-butanoic acid TIFF0007670618000074.tif35170

[0153] A solution of LiOH (1.45 g, 63.1 mmol) in water (15 mL) was added to a solution of [(2R)-2-(tert-butoxycarbonylamino)propyl](2R,3R)-4-[(1S,5R)-3-bicyclo[3.2.1]octanyl]-3-[(2R)-2-(tert-butoxycarbonylamino)propoxy]-2-methyl-butanoate (3.41 g, 6.31 mmol) in MeOH (35 mL) at room temperature. The resulting solution was stirred overnight at 30 °C. MeOH was evaporated under reduced pressure, and the remaining aqueous solution was acidified to pH 4-5 with 1 N HCl solution. Ethyl acetate (200 mL) was added, and the phases were separated. The aqueous phase was extracted with ethyl acetate (2x). The organic layers were combined, dried over anhydrous sodium sulfate, and concentrated in vacuo. The residue was purified by flash chromatography on silica gel eluting with ethyl acetate and petroleum ether (1:6) to give the title compound (2.04 g, 84.3% yield) as a colorless oil. LCMS (ESI): [M+H] + =384.3. Step 6. (2R,3R)-3-((R)-2-((((9H-fluoren-9-yl)methoxy)carbonyl)amino)propoxy)-4-((1R,3s,5S)-bicyclo[3.2.1]octan-3-yl)-2-methylbutanoic acid

[0154] A solution of HCl / dioxane (32 mL, 4 M, 160 mmol) was added to a solution of (2R,3R)-4-[(1S,5R)-3-bicyclo[3.2.1]octanyl]-3-[(2R)-2-(tert-butoxycarbonylamino)propoxy]-2-methyl-butanoic acid (1.84 g, 4.81 mmol) in DCM (8 mL). The mixture was stirred at room temperature for 1 hour and concentrated in vacuo. The residue was then dissolved in water (10 mL) and THF (30 mL). LiOH (331.8 mg, 14.43 mmol) (dissolved in 6 mL of water) was added at 0° C. Fluorenylmethyloxycarbonyl chloride (1.49 g, 5.77 mmol) (dissolved in 20 mL of THF) was added dropwise with stirring at 0° C. The mixture was stirred at 25° C. for 1 hour. The THF was evaporated under reduced pressure, and the remaining aqueous solution was acidified to pH 5-6 with 1N HCl. Ethyl acetate (100 mL) was added, and the phases were separated. The aqueous phase was extracted with ethyl acetate (2x). The organic layers were combined, dried over anhydrous sodium sulfate, and concentrated in vacuo. The residue was purified by flash chromatography on silica gel eluting with DCM / MeOH (49 / 1) to give the title compound (2.24 g, 4.43 mmol, 92.2% yield) as a white solid. LCMS (ESI): [M+1] + =506.3; 1 H NMR (400MHz, DMDO-d6): δ12.17(s,1H), 7.88(d,J=7.2Hz,2H), 7.70(d,J=7.6Hz ,2H), 7.43-7.39(m,2H), 7.34-7.30(m,2H), 7.15(d,J=8.0Hz,1H), 4.29-4.18(m ,3H), 3.64-3.56(m,2H), 3.34-3.18(m,2H), 2.74-2.70(m,1H), 2.20-2.10(m,2 H), 1.73-1.62(m,1H), 1.56-1.41(m,3H), 1.39-1.22(m,6H), 1.08-0.92(m,9H). Intermediate 6. N-(((9H-Fluoren-9-yl)methoxy)carbonyl)-O-(3-((tert-butoxycarbonyl)amino)propyl)-L-serine TIFF0007670618000075.tif44170Step 1. Methyl N-((benzyloxy)carbonyl)-O-(3-nitropropyl)-L-serinate TIFF0007670618000076.tif26170

[0155] To a solution of 1-benzyl 2-methyl(S)-aziridine-1,2-dicarboxylate (11.32 g, 48.14 mmol) and 3-nitropropan-1-ol (5.06 g, 48.12 mmol) in chloroform (150 mL) at 0 °C under nitrogen was added BF3.Et2O (0.1 mL). The reaction mixture was allowed to warm to room temperature and stirred at room temperature for 16 hours. The solvent was removed under reduced pressure. The residue was purified by flash chromatography on silica gel eluting with petroleum ether / ethyl acetate (5 / 1) to give the title compound (11.02 g, 67% yield) as a colorless oil. LCMS (ESI): [M+H] + =341.1. Step 2. Methyl N-((benzyloxy)carbonyl)-O-(3-((tert-butoxycarbonyl)amino)propyl)-L-serinate TIFF0007670618000077.tif30170

[0156] To a solution of methyl N-((benzyloxy)carbonyl)-O-(3-nitropropyl)-L-serinate (11.02 g, 32.4 mmol) in ethanol (80 mL) and water (80 mL) was added NH4Cl (8.75 g, 162 mmol), followed by iron powder (9.07 g, 162 mmol). The reaction mixture was stirred at reflux for 4 h. The solids were filtered off and washed with ethanol. The filtered solution was concentrated under reduced pressure to remove approximately half of the volume. The residue was partitioned between water and ethyl acetate. The aqueous phase was extracted with ethyl acetate (2x). The combined organic phases were dried over sodium sulfate and concentrated under reduced pressure to give a residue (8.89 g), which was redissolved in DCM (100 mL) and then di-tert-butyl dicarbonate (9.36 g, 42.95 mmol) was added, followed by triethylamine (8.68 g, 85.89 mmol). The reaction mixture was stirred at ambient temperature for 2 hours. The solvent was evaporated under reduced pressure. The residue was purified by flash chromatography on silica gel eluting with ethyl acetate / petroleum ether (0-50%) to give the title compound (9.22 g, 78% yield) as a yellow oil. LCMS (ESI): [M+H] + =411.2. Step 3. Methyl O-(3-((tert-butoxycarbonyl)amino)propyl)-L-serinate TIFF0007670618000078.tif23170

[0157] To a solution of methyl N-((benzyloxy)carbonyl)-O-(3-((tert-butoxycarbonyl)amino)propyl)-L-serinate (9.22 g, 22.48 mmol) in ethyl acetate (100 mL) under nitrogen at 0° C. was added palladium (10 wt% on carbon, 0.93 g). The flask was evacuated and filled with hydrogen, then stirred under a hydrogen balloon at 25° C. for 3 h. The reaction mixture was filtered through Celite, and the filtrate was concentrated under reduced pressure to give the title compound. LCMS (ESI) [M+H] + =277.2. Step 4. N-(((9H-fluoren-9-yl)methoxy)carbonyl)-O-(3-((tert-butoxycarbonyl)amino)propyl)-L-serine

[0158] To a solution of methyl O-(3-((tert-butoxycarbonyl)amino)propyl)-L-serinate (5.90 g, 21.35 mmol) in THF (150 mL) and water (150 mL) was added lithium hydroxide (2.05 g, 85.41 mmol) at ambient temperature. The resulting mixture was stirred at 25° C. for 3 hours, cooled to 0° C., and then a solution of 9-fluorenylmethyl chloroformate (8.71 g, 33.74 mmol) in THF (10 mL) was added dropwise. The reaction mixture was stirred at 0° C. for 2 hours. The reaction mixture was acidified to about pH 6 by the addition of 1 M HCl and extracted with ethyl acetate (3×100 mL). The combined organic phases were dried over Na2SO4, filtered, and concentrated under reduced pressure. The residue was purified by flash chromatography on silica gel eluting with petroleum ether / ethyl acetate (5 / 1) to give the title compound (6.12 g, 56% yield) as a white solid. LCMS (ESI): [M+H] + =485.1. 1 H NMR(300MHz,DMSO-d6)δ12.78(s,1H), 7.90(d,J=7.5Hz,2H), 7.75(d,J=7.5Hz,2H), 7.64(d,J=8.1Hz,1H), 7.45-7.37(m,2H), 7.32-7.30( m,2H), 6.79(t,J=5.7Hz,1H), 4.30-4.09(m,4H), 3.67-3.61(m,2H), 3.46-3.39(m,2H), 3.00-2.94(m,2H), 1.63-1.56(m,2H), 1.37(s,9H). Intermediate 7. N-(((9H-Fluoren-9-yl)methoxy)carbonyl)-O-((1r,3S)-3-((tert-butoxycarbonyl)amino)cyclobutyl)-L-serine TIFF0007670618000079.tif47170Step 1. Methyl N-((benzyloxy)carbonyl)-O-((1r,3S)-3-((tert-butoxycarbonyl)amino)cyclobutyl)-L-serinate TIFF0007670618000080.tif42170

[0159] A solution of trans-tert-butyl N-(3-hydroxycyclobutyl)carbamate (7.95 g, 42.46 mmol) and BF3.Et2O (1.23 g, 8.54 mmol) in chloroform (100 mL) was stirred at 0 °C. Then, a solution of 1-benzyl 2-methyl(S)-aziridine-1,2-dicarboxylate (10.0 g, 42.51 mmol) in chloroform (10 mL) was added dropwise at 0 °C. The reaction mixture was then stirred at 25 °C for 20 h and concentrated under reduced pressure. The residue was purified by flash chromatography on silica gel eluting with petroleum ether / ethyl acetate (2 / 1) to give the title compound (2.5 g, 13% yield) as a white solid. LCMS (ESI): [M+H] + =423.2. Step 2. N-(((9H-fluoren-9-yl)methoxy)carbonyl)-O-((1r,3S)-3-((tert-butoxycarbonyl)amino)cyclobutyl)-L-serine

[0160] To a solution of methyl (2S)-2-(benzyloxycarbonylamino)-3-[3-(tert-butoxycarbonylamino)cyclobutoxy]propanoate (2.5 g, 5.92 mmol) in ethyl acetate (30 mL) was added palladium (10 wt% on carbon, 1.0 g) under nitrogen. The reaction mixture was evacuated, filled with hydrogen, and stirred under a hydrogen balloon at 25 °C for 1 hour. The catalyst was removed by filtration and rinsed with ethyl acetate. The filtered solution was concentrated under reduced pressure to give methyl (2S)-2-amino-3-[3-(tert-butoxycarbonylamino)cyclobutoxy]propanoate (1.7 g, crude) as a yellow oil. To 1.5 g of this material was added a solution of lithium hydroxide (250.0 mg, 10.42 mmol) in water (5 mL) and dioxane (15 mL). The resulting solution was stirred at 25°C for 1 hour. 9-Fluorenylmethyl chloroformate (1.61 g, 6.24 mmol) was then added. The mixture was stirred at 25°C for 1 hour, diluted with water (20 ml), and acidified to pH 6 with 1N HCl. The resulting solution was extracted with ethyl acetate (3 x 100 mL). The combined organic layers were dried over anhydrous sodium sulfate, filtered, and concentrated under reduced pressure. The residue was purified by flash chromatography on silica gel eluting with petroleum ether / ethyl acetate (4 / 1) to give the title compound (1.85 g, 71% yield) as a white solid. LCMS (ESI): [M+H] + =497.2; 1 H NMR(400MHz,DMSO-d6)δ12.76(s,1H), 7.89(d,J=7.6Hz,2H), 7.76(d,J=7.6Hz,2H), 7.64(d,J=8.4Hz,1H), 7.44-7.40(m,2H), 7. 35-7.31(m,2H), 7.16(d,J=7.2Hz,1H), 4.29-4.17(m,4H), 4.04-3.98(m,2H), 3.58-3.49(m,2H), 2.17-2.01(m,4H), 1.37(s,9H). JPEG0007670618000081.jpg247170 Intermediate 8. N-(((9H-Fluoren-9-yl)methoxy)carbonyl)-O-(2-(tert-butoxycarbonyl)-2-azaspiro[3.3]heptan-6-yl)-L-serine TIFF0007670618000082.tif54170Step 1. tert-Butyl (S)-6-(2-(((benzyloxy)carbonyl)amino)-3-methoxy-3-oxopropoxy)-2-azaspiro[3.3]heptane-2-carboxylate TIFF0007670618000083.tif54170

[0161] tert-Butyl 6-hydroxy-2-azaspiro[3.3]heptane-2-carboxylate (2.511 g, 11.18 mmol) and chloroform (30 mL) were added to a 200 mL flask, followed by boron trifluoride diethyl etherate (0.30 mL, 2.4 mmol), followed by dropwise addition of 1-benzyl 2-methyl(S)-aziridine-1,2-dicarboxylate (2.2 mL, 11 mmol). The resulting mixture was stirred at room temperature for 6 h, then quenched with methanol and evaporated in vacuo. The crude product was purified by flash chromatography on silica gel (200 g silica, solvent gradient: 20-80% isopropyl acetate in heptane) to afford 1.093 g (22%) of the title compound. LCMS (ESI): [M+H-Boc] + =349.15; 1 H NMR(400MHz,DMSO-d6)δ7.69(d,J=8.0Hz,1H), 7.40-7.27(m,5H), 5.04(s,2H), 4.25(m,1H), 3. 85-3.69(m,5H), 3.64(s,3H), 3.54-3.42(m,2H), 2.41-2.29(m,2H), 1.99(s,2H), 1.35(s,9H). Step 2. N-((benzyloxy)carbonyl)-O-(2-(tert-butoxycarbonyl)-2-azaspiro[3.3]heptan-6-yl)-L-serine TIFF0007670618000084.tif52170

[0162] To a solution of tert-butyl (S)-6-(2-(((benzyloxy)carbonyl)amino)-3-methoxy-3-oxopropoxy)-2-azaspiro[3.3]heptane-2-carboxylate (1.093 g, 2.436 mmol) in 1,4-dioxane (12 mL) was added lithium hydroxide (1.0 M in water, 4 mL, 4.0 mmol). The reaction mixture was stirred at room temperature for 90 minutes. The reaction mixture was poured into 10% aqueous citric acid and extracted with DCM (2 x 50 mL). The combined DCM extracts were dried over magnesium sulfate, filtered, and evaporated in vacuo to give 1.0618 g (quantitative) of the title compound. LCMS (ESI): [M+H] + =435.3. Step 3. N-(((9H-fluoren-9-yl)methoxy)carbonyl)-O-(2-(tert-butoxycarbonyl)-2-azaspiro[3.3]heptan-6-yl)-L-serine

[0163] To a solution of N-((benzyloxy)carbonyl)-O-(2-(tert-butoxycarbonyl)-2-azaspiro[3.3]heptan-6-yl)-L-serine in ethanol (10 mL) was added palladium (10 wt % on carbon) (250.5 mg, 0.2354 mmol). The reaction mixture was stirred under a hydrogen balloon at room temperature for 4 hours. The reaction mixture was filtered through Celite, rinsed with 2×100 mL of ethanol, and evaporated in vacuo to give a quantitative yield of O-(2-(tert-butoxycarbonyl)-2-azaspiro[3.3]heptan-6-yl)-L-serine. LCMS (ESI): [M+H-tBu] + =245.2.

[0164] The resulting material was dissolved in 1,4-dioxane (10 mL) and water (10 mL) and treated with sodium bicarbonate (626.8 mg, 7.44 mmol) and (9H-fluoren-9-yl)methyl (2,5-dioxopyrrolidin-1-yl)carbonate (1.192 g, 3.534 mmol). The reaction mixture was stirred at room temperature for 15 hours. The reaction mixture was poured into 10% aqueous citric acid and extracted with DCM (3 x 100 mL). The combined organic extracts were dried over magnesium sulfate, filtered, and evaporated in vacuo. The crude product was purified by flash chromatography on silica gel (80 g silica, solvent gradient: 0-100% isopropyl acetate in heptane) to afford 779.1 mg (61%) of the title compound as a white foam. LCMS (ESI): [M+H] + =523.3. Intermediate 9. (2S)-2-((((9H-Fluoren-9-yl)methoxy)carbonyl)amino)-4-(4-((benzyloxy)carbonyl)piperazin-1-yl)-3-((tert-butyldimethylsilyl)oxy)butanoic acid TIFF0007670618000085.tif59170Step 1. Benzyl (S)-2-((tert-butoxycarbonyl)amino)but-3-enoate TIFF0007670618000086.tif34170

[0165] A mixture of (S)-2-aminobut-3-enoic acid hydrochloride (1.0 g, 7.3 mmol), di-tert-butyl dicarbonate (1.6 g, 7.3 mmol), and sodium bicarbonate (1.2 g, 14.6 mmol) in THF:water (1:1, 6 mL) was heated at 60 °C for 2 h. The reaction mixture was cooled and concentrated. The residue was dissolved in DMF (3 mL), benzyl bromide (1.5 g, 8.7 mmol) was added, and the mixture was stirred at room temperature overnight. The reaction mixture was diluted with water and extracted with 20% heptane / IPAC. The organic layer was washed with brine, dried over sodium sulfate, and concentrated. The residue was purified by flash chromatography on silica gel eluting with heptane / IPAC (0-50%) to give the title compound (0.87 g, 55%). 1H NMR (400 MHz, chloroform-d) δ 7.42-7.31 (m, 5H), 5.91 (ddd, J = 16.3, 10.2, 5.3 Hz, 1H), 5.34 (ddd, J = 17.2, 1.9, 0.7 Hz, 1H), 5.25 (ddd, J = 10.3, 1.8, 0.6 Hz, 1H), 5.23-5.15 (m, 3H), 4.92 (s, 1H), 1.44 (s, 9H); LCMS (ESI): [M+H] + =292. Step 2. Benzyl (2S)-2-((tert-butoxycarbonyl)amino)-2-(oxiran-2-yl)acetate TIFF0007670618000087.tif37170

[0166] A mixture of (2S)-2-(tert-butoxycarbonylamino)but-3-enoate (1.35 g, 4.6 mmol) and m-CPBA (2.0 g, 70%) in DCM (20 mL) was stirred at room temperature overnight. The reaction mixture was concentrated and triturated with 10% DCM / heptane. The solid was removed by filtration and washed thoroughly with 10% DCM / heptane. The filtered solution was concentrated, and the residue was purified by flash chromatography (silica gel, 0-100% heptane / IPAC) to give the title compound (0.9 g, 63%). LCMS (ESI): [M+H] + = 308 and [M+Na] + =330.1; 1 H NMR(400MHz,DMSO-d6)δ7.52(d,J=7.9Hz,1H), 7.42-7.22(m,5H), 5.89(ddd,J=17.0,10.4,6.4Hz,1H), 5.33(d ,J=17.3Hz,1H), 5.22(d,J=10.4Hz,1H), 5.14(q,J=12.7Hz,2H), 4.66(t,J=7.2Hz,1H), 1.38(d,J=2.4Hz,9H). Step 3. Benzyl 4-((3S)-4-(benzyloxy)-3-((tert-butoxycarbonyl)amino)-2-hydroxy-4-oxobutyl)piperazine-1-carboxylate TIFF0007670618000088.tif32170

[0167] A mixture of benzyl (2S)-2-(tert-butoxycarbonylamino)-2-(oxiran-2-yl)acetate (0.8 g, 2.6 mmol) and benzyl piperazine-1-carboxylate (0.58 g, 2.6 mmol) in THF (5 mL) was heated at 70° C. for 4 h, then cooled and concentrated. The residue was purified by flash chromatography (silica gel; 0-10% methanol / DCM) to give the title compound (0.92 g, 67%). LCMS (ESI): [M+H]=528.4; 1 H NMR(400MHz,DMSO-d6)δ7.43-7.25(m,10H), 6.59(d,J=9.1Hz,1H), 5.23-5.12(m,2H), 5.06(s,2H), 4.3 0(dd,J=9.1,3.0Hz,1H), 4.05(q,J=6.3,5.3Hz,1H), 3.50-3.34(m,4H), 2.44-2.22(m,5H), 1.38(s,9H). Step 4. Benzyl 4-[(3S)-4-benzyloxy-3-(tert-butoxycarbonylamino)-2-[tert-butyl(dimethyl)silyl]oxy-4-oxo-butyl]piperazine-1-carboxylate TIFF0007670618000089.tif44170

[0168] To benzyl 4-((3 S)-4-(benzyloxy)-3-((tert-butoxycarbonyl)amino)-2-hydroxy-4-oxobutyl)piperazine-1-carboxylate (0.9 g, 1.7 mmol) in anhydrous THF (10 mL) was added 2,6-lutidine (0.6 mL, 5.0 mmol) followed by tert-butyldimethylsilyl trifluoromethanesulfonate (0.9 g, 3.4 mmol), and the reaction mixture was stirred for 1 h, then diluted with water and extracted with IPAC. The organic layer was washed with brine, dried over sodium sulfate, and concentrated. The residue was purified by flash chromatography (silica gel, 0-5% MeOH / DCM) to give the title compound (0.53 g, 50%). LCMS (ESI): [M+H]=642.5; 1 H NMR(400MHz,DMSO-d6)δ7.49-7.27(m,10H), 6.49(d,J=9.3Hz,1H), 5.17(d,J=6.9Hz,2H), 5.11(s,2H), 4.43(td,J=9.2,3. 2Hz,1H), 4.30-4.14(m,1H), 3.48-3.38(m,4H), 2.50-2.27(m,6H), 1.42(s,9H), 0.84(d,J=1.2Hz,9H), 0.13-0.05(m,6H). Step 5. (2S)-2-Amino-4-(4-((benzyloxy)carbonyl)piperazin-1-yl)-3-((tert-butyldimethylsilyl)oxy)butanoic acid TIFF0007670618000090.tif36170

[0169] Benzyl 4-[(3S)-4-benzyloxy-3-(tert-butoxycarbonylamino)-2-[tert-butyl(dimethyl)silyl]oxy-4-oxo-butyl]piperazine-1-carboxylate (0.5 g, 0.78 mmol) was dissolved in THF:methanol (4 mL; 3:1), LiOH (1N, 2 equivalents) was added, and the mixture was stirred for 1 hour. The reaction mixture was acidified with a minimum amount of aqueous citric acid and extracted with IPAC. The organic layer was dried over sodium sulfate and concentrated. The residue was dissolved in DCM (2 mL), HCl (4 mL, 4N in dioxane) was added, and the reaction mixture was stirred for 1 hour. The reaction mixture was concentrated and used directly in the next step. Step 6. (2S)-2-((((9H-Fluoren-9-yl)methoxy)carbonyl)amino)-4-(4-((benzyloxy)carbonyl)piperazin-1-yl)-3-((tert-butyldimethylsilyl)oxy)butanoic acid

[0170] (2S)-2-Amino-4-(4-((benzyloxy)carbonyl)piperazin-1-yl)-3-((tert-butyldimethylsilyl)oxy)butanoic acid (used directly from step 5) was dissolved in dioxane:water (5 mL, 1:1), and sodium bicarbonate (0.2 g, 2.3 mmol) and Fmoc-OSu (0.32 g, 0.93 mmol) were added. The mixture was stirred overnight, acidified with citric acid, and extracted with IPAC. The organic layer was washed with brine, dried over sodium sulfate, and concentrated. The residue was purified by flash chromatography (0-10% MeOH / DCM) to give 0.38 g of the title compound. LCMS (ESI): [M+H] + =674.5. Intermediate 10. Fmoc-Allothreonine(Bzl)-OH TIFF0007670618000091.tif44170Step 1. O-Benzyl-N-(tert-butoxycarbonyl)-L-allothreonine TIFF0007670618000092.tif51170

[0171] Into a 2000 mL three-necked round-bottom flask purged and maintained with an inert atmosphere of nitrogen, (2S,3S)-2-amino-3-hydroxybutanoic acid (85 g, 714 mmol), methanol (450 mL), water (450 mL), and sodium bicarbonate (150 g, 1.79 mol) were placed. This was followed by the portionwise addition of BocO (187 g, 856.82 mmol, 1.20 equiv.) over a period of 60 minutes at 0°C. The resulting solution was stirred at 25°C for 12 hours. The resulting mixture was concentrated under reduced pressure. The resulting solution was diluted with 500 mL of water. The resulting solution was extracted with 3 x 500 mL of ether, and the aqueous layers were combined. The pH of the aqueous solution was adjusted to 3 with hydrogen chloride (1 mol / L). The resulting solution was extracted with 3 x 500 mL of ethyl acetate, and the organic layers were combined, dried over anhydrous sodium sulfate, and concentrated under reduced pressure. This gave 110 g (67%) of (2S,3S)-2-[[(tert-butoxy)carbonyl]amino]-3-hydroxybutanoic acid.

[0172] In a 2000 mL three-necked round-bottom flask purged and maintained with an inert atmosphere of nitrogen, (2S,3S)-2-[[(tert-butoxy)carbonyl]amino]-3-hydroxybutanoic acid (110 g, 501.74 mmol) and DMF (1100 mL) were placed. Subsequently, sodium hydride (84.4 g, 3.52 mol) was added portionwise over 60 minutes at 0°C. The resulting solution was stirred at 0°C for 2 hours. To this, benzyl bromide (85.4 g, 499.33 mmol) was added dropwise with stirring over 30 minutes at 0°C. The resulting solution was stirred at 0°C for 5 hours. The reaction was then quenched by the addition of 1000 mL of water / ice. The resulting solution was extracted with 3 x 2000 mL of ethyl acetate, and the aqueous layers were combined. The pH of the solution was adjusted to 3 with aqueous hydrogen chloride (1 mol / L). The resulting solution was extracted with 3 x 2000 mL of ethyl acetate, and the organic layers were combined, dried over anhydrous sodium sulfate, and concentrated under vacuum. The crude product was recrystallized from petroleum ether:ethyl acetate in a 10:1 ratio to give 60 g (37%) of the title compound. Step 2. Fmoc-allothreonine(Bzl)-OH A 2000 mL three-necked round-bottom flask was charged with O-benzyl-N-(tert-butoxycarbonyl)-L-allothreonine (60 g, 193.95 mmol), dioxane (600 mL), and hydrogen chloride (4 N solution in dioxane, 600 mL). The resulting solution was stirred at 25° C. for 12 hours. The resulting mixture was concentrated in vacuo. This afforded 50 g of crude O-benzyl-L-allothreonine hydrochloride as a white solid.

[0173] In a 2000 mL three-necked round-bottom flask purged with nitrogen and maintained under an inert atmosphere, O-benzyl-L-allothreonine hydrochloride (50 g, 203.50 mmol), dioxane (500 mL), water (500 mL), sodium carbonate (43.3 g, 408.53 mmol), and FmocONSu (69 g, 1.00 equiv.) were added. The resulting solution was stirred at 25° C. for 12 hours. The pH of the solution was adjusted to 3 with hydrogen chloride (1 mol / L). The resulting solution was extracted with 3×500 mL of ethyl acetate, and the organic layers were combined and concentrated under vacuum. The residue was purified by silica gel chromatography using ethyl acetate / petroleum ether (1:1). This afforded 53.4 g (58% yield over two steps) of the title compound as a white solid. LCMS (ESI): [M+H] + =432; 1 H NMR(300MHz,CDCl3)δ12.85(1H,s), 7.88-7.90(2H,m), 7.74-7.77(2H,m), 7.64-7.67(1H,m), 7.41-7.44(2H,m ), 7.22-7.39(7H,m), 4.46-4.50(2H,m), 4.34-4.38(1H,m), 4.20-4.30(3H,m), 3.89-3.93(1H,m), 1.16(3H,s). Intermediate 11. N-(((9H-Fluoren-9-yl)methoxy)carbonyl)-O-(4-(((tert-butoxycarbonyl)amino)methyl)-2-methylphenyl)-L-serine TIFF0007670618000093.tif55170Step 1. tert-Butyl N-(4-bromo-3-methyl-phenyl)methyl-carbamate TIFF0007670618000094.tif24170

[0174] To a solution of (4-bromo-3-methyl-phenyl)methanamine (5.00 g, 25.0 mmol) and DIPEA (6.48 g, 50.14 mmol) in DCM (100 mL) was added dropwise a solution of BocO (8.22 g, 37.7 mmol) in 10 mL of DCM. The resulting solution was stirred at room temperature for 4 hours and then concentrated in vacuo. The residue was purified by flash chromatography on silica gel eluting with PE / EA (5% EA) to give the title compound (6.04 g, 80.5% yield) as a pale yellow solid. 1 H NMR (400MHz, DMSO-d6): δ7.36-7.33(m,3H), 7.11(d,J=8.0Hz,1H), 4.05(d,J=6.0Hz,2H), 2.25(s,3H), 1.39(s,9H). Step 2. tert-Butyl N-[[4-[(2S)-2-(benzyloxycarbonylamino)-3-[methoxy(methyl)amino]-3-oxo-propoxy]-3-methyl-phenyl]methyl]carbamate TIFF0007670618000095.tif29170

[0175] To a solution of tert-butyl N-[(4-bromo-3-methyl-phenyl)methyl]carbamate (1.00 g, 3.33 mmol) in toluene (15 mL), benzyl N-[(1S)-1-(hydroxymethyl)-2-[methoxy(methyl)amino]-2-oxo-ethyl]carbamate (943 mg, 3.34 mmol), [Pd(allyl)Cl] (61.0 mg, 0.167 mmol), t-BuBrettPhos (162 mg, 0.334 mmol), and CsCO (2.18 g, 6.65 mmol) were added under nitrogen. The reaction was stirred at 60 °C for 5 h under microwave irradiation. The reaction was repeated in parallel on the same scale. The two runs were combined and concentrated in vacuo. The residue was purified by flash chromatography on silica gel eluting with PE / EA (35% EA) to give the title compound (895 mg, 26.7% yield) as a pale yellow oil. LCMS (ESI): [M+Na] + =524.2. Step 3. tert-Butyl N-[[4-[(2S)-2-amino-3-[methoxy(methyl)amino]-3-oxo-propoxy]-3-methyl-phenyl]methyl]carbamate TIFF0007670618000096.tif28170

[0176] A mixture of tert-butyl N-[[4-[(2S)-2-(benzyloxycarbonylamino)-3-[methoxy(methyl)amino]-3-oxo-propoxy]-3-methyl-phenyl]methyl]carbamate (950 mg, 1.89 mmol) and 10% Pd / C (500 mg) in ethyl acetate (190 mL) was stirred under a hydrogen atmosphere at room temperature for 45 minutes. The catalyst was filtered off and the filtered solution was concentrated in vacuo to give the title compound (668 mg, 96% yield) as a yellow oil. LCMS (ESI): [M+H] + =368.2. Step 4. N-(((9H-fluoren-9-yl)methoxy)carbonyl)-O-(4-(((tert-butoxycarbonyl)amino)methyl)-2-methylphenyl)-L-serine

[0177] A solution of LiOH (173 mg, 7.21 mmol) in water (5.0 mL) was added to a solution of tert-butyl N-[[4-[(2S)-2-amino-3-[methoxy(methyl)amino]-3-oxo-propoxy]-3-methyl-phenyl]methyl]carbamate (662.0 mg, 1.8 mmol) in THF (21 mL) and HO (7.0 mL) at 0 °C. The reaction mixture was stirred at room temperature overnight and cooled to 0 °C. Fluorenylmethyloxycarbonyl chloride (930 mg, 3.6 mmol) (dissolved in 5 mL of THF) was then added dropwise at 0 °C. The reaction mixture was stirred at room temperature for 1 hour. Most of the THF was stripped off under vacuum. The remaining solution was diluted with water (3.0 mL) and adjusted to pH 6 with 1 M HCl. The resulting solution was concentrated in vacuo and the residue was purified by flash chromatography on silica gel eluting with DCM / MeOH (10 / 1) to give the title compound (648 mg, 65.8% yield) as a white solid. LCMS (ESI): [M+1] + =547.3. 1 H NMR (400MHz, DMSO-d6): δ7.92(d,J=7.6Hz,2H), 7.85(d,J=8.0Hz,2H), 7.85(d,J=7.6Hz,1H), 7.69-7.64(m,2H), 7.45-7.40(m,2H), 7.36(d,J= 7.2Hz,1H), 7.32(dd,J=7.2,3.2Hz,1H), 6.75-6.71(m,2H), 4.43-4.40( m,1H), 4.39-4.21(m,4H), 4.43-4.40(m,3H), 2.24(s,3H), 1.39(s,9H). Intermediate 12. (2S)-4-[3-(benzyloxycarbonylamino)propoxy]-2-(9H-fluoren-9-ylmethoxycarbonylamino)butanoic acid TIFF0007670618000097.tif45170Step 1. (2S)-4-Allyloxy-2-(tert-butoxycarbonylamino)butanoic acid TIFF0007670618000098.tif27170

[0178] To a suspension of sodium hydride (228 mg, 60% dispersion in mineral oil, 5.70 mmol) in DMF (20 mL) was added dropwise with stirring at 0°C a solution of Boc-L-homoserine (500 mg, 2.28 mmol) in DMF (5 mL). Upon complete addition of Boc-L-homoserine, the reaction mixture was stirred under nitrogen at 0-10°C for an additional 10 min. To this solution was added allyl bromide (304 mg, 2.51 mmol) in 5.0 mL of DMF. The reaction mixture was stirred at room temperature for 2 h and concentrated in vacuo. Water (10 mL) was added, and the mixture was acidified to approximately pH 5 with 1 M HCl and extracted with ethyl acetate (3 × 20 mL). The organic layers were combined, dried over anhydrous sodium sulfate, and concentrated in vacuo. The residue was purified by flash chromatography on silica gel eluting with 30% ethyl acetate in petroleum ether to give the title compound (350 mg, 59.2% yield) as a colorless oil. LCMS (ESI): [M+H-Boc] + =533.2; 1 H NMR (300MHz, DMSO-d6): δ12.47(s,1H), 7.06(d,J=8.1Hz,1H), 5.94-5.80(m,1H), 5.29-5.10(m,2H) , 4.07-3.90(m,3H), 3.40(dd,J=7.2,5.1Hz,2H), 1.97-1.88(m,1H), 1.80-1.69(m,1H), 1.41(s,9H). Step 2. Methyl (2S)-4-allyloxy-2-(tert-butoxycarbonylamino)butanoate TIFF0007670618000099.tif27170

[0179] (2S)-4-Allyloxy-2-(tert-butoxycarbonylamino)butanoic acid (3.50 g, 13.5 mmol) was treated with trimethylsilyldiazomethane (13.5 mL, 2.0 M in hexanes, 27.0 mmol) in DCM (60 mL) and MeOH (30 mL) at 0° C. The reaction mixture was stirred at room temperature for 30 minutes and then concentrated under reduced pressure. The residue was purified by flash chromatography on silica gel eluting with PE / EA 2 / 1 to give the title compound (3.65 g, 13.4 mmol, 99% yield) as a colorless oil. 1 H NMR (300MHz, DMSO-d6): δ7.23(d,J=7.8Hz,1H), 5.91-5.78(m,1H), 5.26-5.09(m,2H), 4.12-4.06(m,1H), 3.9 1-3.86(m,2H), 3.62(s,3H), 3.40(dd,J=6.9,5.1Hz,2H), 1.96-1.85(m,1H), 1.83-1.71(m,1H), 1.38(s,9H). Step 3. Methyl (2S)-2-(tert-butoxycarbonylamino)-4-(3-hydroxypropoxy)butanoate TIFF0007670618000100.tif32170

[0180] To a solution of methyl (2S)-4-allyloxy-2-(tert-butoxycarbonylamino)butanoate (4.03 g, 14.7 mmol) in THF (10 mL) was added dropwise a solution of 1 M BH3.THF in THF (12.5 mL, 12.5 mmol) under nitrogen at 0 °C. The reaction mixture was stirred at room temperature for 2 h. Water (0.5 mL) was added at 0 °C, followed by 3 N NaOH (2.5 mL, 7.5 mmol) and 30% HO (10 mL). The reaction mixture was stirred at room temperature for 2 h. 50 mL of water was added, and a small amount of Na2SO3 was added to consume excess HO. EtOAc (100 mL) was added, and the phases were separated. The aqueous phase was extracted with EtOAc (2x), and the organic phases were combined, dried over Na2SO4, filtered, and concentrated under reduced pressure. The residue was purified by flash chromatography on silica gel eluting with PE / EA (1 / 1) to give the title compound (1.77 g, 6.09 mmol, 41.3% yield) as a colorless oil. 1 H NMR (400MHz, DMSO-d6): δ7.21(d,J=7.6Hz,1H), 4.37(br,1H), 4.09-4.00(m,1H), 3.62(s,3H) ), 3.44-3.32(m,6H), 1.89-1.85(m,1H), 1.78-1.71(m,1H), 1.65-1.58(m,2H), 1.38(s,9H). Step 4. (S)-Methyl 4-(3-(((benzyloxy)carbonyl)amino)propoxy)-2-((tert-butoxycarbonyl)amino)butanoate TIFF0007670618000101.tif38170

[0181] Methanesulfonyl chloride (1.19 g, 10.4 mmol) was added to a solution of methyl (2S)-2-(tert-butoxycarbonylamino)-4-(3-hydroxypropoxy)butanoate (2.01 g, 6.91 mmol) and triethylamine (2.09 g, 20.7 mmol) in DCM (40 mL) at 0° C. The reaction mixture was stirred at room temperature for 2 h. Water (50 mL) was added and the phases were separated. The aqueous phase was extracted with DCM (2×), and the organic phases were combined, dried over Na2SO4, and concentrated in vacuo.

[0182] The resulting residue was dissolved in DMF (40 mL) and treated with sodium azide (2.34 g, 36.0 mmol) at 0° C. The reaction mixture was stirred at room temperature overnight. Water (30 mL) was added, and the resulting mixture was extracted with ethyl acetate (3 times), and the combined extracts were washed with brine, dried over Na2SO4, filtered, and concentrated in vacuo.

[0183] The resulting residue was dissolved in ethyl acetate (40 mL), and 10% Pd / C (500 mg) was added to the mixture. The mixture was stirred under a hydrogen atmosphere at room temperature for 2 hours. The catalyst was removed by filtration, and the filtered solution was concentrated under reduced pressure.

[0184] The resulting residue was combined with triethylamine (2.1 g, 20.8 mmol) in DCM (40 mL), and to this mixture was added Cbz-Cl (1.77 g, 10.4 mmol) at 0° C. The reaction was stirred at room temperature for 2 hours and then concentrated in vacuo. The residue was purified by flash chromatography on silica gel eluting with PE / EA (1 / 1) to give the title compound (1.02 g, 34.7% yield over four steps) as a colorless oil. LCMS (ESI): [M+H] + =425.4. 1 H NMR (300MHz, DMSO-d6): δ7.39-7.27(m,5H), 7.25-7.21(m,2H), 5.00(s,2H), 4.11-4.01(m,1H), 3.62(s,3H), 3.44-3.26(m,4H), 3.07-3.00(m,2H), 1.94-1.84(m,1H), 1.81-1.71(m,1H), 1.65-1.58(m,2H), 1.38(m,9H). Step 5. (2S)-4-[3-(benzyloxycarbonylamino)propoxy]-2-(9H-fluoren-9-ylmethoxycarbonylamino)butanoic acid

[0185] A solution of HCl / dioxane (20 mL, 80 mmol, 4 M) was added to a solution of methyl (2S)-4-[3-(benzyloxycarbonylamino)propoxy]-2-(tert-butoxycarbonylamino)butanoate (950 mg, 2.24 mmol) in DCM (20 mL) at 0° C. The reaction was stirred at room temperature for 2 hours and then concentrated under reduced pressure. THF (60 mL) was added, followed by a solution of LiOH (163 mg, 6.78 mmol) in water (20 mL) at 0° C. The resulting mixture was stirred at room temperature for 1 hour, followed by the addition of fluorenylmethyloxycarbonyl chloride (693 mg, 2.68 mmol) in 10 mL of THF at 0° C. The reaction mixture was stirred at room temperature for 1 hour and acidified to approximately pH 5 with 1 N HCl. Ethyl acetate (100 mL) was added, and the phases were separated. The aqueous phase was extracted with ethyl acetate (2×). The organic phases were combined, washed with brine, dried over NaSO, and concentrated under reduced pressure. The residue was purified by flash chromatography on silica gel eluting with [DCM] / [MeOH] (10 / 1) to give the title compound (660 mg, 55.4% yield) as a white solid. LCMS (ESI): [M+H] + =533.2; 1 H NMR (300MHz, DMSO-d6): δ12.59(s,1H), 7.89(d,J=7.5Hz,2H), 7.89(d,J=7.5Hz,2H), 7.62(d,J=8.4Hz,1H), 7.44-7.37(m,2H), 7.34-7.21(m,8H), 4.99(s,2H), 4.30-4.19(m,3H), 4.09-4.02(m,1H), 3.41-3.31(m,4H), 3. 07-3.01(m,2H), 1.96-1.88(m,1H), 1.81-1.73(m,1H), 1.66-1.57(m,2H). Intermediate 13. (S)-2-((((9H-Fluoren-9-yl)methoxy)carbonyl)amino)-2-cycloheptylacetic acid TIFF0007670618000102.tif41170

[0186] To a solution of (2S)-2-amino-2-cycloheptyl-acetic acid (1.0 g, 5.84 mmol) in DMF (25 mL) and water (25 mL) was added NaHCO3 (982.0 mg, 11.69 mmol). A solution of 9-fluorenylmethyl chloroformate (1.8 g, 6.98 mmol) in THF (5 mL) was added dropwise at 0 °C. The reaction mixture was warmed to room temperature and stirred at 25 °C for 24 h. Aqueous HCl (1 mol / L) was carefully added until the pH reached approximately 6. The resulting solution was extracted with ethyl acetate (3 × 100 mL). The organic layers were combined, washed with water (5 ×), dried over sodium sulfate, and concentrated under reduced pressure. The residue was purified by flash chromatography on silica gel eluting with DCM / MeOH (10 / 1) to give the title compound (1.77 g, 77% yield) as a white solid. LCMS (ESI): [M+H] + =394.2. 1 H NMR(400MHz,DMSO-d6)12.58(s,1H),7.89(d,J=7.6Hz,2H),7.76(d,J=7.6Hz,2H),7.59(d, J=8.8Hz,1H), 7.44-7.40(m,2H), 7.35-7.30(m,2H), 4.31-3.93(m,4H), 1.95-1.33(m,13H). Intermediate 14. (2R,3R)-3-((R)-2-((((9H-Fluoren-9-yl)methoxy)carbonyl)amino)propoxy)-2-(but-3-en-1-yl)non-8-enoic acid TIFF0007670618000103.tif41170

[0187] The title compound was prepared following a similar method as described for Intermediate 3. LCMS (ESI): [M+1]+ = 506.3. Intermediate 15. O-Benzyl-N-((S)-3-(((benzyloxy)carbonyl)amino)-2-((S)-2-cyclohexyl-2-((S)-4-methyl-2-(methylamino)pentanamido)acetamido)propanoyl)-L-allothreonine-(2-chlorotrityl resin) JPEG0007670618000104.jpg50170

[0188] 2-Chlorotrityl chloride resin (loading 0.956 mmol / g, 5 g) was swollen in a mixture of DMF:DCM (1:1, 20 mL) for 40 min. The resin was drained, and then a solution of (2S,3S)-3-benzyloxy-2-(9H-fluoren-9-ylmethoxycarbonylamino)butanoic acid (Intermediate 10) (2.461 g, 5.70 mmol), DMF (10 mL), and DIPEA (495.6 mg, 3.84 mmol) was added. The resin was agitated with nitrogen bubbling for 4 h, drained, and rinsed sequentially with 10 mL of DCM / MeOH / DIPEA (10 / 10 / 1 = V / V / V), 10 mL of DMF, 10 mL of DCM, and 10 mL of DMF.

[0189] The resin was treated with 25 mL of 20% piperidine in DMF and agitated for 1.5 hours with a nitrogen sparge. The resin was drained and rinsed sequentially with DMF (3 × 25 mL) and DCM (3 × 25 mL). The resin was treated with a solution of (2S)-3-(benzyloxycarbonylamino)-2-(9H-fluoren-9-ylmethoxycarbonylamino)propanoic acid (4.372 g, 9.494 mmol), a pre-prepared stock solution of HOBt / HATU (1:1) in DMF (0.4 mmol / mL, 25 mL), and DIPEA (2.4 g, 19 mmol). The resin was agitated for 2.5 hours with a nitrogen sparge, drained, and rinsed sequentially with 10 mL of DMF, 10 mL of DCM, and 10 mL of DMF. The resin was treated with 25 mL of 20% piperidine in DMF and agitated for 1.5 hours with a nitrogen sparge. The resin was drained and rinsed sequentially with DMF (3 × 25 mL) and DCM (3 × 25 mL). The resin was treated with a solution of (S)-2-((((9H-Fluoren-9-yl)methoxy)carbonyl)amino)-2-cyclohexylacetic acid (3.61 g, 9.52 mmol), a previously prepared stock solution of HOBt / HATU (1:1) in DMF (0.4 mmol / mL, 25 mL), and DIPEA (2.4 g, 19 mmol). The resin was agitated with a nitrogen sparge for 2.5 h, drained, and rinsed sequentially with 10 mL of DMF, 10 mL of DCM, and 10 mL of DMF. The resin was treated with 25 mL of 20% piperidine in DMF and agitated with a nitrogen sparge for 1.5 h. The resin was drained and rinsed sequentially with DMF (3 × 25 mL) and DCM (3 × 25 mL). The resin was treated with a solution of (2S)-2-{[(9H-fluoren-9-ylmethoxy)carbonyl](methyl)amino}-4-methylpentanoic acid (3.492 g, 9.50 mmol), a previously prepared stock solution of HOBt / HATU (1:1) in DMF (0.4 mmol / mL, 25 mL), and DIPEA (2.4 g, 19 mmol). The resin was agitated with a nitrogen sparge for 2.5 h, drained, and rinsed sequentially with 10 mL of DMF, 10 mL of DCM, and 10 mL of DMF. The resin was treated with 25 mL of 20% piperidine in DMF and agitated with a nitrogen sparge for 1.5 h.The resin was drained and rinsed with DMF (3 x 25 mL) and DCM (3 x 25 mL) to give the resin-bound tetrapeptide with a loading of 0.44 mmol / g. Intermediate T1 (Method A). (S)-2-((2-((tert-butoxycarbonyl)amino)ethyl)(butyl)amino)hexanoic acid TIFF0007670618000105.tif34170Step 1. Methyl (2S)-2-[2-(tert-butoxycarbonylamino)ethylamino]hexanoate TIFF0007670618000106.tif35170

[0190] A solution of methyl (2S)-2-aminohexanoate hydrochloride (8.02 g, 44.2 mmol) and tert-butyl N-(2-oxoethyl)carbamate (6.68 g, 42.0 mmol) and AcOH (2.65 g, 44.2 mmol) in ethyl acetate (300 mL) was stirred at room temperature for 2 hours. Sodium triacetoxyborohydride (14.1 g, 66.3 mmol) was then added. The resulting reaction mixture was stirred at room temperature overnight and washed with water (2x) and brine. The organic phase was dried over anhydrous sodium sulfate and concentrated under vacuum. The residue was purified by flash chromatography on silica gel eluting with petroleum ether and ethyl acetate (1 / 1) to give the title compound (7.82 g, 61.4% yield) as a yellow oil. LCMS (ESI): [M+H] + =289.3. Step 2. Methyl (2S)-2-[2-(tert-butoxycarbonylamino)ethyl-butyl-amino]hexanoate TIFF0007670618000107.tif36170

[0191] To a solution of methyl (2S)-2-[2-(tert-butoxycarbonylamino)ethylamino]hexanoate (7.81 g, 27.1 mmol) in ethyl acetate (300 mL) was added butyraldehyde (2.93 g, 40.6 mmol) and AcOH (1.63 g, 27.1 mmol). The reaction mixture was stirred at room temperature for 2 hours. Sodium triacetoxyborohydride (8.62 g, 40.65 mmol) was then added. The reaction mixture was stirred at room temperature overnight and washed successively with water and brine. The organic phase was dried and concentrated under vacuum. The residue was purified by flash chromatography on silica gel eluting with petroleum ether and ethyl acetate (5 / 1) to give the title compound (2.32 g, 24.9% yield) as a yellow oil. LCMS (ESI): [M+H] + =345.3. Step 3. (2S)-2-[2-(tert-butoxycarbonylamino)ethyl-butyl-amino]hexanoic acid TIFF0007670618000108.tif33170

[0192] To a solution of methyl (2S)-2-[2-(tert-butoxycarbonylamino)ethyl-butyl-amino]hexanoate (4.04 g, 11.7 mmol) in MeOH (100 mL) and water (30 mL) was added lithium hydroxide (1.13 g, 47.0 mmol) at room temperature. The reaction mixture was stirred overnight at room temperature, acidified to pH 4 with 1N HCl, and concentrated under reduced pressure. The residue was purified by flash chromatography on silica gel eluting with DCM and MeOH (5 / 1) to afford the title acid (2.56 g, 66.1% yield) as a yellow solid. LCMS (ESI): [M+H] + =331.3; 1 H NMR (300MHz, DMSO-d6): δ12.11(s,1H), 6.74(t,J=5.9Hz,1H), 3.40-3.38(m,1H), 3.03(t,J=6.3Hz,2 H), 2.82-2.80(m,1H), 2.69-2.66(m,3H), 1.65-1.60(m,2H), 1.46-1.30(m,17H), 0.90-0.85(m,6H). Intermediate T2 (Method B). 2-((2-((tert-butoxycarbonyl)amino)ethyl)(methyl)amino)octanoic acid TIFF0007670618000109.tif34170

[0193] To a solution of tert-butyl (2-(methylamino)ethyl)carbamate (1.75 g, 10.0 mmol) and DIPEA (3.32 mL, 19.1 mmol) in CHCN (30 mL) was added 2-bromooctanoic acid (2.06 g, 9.22 mmol) at room temperature. The reaction mixture was stirred at 60° C. overnight and then concentrated in vacuo. The residue was purified by flash chromatography on silica gel eluting with MeOH / DCM (1 / 9) to give the title compound (2.065 g, 70.8% yield) as a colorless oil. LCMS (ESI): [M+H] + =317.3; 1 H NMR (300 MHz, DMSO- d 6):δ10.60(s,1H), 6.59(t,J=5.7Hz,1H), 3.08-2.92(m,3H), 2.64-2.57(m,1H), 2.44-2.37(m ,1H), 2.24(s,3H), 1.60-1.56(m,2H), 1.47(s,9H), 1.45-1.22(m,8H), 0.84(t,J=6.3Hz,3H). Intermediate T12 (Method C). (S)-2-((2-((((9H-Fluoren-9-yl)methoxy)carbonyl)amino)ethyl)(butyl)amino)hexanoic acid TIFF0007670618000110.tif37170

[0194] To a solution of (2S)-2-[2-(tert-butoxycarbonylamino)ethyl-butyl-amino]hexanoic acid (2.50 g, 7.57 mmol) in dioxane (10 mL) was added HCl / dioxane (4.0 M, 40 mL) at room temperature. The reaction mixture was stirred at room temperature for 1 hour and then concentrated in vacuo. The resulting residue was dissolved in water (10 mL) and THF (30 mL). LiOH (545 mg, 22.71 mmol) (dissolved in water) was then added at 0°C, followed by fluorenylmethyloxycarbonyl chloride (2.3 g, 8.91 mmol) (dissolved in THF) at this temperature. The reaction mixture was stirred at 25°C for 1 hour. THF was evaporated in vacuo. The remaining aqueous solution was acidified to pH 4-5 with 1 M aqueous HCl and concentrated in vacuo. The residue was purified by flash chromatography on silica gel eluting with DCM / MeOH (10 / 1) to give the title compound (3.15 g, 92% yield) as a white solid. LCMS (ESI): [M+H] + =453.3; 1 H NMR (400MHz, DMSO-d6): δ12.09(s,1H), 7.88(d,J=7.6Hz,2H), 7.68(d,J=7.6Hz ,2H), 7.43-7.41(m,2H), 7.34-7.30(m,2H), 7.19(t,J=5.8Hz,1H), 4.34-4.18(m ,3H), 3.07-3.02(m,1H), 3.00-2.95(m,2H), 2.79-2.72(m,1H), 2.60-2.55(m,3 H), 1.65-1.60(m,1H), 1.59-1.53(m,1H), 1.45-1.15(m,8H), 0.88-0.82(m,6H). Intermediate T13 (Method D). (S)-2-(((R)-2-((((9H-Fluoren-9-yl)methoxy)carbonyl)amino)propyl)(butyl)amino)hexanoic acid TIFF0007670618000111.tif43170Step 1. Methyl (2S)-2-[[(2R)-2-(tert-butoxycarbonylamino)propyl]amino]hexanoate TIFF0007670618000112.tif37170

[0195] A solution of tert-butyl (4R)-4-methyl-2,2-dioxo-oxathiazolidine-3-carboxylate (16.5 g, 69.3 mmol) and methyl (2S)-2-aminohexanoate (40.3 g, 277 mmol) in CHCN (500 mL) was stirred at 60° C. for 5 hours. The reaction mixture was cooled to room temperature, concentrated under reduced pressure, and the residue was purified by flash chromatography on silica gel eluting with petroleum ether and ethyl acetate (2 / 1) to give the title compound (17.6 g, 83.7% yield) as a yellow oil. LCMS (ESI): [M+H] + =303.3; 1 H NMR (300 MHz, DMSO-d6): δ 6.50 (d, J = 8.4 Hz, 1H), 3.60 (s, 3H), 3.47-3.42 (m, 1H), 3.13-3.10 (m, 1H), 2.45-2.42 (m, 1H), 2.26-2.20 (m, 1H), 1.50-1.44 (m, 2H), 1.35 (s, 9H), 1.25-1.13 (m, 4H), 0.96 (d, J = 6.6 Hz, 3H), 0.87-0.80 (m, 3H). Step 2. Methyl (2S)-2-[[(2R)-2-(tert-butoxycarbonylamino)propyl]-butyl-amino]hexanoate TIFF0007670618000113.tif34170

[0196] A solution of methyl (2S)-2-[[(2R)-2-(tert-butoxycarbonylamino)propyl]amino]hexanoate (17.6 g, 58.0 mmol) and butyraldehyde (8.36 g, 116 mmol) in DCM (200 mL) was stirred at room temperature for 2 hours. Sodium triacetoxyborohydride (25.0 g, 118 mmol) was then added. The reaction mixture was stirred at room temperature overnight. Water (100 mL) was added and the phases were separated. The organic phase was dried over sodium sulfate and concentrated in vacuo. The residue was purified by flash chromatography on silica gel eluting with petroleum ether and ethyl acetate (5 / 1) to give the title compound (20.1 g, 96.7% yield) as a yellow oil. LCMS (ESI): [M+H] + =359.3. Step 3. (2S)-2-[[(2R)-2-(tert-butoxycarbonylamino)propyl]-butyl-amino]hexanoic acid TIFF0007670618000114.tif29170

[0197] A solution of LiOH (5.44 g, 227 mmol) in water (100 mL) was added to a solution of methyl (2S)-2-[[(2R)-2-(tert-butoxycarbonylamino)propyl]-butyl-amino]hexanoate (20.3 g, 56.7 mmol) in MeOH (300 mL). The reaction mixture was stirred overnight at room temperature, acidified to pH ∼5 with 1N HCl, and concentrated in vacuo. The residue was purified by flash chromatography on silica gel eluting with DCM and methanol (5 / 1) to give the title compound (15.5 g, 79.2% yield) as a yellow solid. LCMS (ESI): [M+H] + =345.3. Step 4. (S)-2-(((R)-2-((((9H-Fluoren-9-yl)methoxy)carbonyl)amino)propyl)(butyl)amino)hexanoic acid

[0198] A solution of 4N HCl / dioxane (150 mL, 600 mmol) was added to a solution of (2S)-2-[[(2R)-2-(tert-butoxycarbonylamino)propyl]-butyl-amino]hexanoic acid (15.5 g, 44.9 mmol) in DCM (150 mL) at 0° C. The reaction mixture was stirred at room temperature for 2 hours and concentrated in vacuo. The residue was dissolved in THF (150 mL), and a solution of LiOH (3.02 g, 125.71 mmol) in water (50 mL) was added at 0° C. Fluorenylmethyloxycarbonyl chloride (13.0 g, 50.3 mmol) dissolved in 20 mL of THF was added at 0° C. The reaction mixture was stirred at room temperature for 1 hour and then acidified to pH 5 with 1N HCl. Most of the THF was evaporated in vacuo. The remaining solution was extracted with ethyl acetate (2x50mL), and the combined organic phases were dried over anhydrous sodium sulfate and concentrated in vacuo. The residue was purified by flash chromatography on silica gel eluting with DCM and methanol (5 / 1) to give the title compound (16.5g, 84.4% yield) as a yellow solid. LCMS (ESI): [M+H] + =467.3; 1 H NMR (300MHz, DMSO-d6): δ12.08(s,1H), 7.88(d,J=7.5Hz,2H), 7.67(d,J=7.5Hz,2H), 7.43-7.40(m,2H), 7.33-7.28(m,2H), 7.16(br,1H), 4.30-4.1 7(m,3H), 3.76-3.71(m,2H), 2.88-2.65(m,4H), 1.68-1.64(m,2H), 1.58- 1.50(m,2H), 1.28-1.21(m,6H), 1.05(d,J=6.3Hz,3H), 0.86-0.80(m,6H). Intermediate T14 (Method E). N-((R)-2-((((9H-Fluoren-9-yl)methoxy)carbonyl)amino)propyl)-N-(hex-3-yn-1-yl)-L-alanine TIFF0007670618000115.tif41170Step 1. Benzyl (2S)-2-[[(2R)-2-(tert-butoxycarbonylamino)propyl]amino]propanoate TIFF0007670618000116.tif33170

[0199] A mixture of benzyl (2S)-2-aminopropanoate (11.4 g, 63.7 mmol) and tert-butyl (4R)-4-methyl-2,2-dioxo-oxathiazolidine-3-carboxylate (5.04 g, 21.2 mmol) and benzyl (2S)-2-aminopropanoate (11.4 g, 63.7 mmol) in CHCN (150 mL) was stirred at 60° C. overnight. The reaction mixture was concentrated in vacuo, and the residue was purified by flash chromatography on silica gel eluting with petroleum ether and ethyl acetate to give the title compound (5.66 g, 79.2% yield) as a yellow oil. LCMS (ESI): [M+H] + =337.2. Step 2. Benzyl (2S)-2-[[(2R)-2-(tert-butoxycarbonylamino)propyl]-hex(hex)-3-ynyl-amino]propanoate TIFF0007670618000117.tif30170

[0200] To a solution of TfO (34.5 g, 122 mmol) in DCM (100 mL) was added a mixture of 3-hexyn-1-ol (10.0 g, 102.1 mmol) and pyridine (12.1 g, 154 mmol) in DCM (60 mL) at 0 °C. The reaction was stirred at this temperature for 30 min. Water (100 mL) was added and the phases were separated. The organic phase (containing hex-3-ynyl trifluoromethanesulfonate) was dried over NaSO and used directly in the next step.

[0201] To a mixture of benzyl (2S)-2-[[(2R)-2-(tert-butoxycarbonylamino)propyl]amino]propanoate (3.10 g, 9.21 mmol) and pyridine (1.46 g, 18.4 mmol) in DCM (40 mL) at 0 °C was added the solution of hex-3-ynyl trifluoromethanesulfonate from the previous procedure. The reaction was stirred at 0 °C for 4 h, allowed to warm to room temperature, and stirred at room temperature overnight. The reaction mixture was concentrated under reduced pressure, and the residue was purified by flash chromatography on silica gel eluting with petroleum ether and ethyl acetate (5 / 1) to afford the title compound (1.21 g, 31.7% yield) as a yellow solid. LCMS (ESI): [M+H] = 417.4; 1 H NMR (300MHz, DMSO-d6): δ7.38-7.30(m,5H), 6.43(d,J=8.4Hz,1H), 5.10(s,2H), 3.58(q,J=6.9Hz,1H), 3.48-3.42(m, 1H), 2.65-2.62(m,2H), 2.49-2.38(m,2H), 2.18-2.06(m,4H), 1.36(s,9H), 1.18(d,J=7.2Hz,3H), 1.04-0.94(m,6H). Step 3. (2S)-2-[[(2R)-2-(tert-butoxycarbonylamino)propyl]-hex-3-ynyl-amino]propanoic acid TIFF0007670618000118.tif30170

[0202] A solution of LiOH (476 mg, 19.8 mmol) in HO (20 mL) was added to benzyl (2S)-2-[[(2R)-2-(tert-butoxycarbonylamino)propyl]-hex-3-ynyl-amino]propanoate (2.02 g, 4.85 mmol) in MeOH (60 mL) at 0 °C. The reaction mixture was stirred at room temperature overnight and then acidified to pH ∼5 with 1 M HCl at 0 °C. Most of the MeOH was evaporated under vacuum. The remaining solution was extracted with ethyl acetate (3 × 50 mL). The organic layers were combined, dried over NaSO and concentrated under reduced pressure. The residue was purified by flash chromatography on silica gel eluting with dichloromethane and methanol (5 / 1) to give the title compound (681 mg, 43% yield) as a yellow oil. LCMS (ESI): [M+H] + =327.4. Step 4. N-((R)-2-((((9H-fluoren-9-yl)methoxy)carbonyl)amino)propyl)-N-(hex-3-yn-1-yl)-L-alanine

[0203] (2S)-2-[[(2R)-2-(tert-butoxycarbonylamino)propyl]-hex-3-ynyl-amino]propanoic acid (980 mg, 3.01 mmol) was treated with TFA (20 mL) in DCM (20 mL) at room temperature for 1 hour. The reaction mixture was then concentrated under reduced pressure. THF (30 mL) was added to the residue, followed by a solution of LiOH (216 mg, 9.01 mmol) in HO (10 mL) at 0° C. A solution of fluorenylmethyloxycarbonyl chloride (930 mg, 3.60 mmol) in THF (10 mL) was added at 0° C. The reaction mixture was stirred at room temperature for 1 hour and acidified to pH ∼5 with 1 M HCl. Ethyl acetate (50 mL) was added, and the phases were separated. The aqueous phase was extracted with ethyl acetate (2 × 50 mL). The organic layers were combined, dried over NaSO, and concentrated under reduced pressure. The residue was purified by flash chromatography on silica gel eluting with DCM and methanol (5 / 1) to give the title compound (1.12 g, 83.4% yield) as a white solid. LCMS (ESI): [M+H] + =449.2;1 H NMR (300MHz, DMSO-d6): δ7.89(d,J=7.5Hz,2H), 7.68(d,J=7.5Hz,2H), 7.44-7.39(m,2H), 7.35-7.30(m,2H), 7.14(br,1H), 4.29- 4.21(m,3H), 3.62-3.45(m,2H), 2.80-2.63(m,3H), 2.32-2.21(m,3H), 2.13-2.06(m,2H), 1.20(J=6.9Hz,3H), 1.06-0.98(m,6H). JPEG0007670618000119.jpg151170JPEG0007670618000120.jpg232170JPEG0007670618000121.jpg255170[Example 1] (3S,6S,9S,12S,15S)-6-(aminomethyl)-15,16-dibutyl-9-cyclohexyl-3-(hydroxymethyl)-12-isobutyl-13-methyl-1,4,7,10,13,16-hexaazacyclooctadecane-2,5,8,11,14-pentane (trifluoroacetate) TIFF0007670618000122.tif49170Step 1. Methyl (S)-2-((2-((tert-butoxycarbonyl)amino)ethyl)(butyl)amino)hexanoate TIFF0007670618000123.tif42170

[0204] To a mixture of methyl (2S)-2-aminohexanoate hydrochloride (1.55 g, 7.68 mmol) and methanol (20 mL), sodium acetate (1.279 g, 15.59 mmol) was added, followed by butyraldehyde (0.70 mL, 7.8 mmol) and sodium cyanoborohydride (1.07 g, 16.2 mmol). The reaction mixture was stirred at room temperature for 2 hours. To the reaction mixture, N-Boc-2-aminoacetaldehyde (1.628 g, 9.72 mmol) and sodium cyanoborohydride (1.03 g, 15.6 mmol) were added, and the reaction mixture was continued at room temperature for 2.5 hours, then at 50° C. for 18 hours. To the reaction mixture, N-Boc-2-aminoacetaldehyde (1.098 g, 6.556 mmol) and sodium cyanoborohydride (1.13 g, 17.1 mmol) were added, and the reaction mixture was continued to stir at room temperature for 20 h. The reaction mixture was quenched with saturated aqueous sodium bicarbonate and extracted with DCM (3 × 100 mL). The DCM portion was dried over magnesium sulfate, filtered, and evaporated in vacuo. The crude product was purified by flash chromatography on silica gel (80 g silica, solvent gradient: 0–50% ethyl acetate in heptane) to afford 1.491 g (56%) of the title compound as a clear, colorless oil. LCMS (ESI): [M+H] + =345.2. Step 2. (S)-2-((2-((tert-butoxycarbonyl)amino)ethyl)(butyl)amino)hexanoic acid TIFF0007670618000124.tif41170

[0205] To a solution of methyl (S)-2-((2-((tert-butoxycarbonyl)amino)ethyl)(butyl)amino)hexanoate (1.488 g, 4.319 mmol) in THF (12 mL) was added lithium hydroxide (1.0 M in water, 9 mL, 9 mmol). The resulting mixture was stirred at room temperature for 20 hours, then at 50° C. for 4 hours. Additional lithium hydroxide (1.0 M in water, 4 mL, 4 mmol) was added, and the reaction mixture was stirred at room temperature for 3 days. The reaction mixture was evaporated in vacuo, and the crude product was purified by flash chromatography on silica gel (40 g silica, solvent gradient: 5-10% methanol + 0.5% formic acid in dichloromethane) to afford 1.3437 g (94%) of the title compound. LCMS (ESI): [M+H] + =331.15. Step 3. (3S,6S,9S,12S,15S)-6-(aminomethyl)-15,16-dibutyl-9-cyclohexyl-3-(hydroxymethyl)-12-isobutyl-13-methyl-1,4,7,10,13,16-hexaazacyclooctadecane-2,5,8,11,14-pentane The title compound was prepared as the trifluoroacetic acid salt using (S)-2-((2-((tert-butoxycarbonyl)amino)ethyl)(butyl)amino)hexanoic acid following a procedure similar to that described for Intermediate 15 (substituting the appropriate Fmoc-protected amino acid) and Example 9, steps 2-5. LCMS (ESI):R T (min)=3.503, [M+H] + =652.5, method=A. [Example 2] (3S,6S,9S,12S,15S)-6-(aminomethyl)-16-(4-butylbenzyl)-9-cyclohexyl-3-(hydroxymethyl)-12-isobutyl-13,15-dimethyl-1,4,7,10,13,16-hexaazacyclooctadecane-2,5,8,11,14-pentane (trifluoroacetate) TIFF0007670618000125.tif47170 Step 1. Benzyl (((2S,5S,11S,14S,17S)-5-((benzyloxy)methyl)-10-(4-butylbenzyl)-17-cyclohexyl-14-isobutyl-11,13 dimethyl-3,6,12,15,18 pentaoxo-1,4,7,10,13,16-hexaazacyclooctadecan-2-yl)methyl)carbamate TIFF0007670618000126.tif59170

[0206] The title compound was prepared following a procedure similar to that described for Example 4. LCMS (ESI): [M+H] + =924.45. Step 2. (3S,6S,9S,12S,15S)-6-(aminomethyl)-16-(4-butylbenzyl)-9-cyclohexyl-3-(hydroxymethyl)-12-isobutyl-13,15-dimethyl-1,4,7,10,13,16-hexaazacyclooctadecane-2,5,8,11,14-pentane

[0207] To a solution of benzyl (((2S,5S,11S,14S,17S)-5-((benzyloxy)methyl)-10-(4-butylbenzyl)-17-cyclohexyl-14-isobutyl-11,13-dimethyl-3,6,12,15,18-pentaoxo-1,4,7,10,13,16-hexaazacyclooctadecan-2-yl)methyl)carbamate (198 mg, 0.214 mmol) in DCM (2.0 mL, 31 mmol) at 0° C. was added boron tribromide (1 mol / L in DCM) (0.52 mL, 0.52 mmol). The reaction mixture was stirred at 0° C. for 2 hours, after which additional boron tribromide (1 mol / L in dichloromethane) (0.30 mL, 0.30 mmol) was added. The reaction mixture was stirred at 0° C. for 1 hour and at room temperature for 2 hours. The reaction mixture was quenched with methanol, diluted with DCM, and washed with saturated aqueous sodium bicarbonate. The layers were separated and the aqueous layer was extracted with additional DCM. The combined DCM layers were dried over magnesium sulfate, filtered, and evaporated in vacuo. The crude product was purified by reverse-phase HPLC and lyophilized to give 36.4 mg (24%) of the title compound as its trifluoroacetate salt. LCMS (ESI):R T (minutes)=4.586, [M+H] + =700.5, method=A. [Example 3] (3S,6S,9S,12S,15S)-6-(aminomethyl)-9-cyclohexyl-15-(2-fluoroethyl)-16-hexyl-3-((S)-1-hydroxyethyl)-12-isobutyl-13-methyl-1,4,7,10,13,16-hexaazacyclooctadecane-2,5,8,11,14-pentane (trifluoroacetate) TIFF0007670618000127.tif49170Step 1. Benzyl (tert-butoxycarbonyl)-L-homoserinate TIFF0007670618000128.tif38170

[0208] To a solution of (3S)-4-benzyloxy-3-(tert-butoxycarbonylamino)-4-oxobutanoic acid (1.016 g, 3.143 mmol) dissolved in 1,2-dimethoxyethane (3 mL) at 0 °C, 4-methylmorpholine (0.35 mL, 3.2 mmol) was added, followed by the slow addition of isobutyl chloroformate (0.44 mL, 3.4 mmol). The reaction mixture was stirred at 0 °C for 30 minutes, then filtered through Celite and rinsed with 1,2-dimethoxyethane (3 mL). The filtered solution was cooled to 0 °C and treated with the slow addition of a solution of sodium borohydride (158 mg, 4.135 mmol) in water (1.5 mL). After 5 minutes, water (15 mL) was added. The resulting suspension was extracted with ethyl acetate. The organic portion was dried over brine and magnesium sulfate, filtered, and evaporated in vacuo. The crude product was purified by flash chromatography on silica gel (40 g silica, solvent gradient: 0-100% ethyl acetate in DCM) to give 485.6 mg (50%) of the title compound as a clear, colorless oil. LCMS (ESI): [M+HtButyl] + =254.1, [M+H-Boc] + =210.1. Step 2. Benzyl (S)-2-((tert-butoxycarbonyl)amino)-4-fluorobutanoate TIFF0007670618000129.tif37170

[0209] To a solution of benzyl (tert-butoxycarbonyl)-L-homoserinate (550.2 mg, 1.778 mmol) in 1,4-dioxane (3.5 mL) was added pyridine-2-sulfonyl fluoride (324.6 mg, 1.913 mmol), followed by 1,3,4,6,7,8-hexahydro-1-methyl-2H-pyrimido[1,2-a]pyrimidine (0.52 mL, 3.6 mmol). The resulting mixture was stirred in a sealed vial under air at room temperature for 19 hours. The reaction mixture was diluted with ethyl acetate, washed with water (2x) and brine, dried over magnesium sulfate, filtered, and evaporated in vacuo. Purification of the crude product by flash chromatography on silica gel (24 g silica, solvent gradient: 0-40% ethyl acetate in heptane) afforded 356.6 mg (64%) of the title compound as a clear, colorless oil that crystallized after standing overnight at room temperature. LCMS (ESI): [M+H-Boc] + =212.1; 1 H NMR (400MHz, DMSO-d6) δ7.43-7.30(m,6H), 5.23-5.05(m,2H), 4.60-4.34(m,2H), 4.12(td,J=8.9,4.8Hz,1H), 2.17-1.87(m,2H), 1.37(s,9H). Step 3. Benzyl (S)-2-amino-4-fluorobutanoate hydrochloride TIFF0007670618000130.tif29170

[0210] To a solution of benzyl (2S)-2-(tert-butoxycarbonylamino)-4-fluoro-butanoate (355 mg, 1.14 mmol) in DCM (3 mL) was added hydrogen chloride (4.0 mol / L in dioxane, 3.0 mL, 12 mmol). The reaction mixture was stirred at room temperature for 5 hours. The reaction mixture was evaporated in vacuo to give the title compound as the HCl salt, which was carried forward without purification. LCMS (ESI): [M+H] + =LCMS m / z=212.05. Step 4. (3S,6S,9S,12S,15S)-6-(aminomethyl)-9-cyclohexyl-15-(2-fluoroethyl)-16-hexyl-3-((S)-1-hydroxyethyl)-12-isobutyl-13-methyl-1,4,7,10,13,16-hexaazacyclooctadecane-2,5,8,11,14-pentane

[0211] The title compound was prepared as the trifluoroacetate salt following a procedure similar to that described for Example 4 using benzyl (S)-2-amino-4-fluorobutanoate hydrochloride. LCMS (ESI): R T (min)=7.733, [M+H] + =684.5, method=B; 1 H NMR (400MHz, methanol-d4) δ4.66-4.33(m,4H), 4.19-3.96(m,3H), 3.69-3.36(m, 7H), 2.83(s,2H), 2.11-1.54(m,12H), 1.54-1.13(m,15H), 1.13-0.90(m,12H). [Example 4] (3S,6S,9S,12S,15S)-6-(aminomethyl)-9-cyclohexyl-16-hexyl-3-((S)-1-hydroxyethyl)-12-isobutyl-13-methyl-15-(2,2,2-trifluoroethyl)-1,4,7,10,13,16-hexaazacyclooctadecane-2,5,8,11,14-pentane (trifluoroacetate) TIFF0007670618000131.tif50170Step 1. Benzyl (S)-2-((2-((tert-butoxycarbonyl)amino)ethyl)(hexyl)amino)-4,4,4-trifluorobutanoate TIFF0007670618000132.tif38170

[0212] To a mixture of benzyl (2S)-2-amino-4,4,4-trifluorobutanoate hydrochloride (275.4 mg, 0.9708 mmol) and N-Boc-2-aminoacetaldehyde (175.0 mg, 1.04 mmol) in ethyl acetate (10.0 mL) was added sodium triacetoxyborohydride (421 mg, 1.98 mmol). The reaction mixture was stirred at room temperature for 3 days. To the reaction mixture was added hexanal (0.25 mL, 2.1 mmol) and sodium triacetoxyborohydride (439 mg, 2.0713 mmol). The reaction mixture was heated at 50° C. for 2 hours, after which hexanal (0.30 mL, 2.5 mmol) and sodium triacetoxyborohydride (380 mg, 1.79 mmol) were added. After an additional 3.5 hours, the reaction mixture was diluted with ethyl acetate, washed with aqueous sodium bicarbonate and brine, dried over magnesium sulfate, filtered, and evaporated in vacuo. The crude product was purified by flash chromatography on silica gel (40 g silica, solvent gradient: 0-75% ethyl acetate in heptane) to afford 350.8 mg (76%) of the title compound as a clear, colorless oil. LCMS (ESI): [M+H] + =475.25. Step 2. (3S,6S,9S,12S,15S)-6-(aminomethyl)-9-cyclohexyl-16-hexyl-3-((S)-1-hydroxyethyl)-12-isobutyl-13-methyl-15-(2,2,2-trifluoroethyl)-1,4,7,10,13,16-hexaazacyclooctadecane-2,5,8,11,14-pentane

[0213] The title compound was prepared as the trifluoroacetate salt following a procedure similar to that described for Example 9, steps 2-5, using benzyl (S)-2-((2-((tert-butoxycarbonyl)amino)ethyl)(hexyl)amino)-4,4,4-trifluorobutanoate. LCMS (ESI): R T (min)=3.00, [M+H] + =720.47, method=C; 1H NMR(400MHz,DMSO-d6)δ9.40(s,1H), 8.22(d,J=8.4Hz,1H), 7.89-7.66(m,3H), 7.48-7.21(m,1H), 4.44(dd,J=9.8,4.9Hz,1H), 4. 30-3.77(m,3H), 3.71(d,J=9.4Hz,1H), 3.24-2.77(m,7H), 2.71(s,2H), 2.68-2.57(m,1H), 2.41-2.30(m,2H), 1.92-0.81(m,36H). [Example 5] (3S,6S,9S,12S,15S)-6-(aminomethyl)-15-butyl-9-cyclohexyl-16-(3-fluoropropyl)-3-((S)-1-hydroxyethyl)-12-isobutyl-13-methyl-1,4,7,10,13,16-hexaazacyclooctadecane-2,5,8,11,14-pentane (trifluoroacetate) TIFF0007670618000133.tif49170Step 1. Benzyl (S)-2-((tert-butoxycarbonyl)amino)hexanoate TIFF0007670618000134.tif42170

[0214] To a solution of (2S)-2-(tert-butoxycarbonylamino)hexanoic acid (5.1549 g, 22.288 mmol) in DMF (50 mL) was added sodium bicarbonate (2.34 g, 27.8 mmol) and benzyl bromide (3.0 mL, 25 mmol). The reaction mixture was stirred at room temperature for 18 hours, after which sodium bicarbonate (1.607 g, 19.06 mmol) and benzyl bromide (1.5 mL, 12.5 mmol) were added. The reaction mixture was stirred at room temperature for an additional 4 hours, then Diluted with ethyl acetate, washed with water (2x) and brine, dried over magnesium sulfate, filtered, and evaporated in vacuo. The crude product was purified by flash chromatography on silica gel (220 g silica, solvent gradient: 0-100% ethyl acetate in heptane) to afford 5.2931 g (74%) of the title compound as a clear, colorless oil. LCMS (ESI): [M+HtButyl] + =266.15; 1H NMR(400MHz,DMSO-d6)δ7.53-7.28(m,5H), 7.26(d,J=7.7Hz,1H), 5.15(d,J=12.7Hz,1H), 5.07(d,J=1 2.7Hz,1H), 4.00-3.91(m,1H), 1.78-1.48(m,2H), 1.37(s,9H), 1.28-1.20(m,4H), 0.89-0.77(m,3H). Step 2. Benzyl (S)-2-((2-((tert-butoxycarbonyl)amino)ethyl)amino)hexanoate TIFF0007670618000135.tif40170

[0215] Benzyl (S)-2-(tert-butoxycarbonylamino)hexanoate (5.2931 g, 16.47 mmol) was dissolved in DCM (50 mL) and treated with hydrogen chloride (4 M solution in dioxane, 21 mL, 84 mmol). The resulting mixture was stirred at room temperature for 15 hours and then evaporated in vacuo. The resulting residue was combined with N-Boc-2-aminoacetaldehyde (2.762 g, 16.5 mmol) in ethyl acetate (50.0 mL) at 40° C., followed by the addition of sodium triacetoxyborohydride (32 mmol, 32 mmol) in three portions over 2.5 hours. The reaction mixture was stirred at 40° C. for 16 hours. The reaction mixture was diluted with ethyl acetate, washed with aqueous sodium bicarbonate and brine, dried over magnesium sulfate, filtered, and evaporated in vacuo. The crude product was purified by flash chromatography on silica gel (220 g silica, solvent gradient: 0-5% methanol in DCM) to give 2.2081 g (37%) of the title compound as a yellow oil. LCMS (ESI): [M+H] + =365.2; 1H NMR(400MHz,DMSO-d6)δ7.40-7.29(m,5H), 6.75-6.63(m,1H), 5.12(d,J=2.0Hz,2H), 3.20(t,J=6.6Hz,1H), 2.95(h,J=6.6Hz,2H), 2.55(dd,J=11.6,6.4Hz,1H), 2.40(dt,J=11.5,6.3Hz,1H), 1.59-1.44(m,2H), 1.37(s,9H), 1.28-1.17(m,4H), 0.85-0.77(m,3H). Step 3. Benzyl (S)-2-((2-((tert-butoxycarbonyl)amino)ethyl)(3-((tert-butyldimethylsilyl)oxy)propyl)amino)hexanoate TIFF0007670618000136.tif41170

[0216] To a mixture of benzyl (S)-2-((2-((tert-butoxycarbonyl)amino)ethyl)amino)hexanoate (0.6066 g, 1.664 mmol) and 3-[(tert-butyldimethylsilyl)]-1-propanal (0.6 mL, 3 mmol) in ethyl acetate (8.0 mL) was added sodium triacetoxyborohydride (855 mg, 4.034 mmol). The reaction mixture was stirred at 50 °C for 2 hours. The reaction mixture was diluted with ethyl acetate, washed with aqueous sodium bicarbonate and brine, dried over magnesium sulfate, filtered, and evaporated in vacuo. The crude product was purified by flash chromatography on silica gel (40 g silica, solvent gradient: 0-50% ethyl acetate in heptane) to afford 618.0 mg (69%) of the title compound as a clear, colorless oil. LCMS (ESI): [M+H] + =537.35. Step 4. Benzyl (S)-2-((2-((tert-butoxycarbonyl)amino)ethyl)(3-hydroxypropyl)amino)hexanoate TIFF0007670618000137.tif40170

[0217] To a solution of benzyl (S)-2-((2-((tert-butoxycarbonyl)amino)ethyl)(3-((tert-butyldimethylsilyl)oxy)propyl)amino)hexanoate (618.0 mg, 1.151 mmol) in THF (5 mL) was added tetrabutylammonium fluoride (1 mol / L in THF, 1.4 mL, 1.4 mmol). The reaction mixture was stirred at room temperature for 2 hours. The reaction material was quenched with saturated aqueous ammonium chloride, diluted with ethyl acetate, and washed with saturated aqueous sodium bicarbonate (2 x 50 mL). The organic portion was dried over magnesium sulfate, filtered, and evaporated in vacuo. The crude product was purified by flash chromatography on silica gel (24 g silica, solvent gradient: 0-5% methanol in dichloromethane, monitored at 210 nm) to afford 321.0 mg (66%) of the title compound. LCMS (ESI): [M+H] + =423.25; 1 H NMR(400MHz,DMSO-d6)δ7.41-7.29(m,4H), 6.57(t,J=5.7Hz,1H), 5.11(s,2H), 4.30(t,J=5.0Hz,1H), 3.46-3.33(m,2H), 2.97 -2.79(m,2H), 2.76-2.57(m,2H), 2.47-2.32(m,2H), 1.68-1.42(m,4H), 1.36(s,9H), 1.32-1.19(m,3H), 0.85(t,J=7.0Hz,3H). Step 5. Benzyl (S)-2-((2-((tert-butoxycarbonyl)amino)ethyl)(3-fluoropropyl)amino)hexanoate TIFF0007670618000138.tif41170

[0218] To a solution of benzyl (S)-2-((2-((tert-butoxycarbonyl)amino)ethyl)(3-hydroxypropyl)amino)hexanoate (321.0 mg, 0.7596 mmol) in 1,4-dioxane (1.5 mL) was added pyridine-2-sulfonyl fluoride (150.8 mg, 0.8890 mmol), followed by 1,3,4,6,7,8-hexahydro-1-methyl-2H-pyrimido[1,2-a]pyrimidine (0.24 mL, 1.7 mmol). The resulting mixture was stirred in a sealed vial (under air) at room temperature for 63 hours. The reaction mixture was diluted with ethyl acetate, washed with water (2×) and brine, dried over magnesium sulfate, filtered, and evaporated in vacuo. The crude product was purified by flash chromatography on silica gel (12 g silica, solvent gradient: 0-80% ethyl acetate in heptane) to give 233.1 mg (72%) of the title compound as a clear, colorless oil. LCMS (ESI): [M+H] + =425.2. Step 6. (3S,6S,9S,12S,15S)-6-(aminomethyl)-15-butyl-9-cyclohexyl-16-(3-fluoropropyl)-3-((S)-1-hydroxyethyl)-12-isobutyl-13-methyl-1,4,7,10,13,16-hexaazacyclooctadecane-2,5,8,11,14-pentane

[0219] The title compound was prepared following a procedure similar to that described for Example 4 using benzyl (S)-2-((2-((tert-butoxycarbonyl)amino)ethyl)(3-fluoropropyl)amino)hexanoate. LCMS (ESI): R T (min)=8.210, [M+H] + =670.5, method=B. [Example 6] (3S,6S,9S,12S,15S)-6-(aminomethyl)-15-butyl-9-cyclohexyl-16-(2-fluoroethyl)-3-((S)-1-hydroxyethyl)-12-isobutyl-13-methyl-1,4,7,10,13,16-hexaazacyclooctadecane-2,5,8,11,14-pentane (trifluoroacetate) TIFF0007670618000139.tif49170Step 1. Benzyl (S)-2-((2-((tert-butoxycarbonyl)amino)ethyl)(2-fluoroethyl)amino)hexanoate TIFF0007670618000140.tif36170

[0220] To a solution of benzyl (S)-2-((tert-butoxycarbonyl)amino)hexanoate (Example 5, Step 1) (702 mg, 1.93 mmol) in DMF (4.0 mL) was added triethylamine (3.0 mL, 21 mmol) and 1-fluoro-2-iodoethane (2.0 mL, 25 mmol). The reaction mixture was stirred at 60° C. for 17 hours in a sealed vial. 1-Bromo-2-fluoroethane (1.0 mL, 13 mmol) was added to the reaction mixture, and the mixture was heated at 60° C. for 24 hours. Trimethylamine (2.0 mL, 14 mmol) and 1-bromo-2-fluoroethane (1.0 mL, 13 mmol) were added to the reaction mixture, and the mixture was heated at 60° C. for 24 hours. The reaction mixture was diluted with ethyl acetate, washed with water (2×) and brine, dried over magnesium sulfate, filtered, and evaporated in vacuo. The crude product was purified by flash chromatography on silica gel (40 g silica, solvent gradient: 0-100% ethyl acetate in DCM) to afford 391.7 mg (50%) of the title compound as an orange oil. LCMS (ESI): [M+H] + =411.25 Step 2. (3S,6S,9S,12S,15S)-6-(aminomethyl)-15-butyl-9-cyclohexyl-16-(2-fluoroethyl)-3-((S)-1-hydroxyethyl)-12-isobutyl-13-methyl-1,4,7,10,13,16-hexaazacyclooctadecane-2,5,8,11,14-pentane The title compound was prepared following a procedure similar to that described for Example 4 using benzyl (S)-2-((2-((tert-butoxycarbonyl)amino)ethyl)(2-fluoroethyl)amino)hexanoate. LCMS (ESI): R T (min)=6.69, [M+H]+ =656.56, method=D. [Example 7] (3S,6S,9S,12S,15S,18R)-6-(aminomethyl)-15,16-dibutyl-9-cyclohexyl-3-((S)-1-hydroxyethyl)-13,18-dimethyl-12-propyl-1,4,7,10,13,16-hexaazacyclooctadecane-2,5,8,11,14-pentane (trifluoroacetate) TIFF0007670618000141.tif49170Step 1. N-Me-Norvaline-Cyclohexylglycine-Dap(Cbz)-Allothreonine(Bzl)-(2-Chlorotrityl Resin) TIFF0007670618000142.tif60170

[0221] Commercially available 2-chlorotrityl resin (resin loading 0.95 mmol / g, 5.0 g) was swollen in 40 mL of DMF / DCM (1:1) for 40 min. The resin was drained and treated with a solution of Fmoc-allothreonine(Bzl)-OH (2.46 g, 5.7 mmol) and DIPEA (2.5 g, 19.4 mmol) in DMF (15 mL). The resin was agitated with a nitrogen sparge for 1.5 h, then drained and rinsed sequentially with DCM / MeOH / DIPEA (10:10:1, 15 mL), DCM (3 × 15 mL), and DMF (3 × 15 mL).

[0222] The resin was treated with 25 mL of 20% piperidine in DMF and agitated with a nitrogen sparge for 1 hour. The resin was drained and rinsed with DMF (3 × 10 mL) and DCM (3 × 10 mL). The resin was treated with a solution of (S)-2-(Fmoc-amino)-3-(((benzyloxy)carbonyl)amino)propanoic acid (4.37 g, 9.49 mmol), HATU (3.61 g, 9.50 mmol), HOBt (1.29 g, 9.53 mmol), and DIPEA (3.31 mL, 19.03 mmol) in DMF (50 mL) (premixed for 15 min at room temperature). The resin was shaken overnight on a rotator, drained, and rinsed with DMF (2 × 15 mL) and DCM (3 × 15 mL).

[0223] The resin was treated with 10 mL of 20% piperidine in DMF and agitated with a nitrogen sparge for 60 minutes. The resin was drained and rinsed with DMF (3 × 15 mL) and DCM (3 × 15 mL). The resin was treated with a solution of Fmoc-S-cyclohexylglycine-OH (3.61 g, 9.50 mmol) (premixed at room temperature for 15 minutes), a stock solution of HATU / HOBt (1 / 1) (0.4 M in DMF, 24 mL, 9.6 mmol), and DIPEA (2.45 g, 19 mmol). The resin was agitated with a nitrogen sparge for 2 hours, then drained and rinsed with DMF (3 × 15 mL) and DCM (3 × 15 mL).

[0224] The resin was treated with 25 mL of 20% piperidine in DMF and agitated for 60 min with a nitrogen sparge. The resin was drained and rinsed with DMF (3 × 15 mL) and DCM (3 × 15 mL). The resin was treated with a premixed solution of (S)-Fmoc-N-Me-norvaline-OH (3.37 g, 9.52 mL, 0.4 M HATU / HOBt (1 / 1) in DMF, 24 mL, 9.6 mmol) and DIPEA (2.53 g, 19.6 mmol). The resin was agitated for 3 h, drained, and rinsed with DMF (3 × 15 mL) and DCM (3 × 15 mL).

[0225] The resin was treated with 25 mL of 20% piperidine in DMF, the vial was placed on a rotator at room temperature for 3 h, then filtered, rinsed with DMA (3 × 15 mL) and DCM (3 × 15 mL), and dried under vacuum to give the resin-bound peptide (7.58 g, estimated resin loading 0.5 mmol / g). Step 2. (6R,9S,12S,15S,18S,21S)-18-((((benzyloxy)carbonyl)amino)methyl)-21-((S)-1-(benzyloxy)ethyl)-8,9-dibutyl-15-cyclohexyl-2,2,6,11-tetramethyl-4,10,13,16,19-pentaoxo-12-propyl-3-oxa-5,8,11,14,17,20-hexaazadocosanoic acid TIFF0007670618000143.tif60170

[0226] The resin-bound peptide from the previous step (1.00 g, approximately 0.5 mmol / g resin loading) was treated with a premixed solution of (2S)-2-[[(2R)-2-(tert-butoxycarbonylamino)propyl]-butyl-amino]hexanoic acid (447 mg, 1.29 mmol, Intermediate T5), PyAOP (782 mg, 1.51 mmol), and DIPEA (387 mg, 3 mmol) in DMF (10 mL). The resin was shaken on a rotator for 24 h. The resin was drained and rinsed with DMF (3 × 10 mL) and DCM (3 × 10 mL). The resin was treated with a 1 / 4 HFIP / DCM solution (enough to cover the resin 1.5 times), shaken at room temperature for 1 h, and then filtered. The liquid phase was collected. The process was repeated with 1 / 4 HFIP / DCM, and the liquid phase was collected. The resin was washed with DCM (3 x 10 mL). The filtered liquids were combined and concentrated in vacuo. The residue was purified by reversed-phase flash chromatography on a C18 column to give the title compound (364 mg, 0.361 mmol). LCMS (ESI): [M+H] + =1008.6. Step 3. (2S,5S,8S,11S,14S)-15-((R)-2-aminopropyl)-5-((((benzyloxy)carbonyl)amino)methyl)-2-((S)-1-(benzyloxy)ethyl)-14-butyl-8-cyclohexyl-12-methyl-4,7,10,13-tetraoxo-11-propyl-3,6,9,12,15-pentaazanonadecano-1-oic acid TIFF0007670618000144.tif60170

[0227] (6R,9S,12S,15S,18S,21S)-18-((((benzyloxy)carbonyl)amino)methyl)-21-((S)-1-(benzyloxy)ethyl)-8,9-dibutyl-15-cyclohexyl-2,2,6,11-tetramethyl-4,10,13,16,19-pentaoxo-12-propyl-3-oxa-5,8,11,14,17,20-hexaazadocosan-22-oic acid (204 mg, 0.202 mmol) was dissolved in TFA (20 mL) at 0° C. The resulting mixture was stirred at 0° C. for 1 hour and then diluted with toluene (40 mL). The resulting mixture was concentrated in vacuo. An additional 40 mL of toluene was added and the resulting mixture was concentrated in vacuo to provide the title compound as a TFA salt (180 mg). LCMS(ESI):[M+H] + =908.5. Step 4. Benzyl (((2S,5S,8R,11S,14S,17S)-5-((S)-1-(benzyloxy)ethyl)-10,11-dibutyl-17-cyclohexyl-8,13-dimethyl-3,6,12,15,18-pentaoxo-14-propyl-1,4,7,10,13,16-hexaazacyclooctadecan-2-yl)methyl)carbamate TIFF0007670618000145.tif60170

[0228] (2S,5S,8S,11S,14S)-15-((R)-2-aminopropyl)-5-((((benzyloxy)carbonyl)amino)methyl)-2-((S)-1-(benzyloxy)ethyl)-14-butyl-8-cyclohexyl-12-methyl-4,7,10,13-tetraoxo-11-propyl-3,6,9,12,15-pentaazanonadecanoic acid (184 mg, 0.202 mmol) was dissolved in THF (160 mL) and DIPEA (78 mg, 0.60 mmol) was added. The resulting solution was cooled to 0 °C. A solution of HATU (76.9 mg, 0.200 mmol) and HOBt (27.0 mg, 0.200 mmol) in DMF (2.0 mL) was diluted with THF (20 mL), and the resulting THF / DMF solution was then added dropwise to the cold peptide solution. After the reaction was complete, the reaction mixture was concentrated in vacuo and the THF was stripped off. The residue was diluted with 3 mL of DMF and added dropwise to water (60 mL) with vigorous stirring. The mixture was stirred for an additional 15 min. The solid was collected by filtration, washed with water, and dried under vacuum to give the crude cyclic peptide (170 mg), which was used directly without further purification. LCMS (ESI): [M+H] + =890.5. Step 4. (3S,6S,9S,12S,15S,18R)-6-(aminomethyl)-15,16-dibutyl-9-cyclohexyl-3-[(1S)-1-hydroxyethyl]-13,18-dimethyl-12-propyl-1,4,7,10,13,16-hexaazacyclooctadecane-2,5,8,11,14-pentone

[0229] A mixture of the crude cyclic peptide from step 3 (132 mg) and 10% Pd / C (232 mg) in ethyl acetate (20 mL) was stirred under hydrogen gas (1 atm) at 25° C. for 24 hours. The catalyst was removed by filtration, and the filtered solution was concentrated in vacuo. The residue was purified by preparative LCMS to give the title compound (13.0 mg) as a TFA salt as a white solid. LCMS (ESI): [M+H] + =666.5, R t =2.38 minutes, Method J. 1H NMR (400MHz, DMSO-d6): δ8.74-8.64(m,2H), 8.05(br,3H), 7.92-7.04(m,2H), 6.09(br,1 H), 5.22-5.14(m,1H), 4.81-4.73(m,2H), 4.47-4.41(m,1H), 4.33-4.28(m,1H), 4.03-3.8 7(m,3H), 3.78-3.73(m,1H), 3.21(s,2H), 3.19-3.10(m,3H), 2.99-2.92(m,1H), 2.05-1. 95(m,2H), 1.88-1.76(m,3H), 1.72-1.62(m,6H), 1.44-1.06(m,16H), 0.97-0.84(m,15H). [Example 8] (1R,4S,7S,10S,13S,16R,19R)-10-(aminomethyl)-7-cyclohexyl-13-[(1S)-1-hydroxyethyl]-4-isobutyl-3,16-dimethyl-18-oxa-3,6,9,12,15-pentazabicyclo[17.8.0]heptacosane-2,5,8,11,14-pentone (trifluoroacetate) TIFF0007670618000146.tif47170Step 1. N-Me-Leucine-Cyclohexylglycine-Dap(Boc)-Allothreonine(t-Bu)-(2-Chlorotrityl Resin) TIFF0007670618000147.tif51170

[0230] The resin-bound tetrapeptide was prepared according to a method similar to that described for Example 7. Step 2. (2S,5S,8S,11S,14R,15R,18R)-18-amino-14-(but-3-en-1-yl)-2-((S)-1-(tert-butoxy)ethyl)-5-(((tert-butoxycarbonyl)amino)methyl)-8-cyclohexyl-15-(hex-5-en-1-yl)-11-isobutyl-12-methyl-4,7,10,13-tetraoxo-16-oxa-3,6,9,12-tetraazanonadecano-1-oic acid TIFF0007670618000148.tif54170

[0231] The resin-bound tetrapeptide from step 1 (1.0 g, approximately 0.45 mmol / g loading) was treated with THF (5 mL) and DIPEA (308 mg, 2.39 mmol) and allowed to swell for 30 min. Separately, a solution of (2R,3R)-2-but-3-enyl-3-[(2R)-2-(9H-fluoren-9-ylmethoxycarbonylamino)propoxy]non-8-enoic acid (364 mg, 0.720 mmol, Intermediate 14) in THF (2.0 mL) was added to a solution of triphosgene (71.5 mg, 0.240 mmol) in THF (2.0 mL). 2,4,6-Trimethylpyridine (1.25 g, 10.4 mmol) was added dropwise, causing the mixture to heat up and the formation of a colorless precipitate. The suspension was gently shaken for approximately 1 minute and then added to the pretreated resin. The mixture was placed on a rotator at room temperature for 3 hours, drained, and then rinsed with DMF (10 mL) and DCM (2 × 10 mL). The resin was treated with 10 mL of 20% piperidine in DMF, agitated with a nitrogen stream for 1 hour, drained, and rinsed with DMF (2 × 10 mL) and DCM (2 × 10 mL). The resin was then treated with a 1 / 4 HFIP / DCM solution (20 mL), shaken on a rotator at room temperature for 1 hour, and then filtered. The liquid phase was collected. This process was repeated with 1 / 4 HFIP / DCM, and the liquid phase was collected. The resin was washed with DCM (3 × 10 mL). The filtered liquids were combined and concentrated under vacuum. The residue was purified by reverse-phase flash chromatography on a C18 column to give the title compound as a yellow solid (300 mg, 62%). LCMS(ESI):[M+H] + =893.6. Step 3. tert-Butyl N-[[(3R,6S,9S,12S,15S,18R,19R)-18-but-3-enyl-6-[(1S)-1-tert-butoxyethyl]-12-cyclohexyl-19-hex-5-enyl-15-isobutyl-3,16-dimethyl-5,8,11,14,17-pentaoxo-1-oxa-4,7,10,13,16-pentaazacyclononadequa-9-yl]methyl]carbamate TIFF0007670618000149.tif54170

[0232] To a mixture of the peptide from step 2 (300 mg, 0.340 mmol) and DIPEA (175 mg, 1.35 mmol) in THF (200 mL) at 0 °C was added dropwise a solution of HATU (129 mg, 0.340 mmol) and HOBt (129 mg, 0.34 mmol) in DMF (2.0 mL). The reaction mixture was stirred at 0 °C for 2 h, then concentrated in vacuo to strip off THF. The residue was diluted with DMF (3 mL) and added dropwise to 60 mL of water with vigorous stirring. The resulting solid was collected by filtration, washed with water (3 × 10 mL), and dried to give the crude product (271 mg) as a yellow solid. LCMS (ESI): [M+H] + =875.5. Step 4. tert-Butyl N-[[(1R,4S,7S,10S,13S,16R,19R,24Z)-13-[(1S)-1-tert-butoxyethyl]-7-cyclohexyl-4-isobutyl-3,16-dimethyl-2,5,8,11,14-pentaoxo-18-oxa-3,6,9,12,15-pentazabicyclo[17.8.0]heptacos-24-en-10-yl]methyl]carbamate TIFF0007670618000150.tif54170

[0233] To a solution of the cyclic peptide from step 3 (110 mg, 0.130 mmol) in DCE (70 mL) was added dropwise a solution of dichloro-[(2-isopropoxyphenyl)methylene]-(tricyclohexyl-λ^{5}-phosphanyl)ruthenium (43 mg, CAS 203714-71-0) in DCE (1.0 mL) under nitrogen at 0°C. The reaction was stirred overnight at 80°C, and then additional catalyst (43 mg, CAS 203714-71-0) was added. The reaction was stirred at 80°C for an additional 8 hours, and additional catalyst (43 mg, CAS 203714-71-0) was added. This process was repeated two more times. The total catalyst added was 5 × 43 mg, and the total reaction time was 3 days. The reaction mixture was concentrated under reduced pressure. The residue was dissolved in 2.0 mL of DCE, and the DCE solution was added dropwise to 40 mL of n-hexane. The mixture was stirred overnight at room temperature. The solid was collected by filtration, washed with n-hexane (3 × 10 mL), and dried to give the crude product (100 mg) as a brown solid, which was carried on to the next step without further purification. Step 5. tert-Butyl N-[[(1R,4S,7S,10S,13S,16R,19R)-13-[(1S)-1-tert-butoxyethyl]-7-cyclohexyl-4-isobutyl-3,16-dimethyl-2,5,8,11,14-pentaoxo-18-oxa-3,6,9,12,15-pentaazabicyclo[17.8.0]heptacosan-10-yl]methyl]carbamate TIFF0007670618000151.tif54170

[0234] A mixture of the crude bicyclic olefin from step 4 and 10% Pd / C (433 mg) in ethyl acetate (50 mL) was stirred under a hydrogen gas atmosphere at 25° C. for 15 hours. The catalyst was removed by filtration, and the filtrate was concentrated under reduced pressure. The residue was purified by reverse-phase flash chromatography on a C18 column to give the title compound (23.2 mg) as a yellow oil. LCMS (ESI): [M+H] + =849.5. Step 6. (1R,4S,7S,10S,13S,16R,19R)-10-(aminomethyl)-7-cyclohexyl-13-[(1S)-1-hydroxyethyl]-4-isobutyl-3,16-dimethyl-18-oxa-3,6,9,12,15-pentazabicyclo[17.8.0]heptacosane-2,5,8,11,14-pentone TIFF0007670618000152.tif47170

[0235] The product from step 5 (23.2 mg) was treated with TFA (5.0 mL) at 0° C. for 1 hour, then concentrated under reduced pressure. The residue was purified by preparative LCMS (Column: XBridge Prep C18 OBD Column, 5 μm, 19*150 mm; Mobile phase A: Water (0.05% TFA), Mobile phase B: ACN; Flow rate: 25 mL / min; Gradient: 20% B to 50% B in 20 min; 220 nm; Rt: 18.74 min) to give the trifluoroacetate salt of the title compound (5.3 mg) as a yellow solid. LCMS (ESI): [M+H] + =693.5, R t =2.5 minutes, method H. 1 H NMR (400MHz, DMSO-d6): δ8.72-8.67(m,1H), 8.40-8.25(m,1H), 7.94-7.88(m,3H) , 7.64-7.45(m,1H), 7.24-6.95(m,1H), 4.63-4.54(m,2H), 4.18-4.07(m,2H), 3.95 -3.86(m,2H), 3.67-3.60(m,2H), 3.50-3.43(m,1H), 3.21-3.03(m,4H), 2.78(s,2) H), 2.14-2.07(m,1H), 1.83-1.43(m,24H), 1.24-1.09(m,6H), 1.01-0.88(m,12H). [Example 9] (3S,6S,9S,12S,15S,18R)-6-(aminomethyl)-9-cyclohexyl-16-(cyclopentylmethyl)-18-ethyl-3-((S)-1-hydroxyethyl)-12-isobutyl-13,15-dimethyl-1,4,7,10,13,16-hexaazacyclooctadecane-2,5,8,11,14-pentane (trifluoroacetate) TIFF0007670618000153.tif49170Step 1. Benzyl N-((R)-2-((tert-butoxycarbonyl)amino)butyl)-N-(cyclopentylmethyl)-L-alaninate TIFF0007670618000154.tif36170

[0236] To a solution of tert-butyl N-[(1R)-1-(hydroxymethyl)propyl]carbamate (1.037 g, 5.480 mmol) in DCM (16 mL, 249.6 mmol) was added Dess-Martin periodinane (1.2 equiv.). The reaction mixture was stirred at room temperature for 4 hours. To the reaction mixture were added benzyl (2S)-2-aminopropanoate hydrochloride (911 mg, 4.22 mmol), sodium triacetoxyborohydride (1.816 g, 8.568 mmol), and ethyl acetate (10 mL). The reaction mixture was stirred at room temperature for 2 hours. Then, to this mixture were added cyclopentanecarbaldehyde (0.80 mL, 7.6 mmol) and sodium triacetoxyborohydride (1.65 g, 7.79 mmol), and the reaction was continued at room temperature for 18 hours. The reaction mixture was diluted with ethyl acetate, washed with aqueous sodium bicarbonate and brine, dried over magnesium sulfate, filtered, and evaporated in vacuo. The crude product was purified by flash chromatography on silica gel (80 g silica, solvent gradient: 0-70% isopropyl acetate in heptane) to give 1.025 g (56%) of the title compound. LCMS (ESI): [M+H] + =433.3 Step 2. N-((R)-2-((tert-butoxycarbonyl)amino)butyl)-N-(cyclopentylmethyl)-L-alanine TIFF0007670618000155.tif33170Benzyl N-((R)-2-((tert-butoxycarbonyl)amino)butyl)-N-(cyclopentylmethyl)-L-alaninate in ethanol (10 mL) under nitrogen

[0237] To a solution of (1.1878 g, 2.746 mmol) was added palladium (10 wt% on carbon) (154 mg, 0.1447 mmol). The reaction vessel was purged with nitrogen then hydrogen and stirred under a balloon of hydrogen at room temperature for 4 hours. The reaction mixture was filtered through Celite, rinsed with methanol, and evaporated in vacuo to give the title compound, which was carried forward without purification. LCMS (ESI): [M+H] + =343.25. Step 3. N-((S)-2-((S)-2-((S)-2-((S)-2-(((R)-2-aminobutyl)(cyclopentylmethyl)amino)-N-methylpropanamido)-4-methylpentanamido)-2-cyclohexylacetamido)-3-(((benzyloxy)carbonyl)amino)propanoyl)-O-benzyl-L-allothreonine TIFF0007670618000156.tif51170

[0238] To a filter tube was added Intermediate 15 (0.770 g, estimated 0.47 mmol / g) and 8 mL of DMA. The resin was allowed to swell at room temperature for 90 minutes and then drained. A solution of N-((R)-2-((tert-butoxycarbonyl)amino)butyl)-N-(cyclopentylmethyl)-L-alanine (231 mg, 0.674 mmol) and PyAOP (789 mg, 1.453 mmol) in DMF (4 mL) was added to the resin, followed by DIPEA (0.40 mL, 2.3 mmol). The resulting mixture was placed on a rotator at room temperature for 16 hours. The resin was drained and rinsed with DMA (5x) and DCM (5x). 6 mL of 7:2:1 DCM:AcOH:trifluoroethanol was added to the resin. The mixture was placed on a rotator at room temperature for 2 hours, then filtered and rinsed with DCM. The filtered solution was evaporated in vacuo and azeotroped with toluene (3 x 2 mL). The resulting residue was dissolved in DCM (5 mL) and treated with hydrogen chloride (4.0 mol / L in dioxane, 1.0 mL, 4.0 mmol). The mixture was stirred at room temperature for 90 minutes and then evaporated in vacuo to give the crude peptide (177.7 mg, 51% crude yield), which was carried on to the next step without purification. LCMS (ESI): [M+H] + =920.7. Step 4. Benzyl (((2S,5S,8R,11S,14S,17S)-5-((S)-1-(benzyloxy)ethyl)-17-cyclohexyl-10-(cyclopentylmethyl)-8-ethyl-14-isobutyl-11,13-dimethyl-3,6,12,15,18-pentaoxo-1,4,7,10,13,16-hexaazacyclooctadecan-2-yl)methyl)carbamate TIFF0007670618000157.tif59170

[0239] To a stirred solution of DIPEA (0.2 mL, 1 mmol) and HATU (167 mg, 0.430 mmol) in THF (100 mL) at room temperature was added a solution of N-((S)-2-((S)-2-((S)-2-((S)-2-(((R)-2-aminobutyl)(cyclopentylmethyl)amino)-N-methylpropanamido)-4-methylpentanamido)-2-cyclohexylacetamido)-3-(((benzyloxy)carbonyl)amino)propanoyl)-O-benzyl-L-allothreonine (177.7 mg, 0.1858 mmol) in THF (50 mL) dropwise over 1 h. After a further 90 min, the reaction mixture was evaporated in vacuo. The resulting residue was diluted with ethyl acetate, washed with water (2×) and brine, dried over magnesium sulfate, filtered, and evaporated in vacuo. The crude product was purified by flash chromatography on silica gel (12 g silica, solvent gradient: 0-10% methanol in DCM) to give 76.6 mg of the title compound. LCMS (ESI): [M+H] + =902.7. Step 5. (3S,6S,9S,12S,15S,18R)-6-(aminomethyl)-9-cyclohexyl-16-(cyclopentylmethyl)-18-ethyl-3-((S)-1-hydroxyethyl)-12-isobutyl-13,15-dimethyl-1,4,7,10,13,16-hexaazacyclooctadecane-2,5,8,11,14-pentane

[0240] Benzyl (((2S,5S,8R,11S,14S,17S)-5-((S)-1-(benzyloxy)ethyl)-17-cyclohexyl-10-(cyclopentylmethyl)-8-ethyl-14-isobutyl-11,13-dimethyl-3,6,12,15,18-pentaoxo-1,4,7,10,13,16-hexaazacyclooctadecan-2-yl)methyl)carbamate (76.6 mg, 0.085 mmol) and ethyl acetate (5 mL) were charged to a flask, which was then purged with nitrogen. After the addition of trifluoroacetic acid (25 μL, 0.323 mmol) and palladium (10 wt % on carbon) (104 mg, 0.0977 mmol), the reaction mixture was evacuated and refilled with hydrogen and then stirred under a hydrogen balloon at room temperature for 18 hours. The reaction mixture was filtered through Celite, rinsed with 100 mL of 5% AcOH in ethanol, and the filtered solution evaporated in vacuo and azeotroped with toluene (3 x 1 mL). The crude product was purified by reverse-phase HPLC and lyophilized to give 27.6 mg (48%) of the title compound as its trifluoroacetate salt. LCMS (ESI):R T (min)=7.920, [M+H] + =762.5, method=B; 1 H NMR (400MHz, methanol-d4) δ4.93(d,J=15.6Hz,1H), 4.64-4.17(m,4H), 4.15-3.94(m,2H), 3.79-3.44(m,4H), 3.38(s,1H) ), 3.26(d,J=11.8Hz,2H), 3.13-2.92(m,1H), 2.08(dd,J=42.7,18.2Hz,4H), 1.99-1.11(m,27H), 1.11-0.90(m,11H). [Example 10] (3R,6S,9S,12S,15S,18R,19R)-9-(aminomethyl)-6-(((1r,3S)-3-(aminomethyl)cyclobutoxy)methyl)-12-cycloheptyl-3,16,18-trimethyl-15-propyl-19-(spiro[3.3]heptan-2-ylmethyl)-1-oxa-4,7,10,13,16-pentaazacyclononadecane-5,8,11,14,17-pentaone (trifluoroacetate) JPEG0007670618000158.jpg64170 Step 1. N-Me-Norvaline-Cycloheptylglycine-Dap(Boc)-trans-tert-butoxycarbonyl)amino)methyl)cyclobutyl)-serine-(2-chlorotrityl resin) JPEG0007670618000159.jpg57170

[0241] The resin-bound tetrapeptide was prepared according to a method similar to that described in Example 7. Step 2. (3R,6S,9S,12S,15S,18R,19R)-9-(aminomethyl)-6-[[3-(aminomethyl)cyclobutoxy]methyl]-12-cycloheptyl-3,16,18-trimethyl-15-propyl-19-(spiro[3.3]heptan-2-ylmethyl)-1-oxa-4,7,10,13,16-pentaazacyclononadecane-5,8,11,14,17-pentone

[0242] The title compound was prepared as a TFA salt following a similar method as described in Example 13, steps 2-4, using the intermediate from step 1 and (2R,3R)-3-((R)-2-((((9H-fluoren-9-yl)methoxy)carbonyl)amino)propoxy)-2-methyl-4-(spiro[3.3]heptan-2-yl)butanoic acid (intermediate 4). LCMS (ESI): [M+H] + =774.6, R t =2.21 minutes, method H. 1H NMR (400MHz, DMSO-d6): δ8.52-8.12(m,2H), 8.08-8.02(m,3H), 7.76(br, 3H), 7.46-7.38(m,1 H), 4.68-4.56(m,7H), 4.36-4.28(m,2H), 4.11-4.02(m,2H), 3.96-3.82(m,2H), 3.76-3.68(m ,1H), 3.48-3.40(m,1H), 3.38-3.12(m,4H), 3.07(s,1H), 2.90-2.87(m,3H), 2.73(s,2H), 2.3 9-2.28(m,2H), 2.07-1.94(m,8H), 1.83-1.70(m,5H), 1.67-1.27(m,15H), 1.02-0.87(m,8H). [Example 11] (3R,6S,9S,12S,15S,18R,19R)-6-(((2-azaspiro[3.3]heptan-6-yl)oxy)methyl)-9-(aminomethyl)-12-cycloheptyl-19-hexyl-3,16,18-trimethyl-15-propyl-1-oxa-4,7,10,13,16-pentaazacyclononadecane-5,8,11,14,17-pentaone (trifluoroacetate) TIFF0007670618000160.tif65170 Step 1. N-((S)-2-((S)-2-((S)-2-((2R,3R)-3-((R)-2-aminopropoxy)-N,2-dimethylnonanamido)pentanamido)-2-cycloheptylacetamido)-3-(((benzyloxy)carbonyl)amino)propanoyl)-O-(2-(tert-butoxycarbonyl)-2-azaspiro[3.3]heptan-6-yl)-L-serine TIFF0007670618000161.tif62170

[0243] Commercially available 2-chlorotrityl chloride resin (1.255 g, resin loading 1.5 mmol / g) was swollen in 10 mL of DMA for 4.5 h. The resin was drained and treated with a solution of N-(((9H-fluoren-9-yl)methoxy)carbonyl)-O-(2-(tert-butoxycarbonyl)-2-azaspiro[3.3]heptan-6-yl)-L-serine (Intermediate 8) (0.775 g, 1.48 mmol) and DIPEA (0.60 mL, 3.4 mmol) in DMA (5 mL). The resin was placed on a rotator at room temperature overnight. The resin was drained and rinsed with DMA (2 × 8 mL), after which the unreacted resin was capped by treating with 6 mL of 2:1 DCM:methanol for 15 min. The resin was drained and rinsed with DMA (3 × 8 mL).

[0244] Fmoc-allothreonine(Bzl)-OH (3.267 g, 7.571 mmol), HBTU (2.904 g, 7.657 mmol), DIPEA (1.9 mL, 11 mmol), and DMA (25 mL) were mixed (premixed at room temperature for 15 minutes). The resin was agitated with nitrogen for 2.5 hours, then drained and rinsed with DMA. Unreacted starting material was capped by treating the resin with 25 mL of DMA, 1 mL of DIPEA, and 1 mL of acetic anhydride. The resin was agitated with nitrogen for 5 minutes, then drained and rinsed with DMA (2 × 25 mL) and DCM (3 × 25 mL). The resin was treated with 8 mL of 20% piperidine in DMF for 2.5 hours. The resin was drained and rinsed sequentially with DMA (3 × 8 mL), DCM (3 × 8 mL), and DMA (2 × 8 mL).

[0245] The resin was treated with a solution of Fmoc-Dap(Cbz)-OH (1.384 g, 3.005 mmol), HBTU (1.14 g, 3.01 mmol), and DIPEA (0.8 mL, 5 mmol) in DMA (5 mL) (premixed for 20 min at room temperature). The resin was placed on a rotator at room temperature for 1.75 h, then drained and rinsed sequentially with 3 × 50 mL of DMA and 3 × 50 mL of DCM. The resin was treated with 8 mL of 20% piperidine in DMF for 90 min. The resin was drained and rinsed sequentially with DMA (4 × 8 mL) and DCM (4 × 8 mL).

[0246] The resin was treated with a solution of (2S)-2-cycloheptyl-2-(9H-fluoren-9-ylmethoxycarbonylamino)acetic acid (Intermediate 11, 824 mg, 2.09 mmol), HBTU (1.14 g, 3.01 mmol), and DIPEA (0.8 mL, 5 mmol) in DMA (5 mL) (premixed for 15 min at room temperature). The resin was placed on a rotator at room temperature for 2 h, then drained and rinsed with DMA (3 × 8 mL). The resin was treated with 8 mL of 20% piperidine in DMF for 1 h. The resin was drained and rinsed sequentially with DMA (3 × 8 mL), DCM (3 × 8 mL), and DMA (2 × 8 mL).

[0247] The resin was treated with a solution of Fmoc-N-Me-norvaline (1.06 g, 3.00 mmol), HBTU (1.12 g, 2.95 mmol), and DIPEA (0.8 mL, 5 mmol) in DMA (5 mL) (premixed for 15 min at room temperature). The resin was placed on a rotator at room temperature for 2 h, then drained and rinsed with DMA (3 × 8 mL) and DCM (3 × 8 mL). The resin was treated with 8 mL of 20% piperidine in DMF for 2 h. The resin was drained and rinsed sequentially with DMA (4 × 8 mL) and DCM (4 × 8 mL).

[0248] The resin was swollen in DMA, then drained and treated with a mixture of (2R,3R)-3-((R)-2-((((9H-Fluoren-9-yl)methoxy)carbonyl)amino)propoxy)-2-methylnonanoic acid (Intermediate 3) (644.0 mg, 1.377 mmol), PyAOP (980.4 mg, 1.805 mmol), and DIPEA (0.45 mL, 2.6 mmol) in DMF (5 mL, 64.4 mmol). The resulting mixture was placed on a rotator at room temperature for 18 h. The resin was filtered and rinsed with DMA (3 × 8 mL) and DCM (3 × 8 mL).

[0249] 8 mL of 20% piperidine in DMF was added to the resin. The reaction vessel was placed on a rotator at room temperature for 2.5 hours, then drained and rinsed sequentially with DMA (4 x 8 mL) and DCM (4 x 8 mL).

[0250] DCM (6 mL, 93.61 mmol) and HFIP (2 mL, 19.07 mmol) were added to the resin. The reaction vessel was placed on a rotator at room temperature for 2.5 hours. The resin was filtered, rinsed with DCM, and the filtered solution was evaporated in vacuo to give the crude peptide. LCMS (ESI): [M+H] + =1014.8. Step 2. tert-Butyl 6-(((3R,6S,9S,12S,15S,18R,19R)-9-((((benzyloxy)carbonyl)amino)methyl)-12-cycloheptyl-19-hexyl-3,16,18-trimethyl-5,8,11,14,17-pentaoxo-15-propyl-1-oxa-4,7,10,13,16-pentaazacyclononadecan-6-yl)methoxy)-2-azaspiro[3.3]heptane-2-carboxylate TIFF0007670618000162.tif70170

[0251] To a stirred solution of DIPEA (0.20 mL, 1.1 mmol) and HATU (314 mg, 0.809 mmol) in THF (200 mL) at room temperature was added a solution of N-((S)-2-((S)-2-((S)-2-((2R,3R)-3-((R)-2-aminopropoxy)-N,2-dimethylnonanamido)pentanamido)-2-cycloheptylacetamido)-3-(((benzyloxy)carbonyl)amino)propanoyl)-O-(2-(tert-butoxycarbonyl)-2-azaspiro[3.3]heptan-6-yl)-L-serine (412.0 mg, 0.4062 mmol) in THF (100 mL) dropwise over 90 minutes. After an additional hour, the reaction mixture was evaporated in vacuo. The resulting residue was diluted with ethyl acetate, washed with water (2x) and brine, dried over magnesium sulfate, filtered, and evaporated in vacuo. The crude product was purified by flash chromatography on silica gel (24 g silica, solvent gradient: 0-10% methanol in DCM) to give 157.4 mg (39%) of the title compound. LCMS (ESI): [2M+H] + =1992.15. Step 3. (3R,6S,9S,12S,15S,18R,19R)-6-(((2-azaspiro[3.3]heptan-6-yl)oxy)methyl)-9-(aminomethyl)-12-cycloheptyl-19-hexyl-3,16,18-trimethyl-15-propyl-1-oxa-4,7,10,13,16-pentaazacyclononadecane-5,8,11,14,17-pentaone

[0252] tert-Butyl 6-(((3R,6S,9S,12S,15S,18R,19R)-9-((((benzyloxy)carbonyl)amino)methyl)-12-cycloheptyl-19-hexyl-3,16,18-trimethyl-5,8,11,14,17-pentaoxo-15-propyl-1-oxa-4,7,10,13,16-pentaazacyclononadecan-6-yl)methoxy)-2-azaspiro[3.3]heptane-2-carboxylate (157.4 mg, 0.1580 mmol) and ethyl acetate (5 mL) were charged to a flask, and the flask was purged with nitrogen. After the addition of trifluoroacetic acid (25 μL, 0.323 mmol) and palladium (10 wt % on carbon) (187 mg, 0.176 mmol), the reaction mixture was evacuated, refilled with hydrogen, and stirred under a hydrogen balloon at room temperature for 16 h. The reaction mixture was filtered through Celite, rinsed with 100 mL of 5% acetic acid in ethanol, and the filtered solution evaporated in vacuo and azeotroped with toluene (3 × 1 mL). To the resulting residue at 0 °C were added DCM (2.0 mL) and trifluoroacetic acid (0.2 mL, 3 mmol). The mixture was stirred at 0 °C for 5 h, then evaporated in vacuo and azeotroped with toluene (2 × 2 mL). The resulting residue was purified by reverse-phase HPLC and lyophilized to afford 30.1 mg (21%) of the title compound as its trifluoroacetate salt. LCMS (ESI):R T (minutes)=10.073, [M+H] + =762.5, method=B; 1 H NMR(400MHz,DMSO-d6)δ8.66-8.27(m,3H), 8.18-7.69(m,5H), 7.37(d,J=8.4Hz,1H), 4.56(dd,J=10.6,4.7Hz,1H), 4.47-4.13(m,2H), 4.04(d, J=14.5Hz,1H), 3.99-3.72(m,8H), 3.64(d,J=13.2Hz,2H), 3.29-2.70(m,9H), 2.11(d,J=70.9Hz,4H), 1.87-1.11(m,25H), 1.11-0.79(m,12H). [Example 12] (3S,6S,9S,12S,15S,18R)-6-(aminomethyl)-3-((3-aminopropoxy)methyl)-15,16-dibutyl-9-cyclohexyl-12-isobutyl-13,18-dimethyl-1,4,7,10,13,16-hexaazacyclooctadecane-2,5,8,11,14-pentane (trifluoroacetate) TIFF0007670618000163.tif54170Step 1. N-Me-Leucine-Cyclohexylglycine-Dap(Boc)-3-((tert-butoxycarbonyl)amino)propyl)-Serine-(2-chlorotrityl resin) JPEG0007670618000164.jpg55170

[0253] The resin-bound tetrapeptide was prepared following a method similar to that described for Example 7, step 1. Step 2. (2S,5S,8S,11S,14S)-15-((R)-2-aminopropyl)-5-(((tert-butoxycarbonyl)amino)methyl)-2-((3-((tert-butoxycarbonyl)amino)propoxy)methyl)-14-butyl-8-cyclohexyl-11-isobutyl-12-methyl-4,7,10,13-tetraoxo-3,6,9,12,15-pentaazanonadecan-1-oic acid TIFF0007670618000165.tif70170

[0254] The resin-bound tetrapeptide from step 1 (1.50 g, resin loading approximately 0.6 mmol / g) was treated with a mixture of (2S)-2-[butyl-[(2R)-2-(9H-fluoren-9-ylmethoxycarbonylamino)propyl]amino]hexanoic acid (672 mg, 1.44 mmol, Intermediate T13), PyAOP (1.88 g, 3.60 mmol), and DIPEA (0.94 mL, 5.4 mmol) in 10 mL of DMF (premixed at room temperature for 10 min). The resin was shaken on a rotator for 16 h, drained, and rinsed with DMF (4 × 10 mL) and DCM (2 × 10 mL). The resin was then treated with 15 mL of 20% piperidine in DMF, shaken on a rotator for 1 h, then drained and rinsed with DMF (4 × 10 mL) and DCM (2 × 10 mL). The resin was treated with 10 mL of a HFIP / DCM (1 / 4) mixture and stirred with a nitrogen sparge for 1 hour. The liquid phase was collected and the resin was treated with 10 mL of HFIP / DCM (1 / 4) twice more for 1 hour each. The liquid phases were combined and the resin was washed with DCM (2 x 10 mL). The filtered liquid was combined and concentrated under vacuum. The residue was purified by flash chromatography on C18 eluting with CH3CN / H2O to give the title compound (330 mg, 39% yield) as a yellow solid. LCMS (ESI): [M+H] + =941.7. Step 3. N-[3-[[(2S,5R,8S,11S,14S,17S)-17-[(tert-butoxycarbonylamino)methyl]-7,8-dibutyl-14-cyclohexyl-11-isobutyl-5,10-dimethyl-3,9,12,15,18-pentaoxo-1,4,7,10,13,16-hexaazacyclooctadec-2-yl]methoxy]propyl]carbamate TIFF0007670618000166.tif54170

[0255] The title compound was prepared following a similar method as described in Example 7, Step 3, using the material from Step 2. LCMS (ESI): [M+H] + =923.7. Step 4. (3S,6S,9S,12S,15S,18R)-6-(aminomethyl)-3-(3-aminopropoxymethyl)-15,16-dibutyl-9-cyclohexyl-12-isobutyl-13,18-dimethyl-1,4,7,10,13,16-hexaazacyclooctadecane-2,5,8,11,14-pentone

[0256] The title compound was prepared as the trifluoroacetic acid salt using material from Step 3 similar to that described for Example 7, Step 4 and the following method: LCMS (ESI): [M+H] + =723.6, R t =2.19 minutes, method=J. 1 H NMR (400MHz, DMSO-d6): δ9.01-8.64(m,2H), 8.21-8.05(m,3H), 7.85-7.72(m,3H), 7.60 -7.22(m,2H), 4.82-4.72(m,1H), 4.44-4.31(m,2H), 4.09-3.92(m,2H), 3.49-3.32(m,10 H), 3.23-3.12(m,4H), 2.92-2.80(m,3H), 2.21-2.11(m,1H), 2.06-1.93(m,2H), 1.81-1. 73(m,3H), 1.70-1.55(m,8H), 1.42-1.26(m,6H), 1.19-1.10(m,5H), 1.07-0.83(m,15H). [Example 13] (3R,6S,9S,12S,15S,18R,19R)-6-(((1r,3S)-3-aminocyclobutoxy)methyl)-9-(aminomethyl)-19-((1R,3s,5S)-bicyclo[3.2.1]octan-3-ylmethyl)-12-cycloheptyl-3,16,18-trimethyl-15-propyl-1-oxa-4,7,10,13,16-pentaazacyclononadecane-5,8,11,14,17-pentaone (trifluoroacetate) JPEG0007670618000167.jpg62170 Step 1. N-Me-Norvaline-Cycloheptylglycine-Dap(Boc)-trans-tert-butoxycarbonyl)amino-cyclobutyl)-serine-(2-chlorotrityl resin) JPEG0007670618000168.jpg57170

[0257] The resin-bound tetrapeptide was prepared according to a method similar to that described in Example 7, step 1. Step 2. (2S,5S,8S,11S,14R,15R,18R)-18-amino-15-((1R,3s,5S)-bicyclo[3.2.1]octan-3-ylmethyl)-2-(((1r,3S)-3-((tert-butoxycarbonyl)amino)cyclobutoxy)methyl)-5-(((tert-butoxycarbonyl)amino)methyl)-8-cycloheptyl-12,14-dimethyl-4,7,10,13-tetraoxo-11-propyl-16-oxa-3,6,9,12-tetraazanonadecano-1-oic acid TIFF0007670618000169.tif69170

[0258] The resin-bound tetrapeptide from step 1 (600 mg, 0.5 mmol / g) was treated with THF (3 mL) and DIPEA (0.31 mL, 1.8 mmol) and allowed to swell for 30 min. Separately, a solution of (2R,3R)-4-[(1S,5R)-3-bicyclo[3.2.1]octanyl]-3-[(2R)-2-(9H-fluoren-9-ylmethoxycarbonylamino)propoxy]-2-methyl-butanoic acid (243 mg, 0.478 mmol, Intermediate 5) in THF (6.0 mL) was added to a solution of triphosgene (55.2 mg, 0.19 mmol) in THF (3.0 mL). 2,4,6-Trimethylpyridine (0.87 mL, 6.6 mmol) was added dropwise, causing the mixture to heat up and the formation of a colorless precipitate. The suspension was gently shaken for approximately 1 minute and then added to the pretreated resin. The mixture was placed on a rotator at room temperature for 3 hours, drained, and then rinsed with DMF (10 mL) and DCM (2 x 10 mL).

[0259] The resin was treated with 10 mL of 20% piperidine in DMF, agitated with a nitrogen stream for 1 hour, then drained and rinsed with DMF (2 x 10 mL) and DCM (2 x 10 mL). The resin was then treated with HFIP / DCM = 1 / 4 solution (20 mL), shaken on a rotator at room temperature for 1 hour, and then filtered. The liquid phase was collected. This process was repeated with HFIP / DCM = 1 / 4, and the liquid phase was collected. The resin was washed with DCM (3 x 10 mL). The liquid phase and the filtered liquid were combined and concentrated under vacuum. The residue was purified by reverse-phase flash chromatography on a C18 column to give the title compound (262 mg) as a yellow solid. LCMS (ESI): [M+H] + =992.7. Step 3. tert-Butyl N-[[(3R,6S,9S,12S,15S,18R,19R)-19-[[(1S,5R)-3-bicyclo[3.2.1]octanyl]methyl]-6-[[3-(tert-butoxycarbonylamino)cyclobutoxy]methyl]-12-cycloheptyl-3,16,18-trimethyl-5,8,11,14,17-pentaoxo-15-propyl-1-oxa-4,7,10,13,16-pentaazacyclononadequa-9-yl]methyl]carbamate TIFF0007670618000170.tif65170

[0260] The product from step 2 (240 mg, 0.242 mmol) was dissolved in THF (240 mL) containing DIPEA (0.17 mL, 0.97 mmol). The solution was cooled to 0° C. To this solution was added a premixed solution of HATU (138 mg, 0.36 mmol) and HOBt (49.1 mg, 0.36 mmol) in DMF (2 mL) and THF (8 mL) dropwise at 0° C. The mixture was stirred at 0° C. for 1 h. The mixture was concentrated in vacuo to remove THF. The residue was diluted with DMF (3 mL) and added dropwise to water (300 mL) with vigorous stirring. The resulting solid was collected by filtration, washed with water (2×10 mL), and dried to give the title compound (185 mg, crude) as a colorless solid. LCMS (ESI): [M+H] + =974.6. Step 4. (3R,6S,9S,12S,15S,18R,19R)-6-(((1r,3S)-3-aminocyclobutoxy)methyl)-9-(aminomethyl)-19-((1R,3s,5S)-bicyclo[3.2.1]octan-3-ylmethyl)-12-cycloheptyl-3,16,18-trimethyl-15-propyl-1-oxa-4,7,10,13,16-pentaazacyclononadecane-5,8,11,14,17-pentaone

[0261] The product from step 3 (175 mg, crude) was treated with trifluoroacetic acid (3.0 mL) and stirred at 0° C. for 2 hours. The reaction mixture was concentrated in vacuo. The residue was purified by preparative HPLC using the following conditions: Column: XSelect CSH Prep C18 OBD Column, 19*250 mm, 5 um; Mobile phase A: Water (0.05% TFA), Mobile phase B: ACN; Flow rate: 25 mL / min; Gradient: 20% B to 40% B in 20 min; 210 nm; Rt: 19.57 min to give the title compound as a TFA salt (46.1 mg) as a white solid. LCMS (ESI): [M+H] + =774.6, R t =1.97 minutes, method H. 1 H NMR (400MHz, DMSO-d6): δ8.52-8.12(m,2H), 8.03-7.93(m,6H), 7.65-7.33(m,1H), 4.56-4.15(m,4H), 3.84-3.60(m,4H), 3.28 -2.82(m,9H), 2.74(s,2H), 2.24-2.02(m,8H), 1.79-1.58(m,10H), 1.52-1.39(m,10H), 1.37-1.26(m,5H), 1.03-0.93(m,11H). [Example 14] (3R,6S,9S,12S,15S,18R,19R)-9-(aminomethyl)-6-[(3-aminopropylamino)methyl]-12-cycloheptyl-19-hexyl-3,16,18-trimethyl-15-propyl-1-oxa-4,7,10,13,16-pentaazacyclononadecane-5,8,11,14,17-pentone (trifluoroacetate) TIFF0007670618000171.tif63170 Step 1. Benzyl tert-butyl (((3R,6S,9S,12S,15S,18R,19R)-12-cycloheptyl-19-hexyl-3,16,18-trimethyl-5,8,11,14,17-pentaoxo-15-propyl-1-oxa-4,7,10,13,16-pentaazacyclononadecane-6,9-diyl)bis(methylene)) dicarbamate TIFF0007670618000172.tif55170

[0262] The title compound was prepared following a similar method to that described in Example 13, steps 1-3. Step 2. tert-Butyl N-[[(3R,6S,9S,12S,15S,18R,19R)-6-(aminomethyl)-12-cycloheptyl-19-hexyl-3,16,18-trimethyl-5,8,11,14,17-pentaoxo-15-propyl-1-oxa-4,7,10,13,16-pentaazacyclononadece-9-yl]methyl]carbamate TIFF0007670618000173.tif46170

[0263] The product of step 1 (905 mg, 1.01 mmol) and 10% Pd / C (450 mg) in isopropanol (100 mL) were stirred under hydrogen gas (1 atm) at room temperature for 16 hours. The catalyst was removed by filtration, and the filtered solution was concentrated under reduced pressure. The residue was purified by reverse-phase chromatography on a C18 column (0-46% acetonitrile / water with 0.05% TFA) to give the title compound (350 mg) as a white solid. LCMS (ESI): [M+H] + =766.6. Step 3. tert-Butyl N-[3-[[(3R,6S,9S,12S,15S,18R,19R)-9-[(tert-butoxycarbonylamino)methyl]-12-cycloheptyl-19-hexyl-3,16,18-trimethyl-5,8,11,14,17-pentaoxo-15-propyl-1-oxa-4,7,10,13,16-pentaazacyclononadece-6-yl]methylamino]propyl]carbamate TIFF0007670618000174.tif59170

[0264] To a solution of the product from step 2 (420 mg, 0.548 mmol) in DCM (400 mL) was added tert-butyl N-(3-oxopropyl)carbamate (95.7 mg, 0.55 mmol). The solution was stirred at room temperature for 2 hours, and then sodium triacetoxyborohydride (233.6 mg, 1.1 mmol) was added. The resulting mixture was stirred at room temperature for 2 hours and then washed with water and brine. The organic phase was dried over anhydrous sodium sulfate and concentrated in vacuo. The residue was purified by reverse-phase flash chromatography on a C18 column eluting with CH3CN / HO (45 / 55) to give the title compound (235 mg, 46.3% yield) as a white solid. LCMS (ESI): [M+H] + =923.6. Step 4. (3R,6S,9S,12S,15S,18R,19R)-9-(aminomethyl)-6-[(3-aminopropylamino)methyl]-12-cycloheptyl-19-hexyl-3,16,18-trimethyl-15-propyl-1-oxa-4,7,10,13,16-pentaazacyclononadecane-5,8,11,14,17-pentone

[0265] The product from step 3 (100.4 mg, 0.11 mmol) was treated with TFA (5.0 mL) at 0° C. for 30 minutes and then concentrated in vacuo. The residue was purified by preparative LCMS to give the title compound as its TFA salt (45.5 mg) as a white solid. LCMS (ESI): [M+H] + =723.6, R t =2.2 minutes, method=K. 1H NMR (400MHz, DMSO-d6): δ8.93-8.56(m,3H), 8.38-7.90(m,7H), 7.52-7.44(m,1H), 4.62-4.55(m,2H), 4.28-4.09(m,2H), 3.91-3.74(m,3H), 3.65 -3.48(m,4H), 3.23-3.03(m,5H), 2.98-2.79(m,6H), 2.72(s,1H), 2.13- 1.90(m,5H), 1.80-1.62(m,6H), 1.57-1.26(m,16H), 1.05-0.84(m,11H). [Example 15] (3S,6S,9S,12S,15S,17S)-6-(aminomethyl)-9-cyclohexyl-16-(cyclopentylmethyl)-3-((S)-1-hydroxyethyl)-12-isobutyl-13,15,17-trimethyl-1,4,7,10,13,16-hexaazacyclooctadecane-2,5,8,11,14-pentane (trifluoroacetate) TIFF0007670618000175.tif54170 Step 1. (2S,5S,8S,11S,14S,16S)-17-amino-2-((S)-1-(tert-butoxy)ethyl)-5-(((tert-butoxycarbonyl)amino)methyl)-8-cyclohexyl-15-(cyclopentylmethyl)-11-isobutyl-12,14,16-trimethyl-4,7,10,13-tetraoxo-3,6,9,12,15-pentaazaheptadecano-1-oic acid TIFF0007670618000176.tif68170

[0266] The title compound was prepared from N-Me-Leucine-Cyclohexylglycine-Dap(Boc)-Allothreonine(t-Bu)-(2-Chlorotrityl Resin) (Example 8, Step 1) and (2S)-2-[Cyclopentylmethyl-[(1S)-2-(9H-Fluoren-9-ylmethoxycarbonylamino)-1-methyl-ethyl]amino]propanoic acid (Intermediate T21) following a similar method as described in Example 12, Step 2. LCMS (ESI): [M+H] + =838.6. Step 2. tert-Butyl (((2S,5S,9S,11S,14S,17S)-5-((S)-1-(tert-butoxy)ethyl)-17-cyclohexyl-10-(cyclopentylmethyl)-14-isobutyl-9,11,13-trimethyl-3,6,12,15,18-pentaoxo-1,4,7,10,13,16-hexaazacyclooctadecan-2-yl)methyl)carbamate TIFF0007670618000177.tif60170

[0267] The title compound was prepared using the product of Step 1 and following a similar method as described in Example 12, Step 3. LCMS (ESI): [M+H] + =820.6. Step 3. (3S,6S,9S,12S,15S,17S)-6-(aminomethyl)-9-cyclohexyl-16-(cyclopentylmethyl)-3-((S)-1-hydroxyethyl)-12-isobutyl-13,15,17-trimethyl-1,4,7,10,13,16-hexaazacyclooctadecane-2,5,8,11,14-pentane

[0268] The product from step 2 (192 mg, 0.234 mmol) was treated with TFA (10 mL) at 0° C. for 1 hour. The mixture was evaporated under reduced pressure and the residue was purified by preparative LCMS: Column: XBridge Prep C18 OBD Column, 5 μm, 19*150 mm; Mobile phase A: Water (0.05% TFA), Mobile phase B: ACN; Flow rate: 25 mL / min; Gradient: 10% B to 35% B in 20 min; 210 nm; Rt: 14.86 min to give the title compound as its TFA salt (14.6 mg) as a white solid. LCMS (ESI): [M+H] + =664.5, R t =2.18 minutes, method=J. 1H NMR (300MHz, DMSO-d6): δ9.00-8.59(m,1H), 8.09-7.95(m,3H), 7.84-7.24(m,2H), 4.98-4.86(m,1H), 4.65-4.57(m,1H) , 4.06-3.99(m,3H), 3.91-3.65(m,1H), 3.21-2.69(m,11H), 2.35-2.12(m,4H), 1.81-1.47(m,14H), 1.36-0.88(m,20H). [Example 16] N-[[(2S,5R,8S,11S,14S,17S)-17-(aminomethyl)-7,8-dibutyl-14-cyclohexyl-5,10-dimethyl-3,9,12,15,18-pentaoxo-11-propyl-1,4,7,10,13,16-hexaazacyclooctadec-2-yl]methyl]-2-azaspiro[3.3]heptane-6-carboxamide (trifluoroacetate) TIFF0007670618000178.tif58170 Step 1. tert-Butyl N-[[(2S,5R,8S,11S,14S,17S)-5-(benzyloxycarbonylaminomethyl)-10,11-dibutyl-17-cyclohexyl-8,13-dimethyl-3,6,12,15,18-pentaoxo-14-propyl-1,4,7,10,13,16-hexaazacyclooctadec-2-yl]methyl]carbamate TIFF0007670618000179.tif59170

[0269] The title compound was prepared according to a similar method as described in Example 7, steps 1-4. LCMS (ESI): [M+H] + =885.5. Step 2. tert-Butyl (((2S,5R,8S,11S,14S,17S)-5-(aminomethyl)-10,11-dibutyl-17-cyclohexyl-8,13-dimethyl-3,6,12,15,18-pentaoxo-14-propyl-1,4,7,10,13,16-hexaazacyclooctadecan-2-yl)methyl)carbamate TIFF0007670618000180.tif52170

[0270] A mixture of the product from step 1 (181 mg, 0.205 mmol) and Pd / C (200 mg, 10% Pd on carbon) in ethyl acetate (17 mL) was stirred under a hydrogen atmosphere at 40° C. for 6 hours. The catalyst was removed by filtration and the filtrate was concentrated under reduced pressure. The crude product was used directly in the next step. LCMS (ESI): [M+H] + =751.5. Step 3. tert-Butyl 6-[[(2S,5R,8S,11S,14S,17S)-17-[(tert-butoxycarbonylamino)methyl]-7,8-dibutyl-14-cyclohexyl-5,10-dimethyl-3,9,12,15,18-pentaoxo-11-propyl-1,4,7,10,13,16-hexaazacyclooctadec-2-yl]methylcarbamoyl]-2-azaspiro[3.3]heptane-2-carboxylate TIFF0007670618000181.tif69170

[0271] tert-Butyl (((2S,5R,8S,11S,14S,17S)-5-(aminomethyl)-10,11-dibutyl-17-cyclohexyl-8,13-dimethyl-3,6,12,15,18-pentaoxo-14-propyl-1,4,7,10,13,16-hexaazacyclooctadecan-2-yl)methyl)carbamate (100 mg, 0.13 To a solution of 2-tert-butoxycarbonyl-2-azaspiro[3.3]heptane-6-carboxylic acid (52.7 mg, 0.220 mmol) and 2-tert-butoxycarbonyl-2-azaspiro[3.3]heptane-6-carboxylic acid (52.7 mg, 0.220 mmol) and DIPEA (65.8 mg, 0.510 mmol) was added dropwise a solution of HATU (77.5 mg, 0.204 mmol) and HOBt (21.4 mg, 0.159 mmol) in DMF (1.0 mL) at 0° C. The mixture was stirred at 0° C. for 1 h. Water (30 mL) was added with stirring. The resulting solid was collected and washed with water (3 times) to give the crude title compound (120.5 mg) as a yellow solid. LCMS (ESI): [M+H] + =974.6. Step 4. N-[[(2S,5R,8S,11S,14S,17S)-17-(aminomethyl)-7,8-dibutyl-14-cyclohexyl-5,10-dimethyl-3,9,12,15,18-pentaoxo-11-propyl-1,4,7,10,13,16-hexaazacyclooctadec-2-yl]methyl]-2-azaspiro[3.3]heptane-6-carboxamide

[0272] The product of step 3 (110 mg, 0.113 mmol) was treated with TFA (6.0 mL) at 0° C. for 1 hour and then concentrated in vacuo. The residue was purified by preparative HPLC to give the title compound (15.9 mg) as its TFA salt as a white solid. LCMS (ESI): [M+H] + =774.6, R t =2.05 minutes, method=J. 1 H NMR (400MHz, DMSO-d6): δ8.91-8.49(m,3H), 8.16-8.03(m,4H), 7.96-7.86(m,1H), 7.62 -6.96(m,2H), 4.79-4.70(m,1H), 4.58-4.42(m,2H), 4.31-4.15(m,2H), 4.02-3.87(m,8 H), 3.22-3.01(m,8H), 2.91-2.82(m,2H), 2.35-2.21(m,5H), 2.04-1.84(m,3H), 1.67-1 .54(m,7H), 1.47-1.23(m,9H), 1.15-1.06(m,4H), 1.02-0.93(m,8H), 0.90-0.83(m,4H). [Example 17 and Example 18] (3R,6S,9S,12S,15S,18R,19R)-9-(aminomethyl)-12-cyclohexyl-19-hexyl-15-isobutyl-3,16,18-trimethyl-6-(piperazin-1-ylmethyl)-1-oxa-4,7,10,13,16-pentaazacyclononadecane-5,8,11,14,17-pentaone (trifluoroacetate) and (3R,6S,9S,12S,15S,18R,19R)-9-(aminomethyl)-12-cyclohexyl-19-hexyl-15-isobutyl-3,16,18-trimethyl-6-(piperazin-1-ylmethyl)-1-oxa-4,7,10,13,16-pentaazacyclononadecane-5,8,11,14,17-pentaone (trifluoroacetate), a single unknown stereoisomer TIFF0007670618000182.tif62170 Step 1. Benzyl 4-(((3R,9S,12S,15S,18R,19R)-9-((((benzyloxy)carbonyl)amino)methyl)-12-cyclohexyl-19-hexyl-15-isobutyl-3,16,18-trimethyl-5,8,11,14,17-pentaoxo-1-oxa-4,7,10,13,16-pentaazacyclononadecan-6-yl)methyl)piperazine-1-carboxylate TIFF0007670618000183.tif74170

[0273] The title compound was prepared following procedures similar to those described for Examples 25 and 11. The two diastereomers were separated by flash chromatography on silica gel (12 g silica, solvent gradient: 0-10% methanol in DCM) to give two single unknown stereoisomers. Stereoisomer 1: Yield = 46.5 mg; LCMS (ESI): [M+H] + =1003.7 Stereoisomer 2: Yield = 41.2 mg; LCMS (ESI): [M+H] + =1003.8 Step 2. (3R,6S,9S,12S,15S,18R,19R)-9-(aminomethyl)-12-cyclohexyl-19-hexyl-15-isobutyl-3,16,18-trimethyl-6-(piperazin-1-ylmethyl)-1-oxa-4,7,10,13,16-pentaazacyclononadecane-5,8,11,14,17-pentaone (trifluoroacetate) and and (3R,6S,9S,12S,15S,18R,19R)-9-(aminomethyl)-12-cyclohexyl-19-hexyl-15-isobutyl-3,16,18-trimethyl-6-(piperazin-1-ylmethyl)-1-oxa-4,7,10,13,16-pentaazacyclononadecane-5,8,11,14,17-pentaone (trifluoroacetate)

[0274] The title compounds were each prepared as a single unknown stereoisomer following a procedure similar to that described for Example 11 using the isomers isolated in Step 1. [Example 17] LCMS (ESI):R T (min)=9.98, [M+H] + =735.6, method=B. 1 H NMR(400MHz,DMSO-d6)δ8.53(s,2H), 8.15(d,J=4.7Hz,1H), 7.98-7.81(m,4H), 7.68(d,J=8.2Hz,1H), 4.75 -4.60(m,1H), 4.28-4.14(m,1H), 3.98-3.76(m,3H), 3.15(s,4H), 3.10-2.57(m,13H), 1.90-0.74(m,41H). [Example 18] LCMS (ESI):R T (min)=10.12, [M+H] + =735.6, method=B; 1 H NMR (400MHz, DMSO-d6) δ8.63(d,J=93.7Hz,3H), 8.34-8.09(m,1H), 8.02-7.59(m,5H), 4.75-3.64(m,8H), 3.23-2.55(m,17H), 2.21-0.71(m,39H). [Example 19] (3R,6S,9S,12S,15S,18R,19R)-9-(aminomethyl)-6-((3-(((3-aminopropyl)amino)methyl)phenoxy)methyl)-12-cycloheptyl-19-hexyl-3,16,18-trimethyl-15-propyl-1-oxa-4,7,10,13,16-pentaazacyclononadecane-5,8,11,14,17-pentaone (trifluoroacetate) TIFF0007670618000184.tif65170Step 1. tert-Butyl N-[3-[[3-[(2S)-2-(benzyloxycarbonylamino)-3-[methoxy(methyl)amino]-3-oxo-propoxy]phenyl]methylamino]propyl]carbamate TIFF0007670618000185.tif41170

[0275] To a solution of (2S)-2-(benzyloxycarbonylamino)-3-hydroxy-propanoic acid (10.0 g, 41.9 mmol) in DMF (120 mL) were added HATU (23.9 g, 63.0 mmol), N,O-dimethylhydroxylamine hydrochloride (6.15 g, 63.0 mmol), and DIPEA (10.8 g, 83.9 mmol) at 0 °C. The reaction mixture was stirred at room temperature for 2 h. The system was diluted with ethyl acetate (500 mL) and washed successively with 0.5 M aqueous NaOH (3 × 250 mL) and brine (250 mL). The organic layer was dried over anhydrous sodium sulfate, concentrated under vacuum, and the residue was purified by chromatography on silica gel eluting with (EtOAc / PE=8 / 2) to give benzyl N-[(1S)-1-(hydroxymethyl)-2-[methoxy(methyl)amino]-2-oxo-ethyl]carbamate (11.0 g, 93.1% yield) as a pale yellow oil. LCMS (ESI): [M+H] + =283.1.

[0276] To a solution of benzyl N-[(1S)-1-(hydroxymethyl)-2-[methoxy(methyl)amino]-2-oxo-ethyl]carbamate (860 mg, 3.05 mmol) in toluene (10 mL) under nitrogen was added 3-bromobenzaldehyde (673 mg, 3.64 mmol), CsCO (1.99 g, 6.11 mmol), t-BuBrettphos (148 mg, 0.305 mmol), and [Pd(allyl)Cl] (55.5 mg, 0.152 mmol). The reaction mixture was stirred at 60 °C overnight and concentrated in vacuo. The residue was purified by flash chromatography on silica gel eluting with (EtOAc / PE=4 / 6) to give benzyl N-[(1S)-1-[(3-formylphenoxy)methyl]-2-[methoxy(methyl)amino]-2-oxo-ethyl]carbamate (140 mg, 11.9% yield) as a yellow oil. LCMS (ESI): [M+Na] + =409.1.

[0277] To a solution of tert-butyl (3-aminopropyl)carbamate (336 mg, 1.93 mmol) in DCM (12 mL) was added a solution of benzyl N-[(1S)-1-[(3-formylphenoxy)methyl]-2-[methoxy(methyl)amino]-2-oxo-ethyl]carbamate (670 mg, 1.73 mmol) in DCM (3.0 ml) at 0° C. The mixture was stirred at room temperature for 2 hours. Sodium triacetoxyborohydride (818 mg, 3.86 mmol) was added portionwise at 0° C. The mixture was stirred at room temperature for 1 hour and then concentrated under vacuum. The residue was loaded onto a silica gel column eluted with (DCM / MeOH=91 / 9) to give the title compound (810 mg, 77.2% yield) as a yellow oil. LCMS (ESI): [M+H] + =545.4. Step 2. tert-Butyl N-[[3-[(2S)-2-(benzyloxycarbonylamino)-3-[methoxy(methyl)amino]-3-oxo-propoxy]phenyl]methyl]-N-[3-(tert-butoxycarbonylamino)propyl]carbamate TIFF0007670618000186.tif45170

[0278] To a solution of tert-butyl N-[3-[[3-[(2S)-2-(benzyloxycarbonylamino)-3-[methoxy(methyl)amino]-3-oxo-propoxy]phenyl]methylamino]propyl]carbamate (710 mg, 1.3 mmol) and DIPEA (336.7 mg, 2.61 mmol) in DCM (20 mL) was added a solution of (Boc)2O (569.7 mg, 2.61 mmol) in DCM (5 mL) at 0° C. The reaction mixture was stirred at 25° C. for 1.5 h and then concentrated in vacuo. The residue was purified by flash chromatography on silica gel eluting with PE / EtOAc (2 / 1) to give the title compound (692 mg, 82.3% yield) as a yellow oil. LCMS (ESI): [M+H] + =645.3. Step 3. tert-Butyl N-[[3-[(2S)-2-amino-3-[methoxy(methyl)amino]-3-oxo-propoxy]phenyl]methyl]-N-[3-(tert-butoxycarbonylamino)propyl]carbamate TIFF0007670618000187.tif47170

[0279] A mixture of tert-butyl N-[[3-[(2S)-2-(benzyloxycarbonylamino)-3-[methoxy(methyl)amino]-3-oxo-propoxy]phenyl]methyl]-N-[3-(tert-butoxycarbonylamino)propyl]carbamate (588 mg, 0.909 mmol) and Pd (10 wt% on carbon, 600 mg) in EtOAc (120 mL) was stirred under a hydrogen atmosphere at 25° C. for 45 min. The catalyst was filtered off and the filtrate was concentrated in vacuo to give the title compound (460 mg) as a yellow oil. LCMS (ESI): [M+H] + =511.3. Step 4. (2S)-3-[3-[[tert-butoxycarbonyl-[3-(tert-butoxycarbonylamino)propyl]amino]methyl]phenoxy]-2-(9H-fluoren-9-ylmethoxycarbonylamino)propanoic acid TIFF0007670618000188.tif62170

[0280] To a solution of tert-butyl N-[[3-[(2S)-2-amino-3-[methoxy(methyl)amino]-3-oxo-propoxy]phenyl]methyl]-N-[3-(tert-butoxycarbonylamino)propyl]carbamate (460 mg, 0.90 mmol) in THF (24 mL) and water (8 mL) was added a solution of LiOH (108 mg, 4.5 mmol) in water (2 mL) at room temperature. The reaction mixture was stirred at 25°C for 3 days and then cooled to 0°C. 9-Fluorenylmethyl chloroformate (465.4 mg, 1.8 mmol) dissolved in 5 mL of THF was added dropwise at 0°C. The reaction mixture was stirred at 25°C for 0.5 hours. THF was evaporated under vacuum. The residue was diluted with water (2 mL) and the pH was adjusted to about 6 with 1 M aqueous HCl. The resulting solution was concentrated in vacuo and the residue was purified by flash chromatography on silica gel eluting with DCM / MeOH (10 / 1) to give the title compound (241.5 mg, 38.9% yield) as a white solid. LCMS (ESI): [M+H] + =690.2. 1 H NMR (300MHz, DMSO-d6): δ13.16(s,1H), 7.88(d,J=8.1Hz,2H), 7.73-7.62(m,3H), 7.44-7.41(m,2H), 7.33-7.20(m, 3H), 6.84-6.77(m,4H), 4.38-4.21(m,8H), 3.10-3.07(m,2H), 2.88-2.85(m,2H), 1.60-1.57(m,2H), 1.35(s,18H). Step 5. O-(3-(((tert-butoxycarbonyl)(3-((tert-butoxycarbonyl)amino)propyl)amino)methyl)phenyl)-N-((S)-3-((tert-butoxycarbonyl)amino)-2-((S)-2-cycloheptyl-2-((S)-2-(methylamino)pentanamido)acetamido)propanoyl)-L-serine-(2-chlorotrityl resin) JPEG0007670618000189.jpg57170

[0281] The resin-bound tetrapeptide was prepared following a procedure similar to that described in Example 7, step 1. Step 6. (3R,6S,9S,12S,15S,18R,19R)-9-(aminomethyl)-6-((3-(((3-aminopropyl)amino)methyl)phenoxy)methyl)-12-cycloheptyl-19-hexyl-3,16,18-trimethyl-15-propyl-1-oxa-4,7,10,13,16-pentaazacyclononadecane-5,8,11,14,17-pentaone

[0282] The title compound was prepared following a procedure similar to that described in Example 13, steps 2-4, using the resin-bound tetrapeptide from step 5. LCMS (ESI): [M+H] + =829.6, R t =2.78 minutes, method=J. 1 H NMR (400MHz, DMSO-d6): δ9.05-8.34(m,4H), 8.10-7.90(m,7H), 7.53-6.96(m,5H ), 4.62-4.58(m,1H), 4.39-4.37(m,1H), 4.28-4.23(m,1H), 4.14-4.05(m,2H), 3 .88-3.82(m,2H), 3.29-3.15(m,4H), 3.11(s,1H), 3.01-2.91(m,4H), 2.87-2.78 (m,4H), 2.70(s,2H), 2.07-1.91(m,4H), 1.79-1.23(m,25H), 1.04-0.83(m,12H). [Example 20] (3S,6S,9S,12S,15S,18R)-6-(aminomethyl)-9-cyclohexyl-18-(2-fluoroethyl)-16-hexyl-3-((S)-1-hydroxyethyl)-12-isobutyl-13,15-dimethyl-1,4,7,10,13,16-hexaazacyclooctadecane-2,5,8,11,14-pentane TIFF0007670618000190.tif48170Step 1. Benzyl (tert-butoxycarbonyl)-D-homoserinate TIFF0007670618000191.tif27170

[0283] A mixture of Boc(D)-homoserine (1.0 g, 4.3 mmol), benzyl bromide (1.1 g, 6.5 mmol), and potassium carbonate (0.8 g, 8.7 mmol) in DMF (5 mL) was stirred overnight. The reaction mixture was diluted with water and extracted with an IPAC / heptane mixture. The organic layer was washed with brine, dried over sodium sulfate, and concentrated. The residue was purified by flash chromatography (silica gel, 0-100% IPAC / heptane) to give 1.2 g (91%) of the title compound. Step 2. Benzyl (R)-2-((tert-butoxycarbonyl)amino)-4-fluorobutanoate TIFF0007670618000192.tif26170

[0284] To a solution of benzyl (tert-butoxycarbonyl)-D-homoserinate (1.2 g, 3.9 mmol) in anhydrous 1,4-dioxane (5 mL) was added pyridine-2-sulfonyl fluoride (0.7 g, 3.9 mmol) and 1,3,4,6,7,8-hexahydro-1-methyl-2H-pyrimido[1,2-a]pyrimidine (1.2 g, 7.8 mmol), and the mixture was stirred at room temperature overnight. The reaction mixture was diluted with water and extracted with an IPAC / heptane mixture. The organic layer was washed with brine, dried over sodium sulfate, and concentrated. The residue was purified by flash chromatography (silica gel, 0-50% IPAC / heptane) to give 0.87 g (72%) of the title compound. 1 H NMR (400MHz, DMSO-d6) δ7.47-7.32(m,5H), 5.29-4.94(m,2H), 4.50(dt,J=46.9,5.3Hz,2H), 4.24-4.01(m,1H), 2.25-1.83(m,2H), 1.38(s,9H). Step 3. tert-Butyl (R)-(4-fluoro-1-hydroxybutan-2-yl)carbamate TIFF0007670618000193.tif20170

[0285] To an ice-cold solution of benzyl (R)-2-((tert-butoxycarbonyl)amino)-4-fluorobutanoate (2.2 g, 7.1 mmol) was added lithium aluminum hydride (3.5 mL, 2 M in THF), and the mixture was stirred for 1 h. The reaction mixture was quenched with saturated aqueous ammonium chloride, and the solid was removed by filtration, washing thoroughly with IPAC. The filtered solution was dried over sodium sulfate and concentrated. The residue was purified by flash chromatography (silica gel, 0-5% MeOH / DCM) to afford 1.0 g (68%) of the title compound. H NMR (400 MHz, chloroform-d) δ 5.12-4.89 (m, 1H), 4.56 (dt, J = 47.2, 5.8 Hz, 2H), 3.87-3.59 (m, 3H), 2.77 (s, 1H), 2.09-1.77 (m, 2H), 1.45 (d, J = 4.2 Hz, 9H). Step 4. tert-Butyl (R)-(4-fluoro-1-oxobutan-2-yl)carbamate TIFF0007670618000194.tif21170

[0286] A mixture of tert-butyl (R)-(4-fluoro-1-hydroxybutan-2-yl)carbamate (0.5 g, 2.4 mmol), TEMPO (75 mg, 0.42 mmol), and iodobenzene diacetate (1.2 g, 3.6 mmol) in DCM (20 mL) was stirred at room temperature for 4 h. The reaction mixture was concentrated and triturated with 20% DCM / heptane. The solid was removed by filtration and washed thoroughly with 20% DCM / heptane. The filtered solution was concentrated, and the residue was purified by flash chromatography (silica gel, 0-5% MeOH / DCM) to give 0.35 g (70%) of the title compound. 1 H NMR (400 MHz, chloroform-d) δ 1 H NMR (400 MHz, chloroform-d) δ 9.61 (d, J = 2.1 Hz, 1H), 5.34 (s, 1H), 4.59 (dt, J = 47.0, 6.0 Hz, 2H), 4.31 (s, 1H), 2.49-2.08 (m, 2H), 1.46 (s, 9H). Step 5. Benzyl N-((R)-2-((tert-butoxycarbonyl)amino)-4-fluorobutyl)-N-hexyl-L-alaninate TIFF0007670618000195.tif35170

[0287] A mixture of tert-butyl (R)-(4-fluoro-1-oxobutan-2-yl)carbamate (0.25 g, 1.2 mmol) and benzyl (2S)-2-aminopropanoate hydrochloride (0.25 g, 1.2 mmol) in 3 mL of ethyl acetate was stirred for 1 hour. Triacetoxyborohydride (0.38 g, 1.8 mmol) was then added, and the mixture was stirred at room temperature overnight. To this mixture, hexanal (0.24 g, 2.4 mmol) and triacetoxyborohydride (0.5 g, 2.4 mmol) were added, and the mixture was heated at 50° C. overnight. The reaction mixture was cooled, diluted with IPAC, and stirred over aqueous sodium bicarbonate. The organic layer was separated, washed with brine, dried over sodium sulfate, and concentrated. The residue was purified by flash chromatography (silica gel, 0-50% IPAC / heptane) to give 0.2 g (30%) of the title compound. LCMS (ESI): [M+H] + =453.5. Step 6. Benzyl N-((R)-2-((((9H-fluoren-9-yl)methoxy)carbonyl)amino)-4-fluorobutyl)-N-hexyl-L-alaninate TIFF0007670618000196.tif46170

[0288] ((R)-2-((tert-butoxycarbonyl)amino)-4-fluorobutyl)-L-alaninate (0.4 g, 0.9 mmol) was dissolved in 2 mL of DCM. HCl (1 mL, 4N in dioxane) was added, and the reaction mixture was stirred for 1 h and then concentrated. The residue was dissolved in dioxane:water (1:1 6 mL), Fmoc-OSu (0.3 g, 0.9 mmol) and sodium bicarbonate (0.22 g, 2.7 mmol) were added, and the mixture was stirred for 1 h. The reaction mixture was diluted with IPAC, washed with water, brine, dried over sodium sulfate, and concentrated. The residue was purified by chromatography on silica gel (solvent gradient 0-50% IPAC / heptane) to give 0.46 g (90%) of the title compound. 1 H NMR (400MHz, DMSO-d6) δ7.89(d,J=7.6Hz,2H), 7.68(d,J=7.5Hz,2H), 7.48-7.26(m,9H), 7.04(dd,J=32.9,8.8Hz,1H), 5.16-5 .00(m,2H), 4.52-4.14(m,5H), 3.55(d,J=9.5Hz,2H), 2.60-2.27(m,2H), 2.05-1.44(m,2H), 1.37-1.04(m,13H), 0.80(m,3H). LCMS(ESI):[M+H] + =575.2. Step 7. N-((R)-2-((((9H-fluoren-9-yl)methoxy)carbonyl)amino)-4-fluorobutyl)-N-hexyl-L-alanine TIFF0007670618000197.tif44170

[0289] Benzyl (2S)-2-[[(2R)-2-(9H-fluoren-9-ylmethoxycarbonylamino)-4-fluoro-butyl]-hexylamino]propanoate (0.46 g, 0.8 mmol) was dissolved in ethanol (5 mL) and Pd (0.05 g, 10% on carbon) was added. The reaction mixture was purged with hydrogen and evacuated three times, then stirred under a hydrogen balloon for 30 minutes. The catalyst was removed by filtration and the filtered solution was concentrated to give the title compound (0.40 g): 1H NMR(400MHz,DMSO-d6D2O)δ7.88(t,J=6.7Hz,2H), 7.69(dd,J=7.5,2.3Hz,2H), 7.42(t,J=7.5Hz,2H), 7.37-7.29(m,2H), 7.04(d,J=8.9H) z,1H), 4.54-4.46(m,1H), 4.42-4.18(m,3H), 3.61(s,2H), 2.68-2.41(m,2H), 1.42-1.33(m,2H), 1.28-1.09(m,13H), 0.91-0.76(m,3H). LCMS(ESI):[M+H] + =485.4. Step 8. N-((S)-2-((S)-2-((S)-2-((S)-2-(((R)-2-amino-4-fluorobutyl)(hexyl)amino)-N-methylpropanamido)-4-methylpentanamido)-2-cyclohexylacetamido)-3-(((benzyloxy)carbonyl)amino)propanoyl)-O-benzyl-L-allothreonine TIFF0007670618000198.tif59170

[0290] To a filter tube was added Intermediate 15 (0.75 g, estimated 0.45 mmol / g) and 8 mL of DMF. The resin was allowed to swell at room temperature for 90 minutes and then drained. A solution of N-((R)-2-((((9H-Fluoren-9-yl)methoxy)carbonyl)amino)-4-fluorobutyl)-N-hexyl-L-alanine (0.33 g, 0.68 mmol) and PyAOP (0.36 mg, 0.68 mmol) in DMF (4 mL) was added to the resin, followed by DIPEA (0.35 mL, 2.0 mmol). The resulting mixture was placed on a rotator at room temperature for 16 hours. The resin was drained and rinsed with DMA (5x) and DCM (5x). 6 mL of 20% piperidine / DMF solution was added to the resin and stirred for 1 hour, then drained and rinsed with DCM (5x). The resin was then suspended in 6 mL of a 1:3 mixture of HFIP:DCM and stirred for 1 h. The mixture was then filtered and rinsed with DCM. The filtered solution was evaporated in vacuo and azeotroped with toluene (3 x 2 mL) to give 140 mg (43% crude yield) of crude peptide, which was carried on to the next step without further purification. LCMS (ESI): [M+H] + =940.9. Step 9. Benzyl (((2S,5S,8R,11S,14S,17S)-5-((S)-1-(benzyloxy)ethyl)-17-cyclohexyl-8-(2-fluoroethyl)-10-hexyl-14-isobutyl-11,13-dimethyl-3,6,12,15,18-pentaoxo-1,4,7,10,13,16-hexaazacyclooctadecan-2-yl)methyl)carbamate TIFF0007670618000199.tif63170

[0291] To a stirred solution of DIPEA (0.2 mL, 1 mmol) and HATU (0.026 mg, 0.43 mmol) in THF (10 mL) at room temperature was added a solution of N-((S)-2-((S)-2-((S)-2-((S)-2-(((R)-2-amino-4-fluorobutyl)(hexyl)amino)-N-methylpropanamido)-4-methylpentanamido)-2-cyclohexylacetamido)-3-(((benzyloxy)carbonyl)amino)propanoyl)-O-benzyl-L-allothreonine (0.14 g, 0.15 mmol) in THF (100 mL) dropwise over 4 hours and stirred overnight. The reaction mixture was evaporated in vacuo. The resulting residue was diluted with ethyl acetate, washed with water (2×) and brine, dried over sodium sulfate, filtered, and evaporated in vacuo. The crude product was purified by flash chromatography on silica gel (12 g silica, solvent gradient: 0-10% methanol in DCM) to give 60 mg of the title compound. LCMS (ESI): [M+H] + =922.9. Step 10. (3S,6S,9S,12S,15S,18R)-6-(aminomethyl)-9-cyclohexyl-18-(2-fluoroethyl)-16-hexyl-3-((S)-1-hydroxyethyl)-12-isobutyl-13,15-dimethyl-1,4,7,10,13,16-hexaazacyclooctadecane-2,5,8,11,14-pentane

[0292] A flask was charged with the above residue (60 mg, 0.065 mmol) and ethyl acetate (5 mL), and the flask was purged with nitrogen. After the addition of trifluoroacetic acid (25 μL, 0.323 mmol) and palladium (10 wt % on carbon) (50 mg), the reaction mixture was evacuated, refilled with hydrogen, and then stirred under a hydrogen balloon at room temperature for 18 hours. The reaction mixture was filtered through Celite, rinsed with 100 mL of 5% AcOH in methanol, and the filtered solution evaporated in vacuo and azeotroped with toluene (3 × 1 mL). The crude product was purified by reverse-phase HPLC and lyophilized to afford 19.6 mg (37%, 87% purity) of the title compound as its trifluoroacetate salt. LCMS (ESI):RT (min)=8.33, [M+H] + =698.5, method=B; 1 H NMR(400MHz,DMSO-d6)δ8.69-8.55(m,1H), 8.41-8.01(m,1H), 8.01-7.74(m,3H), 7.66-7.41(m,1H), 5.38-4.69(m,1H), 4.69-4. 18(m,4H), 4.18-3.44(m,7H), 3.25-3.03(m,4H), 3.05-2.85(m,1H), 2.16-1.89(m,2H), 1.89-1.36(m,11H), 1.36-0.77(m,25H). [Example 21] 4-(2-aminoethyl)-N-[[(3R,6S,9S,12S,15S,18R,19R)-9-(aminomethyl)-12-cycloheptyl-19-hexyl-3,16,18-trimethyl-5,8,11,14,17-pentaoxo-15-propyl-1-oxa-4,7,10,13,16-pentaazacyclononadecate-6-yl]methyl]benzamide (trifluoroacetate) TIFF0007670618000200.tif59170 Step 1. tert-Butyl N-[[(3R,6S,9S,12S,15S,18R,19R)-6-[[[4-[2-(tert-butoxycarbonylamino)ethyl]benzoyl]amino]methyl]-12-cycloheptyl-19-hexyl-3,16,18-trimethyl-5,8,11,14,17-pentaoxo-15-propyl-1-oxa-4,7,10,13,16-pentaazacyclononadequa-9-yl]methyl]carbamate TIFF0007670618000201.tif58170

[0293] To a solution of 4-(2-((tert-butoxycarbonyl)amino)ethyl)benzoic acid (82.4 mg, 0.31 mmol), HATU (117.8 mg, 0.31 mmol), HOBt (41.8 mg, 0.31 mmol), and DIPEA (0.11 mL, 0.63 mmol) in THF (120 mL) was added N-[[(3R,6S,9S,12S,15 A solution of (S,18R,19R)-6-(aminomethyl)-12-cycloheptyl-19-hexyl-3,16,18-trimethyl-5,8,11,14,17-pentaoxo-15-propyl-1-oxa-4,7,10,13,16-pentaazacyclononadece-9-yl]methyl]carbamate (121 mg, 0.158 mmol, Example 14, Step 2) was added at 0° C. The mixture was stirred for 1 hour and then concentrated in vacuo. The residue was purified by reverse-phase chromatography (acetonitrile 0-80 / 0.05% TFA in water) to afford the title compound (95.4 mg, 59.8% yield) as a white solid. LCMS (ESI): [M+H] + =1013.7. Step 2. 4-(2-aminoethyl)-N-[[(3R,6S,9S,12S,15S,18R,19R)-9-(aminomethyl)-12-cycloheptyl-19-hexyl-3,16,18-trimethyl-5,8,11,14,17-pentaoxo-15-propyl-1-oxa-4,7,10,13,16-pentaazacyclononadequa-6-yl]methyl]benzamide

[0294] The product of step 1 (95.2 mg, 0.090 mmol) was treated with TFA (5.0 mL) at 0° C. for 30 minutes and then concentrated in vacuo. The residue was purified by preparative LCMS to give the title compound as its TFA salt (25.5 mg) as a white solid. LCMS (ESI): [M+H] + =813.6, R t =2.89 minutes, method=K; 1H NMR (400MHz, DMSO-d6): δ8.85-8.12(m,2H), 7.99-7.90(m,3H), 7.88-7.72(m,5H) , 7.56-7.04(m,4H), 4.62-4.43(m,1H), 4.22-4.03(m,1H), 3.98-3.82(m,2H), 3.3 2-3.15(m,4H), 3.12-3.05(m,5H), 3.02-2.89(m,5H), 2.80(s,2H), 2.21-1.94(m, 3H), 1.79-1.52(m,8H), 1.48-1.25(m,17H), 1.03-0.90(m,7H), 0.88-0.82(m,4H). [Example 22] (3S,6S,9S,12S,15S,18R)-6-(aminomethyl)-3-((3-aminopropoxy)methyl)-15-butyl-9-cyclohexyl-16-(cyclopropylmethyl)-12-isobutyl-13,18-dimethyl-1,4,7,10,13,16-hexaazacyclooctadecane-2,5,8,11,14-pentane (trifluoroacetate) TIFF0007670618000202.tif55170Step 1. Benzyl (2S)-2-[[(2R)-2-(tert-butoxycarbonylamino)propyl]-(cyclopropylmethyl)amino]hexanoate TIFF0007670618000203.tif37170

[0295] A mixture of benzyl (2S)-2-aminohexanoate hydrochloride (1.0 g, 3.88 mmol) and tert-butyl (R)-(1-oxopropan-2-yl)carbamate (0.63 g, 3.5 mmol) in ethyl acetate (10 mL) was stirred for 2 hours, and then sodium triacetoxyborohydride (1.23 g, 5.8 mmol) was added, and the mixture was stirred at room temperature overnight. Cyclopropylcarboxaldehyde (0.81 g, 5.82 mmol) and sodium triacetoxyborohydride (1.0 g, 3.5 mmol) were then added to the mixture, and the mixture was stirred at room temperature for 24 hours. The reaction mixture was diluted with IPAC and stirred over sodium bicarbonate solution. The organic layer was separated, washed with brine, dried over sodium sulfate, and concentrated. The residue was purified by column chromatography (silica gel, 0-50% IPAC / heptane) to give 1.1 g (65%) of the title compound. LCMS (ESI): [M+H] + =433.3. 1 H NMR(400MHz,DMSO-d6)δ7.46-7.26(m,5H), 6.32(d,1H), 5.18-4.99(m,2H), 3.52(t, J=7.4Hz,2H), 2.60(dd,J=13.4,8.5Hz,1H), 2.45(dd,J=12.4,5.9Hz,1H), 2.38-2.17 (m,2H), 1.65-1.53(m,2H), 1.36(s,10H), 1.34-1.18(m,3H), 0.98(d,3H), 0.90-0.83 (m,3H), 0.82-0.73(m,1H), 0.48-0.39(m,1H), 0.38-0.29(m,1H), 0.12-0.02(m,2H). Step 2. (S)-2-(((R)-2-((((9H-Fluoren-9-yl)methoxy)carbonyl)amino)propyl)(cyclopropylmethyl)amino)hexanoic acid TIFF0007670618000204.tif41170

[0296] To a solution of benzyl (2S)-2-[[(2R)-2-(tert-butoxycarbonylamino)propyl]-(cyclopropylmethyl)amino]hexanoate (1.1 g, 2.5 mmol) in 4 mL of DCM, HCl (4 mL, 4 M in dioxane) was added, and the resulting mixture was stirred for 2 h and then concentrated. The residue was dissolved in a 1:1 mixture of dioxane:water, Fmoc-OSu (1.7 g, 4.65 mmol) and sodium carbonate (0.65 g, 5.00 mmol) were added, and the mixture was stirred at room temperature overnight. The reaction mixture was diluted with IPAC, and the organic layer was separated, washed with brine, dried over sodium sulfate, and concentrated. The residue was purified by chromatography (silica gel, 0-50% IPAC / heptane) to give the benzyl ester (1.4 g). LCMS (ESI) [M+H] + 556.8.

[0297] The residue was dissolved in ethanol (30 mL) with palladium (0.12 g, 10 wt % on carbon) and hydrogenated under a H balloon for 30 min. The catalyst was removed by filtering through Celite, and the filtered solution was concentrated. The residue was dried in a vacuum oven at 40 °C overnight to give the title compound (1.1 g). LCMS (ESI) [M+H] + =465.1; 1 H NMR(400MHz,DMSO-d6)δ7.96-7.85(m,2H), 7.74-7.66(m,2H), 7.41(m,2H) , 7.32(m,2H), 6.92(d,J=8.3Hz,1H), 4.35-4.12(m,3H), 3.61(d,J=14.4Hz ,1H), 3.36(q,J=8.8,7.4Hz,1H), 2.75-2.58(m,1H), 2.48-2.26(m,3H), 1. 66-1.00(m,10H), 0.90-0.76(m,4H), 0.54-0.28(m,2H), 0.17-0.01(m,2H). Step 3. (11S,14R,17S,20S,23S,26S)-11-amino-26-((((benzyloxy)carbonyl)amino)methyl)-17-butyl-23-cyclohexyl-16-(cyclopropylmethyl)-20-isobutyl-2,2,14,19-tetramethyl-4,12,18,21,24-pentaoxo-3,9-dioxa-5,13,16,19,22,25-hexaazaheptacosane-27-oic acid TIFF0007670618000205.tif49170

[0298] (S)-3-(((benzyloxy)carbonyl)amino)-2-((S)-2-cyclohexyl-2-((S)-4-methyl-2-(methylamino)pentanamido)acetamido)propanoic acid-(2-chlorophenyltrityl-resin) (0.6 g, 0.47 mmol / g, prepared using a method similar to that described in the synthesis of Intermediate 15) was swollen in DMF (3 mL) for 30 min and then treated with a premixed solution of (S)-2-(((R)-2-((((9H-fluoren-9-yl)methoxy)carbonyl)amino)propyl)(cyclopropylmethyl)amino)hexanoic acid (0.26 g, 0.56 mmol), PyAOP (0.3 g, 0.56 mmol), and DIPEA (0.15 mL, 0.85 mmol) in DMF (4 mL). The resin and solution were agitated for 20 hours, then drained and washed thoroughly with DMF and DCM. The resin was suspended in a 20% piperidine / DMF mixture (8 mL) and agitated for 1 hour. The resin was then drained and washed thoroughly with DMF and DCM. The resin was treated with a premixed solution of Intermediate 6 (0.26 g, 0.6 mmol), PyAOP (0.31 g, 0.6 mmol), and DIPEA (0.1 mL, 0.8 mmol) in DMF (4 mL) and agitated for 3 hours, then drained and washed thoroughly with DCM. The resin was treated with 20% piperidine / DMF (8 mL) and agitated for 1 hour. The liquid was removed by filtration, the resin was washed thoroughly with DCM, and suspended in a 1:3 mixture of HFIP and DCM (8 mL) and agitated for 1 hour. The liquid was collected by filtration while washing the resin with DCM. The combined filtered liquid was concentrated while azeotroping with toluene (3 times) to give the crude peptide (0.2 g). LCMS (ESI): [M+H] + =974.8. Step 4. tert-Butyl (3-(((2S,5R,8S,11S,14S,17S)-17-((((benzyloxy)carbonyl)amino)methyl)-8-butyl-14-cyclohexyl-7-(cyclopropylmethyl)-11-isobutyl-5,10-dimethyl-3,9,12,15,18-pentaoxo-1,4,7,10,13,16-hexaazacyclooctadecan-2-yl)methoxy)propyl)carbamate TIFF0007670618000206.tif65170

[0299] To a mixture of HATU (0.16 g, 0.4 mmol) and DIPEA (0.11 mL, 0.62 mmol), the crude peptide from step 3 (0.2 g, 0.2 mmol) in anhydrous THF (175 mL) was added dropwise over 3 h and then stirred overnight. The reaction mixture was concentrated, dissolved in IPAC, washed with water, brine, dried over sodium sulfate, and concentrated. The residue was purified by column chromatography (silica gel, 0-10% MeOH / DCM) to give the cyclic peptide (60 mg, 30%). LCMS (ESI): [M+H] + =955.8. Step 5. (3S,6S,9S,12S,15S,18R)-6-(aminomethyl)-3-((3-aminopropoxy)methyl)-15-butyl-9-cyclohexyl-16-(cyclopropylmethyl)-12-isobutyl-13,18-dimethyl-1,4,7,10,13,16-hexaazacyclooctadecane-2,5,8,11,14-pentane

[0300] The product of step 4 was dissolved in ethyl acetate (3 mL), Pd (10 wt% on carbon, 60 mg) and trifluoroacetic acid (0.2 μL) were added, the flask was evacuated, purged with hydrogen, and then stirred under a hydrogen balloon for 3 hours. The catalyst was removed by filtration and washed with 5% AcOH / methanol. The filtered solution was concentrated, and the resulting residue was treated with 1 mL of HCl (4N in dioxane) for 1 hour, then concentrated and azeotroped with toluene (3 times). The resulting residue was purified by reverse-phase HPLC and lyophilized to afford 10.3 mg (6%, 87% purity) of the title compound as its trifluoroacetic acid salt. LCMS (ESI):R T (min)=6.00, [M+H] + =721.5, method=B. 1H NMR(400MHz,DMSO-d6)δ9.01-8.61(m,2H), 8.00(t,J=32.4Hz,5H), 7.69(d ,J=23.8Hz,3H), 7.33(d,J=144.9Hz,2H), 4.89-4.57(m,2H), 4.57-3.87(m, 7H), 3.59(d,J=18.1Hz,6H), 3.28-3.01(m,3H), 2.83(s,2H), 2.05(d,J=50 .5Hz,2H), 1.91-1.41(m,11H), 1.41-0.60(m,25H), 0.43(d,J=41.0Hz,2H). [Example 23] 2-((3R,6S,9S,12S,15S,18R,19R)-9-(aminomethyl)-12-cycloheptyl-19-hexyl-3,16,18-trimethyl-5,8,11,14,17-pentaoxo-15-propyl-1-oxa-4,7,10,13,16-pentaazacyclononadecan-6-yl)acetamide (trifluoroacetate) TIFF0007670618000207.tif58170 Process 1. tert-Butyl-2-[(3R,6S,9S,12S,15S,18R,19R)-9-(benzyloxycarbonylaminomethyl)-12-cycloheptyl-19-hexyl-3,16,18-trimethyl-5,8,11,14,17-pentaoxo-15-propyl-1-oxa-4,7,10,13,16-pentaazacyclononadequa-6-yl]acetate TIFF0007670618000208.tif63170

[0301] The title compound was prepared following a similar method as described for Example 8, steps 1-3. LCMS (ESI): [M+H] + =885.6. Step 2. 2-((3R,6S,9S,12S,15S,18R,19R)-9-((((benzyloxy)carbonyl)amino)methyl)-12-cycloheptyl-19-hexyl-3,16,18-trimethyl-5,8,11,14,17-pentaoxo-15-propyl-1-oxa-4,7,10,13,16-pentaazacyclononadecan-6-yl)acetic acid TIFF0007670618000209.tif59170

[0302] The product from step 1 (1.04 g, 1.17 mmol) was treated with TFA (15 mL) at 0° C. for 1 h and then concentrated in vacuo. The residue was purified by reverse-phase chromatography (solvent gradient: 0-70% acetonitrile / 0.05% TFA in water) to give the title compound (403 mg, 41.4% yield). LCMS (ESI): [M+H] + =829.5. Step 3. Benzyl-N-[[(3R,6S,9S,12S,15S,18R,19R)-6-(2-amino-2-oxo-ethyl)-12- Cycloheptyl-19-hexyl-3,16,18-trimethyl-5,8,11,14,17-pentaoxo-15-propanol Pyr-1-oxa-4,7,10,13,16-pentaazacyclononadequa-9-yl]methyl]carbamate TIFF0007670618000210.tif52170

[0303] To a solution of 2-((3R,6S,9S,12S,15S,18R,19R)-9-(((benzyloxy)carbonyl)amino)methyl)-12-cycloheptyl-19-hexyl-3,16,18-trimethyl-5,8,11,14,17-pentaoxo-15-propyl-1-oxa-4,7,10,13,16-pentaazacyclononadecan-6-yl)acetic acid (101 mg, 0.120 mmol) in DMF (2.0 mL) was added HATU (92.0 mg, 0.240 mmol) and DIPEA (62.0 mg, 0.48 mmol). The reaction mixture was stirred for 5 minutes. To the mixture was added ammonia (1.0 mL, 0.5 M in THF) and ammonium chloride (13.0 mg, 0.24 mmol). The reaction mixture was stirred for 1 hour and then concentrated in vacuo. The residue was purified by reverse phase chromatography (solvent gradient: 0-68% acetonitrile / 0.05% TFA in water) to give the title compound (67.0 mg, 66.4% yield). LCMS (ESI): [M+H] + =828.6. Step 4. 2-[(3R,6S,9S,12S,15S,18R,19R)-9-(aminomethyl)-12-cycloheptyl-19-hexyl-3,16,18-trimethyl-5,8,11,14,17-pentaoxo-15-propyl-1-oxa-4,7,10,13,16-pentaazacyclononadequa-6-yl]acetamide

[0304] A mixture of benzyl-N-[[(3R,6S,9S,12S,15S,18R,19R)-6-(2-amino-2-oxo-ethyl)-12-cycloheptyl-19-hexyl-3,16,18-trimethyl-5,8,11,14,17-pentaoxo-15-propyl-1-oxa-4,7,10,13,16-pentaazacyclononadequa-9-yl]methyl]carbamate (67.0 mg, 0.080 mmol) and Pd / C (100 mg, 10% Pd on carbon) in 2-propanol (10 mL) was stirred under a hydrogen atmosphere at room temperature for 2.5 hours. The catalyst was removed by filtration, and the filtered solution was evaporated in vacuo. The residue was purified by preparative HPLC and lyophilized to give the title compound as its TFA salt (12.3 mg) as a white solid. LCMS (ESI): [M+H] + =694.5, R t =2.43 minutes, method=H. 1 H NMR (400MHz, DMSO-d6): δ8.62-8.18(m,2H), 8.02-7.92(m,4H), 7.48-7.21(m,2H ), 6.93-6.86(m,1H), 4.62-4.41(m,1H), 4.32-4.22(m,1H), 4.18-4.10(m,1H), 3 .86-3.78(m,1H), 3.30-3.22(m,3H), 3.10(s,2H), 3.02-2.91(m,2H), 2.71(s,1H) ), 2.21-2.02(m,2H), 1.76-1.52(m,8H), 1.48-1.22(m,20H), 1.05-0.85(m,13H). [Example 24] (3R,6S,9S,12S,15S,18R,19R)-9-(aminomethyl)-6-((3-((3-aminopropyl)amino)propoxy)methyl)-19-((1R,3s,5S)-bicyclo[3.2.1]octan-3-ylmethyl)-12-cycloheptyl-3,16,18-trimethyl-15-propyl-1-oxa-4,7,10,13,16-pentaazacyclononadecane-5,8,11,14,17-pentaone (trifluoroacetate) TIFF0007670618000211.tif63170 Step 1. N-Me-Norvaline-Cycloheptylglycine-Dap(Cbz)-3-((tert-butoxycarbonyl)amino)propyl)-serine-(2-chlorotrityl resin) JPEG0007670618000212.jpg61170

[0305] The resin-bound tetrapeptide was prepared according to a method similar to that described in Example 7, step 1. Step 2. tert-Butyl N-[3-[[(3R,6S,9S,12S,15S,18R,19R)-9-(benzyloxycarbonylaminomethyl)-19-[[(1S,5R)-3-bicyclo[3.2.1]octanyl]methyl]-12-cycloheptyl-3,16,18-trimethyl-5,8,11,14,17-pentaoxo-15-propyl-1-oxa-4,7,10,13,16-pentaazacyclononadece-6-yl]methoxy]propyl]carbamate TIFF0007670618000213.tif63170

[0306] The title compound was prepared using the resin-bound tetrapeptide from step 1 and (2R,3R)-4-[(1S,5R)-3-bicyclo[3.2.1]octanyl]-3-[(2R)-2-(9H-fluoren-9-ylmethoxycarbonylamino)propoxy]-2-methyl-butanoic acid (Intermediate 5) according to the following method similar to that described in Example 13. LCMS (ESI): [M+H] + =996.6. Step 3. Benzyl N-[[(3R,6S,9S,12S,15S,18R,19R)-6-(3-aminopropoxymethyl)-19-[[(1S,5R)-3-bicyclo[3.2.1]octanyl]methyl]-12-cycloheptyl-3,16,18-trimethyl-5,8,11,14,17-pentaoxo-15-propyl-1-oxa-4,7,10,13,16-pentaazacyclononadequa-9-yl]methyl]carbamate TIFF0007670618000214.tif59170

[0307] The product of step 2 (430 mg, 0.43 mmol) was treated with TFA (12 mL) at 0° C. for 1 hour. Toluene (10 mL) was added and the resulting solution was concentrated in vacuo. The residue was partitioned between 100 mL of DCM and 5% Na2CO3 solution (20 mL). The organic phase was washed with brine (20 mL), dried over anhydrous sodium sulfate, and concentrated in vacuo to give the title compound (385 mg) as a pale yellow solid. LCMS (ESI): [M+H] + =896.6. Step 4. tert-Butyl N-[3-[3-[[(3R,6S,9S,12S,15S,18R,19R)-9-(benzyloxycarbonylaminomethyl)-19-[[(1S,5R)-3-bicyclo[3.2.1]octanyl]methyl]-12-cycloheptyl-3,16,18-trimethyl-5,8,11,14,17-pentaoxo-15-propyl-1-oxa-4,7,10,13,16-pentaazacyclononadecu-6-yl]methoxy]propylamino]propyl]carbamate TIFF0007670618000215.tif64170

[0308] The product of step 3 (385 mg, 0.430 mmol) and tert-butyl N-(3-oxopropyl)carbamate (66.9 mg, 0.39 mmol) in DCM (30 mL) were stirred at room temperature for 2 hours. The mixture was cooled to 0°C, and sodium triacetoxyborohydride (137 mg, 0.646 mmol) was added at this temperature. The reaction mixture was allowed to warm to room temperature and then stirred at room temperature for 3 hours. The reaction was quenched with 5% aqueous NH4Cl (30 mL), and the phases were separated. The aqueous phase was extracted with DCM (100 mL). The combined organic layers were washed with brine (30 mL), dried over anhydrous sodium sulfate, and concentrated under vacuum. The residue was purified by reverse-phase flash chromatography on a C18 column to give the title compound (194 mg, 42.9% yield) as a white solid. LCMS (ESI): [M+H] + =1053.7. Step 5. tert-Butyl N-[3-[3-[[(3R,6S,9S,12S,15S,18R,19R)-9-(aminomethyl)-19-[[(1S,5R)-3-bicyclo[3.2.1]octanyl]methyl]-12-cycloheptyl-3,16,18-trimethyl-5,8,11,14,17-pentaoxo-15-propyl-1-oxa-4,7,10,13,16-pentaazacyclononadequa-6-yl]methoxy]propylamino]propyl]carbamate TIFF0007670618000216.tif64170

[0309] To a solution of N-[3-[3-[[(3R,6S,9S,12S,15S,18R,19R)-9-(benzyloxycarbonylaminomethyl)-19-[[(1S,5R)-3-bicyclo[3.2.1]octanyl]methyl]-12-cycloheptyl-3,16,18-trimethyl-5,8,11,14,17-pentaoxo-15-propyl-1-oxa-4,7,10,13,16-pentaazacyclononadequa-6-yl]methoxy]propylamino]propyl]carbamate (194 mg, 0.184 mmol) in isopropanol (30 mL) was added palladium (10 wt% on carbon, 194 mg). The mixture was stirred under a hydrogen atmosphere at room temperature for 2.5 hours. The catalyst was removed by filtration and the filtered solution was concentrated in vacuo to give the title compound (159 mg, 93.9% yield) as a white solid, which was used in the next step without further purification. LCMS (ESI): [M+H] + =919.7. Step 6. (3R,6S,9S,12S,15S,18R,19R)-9-(aminomethyl)-6-((3-((3-aminopropyl)amino)propoxy)methyl)-19-((1R,3s,5S)-bicyclo[3.2.1]octan-3-ylmethyl)-12-cycloheptyl-3,16,18-trimethyl-15-propyl-1-oxa-4,7,10,13,16-pentaazacyclononadecane-5,8,11,14,17-pentaone

[0310] The product of step 5 (159 mg, 0.173 mmol) was treated with TFA (10 mL) at 0° C. for 1 hour. Toluene (10 mL) was added, and the mixture was concentrated under vacuum. The residue was purified by preparative HPLC. Column: XSelect CSH Prep C18 OBD Column, 5 μm, 19*150 mm; Mobile phase A: Water (0.05% TFA), Mobile phase B: MeOH-----preparative; Flow rate: 25 mL / min; Gradient: 37% B to 67% B in 20 min; 254 / 220 nm; Rt: 17.09 min to give the title compound as its TFA salt (65 mg) as a white solid. LCMS (ESI): [M+H] + =819.6, R t=2.64 minutes, method=I. 1 H NMR (400MHz, DMSO-d6): δ8.72-8.42(m,3H), 8.20-8.02(m,5H), 8.00-7.82(m,3H), 7.56-7.44(m,1H), 4.58-4.45(m,1H), 4.38-4.11(m,2H) ), 3.89-3.65(m,4H), 3.38-3.12(m,4H), 3.09-2.72(m,11H), 2.22-1.99(m,4H), 1.89-1.52(m,15H), 1.49-1.21(m,16H), 1.09-0.87(12H). [Example 25] (3R,9S,12S,15S,18R,19R)-9-(aminomethyl)-6-((3-(aminomethyl)azetidin-1-yl)methyl)-12-cyclohexyl-19-hexyl-15-isobutyl-3,16,18-trimethyl-1-oxa-4,7,10,13,16-pentaazacyclononadecane-5,8,11,14,17-pentaone (trifluoroacetate) TIFF0007670618000217.tif60170Step 1. Methyl 2-(((benzyloxy)carbonyl)amino)-3-(3-(((tert-butoxycarbonyl)amino)methyl)azetidin-1-yl)propanoate TIFF0007670618000218.tif37170

[0311] To a mixture of methyl (2S)-2-(benzyloxycarbonylamino)-3-hydroxypropanoate (2.5191 g, 9.9471 mmol) and triethylamine (3.0 mL, 21 mmol) in DCM (20 mL) at 0 °C was added methanesulfonyl chloride (0.90 mL, 12 mmol). After stirring at room temperature for 2 hours, tert-butyl (azetidin-3-ylmethyl)carbamate hydrochloride (2.5107 g, 10.935 mmol) and triethylamine (4.0 mL, 28 mmol) were added. The reaction mixture was stirred at room temperature for 16 hours. The reaction mixture was washed with saturated aqueous NaHCO3, and the aqueous layer was extracted with an additional 100 mL of DCM. The combined organic extracts were dried over MgSO4, filtered, and evaporated in vacuo. The crude product was purified by flash chromatography on silica gel (120 g silica, solvent gradient: 0-10% MeOH in DCM). The resulting thick gum was dried under vacuum overnight to give 3.4092 g (81%) of the title compound. LCMS (ESI): [M+H] + =422.25; 1 H NMR(400MHz,DMSO-d6)δ7.53(d,J=7.8Hz,1H), 7.40-7.25(m,5H), 6.84(d,J=6.3Hz,1H), 5.03(s,2H), 4.02-3.93(m,1H), 3.62( s,3H), 3.17(q,J=7.7Hz,2H), 3.03(t,J=6.3Hz,2H), 2.80(q,J=6.7Hz,2H), 2.70-2.55(m,2H), 2.42-2.33(m,1H), 1.37(s,9H). Step 2. 2-((((9H-Fluoren-9-yl)methoxy)carbonyl)amino)-3-(3-(((tert-butoxycarbonyl)amino)methyl)azetidin-1-yl)propanoic acid TIFF0007670618000219.tif42170

[0312] To a solution of methyl 2-(((benzyloxy)carbonyl)amino)-3-(3-(((tert-butoxycarbonyl)amino)methyl)azetidin-1-yl)propanoate (3.4092 g, 8.088 mmol) in 1,4-dioxane (20 mL) was added lithium hydroxide (1.0 M in water, 10.0 mL, 10 mmol). The reaction mixture was stirred at room temperature for 6 hours. The reaction mixture was poured into 100 mL of DCM and washed with 10% aqueous citric acid. The aqueous portion was extracted with an additional 3×100 mL of DCM, and the combined organic extracts were dried over magnesium sulfate, filtered, evaporated in vacuo, and dried under vacuum overnight. The resulting material was dissolved in ethanol (20 mL). Under a nitrogen atmosphere, palladium (10 wt % on carbon) (271.7 mg, 0.2554 mmol) was added. The reaction mixture was then stirred under a hydrogen balloon at room temperature for 4 hours, then filtered through Celite, rinsed with 200 mL of ethanol, and evaporated in vacuo. The resulting material was dissolved in water (15 mL) and 1,4-dioxane (15 mL) to which sodium bicarbonate (2.066 g, 24.5 mmol) and Fmoc-OSu (3.274 g, 9.706 mmol) were added. The reaction mixture was stirred at room temperature for 2 hours, then diluted with 100 mL of DCM and washed with 10% aqueous citric acid. The aqueous portion was extracted with an additional 100 mL of DCM, and the combined organic extracts were dried over magnesium sulfate, filtered, and evaporated in vacuo. The crude product was purified by flash chromatography on silica gel (120 g silica, solvent gradient: 0-15% methanol in DCM) to afford 2.3477 g (59%) of the title compound. LCMS (ESI): [M+H] + =496.25. Step 3. (3R,9S,12S,15S,18R,19R)-9-(aminomethyl)-6-((3-(aminomethyl)azetidin-1-yl)methyl)-12-cyclohexyl-19-hexyl-15-isobutyl-3,16,18-trimethyl-1-oxa-4,7,10,13,16-pentaazacyclononadecane-5,8,11,14,17-pentaone

[0313] The title compound (mixture of two diastereomers) was prepared following a procedure similar to that described for Example 11 using 2-((((9H-fluoren-9-yl)methoxy)carbonyl)amino)-3-(3-(((tert-butoxycarbonyl)amino)methyl)azetidin-1-yl)propanoic acid. LCMS (ESI):R T (min) = 8.55 and 8.70, [M+H] + =735.6, method=B; 1 H NMR(400MHz,DMSO-d6)δ8.80-8.51(m,1H), 8.38-8.17(m,1H), 8.17-7.91(m,3H), 7.91-7.56(m,4H) , 4.73-3.64(m,8H), 3.02(d,J=104.9Hz,10H), 2.73(s,1H), 2.09-1.45(m,10H), 1.45-0.73(m,33H). [Example 26] (2S)-2,3-Diamino-N-[[(3R,6S,9S,12S,15S,18R,19R)-9-(aminomethyl)-12-cyclohexyl-19-hexyl-15-isobutyl-3,16,18-trimethyl-5,8,11,14,17-pentaoxo-1-oxa-4,7,10,13,16-pentaazacyclononadequa-6-yl]methyl]propanamide (trifluoroacetate) TIFF0007670618000220.tif69170Step 1. (2S)-2-[[(2S)-2-[[(2S)-2-[[(2S)-2-[[(2R,3R)-3-[(2R)-2-aminopropoxy]-2-methyl-nonanoyl]-methyl-amino]-4-methyl-pentanoyl]amino]-2-cyclohexyl-acetyl]amino]-3-(benzyloxycarbonylamino)propanoyl]amino]-3-(tert-butoxycarbonylamino)propanoic acid TIFF0007670618000221.tif57170

[0314] The title compound was prepared following a procedure similar to that described for Example 11, Step 1, using an appropriately protected amino acid. LCMS (ESI): [M+H] + =918.5. Step 2. tert-Butyl N-[[(3R,6S,9S,12S,15S,18R,19R)-9-(benzyloxycarbonylaminomethyl)-12-cyclohexyl-19-hexyl-15-isobutyl-3,16,18-trimethyl-5,8,11,14,17-pentaoxo-1-oxa-4,7,10,13,16-pentaazacyclononadequa-6-yl]methyl]carbamate TIFF0007670618000222.tif65170

[0315] (2S)-2-[[(2S)-2-[[(2S)-2-[[(2S)-2-[[(2R,3R)-3-[(2R)-2-aminopropoxy]-2-methyl-nonanoyl]-methyl-amino]-4-methyl-pentanoyl]amino]-2-cyclohexyl-acetyl]amino]-3-(benzyloxycarbonylamino)propanoyl]amino]-3-(tert-butoxycarbonylamino)propanoic acid (1.2 g, 1.3 mmol) was dissolved in 1.2 L of THF and DIPEA (0.69 mL, 4 mmol) was added. HATU (1.1 g, 2.6 mmol) was dissolved in 6 mL of DMF and added to the mixture. The mixture was then stirred overnight at room temperature and evaporated in vacuo. The residue was then diluted with ethyl acetate and water, vigorously shaken, and partitioned. The organic portion was then concentrated in vacuo and the residue purified by silica gel flash chromatography (125 g silica, 0-10% MeOH in DCM) to give the title compound (650 mg, 0.72 mmol, 55% yield). LCMS (ESI): [M+H] + =900.5. Step 3. Benzyl N-[[(3R,6S,9S,12S,15S,18R,19R)-6-(aminomethyl)-12-cyclohexyl-19-hexyl-15-isobutyl-3,16,18-trimethyl-5,8,11,14,17-pentaoxo-1-oxa-4,7,10,13,16-pentaazacyclononadequa-9-yl]methyl]carbamate TIFF0007670618000223.tif48170

[0316] tert-Butyl N-[[(3R,6S,9S,12S,15S,18R,19R)-9-(benzyloxycarbonylaminomethyl)-12-cyclohexyl-19-hexyl-15-isobutyl-3,16,18-trimethyl-5,8,11,14,17-pentaoxo-1-oxa-4,7,10,13,16-pentaazacyclononadequa-6-yl]methyl]carbamate (600 mg, 0.66 mmol) was dissolved in 4 mL of DCM, and 1.6 mL of triisopropylsilane and 0.8 mL of TFA were added. The mixture was then stirred at room temperature for 1 hour, diluted with toluene, and concentrated in vacuo. The residue was then diluted with EtOAc, charged with 0.10 mL of TEA, and diluted with water. The mixture was then vigorously shaken and partitioned. The organics were then concentrated in vacuo to give the title compound (540 mg, 0.66 mmol, 100% yield). LCMS (ESI): [M+H] + =800.5. Step 4. tert-Butyl N-[(1S)-2-[[(3R,6S,9S,12S,15S,18R,19R)-9-(benzyloxycarbonylaminomethyl)-12-cyclohexyl-19-hexyl-15-isobutyl-3,16,18-trimethyl-5,8,11,14,17-pentaoxo-1-oxa-4,7,10,13,16-pentaazacyclononadece-6-yl]methylamino]-1-[(tert-butoxycarbonylamino)methyl]-2-oxo-ethyl]carbamate TIFF0007670618000224.tif69170

[0317] (2S)-2,3-bis(tert-butoxycarbonylamino)propanoic acid (190 mg, 0.63 mmol) was dissolved in 2 mL of DMF and charged with HATU (243 mg, 0.63 mmol) and TEA (174 uL, 1.25 mmol). The mixture was then stirred at room temperature for 5 minutes and added to benzyl N-[[(3R,6S,9S,12S,15S,18R,19R)-6-(aminomethyl)-12-cyclohexyl-19-hexyl-15-isobutyl-3,16,18-trimethyl-5,8,11,14,17-pentaoxo-1-oxa-4,7,10,13,16-pentaazacyclononadequa-9-yl]methyl]carbamate (290 mg, 0.36 mmol) dissolved in 0.30 mL of DMF. The mixture was then stirred at room temperature for 30 minutes. 1 mL of saturated aqueous NaHCO3 was then added to the mixture and stirred at room temperature for 5 minutes. The mixture was then diluted with EtOAc and water, shaken vigorously, and partitioned. The organic portion was then washed twice with water and concentrated in vacuo to give the title compound (400 mg, 0.36 mmol, 100% yield). LCMS (ESI): [M+H] + =1086.7. Step 5. (2S)-2,3-Diamino-N-[[(3R,6S,9S,12S,15S,18R,19R)-9-(aminomethyl)-12-cyclohexyl-19-hexyl-15-isobutyl-3,16,18-trimethyl-5,8,11,14,17-pentaoxo-1-oxa-4,7,10,13,16-pentaazacyclononadequa-6-yl]methyl]propanamide (trifluoroacetate)

[0318] tert-Butyl N-[(1S)-2-[[(3R,6S,9S,12S,15S,18R,19R)-9-(benzyloxycarbonylaminomethyl)-12-cyclohexyl-19-hexyl-15-isobutyl-3,16,18-trimethyl-5,8,11,14,17-pentaoxo-1-oxa-4,7,10,13,16-pentaazacyclononadequa-6-yl]methylamino]-1-[(tert-butoxycarbonylamino)methyl]-2-oxo-ethyl]carbamate (400 mg, 0.36 mmol) was dissolved in 40 mL of 1:1 isopropanol:EtOAc and subjected to hydrogenation via an HCube apparatus (10% Pd / C, 50 °C, 75 psi). The mixture was then concentrated in vacuo.

[0319] The residue was then dissolved in 4 mL of DCM and charged with 1 mL of triisopropylsilane and 0.80 mL of TFA. The mixture was then stirred at room temperature for 1 hour, concentrated in vacuo, and azeotroped twice with dioxane. The residue was then purified by reverse-phase HPLC to give the title compound (88 mg, 0.11 mmol, 36% yield). LCMS (ESI):R T (min)=9.11, [M+H] + =753.8, method=B. 1 H NMR(400MHz,DMSO-d6)δ8.67(s,1H), 8.44(s,1H), 8.35(s,1H), 8.22(d,J=5.7Hz,1H), 8.11(s,1H) , 8.02(s,1H), 7.96(d,J=4.9Hz,1H), 7.69(d,J=8.3Hz,1H), 7.54(d,J=8.1Hz,1H), 4.66(t,J=7.7H z,1H), 4.25(s,1H), 4.05(s,1H), 3.85(s,2H), 3.72(s,1H), 3.20(s,1H), 3.11(d,J=6.2Hz,1H), 2. 98-2.70(m,5H), 1.87(s,2H), 1.65-1.58(m,14H), 1.27(s,13H), 1.11(s,1H), 1.06-0.81(m,17H). [Example 27] 3-(aminomethyl)-N-[[(3R,6S,9S,12S,15S,18R,19R)-9-(aminomethyl)-12-cycloheptyl-19-hexyl-3,16,18-trimethyl-5,8,11,14,17-pentaoxo-15-propyl-1-oxa-4,7,10,13,16-pentaazacyclononadecate-6-yl]methyl]azetidine-1-carboxamide (trifluoroacetate) TIFF0007670618000225.tif62170 Step 1. tert-Butyl N-[[(3R,6S,9S,12S,15S,18R,19R)-6-[[[3-[(tert-butoxycarbonylamino)methyl]azetidine-1-carbonyl]amino]methyl]-12-cycloheptyl-19-hexyl-3,16,18-trimethyl-5,8,11,14,17-pentaoxo-15-propyl-1-oxa-4,7,10,13,16-pentaazacyclononadequa-9-yl]methyl]carbamate TIFF0007670618000226.tif59170

[0320] A solution of N-[[(3R,6S,9S,12S,15S,18R,19R)-6-(aminomethyl)-12-cycloheptyl-19-hexyl-3,16,18-trimethyl-5,8,11,14,17-pentaoxo-15-propyl-1-oxa-4,7,10,13,16-pentaazacyclononadequa-9-yl]methyl]carbamate (70.1 mg, 0.090 mmol, Example 14, Step 2) and 1,1'-carbonyldiimidazole (29.7 mg, 0.18 mmol) in THF (5 mL) was stirred at room temperature for 2 hours. tert-Butyl (azetidin-3-ylmethyl)carbamate (34.4 mg, 0.18 mmol) was then added. The resulting mixture was stirred at room temperature for 2 hours and then concentrated in vacuo. The residue was purified by reverse-phase flash chromatography on a C18 column eluting with 0-53% acetonitrile / 0.1% TFA in water to give the title compound (65.4 mg, 73.1% yield) as a white solid. LCMS (ESI): [M+H] + =978.8. Step 2. 3-(aminomethyl)-N-[[(3R,6S,9S,12S,15S,18R,19R)-9-(aminomethyl)-12-cycloheptyl-19-hexyl-3,16,18-trimethyl-5,8,11,14,17-pentaoxo-15-propyl-1-oxa-4,7,10,13,16-pentaazacyclononadecc-6-yl]methyl]azetidine-1-carboxamide

[0321] The product of step 1 (93.2 mg, 0.10 mmol) was treated with TFA (3 mL) with stirring at 0° C. for 30 minutes. The mixture was concentrated in vacuo. The residue was purified by preparative LCMS to give the title compound as its TFA salt (28.4 mg) as a white solid. LCMS (ESI): [M+H] + =778.6, R t =2.7 minutes, method=I. 1 H NMR (400MHz, DMSO-d6): δ8.75-8.55(m,1H), 8.21-7.75(m,8H), 7.45-7.26(m,1H), 4.65-4.17(m,3H), 3.93-3.85(m,5H), 3.61-3.44(m,4H) ), 3.39-3.35(m,3H), 3.15-3.03(m,4H), 2.93-2.82(m,3H), 2.19-1.98(m,2H), 1.86-1.65(m,8H), 1.55-1.25(m,18H), 1.00-0.88(m,12H). [Example 28] (3S,6S,9S,12S,15S)-6-(aminomethyl)-9-cyclohexyl-16-hexyl-3-(1-hydroxy-2-(piperazin-1-yl)ethyl)-12-isobutyl-13,15-dimethyl-1,4,7,10,13,16-hexaazacyclooctadecane-2,5,8,11,14-pentane (trifluoroacetate) TIFF0007670618000227.tif62170 Step 1. Benzyl 4-(2-((2S,8S,11S,14S,17S)-17-((((benzyloxy)carbonyl)amino)methyl)-14-cyclohexyl-7-hexyl-11-isobutyl-8,10-dimethyl-3,9,12,15,18-pentaoxo-1,4,7,10,13,16-hexaazacyclooctadecan-2-yl)-2-((tert-butyldimethylsilyl)oxy)ethyl)piperazine-1-carboxylate TIFF0007670618000228.tif68170

[0322] The title compound was obtained following a procedure similar to that described for Example 22 using Intermediate 9 and Intermediate T6. Step 2. (3S,6S,9S,12S,15S)-6-(aminomethyl)-9-cyclohexyl-16-hexyl-3-(1-hydroxy-2-(piperazin-1-yl)ethyl)-12-isobutyl-13,15-dimethyl-1,4,7,10,13,16-hexaazacyclooctadecane-2,5,8,11,14-pentane (trifluoroacetate)

[0323] To the material from step 1 (50 mg) dissolved in THF (1 mL) was added tetrabutylammonium fluoride (1 mL, 1N in THF), and the mixture was stirred overnight. The reaction mixture was diluted with IPAC, washed with water and brine, dried over sodium sulfate, and concentrated to give crude benzyl 4-(2-((2S,8S,11S,14S,17S)-17-((((benzyloxy)carbonyl)amino)methyl)-14-cyclohexyl-7-hexyl-11-isobutyl-8,10-dimethyl-3,9,12,15,18-pentaoxo-1,4,7,10,13,16-hexaazacyclooctadecan-2-yl)-2-hydroxyethyl)piperazine-1-carboxylate. This material was converted to the title compound using a procedure similar to that described in Example 22 (Step 6), and then purified by reverse phase HPLC to give the title compound as the trifluoroacetic acid salt. LCMS (ESI):R T(minutes)=5.95;[M+H] + =736.5, method=B. [Example 29] (3S,6S,9S,12S,15S,17S)-6-(aminomethyl)-3-((3-aminopropoxy)methyl)-9-cyclohexyl-16-(cyclopentylmethyl)-13,15,17-trimethyl-12-propyl-1,4,7,10,13,16-hexaazacyclooctadecane-2,5,8,11,14-pentane (trifluoroacetate) TIFF0007670618000229.tif65170Step 1. N-Me-Norvaline-Cyclohexylglycine-Dap(Boc)-3-((tert-butoxycarbonyl)amino)propyl)-serine-(2-chlorotrityl resin) JPEG0007670618000230.jpg57170

[0324] The resin-bound tetrapeptide was prepared according to a method similar to that described in Example 7, step 1. Step 2. (3S,6S,9S,12S,15S,17S)-6-(aminomethyl)-3-((3-aminopropoxy)methyl)-9-cyclohexyl-16-(cyclopentylmethyl)-13,15,17-trimethyl-12-propyl-1,4,7,10,13,16-hexaazacyclooctadecane-2,5,8,11,14-pentane

[0325] The title compound was prepared as its TFA salt using the resin-bound tetrapeptide from Step 1 and (2S)-2-[cyclopentylmethyl-[(1S)-2-(9H-fluoren-9-ylmethoxycarbonylamino)-1-methyl-ethyl]amino]propanoic acid (Intermediate T21) following a similar method to that described in Example 12, Steps 2-4. LCMS (ESI): [M+H] + =693.5, R t =1.9 minutes, method=J. 1H NMR (400MHz, DMSO-d6): δ8.92-8.75(m,1H), 8.18-7.92(m,4H), 7.81-7.70(m,4H), 4.85-4.72(m,1H), 4.58-4.50(m,1H), 4.24-4.07(m,3H), 3.95-3.77(m,2H), 3.28-3 .09(m,8H), 2.91-2.82(m,4H), 2.14-1.92(m,5H), 1.80-1.71(m,5H), 1.67-1.58(m ,6H), 1.57-1.50(m,5H), 1.44-1.40(m,2H), 1.29-1.02(m,10H), 0.99-0.88(m,4H). [Example 30] (3R,6S,9S,12S,15S,18R,19R)-9-(aminomethyl)-6-[[3-aminopropyl(methyl)amino]methyl]-12-cycloheptyl-19-hexyl-3,16,18-trimethyl-15-propyl-1-oxa-4,7,10,13,16-pentaazacyclononadecane-5,8,11,14,17-pentone (trifluoroacetate) TIFF0007670618000231.tif64170 Step 1. tert-Butyl N-[3-[[(3R,6S,9S,12S,15S,18R,19R)-9-[(tert-butoxycarbonylamino)methyl]-12-cycloheptyl-19-hexyl-3,16,18-trimethyl-5,8,11,14,17-pentaoxo-15-propyl-1-oxa-4,7,10,13,16-pentaazacyclononadequa-6-yl]methyl-methyl-amino]propyl]carbamate TIFF0007670618000232.tif58170

[0326] To a solution of tert-butyl N-[3-[[(3R,6S,9S,12S,15S,18R,19R)-9-[(tert-butoxycarbonylamino)methyl]-12-cycloheptyl-19-hexyl-3,16,18-trimethyl-5,8,11,14,17-pentaoxo-15-propyl-1-oxa-4,7,10,13,16-pentaazacyclononadece-6-yl]methylamino]propyl]carbamate (Example 14, Step 3) (201 mg, 0.218 mmol) in ethanol (5 mL) was added paraformaldehyde (71.7 mg, 2.39 mmol) and acetic acid (26.1 mg, 0.435 mmol). The reaction mixture was stirred at room temperature for 2 hours, after which sodium cyanoborohydride (54.7 mg, 0.87 mmol) was added. The reaction mixture was stirred at room temperature for 3 days and then concentrated in vacuo. The residue was purified by reverse-phase chromatography on a C18 column (0-50% acetonitrile / water with 0.1% TFA) to give the title compound (119 mg) as a white solid. LCMS (ESI): [M+H] + =937.8. Step 2. (3R,6S,9S,12S,15S,18R,19R)-9-(aminomethyl)-6-[[3-aminopropyl(methyl)amino]methyl]-12-cycloheptyl-19-hexyl-3,16,18-trimethyl-15-propyl-1-oxa-4,7,10,13,16-pentaazacyclononadecane-5,8,11,14,17-pentone

[0327] The product of step 1 (119 mg, 0.126 mmol) was treated with TFA (5.0 mL) at 0° C. for 30 minutes and then concentrated in vacuo. The residue was purified by preparative LCMS to give the title compound (9.90 mg) as its TFA salt as a white solid. LCMS (ESI): [M+H] + =737.6, R t =2.5 minutes, method=I. 1H NMR (400MHz, DMSO-d6): δ9.62-9.45(m,1H), 8.62-8.35(m,1H), 8.18-8.12(m,1H), 8.02-7.92 (m,2H), 7.88-7.74(m,3H), 7.52-7.41(m,1H), 7.29-6.95(m,1H), 4.62-4.55(m,1H), 4.30-4. 22(m,1H), 4.09-4.02(m,1H), 3.92-3.80(m,2H), 3.30-3.08(m,10H), 2.97-2.74(m,8H), 2.15 -2.07(m,2H), 1.98-1.88(m,4H), 1.75-1.59(m,7H), 1.45-1.24(m,17H), 1.05-0.85(m,11H). [Example 31] 1-((1S,3r)-3-(((3R,6S,9S,12S,15S,18R,19R)-9-(aminomethyl)-12-cycloheptyl-3,16,18-trimethyl-5,8,11,14,17-pentaoxo-15-propyl-19-(spiro[3.3]heptan-2-ylmethyl)-1-oxa-4,7,10,13,16-pentaazacyclononadecan-6-yl)methoxy)cyclobutyl)guanidine (trifluoroacetate) TIFF0007670618000233.tif75170 Step 1. tert-Butyl N-[3-[[(3R,6S,9S,12S,15S,18R,19R)-9-(benzyloxycarbonylaminomethyl)-12-cycloheptyl-3,16,18-trimethyl-5,8,11,14,17-pentaoxo-15-propyl-19-(spiro[3.3]heptan-2-ylmethyl)-1-oxa-4,7,10,13,16-pentaazacyclononadequa-6-yl]methoxy]cyclobutyl]carbamate TIFF0007670618000234.tif59170

[0328] The title compound was prepared according to a method similar to that described in Example 8, steps 1-3. LCMS (ESI): [M+H] + =994.6. Step 2. Benzyl (((3R,6S,9S,12S,15S,18R,19R)-6-(((1r,3S)-3-aminocyclobutoxy)methyl)-12-cycloheptyl-3,16,18-trimethyl-5,8,11,14,17-pentaoxo-15-propyl-19-(spiro[3.3]heptan-2-ylmethyl)-1-oxa-4,7,10,13,16-pentaazacyclononadecan-9-yl)methyl)carbamate TIFF0007670618000235.tif68170

[0329] The product of step 1 (577 mg, 0.580 mmol) was treated with TFA (10 mL) at 0° C. for 1 h and then concentrated in vacuo. The residue was co-evaporated with toluene (2×10 mL) to remove residual TFA. The crude product was used directly as the TFA salt in the next step without further purification. LCMS (ESI): [M+H] + =894.6. Step 3. tert-Butyl (NZ)-N-[[[3-[[(3R,6S,9S,12S,15S,18R,19R)-9-(benzyloxycarbonylaminomethyl)-12-cycloheptyl-3,16,18-trimethyl-5,8,11,14,17-pentaoxo-15-propyl-19-(spiro[3.3]heptan-2-ylmethyl)-1-oxa-4,7,10,13,16-pentaazacyclononadece-6-yl]methoxy]cyclobutyl]amino]-(tert-butoxycarbonylamino)methylene]carbamate TIFF0007670618000236.tif85170

[0330] To a solution of the crude product from step 2 (519 mg) in ethanol (60 mL) was added (Z)-tert-butyl(((tert-butoxycarbonyl)amino)(1H-pyrazol-1-yl)methylene)carbamate (360 mg, 1.16 mmol) and DIPEA (0.40 mL, 2.3 mmol) at room temperature. The reaction mixture was stirred at room temperature for 4 hours and then concentrated in vacuo. The residue was dissolved in DCM (150 mL) and washed with 10% ammonium chloride in water (2×150 mL). The organic layer was dried over anhydrous sodium sulfate and concentrated in vacuo. The residue was purified by reverse-phase chromatography on a C18 column (0-100% acetonitrile / 0.05% TFA in water) to give the title compound (281 mg) as a colorless solid. LCMS (ESI): [M+H] + =1136.7. Step 4. tert-Butyl (NZ)-N-[[[3-[[(3R,6S,9S,12S,15S,18R,19R)-9-(aminomethyl)-12-cycloheptyl-3,16,18-trimethyl-5,8,11,14,17-pentaoxo-15-propyl-19-(spiro[3.3]heptan-2-ylmethyl)-1-oxa-4,7,10,13,16-pentaazacyclononadece-6-yl]methoxy]cyclobutyl]amino]-(tert-butoxycarbonylamino)methylene]carbamate TIFF0007670618000237.tif79170

[0331] The product from step 3 (281 mg, 0.247 mmol) was combined with Pd / C (280 mg, 10% Pd on carbon) in 2-propanol (35 mL) and stirred under a hydrogen atmosphere at room temperature for 6 hours. The catalyst was removed by filtration, and the filtered solution was concentrated under reduced pressure to give the title compound (220 mg, 88.8% yield) as a colorless solid. LCMS (ESI): [M+H] + =1002.8. Step 5. 1-((1S,3r)-3-(((3R,6S,9S,12S,15S,18R,19R)-9-(aminomethyl)-12-cycloheptyl-3,16,18-trimethyl-5,8,11,14,17-pentaoxo-15-propyl-19-(spiro[3.3]heptan-2-ylmethyl)-1-oxa-4,7,10,13,16-pentaazacyclononadecan-6-yl)methoxy)cyclobutyl)guanidine

[0332] The product from step 4 (220 mg, 0.220 mmol) was treated with TFA (4.0 mL) at 0° C. for 4 hours and then concentrated in vacuo. The residue was co-evaporated with toluene (2×10 mL). The crude product was purified by preparative HPLC to give the title compound (45.3 mg) as its TFA salt as a white solid. LCMS (ESI): [M+H] + =802.6, R t =2.72 minutes, method=I. 1 H NMR (400MHz, DMSO-d6): δ8.56-8.31(m,1H), 8.18-8.12(m,2H), 8.08-7.99(m,4H), 7.92-7.85(m,1H), 7.34-7.06(m,4 H), 4.52-4.35(m,2H), 4.20-4.08(m,2H), 4.02-3.92(m,2H), 3.88-3.80(m,1H), 3.68-3.60(m,1H), 3.52-3.42(m,1H), 3.41-3.34(m,1H), 3.32-3.12(m,6H), 3.15(s,1H), 2.73(s,2H), 2.31-2.23(m,3H), 2.16-1.98(m,8H), 1.88-1.81(m,2 H), 1.78-1.72(m,3H), 1.69-1.55(m,8H), 1.53-1.45(m,6H), 1.32-1.21(m,3H), 0.99-0.89(m,6H), 0.88-0.83(m,3H). [Example 32] (3S,6S,9S,12S,15S,18S)-6-(aminomethyl)-16-butyl-9-cyclohexyl-3-((S)-1-hydroxyethyl)-18-(hydroxymethyl)-12-isobutyl-13,15-dimethyl-1,4,7,10,13,16-hexaazacyclooctadecane-2,5,8,11,14-pentane TIFF0007670618000238.tif44170Step 1: Methyl N-(tert-butoxycarbonyl)-O-(tert-butyldiphenylsilyl)-D-serinate TIFF0007670618000239.tif38170

[0333] To a suspension of methyl (2R)-2-amino-3-hydroxypropanoate hydrochloride (2.8 g, 18 mmol) in a THF / water mixture, di-tert-butyl carbonate (3.8 g, 22 mmol) and sodium bicarbonate (1.5 g, 18 mmol) were added, and the mixture was stirred overnight. The reaction mixture was diluted with IPAC, washed with brine, dried over sodium sulfate, and concentrated. The residue was dissolved in DCM (30 mL), and tert-butylchlorodiphenylsilane (5.9 g, 22 mmol) and imidazole (2.5 g, 36 mmol) were added, and the mixture was stirred overnight. The reaction mixture was diluted with IPAC, washed with water and brine, dried over sodium sulfate, and concentrated. The residue was purified by chromatography (silica gel 0-50% IPAC / heptane) to give the title compound (8.1 g, 98%). 1 H NMR (400MHz, chloroform-d) δ7.61-7.53(m,3H), 7.48-7.34(m,7H), 5.41(d,J=8.8Hz,1H), 4.50-4.30(m,1) H), 4.06(dd,J=10.1,3.0Hz,1H), 3.89(dd,J=10.1,3.1Hz,1H), 3.74(s,3H), 1.46(s,9H), 1.03(s,9H). Step 2: Benzyl N-((S)-2-((tert-butoxycarbonyl)amino)-3-((tert-butyldiphenylsilyl)oxy)propyl)-N-hexyl-L-alaninate TIFF0007670618000240.tif49170

[0334] The residue from step 1 was dissolved in anhydrous THF (30 mL) and cooled in an ice bath. Lithium aluminum hydride (8.85 mL, 2 M in THF) was added, and the reaction mixture was allowed to warm to room temperature over 2 hours. The reaction mixture was quenched with saturated aqueous ammonium chloride, and the solids were removed by filtering through Celite. The filtered solution was dried over sodium sulfate and concentrated to give crude tert-butyl (S)-(1-((tert-butyldiphenylsilyl)oxy)-3-hydroxypropan-2-yl)carbamate.

[0335] This material was converted to the aldehyde as described in Step 4 of Example 20 to give tert-butyl N-[(1R)-1-[[tert-butyl(diphenyl)silyl]oxymethyl]-2-oxo-ethyl]carbamate (3.5 g, 54%). 1 H NMR (400 MHz, chloroform-d) δ 9.66 (s, 1H), 7.69-7.56 (m, 4H), 7.51-7.35 (m, 6H), 5.40 (d, J = 7.6 Hz, 1H), 4.42-3.88 (m, 3H), 1.45 (d, J = 9.5 Hz, 9H), 1.03 (s, 9H).

[0336] Following the procedure described in Step 5 of Example 20, this material was converted to the title compound (1.75 g, 55%). 1 H NMR (400MHz, chloroform-d) δ7.70-7.52(m,4H), 7.48-7.30(m,11H), 5.10(s,2H), 4.73(s,1H), 3.79-3.50(m,4 H), 2.88-2.67(m,2H), 2.63-2.40(m,2H), 1.43(s,9H), 1.38-1.18(m,11H), 1.05(s,9H), 0.94-0.81(m,3H). Step 3. N-((S)-2-((((9H-fluoren-9-yl)methoxy)carbonyl)amino)-3-((tert-butyldiphenylsilyl)oxy)propyl)-N-hexyl-L-alanine TIFF0007670618000241.tif58170

[0337] Benzyl N-((S)-2-((tert-butoxycarbonyl)amino)-3-((tert-butyldiphenylsilyl)oxy)propyl)-N-hexyl-L-alaninate (1.75 g) was converted to the title compound (1.0 g, 54%) as described in Steps 6 and 7 of Example 20. 1 H NMR (400MHz, chloroform-d) δ7.74(d,J=7.6Hz,2H), 7.60(dd,J=18.3,7.4Hz,6H), 7.46- 7.34(m,8H), 7.32-7.19(m,2H), 5.98(d,J=7.4Hz,1H), 4.33(d,J=8.5Hz,2H), 4.18( t,J=7.2Hz,1H), 3.97-3.59(m,4H), 3.07-2.93(m,2H), 2.73(d,J=56.2Hz,2H), 1.51 (s,2H), 1.42-1.35(m,3H), 1.22(d,J=11.7Hz,6H), 1.08(s,9H), 0.88-0.78(m,3H). Step 4. Benzyl (((2S,5S,8S,11S,14S,17S)-5-((S)-1-(benzyloxy)ethyl)-17-cyclohexyl-8-((((2,2-dimethyl-1,1-diphenylpropyl)dimethylsilyl)oxy)methyl)-10-hexyl-14-isobutyl-11,13-dimethyl-3,6,12,15,18-pentaoxo-1,4,7,10,13,16-hexaazacyclooctadecan-2-yl)methyl)carbamate TIFF0007670618000242.tif49170

[0338] The title compound (0.30 g) was prepared using the intermediate from step 2 and following a procedure similar to that described in steps 8 and 9 of example 20. LCMS (ESI): [M+H] + =1145.8. Step 5. (3S,6S,9S,12S,15S,18S)-6-(aminomethyl)-16-butyl-9-cyclohexyl-3-((S)-1-hydroxyethyl)-18-(hydroxymethyl)-12-isobutyl-13,15-dimethyl-1,4,7,10,13,16-hexaazacyclooctadecane-2,5,8,11,14-pentane

[0339] The title compound was prepared as its TFA salt using the intermediate from step 4 following a procedure similar to that described in example 28, step 2. LCMS (ESI):R T (min)=7.807, [M+H] + =682.5, Method B (purity: 82%). [Example 33] N-[[(3R,6S,9S,12S,15S,18R,19R)-9-(aminomethyl)-12-cyclohexyl-19-hexyl-15-isobutyl-3,16,18-trimethyl-5,8,11,14,17-pentaoxo-1-oxa-4,7,10,13,16-pentaazacyclononadequa-6-yl]methyl]methanesulfonamide (trifluoroacetate) TIFF0007670618000243.tif63170

[0340] Benzyl N-[[(3R,6S,9S,12S,15S,18R,19R)-6-(aminomethyl)-12-cyclohexyl-19-hexyl-15-isobutyl-3,16,18-trimethyl-5,8,11,14,17-pentaoxo-1-oxa-4,7,10,13,16-pentaazacyclononadequa-9-yl]methyl]carbamate (Example 26, Step 3) (60 mg, 0.68 mmol) was dissolved in 0.20 mL of DCM, and DIPEA (26 μL, 0.15 mmol) and methanesulfonyl chloride (6 μL, 0.075 mmol) were added. The mixture was then stirred at room temperature for 1 hour. The mixture was then diluted with EtOAc and water, shaken vigorously, and partitioned. The organic portion was then washed once with water and concentrated in vacuo.

[0341] The residue was then dissol...

Claims

1. Formula (I): During the ceremony: R １ａ , R ２ａ , R ４ａ , R ５ａ and R ６ａ are each independently hydrogen or an optionally substituted C １ ~C ６ is alkyl, R ３ａ is hydrogen, optionally substituted C １ ~C ２０ Alkyl, optionally substituted C ２ ~C ２０ Alkenyl, optionally substituted C ２ ~C ２０ Alkynyl, optionally substituted cycloalkyl, optionally substituted (C １ ~C ６ alkyl)cycloalkyl, optionally substituted heterocycloalkyl, optionally substituted (C １ ~C ６ alkyl)heterocycloalkyl, optionally substituted aryl, optionally substituted (C １ ~C ６ alkyl)aryl, optionally substituted heteroaryl, or optionally substituted (C １ ~C ６ alkyl)heteroaryl; R ２０ is -NR １ R ２ and R １ and R ２ is hydrogen, R ３ and R ４ is hydrogen, R ５ is hydrogen, R ６ and R ７ are each independently hydrogen, halogen, or optionally substituted C １ ~C ６ Alkyl, optionally substituted C ２ ~C ６ Alkenyl, optionally substituted C ２ ~C ６ Alkynyl, optionally substituted cycloalkyl, optionally substituted (C １ ~C ６ alkyl)cycloalkyl, optionally substituted heterocycloalkyl, optionally substituted (C １ ~C ６ alkyl)heterocycloalkyl, optionally substituted aryl, optionally substituted (C １ ~C ６ alkyl)aryl, optionally substituted heteroaryl, or optionally substituted (C １ ~C ６ alkyl)heteroaryl; R ８ and R ９ are each independently hydrogen, halogen, or optionally substituted C １ ~C ６ Alkyl, optionally substituted C ２ ~C ６ Alkenyl, optionally substituted C ２ ~C ６ Alkynyl, optionally substituted cycloalkyl, optionally substituted (C １ ~C ６ alkyl)cycloalkyl, optionally substituted heterocycloalkyl, optionally substituted (C １ ~C ６ alkyl)heterocycloalkyl, optionally substituted aryl, optionally substituted (C １ ~C ６ alkyl), aryl, optionally substituted heteroaryl, or optionally substituted (C １ ~C ６ alkyl)heteroaryl; Or, R ３ａ and R ８ together with the atom to which they are attached form an optionally substituted heterocycloalkyl; R １０ and R １１ are each independently hydrogen, halogen, or optionally substituted C １ ~C ６ Alkyl, optionally substituted C ２ ~C ２０ Alkenyl, optionally substituted C ２ ~C ６ Alkynyl, optionally substituted cycloalkyl, optionally substituted (C １ ~C ６ alkyl)cycloalkyl, optionally substituted heterocycloalkyl, optionally substituted (C １ ~C ６ alkyl)heterocycloalkyl, optionally substituted aryl, optionally substituted (C １ ~C ６ alkyl)aryl, optionally substituted heteroaryl, or optionally substituted (C １ ~C ６ alkyl)heteroaryl; R １２ and R １３ are each independently hydrogen, halogen, or optionally substituted C １ ~C ２０ Alkyl, optionally substituted C ２ ~C ６ Alkenyl, optionally substituted C ２ ~C ６ Alkynyl, optionally substituted cycloalkyl, optionally substituted (C １ ~C ６ alkyl)cycloalkyl, optionally substituted heterocycloalkyl, optionally substituted (C １ ~C ６ alkyl)heterocycloalkyl, optionally substituted aryl, optionally substituted (C １ ~C ６ alkyl)aryl, optionally substituted heteroaryl, or optionally substituted (C １ ~C ６ alkyl)heteroaryl; R １４ and R １５ are each independently hydrogen, halogen, or optionally substituted C １ ~C ６ Alkyl, optionally substituted C ２ ~C ６ Alkenyl, optionally substituted C ２ ~C ６ Alkynyl, optionally substituted cycloalkyl, optionally substituted (C １ ~C ６ alkyl)cycloalkyl, optionally substituted heterocycloalkyl, optionally substituted (C １ ~C ６ alkyl)heterocycloalkyl, optionally substituted aryl, optionally substituted (C １ ~C ６ alkyl)aryl, optionally substituted heteroaryl, or optionally substituted (C １ ~C ６ alkyl)heteroaryl; R １６ and R １７ are each independently hydrogen, halogen, or optionally substituted C １ ~C ６ Alkyl, optionally substituted C ２ ~C ６ Alkenyl, optionally substituted C ２ ~C ６ Alkynyl, optionally substituted cycloalkyl, optionally substituted (C １ ~C ６ alkyl)cycloalkyl, optionally substituted heterocycloalkyl, optionally substituted (C １ ~C ６ alkyl)heterocycloalkyl, optionally substituted aryl, optionally substituted (C １ ~C ６ alkyl)aryl, optionally substituted heteroaryl, or optionally substituted (C １ ~C ６ alkyl)heteroaryl; or a pharma- ceutically acceptable salt, solvate or stereoisomer thereof, wherein the compound of formula (I) has the formula (Ib): R １ , R ２ , R ３ , R ４ , R １ａ , R ２ａ , R ４ａ , R ５ａ , R ６ａ , R ９ , R １０ and R １１ is hydrogen, R 6 is and R 16 is or a pharma- ceutically acceptable salt, solvate or stereoisomer thereof.

2. R ８ is hydrogen or C １ ~C ６ The compound of claim 1 , wherein the aryl group is an alkyl group.

3. R ３ａ and R ８ 13. The compound of claim 1, wherein together with the atom to which they are attached form an azetidinyl, pyrrolidinyl, or piperidinyl.

4. R ６ but, The compound according to any one of claims 1 to 3,

5. R １２ but, The compound according to any one of claims 1 to 4,

6. R １４ is optionally substituted C １ ~C ６ Alkyl or optionally substituted (C １ ~C ６ 6. The compound according to any one of claims 1 to 5, wherein the aryl group is aryl)cycloalkyl.

7. R １６ The compound according to any one of claims 1 to 6, wherein is cyclohexyl.

8. R ３ａ but, The compound according to any one of claims 1, 2 or 4 to 7, 9. The compound of claim 1, or a pharma- ceutically acceptable salt, solvate or stereoisomer thereof, selected from the group consisting of:

10. A pharmaceutical composition comprising a compound according to any one of claims 1 to 9 or a pharma- ceutically acceptable salt, solvate or stereoisomer thereof, and a pharma- ceutically acceptable excipient.

11. 10. A compound according to any one of claims 1 to 9, or a pharma- ceutically acceptable salt, solvate or stereoisomer thereof, for use in a method of treating a bacterial infection in a mammal, said method comprising administering to said mammal an effective amount of a compound according to any one of claims 1 to 9, or a pharma- ceutically acceptable salt, solvate or stereoisomer thereof, at a frequency and for a period of time sufficient to provide a beneficial effect to said mammal, said bacterial infection being selected from the group consisting of Pseudomonas aeruginosa, Pseudomonas fluorescens, Pseudomonas acidovorans, Pseudomonas alcaligenes, Pseudomonas putida, Pseudomonas erythropoeis ... putida, Stenotrophomonas maltophilia, Burkholderia cepacia, Aeromonas hydrophila, Escherichia coli, Citrobacter freundii, Salmonella typhimurium, Salmonella typhi, Salmonella paratyphi, Salmonella enteritidis enteritidis, Shigella dysenteriae, Shigella flexneri, Shigella sonnei, Enterobacter cloacae, Enterobacter aerogenes, Klebsiella pneumoniae, Klebsiella oxytoca, Serratia marcescensmarcescens, Francisella tularensis, Morganella morganii, Proteus mirabilis, Proteus vulgaris, Providencia alcalifaciens, Providencia rettgeri, Providencia stuartii, Acinetobacter baumannii baumannii, Acinetobacter calcoaceticus, Acinetobacter haemolyticus, Yersinia enterocolitica, Yersinia pestis, Yersinia pseudotuberculosis, Yersinia intermedia, Bordetella pertussis, Bordetella parapertussis parapertussis, Bordetella bronchiseptica, Haemophilus influenzae, Haemophilus parainfluenzae, Haemophilus haemolyticus, Haemophilus parahaemolyticus, Haemophilus ducreyi, Pasteurella multocida multocida, Pasteurella haemolytica, Branhamella catarrhaliscatarrhalis, Helicobacter pylori, Campylobacter fetus, Campylobacter jejuni, Campylobacter coli, Borrelia burgdorferi, Vibrio cholerae, Vibrio parahaemolyticus, Legionella pneumophila, Listeria monocytogenes monocytogenes, Neisseria gonorrhoeae, Neisseria meningitidis, Kingella, Moraxella, Gardnerella vaginalis, Bacteroides fragilis, Bacteroides distasonis, Bacteroides 3452A homology group, Bacteroides vulgatus or a pharmaceutically acceptable salt, solvate or stereoisomer thereof, wherein the infection is an infection involving Bacteroides vulgatus, Bacteroides ovalus, Bacteroides thetaiotaomicron, Bacteroides uniformis, Bacteroides eggerthii or Bacteroides splanchnicus.

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