Rapafusine derivative compounds and methods of use thereof

Abstract

This disclosure provides macrocycles inspired by the natural product FK506 and rapamycin, a family of immunophilin ligands. The generation of a rapafusin library of macrocycles containing the FK506 and rapamycin binding domains is expected to hold great potential as new leads for the development of drugs for the treatment of diseases.

Description

CROSS-REFERENCE TO RELATED APPLICATIONS

[0001] This application claims the benefit of priority under 35 U.S.C. § 119(e) of U.S. Serial No. 62 / 909,008, filed October 1, 2019, the entire contents of which are incorporated by reference. Government support announcement

[0002] This invention was made with government support under CA174428 awarded by the National Institutes of Health. The government has certain rights in this invention. Background information Summary of the Invention [Problem to be solved by the invention]

[0003] The macrocyclic natural products FK506 and rapamycin are approved immunosuppressive drugs with significant biological activity. Both have been shown to suppress T cell activation, albeit through distinct mechanisms. Furthermore, rapamycin has been shown to have potent antiproliferative activity. FK506 and rapamycin share a distinct mode of action; they act by recruiting the abundant and ubiquitously expressed intracellular protein, prolyl cis-trans isomerase FKBP, and the resulting binary complexes subsequently bind to and allosterically inhibit their target proteins, calcineurin and mTOR, respectively. Structurally, FK506 and rapamycin share a similar FKBP-binding domain but differ in their effector domains. Properties of FK506 and rapamycin suggest that swapping the effector domain of FK506 with that of rapamycin can shift the target from calcineurin to mTOR. The generation of a Rapafucin library of macrocyles containing FK506 and rapamycin binding domains should hold great potential as new leads for the development of drugs to treat disease.

[0004] With the completion of the sequencing and annotation of the human genome, a complete catalog of all human proteins encoded in the genome is now available. However, the functions of the majority of these proteins remain unknown. One approach to elucidating these protein functions is to find small molecule ligands that specifically bind to proteins of interest and perturb their biochemical and cellular functions. Thus, a major challenge for chemical biologists today is to discover new small molecule probes for novel proteins to facilitate elucidation of their functions. Recent advances in the development of protein chips have provided exciting new opportunities for simultaneously screening chemical libraries against nearly the entire human proteome. A single chip, in the form of a glass slide, is sufficient to display the entire proteome in a dual array. Recently, a protein chip with 17,000 human proteins displayed on a single slide was produced. A major benefit of using human protein chips for screening is that the entire displayed proteome can be interrogated at once in a small volume (<3 mL) of assay buffer. However, screening of human protein chips is not yet feasible with most, if not all, existing chemical libraries due to the lack of a universal readout for detecting ligand binding to proteins on these chips. While it is possible to add artificial tags to individual compounds in synthetic libraries, the added tags themselves often interfere with the activity of the ligand. Thus, there remains a need for new compounds and methods for screening chemical libraries against the human proteome. [Means for solving the problem]

[0005] Overview The present disclosure is directed to libraries of rapafucine compounds, methods for making these compounds, and methods for using them. The present disclosure is further directed to DNA-encoded libraries of hybrid cyclic molecules, and more specifically, DNA-encoded libraries of hybrid cyclic compounds based on the natural product FK506 and the rapamycyin family of immunophilin ligands.

[0006] Also provided herein is a macrocyclic compound of formula (XIV) or a pharmaceutically acceptable salt, solvate, or stereoisomer thereof: [ka]

[0007] Each n, m, and p can be independently an integer selected from 0 to 5 inclusive.

[0008] Each R1, R2, and R3 is H, F, Cl, Br, CF3, CN, N3, -N(R 12 )2, -N(R 12 )3, -CON(R 12 )2, NO2, OH, OCH3, methyl, ethyl, propyl, -COOH, -SO3H, -PO(OR 12 )2, -OPO(OR 12 )2, -(CH2) q COOH, -O-(CH2) q COOH, -S-(CH2) q COOH, -CO-(CH2) q COOH, -NR 12 -(CH2) q COOH, -(CH2) q SO3H, -O-(CH2) q SO3H, -S-(CH2) q SO3H, -CO-(CH2) q SO3H, -NR 12 -(CH2) q SO3H, -(CH2) q N(R 12 )2, -O-(CH2) qN(R 12 )2, -S-(CH2) q N(R 12 )2, -CO-(CH2) q N(R 12 )2, -(CH2) q N(R 12 )3, -O-(CH2) q N(R 12 )3, -S-(CH2) q N(R 12 )3, -CO-(CH2) q N(R 12 )3, -NR 12 -(CH2) q N(R 12 )3, -(CH2) q CON(R 12 )2, -O-(CH2) q CON(R 12 )2, -S-(CH2) q CON(R 12 )2, -CO-(CH2) q CON(R 12 )2, -(CH2) q PO(OR 12 )2, -O(CH2) q PO(OR 12 )2, -S(CH2) q PO(OR 12 )2, -CO(CH2) q PO(OR 12 )2, -NR 12 (CH2) q PO(OR 12 )2, -(CH2) q OPO(OR 12 )2, -O(CH2) q OPO(OR 12 )2, -S(CH2) q OPO(OR 12 )2, -CO(CH2) q OPO(OR 12 )2, and -NR 12 (CH2) q OPO(OR 12 )2.

[0009] In one embodiment, q can be an integer selected from 0 to 5. Each of R, R, R, R, R, and R 11 may be independently selected from the group consisting of H, methyl, ethyl, propyl, and isopropyl.

[0010] In another embodiment, each R and R 10 is H, halogen, hydroxyl, C 1-20 Alkyl, N3, NH2, NO2, CF3, OCF3, OCHF2, COC 1-20 Alkyl, CO2C 1-20 Alkyl, a 5- or 6-membered ring moiety formed with the adjacent nitrogen, -N(R 12 )2, -N(R 12 )3, -CON(R 12 )2, -COOH, -SO3H, -PO(OR 12 )2, -OPO(OR 12 )2, -(CH2) q COOH, -O-(CH2) q COOH, -S-(CH2) q COOH, -CO-(CH2) q COOH, -NR 12 -(CH2) q COOH, -(CH2) q SO3H, -O-(CH2) q SO3H, -S-(CH2) q SO3H, -CO-(CH2) q SO3H, -NR 12 -(CH2) q SO3H, -(CH2) q N(R 12 )2, -O-(CH2) q N(R 12 )2, -S-(CH2) q N(R 12 )2, -CO-(CH2) q N(R 12 )2, -(CH2) q N(R 12 )3, -O-(CH2) q N(R 12 )3, -S-(CH2) q N(R 12)3, -CO-(CH2) q N(R 12 )3, -NR 12 -(CH2) q N(R 12 )3, -(CH2) q CON(R 12 )2, -O-(CH2) q CON(R 12 )2, -S-(CH2) q CON(R 12 )2, -CO-(CH2) q CON(R 12 )2, -(CH2) q PO(OR 12 )2, -O(CH2) q PO(OR 12 )2, -S(CH2) q PO(OR 12 )2, -CO(CH2) q PO(OR 12 )2, -NR 12 (CH2) q PO(OR 12 )2, -(CH2) q OPO(OR 12 )2, -O(CH2) q OPO(OR 12 )2, -S(CH2) q OPO(OR 12 )2, -CO(CH2) q OPO(OR 12 )2, and -NR 12 (CH2) q OPO(OR 12 )2.

[0011] Each R 12 may be independently selected from the group consisting of H, methyl, ethyl, propyl, and isopropyl.

[0012] However, R2, R3, R8 and R 10 At least one of -N(R 12 )2, -N(R 12 )3, -CON(R 12 )2, -COOH, -SO3H, -PO(OR 12)2、-OPO(OR 12 )2、-(CH2) q COOH、-O-(CH2) q COOH、-S-(CH2) q COOH、-CO-(CH2) q COOH、-NR 12 -(CH2) q COOH、-(CH2) q SO3H、-O-(CH2) q SO3H、-S-(CH2) q SO3H、-CO-(CH2) q SO3H、-NR 12 -(CH2) q SO3H、-(CH2) q N(R 12 )2、-O-(CH2) q N(R 12 )2、-S-(CH2) q N(R 12 )2、-CO-(CH2) q N(R 12 )2、-(CH2) q N(R 12 )3、-O-(CH2) q N(R 12 )3、-S-(CH2) q N(R 12 )3、-CO-(CH2) q N(R 12 )3、-NR 12 -(CH2) q N(R 12 )3、-(CH2) q CON(R 12 )2、-O-(CH2) q CON(R 12 )2、-S-(CH2) q CON(R 12 )2、-CO-(CH2) q CON(R 12 )2、-(CH2) q PO(OR 12 )2、-O(CH2) q PO(OR 12 )2、-S(CH2) q PO(OR 12 )2、-CO(CH2) q PO(OR12 )2, -NR 12 (CH2) q PO(OR 12 )2, -(CH2) q OPO(OR 12 )2, -O(CH2) q OPO(OR 12 )2, -S(CH2) q OPO(OR 12 )2, -CO(CH2) q OPO(OR 12 )2, and -NR 12 (CH2) q OPO(OR 12 )2 is selected.

[0013] In certain embodiments, R1 can be H, R2 can be H, R3 can be —O—CH2COOH, and p can be 1. In certain embodiments, disclosed herein is compound 1593, having the following structure: [ka]

[0014] Also disclosed herein is a pharmaceutical composition comprising an effective amount of a compound according to Formula (XIV) and a pharmaceutically acceptable carrier. Also disclosed herein is a method of treating a disease in a subject, the method can include administering an effective amount of a compound according to Formula (XIV). In some embodiments, the disease can be selected from acute kidney injury, cerebral ischemia, hepatic ischemia-reperfusion injury, and organ transplant transport solution. In some embodiments, the compound can be administered intravenously.

[0015] Also disclosed herein are methods for synthesizing macrocycles, the methods including attaching a linker having an amine-terminated structure to a resin; sequentially reacting the linker-modified resin with different amino acids to obtain a polypeptide-modified resin; removing the resin to obtain a polypeptide intermediate; subjecting the polypeptide intermediate to reverse-phase chromatography to obtain a pure diastereomer of the polypeptide intermediate; reacting the pure diastereomer of the polypeptide intermediate with an FKBP binding domain (FKBD); and performing a macrocyclization reaction via olefin metathesis or lactamization. In some embodiments, four amino acids are used to obtain a tetrapeptide intermediate. In some embodiments, the R stereoisomer is obtained. [Brief explanation of the drawings]

[0016] [Figure 1] Figure 1 shows urea levels in a rat kidney ischemia-reperfusion model after 24 hours of administration: dipyridamole (DPA) was administered at 10 mg / kg; compound 1593 was administered at 12 mg / kg or 4 mg / kg; and compound 1594 was administered at 4 mg / kg. [Figure 2] Creatinine levels in a rat kidney ischemia-reperfusion model after 24 hours of administration: dipyridamole (DPA) administered at 10 mg / kg; compound 1593 administered at 12 mg / kg or 4 mg / kg; and compound 1594 administered at 4 mg / kg. [Figure 3] Figure 1 shows kidney injury molecule-1 (KIM-1) levels in a rat renal ischemia-reperfusion model after 24 hours of administration of dipyridamole (DPA) at 10 mg / kg, compound 1593 at 12 mg / kg or 4 mg / kg, and compound 1594 at 4 mg / kg. [Figure 4] Neutrophil gelatinase-associated lipocalin-1 (NGAL-1) levels in a rat kidney ischemia-reperfusion model after 24 hours of administration: dipyridamole (DPA) administered at 10 mg / kg; compound 1593 administered at 12 mg / kg or 4 mg / kg; and compound 1594 administered at 4 mg / kg. DETAILED DESCRIPTION OF THE INVENTION

[0017] Thanks to billions of years of evolution, nature is a rich source of bioactive small molecules that exhibit a dizzying array of cellular activities. Rapamycin and FK506 constitute a unique structural family of macrocyclic natural products with extraordinary modes of action. Once inside cells, both compounds form binary complexes with FKBP12 and other members of the FKBP family. The FKBP12-rapamycin complex can then bind to mTOR and block its kinase activity against downstream substrates, such as p70S6K and 4E-BP. Meanwhile, the FKBP12-FK506 complex interacts with calcineurin, a protein phosphatase, and its inhibition prevents calcium-dependent signaling and T cell activation. The ability of rapamycin and FK506 to bind to FKBPs offers multiple benefits for their use as small molecule probes in biology and as drugs in medicine. First, the binding of both rapamycin and FK506 to FKBP dramatically increases their effective size, enabling allosteric blockade of substrates to the active sites of mTOR or calcineurin through indirect disruption of protein-protein interactions. Second, the abundant and ubiquitous expression of intracellular FKBPs helps to concentrate and maintain the stability of rapamycin and FK506 in intracellular compartments. Third, as macrocycles, FK506 and rapamycin are capable of interacting with a wider range of proteins than much smaller molecules, regardless of their ability to bind FKBP. Finally, but particularly, the high expression of FKBPs in blood cells makes them drug reservoirs and carriers for efficient in vivo delivery. Thus, it is not surprising that both rapamycin and FK506 have become widely used drugs in their natural forms without further chemical modification.

[0018] Both rapamycin and FK506 can be divided into two structural and functional domains: the FKBP-binding domain (FKBD) and the effector domain that mediates interaction with mTOR or calcineurin, respectively. While the structures of the FKBDs of rapamycin and FK506 are very similar, their effector domains differ, explaining their exclusive target specificities. The presence of separable and modular structural domains in FK506 and rapamycin has been extensively exploited to generate new analogs of both FK506 and rapamycin, including chemical inducers of dimerization and numerous rapamycin analogs, known as rapalogs, to alter rapamycin's specificity for mutated FKBP-rapamycin binding domains of mTOR and to improve its toxicity and solubility profile. The existence of two distinct FKBD-containing macrocycles with distinct target specificities also raised the intriguing question of whether substitution of the effector domains of rapamycin or FK506 could further expand the target repertoire of the resulting macrocycles. In their pioneering work, Chakraborty and colleagues synthesized several rapamycin-peptide hybrid molecules that retained high affinity for FKBP but exhibited no biological activity. More recently, we and others have independently explored this possibility by creating even larger libraries of FKBD-containing macrocycles. In one study, a much larger library of FKBD-containing macrocycles was created by synthetic mimicking of FKBD, but the resulting macrocycles suffered a significant loss in binding affinity for FKBP12, likely explaining the lack of biologically active compounds from that library. Using the native FKBD extracted from rapamycin, we also observed a significant loss in FKBP binding affinity upon construction of the macrocycle (see below). [ka] Scheme 1. Structures of rapamycin and FK506 highlighting the FKBD moiety.

[0019] A rapafucine library was synthesized as described in WO 2017 / 136708 (International Publication No. 2017 / 136708). Rapadocin compounds and their analogs are disclosed in WO 2017 / 136717, where they are used to inhibit human equilibrative nucleoside transporter 1 (ENT1). Rapaglutins and their analogs are disclosed in WO 2017 / 136731, where they are used as inhibitors of cell proliferation and are useful for the treatment of cancer. Approximately 45,000 compounds were generated, and ongoing screening of the library, as described in WO 2018 / 045250, identified several compounds as inhibitors of MIF nuclease activity. All of these references are incorporated herein by reference.

[0020] In a continuing effort to explore the possibility of using FKBD-containing macrocycles to target new proteins, we have attempted and succeeded in optimizing FKBDs that enable significant retention of binding affinity for FKBP12 upon incorporation into macrocycles, and we have established a facile synthetic route for the parallel synthesis of multiple FKBD-containing macrocycles.

[0021] The following are some acronyms used in this disclosure: 2-MeTHF refers to 2-methyltetrahydrofuran; DMF refers to dimethylformamide; DMSO refers to dimethylsulfoxide; DCM refers to dichloromethane; HATU refers to 1-[bis(dimethylamino)methylene]-1H-1,2,3-triazolo[4,5-b]pyridinium 3-oxide hexafluorophosphate; DIEA refers to N,N-diisopropylethylamine; TFA refers to trifluoroacetic acid; Fmoc refers to fluorenylmethyloxycarbonyl; MeOH refers to methyl methyl ether; COMU-PF6 refers to (1-cyano-2-ethoxy-2-oxoethylidenaminooxy)dimethylamino-morpholinocarbenium hexafluorophosphate; CAN refers to acetonitrile; Oxyma refers to ethyl cyanohydroxyiminoacetate; LC-MS refers to liquid chromatography-mass spectrometry; T3P refers to n-propanephosphonic anhydride; and SPPS refers to solid phase peptide synthesis.

[0022] The following explanations of terms and methods are provided to better explain the present disclosure and to guide those skilled in the art in practicing the present disclosure. The singular terms "a," "an," and "the" include plural referents unless the context clearly dictates otherwise. Similarly, the word "or" is intended to include "and" unless the context clearly dictates otherwise. The term "comprises" means "includes." Thus, "including A or B" means "including A, B, or A and B," without excluding additional elements. The term "about" would be understood by a person of ordinary skill in the art. Whether or not the term "about" is explicitly used, all quantities given herein are intended to refer to the actual given value and to approximations to such given value that would be reasonably inferred based on one of ordinary skill in the art.

[0023] Further, it will be understood that all base or amino acid sizes, and all molecular weight or molecular mass values ​​given for nucleic acids or polypeptides are approximate and are provided for illustrative purposes. Methods and materials similar or equivalent to those described herein can be used in the practice or testing of this disclosure, suitable methods and materials are described below.

[0024] Unless otherwise explained, all technical and scientific terms used herein have the same meaning as commonly understood by one of ordinary skill in the art to which this disclosure belongs. Definitions of common terms in molecular biology can be found in Benjamin Lewin, Genes V, Oxford University Press, published by 1994 (ISBN 0-19-854287-9); Kendrew et al. (eds.), The Encyclopedia of Molecular Biology, published by Blackwell Science Ltd., 1994 (ISBN 0-632-02182-9); and Robert A. Meyers (ed.), Molecular Biology and Biotechnology: a Comprehensive Desk Reference, published by VCH Publishers, Inc., 1995 (ISBN 1-56081-569-8).

[0025] Unless otherwise indicated, the nomenclature of substituents not explicitly defined herein is arrived at by naming the terminal portion of the functionality followed by the adjacent functionality toward the point of attachment. A person of ordinary skill in the art will recognize that the above definitions are not intended to include impermissible substitution patterns (e.g., methyl substituted with five different groups, a pentavalent carbon, and the like). Such impermissible substitution patterns would be readily recognized by a person of ordinary skill in the art. All publications, patent applications, patents, and other references mentioned herein are incorporated by reference in their entirety. All sequences disclosed and provided with GenBank accession numbers are incorporated by reference. In case of conflict, the present specification, including explanations of terms, will control. Furthermore, the materials, methods, and examples are illustrative only and are not intended to be limiting.

[0026] An alkyl group refers to a monovalent group derived from an alkane by removing a hydrogen atom from any carbon atom, and includes straight-chain and branched-chain groups having 1 to 12 carbon atoms, and typically 1 to about 10 carbon atoms or, in some embodiments, 1 to about 6 carbon atoms, or in other embodiments, 1, 2, 3, or 4 carbon atoms. Examples of straight-chain alkyl groups include, but are not limited to, methyl, ethyl, n-propyl, n-butyl, n-pentyl, and n-hexyl groups. Examples of branched-chain alkyl groups include, but are not limited to, isopropyl, isobutyl, sec-butyl, and tert-butyl groups. Alkyl groups can be substituted or unsubstituted. Representative substituted alkyl groups can be mono-substituted or substituted more than once, such as, but not limited to, mono-, di-, or tri-substituted. As used herein, the term alkyl refers to both cyclic and non-cyclic groups, unless otherwise stated.

[0027] The term "cyclic alkyl" or "cycloalkyl" refers to a monovalent group derived from a cycloalkane by removing a hydrogen atom from a ring carbon atom. Cycloalkyl groups are saturated or partially saturated non-aromatic structures having single or multiple rings, including isolated, fused, bridged, and spiro ring systems, and have from 3 to 14 carbon atoms, or in some embodiments, from 3 to 12, or from 3 to 10, or from 3 to 8, or from 3, 4, 5, 6, or 7 carbon atoms. Cycloalkyl groups can be substituted or unsubstituted. Representative substituted cycloalkyl groups can be mono-substituted or substituted more than once, such as, but not limited to, mono-, di-, or tri-substituted. Examples of monocyclic cycloalkyl groups include, but are not limited to, cyclopropyl, cyclobutyl, cyclopentyl, and cyclohexyl groups. Examples of multi-cyclic ring systems include, but are not limited to, bicycle[4.4.0]decane, bicycle[2.2.1]heptane, spiro[2.2]pentane, and the like. (Cycloalkyl)oxy refers to -O-cycloalkyl. (Cycloalkyl)thio refers to -S-cycloalkyl. This term also includes oxidized forms of sulfur, such as -S(O)-cycloalkyl, or -S(O)2-cycloalkyl.

[0028] Alkenyl refers to straight-chain and branched-chain and cycloalkenyl groups, as defined above, having one or more double bonds between two carbon atoms. Alkenyl groups can have from 2 to about 12 carbon atoms, or in some embodiments, from 1 to about 10 carbon atoms, or in other embodiments, from 1 to about 6 carbon atoms, or in other embodiments, 1, 2, 3, or 4 carbon atoms. Alkenyl groups can be substituted or unsubstituted. Representative substituted alkenyl groups can be mono-substituted or substituted more than once, such as mono-, di-, or tri-substituted. Examples of alkenyl groups include, but are not limited to, vinyl, allyl, -CH=CH(CH), -CH=C(CH), -C(CH)=CH, cyclopentenyl, cyclohexenyl, butadienyl, pentadienyl, and hexadienyl, among others.

[0029] Alkynyl groups refer to straight-chain and branched-chain alkynyl groups, as defined above, and have one or more triple bonds between two carbon atoms. Alkynyl groups can have from 2 to about 12 carbon atoms, or in some embodiments, from 1 to about 10 carbon atoms, or in other embodiments, from 1 to about 6 carbon atoms, or in other embodiments, 1, 2, 3, or 4 carbon atoms. Alkynyl groups can be substituted or unsubstituted. Representative substituted alkynyl groups can be mono-substituted or substituted more than once, such as, but not limited to, mono-, di-, or tri-substituted. Representative alkynyl groups include, but are not limited to, ethynyl, propargyl, and -C≡C(CH), among others.

[0030] Aryl groups are cyclic aromatic hydrocarbons, including single- and multiple-ring compounds, including multiple-ring compounds containing separated and / or fused aryl groups. Aryl groups can contain from 6 to about 18 ring carbons, or in some embodiments, from 6 to 14 ring carbons, or even from 6 to 10 ring carbons in other embodiments. Aryl groups also include heteroaryl groups, which are aromatic ring compounds containing five or more ring members in which one or more ring carbon atoms are replaced with heteroatoms, such as, but not limited to, N, O, and S. Aryl groups can be substituted or unsubstituted. Representative substituted aryl groups can be mono-substituted or substituted more than once, such as, but not limited to, mono-, di-, tri-substituted, and the like. Aryl groups include, but are not limited to, phenyl, biphenylenyl, triphenylenyl, naphthyl, anthryl, and pyrenyl groups. Aryloxy refers to -O-aryl. Arylthio refers to -S-aryl, where aryl is as defined herein. This term also includes oxidized forms of sulfur, such as -S(O)-aryl, or -S(O)-aryl. Heteroaryloxy refers to -O-heteroaryl. Heteroarylthio refers to -S-heteroaryl. This term also includes oxidized forms of sulfur, such as -S(O)-heteroaryl, or -S(O)-heteroaryl.

[0031] Suitable heterocyclyl groups include cyclic groups having atoms of at least two different elements as members of its ring, one or more of which is a heteroatom, such as, but not limited to, N, O, or S. Heterocyclyl groups can contain from 3 to about 20 ring members, or in some embodiments, from 3 to 18, or from about 3 to 15, from 3 to 12, from 3 to 10, or from 3 to 6 ring members. The ring system in a heterocyclyl group can be unsaturated, partially saturated, and / or saturated. Heterocyclyl groups can be substituted or unsubstituted. Representative substituted heterocyclyl groups can be mono-substituted or substituted more than once, such as, but not limited to, mono-, di-, or tri-substituted. Exemplary heterocyclyl groups include, but are not limited to, pyrrolidinyl, tetrahydrofuryl, dihydrofuryl, tetrahydrothienyl, tetrahydrothiopyranyl, piperidyl, morpholinyl, thiomorpholinyl, thioxanyl, piperazinyl, azetidinyl, aziridinyl, imidazolidinyl, pyrazolidinyl, thiazolidinyl, tetrahydrothiophenyl, tetrahydrofuranyl, dioxolyl, furanyl, thiophenyl, pyrrolyl, imidazolyl. , pyrazolyl, pyrazolinyl, triazolyl, tetrazolyl, oxazolyl, isoxazolyl, thiazolyl, thiazolinyl, oxetanyl, thietanyl, homopiperidyl, oxepanyl, thiepanyl, oxazepinyl, diazepinyl, thiazepinyl, 1,2,3,6-tetrahydropyridyl, indolinyl, 2H-pyranyl, 4H-pyranyl, dioxolanyl, dioxanyl, purinyl, quinolidinyl, cinnolinyl, phthalazinyl, pteridinyl, and benzothiazolyl groups. Heterocyclyloxy refers to -O-heterocyclyl. Heterocyclylthio refers to -S-heterocyclyl. This term also includes oxidized forms of sulfur, such as -S(O)-heterocyclyl, or -S(O)2-heterocyclyl, etc.

[0032] Polycyclic or polycyclyl groups refer to two or more rings, where two or more carbons are common to two adjacent rings, where the rings are "fused rings," and where the rings are joined by one common carbon atom, they are "spiro" ring systems. Rings joined through non-adjacent atoms are "bridged" rings. Polycyclic groups may be substituted or unsubstituted. Representative polycyclic groups may be substituted one or more times.

[0033] Halogen groups include F, Cl, Br, and I; nitro groups refer to -NO; cyano groups refer to -CN; isocyano groups refer to -N≡C; epoxy groups encompass structures in which an oxygen atom is attached directly to two adjacent or non-adjacent carbon atoms of a carbon chain or ring system, which is essentially a cyclic ether structure. Epoxides are cyclic ethers with a three-atom ring.

[0034] An alkoxy group is a substituted or unsubstituted alkyl group, as defined above, single-bonded to oxygen. The alkoxy group may be substituted or unsubstituted. Representative substituted alkoxy groups may be substituted one or more times. Exemplary alkoxy groups include, but are not limited to, methoxy, ethoxy, propoxy, butoxy, pentoxy, hexoxy, isopropoxy, sec-butoxy, tert-butoxy, cyclopropyloxy, cyclobutyloxy, cyclopentyloxy, and cyclohexyloxy groups.

[0035] Thiol refers to -SH. Thiocarbonyl refers to (=S). Sulfonyl refers to -SO2-alkyl, -SO2-substituted alkyl, -SO2-cycloalkyl, -SO2-substituted cycloalkyl, -SO2-aryl, -SO2-substituted aryl, -SO2-heteroaryl, -SO2-substituted heteroaryl, -SO2-heterocyclyl, and -SO2-substituted heterocyclyl. Sulfonylamino refers to -NR a SO2 alkyl, -NR a SO2-substituted alkyl, -NR a SO2 cycloalkyl, --NR a SO2-substituted cycloalkyl, -NR a SO2 aryl, -NR aSO2-substituted aryl, --NR a SO2 heteroaryl, -NR a SO2-substituted heteroaryl, -NR a SO2 heterocyclyl, -NR a Refers to a SO2-substituted heterocyclyl, where each R a are as defined herein without regard to

[0036] Carboxyl refers to -COOH or a salt thereof. Carboxy ester refers to -C(O)O-alkyl, -C(O)O-substituted alkyl, -C(O)O-aryl, -C(O)O-substituted aryl, -C(O)β-cycloalkyl, -C(O)O-substituted cycloalkyl, -C(O)O-heteroaryl, -C(O)O-substituted heteroaryl, -C(O)O-heterocyclyl, and -C(O)O-substituted heterocyclyl. (Carboxy ester)amino refers to -NR a -C(O)O-alkyl, -NR a -C(O)O-substituted alkyl, -NR a -C(O)O-aryl, -NR a -C(O)O-substituted aryl, -NR a -C(O)β-cycloalkyl, --NR a -C(O)O-substituted cycloalkyl, -NR a —C(O)O-heteroaryl, —NR a -C(O)O-substituted heteroaryl, -NR a -C(O)O-heterocyclyl, and -NR a refers to a —C(O)O-substituted heterocyclyl, where R a are as listed herein. (Carboxy ester)oxy refers to -OC(O)O-alkyl, -OC(O)O-substituted alkyl, -OC(O)O-aryl, -OC(O)O-substituted aryl, -OC(O)β-cycloalkyl, -OC(O)O-substituted cycloalkyl, -OC(O)O-heteroaryl, -OC(O)O-substituted heteroaryl, -OC(O)O-heterocyclyl, and -OC(O)O-substituted heterocyclyl. Oxo refers to (=O).

[0037] The terms "amine" and "amino" refer to derivatives of ammonia in which one or more hydrogen atoms are replaced by substituents, including, but not limited to, alkyl, alkenyl, aryl, and heterocyclyl groups. A carbamate group refers to -O(C=O)NR1R2, where R1 and R2 are independently hydrogen, an aliphatic group, an aryl group, or a heterocyclyl group.

[0038] Aminocarbonyl is -C(O)N(R b )2, where each R b are independently selected from hydrogen, alkyl, substituted alkyl, aryl, substituted aryl, cycloalkyl, substituted cycloalkyl, heteroaryl, substituted heteroaryl, heterocyclyl, and substituted heterocyclyl; and each R b Both R b may optionally be joined together with the nitrogen attached thereto to form a heterocyclyl or substituted heterocyclyl group, provided that neither of the nitrogen atoms is hydrogen. Aminocarbonylalkyl is an -alkylC(O)N(R b )2, where each R b are independently selected from hydrogen, alkyl, substituted alkyl, aryl, substituted aryl, cycloalkyl, substituted cycloalkyl, heteroaryl, substituted heteroaryl, heterocyclyl, and substituted heterocyclyl; and each R b Both R b may optionally be joined together with the nitrogen attached thereto to form a heterocyclyl or substituted heterocyclyl group, provided that neither of the nitrogen atoms is hydrogen. a C(O)N(R b )2, where R a and each R b is as defined herein. Aminodicarbonylamino is -NR a C(O)C(O)N(R b )2, where R a and each R b is as defined herein. Aminocarbonyloxy is —OC(O)N(R b )2, where each R bis defined herein independently. Aminosulfonyl is -SO2N(R b )2, where each R b appears to be defined here irrespectively.

[0039] Imino is -N=R c He mentioned that R c may be selected from hydrogen, aminocarbonylalkyloxy, substituted aminocarbonylalkyloxy, aminocarbonylalkylamino, and substituted aminocarbonylalkylamino.

[0040] The term "optionally substituted" means that the preceding group can be substituted or unsubstituted. When substituted, the substituents of the "optionally substituted" group can include, without limitation, the following groups: lower alkyl, lower alkenyl, lower alkynyl, lower alkanoyl, lower heteroalkyl, lower heterocycloalkyl, lower haloalkyl, lower haloalkenyl, lower haloalkynyl, lower perhaloalkyl, lower perhaloalkoxy, lower cycloalkyl, phenyl, aryl, aryloxy, lower alkoxy, lower haloalkoxy, oxo, lower acyloxy, carbonyl, carboxyl, lower alkylcarbonyl, lower carboxyl ... The substituents may include, alone or in combination, one or more substituents independently selected from the set of carboxyesters, lower carboxamido, cyano, hydrogen, halogen, hydroxy, amino, lower alkylamino, arylamino, amido, nitro, thiol, lower alkylthio, lower haloalkylthio, lower perhaloalkylthio, arylthio, sulfonate, sulfonic acid, trisubstituted silyl, N, SH, SCH, C(O)CH, COCH, COH, pyridinyl, thiophene, furanyl, lower carbamates, and lower ureas, or a specific specified group. Two substituents may be joined together to form a fused 5-, 6-, or 7-membered carbocyclic or heterocyclic ring consisting of zero to three heteroatoms, e.g., methylenedioxy or ethylenedioxy. Optionally substituted groups can be unsubstituted (e.g., -CHCH), fully substituted (e.g., -CFCF), monosubstituted (e.g., -CHCHF), or substituted at any level between fully and monosubstituted (e.g., -CHCF). When substituents are recited without qualification as to substitution, both substituted and unsubstituted forms are encompassed. When a substituent is designated as "substituted," the substituted form is specifically intended. Furthermore, different sets of optional substituents for a particular moiety can be defined as desired; in these cases, the optional substitution will be as defined, often immediately following the phrase "optionally substituted with."

[0041] Pharmaceutically acceptable salts of the compounds described herein include the conventional non-toxic salts or quaternary ammonium salts of the compounds, e.g., from non-toxic organic or inorganic acids. For example, such conventional non-toxic salts include those derived from inorganic acids such as hydrochloric acid, hydrobromic acid, sulfuric acid, sulfamic acid, phosphoric acid, nitric acid, and the like; and organic acids such as acetic acid, propionic acid, succinic acid, glycolic acid, stearic acid, lactic acid, malic acid, tartaric acid, citric acid, ascorbic acid, palmitic acid, maleic acid, hydroxymaleic acid, phenylacetic acid, glutamic acid, benzoic acid, salicylic acid, sulfanilic acid, 2-acetoxybenzoic acid, fumaric acid, toluenesulfonic acid, methanesulfonic acid, ethanedisulfonic acid, oxalic acid, isothionic acid, and the like. In other cases, the described compounds may contain one or more acidic functional groups and are therefore capable of forming pharmaceutically acceptable salts with pharmaceutically acceptable bases. These salts can likewise be prepared in situ during the administration vehicle or dosage form manufacturing process, or by separately reacting the purified compound in its free acid form with a suitable base, such as a hydroxide, carbonate, or bicarbonate of a pharmaceutically acceptable metal cation, with ammonia, or with a pharmaceutically acceptable organic primary, secondary, or tertiary amine. Representative alkali or alkaline earth salts include lithium, sodium, potassium, calcium, magnesium, and aluminum salts, and the like. Representative organic amines useful for forming base addition salts include ethylamine, diethylamine, ethylenediamine, ethanolamine, diethanolamine, piperazine, and the like.

[0042] Herein, the term "treatment" is used interchangeably with the term "therapy" and refers to both 1) therapeutic procedures or measures that cure, slow, alleviate symptoms, and / or halt the progression of a diagnosed pathological condition, disease, or disorder, and 2) prophylactic / preventative measures. Those in need of treatment can include individuals who already have a particular medical disease or disorder as well as those who may eventually acquire the disorder (i.e., those requiring preventative measures).

[0043] As used herein, the term "subject" refers to any individual or patient on whom the subject methods are performed. Generally, subjects are human, although as will be understood by those in the art, subjects can be animals.

[0044] Terms such as "therapeutically effective amount," "effective dose," "therapeutically effective dose," "effective amount," or the like refer to an amount of a subject compound that elicits a desired biological or medical response in a tissue, system, animal, or human upon administration of the compound. Generally, the response is either an improvement in symptoms or a desired biological outcome in the patient. Such an amount should be sufficient to inhibit MIF activity.

[0045] Also disclosed herein are pharmaceutical compositions comprising a compound having the structure of Formula (I). The term "pharmaceutically acceptable carrier" refers to a non-toxic carrier that can be administered to a patient together with a compound of the present disclosure and that does not destroy its pharmacological activity. Pharmaceutically acceptable carriers that can be used in these compositions include, but are not limited to, ion exchangers, alumina, aluminum stearate, lecithin, serum proteins such as human serum albumin, buffer substances such as phosphates, glycine, sorbic acid, potassium sorbate, partial glyceride mixtures of saturated vegetable fatty acids, water, salts or electrolytes such as protamine sulfate, disodium hydrogen phosphate, potassium hydrogen phosphate, sodium chloride, zinc salts, colloidal silica, magnesium trisilicate, polyvinylpyrrolidone, cellulose-based materials, polyethylene glycol, sodium carboxymethylcellulose, polyacrylates, waxes, polyethylene-polyoxypropylene block polymers, polyethylene glycol, and wool fat.

[0046] Pharmaceutically acceptable carriers that may be used in the pharmaceutical compositions of the present disclosure include, but are not limited to, ion exchangers, alumina, aluminum stearate, lecithin, serum proteins such as human serum albumin, buffer substances such as phosphates, glycine, sorbic acid, potassium sorbate, partial glyceride mixtures of saturated vegetable fatty acids, water, salts or electrolytes such as protamine sulfate, disodium hydrogen phosphate, potassium hydrogen phosphate, sodium chloride, zinc salts, colloidal silica, magnesium trisilicate, polyvinylpyrrolidone, cellulose-based materials, polyethylene glycol, sodium carboxymethylcellulose, polyacrylates, waxes, polyethylene-polyoxypropylene-block polymers, wool fat and self-emulsifying drug delivery systems (SEDDS) such as α-tocopherol, polyethylene glycol 1000 succinate, or other similar polymer delivery matrices.

[0047] For pharmaceutical compositions containing only the compounds described herein as the active ingredient, methods for administering these compositions may additionally include administering to a subject an additional drug or therapy. Such therapies include, but are not limited to, anemia therapy, diabetes therapy, hypertension therapy, cholesterol therapy, neuropharmacological drugs, drugs that regulate cardiovascular function, drugs that regulate inflammation, immune function, or blood cell production; hormones and antagonists, drugs that affect gastrointestinal function, chemotherapeutics for microbial diseases, and / or chemotherapeutics for neoplastic diseases. Other pharmacological therapies may include any other drug or biologic found in any drug class. For example, other drug classes may include allergy / cold / ENT therapies, analgesics, anesthetics, anti-inflammatory drugs, antimicrobials, antivirals, asthma / pulmonary therapies, cardiovascular therapies, dermatological therapies, endocrine / metabolic therapies, gastrointestinal therapies, cancer therapies, immunological therapies, neurological therapies, ophthalmological therapies, psychiatric therapies, or rheumatological therapies. Other examples of drugs or therapies that can be administered in conjunction with the compounds described herein include matrix metalloproteinase inhibitors, lipoxygenase inhibitors, cytokine antagonists, immunosuppressants, cytokines, growth factors, immunomodulators, prostaglandins, or anti-vascular hyperproliferation compounds.

[0048] As used herein, the term "therapeutically effective amount" refers to an amount of an active compound or pharmaceutical agent that elicits the biological or medicinal response in a tissue, system, animal, individual, or human that is desired by a researcher, veterinarian, medical doctor, or other clinician, including one or more of the following: (1) disease prevention; e.g., preventing a disease, condition, or disorder in an individual who may be predisposed to the disease, condition, or disorder but who has not yet experienced or displayed the pathology or symptoms of the disease; (2) disease inhibition; e.g., inhibiting a disease, condition, or disorder (i.e., preventing further development of the pathology and / or symptomatology) in an individual who is experiencing or displaying the pathology or symptomatology of the disease, condition, or disorder; and (3) disease amelioration; e.g., ameliorating a disease, condition, or disorder (i.e., reversing the pathology and / or symptomatology) in an individual who is experiencing or displaying the pathology or symptomatology of the disease, condition, or disorder.

[0049] As used herein, the terms "combination," "combining," and related terms refer to the simultaneous or sequential administration of therapeutic agents according to this disclosure. For example, a described compound may be administered simultaneously or sequentially with another therapeutic agent in separate unit dosage forms, or together in a single unit dosage form. Accordingly, the present disclosure provides a single unit dosage form comprising a described compound, an additional therapeutic agent, and a pharmaceutically acceptable carrier, adjuvant, or vehicle. Two or more agents are typically considered to be administered "in combination" when a patient or individual is exposed to both agents simultaneously. In many embodiments, two or more agents are considered to be administered "in combination" when a patient or individual simultaneously exhibits therapeutically relevant levels of the agents in specific target tissues or samples (e.g., in the brain, in serum, etc.).

[0050] When the compounds of this disclosure are administered in combination therapy with other agents, they may be administered sequentially or concurrently to a patient. Alternatively, a pharmaceutical or prophylactic composition according to this disclosure includes a combination of ivermectin, or any other compound described herein, and another therapeutic or prophylactic agent. Additional therapeutic agents that are normally administered to treat a particular disease or condition may be referred to as "agents appropriate for the disease or condition being treated."

[0051] The compounds utilized in the compositions and methods of this disclosure can also be modified by attaching appropriate functional groups to enhance selective biological properties. Such modifications are known in the art and may increase biological penetration into a given biological system (e.g., blood, lymphatic system, or central nervous system), increase oral availability, increase solubility to allow administration by injection, alter metabolism, and / or alter excretion rate.

[0052] According to a preferred embodiment, the compositions of this disclosure are formulated for pharmaceutical administration to a subject or patient, e.g., a mammal, preferably a human being. Such pharmaceutical compositions are used to ameliorate, treat, or prevent any of the disorders described herein in a subject.

[0053] The agents of the present disclosure are often administered as pharmaceutical compositions containing an active therapeutic agent, i.e., and various other pharmaceutically acceptable ingredients. See Remington's Pharmaceutical Science (15th ed., Mack Publishing Company, Easton, Pa., 1980). The preferred form depends on the intended mode of administration and therapeutic application. Depending on the desired formulation, the composition may also contain pharmaceutically acceptable, non-toxic carriers or diluents, which are defined as vehicles commonly used to formulate pharmaceutical compositions for administration to animals or humans. The diluent is selected so as not to affect the biological activity of the combination. Examples of such diluents are distilled water, physiological phosphate-buffered saline, Ringer's solution, dextrose solution, and Hank's solution. In addition, the pharmaceutical composition or formulation may also include other carriers, adjuvants, or nontoxic, nontherapeutic, nonimmunogenic stabilizers and the like.

[0054] In some embodiments, the present disclosure provides pharmaceutically acceptable compositions comprising a therapeutically effective amount of one or more of the described compounds, formulated together with one or more pharmaceutically acceptable carriers (excipients) and / or diluents, for use in treating the diseases described herein, including, but not limited to, stroke, ischemia, Alzheimer's disease, ankylosing spondylitis, arthritis, osteoarthritis, rheumatoid arthritis, psoriatic arthritis, asthma atherosclerosis, Crohn's disease, colitis, dermatitis diverticulitis, fibromyalgia, hepatitis, irritable bowel syndrome, systemic lupus erythematous, nephritis, ulcerative colitis, and Parkinson's disease. While the described compounds can be administered alone, they are preferably administered as pharmaceutical formulations (compositions) as described herein. The described compounds, by analogy with other drugs, may be formulated for administration in any convenient way for use in human or veterinary medicine.

[0055] As described in detail, the pharmaceutical compositions of the present disclosure may be specially formulated for administration in solid or liquid form, including those adapted for oral administration, e.g., drenches (aqueous or non-aqueous solutions or suspensions), tablets, e.g., buccal, sublingual, and those targeted for systemic absorption, boluses for application to the tongue, powders, granules, pastes; parenteral administration, e.g., by subcutaneous, intramuscular, intravenous, or epidural injection, e.g., as a sterile solution or suspension, or sustained-release formulation; topical administration, e.g., as a cream, ointment, or controlled-release patch or spray applied to the skin, lungs, or oral cavity; intravaginally or rectally, e.g., as a pessary, cream, or foam; sublingually; ophthalmically; transdermally; or nasally, to the lungs and other mucosal surfaces.

[0056] Wetting agents, emulsifying agents and lubricating agents, 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.

[0057] Examples of pharmaceutically acceptable antioxidants include: water-soluble antioxidants such as ascorbic acid, cysteine ​​hydrochloride, sodium bisulfate, sodium metabisulfite, sodium sulfite, and the like; oil-soluble antioxidants such as ascorbyl palmitate, butylhydroxyanisole (BHA), butylhydroxytoluene (BHT), lecithin, propyl gallate, alpha tocopherol, and the like; and metal chelating agents such as citric acid, ethylenediaminetetraacetic acid (EDTA), sorbitol, tartaric acid, phosphoric acid, and the like.

[0058] Formulations for use in accordance with the present disclosure include those suitable for oral, nasal, topical (including buccal and sublingual), rectal, vaginal, and / or parenteral administration. The formulations may conveniently be presented in unit dosage form and may be prepared by any method well known in the art of pharmacy. The amount of active ingredient that can be combined with a carrier material to produce a single dosage form will vary depending on the host treated and the particular mode of administration. The amount of active ingredient that can be combined with a carrier material to produce a single dosage form will generally be that amount of compound that produces a therapeutic effect. This amount often ranges from about 1% to about 99% of the active ingredient. In some embodiments, this amount will range from about 5% to about 70%, from about 10% to about 50%, or from about 20% to about 40%.

[0059] In certain embodiments, the formulations described herein comprise an excipient selected from the group consisting of cyclodextrins, liposomes, micelle-forming agents, e.g., bile acids, and polymeric carriers, e.g., polyesters and polyanhydrides; and a compound of the present disclosure. In certain embodiments, the formulations described herein make the compounds of the present disclosure orally bioavailable.

[0060] Methods of preparing formulations or compositions containing the described compounds include the step of bringing into association a compound of the present disclosure with the carrier and, optionally, one or more accessory ingredients. In general, the formulations may be prepared by uniformly and intimately bringing into association a compound of the present disclosure with liquid carriers, or finely divided solid carriers, or both, and then, if necessary, shaping the product.

[0061] The pharmaceutical compositions may be in the form of sterile injectable preparations, for example, as sterile injectable aqueous or oleaginous suspensions. These suspensions may be formulated according to techniques known in the art using suitable dispersing or wetting agents (e.g., Tween 80, Cremophor RH40, Cremophor E1, and the like) and suspending agents. The sterile injectable preparation may also be a sterile injectable solution or suspension in a non-toxic, parenterally acceptable diluent or solvent, for example, as a solution in 1,3-butanediol. Among the acceptable vehicles and solvents that may be employed are mannitol, water, Ringer's solution, and isotonic sodium chloride solution. Additionally, sterile, fixed oils are conventionally employed as solvents or suspending media. For this purpose, any bland fixed oil, including synthetic mono- or diglycerides, may be employed. Fatty acids, such as oleic acid and its glyceride derivatives, are useful in the preparation of injectables, as are natural pharmaceutically acceptable oils, such as olive oil or castor oil, especially their polyoxyethylated versions. These oil solutions or suspensions may also contain long-chain alcohol diluents or dispersants, such as those listed in the Pharmacopeia Helvetica (Swiss Pharmacopoeia), or similar alcohols. Other commonly used surfactants commonly used in the manufacture of pharmaceutically acceptable solid, liquid, or other dosage forms, such as Tweens, Spans, and other emulsifiers or bioavailability enhancers, may also be used for formulation purposes.

[0062] In some cases, in order to prolong the effect of a drug, it may be desirable to delay the absorption of the drug from subcutaneous or intramuscular injection.This can be achieved by using a liquid suspension of crystalline or amorphous material with low water solubility.The absorption rate of the drug then depends on its dissolution rate, which may further depend on the crystal size and crystalline form.Alternatively, delayed absorption of parenterally administered drug forms can be achieved by dissolving or suspending the drug in an oil vehicle.

[0063] Injectable depot forms are made by forming microencapsule matrices of the described compounds in biodegradable polymers, such as polylactide-polyglycolide. The drug release rate can be controlled by the ratio of drug to polymer and the properties of the particular polymer employed. Examples of other biodegradable polymers include poly(orthoesters) and poly(anhydrides). Depot injectable formulations can also be prepared by entrapping the drug in liposomes or microemulsions that are compatible with body tissues.

[0064] The pharmaceutical composition of this disclosure can be orally administered in any orally acceptable dosage form, including, but not limited to, capsules, tablets, and aqueous suspensions and solutions.For oral tablets, carriers are commonly used and include lactose and cornstarch.Lubricants, such as magnesium stearate, are also typically added.For oral administration in capsule form, useful diluents include lactose and dried cornstarch.When aqueous suspensions and solutions and propylene glycol are orally administered, the active ingredient is combined with an emulsifier and suspending agent.If desired, certain sweeteners and / or flavorings and / or colorings can be added.

[0065] Formulations described herein suitable for oral administration may be in the form of capsules, cachets, pills, tablets, lozenges (using a flavored basis, usually sucrose and acacia or tragacanth), powder, granules, or as a solution or suspension in an aqueous or non-aqueous liquid, or as an oil-in-water or water-in-oil liquid emulsion, or as an elixir or syrup, or as a pastille (using an inert base such as gelatin and glycerin, or sucrose and acacia), and / or as mouthwashes and the like, each containing a predetermined amount of a compound of the present disclosure as an active ingredient. The compounds described herein may also be administered as a bolus, electuary, or paste.

[0066] In solid dosage forms for oral administration (such as capsules, tablets, pills, dragees, powders, granules, and the like), the active ingredient may be incorporated into one or more pharmaceutically acceptable carriers, such as sodium citrate or dicalcium phosphate, and / or any of the following: fillers or extenders, such as starch, lactose, sucrose, glucose, mannitol, and / or silicic acid; binders, such as carboxymethylcellulose, alginic acid, gelatin, polyvinylpyrrolidone, sucrose, and / or acacia; humectants, such as glycerol; disintegrating agents, such as agar-agar, calcium carbonate, potato or tapioca starch, alginic acid, certain silicates, and sodium carbonate; solution retarding agents, such as PEG-100, PEG-10 ... The pharmaceutical compositions may be mixed with additives such as sorbent, sorbent powders, sorbent additives, sorbent additives (e.g., sorbent additives ...

[0067] Tablets may be made by compression or molding, optionally with one or more accessory ingredients. Compressed tablets may be prepared using binders (e.g., gelatin or hydroxypropylmethylcellulose), lubricants, inert diluents, preservatives, disintegrants (e.g., sodium starch glycolate or cross-linked carboxymethylcellulose), surfactants, or dispersing agents. Molded tablets may be made in a suitable machine by moistening a mixture of powdered compounds with an inert liquid diluent. When a solid carrier is used, the preparation may be in the form of a tablet, in a hard gelatin capsule in powder or pellet form, or in the form of a troche or lozenge. The amount of solid carrier may, for example, vary from about 25 to 800 mg, preferably from about 25 to 400 mg. When a liquid carrier is used, the preparation may be in the form of, for example, a syrup, emulsion, soft gelatin capsule, sterile injectable liquid such as an ampule, or non-aqueous liquid suspension. Where the composition is in the form of a capsule, any routine encapsulation is suitable, for example using the carriers described above in a hard gelatin capsule shell.

[0068] Tablets and other solid dosage forms, such as dragees, capsules, pills, and granules, can be optionally scored or prepared with coatings and shells, such as enteric coatings and other coatings well known in the pharmaceutical compounding art. They can alternatively or additionally be formulated to provide sustained or controlled release of the active ingredient therein, for example, using hydroxypropylmethylcellulose, other polymer matrices, liposomes, and / or microspheres, with proportions varied to provide a desired release profile. They can be formulated for rapid release, e.g., lyophilized. They can be sterilized, for example, by filtration through a bacteria-retaining filter, or by incorporating a sterilizing agent in the form of a sterile solid composition that can be dissolved in sterile water or some other sterile injectable medium immediately before use. These compositions can also optionally contain opacifying agents and can be of a composition that releases the active ingredient(s) only, or preferentially, in a certain part of the digestive tract, optionally in a delayed manner. Examples of embedding compositions that can be used include polymeric substances and waxes. The active ingredient can also be in microencapsulated form, if appropriate, with one or more of the above-mentioned excipients.

[0069] The liquid dosage form for oral administration of the compounds of the present disclosure includes pharmaceutically acceptable emulsions, microemulsions, solutions, suspensions, syrups and elixirs.In addition to the active ingredient, the liquid dosage form includes the inert diluent commonly used in the art, such as water or other solvents, solubilizers and emulsifiers, such as ethyl alcohol, isopropyl alcohol, ethyl carbonate, ethyl acetate, benzyl alcohol, benzyl benzoate, propylene glycol, 1,3-butylene glycol, oils (especially cottonseed, groundnut, corn, germ, olive, castor and sesame oils), glycerol, tetrahydrofuryl alcohol, polyethylene glycol and fatty acid esters of sorbitan, and mixtures thereof.

[0070] Besides inert diluents, the oral compositions can also include adjuvants such as wetting agents, emulsifying and suspending agents, sweetening, flavoring, coloring, perfuming and preservative agents.

[0071] Suspensions may contain, in addition to the active compound, suspending agents such as, for example, ethoxylated isostearyl alcohols, polyoxyethylene sorbitol and sorbitan esters, microcrystalline cellulose, aluminum metahydroxide, bentonite, agar-agar, and tragacanth, and mixtures thereof.

[0072] The pharmaceutical composition of this disclosure can also be administered in the form of suppositories for rectal administration.These compositions can be prepared by mixing the compound of this disclosure with a suitable non-irritating excipient that is solid at room temperature but liquid at rectal temperature, and therefore melts in the rectum to release the active ingredient.Such materials include, but are not limited to, cocoa butter, beeswax and polyethylene glycol.

[0073] Topical administration of the pharmaceutical compositions of this disclosure is particularly useful when the desired treatment involves areas or organs easily accessible by topical application. For topical application to the skin, the pharmaceutical composition should be formulated with a suitable ointment containing the active ingredient suspended or dissolved in a carrier. Carriers for topical administration of the compounds of this disclosure include, but are not limited to, mineral oil, liquid petroleum, white petrolatum, propylene glycol, polyoxyethylene polyoxypropylene compounds, emulsifying wax, and water. Alternatively, the pharmaceutical composition can be formulated with a suitable lotion or cream containing the active compound suspended or dissolved in a carrier. Suitable carriers include, but are not limited to, mineral oil, sorbitan monostearate, polysorbate 60, cetyl esters wax, cetearyl alcohol, 2-octyldodecanol, benzyl alcohol, and water. The pharmaceutical compositions of this disclosure can also be topically applied to the lower intestinal tract by rectal suppository formulation or in a suitable enema formulation. Topically administered transdermal patches are also included in this disclosure.

[0074] The pharmaceutical compositions of this disclosure may be administered by nasal aerosol or inhalation. Such compositions are prepared according to techniques well known in the pharmaceutical formulation art and may be prepared as solutions in saline using benzyl alcohol or other suitable preservatives, absorption enhancers to enhance bioavailability, fluorocarbons, and / or other solubilizing or dispersing agents well known in the art.

[0075] For ophthalmic use, the pharmaceutical composition may be formulated as a micronized suspension in isotonic, pH-adjusted sterile saline, or preferably as a solution in isotonic, pH-adjusted sterile saline, either with or without a preservative, such as benzylalkonium chloride. Alternatively, for ophthalmic use, the pharmaceutical composition may be formulated in an ointment, such as petrolatum.

[0076] Transdermal patches have the additional benefit of providing controlled delivery of the compounds of the present disclosure into the body.The compound can be dissolved or dispersed in a suitable medium to produce such a dosage form.Absorption enhancers can also be used to increase the flux of the compound across the skin.The rate of such flux can also be controlled by providing a rate-controlling membrane or dispersing the compound in a polymer matrix or gel.

[0077] Examples of suitable aqueous and non-aqueous carriers that may be employed in the pharmaceutical compositions of the present disclosure include water, ethanol, polyols (such as, for example, glycerol, propylene glycol, polyethylene glycol, and the like) and suitable mixtures thereof, vegetable oils such as olive oil, and injectable organic esters such as ethyl oleate. Proper fluidity can be maintained by the use of coating materials, such as lecithin, by the maintenance of the required particle size in the case of dispersions, and by the use of surfactants.

[0078] Such compositions may also contain adjuvants, such as preservatives, wetting agents, emulsifying agents, and dispersing agents. The inclusion of one or more antibacterial and / or antifungal agents, such as parabens, chlorobutanol, phenol sorbic acid, and the like, may be desirable in certain embodiments. Alternatively, or additionally, it may be desirable to include isotonic agents, such as sugars, sodium chloride, and the like, in the compositions. In addition, prolonged absorption of the injectable pharmaceutical form can be brought about by the inclusion of agents which delay absorption, such as aluminum monostearate and gelatin.

[0079] In certain embodiments, the described compounds or pharmaceutical formulations are administered orally. In other embodiments, the described compounds or pharmaceutical formulations are administered intravenously. Alternative routes of administration include sublingual, intramuscular, and transdermal administration.

[0080] When the compounds described herein are administered to humans and animals as medicines, they can be given per se or as pharmaceutical compositions containing, for example, from 0.1% to 99.5% (more preferably, from 0.5% to 90%) of the active ingredient in combination with a pharmaceutically acceptable carrier.

[0081] The preparations described herein may be given orally, parenterally, topically, or rectally. They are, of course, given in a form appropriate for the relevant route of administration. For example, they may be given in the form of tablets or capsules, as injections, inhalations, eye lotions, ointments, suppositories, etc., administered by injection, infusion, or inhalation; topically by lotion or ointment; and rectally by suppositories. Oral administration is preferred.

[0082] Such compounds may be administered to humans and other animals for therapy by any suitable route of administration, including orally, nasally, rectally, vaginally, parenterally, intracisternally and topically, such as by powder, ointment or drops, including bucally and sublingually.

[0083] Regardless of the route of administration selected, the compounds described herein, which may be used in a suitable hydrated form, and / or pharmaceutical compositions of the present disclosure, are formulated into pharmaceutically acceptable dosage forms by conventional methods known to those skilled in the art.

[0084] Actual dosage levels of the active ingredients in the pharmaceutical compositions of the present disclosure may be varied to obtain an amount of the active ingredient that is effective for a particular patient, composition, and mode of administration to achieve a desired therapeutic response without toxicity to the patient.

[0085] The terms "administration of" and / or "administering" should be understood to mean providing a therapeutically effective amount of a pharmaceutical composition to a subject in need of treatment. The route of administration can be enteral, topical, or parenteral. As such, routes of administration include, but are not limited to, intracutaneous, subcutaneous, intravenous, intraperitoneal, intraarterial, intrathecal, intracapsular, intraorbital, intracardiac, intradermal, transdermal, transtracheal, subcuticular, intraarticular, subcapsular, subarachnoid, intrathecal, and intrasternal, oral, sublingual, buccal, rectal, vaginal, nasal, ocular administration, as well as injection, inhalation, and nebulization.

[0086] The crystal structures of the FKBP-FK506-calcineurin and FKBP-rapamycin-TOR complexes revealed that both FK506 and rapamycin can be divided into two functional domains, the "FKBP-binding domain" (FKBD) and the "effector" domain, which mediate their interactions with calcineurin and TOR, respectively. While extensive protein-protein interactions between FKBP and calcineurin exist in their ternary complexes, there are far fewer interactions between FKBP and TOR, suggesting that a key role of FKBP in rapamycin-mediated TOR inhibition is to bind the drug's FKBD and present its effector domain to TOR.

[0087] Comparison of the structures of FK506 and rapamycin revealed that they share a nearly identical FKBD but each possesses a distinct effector domain. By swapping the effector domain of FK506 with that of rapamycin, the target could be changed from calcineurin to TOR, which bears no sequence, function, or structural similarity to either. Additionally, other proteins could be targeted by grafting new structures onto the FKBDs of FK506 and rapamycin. Thus, the generation of new compounds with novel target specificities could be achieved by grafting sufficiently large combinatorial libraries onto the FKBD in combination with proteome-wide screening, where each compound in the library is tested against every protein in the human proteome. [ka]

[0088] In some embodiments, provided herein are macrocycles according to Formula (I), which include an FKBD, an effector domain, a first linker, and a second linker, wherein the FKBD, the effector domain, the first linker, and the second linker together form a macrocycle.

[0089] In some embodiments, provided herein are macrocyclic compounds according to Formula (II), or optically pure stereoisomers or pharmaceutically acceptable salts thereof. [ka]

[0090] B can be CH, NH, NMe, O, S, or S(O)2; X can be O, NH, or NMe; E can be CH or N; n is an integer selected from 0 to 4; and m is an integer selected from 1 to 10. In this formula, AA represents natural and unnatural amino acids, each of which can be selected from Table 4 below.

[0091] In some embodiments, m can be 1. In some embodiments, m can be 2. In some embodiments, m can be 3. In some embodiments, m can be 4. In some embodiments, m can be 5. In some embodiments, m can be 6. In some embodiments, m can be 7. In some embodiments, m can be 8. In some embodiments, m can be 9. In some embodiments, m can be 10. In specific embodiments, m is 3 or 4.

[0092] Each R 1 is H, halogen, hydroxyl, C 1-20 Alkyl, N3, NH2, NO2, CF3, OCF3, OCHF2, COC 1-20 Alkyl, and CO2C 1-20 alkyl. R 2 are H, halogen, hydroxyl, N3, NH2, NO2, CF3, C 1-10 Alkyl, substituted C 1-10 Alkyl, C 1-10 Alkoxy, substituted C 1-10 Alkoxy, acyl, acylamino, acyloxy, acyl C 1-10 Alkyloxy, amino, substituted amino, aminoacyl, aminocarbonyl C 1-10 Alkyl, aminocarbonylamino, aminodicarbonylamino, aminocarbonyloxy, aminosulfonyl, C 6-15 Aryl, substituted C 6-15 Aryl, C 6-15 Aryloxy, substituted C 6-15 Aryloxy, C 6-15Arylthio, substituted C 6-15 Arylthio, carboxyl, carboxy ester, (carboxy ester) amino, (carboxy ester) oxy, cyano, C 3-8 Cycloalkyl, substituted C 3-8 Cycloalkyl, (C 3-8 cycloalkyl)oxy, substituted (C 3-8 cycloalkyl)oxy, (C 3-8 cycloalkyl)thio, substituted (C 3-8 cycloalkyl)thio, C 1-10 Heteroaryl, substituted C 1-10 Heteroaryl, C 1-10 Heteroaryloxy, substituted C 1-10 Heteroaryloxy, C 1-10 Heteroarylthio, substituted C 1-10 Heteroarylthio, C 2-10 Heterocyclyl, C 2-10 Substituted heterocyclyl, C 2-10 Heterocyclyloxy, substituted C 2-10 Heterocyclyloxy, C 2-10 Heterocyclylthio, substituted C 2-10 Heterocyclylthio, imino, oxo, sulfonyl, sulfonylamino, thiol, C 1-10 Alkylthio, substituted C 1-10 C optionally substituted with alkylthio and thiocarbonyl 6-15 Aryl and C 1-10 Heteroaryl is selected from the group consisting of:

[0093] V is [ka] Z is a bond, [ka] where R 3 and R 4are each independently selected from the group consisting of hydrogen, hydroxy, halo, alkyl, alkoxy, cycloalkyl, cyano, alkylthio, amino, alkylamino, and dialkylamino; K is O, CHR 5 , C.R. 5 , N and NR 5 where R 5 is hydrogen or alkyl.

[0094] Each L 1 , L 2 , or L 3 can be selected from the group consisting of the structures shown in Table 1 below. [Table 1-1] [Table 1-2] [Table 1-3] [Table 1-4] [Table 1-5] [Table 1-6] [Table 1-7] [Table 1-8] [Table 1-9] [Table 1-10] [Table 1-11] [Table 1-12] [Table 1-13] [Table 1-14] [Table 1-15] *Each R 20 and R 21 is hydrogen, hydroxy, OR 22 , N.R. 23 R 24 , alkyl, arylalkyl, [ka] independently selected from the group consisting of: N is aryl, alkyl, or arylalkyl; where R 22 , R 23 and R 24 are each independently hydrogen or alkyl.

[0095] In some embodiments, the FKBD-containing moiety, prior to incorporation into the macrocycle, has the formula (III): [ka] or an optically pure stereoisomer or a pharmaceutically acceptable salt thereof.

[0096] where L is selected from the structures in Table 1; A is CH, NH, O, or S; each X is independently O, NH, or NMe; E is CH or N; [ka] represents a single bond or a double bond, and n is an integer selected from 0 to 4.

[0097] Each R 1 is H, halogen, hydroxyl, C 1-20Alkyl, N3, NH2, NO2, CF3, OCF3, OCHF2, COC 1-20 Alkyl, and CO2C 1-20 alkyl. R 2 is H, halogen, hydroxyl, C 1-20 Alkyl, N3, NH2, NO2, CF3, OCF3, OCHF2, COC 1-20 Alkyl, and CO2C 1-20 alkyl. R 3 are H, halogen, hydroxyl, N3, NH2, NO2, CF3, C 1-10 Alkyl, substituted C 1-10 Alkyl, C 1-10 Alkoxy, substituted C 1-10 Alkoxy, acyl, acylamino, acyloxy, acyl C 1-10 Alkyloxy, amino, substituted amino, aminoacyl, aminocarbonyl C 1-10 Alkyl, aminocarbonylamino, aminodicarbonylamino, aminocarbonyloxy, aminosulfonyl, C 6-15 Aryl, substituted C 6-15 Aryl, C 6-15 Aryloxy, substituted C 6-15 Aryloxy, C 6-15 Arylthio, substituted C 6-15 Arylthio, carboxyl, carboxy ester, (carboxy ester) amino, (carboxy ester) oxy, cyano, C 3-8 Cycloalkyl, substituted C 3-8 Cycloalkyl, (C 3-8 cycloalkyl)oxy, substituted (C 3-8 cycloalkyl)oxy, (C 3-8 cycloalkyl)thio, substituted (C 3-8 cycloalkyl)thio, C 1-10 Heteroaryl, substituted C 1-10 Heteroaryl, C 1-10 Heteroaryloxy, substituted C 1-10 Heteroaryloxy, C 1-10 Heteroarylthio, substituted C 1-10 Heteroarylthio, C 2-10 Heterocyclyl, C 2-10Substituted heterocyclyl, C 2-10 Heterocyclyloxy, substituted C 2-10 Heterocyclyloxy, C 2-10 Heterocyclylthio, substituted C 2-10 Heterocyclylthio, imino, oxo, sulfonyl, sulfonylamino, thiol, C 1-10 Alkylthio, substituted C 1-10 C optionally substituted with alkylthio and thiocarbonyl 6-15 Aryl and C 1-10 Heteroaryl is selected from the group consisting of:

[0098] V is [ka] Z is a bond, [ka] where R 4 and R 5 are each independently selected from the group consisting of hydrogen, hydroxy, halo, alkyl, alkoxy, cycloalkyl, cyano, alkylthio, amino, alkylamino, and dialkylamino; K is O, CHR 6 , C.R. 6 , N and NR 6 where R 6 is hydrogen or alkyl.

[0099] In some embodiments, the FKBD-containing moiety, prior to incorporation into the macrocycle, can have a structure according to formula (IV), or an optically pure stereoisomer or pharmaceutically acceptable salt thereof. [ka]

[0100] where L is selected from the structures in Table 1; A is CH, NH, O, or S; each X is independently O or NH; E is CH or N; and each R 1 is H, halogen, hydroxyl, C1-20 Alkyl, N3, NH2, NO2, CF3, OCF3, OCHF2, COC 1-20 Alkyl, and CO2C 1-20 alkyl; each R 2 are H, halogen, hydroxyl, N3, NH2, NO2, CF3, C 1-10 Alkyl, substituted C 1-10 Alkyl, C 1-10 Alkoxy, substituted C 1-10 Alkoxy, acyl, acylamino, acyloxy, acyl C 1-10 Alkyloxy, amino, substituted amino, aminoacyl, aminocarbonyl C 1-10 Alkyl, aminocarbonylamino, aminodicarbonylamino, aminocarbonyloxy, aminosulfonyl, C 6-15 Aryl, substituted C 6-15 Aryl, C 6-15 Aryloxy, substituted C 6-15 Aryloxy, C 6-15 Arylthio, substituted C 6-15 Arylthio, carboxyl, carboxy ester, (carboxy ester) amino, (carboxy ester) oxy, cyano, C 3-8 Cycloalkyl, substituted C 3-8 Cycloalkyl, (C 3-8 cycloalkyl)oxy, substituted (C 3-8 cycloalkyl)oxy, (C 3-8 cycloalkyl)thio, substituted (C 3-8 cycloalkyl)thio, C 1-10 Heteroaryl, substituted C 1-10 Heteroaryl, C 1-10 Heteroaryloxy, substituted C 1-10 Heteroaryloxy, C 1-10 Heteroarylthio, substituted C 1-10 Heteroarylthio, C 2-10 Heterocyclyl, C 2-10 Substituted heterocyclyl, C 2-10 Heterocyclyloxy, substituted C 2-10 Heterocyclyloxy, C 2-10 Heterocyclylthio, substituted C 2-10Heterocyclylthio, imino, oxo, sulfonyl, sulfonylamino, thiol, C 1-10 Alkylthio, substituted C 1-10 n is an integer selected from the group consisting of 0 to 4; and m is an integer selected from 0 to 5.

[0101] In some embodiments, the rapafucine compounds of the present disclosure can have a structure according to formula (V), or an optically pure stereoisomer or pharmaceutically acceptable salt thereof. [ka]

[0102] where L is selected from the group in Table 1; A is CH2, NH, NMe, O, S(O)2, or S; each X is independently O, NMe, or NH; and E is CH or N.

[0103] Each R 1 , R 2 , R 3 , and R 4 is H, halogen, hydroxyl, N3, NH2, NO2, CF3, OCF3, OCHF2, COC 1-10 Alkyl, CO2C 1-10 Alkyl, C 3-8 Cycloalkyl, C 2-8 Alkenyl, C 2-8 Alkynyl, C 1-10 Alkoxy, C 6-15 Aryl, C 6-15 Aryloxy, C 6-15 Arylthio, C 2-10 Carboxyl, C 1-10 Alkylamino, thiol, C 1-10 Alkylthio, C 1-10 Alkyl disulfide, C 6-15 Arylthio, C 1-10 Heteroarylthio, (C 3-8 cycloalkyl)thio, C 2-10 Heterocyclylthio, sulfonyl, C 1-10Alkyl sulfonyl, amide, C 1-10 Alkylamide, Selenol, C 1-10 Alkylselenol, C 6-15 Arylselenol, C 1-10 Heteroarylselenols, (C 3-8 cycloalkyl)selenol, C 2-10 Heterocyclylselenol, guanidino, C 1-10 Alkylguanidino, urea, C 1-10 Alkyl urea, ammonium, C 1-10 Alkylammonium, Cyano, C 1-10 Alkyl cyano, C 1-10 Alkyl nitro, adamantine, phosphonate, C 1-10 Alkyl phosphonates, and C 6-15 aryl phosphonates, each of which may be independently selected from the group consisting of H, halogen, hydroxyl, N, NH, NO, CF, C 1-10 Alkyl, substituted C 1-10 Alkyl, C 1-10 Alkoxy, substituted C 1-10 Alkoxy, acyl, acylamino, acyloxy, acyl C 1-10 Alkyloxy, amino, substituted amino, aminoacyl, aminocarbonyl C 1-10 Alkyl, aminocarbonylamino, aminodicarbonylamino, aminocarbonyloxy, aminosulfonyl, C 6-15 Aryl, substituted C 6-15 Aryl, C 6-15 Aryloxy, substituted C 6-15 Aryloxy, C 6-15 Arylthio, substituted C 6-15 Arylthio, carboxyl, carboxy ester, (carboxy ester) amino, (carboxy ester) oxy, cyano, C 3-8 Cycloalkyl, substituted C 3-8 Cycloalkyl, (C 3-8 cycloalkyl)oxy, substituted (C 3-8 cycloalkyl)oxy, (C 3-8 cycloalkyl)thio, substituted (C 3-8cycloalkyl)thio, halo, hydroxyl, C 1-10 Heteroaryl, substituted C 1-10 Heteroaryl, C 1-10 Heteroaryloxy, substituted C 1-10 Heteroaryloxy, C 1-10 Heteroarylthio, substituted C 1-10 Heteroarylthio, C 2-10 Heterocyclyl, C 2-10 Substituted heterocyclyl, C 2-10 Heterocyclyloxy, substituted C 2-10 Heterocyclyloxy, C 2-10 Heterocyclylthio, substituted C 2-10 Heterocyclylthio, imino, oxo, sulfonyl, sulfonylamino, thiol, C 1-10 Alkylthio, substituted C 1-10 It can be optionally substituted with alkylthio, and thiocarbonyl.

[0104] or any R 4 is any R 3 The cyclic structure is formed with H, halogen, hydroxyl, N3, NH2, NO2, CF3, C 1-10 Alkyl, substituted C 1-10 Alkyl, C 1-10 Alkoxy, substituted C 1-10 Alkoxy, acyl, acylamino, acyloxy, acyl C 1-10 Alkyloxy, amino, substituted amino, aminoacyl, aminocarbonyl C 1-10 Alkyl, aminocarbonylamino, aminodicarbonylamino, aminocarbonyloxy, aminosulfonyl, C 6-15 Aryl, substituted C 6-15 Aryl, C 6-15 Aryloxy, substituted C 6-15 Aryloxy, C 6-15 Arylthio, substituted C 6-15 Arylthio, carboxyl, carboxy ester, (carboxy ester) amino, (carboxy ester) oxy, cyano, C 3-8 Cycloalkyl, substituted C 3-8Cycloalkyl, (C 3-8 cycloalkyl)oxy, substituted (C 3-8 cycloalkyl)oxy, (C 3-8 cycloalkyl)thio, substituted (C 3-8 cycloalkyl)thio, halo, hydroxyl, C 1-10 Heteroaryl, substituted C 1-10 Heteroaryl, C 1-10 Heteroaryloxy, substituted C 1-10 Heteroaryloxy, C 1-10 Heteroarylthio, substituted C 1-10 Heteroarylthio, C 2-10 Heterocyclyl, C 2-10 Substituted heterocyclyl, C 2-10 Heterocyclyloxy, substituted C 2-10 Heterocyclyloxy, C 2-10 Heterocyclylthio, substituted C 2-10 Heterocyclylthio, imino, oxo, sulfonyl, sulfonylamino, thiol, C 1-10 Alkylthio, substituted C 1-10 C optionally substituted with alkylthio and thiocarbonyl 2-10 Heterocyclyl and C 1-10 Heteroaryl (C 2-10 heterocyclyl and C 1-10 heteroaryl optionally substituted with

[0105] n is an integer selected from 0 to 4; m is an integer selected from 0 to 5; each p is an integer independently selected from 0 to 2; q is an integer selected from 1 to 10.

[0106] In some embodiments, q can be 1. In some embodiments, q can be 2. In some embodiments, q can be 3. In some embodiments, q can be 4. In some embodiments, q can be 5. In some embodiments, q can be 6. In some embodiments, q can be 7. In some embodiments, q can be 8. In some embodiments, q can be 9. In some embodiments, q can be 10. In certain embodiments, q is 3 or 4.

[0107] In some embodiments, the rapafucine compounds of the present disclosure can have a structure according to formula (VI), or an optically pure stereoisomer or pharmaceutically acceptable salt thereof. [ka]

[0108] Each L 1 , L 2 , or L 3 can be independently selected from the linker structures in Table 1. Each AA1, AA2, AA3, or AA4 can be independently selected from the amino acid monomers shown in Table 3 below. X can be CH2, NH, O, or S; Y can be O, NH, or N-alkyl; E can be CH or N; and n is an integer selected from 0 to 4. The amino acids can be either NC-linked or CN-linked.

[0109] Each R 1 is H, halogen, hydroxyl, C 1-20 Alkyl, N3, NH2, NO2, CF3, OCF3, OCHF2, COC 1-20 Alkyl, and CO2C 1-20 alkyl. R 2 are H, halogen, hydroxyl, N3, NH2, NO2, CF3, C 1-10 Alkyl, substituted C1-10 Alkyl, C 1-10 Alkoxy, substituted C 1-10 Alkoxy, acyl, acylamino, acyloxy, acyl C 1-10 Alkyloxy, amino, substituted amino, aminoacyl, aminocarbonyl C 1-10 Alkyl, aminocarbonylamino, aminodicarbonylamino, aminocarbonyloxy, aminosulfonyl, C 6-15 Aryl, substituted C 6-15 Aryl, C 6-15 Aryloxy, substituted C 6-15 Aryloxy, C 6-15 Arylthio, substituted C 6-15 Arylthio, carboxyl, carboxy ester, (carboxy ester) amino, (carboxy ester) oxy, cyano, C 3-8 Cycloalkyl, substituted C 3-8 Cycloalkyl, (C 3-8 cycloalkyl)oxy, substituted (C 3-8 cycloalkyl)oxy, (C 3-8 cycloalkyl)thio, substituted (C 3-8 cycloalkyl)thio, C 1-10 Heteroaryl, substituted C 1-10 Heteroaryl, C 1-10 Heteroaryloxy, substituted C 1-10 Heteroaryloxy, C 1-10 Heteroarylthio, substituted C 1-10 Heteroarylthio, C 2-10 Heterocyclyl, C 2-10 Substituted heterocyclyl, C 2-10 Heterocyclyloxy, substituted C 2-10 Heterocyclyloxy, C 2-10 Heterocyclylthio, substituted C 2-10 Heterocyclylthio, imino, oxo, sulfonyl, sulfonylamino, thiol, C 1-10 Alkylthio, substituted C 1-10 C optionally substituted with alkylthio and thiocarbonyl 6-15 Aryl and C 1-10 Heteroaryl is selected from the group consisting of:

[0110] V is [ka] Z is a bond, [ka] where R 3 and R 4 are each independently selected from the group consisting of hydrogen, hydroxy, halo, alkyl, alkoxy, cycloalkyl, cyano, alkylthio, amino, alkylamino, and dialkylamino; K is O, CHR 5 , C.R. 5 , N, and NR 5 where R 5 is hydrogen or alkyl.

[0111] Synthetic Routes to Rapafusin. There are several methods for the synthesis of rapafucine, including both solid-phase and solution-phase synthesis. These methods can result in modifications to the linker(s) and / or effector domain, including alkylation, amide bond formation, double bond metathesis, oxadiazole formation, triazole formation, dithiol formation, sulfone formation, Diels-Alder cycloaddition, and others.

[0112] We applied solid-phase peptide synthesis to assemble the effector domain of the polypeptide. A preassembled FKBD capped with a carboxylic acid at one end and an olefin at the other end was later coupled to the polypeptide while still tethered on the bead. To facilitate purification of the newly formed macrocycle, we employed a coupled macrocyclization and cyclic release strategy, whereby macrocyclization is accompanied by the simultaneous release of the macrocyclic product from the solid bead. Those skilled in the art can contemplate different macrocyclization methods for the synthesis of the rapafusin molecules in this disclosure. In some embodiments, ring-closing metathesis / cyclative release (RCM) is used. In some embodiments, macrolactamization can be used for the efficient parallel synthesis of different rapafusins. For the construction of a rapafusin library, a cis-C6 linker can be used. Combination of medium temperature and catalyst loading (140° C., 30 mol % Hoveyda-Grubbs II catalyst) for the subsequent large-scale synthesis of the rapafucine library.

[0113] Other ring-closure methods can be used to synthesize the rapafucine molecules disclosed herein. Exemplary methods include, but are not limited to, aminolysis, chemoenzymatic methods, click chemistry, macrocyclization via ring contraction using auxiliary groups, macrocyclization mediated through sulfur-containing groups, macrocyclization via cycloaddition, macrocyclization via Wittiga or Wittig-like reactions, macrocyclization from multicomponent reactions, metal-assisted macrocyclization, macrocyclization via C-N bond formation, macrocyclization via C-O bond formation, alkylation with or without metal assistance, intramolecular cyclopropanation, oxidative coupling of arenes, side chain cyclization, and oxidative coupling of arenes. Each of these macrocyclization methods can be performed in solid phase or solution phase. Macrocyclization reactions via ring contraction using auxiliary groups include, but are not limited to, the use of hydroxylbenzaldehyde, hydroxylnitrophenol, and nitrovinylphenol. Macrocyclization reactions mediated through sulfur-containing groups can include, but are not limited to, thiazolidine formation O to N acyl transfer, transesterification S to N acyl transfer, ring chain tautomerization S to N acyl transfer, Staudinger ligation ring contraction, bis-thiol-ene macrocyclization, thiol-ene macrocyclization, thiol alkylation, and disulfide formation. Macrocyclization reactions via cycloaddition can include, but are not limited to, phospholene-azide ligation and oxadiazole grafting. Metal-assisted macrocyclizations can include, but are not limited to, C—C bond formation, Suzuki coupling, Sonogashira coupling, Tasuji-Trost reaction, Glaser-Hay coupling, and Nickel-catalyzed macrocyclization.Macrocyclization reactions via C-N bond configurations can include, but are not limited to, Ullmann coupling and Buchwald-Hartwig animation. Macrocyclization reactions via C-O bond configurations can include, but are not limited to, Chan-Lam-Evans coupling, C-H activation, and Ullmann coupling. Macrocyclization reactions via alkylation can include enolate chemistry, Williamson etherification, Mitsunobu reaction, nucleophilic aromatic substitution (SNAr), and Friedel-Crafts alkylation.

[0114] In some embodiments, the rapafucine molecule can be cyclized using the methods described in Marsault, E., & Peterson, M.L. (Eds.) (2017), Practical Medicinal Chemistry with Macrocycles: Design, Synthesis, and Case Studies, which is incorporated herein by reference in its entirety. Some non-limiting examples of macrocyclization methods are shown in Table 2 below, where each n can be an integer independently selected from 0 to 10. [Table 2-1] [Table 2-2] [Table 2-3] [Table 2-4] [Table 2-5] [Table 2-6] [Table 2-7] [Table 2-8] [Table 2-9] [Table 2-10] [Table 2-11] [Table 2-12] [Table 2-13] [Table 2-14] [Table 2-15]

[0115] In some embodiments, the rapafucine compounds of the present disclosure can have a structure according to formula (VII), or an optically pure stereoisomer or pharmaceutically acceptable salt thereof. [ka]

[0116] Each T1 or T2 can be independently selected from the terminal structures as outlined in Table 2 above prior to macrocyclization. Each L1, L2, or L3 can be independently selected from the linker structures in Table 1. Each AA can be independently selected from the amino acid monomers shown in Table 3 below. X can be CH2, NH, O, or S; Y can be O, NH, or N-alkyl; E can be CH or N; and n is an integer selected from 0 to 4. The amino acids can be either NC-linked or CN-linked.

[0117] In some embodiments, m can be 1. In some embodiments, m can be 2. In some embodiments, m can be 3. In some embodiments, m can be 4. In some embodiments, m can be 5. In some embodiments, m can be 6. In some embodiments, m can be 7. In some embodiments, m can be 8. In some embodiments, m can be 9. In some embodiments, m can be 10. In specific embodiments, m is 3 or 4.

[0118] Each R 1 is H, halogen, hydroxyl, C 1-20 Alkyl, N3, NH2, NO2, CF3, OCF3, OCHF2, COC 1-20 Alkyl, and CO2C 1-20 alkyl. R 2 are H, halogen, hydroxyl, N3, NH2, NO2, CF3, C 1-10 Alkyl, substituted C 1-10 Alkyl, C 1-10 Alkoxy, substituted C 1-10 Alkoxy, acyl, acylamino, acyloxy, acyl C 1-10 Alkyloxy, amino, substituted amino, aminoacyl, aminocarbonyl C 1-10 Alkyl, aminocarbonylamino, aminodicarbonylamino, aminocarbonyloxy, aminosulfonyl, C 6-15Aryl, substituted C 6-15 Aryl, C 6-15 Aryloxy, substituted C 6-15 Aryloxy, C 6-15 Arylthio, substituted C 6-15 Arylthio, carboxyl, carboxy ester, (carboxy ester) amino, (carboxy ester) oxy, cyano, C 3-8 Cycloalkyl, substituted C 3-8 Cycloalkyl, (C 3-8 cycloalkyl)oxy, substituted (C 3-8 cycloalkyl)oxy, (C 3-8 cycloalkyl)thio, substituted (C 3-8 cycloalkyl)thio, C 1-10 Heteroaryl, substituted C 1-10 Heteroaryl, C 1-10 Heteroaryloxy, substituted C 1-10 Heteroaryloxy, C 1-10 Heteroarylthio, substituted C 1-10 Heteroarylthio, C 2-10 Heterocyclyl, C 2-10 Substituted heterocyclyl, C 2-10 Heterocyclyloxy, substituted C 2-10 Heterocyclyloxy, C 2-10 Heterocyclylthio, substituted C 2-10 Heterocyclylthio, imino, oxo, sulfonyl, sulfonylamino, thiol, C 1-10 Alkylthio, substituted C 1-10 C optionally substituted with alkylthio and thiocarbonyl 6-15 Aryl and C 1-10 Heteroaryl is selected from the group consisting of:

[0119] V is [ka] Z is a bond, [ka] where R 3 and R 4are each independently selected from the group consisting of hydrogen, hydroxy, halo, alkyl, alkoxy, cycloalkyl, cyano, alkylthio, amino, alkylamino, and dialkylamino; K is O, CHR 5 , C.R. 5 , N, and NR 5 where R 5 is hydrogen or alkyl.

[0120] Table 3 below shows the FKBD moieties with linkers before incorporation into the rapafucine macrocyclic structure. [Table 3-1] [Table 3-2] [Table 3-3] [Table 3-4] [Table 3-5] [Table 3-6] [Table 3-7] [Table 3-8] *This FKBD is reduced and cyclized via lactamization.

[0121] Table 4 below shows the amino acid monomers used for the synthesis of rapafucine macrocyclic compounds in this disclosure. [Table 4-1] [Table 4-2]

Table 4-3

Table 4-4

Table 4-5

Table 4-6

Table 4-7

Table 4-8

Table 4-9

Table 4-10

Table 4-11

Table 4-12

Table 4-13

Table 4-14

Table 4-15

Table 4-16

Table 4-17

Table 4-18

Table 4-19

[0122] The monomers RbAsp, dD, D, and SbAsp have more than one hydroxyl group. In some embodiments, the hydroxyl group that serves as the attachment point to an adjacent residue in each of these monomers is illustrated above in Scheme 2. In some embodiments, other hydroxyl groups in these monomers can be used as attachment points to adjacent residues.

[0123] In some embodiments, disclosed herein is a compound of Formula VIII, or a pharmaceutically acceptable salt or solvate thereof: [ka]

[0124] In some embodiments, R is [ka] It can be R 1 , R 2 , R 3 , R 4 , and R 5 is hydrogen, hydroxyl, alkoxy, cyano, alkylthio, amino, and alkylamino, and [ka] can be independently selected from [ka] can be resin; so 1, 2, 3, or 4 A 1 , A 2 , A 3 , A 4 , and A 5 can be N or P and the rest CH; so 1, 2, 3, or 4 B 1 , B 2 , B 3 and B 4 can be O, N, or S, and the remainder can be CH or CH2, as appropriate; [ka] can be a single or double bond.

[0125] In some embodiments, X1 is O or NR 6 Y can be -C(O)- or [ka] X2 can be (CH2) m , O, OC(O), NR 6 , N.R. 6 C(O); Z can be [ka] W can be O, CH, CH2, CR 9 , or CR 10 R 11 L1 and L2 can each independently be a direct bond, a substituted or unsubstituted -(C1-C6) alkyl-, a substituted or unsubstituted -(CH2) n O(C1-C6) alkyl-, substituted or unsubstituted -(CH2) n C(O)(C1-C6) alkyl-, substituted or unsubstituted -(CH2) n C(O)O(C1-C6) alkyl-, substituted or unsubstituted -(CH2) n OC(O)(C1-C6) alkyl-, substituted or unsubstituted -(CH2) n NH(C1-C6) alkyl-, substituted or unsubstituted -(CH2) n C(O)NH(C1-C6) alkyl-, substituted or unsubstituted -(CH2) n S(C1-C6) alkyl-, substituted or unsubstituted -(CH2) n C(O)(CH2) n S(C1-C6) alkyl-, substituted or unsubstituted -(C2-C6) alkenyl-, substituted or unsubstituted -(CH2) n O(C2-C6)alkenyl-, substituted or unsubstituted -(CH2) n C(O)(C2-C6)alkenyl-, substituted or unsubstituted -(CH2) n C(O)O(C2-C6)alkenyl-, substituted or unsubstituted -(CH2) n OC(O)(C2-C6)alkenyl-, substituted or unsubstituted -(CH2) n NH(C2-C6)alkenyl-, substituted or unsubstituted -(CH2) n C(O)NH(C2-C6)alkenyl-, substituted or unsubstituted -(CH2) n S(C2-C6)alkenyl-, substituted or unsubstituted -(CH2) n C(O)(CH2) n S(C2-C6)alkenyl-, substituted or unsubstituted -(C2-C6)alkynyl-, substituted or unsubstituted -(CH2) nO(C2-C6)alkynyl-, substituted or unsubstituted -(CH2) n C(O)(C2-C6)alkynyl-, substituted or unsubstituted -(CH2) n C(O)O(C2-C6)alkynyl-, substituted or unsubstituted -(CH2) n OC(O)(C2-C6)alkynyl-, substituted or unsubstituted -(CH2) n NH(C2-C6)alkynyl-, substituted or unsubstituted -(CH2) n C(O)NH(C2-C6)alkynyl-, substituted or unsubstituted -(CH2) n S(C2-C6)alkynyl-, substituted or unsubstituted -(CH2) n C(O)(CH2) n S(C2-C6)alkynyl-, substituted or unsubstituted -(C1-C6)alkyl-NR 18 -, substituted or unsubstituted -(CH2) n O(C1-C6)alkyl-NR 18 -, substituted or unsubstituted -(CH2) n C(O)(C1-C6)alkyl-NR 18 -, substituted or unsubstituted -(CH2) n C(O)O(C1-C6)alkyl-NR 18 -, suabstituted or unsubstituted -(CH2) n OC(O)(C1-C6) alkyl-NR 18 -, substituted or unsubstituted -(CH2) n NH(C1-C6) alkyl-NR 18 -, substituted or unsubstituted -(CH2) n C(O)NH(C1-C6)alkyl-NR 18 -, substituted or unsubstituted -(CH2) n S(C1-C6)Alkyl-NR 18 -, substituted or unsubstituted -(CH2) n C(O)(CH2) n S(C1-C6)Alkyl-NR 18 -, substituted or unsubstituted -(C2-C6)alkenyl-NR 18 -, substituted or unsubstituted -(CH2) n O(C2-C6)Alkenyl-NR 18-, substituted or unsubstituted -(CH2) n C(O)(C2-C6)alkenyl-NR 18 -, substituted or unsubstituted -(CH2) n C(O)O(C2-C6)Alkenyl-NR 18 -, substituted or unsubstituted -(CH2) n OC(O)(C2-C6)alkenyl-NR 18 -, substituted or unsubstituted -(CH2) n NH(C2-C6)alkenyl-NR 18 -, substituted or unsubstituted -(CH2) n C(O)NH(C2-C6)alkenyl-NR 18 -, substituted or unsubstituted -(CH2) n S(C2-C6)Alkenyl-NR 18 -, substituted or unsubstituted -(CH2) n C(O)(CH2) n S(C2-C6)Alkenyl-NR 18 -, substituted or unsubstituted -(C2-C6)alkynyl-NR 18 -, substituted or unsubstituted -(CH2) n O(C2-C6)alkynyl-NR 18 -, substituted or unsubstituted -(CH2) n C(O)(C2-C6)alkynyl-NR 18 -, substituted or unsubstituted -(CH2) n C(O)O(C2-C6)alkynyl-NR 18 -, substituted or unsubstituted -(CH2) n OC(O)(C2-C6)alkynyl-NR 18 -, substituted or unsubstituted -(CH2) n NH(C2-C6)alkynyl-NR 18 -, substituted or unsubstituted -(CH2) n C(O)NH(C2-C6)alkynyl-NR 18 -, substituted or unsubstituted -(CH2) n S(C2-C6)alkynyl-NR 18 -, substituted or unsubstituted -(CH2) n C(O)(CH2) n S(C2-C6)alkynyl-NR 18-, substituted or unsubstituted -(C1-C6)alkyl-C(O)-, substituted or unsubstituted -(CH2) n O(C1-C6) alkyl-C(O)-, substituted or unsubstituted -(CH2) n C(O)(C1-C6) alkyl-C(O)-, substituted or unsubstituted -(CH2) n C(O)O(C1-C6) alkyl-C(O)-, substituted or unsubstituted -(CH2) n OC(O)(C1-C6) alkyl-C(O)-, substituted or unsubstituted -(CH2) n NH(C1-C6)alkyl-C(O)-, substituted or unsubstituted -(CH2) n C(O)NH(C1-C6)alkyl-C(O)-, substituted or unsubstituted -(CH2) n S(C1-C6) alkyl-C(O)-, substituted or unsubstituted -(CH2) n C(O)(CH2) n S(C1-C6)alkyl-C(O)-, substituted or unsubstituted -(C2-C6)alkenyl-C(O)-, substituted or unsubstituted -(CH2) n O(C2-C6)alkenyl-C(O)-, substituted or unsubstituted -(CH2) n C(O)(C2-C6)alkenyl-C(O)-, substituted or unsubstituted -(CH2) n C(O)O(C2-C6)alkenyl-C(O)-, substituted or unsubstituted -(CH2) n OC(O)(C2-C6)alkenyl-C(O)-, substituted or unsubstituted -(CH2) n NH(C2-C6)alkenyl-C(O)-, substituted or unsubstituted -(CH2) n C(O)NH(C2-C6)alkenyl-C(O)-, substituted or unsubstituted -(CH2) n S(C2-C6)alkenyl-C(O)-, substituted or unsubstituted -(CH2) n C(O)(CH2) n S(C2-C6)alkenyl-C(O)-, substituted or unsubstituted -(C2-C6)alkynyl-C(O)-, substituted or unsubstituted -(CH2) n O(C2-C6)alkynyl-C(O)-, substituted or unsubstituted -(CH2) nC(O)(C2-C6)alkynyl-C(O)-, substituted or unsubstituted -(CH2) n C(O)O(C2-C6)alkynyl-C(O)-, substituted or unsubstituted -(CH2) n OC(O)(C2-C6)alkynyl-C(O)-, substituted or unsubstituted -(CH2) n NH(C2-C6)alkynyl-C(O)-, substituted or unsubstituted -(CH2) n C(O)NH(C2-C6)alkynyl-C(O)-, substituted or unsubstituted -(CH2) n S(C2-C6)alkynyl-C(O)-, substituted or unsubstituted -(CH2) n C(O)(CH2) n S(C2-C6)alkynyl can be -C(O)-, -O-, -NH-, -S-, -S(O)-, -SO2-, -Si-, and -B-, where each alkyl, alkenyl, and alkynyl group can be optionally substituted with alkyl, alkoxy, amino, hydroxy, sulfhydryl, halogen, carbonyl, oxo, cyano, nitro, or trifluoromethyl.

[0126] L3 is a direct bond, a substituted or unsubstituted -(C1-C6) alkyl-, a substituted or unsubstituted -(CH2) n O(C1-C6) alkyl-, substituted or unsubstituted -(CH2) n C(O)(C1-C6) alkyl-, substituted or unsubstituted -(CH2) n C(O)O(C1-C6) alkyl-, substituted or unsubstituted -(CH2) n OC(O)(C1-C6) alkyl-, substituted or unsubstituted -(CH2) n NH(C1-C6) alkyl-, substituted or unsubstituted -(CH2) n C(O)NH(C1-C6) alkyl-, substituted or unsubstituted -(CH2) n S(C1-C6) alkyl-, substituted or unsubstituted -(CH2) n C(O)(CH2) n S(C1-C6) alkyl-, substituted or unsubstituted -(C2-C6) alkenyl-, substituted or unsubstituted -(CH2) nO(C2-C6)alkenyl-, substituted or unsubstituted -(CH2) n C(O)(C2-C6)alkenyl-, substituted or unsubstituted -(CH2) n C(O)O(C2-C6)alkenyl-, substituted or unsubstituted -(CH2) n OC(O)(C2-C6)alkenyl-, substituted or unsubstituted -(CH2) n NH(C2-C6)alkenyl-, substituted or unsubstituted -(CH2) n C(O)NH(C2-C6)alkenyl-, substituted or unsubstituted -(CH2) n S(C2-C6)alkenyl-, substituted or unsubstituted -(CH2) n C(O)(CH2) n S(C2-C6)alkenyl-, substituted or unsubstituted -(C2-C6)alkynyl-, substituted or unsubstituted -(CH2) n O(C2-C6)alkynyl-, substituted or unsubstituted -(CH2) n C(O)(C2-C6)alkynyl-, substituted or unsubstituted -(CH2) n C(O)O(C2-C6)alkynyl-, substituted or unsubstituted -(CH2) n OC(O)(C2-C6)alkynyl-, substituted or unsubstituted -(CH2) n NH(C2-C6)alkynyl-, substituted or unsubstituted -(CH2) n C(O)NH(C2-C6)alkynyl-, substituted or unsubstituted -(CH2) n S(C2-C6)alkynyl-, substituted or unsubstituted -(CH2) n C(O)(CH2) n S(C2-C6)alkynyl-, substituted or unsubstituted -(C1-C6)alkyl-NR 18 -, substituted or unsubstituted -(CH2) n O(C1-C6)alkyl-NR 18 -, substituted or unsubstituted -(CH2) n C(O)(C1-C6)alkyl-NR 18 -, substituted or unsubstituted -(CH2) n C(O)O(C1-C6)alkyl-NR 18 -, substituted or unsubstituted -(CH2) nOC(O)(C1-C6) alkyl-NR 18 -, substituted or unsubstituted -(CH2) n NH(C1-C6) alkyl-NR 18 -, substituted or unsubstituted -(CH2) n C(O)NH(C1-C6)alkyl-NR 18 -, substituted or unsubstituted -(CH2) n S(C1-C6)Alkyl-NR 18 -, substituted or unsubstituted -(CH2) n C(O)(CH2) n S(C1-C6)Alkyl-NR 18 -, substituted or unsubstituted -(C2-C6)alkenyl-NR 18 -, substituted or unsubstituted -(CH2) n O(C2-C6)Alkenyl-NR 18 -, substituted or unsubstituted -(CH2) n C(O)(C2-C6)alkenyl-NR 18 -, substituted or unsubstituted -(CH2) n C(O)O(C2-C6)Alkenyl-NR 18 -, substituted or unsubstituted -(CH2) n OC(O)(C2-C6)alkenyl-NR 18 -, substituted or unsubstituted -(CH2) n NH(C2-C6)alkenyl-NR 18 -, substituted or unsubstituted -(CH2) n C(O)NH(C2-C6)alkenyl-NR 18 -, substituted or unsubstituted -(CH2) n S(C2-C6)Alkenyl-NR 18 -, substituted or unsubstituted -(CH2) n C(O)(CH2) n S(C2-C6)Alkenyl-NR 18 -, substituted or unsubstituted -(C2-C6)alkynyl-NR 18 -, substituted or unsubstituted -(CH2) n O(C2-C6)alkynyl-NR 18 -, substituted or unsubstituted -(CH2) n C(O)(C2-C6)alkynyl-NR 18-, substituted or unsubstituted -(CH2) n C(O)O(C2-C6)alkynyl-NR 18 -, substituted or unsubstituted -(CH2) n OC(O)(C2-C6)alkynyl-NR 18 -, substituted or unsubstituted -(CH2) n NH(C2-C6)alkynyl-NR 18 -, substituted or unsubstituted -(CH2) n C(O)NH(C2-C6)alkynyl-NR 18 -, substituted or unsubstituted -(CH2) n S(C2-C6)alkynyl-NR 18 -, substituted or unsubstituted -(CH2) n C(O)(CH2) n S(C2-C6)alkynyl-NR 18 -, substituted or unsubstituted -(C1-C6)alkyl-C(O)-, substituted or unsubstituted -(CH2) n O(C1-C6) alkyl-C(O)-, substituted or unsubstituted -(CH2) n C(O)(C1-C6) alkyl-C(O)-, substituted or unsubstituted -(CH2) n C(O)O(C1-C6) alkyl-C(O)-, substituted or unsubstituted -(CH2) n OC(O)(C1-C6) alkyl-C(O)-, substituted or unsubstituted -(CH2) n NH(C1-C6)alkyl-C(O)-, substituted or unsubstituted -(CH2) n C(O)NH(C1-C6)alkyl-C(O)-, substituted or unsubstituted -(CH2) n S(C1-C6) alkyl-C(O)-, substituted or unsubstituted -(CH2) n C(O)(CH2) n S(C1-C6)alkyl-C(O)-, substituted or unsubstituted -(C2-C6)alkenyl-C(O)-, substituted or unsubstituted -(CH2) n O(C2-C6)alkenyl-C(O)-, substituted or unsubstituted -(CH2) n C(O)(C2-C6)alkenyl-C(O)-, substituted or unsubstituted -(CH2) nC(O)O(C2-C6)alkenyl-C(O)-, substituted or unsubstituted -(CH2) n OC(O)(C2-C6)alkenyl-C(O)-, substituted or unsubstituted -(CH2) n NH(C2-C6)alkenyl-C(O)-, substituted or unsubstituted -(CH2) n C(O)NH(C2-C6)alkenyl-C(O)-, substituted or unsubstituted -(CH2) n S(C2-C6)alkenyl-C(O)-, substituted or unsubstituted -(CH2) n C(O)(CH2) n S(C2-C6)alkenyl-C(O)-, substituted or unsubstituted -(C2-C6)alkynyl-C(O)-, substituted or unsubstituted -(CH2) n O(C2-C6)alkynyl-C(O)-, substituted or unsubstituted -(CH2) n C(O)(C2-C6)alkynyl-C(O)-, substituted or unsubstituted -(CH2) n C(O)O(C2-C6)alkynyl-C(O)-, substituted or unsubstituted -(CH2) n OC(O)(C2-C6)alkynyl-C(O)-, substituted or unsubstituted -(CH2) n NH(C2-C6)alkynyl-C(O)-, substituted or unsubstituted -(CH2) n C(O)NH(C2-C6)alkynyl-C(O)-, substituted or unsubstituted -(CH2) n S(C2-C6)alkynyl-C(O)-, substituted or unsubstituted -(CH2) n C(O)(CH2) n It can be S(C2-C6)alkynyl-C(O)-, where each alkyl, alkenyl, and alkynyl group can be optionally substituted with alkyl, alkoxy, amino, hydroxy, sulfhydryl, halogen, carbonyl, oxo, cyano, nitro, or trifluoromethyl.

[0127] Each m can be independently an integer selected from 0, 1, 2, 3, 4, 5, and 6; each n is independently an integer selected from 0, 1, 2, 3, 4, 5, and 6; R 6 is hydrogen or alkyl; R 7and R 8 are each independently selected from hydrogen, hydroxy, alkyl, alkoxy, cyano, alkylthio, amino, and alkylamino, and OPG, where OPG is a protecting group; R 9 , R 10 , and R 11 are each independently selected from hydrogen, hydroxy, alkyl, alkoxy, cyano, alkylthio, amino, and alkylamino, and OPG, where OPG is a protecting group.

[0128] The effector domain can have the formula (A): [ka]

[0129] R 12 , R 14 , R 16 , and R 18 are each independently hydrogen, substituted or unsubstituted alkyl, substituted or unsubstituted alkenyl, substituted or unsubstituted alkynyl, substituted or unsubstituted cycloalkyl, substituted or unsubstituted perfluoroalkyl, substituted or unsubstituted alkoxy, substituted or unsubstituted alkylamino, substituted or unsubstituted aryl, substituted or unsubstituted alkylaryl, substituted or unsubstituted heteroalkyl, substituted or unsubstituted heterocycloalkyl, substituted or unsubstituted heteroaryl, substituted or unsubstituted heteroalkylaryl, (CH) n CN, (CH2) n CF3, (CH2) n It can be C2F5. R 13 , R 15 , and R 17are independently selected from the side chains of naturally occurring amino acids and modified forms thereof, including, but not limited to, D-amino acid configurations, or from hydrogen, halogen, amino, cyano, nitro, trifluoromethyl, substituted or unsubstituted alkyl, substituted or unsubstituted alkenyl, substituted or unsubstituted alkynyl, substituted or unsubstituted cycloalkyl, substituted or unsubstituted perfluoroalkyl, substituted or unsubstituted alkoxy, substituted or unsubstituted alkylamino, substituted or unsubstituted alkylthio, substituted or unsubstituted aryl, substituted or unsubstituted alkylaryl, substituted or unsubstituted heteroalkyl, substituted or unsubstituted heterocycloalkyl, substituted or unsubstituted heteroaryl, substituted or unsubstituted heteroalkylaryl, substituted or unsubstituted (CH n -aryl, substituted or unsubstituted (CH2) n -heteroaryl, (CH2) n CN, (CH2) n CF3, (CH2) n C2F5, (CH2) n OR 19 , (CH2) n C(O)R 19 , (CH2) n C(O)OR 19 , (CH2) n OC(O)R 19 , (CH2) n NR 20 R 21 , (CH2) n C(O)NR 20 R 21 , (CH2) n NR 22 C(O)R 19 , (CH2) n NR 22 C(O)OR 19 , (CH2) n NR 22 C(O)NR 20 R 21 , (CH2) n SR 19 , (CH2) n S(O) j NR 20 R 21 , (CH2) n NR 22 S(O)j R 19 , or -(CH2) n NR 22 S(O) j NR 20 R 21 is.

[0130] R 12 and R 13 , R 14 and R 15 , R 16 and R 17 can be convalently connected to form a substituted or unsubstituted 5-, 6-, or 7-membered heterocycle. Each k can independently be an integer selected from 0, 1, 2, 3, 4, 5, 6, 7, 8, 9, and 10. Each j can independently be an integer selected from 0, 1, and 2. R 19 , R 20 , R 21 , and R 22 can each independently be hydrogen, halogen, amino, cyano, nitro, trifluoromethyl, alkyl, alkenyl, alkynyl, cycloalkyl, perfluoroalkyl, alkoxy, alkylamino, alkylthio, aryl, alkylaryl, heteroalkyl, heterocycloalkyl, heteroaryl, or heteroalkylaryl.

[0131] or R 19 and R 22 is as above, and R 20 and R 21 together with the N to which they are attached form a substituted or unsubstituted 5-, 6-, or 7-membered heterocycloalkyl or a substituted or unsubstituted 5-membered heteroaryl, where R 13 , R 15 , and R 17Each of the above groups listed for is halogen, amino, cyano, nitro, trifluoromethyl, alkyl, alkenyl, alkynyl, cycloalkyl, perfluoroalkyl, alkoxy, alkylamino, alkylthio, aryl, alkylaryl, heteroalkyl, heterocycloalkyl, heteroaryl, heteroalkylaryl, (CH2) n CN, (CH2) n CF3, (CH2) n C2F5, (CH2) n OR 19 , (CH2) n C(O)R 19 , (CH2) n C(O)OR 19 , (CH2) n OC(O)R 19 , (CH2) n NR 20 R 21 , (CH2) n C(O)NR 20 R 21 , (CH2) n NR 22 C(O)R 19 , (CH2) n NR 22 C(O)OR 19 , (CH2) n NR 22 C(O)NR 20 R 21 , (CH2) n SR 19 , (CH2) n S(O) j NR 20 R 21 , (CH2) n NR 22 S(O) j R 19 , or -(CH2) n NR 22 S(O) j NR 20 R 21 may be optionally and independently substituted with from 1 to 3 groups selected from:

[0132] Or the effector domain can have the formula (B): [ka]

[0133] Each k can be independently an integer selected from 0, 1, 2, 3, 4, 5, 6, 7, 8, 9, and 10; R 23 can be hydrogen or alkyl; X3 can be substituted or unsubstituted -(C1-C 30 ) alkyl-, alkenyl-, alkynyl-, and each carbon is individually assumed to be in one of the following redox states: CH2, CH-OH, C(O);

[0134] Or the effector domain can have the formula (C): [ka]

[0135] X4 is substituted or unsubstituted -(C1-C 30 ) alkyl-, alkenyl-, or alkynyl-, with each carbon individually assumed to be in one of the following redox states: CH2, CH-OH, C(O).

[0136] Or the effector domain has the formula (D): [ka]

[0137] R 24 and R 25 are each hydrogen or alkyl; X5 is a substituted or unsubstituted -(C1-C 30 ) alkyl-, alkenyl-, or alkynyl-, with each carbon individually assumed to be in one of the following redox states: CH2, CH-OH, C(O).

[0138] Or the effector domain can be of formula (E): [ka]

[0139] X6 is substituted or unsubstituted -(C1-C 30 ) alkyl-, alkenyl-, or alkynyl-, with each carbon individually assumed to be in one of the following redox states: CH2, CH-OH, C(O).

[0140] In some embodiments, L3 is [ka] Not R 26 is hydrogen or alkyl.

[0141] In some embodiments, R is [ka] Instead, R 3 is hydrogen, hydroxyl, or OPG, where PG is a protecting group, or [ka] and there [ka] is the resin; 2 is hydrogen, hydroxyl, or alkoxy; and 1 , R 4 , and R 5 are each independently free of substitution as defined by hydrogen or chemical bonds; [ka] is a single or double bond.

[0142] In some embodiments, L and L are each independently a direct bond, a substituted or unsubstituted -(C-C) alkyl-, a substituted or unsubstituted -(CH) n O(C1-C6) alkyl-, substituted or unsubstituted -(CH2)n C(O)-, substituted or unsubstituted -(CH2) n C(O)(C1-C6) alkyl-, substituted or unsubstituted -(CH2) n C(O)O(C1-C6) alkyl-, substituted or unsubstituted -(CH2) n NH(C1-C6) alkyl-, substituted or unsubstituted -(CH2) n C(O)NH(C1-C6) alkyl-, substituted or unsubstituted -(CH2) n S(C1-C6) alkyl-, substituted or unsubstituted -(CH2) n C(O)(CH2) n S(C1-C6) alkyl-, substituted or unsubstituted -(C2-C6) alkenyl-, substituted or unsubstituted -(CH2) n O(C2-C6)alkenyl-, substituted or unsubstituted -(CH2) n C(O)(C2-C6)alkenyl-, substituted or unsubstituted -(CH2) n C(O)O(C2-C6)alkenyl-, substituted or unsubstituted -(CH2) n NH(C1-C6)alkenyl-, substituted or unsubstituted -(CH2) n C(O)NH(C2-C6)alkenyl-, substituted or unsubstituted -(CH2) n S(C2-C6)alkenyl-, substituted or unsubstituted -(CH2) n C(O)(CH2) n S(C2-C6)alkenyl-, substituted or unsubstituted -(C2-C6)alkynyl-, substituted or unsubstituted -(CH2) n O(C2-C6)alkynyl-, substituted or unsubstituted -(CH2) n C(O)(C2-C6)alkynyl-, substituted or unsubstituted -(CH2) n C(O)O(C2-C6)alkynyl-, substituted or unsubstituted -(CH2) n NH(C1-C6)alkynyl-, substituted or unsubstituted -(CH2) n C(O)NH(C2-C6)alkynyl-, substituted or unsubstituted -(CH2) n S(C2-C6)alkynyl-, substituted or unsubstituted -(CH2) n C(O)(CH2) nS(C2-C6)alkynyl-, not each independently, where each alkyl, alkenyl, and alkynyl group can be optionally substituted with alkyl, alkoxy, amino, carboxyl, cyano, nitro, or trifluoromethyl.

[0143] In some embodiments, the effector domain is a compound of Formula (F). [ka]

[0144] R 12 , R 14 , R 14′ , R 16 , and R 27 are each independently not hydrogen or alkyl, and R 13 , R 14 , R 14′ , and R 16 are each independently hydrogen, halogen, amino, cyano, nitro, trifluoromethyl, substituted or unsubstituted alkyl, substituted or unsubstituted alkenyl, substituted or unsubstituted alkynyl, substituted or unsubstituted cycloalkyl, substituted or unsubstituted perfluoroalkyl, substituted or unsubstituted alkoxy, substituted or unsubstituted alkylamino, substituted or unsubstituted alkylthio, substituted or unsubstituted aryl, substituted or unsubstituted alkylaryl, substituted or unsubstituted heteroalkyl, substituted or unsubstituted heterocycloalkyl, substituted or unsubstituted heteroaryl, substituted or unsubstituted heteroalkylaryl, (CH n CN, (CH2) n CF3, (CH2) n C2F5, (CH2) n OR 19 , (CH2) n C(O)R 19 , (CH2) n C(O)OR 19 , (CH2) n OC(O)R 19 , (CH2) n NR 20 R 21 , (CH2)n C(O)NR 20 R 21 , (CH2) n NR 22 C(O)R 19 , (CH2) n NR 22 C(O)OR 19 , (CH2) n NR 22 C(O)NR 20 R 21 , (CH2) n S(O) j NR 20 R 21 , (CH2) n NR 22 S(O) j R 19 , or -(CH2) n NR 22 S(O) j NR 20 R 21 n is an integer selected from 0, 1, 2, 3, 4, 5, and 6; and j is an integer selected from 0, 1, and 2.

[0145] R 19 , R 20 , R 21 , and R 22 are each independently hydrogen, halogen, amino, cyano, nitro, trifluoromethyl, alkyl, alkenyl, alkynyl, cycloalkyl, perfluoroalkyl, alkoxy, alkylamino, alkylthio, aryl, alkylaryl, heteroalkyl, heterocycloalkyl, heteroaryl, or heteroalkylaryl; or R 19 and R 22 is as above, and R 20 and R 21 together with the N to which they are attached form a substituted or unsubstituted 5-, 6-, or 7-membered heterocycloalkyl or a substituted or unsubstituted 5-membered heteroaryl.

[0146] R 13 , R 15 , and R 17Each of the above groups listed for is halogen, amino, cyano, nitro, trifluoromethyl, alkyl, alkenyl, alkynyl, cycloalkyl, perfluoroalkyl, alkoxy, alkylamino, alkylthio, aryl, alkylaryl, heteroalkyl, heterocycloalkyl, heteroaryl, heteroalkylaryl, (CH2) n CN, (CH2) n CF3, (CH2) n C2F5, (CH2) n OR 19 , (CH2) n C(O)R 19 , (CH2) n C(O)OR 19 , (CH2) n OC(O)R 19 , (CH2) n NR 20 R 21 , (CH2) n C(O)NR 20 R 21 , (CH2) n NR 22 C(O)R 19 , (CH2) n NR 22 C(O)OR 19 , (CH2) n NR 22 C(O)NR 20 R 21 , (CH2) n SR 19 , (CH2) n S(O) j NR 20 R 21 , (CH2) n NR 22 S(O) j R 19 , or -(CH2) n NR 22 S(O) j NR 20 R 21 may be optionally and independently substituted with from 1 to 3 groups selected from:

[0147] In some embodiments, L3 in Formula (VII) is -CH2CH2- and R is [ka] and;R 1 , R 4 , R 5 and R 6 are hydrogen; R 2 and R 3 are methoxy; m=0; Y is [ka] and X2 is O or NR 6 L1 is -CH2-C(O)- or -(CH2)2C(O)-; Z is [ka] and L2 is -OCO-CH=CH-(CH2)2N(Me)-. In some embodiments, X2 is O and L1 is -CH2-C(O)-. In some embodiments, X2 is NR 6 C(O), and L1 is —(CH2)2C(O)—.

[0148] In some embodiments, the effector domain can be of formula (G): [ka]

[0149] R in the formula 12 , R 14 , R 14′ , and R 16are each independently hydrogen, substituted or unsubstituted alkyl, substituted or unsubstituted alkenyl, substituted or unsubstituted alkynyl, substituted or unsubstituted cycloalkyl, substituted or unsubstituted perfluoroalkyl, substituted or unsubstituted alkoxy, substituted or unsubstituted alkylamino, substituted or unsubstituted aryl, substituted or unsubstituted alkylaryl, substituted or unsubstituted heteroalkyl, substituted or unsubstituted heterocycloalkyl, substituted or unsubstituted heteroaryl, substituted or unsubstituted heteroalkylaryl, (CH) n CN, (CH2) n CF3, (CH2) n It's C2F5.

[0150] R 13 , R 15 , R 15′ and R 17 are independently selected from the side chains of naturally occurring amino acids and modified forms thereof, including, but not limited to, D-amino acid configurations, or from hydrogen, halogen, amino, cyano, nitro, trifluoromethyl, substituted or unsubstituted alkyl, substituted or unsubstituted alkenyl, substituted or unsubstituted alkynyl, substituted or unsubstituted cycloalkyl, substituted or unsubstituted perfluoroalkyl, substituted or unsubstituted alkoxy, substituted or unsubstituted alkylamino, substituted or unsubstituted alkylthio, substituted or unsubstituted aryl, substituted or unsubstituted alkylaryl, substituted or unsubstituted heteroalkyl, substituted or unsubstituted heterocycloalkyl, substituted or unsubstituted heteroaryl, substituted or unsubstituted heteroalkylaryl, substituted or unsubstituted (CH n -aryl, substituted or unsubstituted (CH2) n -heteroaryl, (CH2) n CN, (CH2) n CF3, (CH2) n C2F5, (CH2) n OR 19 , (CH2) n C(O)R 19 , (CH2) n C(O)OR 19 , (CH2) n OC(O)R 19 , (CH2) n NR20 R 21 , (CH2) n C(O)NR 20 R 21 , (CH2) n NR 22 C(O)R 19 , (CH2) n NR 22 C(O)OR 19 , (CH2) n NR 22 C(O)NR 20 R 21 , (CH2) n SR 19 , (CH2) n S(O) j NR 20 R 21 , (CH2) n NR 22 S(O) j R 19 , or -(CH2) n NR 22 S(O) j NR 20 R 21 is. R 12 and R 13 , R 14 and R 15 , R 14′ and R 15 ′ , R 16 and R 17 can be covalently linked to form a substituted or unsubstituted 5-, 6-, or 7-membered heterocycle.

[0151] In some embodiments, disclosed herein are methods of using hybrid cyclic libraries based on the immunophilin ligand family of natural products FK506 and rapamycin to screen for compounds for treating cancer. In some embodiments, disclosed herein are methods of using hybrid cyclic libraries based on the immunophilin ligand family of natural products FK506 and rapamycin to screen for compounds for treating autoimmune diseases.

[0152] The macrocyclic natural products FK506 and rapamycin are approved immunosuppressants with significant biological activity. Both have been shown to inhibit T cell activation, each with distinct mechanisms. Furthermore, rapamycin has been shown to have potent antiproliferative activity. FK506 and rapamycin share a surprising mode of action; they act by recruiting an abundantly and widely expressed intracellular protein, the prolyl cis-trans isomerase FKBP, and a binary complex, which subsequently binds to and allosterically inhibits the target proteins calcineurin and mTOR, respectively. Structurally, FK506 and rapamycin share a similar FKBP-binding domain but differ in their effector domains. The properties of FK506 and rapamycin suggest that swapping the effector domain of FK506 with that of rapamycin could shift the target from calcineurin to mTOR. The generation of a rapafucine library of macrocyles containing FK506 and rapamycin binding domains should hold great potential as new leads for the development of drugs used to treat disease.

[0153] Various methods exist for generating compound libraries for developing and screening compounds potentially useful in treating disease. One such method is the development of encoded libraries, and specific libraries, in which each compound contains an amplifiable tag. Such libraries include DNA-encoded libraries, in which DNA tags that identify library members can be amplified using molecular biology techniques, such as polymerase chain reaction (PCR). The use of such methods to produce libraries of rapafusin macrocycles containing FK506-like and rapamycin-like binding domains has not yet been demonstrated. Thus, there remains a need for DNA-encoded rapafusin libraries of macrocycles containing FK506-like and rapamycin-like binding domains.

[0154] In one embodiment, provided herein is a tagged macrocycle comprising: an FK506 binding protein binding domain (FKBD); an effector domain; a first linking region; and a second linking region; wherein the FKBD, effector domain, first linker, and second linker together form a macrocycle; and wherein at least one of the FKBD, effector domain, first linker, and second linker can be operatively linked to one or more oligonucleotides (D), which can identify the structure of at least one of the FKBD, effector domain, first linker, and second linker.

[0155] In certain embodiments, provided herein are tagged macrocycles of formula (IX): [ka]

[0156] In some embodiments, h, i, j, and k are each independently an integer from 0-20, with the proviso that at least one of h, i, j, and k is not 0; and D is an oligonucleotide capable of identifying at least one of the FKBD, effector domain, linking region A, or linking region Z, where a solid line connecting the FKBD, effector domain, linking region A, and / or linking region Z indicates an operative linkage, and a squiggly line indicates an operative linkage. In certain embodiments, oligonucleotide (D) can be operably linked to at least one of the FKBD, effector domain, linking region A, or linking region Z.

[0157] In some embodiments, provided herein are tagged macrocycles of formula (X), or a pharmaceutically acceptable salt, solvate, or stereoisomer thereof: [ka]

[0158] In some embodiments, Ring A is a 5-10 membered aryl, cycloalkyl, heteroaryl, or heterocycloalkyl, optionally substituted with 1-17 substituents, each of which is hydrogen, hydroxy, halo, alkyl, alkoxy, cyano, haloalkyl, haloalkoxy, alkylthio, oxo, amino, alkylamino, dialkylamino, [ka] and wherein [ka] is the resin; J is

[0159] -C(O)NR at each occurrence 6 -. [ka] independently selected from the group consisting of;

[0160] R in the formula 6 are hydrogen, alkyl, arylalkyl, [ka] and so R N is aryl, alkyl, or arylalkyl; R' is hydrogen, alkyl, arylalkyl, or haloalkyl; D is independently at each occurrence an oligonucleotide; L b and L c is a bond, -O-, -S-, -OC(O)-, -C(O)O-, -(CH2) n C(O)-, -(CH2) n C(O)C(O)-, -(CH2) n NR 5 C(O)C(O)-, -NR 5 (CH2)n C(O)C(O)-, optionally substituted (CH2) n C 1-6 Alkylene (CH2) n -, optional substitution (CH2) n C(O)C 1-6 Alkylene (CH2) n -, optional substitution (CH2) n NR 5 C 1-6 Alkylene (CH2) n -, optional substitution (CH2) n C(O)NR 5 C 1-6 Alkylene (CH2) n -, optionally substituted (CH2)nNR 5 C(O)C 1-6 Alkylene (CH2) n -, optional substitution (CH2) n C(O)OC 1-6 Alkylene (CH2) n -, optional substitution (CH2) n OC(O)C 1-6 Alkylene (CH2) n -, optional substitution (CH2) n O.C. 1-6 Alkylene (CH2) n -, optional substitution (CH2) n NR 5 C 1-6 Alkylene (CH2)n-, optionally substituted (CH2) n -SC 1-6 Alkylene (CH2) n - and optionally substituted (CH2CH2O) n wherein each alkylene is optionally substituted with one or two groups independently selected from the group consisting of halo, hydroxy, haloalkyl, haloalkoxy, alkyl, alkoxy, amino, carboxyl, cyano, nitro, and NHFmoc; and wherein each R 5 is independently hydrogen, alkyl, arylalkyl, [ka] and so R Nis aryl, alkyl, or arylalkyl; X is O, S, or NR 8 where R 8 is hydrogen, hydroxy, OR 9 , N.R. 10 R 11 , alkyl, arylalkyl, [ka] and so R N is aryl, alkyl, or arylalkyl; where R 9 , R 10 and R 11 are each independently hydrogen or alkyl; V 1 and V 2 are independent of each other [ka] and W is [ka] wherein Ring B is a 4-10 membered heterocycloalkyl, optionally substituted with 1-10 substituents, each of which is hydrogen, hydroxy, halo, alkyl, alkoxy, cyano, haloalkyl, haloalkoxy, alkylthio, oxo, amino, alkylamino, dialkylamino, arylalkyl, [ka] where R 12 is aryl, alkyl, or arylalkyl; where R 13 is hydrogen, hydroxy, OR 16 , N.R. 17 R 18 , alkyl, arylalkyl, [ka] and so R N is aryl, alkyl, or arylalkyl; R 14 and R15 are each independently hydrogen, hydroxy, halo, alkyl, alkoxy, haloalkyl, haloalkoxy, aryl, arylalkyl, or heteroaryl; Z is a bond, [ka] where R 16 and R 17 are each independently selected from the group consisting of hydrogen, hydroxy, halo, alkyl, alkoxy, cycloalkyl, cyano, alkylthio, amino, alkylamino, and dialkylamino; K is O, CHR 18 , C.R. 18 , N, or and NR 18 where R 18 is hydrogen or alkyl;

[0161] L a , L 1 , L 2 , L 3 , L 4 , L 5 , L 6 , L 7 and L 8 are independently bonded, -O-, -NR 19 -, -SO-, -SO2-, (CH2) n -,, [ka] or a linking group selected from Table 1; wherein Ring C is a 5-6 membered heteroaryl optionally substituted with 1-4 substituents, each of which is hydrogen, hydroxyl, halo, alkyl, alkoxy, haloalkyl, haloalkoxy, cyano, alkylthio, amino, alkylamino, dialkylamino, and [ka] where each R 19 , R 20 , and R21 is hydrogen, hydroxy, OR 22 , N.R. 23 R 24 , alkyl, arylalkyl, [ka] is independently selected from the group consisting of; therefore, R N is aryl, alkyl, or arylalkyl; where R 22 , R 23 , and R 24 are each independently hydrogen or alkyl;

[0162] N is 0, 1, 2, 3, 4, 5 or 6; and the effector domain has the formula (Xa): [ka]

[0163] In some embodiments, each k a , k b , k c , k d 、 k e , k f , k g , k h 、 and k i is independently 0 or 1; each X a , X b , X c , X d , X e , X f , X g , X h , and X i are independently a bond, -S-, -SS-, -S(O)-, -S(O)2-, substituted or unsubstituted -(C1-C3)alkylene-, -(C2-C4)alkenylene-, -(C2-C4)alkynylene-, or [ka] where ring E is phenyl or 5-6 heteroaryl or heterocycloalkyl; where each w is independently 0, 1, or 2; and where each R 1 , R 1a , R 1b , R 1c , R 1d , R 1e , R 1f , R 1g , R 1h , R 1i , and R 4 is independently hydrogen, alkyl, arylalkyl, or NR 25 where R 25 is hydrogen, hydroxy, OR 26 , N.R. 27 R 28 , alkyl, arylalkyl, [ka] and so R N is aryl, alkyl, or arylalkyl; where R 26 , R 27 , and R 28 are each independently hydrogen or alkyl; 2 , R 3 , R 2a , R 3a , R 2b , R 3b , R 2c , R 3c , R 2d , R 3d , R 2e , R 3e , R 2f , R 3f , R 2g , R 3g , R 2h , R 3h , R 2i , and R 3iis hydrogen, halo, amino, cyano, nitro, haloalkyl, optionally substituted alkyl, optionally substituted alkoxy, optionally substituted alkenyl, optionally substituted alkynyl, optionally substituted cycloalkyl, optionally substituted heterocycloalkyl, optionally substituted alkylamino, optionally substituted dialkylamino, optionally substituted aryl, optionally substituted heteroaryl, optionally substituted arylalkyl, optionally substituted heteroarylalkyl, and [ka] independently selected from the group consisting of;

[0164] or where the effector domain has the formula (Xb): [ka]

[0165] So each AA 1 , A.A. 2 , ..., and AA r is a natural or unnatural amino acid residue; and r is 3, 4, 5, 6, 7, 8, 9, or 10;

[0166] or where the effector domain has the formula (Xc): [ka]

[0167] where each t is an integer independently selected from 0, 1, 2, 3, 4, 5, 6, 7, 8, 9, and 10; R 29 is hydrogen, hydroxy, OR 30 , N.R. 31 R 32 , alkyl, arylalkyl, [ka] and so R N is aryl, alkyl, or arylalkyl; where R 30 , R 31 , and R32 are each independently hydrogen or alkyl; X 3 is a substituted or unsubstituted -(C1-C6)alkylene-, -(C2-C6)alkenylene-, -(C2-C6)alkynylene-, or [ka] wherein ring E is phenyl or a 5-6 heteroaryl or heterocycloalkyl; and wherein each w is independently 0, 1, or 2;

[0168] or where the effector domain has the formula (Xd): [ka]

[0169] So X 4 is a substituted or unsubstituted -(C1-C6)alkylene-, -(C2-C6)alkenylene-, -(C2-C6)alkynylene-, or [ka] wherein ring E is phenyl or a 5-6 heteroaryl or heterocycloalkyl; and wherein each w is independently 0, 1, or 2;

[0170] or where the effector domain has the formula (Xe): [ka]

[0171] So R 33 , R 34 、 R 35 and R 36 are each hydrogen or alkyl; X 5 is a substituted or unsubstituted -(C1-C6)alkylene-, -(C2-C6)alkenylene-, -(C2-C6)alkynylene-, or [ka] wherein ring E is phenyl or a 5-6 heteroaryl or heterocycloalkyl; and wherein each w is independently 0, 1, or 2;

[0172] or where the effector domain has the formula (Xf): [ka]

[0173] X 6 is a substituted or unsubstituted -(C1-C6)alkylene-, -(C2-C6)alkenylene-, -(C2-C6)alkynylene-, or [ka] wherein ring E is phenyl or 5-6 heteroaryl or heterocycloalkyl; wherein each w is independently 0, 1, or 2; provided that R is [ka] When L is ethylene, X is O, and W is [ka] and V is [ka] and Z is [ka] and -L 6 -L 7 -L 8 -teeth [ka] and also -L 1 -L 2 -L 3 -L 4 -L5 -teeth [ka] and; where ring A has at least one [ka] or at least one R 2 , R 3 , R 2a , R 3a , R 2b , R 3b , R 2c , R 3c , R 2d , R 3d , R 2e , R 3e , R 2f , R 3f , R 2g , R 3g , R 2h , R 3h , R 2i , and R 3i teeth [ka] or at least one L a , L 1 , L 2 , L 3 , L 4 , L 5 , L 6 , L 7 and L 8 At least one [ka] or wherein at least one linking group selected from Table 1 is at least one [ka] is replaced by

[0174] In another aspect, provided herein is a compound library comprising a plurality of distinct tagged macrocycles according to any of the above. In certain embodiments, provided herein is a compound library comprising at least about 10 distinct tagged macrocycles according to any of the above. 2 In certain embodiments, provided herein is a compound library comprising about 10 distinct tagged macrocycles according to any of the above. 2 from about 10 10 A compound library containing up to 10 distinct tagged macrocycles.

[0175] In a further aspect, provided herein is a method of making a library of tagged macrocycles disclosed herein, the method comprising synthesizing a plurality of distinct tagged macrocycles according to any of the above.

[0176] In yet a further aspect, provided herein is a method of making a tagged macrocycle disclosed herein, the method comprising operably linking at least one oligonucleotide (D) to at least one of an FKBD, an effector domain, a first linking region, and a second linking region, and forming a macrocycle comprising the FKBD, the effector domain, the first linking region, and the second linking region.

[0177] In certain embodiments, provided herein are methods for making a tagged macrocycle disclosed herein, the methods comprising attaching the macrocycle to at least one oligonucleotide (D), wherein the macrocycle comprises an FKBD, an effector domain, a first linking region, and a second linking region, wherein the FKBD, the effector domain, the first linking region, and the second linking region together form a macrocycle; and wherein the at least one oligonucleotide (D) is capable of identifying the structure of at least one of the FKBD, the effector domain, the first linking region, and the second linking region.

[0178] In yet a further aspect, the method of making a tagged macrocycle comprises the step of: [ka] with a compound of formula (XII): [ka] This includes operably connecting to

[0179] In some embodiments, [ka] independently at each occurrence: bond, -O-, -NR 19 -, -SO-, -SO2-, -(CH2) n -, [ka] or a linking group selected from Table 1, wherein Ring C is a 5-6 membered heteroaryl optionally substituted with 1-4 substituents, each of which is independently selected from the group consisting of hydrogen, hydroxy, halo, alkyl, alkoxy, haloalkyl, haloalkoxy, cyano, alkylthio, amino, alkylamino, and dialkylamino; 19 is hydrogen, hydroxy, OR 22 , N.R. 23 R 24 , alkyl, arylalkyl, [ka] where R N is aryl, alkyl, or arylalkyl; where R 22 , R 23 , and R 24 are each independently hydrogen or alkyl; Q and Q′ are N3, —C≡CH, NR 6 R 7 , -COOH, -ONH2, -SH, -NH2, [ka] , -(C=O)R′, [ka] Each is independently selected from the group consisting of:

[0180] So R 6 and R 7 are independently hydrogen, alkyl, arylalkyl, [ka] and so R N is aryl, alkyl, or arylalkyl; and R' is hydrogen, alkyl, arylalkyl, or haloalkyl; L b and L c a is a bond, -O-, -S-, -OC(O)-, -C(O)O-, -(CH2) n C(O)-, -(CH2) n C(O)C(O)-, -(CH2) n NR 5 C(O)C(O)-, -NR 5 (CH2) n C(O)C(O)-, optionally substituted (CH2) n C 1-6 Alkylene-(CH2) n -, optional substitution (CH2) n C(O)C 1-6 Alkylene-(CH2) n -, optional substitution (CH2) n NR 5 C 1-6 Alkylene-(CH2) n -, optional substitution (CH2) n C(O)NR 5 C 1-6 Alkylene-(CH2) n -, optional substitution (CH2) n NR 5 C(O)C 1-6 Alkylene-(CH2) n -, optional substitution (CH2) n C(O)OC 1-6Alkylene-(CH2) n -, optional substitution (CH2) n OC(O)C 1-6 Alkylene-(CH2) n -, optional substitution (CH2) n O.C. 1-6 Alkylene-(CH2) n -, optional substitution (CH2) n NR 5 C 1-6 Alkylene-(CH2) n -, optional substitution (CH2) n -SC 1-6 Alkylene-(CH2) n - and optionally substituted (CH2CH2O) n wherein each alkylene is optionally substituted with one or two groups independently selected from the group consisting of halo, hydroxy, haloalkyl, haloalkoxy, alkyl, alkoxy, amino, carboxyl, cyano, nitro, NHFmoc; and wherein each R 5 is independently hydrogen, alkyl, arylalkyl, [ka] and so R N is aryl, alkyl, or arylalkyl;

[0181] D is an oligonucleotide; h, i, j, and k are each independently an integer from 0-20, provided that at least one h, i, j, and k is not 0; n is an integer from 1-5; and m is an integer from 1-5.

[0182] In another aspect, provided herein is a method of making a tagged macrocycle, the method comprising: [ka] with a compound of formula (XII): [ka] and operatively connecting the

[0183] Ring A is a 5-10 membered aryl, cycloalkyl, heteroaryl, or heterocycloalkyl, optionally substituted with 1-17 substituents, each of which is hydrogen, hydroxy, halo, alkyl, alkoxy, cyano, haloalkyl, haloalkoxy, alkylthio, oxo, amino, alkylamino, dialkylamino, [ka] and wherein [ka] is resin;

[0184] L b and L c is a bond, -O-, -S-, -OC(O)-, -C(O)O-, -(CH2) n C(O)-, -(CH2) n C(O)C(O)-, -(CH2) n NR 5 C(O)C(O)-, -NR 5 (CH2) n C(O)C(O)-, optionally substituted (CH2) n C 1-6 Alkylene-(CH2) n -, optional substitution (CH2) n C(O)C 1-6 Alkylene-(CH2) n -, optional substitution (CH2) n NR 5 C 1-6 Alkylene-(CH2) n -, optional substitution (CH2) n C(O)NR 5 C 1-6 Alkylene-(CH2) n -, optional substitution (CH2) n NR 5 C(O)C 1-6 Alkylene-(CH2) n -, optional substitution (CH2)n C(O)OC 1-6 Alkylene-(CH2) n -, optional substitution (CH2) n OC(O)C 1-6 Alkylene-(CH2) n -, optional substitution (CH2) n O.C. 1-6 Alkylene-(CH2) n -, optional substitution (CH2) n NR 5 C 1-6 Alkylene-(CH2) n -, optional substitution (CH2) n -SC 1-6 Alkylene-(CH2) n - and optionally substituted (CH2CH2O) n wherein each alkylene is optionally substituted with one or two groups independently selected from the group consisting of halo, hydroxy, haloalkyl, haloalkoxy, alkyl, alkoxy, amino, carboxyl, cyano, nitro, and NHFmoc; and wherein each R 5 is independently hydrogen, alkyl, arylalkyl, [ka] and so R N is aryl, alkyl, or arylalkyl;

[0185] Q and Q′ are -N3, -C≡CH, NR 6 R 7 , -COOH, -ONH2, -SH, -NH2, [ka] -(C=O)R′, [ka] independently selected from the group consisting of;

[0186] So R 6 and R 7 are independently hydrogen, alkyl, arylalkyl, [ka] and so R N is aryl, alkyl, or arylalkyl; and R' is hydrogen, alkyl, arylalkyl, or haloalkyl; X is O, S, or NR 8 where R 8 is hydrogen, hydroxy, OR 9 , N.R. 10 R 11 , alkyl, arylalkyl, [ka] and so R N is aryl, alkyl, or arylalkyl; where R 9 , R 10 and R 11 are each independently hydrogen or alkyl; V 1 and V 2 are independent of each other [ka] and W is [ka] wherein Ring B is a 4-10 membered heterocycloalkyl, optionally substituted with 1-10 substituents, each of which is hydrogen, hydroxy, halo, alkyl, alkoxy, cyano, haloalkyl, haloalkoxy, alkylthio, oxo, amino, alkylamino, dialkylamino, arylalkyl, [ka] where R 12 is aryl, alkyl, or arylalkyl; where R 13 is hydrogen, hydroxy, OR 16 , N.R. 17 R 18 , alkyl, arylalkyl, [ka] and so R N is aryl, alkyl, or arylalkyl; R 14 and R 15 are each independently hydrogen, hydroxy, halo, alkyl, alkoxy, haloalkyl, haloalkoxy, aryl, arylalkyl, or heteroaryl;

[0187] Z is a bond, [ka] where R 16 and R 17 are each independently selected from the group consisting of hydrogen, hydroxy, halo, alkyl, alkoxy, cycloalkyl, cyano, alkylthio, amino, alkylamino, and dialkylamino; K is O, CHR 18 , C.R. 18 , N, and NR 18 where R 18 is hydrogen or alkyl;

[0188] L a , L 1 , L 2 , L 3 , L 4 , L 5 , L 6 , L 7 and L 8 are independently bonded, -O-, -NR 19 -, -SO-, -SO2-, -(CH2) n -, [ka] or a linking group selected from Table 1; wherein Ring C is a 5-6 membered heteroaryl optionally substituted with 1-4 substituents, each of which is hydrogen, hydroxy, halo, alkyl, alkoxy, haloalkyl, haloalkoxy, cyano, alkylthio, amino, alkylamino, dialkylamino, and [ka] are chosen independently from the group consisting of 19 is hydrogen, hydroxy, OR 22 , N.R. 23 R 24 , alkyl, arylalkyl, [ka] where R N is aryl, alkyl, or arylalkyl; where R 22 , R 23 , and R 24 are each independently hydrogen or alkyl;

[0189] n is 0, 1, 2, 3, 4, 5 or 6; and the effector domain has the formula (Xa): [ka]

[0190] each k a , k b , k c , k d 、 k e , k f , k g , k h 、 and k i is independently 0 or 1; each X a , X b , X c , X d , X e , X f , X g , X h , and X i are independently a bond, -S-, -SS-, -S(O)-, -S(O)2-, substituted or unsubstituted -(C1-C3)alkylene-, -(C2-C4)alkenylene-, -(C2-C4)alkynylene-, or [ka] where ring E is phenyl or 5-6 heteroaryl or heterocycloalkyl; where each w is independently 0, 1, or 2; and where each R 1 , R 1a , R 1b , R 1c , R 1d , R 1e , R 1f , R 1g , R 1h , R 1i , and R 4 is independently hydrogen, alkyl, arylalkyl, or NR 25 where R 25 is hydrogen, hydroxy, OR 26 , N.R. 27 R 28 , alkyl, arylalkyl, [ka] and so R N is aryl, alkyl, or arylalkyl; where R 26 , R 27 , and R 28 are each independently hydrogen or alkyl; 2 , R 3 , R 2a , R 3a , R 2b , R 3b , R 2c , R 3c , R 2d , R 3d , R 2e , R 3e , R 2f , R 3f , R 2g , R 3g , R 2h , R 3h , R 2i , and R 3iis hydrogen, halo, amino, cyano, nitro, haloalkyl, optionally substituted alkyl, optionally substituted alkoxy, optionally substituted alkenyl, optionally substituted alkynyl, optionally substituted cycloalkyl, optionally substituted heterocycloalkyl, optionally substituted alkylamino, optionally substituted dialkylamino, optionally substituted aryl, optionally substituted heteroaryl, optionally substituted arylalkyl, optionally substituted heteroarylalkyl, and [ka] independently selected from the group consisting of;

[0191] or where the effector domain has the formula (Xb): [ka]

[0192] So each AA 1 , A.A. 2 , …, and AA r is a natural or unnatural amino acid residue; and r is 3, 4, 5, 6, 7, 8, 9, or 10;

[0193] or where the effector domain has the formula (Xc): [ka]

[0194] Each t is independently an integer selected from 0, 1, 2, 3, 4, 5, 6, 7, 8, 9, and 10; R 29 is hydrogen, hydroxy, OR 30 , N.R. 31 R 32 , alkyl, arylalkyl, [ka] and so R N is aryl, alkyl, or arylalkyl; where R 30 , R 31 , and R 32are each independently hydrogen or alkyl; X 3 is a substituted or unsubstituted -(C1-C6)alkylene-, -(C2-C6)alkenylene-, -(C2-C6)alkynylene-, or [ka] wherein ring E is phenyl or a 5-6 heteroaryl or heterocycloalkyl; and wherein each w is independently 0, 1, or 2;

[0195] or where the effector domain has the formula (Xd): [ka]

[0196] X 4 is a substituted or unsubstituted -(C1-C6)alkylene-, -(C2-C6)alkenylene-, -(C2-C6)alkynylene-, or [ka] wherein ring E is phenyl or a 5-6 heteroaryl or heterocycloalkyl; and wherein each w is independently 0, 1, or 2;

[0197] or where the effector domain has the formula (Xe): [ka]

[0198] R 33 , R 34 、 R 35 and R 36 are each hydrogen or alkyl; X 5 is a substituted or unsubstituted -(C1-C6)alkylene-, -(C2-C6)alkenylene-, -(C2-C6)alkynylene-, or [ka] wherein ring E is phenyl or a 5-6 heteroaryl or heterocycloalkyl; and wherein each w is independently 0, 1, or 2;

[0199] or where the effector domain has the formula (Xf): [ka]

[0200] X 6 is a substituted or unsubstituted -(C1-C6)alkylene-, -(C2-C6)alkenylene-, -(C2-C6)alkynylene-, or [ka] wherein ring E is phenyl or 5-6 heteroaryl or heterocycloalkyl; wherein each w is independently 0, 1, or 2; and with the proviso that ring A is [ka] When L a is ethylene, X is O, and W is [ka] and V 1 teeth [ka] and V 2 teeth [ka] and Z is [ka] and -L 6 -L 7 -L 8 -teeth [ka] and -L 1 -L 2 -L 3 -L 4 -L 5 -teeth [ka] D is an oligonucleotide; wherein ring A is at least one [ka] or at least one R 2 , R 3 , R 2a , R 3a , R 2b , R 3b , R 2c , R 3c , R 2d , R 3d , R 2e , R 3e , R 2f , R 3f , R 2g , R 3g , R 2h , R 3h , R 2i , and R 3i teeth [ka] or at least one L a , L 1 , L 2 , L 3 , L 4 , L 5 , L 6 , L 7 and L 8 At least one [ka] or wherein at least one linking group selected from Table 1 is at least one [ka] is replaced by

[0201] Also provided herein is a macrocyclic compound of formula (XIV) or a pharmaceutically acceptable salt, solvate, or stereoisomer thereof: [ka]

[0202] Each of n, m, and p is independently an integer selected from 0 to 5.

[0203] Each R1, R2, and R3 is H, F, Cl, Br, CF3, CN, N3, -N(R 12 )2, -N(R 12 )3, -CON(R 12 )2, NO2, OH, OCH3, methyl, ethyl, propyl, -COOH, -SO3H, -PO(OR 12 )2, -OPO(OR 12 )2, -(CH2) q COOH, -O-(CH2) q COOH, -S-(CH2) q COOH, -CO-(CH2) q COOH, -NR 12 -(CH2) q COOH, -(CH2) q SO3H, -O-(CH2) q SO3H, -S-(CH2) q SO3H, -CO-(CH2) q SO3H, -NR 12 -(CH2) q SO3H, -(CH2) q N(R 12 )2, -O-(CH2) q N(R 12 )2, -S-(CH2) q N(R 12 )2, -CO-(CH2) q N(R 12 )2, -(CH2) q N(R 12 )3, -O-(CH2) q N(R12 )3, -S-(CH2) q N(R 12 )3, -CO-(CH2) q N(R 12 )3, -NR 12 -(CH2) q N(R 12 )3, -(CH2) q CON(R 12 )2, -O-(CH2) q CON(R 12 )2, -S-(CH2) q CON(R 12 )2, -CO-(CH2) q CON(R 12 )2, -(CH2) q PO(OR 12 )2, -O(CH2) q PO(OR 12 )2, -S(CH2) q PO(OR 12 )2, -CO(CH2) q PO(OR 12 )2, -NR 12 (CH2) q PO(OR 12 )2, -(CH2) q OPO(OR 12 )2, -O(CH2) q OPO(OR 12 )2, -S(CH2) q OPO(OR 12 )2, -CO(CH2) q OPO(OR 12 )2, and -NR 12 (CH2) q OPO(OR 12 )2.

[0204] q can be an integer selected from 0 to 5. Each of R4, R5, R6, R7, R9, and R 11 may be independently selected from the group consisting of H, methyl, ethyl, propyl, and isopropyl.

[0205] Each R8 and R 10 is H, halogen, hydroxyl, C1-20 Alkyl, N3, NH2, NO2, CF3, OCF3, OCHF2, COC 1-20 Alkyl, CO2C 1-20 Alkyl, a 5- or 6-membered ring moiety formed with the adjacent nitrogen, -N(R 12 )2, -N(R 12 )3, -CON(R 12 )2, -COOH, -SO3H, -PO(OR 12 )2, -OPO(OR 12 )2, -(CH2) q COOH, -O-(CH2) q COOH, -S-(CH2) q COOH, -CO-(CH2) q COOH, -NR 12 -(CH2) q COOH, -(CH2) q SO3H, -O-(CH2) q SO3H, -S-(CH2) q SO3H, -CO-(CH2) q SO3H, -NR 12 -(CH2) q SO3H, -(CH2) q N(R 12 )2, -O-(CH2) q N(R 12 )2, -S-(CH2) q N(R 12 )2, -CO-(CH2) q N(R 12 )2, -(CH2) q N(R 12 )3, -O-(CH2) q N(R 12 )3, -S-(CH2) q N(R 12 )3, -CO-(CH2) q N(R 12 )3, -NR 12 -(CH2) q N(R 12 )3, -(CH2) q CON(R 12 )2, -O-(CH2) q CON(R 12 )2, -S-(CH2) qCON(R 12 )2, -CO-(CH2) q CON(R 12 )2, -(CH2) q PO(OR 12 )2, -O(CH2) q PO(OR 12 )2, -S(CH2) q PO(OR 12 )2, -CO(CH2) q PO(OR 12 )2, -NR 12 (CH2) q PO(OR 12 )2, -(CH2) q OPO(OR 12 )2, -O(CH2) q OPO(OR 12 )2, -S(CH2) q OPO(OR 12 )2, -CO(CH2) q OPO(OR 12 )2, and -NR 12 (CH2) q OPO(OR 12 ) 2 independently selected from the group consisting of

[0206] Each R 12 may be independently selected from the group consisting of H, methyl, ethyl, propyl, and isopropyl.

[0207] However, R2, R3, R8 and R 10 At least one of -N(R 12 )2, -N(R 12 )3, -CON(R 12 )2, -COOH, -SO3H, -PO(OR 12 )2, -OPO(OR 12 )2, -(CH2) q COOH, -O-(CH2) q COOH, -S-(CH2) q COOH, -CO-(CH2) q COOH, -NR 12 -(CH2) q COOH, -(CH2) q SO3H, -O-(CH2)q SO3H、-S-(CH2) q SO3H、-CO-(CH2) q SO3H、-NR 12 -(CH2) q SO3H、-(CH2) q N(R 12 )2、-O-(CH2) q N(R 12 )2、-S-(CH2) q N(R 12 )2、-CO-(CH2) q N(R 12 )2、-(CH2) q N(R 12 )3、-O-(CH2) q N(R 12 )3、-S-(CH2) q N(R 12 )3、-CO-(CH2) q N(R 12 )3、-NR 12 -(CH2) q N(R 12 )3、-(CH2) q CON(R 12 )2、-O-(CH2) q CON(R 12 )2、-S-(CH2) q CON(R 12 )2、-CO-(CH2) q CON(R 12 )2、-(CH2) q PO(OR 12 )2、-O(CH2) q PO(OR 12 )2、-S(CH2) q PO(OR 12 )2、-CO(CH2) q PO(OR 12 )2、-NR 12 (CH2) q PO(OR 12 )2、-(CH2) q OPO(OR 12 )2、-O(CH2) q OPO(OR 12 )2、-S(CH2) q OPO(OR 12 )2、-CO(CH2)q OPO(OR 12 )2, and -NR 12 (CH2) q OPO(OR 12 )2 is selected.

[0208] In yet another embodiment, provided herein is a method for identifying one or more compounds that bind to a biological target, the method comprising: (a) incubating the biological target with at least a portion of a plurality of distinct tagged macrocyclic compounds of the compound library of claim 2 to generate at least one binding compound and at least one non-binding compound of the plurality of distinct tagged macrocyclic compounds; (b) removing at least one non-binding compound; and (c) sequencing the oligonucleotide (D) of each of the at least one binding compound.

[0209] In certain embodiments, the DNA-encoded library can be a single-pharmacophore library, where only one chemical moiety can be attached to a single strand of DNA, for example, as described in Neri & Lerner, Annu. Rev. Biochem. (2018) 87:5.1-5.24, which is incorporated herein by reference in its entirety. In certain embodiments, the DNA-encoded library can be a dual-pharmacophore library, where only one chemical moiety can be attached to a double strand, for example, as described in Id; Mannocci et al., Chem. Commun. (2011) 47:12747-53, which is incorporated herein by reference in its entirety.

[0210] In a further aspect, provided herein are methods for generating a library of tagged macrocycles, the method comprising synthesizing a plurality of distinct tagged macrocycles. In certain embodiments, each tagged macrocycle of the plurality of distinct tagged macrocycles comprises a macrocycle operably linked to at least one oligonucleotide (D). In certain embodiments, each compound of the plurality of distinct tagged macrocycles of the compound library comprises a macrocycle operably linked to at least one oligonucleotide (D). In certain embodiments, the macrocycle comprises an FKBD, an effector domain, a first linking region, and a second linking region. In certain embodiments, the FKBD, the effector domain, the first linking region, and the second linking region together form a macrocycle. In certain embodiments, each of the at least one oligonucleotide (D) is capable of identifying at least one of the FKBD, the effector domain, the first linking region, and the second linking region of each of the plurality of distinct tagged macrocycles. In certain embodiments, each compound of the plurality of distinct tagged macrocycles of the compound library comprises a compound of Formula (A) (as defined above). In certain embodiments, each compound of the plurality of distinct tagged macrocycles of the compound library comprises a compound of Formula (I) (as defined herein above). In certain embodiments, each compound of the plurality of distinct tagged macrocycles of the compound library can be a reaction product operatively linking a compound of Formula (B) (as defined herein above) with a compound of Formula (C) (as defined herein above). In certain embodiments, each compound of the plurality of distinct tagged macrocycles of the compound library can be a reaction product operatively linking a compound of Formula (B') (as defined herein above) with a compound of Formula (C) (as defined herein above).

[0211] In certain embodiments, the method for synthesizing the library of compounds can be selected from the group consisting of the split-and-pool method, DNA-templated library synthesis (DTS), encoded self-assembling chemical (ESAC) library synthesis, DNA-recorded library synthesis, DNA-directed library synthesis, DNA routing, and 3-D proximity-based library synthesis (YoctoReactor). As a person of ordinary skill in the art would recognize, various techniques for synthesizing libraries of tagged macrocycles are known, e.g., Neri & Lerner, Annu. Rev. Biochem. (2018) 87:5.1-5.24; Roman et al., SLAS Discovery (2018) 23(5):387-396; Lim, C&EN (Chemical & Engineering News), (2017) 95 (29):10-10; Halford, C&EN, (2017) 95(25): 28-33; Estevez, Tetrahedron: Asymmetry. (2017) 28:837-842; Neri, Chembiochem. (European Journal of Chemical Biology) (2017) 4;18(9):827-828; Yuen & Franzini, Chembiochem. (2017) 4;18(9):829-836; Skopic et al., Chem Sci. (Chemical Science) (2017) 1;8(5):3356-3361; Shi et al., Bioorg Med Chem Lett.Bioorganic & Medicinal Chemistry Letters (2017) 1;27(3):361-69; Zimmermann & Neri, Drug Discovery Today. (2016) 21(11):1828-1834; Satz et al., Bioconjug Chem. (2015) 19;26(8):1623-32; Ding et al., ACS Comb Sci. (2016) 10;18(10):625-629; Arico-Muendel, MedChemComm. (2016) 7(10):1898-1909; Skopic, MedChemComm, (2016) 7(10): 1957-1965; Satz, C.S. Comb. Sci. (2016) 18 (7):415-424; Tian et al., MedChemComm, (2016) 7(7): 1316-1322; Salamon et al., ACS Chem Biol. (2016) 19;11(2):296-307; Satz et al., Bioconjug Chem. (2015) 19;26(8):1623-32; Connors et al., Curr Opin Chem Biol. (2015) 26:42-7; Blakskjaer et al., Curr Opin Chem Biol. (2015) 26:62-71; Scheuermann & Neri, Curr Opin Chem Biol. (2015) 26:99-103; Franzini et al., Angew Chem Int Ed Engl. (2015) 23;54(13):3927-31; Franzini et al., Bioconjug Chem.(2014) 20;25(8):1453-61; Franzini, Neri & Scheuermann, Acc Chem Res. (Accounts of Chemical Research) (2014) 15;47(4):1247-55; Mannocci et al., Chem. Commun. (2011) 47:12747-53; Kleiner et al., Chem Soc Rev. (Chemical Society Reviews) (2011) 40(12): 5707-17; Clark, Curr Opin Chem Biol. (2010) 14(3):396-403; Mannocci et al., Proc Natl Acad Sci US A. (Proceedings of the National Academy of Sciences of the United States of America) (2008) 18;105(46):17670-75; Buller et al., Bioorg Med Chem Lett. (2008) 18(22):5926-31; Scheuermann et al., Bioconjugate Chem. (2008) 19:778-85; Zimmerman et al., ChemBioChem (A European Journal of Chemical Biology) (2017) 18(9):853-57, and Cuozzo et al., ChemBioChem (2017) 18(9):864-71, each of which is incorporated herein by reference in its entirety.

[0212] In some embodiments, the method for synthesizing a library of tagged macrocycles includes DNA-encoded library synthesis, in which encoding and library synthesis occur separately, e.g., as described in Shi et al., Bioorg Med Chem Lett. (2017) 1;27(3):361-369; Kleiner et al., Chem Soc Rev. (2011) 40(12): 5707-17. In certain embodiments, DNA-encoded library synthesis involves split-and-pooling methods, examples of which are described in Krall, Scheuermann & Neri, Angew Chem. Int. Ed Engl. (2013) 28;52(5):1384-402; Mannocci et al., Chem. Commun. (2011) 47:12747-53; and U.S. Patent No. 7,989,395 to Morgan et al., each of which is incorporated herein by reference in its entirety. In certain embodiments, the split-and-pooling method involves sequential chemical ligation of oligonucleotide tags to initial oligonucleotides (or headpieces), which can be covalently linked to the chemically generated entities by successive split-and-pooling steps. In certain embodiments, between each splitting step, a chemical synthesis step can be performed together with an oligonucleotide ligation step.

[0213] In some embodiments, libraries can be synthesized by a series of split-and-pool cycles, in which an initial oligonucleotide (or headpiece) can be reacted with a first set of building blocks (e.g., multiple FKBD building blocks). For each building block of the first set of building blocks (e.g., each FKBD building block), an oligonucleotide (D) can be added to the initial oligonucleotide (or headpiece), and the resulting products can be pooled (or mixed) and later separated into separate reactions. Later, in certain embodiments, a second set of building blocks (e.g., multiple effector domain building blocks) can be added, and an oligonucleotide (D) can be added to each building block of the second set of building blocks. In certain embodiments, each oligonucleotide (D) identifies a distinct building block.

[0214] In some embodiments, methods for synthesizing libraries of tagged macrocycles include DNA-directed library synthesis, where DNA both encodes and templates the library synthesis, examples of which are described in Kleiner et al., Bioconjugate Chem. (2010) 21, 1836-41; and Shi et al., Bioorg Med Chem Lett. (2017) 1;27(3):361-369, each of which is incorporated herein by reference in its entirety. In certain embodiments, DNA-directed library synthesis involves DNA-templated synthesis (DTS) methods, such as those described in Mannocci et al., Chem. Commun. (2011) 47:12747-53; Franzini, Neri & Scheuermann, Acc Chem Res. (2014) 15;47(4):1247-55; and Mannocci et al., Chem. Commun. (2011) 47:12747-53, each of which is incorporated herein by reference in its entirety. In certain embodiments, DTS methods involve DNA oligonucleotides that not only code but also direct the construction of the library. See Buller et al., Bioconjugate Chem. (2010) 21, 1571-80, which is incorporated herein by reference in its entirety. In certain embodiments, different building blocks can be incorporated into molecules using DNA ligation reagents, which can be forced into proximity by base pairing between their DNA tags. See Gartner et al., Science (2004) 305:1601-05, which is incorporated herein by reference in its entirety. In certain embodiments, a library of long oligonucleotides can be first synthesized as a template for a DNA-encoded library. In certain embodiments, oligonucleotides can be subjected to sequence-specific chemical reactions via immobilization on a resin tagged with complementary DNA sequences.See Wrenn & Harbury, Annu. Rev. Biochem. (2007) 76:331-49, which is incorporated herein by reference in its entirety.

[0215] In certain embodiments, DNA-directed library synthesis comprises 3D proximity-based library synthesis, also known as YoctoReactor technology, an example of which is described in Blakskjaer et al., Curr Opin Chem Biol. (2015) 26:62-7, which is incorporated herein by reference in its entirety.

[0216] In certain embodiments, methods for synthesizing libraries of tagged macrocycles include encoded self-assembly chemistry (ESAC) library synthesis, also known as double pharmacophore DNA-encoded chemical libraries, e.g., as described in Mannocci et al., Chem. Commun. (2011) 47:12747-53; Melkko et al., Nat. Biotechnol. (2004) 22(5):568-74; Scheuermann et al., Bioconjugate Chem. (2008) 19:778-85; and U.S. Pat. No. 8,642,215 to Neri et al., each of which is incorporated herein by reference in its entirety. In certain embodiments, synthesizing a library of tagged macrocycles by ESAC synthesis can include, for example, non-covalent combinatorial assembly of complementary oligonucleotide sub-libraries, where each sub-library can include a first oligonucleotide attached to a first building block, where the first oligonucleotide includes a coding domain that identifies the first building block and a hybridization domain that self-assembles to a second oligonucleotide attached to a second building block, where the second oligonucleotide includes a coding domain that identifies the second building block and a hybridization domain that self-assembles to the first oligonucleotide.

[0217] In some embodiments, methods for synthesizing libraries of tagged macrocycles include DNA routing, for example, as described in Clark, Curr Opin Chem Biol. (2010) 14(3):396-403, which is incorporated herein by reference in its entirety.

[0218] In certain embodiments, oligonucleotide ligation can utilize one of several methods that would be recognized by a person of ordinary skill in the art, such as those described in Zimmermann & Neri, Drug Discovery Today. (2016) 21(11):1828-1834; and Keefe et al., Curr Opin Chem Biol. (2015) 26:80-88, each of which is incorporated herein by reference in its entirety. In certain embodiments, oligonucleotide ligation can be enzymatic ligation. In certain embodiments, oligonucleotide ligation can be chemical ligation.

[0219] In certain embodiments, ligation involves base-pairing a short, complementary "adapter" oligonucleotide to the single-stranded oligonucleotide at either end of the ligation site, allowing ligation of a single-stranded DNA tag in each cycle. See Clark et al., Nat. Chem. Biol. (2009) 5:647-54, which is incorporated herein by reference in its entirety. In certain embodiments, oligonucleotide ligation involves utilizing two-base overhangs at the 3' end of the headpiece and of the DNA tag of each building block to form sticky ends for ligation. In certain embodiments, the sequence of the overhangs can depend on the cycle, but not the building blocks, such that any DNA tag can be ligated to any DNA tag from the previous cycle, but not the truncated sequence. See id. In certain embodiments, the oligonucleotide ligation step can utilize an opposite-sense oligonucleotide for subsequent cycles, where the two oligonucleotides have a small overlapping region where they are complementary. In certain embodiments, instead of ligation, DNA polymerase can be used to fill in the remaining complementary sequence, creating a double-stranded oligonucleotide containing both tags. In certain embodiments, the ligation of the oligonucleotides can be chemical. Without wishing to be bound by theory, it is believed that chemical ligation can allow for greater flexibility in terms of solution conditions and can reduce the number of required buffer exchange steps. See Keefe et al., Curr Opin Chem Biol. (2015) 26:80-88, which is incorporated herein by reference in its entirety.

[0220] In certain embodiments, provided herein are methods for identifying one or more compounds that bind to a biological target, the method comprising: (a) incubating the biological target with at least a portion of a plurality of distinct tagged macrocyclic compounds of a compound library to generate at least one binding compound and at least one non-binding compound of the plurality of distinct tagged macrocyclic compounds; (b) removing the at least one non-binding compound; and (c) sequencing at least one oligonucleotide (D) of each of the at least one binding compound. In certain embodiments, each compound of the plurality of distinct tagged macrocyclic compounds of the compound library comprises a macrocyclic compound operably linked to at least one oligonucleotide (D). In certain embodiments, the macrocyclic compound comprises an FKBD, an effector domain, a first linking region, and a second linking region. In certain embodiments, the FKBD, the effector domain, the first linking region, and the second linking region together form a macrocycle. In certain embodiments, each at least one oligonucleotide (D) can identify at least one of the FKBD, effector domain, first linking region, and second linking region of each of the plurality of distinct tagged macrocycles. In certain embodiments, each compound of the plurality of distinct tagged macrocycles of the compound library comprises a compound of Formula (A) (as defined above). In certain embodiments, each compound of the plurality of distinct tagged macrocycles of the compound library comprises a compound of Formula (I) (as defined above). As a person of ordinary skill in the art would recognize, various techniques for synthesizing libraries of tagged macrocycles are described, for example, in Kuai et al., SLAS Discov. (2018) 23(5):405-416; Brown et al., Annu. Rev Biochem. (2018) 87:5.1-5.24; Roman et al., SLAS Discov. (2018) 23(5):387-396; Amigo et al., SLAS Discov. (2018) 23(5):397-404; Shi et al., Bioconjug Chem.(2017) 20;28(9):2293-2301; Machutta et al., Nat Commun. (2017) 8:16081; Li et al., Chembiochem. (2017) 4;18(9):848-852; Satz et al., ACS Comb Sci. (2017) 10;19(4):234-238; Denton & Krusemark, MedChemComm, (2016) 7(10): 2020-2027; Eidam & Satz, MedChemComm, (2016) 7(7): 1323-1331; Bao et al., Anal. Chem. (2016) 88 (10):5498-5506; Decurtins et al., Nat Protoc. (2016) 11(4):764-80; Harris et al., J. Med. Chem. (Journal of Medicinal Chemistry) (2016) 59 (5):2163-78; Satz, ACS Chem Biol. (2016) 16;10(10):2237-45; Chan et al., Curr Opin Chem Biol. (2015) 26:55-61; Franzini et al., Chem Commun. (2015) 11;51(38):8014-16; and Buller et al., Bioorg Med Chem Lett. (2010) 15;20(14):4188-92, each of which is incorporated herein by reference in its entirety.

[0221] In certain embodiments, the incubating step can be carried out under conditions suitable for at least one of the plurality of distinct tagged macrocyclic compounds of the compound library to bind to the biological target. A person of ordinary skill in the art will understand what conditions would be suitable for at least one of the plurality of distinct tagged macrocyclic compounds of the compound library to bind to the biological target.

[0222] In certain embodiments, identifying one or more compounds that bind to a biological target is described, for example, in Ding et al., ACS Med. Chem. Lett. (2015) 7;6(8):888-93, which is incorporated herein by reference in its entirety. In certain embodiments, the incubating step (a) comprises contacting a plurality of tagged compounds in the compound library with a target protein, wherein the target protein can be immobilized on a substrate (e.g., a resin). In certain embodiments, the removing step (b) comprises washing the substrate to remove at least one unbound compound. In certain embodiments, the sequencing step (c) comprises sequencing at least one oligonucleotide (D) to identify which of the plurality of tagged compounds bound to the target protein.

[0223] In certain embodiments, identifying one or more compounds that bind to a biological target includes utilizing an unmodified, non-immobilized target protein. Such a method can utilize a ligate-crosslink-purify strategy, an example of which is described in Shi et al., Bioconjug. Chem. (2017) 20;28(9):2293-2301, which is incorporated herein by reference in its entirety. In certain embodiments, other methods for identifying one or more compounds that bind to a biological target can be utilized. Such methods will be apparent to persons of ordinary skill in the art, and examples of such methods are described, for example, in Machutta et al., Nat. Commun. (2017) 8:16081; Chan et al., Curr. Opin. Chem. Biol. (2015) 26:55-61; Lim, C&EN, (2017) 95 (29):10; Amigo et al., SLAS Discov. (2018) 23(5):397-404; Tian et al., MedChemComm. (2016) 7(7): 1316-1322; Satz, CS Comb. Sci. (2016) 18 (7):415-424, each of which is incorporated herein by reference in its entirety.

[0224] Tables 5-7 below illustrate all rapafucine compounds synthesized and characterized in the instant disclosure. In some embodiments, the disclosure does not include rapafucine compounds having AA2 as dmPhe. In some embodiments, the disclosure does not include rapafucine compounds having AA2 as dPro, dHoPro, or G. In some embodiments, the disclosure does not include rapafucine compounds having AA1 as G, mG, Pro, and dPro. [ka]

[0225] Tables 5-7 below show all rapafucine molecules in the present disclosure, with structural moieties shown according to formula (XIII), and an example of a chemical structure generated from formula (XIII) for compound 1 is shown below. In the case of amino acid monomers and FKBDs, a dehydration reaction occurs, resulting in a peptide bond. An example without specifying monomer 4 is a rapafucine composed of only an FKBD with a linker and three monomers. [ka] [Table 5-1] [Table 5-2] [Table 5-3] [Table 5-4] [Table 5-5] [Table 5-6] [Table 5-7] [Table 5-8] [Table 5-9] [Table 5-10] [Table 5-11] [Table 5-12] [Table 5-13]

Table 5-14

Table 5-15

Table 5-16

Table 5-17

[0226]

Table 6-1

Table 6-2

Table 6-3

Table 6-4

Table 6-5

Table 6-6

Table 6-7

Table 6-8

Table 6-9

Table 6-10

[0227]

Table 7-1

Table 7-2

Table 7-3

Table 7-4

Table 7-5

Table 7-6

Table 7-7

Table 7-8

Table 7-9

Table 7-10

Table 7-11

Table 7-12

Table 7-13

Table 7-14

Table 7-15

Table 7-16

Table 7-17

Table 7-18

Table 7-19

[0228] In treatment, the dosage of the agent optionally ranges from about 0.0001 mg / kg to about 100 mg / kg, from about 0.01 mg / kg to about 5 mg / kg, from about 0.15 mg / kg to about 3 mg / kg, from 0.5 mg / kg to about 2 mg / kg, and from about 1 mg / kg to about 2 mg / kg of the subject's body weight. In other embodiments, the dosage ranges from about 100 mg / kg to about 5 g / kg, from about 500 mg / kg to about 2 mg / kg, and from about 750 mg / kg to about 1.5 g / kg of the subject's body weight. For example, depending on the type and severity of the disease, from about 1 μg / kg to 15 mg / kg (e.g., 0.1-20 mg / kg) of the agent is a candidate dosage for administration to a patient, whether by one or more separate administrations or by continuous infusion, for example. A typical daily dosage ranges from about 1 μg / kg to 100 mg / kg or more, depending on the factors mentioned above. For repeated administration over several days or longer, depending on the symptoms, treatment is continued until a desired suppression of disease symptoms occurs. However, other dosage regimens may also be useful. The unit dose can be in the range, for example, from about 5 mg to 500 mg, such as 50 mg, 100 mg, 150 mg, 200 mg, 250 mg, and 300 mg. The progress of treatment is monitored by conventional techniques and assays.

[0229] In some embodiments, the agent is administered to a human patient in an effective amount (or dose) of less than about 1 μg / kg, e.g., from about 0.35 to about 0.75 μg / kg or from about 0.40 to about 0.60 μg / kg. In some embodiments, the dose of the agent is about 0.35 μg / kg, or about 0.40 μg / kg, or about 0.45 μg / kg, or about 0.50 μg / kg, or about 0.55 μg / kg, or about 0.60 μg / kg, or about 0.65 μg / kg, or about 0.70 μg / kg, or about 0.75 μg / kg, or about 0.80 μg / kg, or about 0.85 μg / kg, or about 0.90 μg / kg, or about 0.95 μg / kg, or 1 μg / kg. In various embodiments, the absolute dose of the agent is from about 2 μg / subject to about 45 μg / subject, or from about 5 to about 40, or from about 10 to about 30, or from about 15 to about 25 μg / subject. In some embodiments, the absolute dose of the agent is about 20 μg, or about 30 μg, or about 40 μg.

[0230] In various embodiments, the dose of the agent can be determined by the weight of the human patient. For example, about 2 μg for a pediatric human patient weighing from about 0 to about 5 kg (e.g., about 0, or about 1, or about 2, or about 3, or about 4, or about 5 kg); or about 3 μg for a pediatric human patient weighing from about 6 to about 8 kg (e.g., about 6, or about 7, or about 8 kg); or about 5 μg for a pediatric human patient weighing from about 9 to about 13 kg (e.g., 9, or about 10, or about 11, or about 12, or about 13 kg); or about 8 μg for a pediatric human patient weighing from about 14 to about 20 kg (e.g., about 14, or about 16, or about 18, or about 20 kg); or about 12 μg for a pediatric human patient weighing from about 21 to about 30 kg (e.g., about 21 kg, or about 23 kg, or about 25 kg, or about 27 kg, or about 30 kg). μg, or about 13 μg for a pediatric human patient weighing from about 31 to about 33 kg (e.g., about 31 kg, or about 32 kg, or about 33 kg), or about 20 μg for an adult human patient weighing from about 34 to about 50 kg (e.g., about 34, or about 36, or about 38, or about 40, or about 42, or about 44, or about 46, or about 48, or about 50 kg), or about 30 μg for an adult human patient weighing from about 51 to about 75 kg (e.g., about 51, or about 55, or about 60, or about 65, or about 70, or about 75 kg), or about 45 μg for an adult human patient weighing more than about 114 kg (e.g., about 114, or about 120, or about 130, or about 140, or about 150 kg).

[0231] In certain embodiments, the agents according to the methods provided herein are administered subcutaneously (sc), intravenously (iv), intramuscularly (im), intranasally, or topically. Administration of the agents described herein can be, independently, from 1 to 4 times per day, or from 1 to 4 times per month, or from 1 to 6 times per year, or once every 2, 3, 4, or 5 years. Administration can be for a period of 1 day, or 1 month, 2 months, 3 months, 6 months, 1 year, 2 years, 3 years, and even for the lifetime of the human patient. Dosages can be administered as a single dose or divided into multiple doses. In some embodiments, the agent is administered from about 1 to about 3 times (e.g., 1, or 2 or 3 times).

[0232] The following examples are provided to further illustrate the benefits and advantages of the present disclosure, but are not intended to limit the scope of the disclosure. While the examples are typical of those that might be used, other procedures, methodologies, or techniques known to those skilled in the art may alternatively be used. [Example]

[0233] General Experiments for Synthesis. Syntheti reagents. Piperidine and N,N-diisopropylethylamine (DIPEA) were purchased from Alfa Aesar. Anhydrous pyridine was purchased from Acros. Solid support resin with 2-chlorotrityl chloride (Cat#: 03498) was purchased from Chem-Impex. HATU was purchased from Chem-Impex. Fmoc-protected amino acid building blocks were purchased from Chem-Impex, Novabiochem, or GL Biochem. Dichloromethane (DCM or CH2Cl2), methanol (MeOH), hexane, ethyl acetate (EtOAc), 1,2-dichloroethane (DCE, anhydrous), N,N'-dimethylformamide (DMF, anhydrous), Hoveyda-Grubbs catalyst second generation, and all other chemical reagents were purchased from Sigma-Aldrich.

[0234] Instrumentation for synthesis and purification. MR spectra were recorded on a Bruker -400 and -500. High-performance liquid chromatography analysis was performed using an Agilent LC-MS system (Agilent 1260 series, mass detector 6120 quadrupole). Orbital shaking for solid-phase reactions was performed on a 96-tube Mettler-Toledo Bohdan MiniBlock system (30–200 mg of resin in SiliCycle tubes) or a VWR Mini Shaker (0.2–2 g of resin in plastic syringes with fritted discs). Reagents were added using an adjustable Rainin 8-channel pipette for the MiniBlock system. Microwave reactions were performed using a Biotage Initiator Plus or a Multiwave Pro with a silicon carbide 24-well block from Anton Parr. Compound purification on a 0.05-50 g scale was performed on a Teledyne Isco CombiFlash Rf 200 or Biotage Isolera One system followed by a Heidolph rotary evaporator. Purification on a 1-50 mg scale was performed on an Agilent HPLC system. The mixture of rapafucine from the 45,000 compound library was purified using a vacuum manifold (Sigma-Aldrich, Visiprep). TMPurification was performed in a high-throughput manner using SPE cartridges (Biotage, 460-0200-C, ISOLUTE, SI 2g / 6mL) on a Visiprep SPE Vacuum Manifold (12-port, Disposable Liner), followed by overnight drying using a custom-designed box (50cm x 50cm x 15cm) that allows for solvent removal by rapid internal airflow. High-throughput weighing of compounds within the library was performed using a Mettler-Toledo analytical balance (Sartorious Entris line with RS232 port) linked to a computer with a custom-coded electronic spreadsheet. FKBD Example 1 4-((3-((R)-1-((S)-1-(4-(acryloyloxy)-3,3-dimethyl-2-oxobutanoyl)piperidine-2-carbonyl)oxy)-3-(3,4-dimethoxyphenyl)propynyl)phenyl)amino)-4-oxobutanoic acid (aFKBD) [ka] Scheme 3. Synthesis of Intermediate 5 ((S)-1-(4-(acryloyloxy)-3,3-dimethyl-2-oxobutanoyl)piperidine-2-carboxylic acid)

[0235] 2-Allyl 1-(tert-butyl)(S)-piperidine-1,2-dicarboxylate (2). To a solution of N-Boc homoproline 1 (6.30 g) in DMF (40 mL) was added CsCO (2.90 g). The resulting suspension was stirred at room temperature for 5 min before the addition of allyl bromide (6.3 g). After stirring at RT for 2 h, the suspension was filtered through a pad of Celite, rinsed with EtOAc (50 mL), and washed with HCl (1 M, 50 mL × 3). The organic layer was dried over NaSO and coevaporated with toluene (30 mL × 2). The crude product (8.10 g) was collected as a yellow oil and was sufficiently pure for the next step without further purification. The crude product (8.10 g) and TFA (4.3 g) were mixed thoroughly in dichloromethane (20 mL) and stirred at RT for 0.5 h. 2-Allyl 1-(tert-butyl)(S)-piperidine-1,2-dicarboxylate 2 (3.00 g) was collected as a yellow oil and was pure enough for the next step without further purification.

[0236] Allyl (S)-1-(4-hydroxy-3,3-dimethyl-2-oxobutanoyl)piperidine-2-carboxylate (3). Compound 2 (3.0 g), dihydro-4,4-dimethyl-2,3-furandione (2.1 g), and DMAP (20 mg) were dissolved in toluene (20 mL) and refluxed for 14 h using an oil bath (120 °C). After removing the solvent, the residue was purified by column chromatography (80-200 mesh) with EtOAc / hexane (1 / 3). 3 (3.50 g) was collected as a yellow oil. 1 H NMR (500MHz, CDCl3)δ 6.04-5.80(m, 1H), 5.36(d, J=17Hz, 1H), 5.31-5.25(m, 2H), 4.68(s, 2H), 3.76-3.56(m, 2H), 3.50(d, J=13Hz, 1 H), 3.40(s, 1H), 3.20(t, J=13Hz, 1H), 2.37(d, J=13Hz, 1H), 1.84-1.61(m, 3H), 1.61-1.34(m, 2H), 1.24(s, 6H). 13C NMR (126MHz, CDCl3)δ 205.9, 170.1, 168.1, 131.4, 119.2, 69.3, 66.3, 51.6, 49.5, 44.2, 26.3, 24.8, 21.3, 21.2, 21.0.

[0237] Allyl (S)-1-(4-(acryloyloxy)-3,3-dimethyl-2-oxobutanoyl)piperidine-2-carboxylate (4). To a mixture of compound 3 (3.50 g) and N,N-diisopropylethylamine (2.0 mL) in 50 mL of CHCl was added acryloyl chloride (0.78 g) in dry CHCl (20 mL) dropwise over 30 min using an ice bath. After the addition, the reaction was stirred at RT for 30 min before being quenched with saturated aqueous NaHCO (20 mL). The organic phase was washed with water, dried over NaSO, concentrated, and purified by column (EtOAc:hexane = 1:5) to give product 4 (2.21 g) as a yellow oil. 1 H NMR(500MHz, CDCl3)δ(dd, J=17, 1.5Hz, 1H), 6.08(dd, J=17, 11Hz, 1H), 5.91(ddt, J=17, 11, 6Hz, 1H), 5.84(dd, J=11 , 1.5Hz, 1H), 5.35(ddd, J=17, 2.5, 1.5Hz, 1H), 5.28-5.25(m, 1H), 5.26(ddd, J=11, 2.5, 1.5Hz, 1H), 4.66(ddd, J=6, 4, 2.5Hz, 2H), 4.37(d, J=11Hz, 1H), 4.27(d, J=11Hz, 1H), 3.52(dd, J=13, 1.5Hz, 1H), 3.23(td, J=13, 3Hz, 1H), 2.34(d, J=14Hz, 1 H), 1.84-1.76(m, 1H), 1.76-1.67(m, 1H), 1.67-1.60(m, 1H), 1.59-1.47(m, 1H), 1.47-1.38(m, 1H), 1.36(s, 3H), 1.35(s, 3H). 13C NMR (126 MHz, CDCl3) δ 204.8, 169.8, 166.7, 165.5, 131.5, 131.2, 128.0, 118.9, 69.5, 69.3, 66.0, 51.3, 46.7, 43.9, 26.4, 24.9, 22.2, 21.5, 21.1. [M+H]+ HRMS calculated for C18H25NO6: 352.1760, observed: 352.1753.

[0238] (S)-1-(4-(acryloyloxy)-3,3-dimethyl-2-oxobutanoyl)piperidine-2-carboxylic acid (5). Compound 4 (4.2 g), Pd(PPh3)4 (230 mg), and N-methylaniline (2.5 mL) were dissolved in THF (40 mL) and stirred at RT for 6 h. The reaction mixture was then diluted with EtOAc (80 mL) and washed with HCl (1 M, 50 mL × 3). The organic phase was separated, dried over Na2SO4, filtered, and concentrated. The crude product was purified using column chromatography (200-400 mesh), where the by-product can be eluted with 2% MeOH in dichloromethane, followed by the desired product with 3% MeOH and 0.1% AcOH in dichloromethane. 5 (2.55 g) was collected as a white solid (66%). 1 H NMR (500MHz, CDCl3)δ 9.96(s, 1H), 6.39(d, J=17Hz, 1H), 6.08(dd, J=17, 10Hz, 1H), 5.85(d, J=10Hz, 1H), 5.30(s, 1H), 4.55-4.30(m, 1H), 4.32(d, J=6Hz, 2 H), 3.53(d, J=12Hz, 1H), 3.24(t, J=12Hz, 1H), 2.35(d, J=13Hz, 1H), 1.91-1.60(m, 2H), 1.60-1.42(m, 2H), 1.36(s, 3H), 1.34(s, 3H). 13 C NMR (126 MHz, CDCl3) δ 204.7, 175.3, 166.8, 165.7, 131.4, 127.8, 69.6, 69.5, 51.2, 46.7, 44.0, 26.2, 24.9, 22.1, 21.8, 21.1. HRMS calculated for [M+H]+ C15H21NO6: 312.1447, observed: 312.1444. [ka] Scheme 4. Synthesis of aFKBD

[0239] (E)-1-(3-Aminophenyl)-3-(3,4-dimethoxyphenyl)prop-2-en-1-one (6). To a solution of 3,4-dimethoxybenzaldehyde (5.10 g) and 3-aminoacetophenone (4.15 g) in EtOH (20 mL, 95%) was added NaOH (0.2 g in 2 mL of water). The reaction mixture was stirred at RT for 6 h, and a yellow precipitate formed. The reaction mixture was then diluted with EtOAc (40 mL) and washed with water (30 mL × 3). After concentration, the crude product 6 (9.0 g) was sufficiently pure for the next step.

[0240] 1-(3-Aminophenyl)-3-(3,4-dimethoxyphenyl)propan-1-one (7). To a solution of α,β-unsaturated ketone 6 (crude, 9.0 g) in MeOH (20 mL) was added Pd / C (10%, 1.61 g). The reaction vessel was repeatedly flushed with hydrogen gas using a hydrogen balloon and high vacuum. The reaction mixture was stirred at RT for 1 h and then filtered through a pad of Celite. Longer reaction times lead to the formation of undesired by-products. The filtrate was concentrated and subjected to column chromatography (50 g silica gel) and eluted with EtOAc / CHCl / hexane (1 / 3 / 3 to 1 / 1 / 1). 7 (2.48 g) was collected as a yellow oil. 1 H NMR (500MHz, CDCl3)δ 7.36-7.16(m, 3H, ar), 6.92-6.71(m, 4H, ar), 3.86(s, 3H, OCH3), 3.85(s, 3H, OCH 3), 3.81(s, 2H, NH2), 3.23(t, J=7.5Hz, 2H, COCH2), 2.99(t, J=7.4Hz, 2H, ArCH2). 13C NMR (126MHz, CDCl3)δ 199.66(C=O), 148.90(ar), 147.38(ar), 146.82(ar), 138.03(ar), 134.03(ar), 129.49(ar), 120.19(ar), 119.61(ar), 118.44(ar), 113.91(ar) , 111.87(ar), 111.35(ar), 55.98(OCH3), 55.87(OCH3), 40.80(COCH2), 29.91(ArCH2). HRMS for [M+H]+C17H19NO3, calculation: 286.1443, observation: 286.1436.

[0241] 4-((3-(3,4-Dimethoxyphenyl)propanoyl)phenyl)amino)-4-oxobutanoic acid (9). Aniline 7 (3.50 g), succinic anhydride (1.0 g), and DMAP (61 mg) were mixed in dichloromethane (30 mL). After stirring at RT for 3 h, the reaction mixture was washed with HCl (1 M, 30 mL × 4). The crude product (3.80 g) was collected as a white solid and used directly in the next step without further purification. Cs2CO3 (1.86 g) was added to a solution of the above crude product (3.80 g) in DMF (20 mL). The resulting suspension was stirred at RT for 10 min, and then allyl bromide (1.50 mL) was added. The reaction mixture was stirred for an additional 2 h. The white precipitate was removed by filtration through a pad of Celite. To the filtrate, EtOAc (40 mL) and H2O (40 mL) were added. After stirring for 10 min, the product precipitated. The product 9 (2.11 g) was obtained by filtration, air-dried as an off-white solid, and used for the next step without further purification.

[0242] (R)-1-(3-(4-(allyloxy)-4-oxobutanamido)phenyl)-3-(3,4-dimethoxyphenyl)propyl (S)-1-(4-(acryloyloxy)-3,3-dimethyl-2-oxobutanoyl)piperidine-2-carboxylate (10). Alcohol 9 (1.65 g) and carboxylic acid 5 (1.26 g, see FKBD Example 1 for synthesis) were dissolved in a mixture of THF (anhydrous, 5 mL) and dichloromethane (anhydrous, 10 mL). Benzoyl chloride (0.60 mL), EtN (1.0 mL), and DMAP (18 mg) were added sequentially, and the resulting suspension was stirred at RT for 2 h. Without further treatment, the mixture was subjected to column chromatography (80-200 mesh) with EtOAc / hexane (1 / 2 a 1 / 1). 10 (2.50 g) was collected as a yellow foam.1H NMR(500MHz、CDCl3)δ 8.08(s、1H)、7.65(d、J=8Hz、1H)、7.46(s、1H)、7.28(t、J=8Hz、1H)、7.01(d、J=8Hz、1H)、6.77(d、J=9Hz、1H)、6.69(d、J=5Hz、1H)、6.67(s、1H)、6.39(dd、J=17、1.5Hz、1H)、6.06(dd、J=17、10.5Hz、1H)、5.90(ddt、J=17、10.5、6Hz、1H)、5.83(dd、J=10.5、1.5Hz、1H)、5.79(ddd、J=10.5、8、3.5Hz、1H)、5.31(dd、J=17、1.5Hz、2H)、5.31(d、J=6Hz、1H)、5.22(dd、J=10.5、1.5Hz、1H)、4.60(dt、J=6、1.5Hz、2H)、4.33(d、J=0.7Hz、2H)、3.86(s、3H)、3.85(s、3H)、3.46(d、J=14Hz、1H)、3.09(dd、J=18, 8Hz、1H)、2.78(t、J=6Hz、2H)、2.70(t、J=6Hz、2H)、2.62-2.48(m、2H)、2.36(d、J=14Hz、1H)、2.30-2.16(m、1H)、2.13-2.00(m、1H)、1.74(d、J=10.5Hz、2H)、1.62(d、J=12Hz、1H)、1.42(d、J=12.6Hz、1H)、1.36(s、6H).13C NMR(126MHz、CDCl3)δ 205.6、172.6、169.8、169.3、166.2、165.6、148.9、147.3、140.7、138.6、133.5、132.0、131.5、129.2、127.8、122.0、120.2、119.3、118.4、117.2、111.7、111.3、76.5、69.2、65.5、55.9、55.9、51.3、46.8、44.1、38.1、31.9、31.1、29.3、26.1、25.1、22.0、21.9、20.9.。

[0243] 4-((3-((R)-1-(((S)-1-(4-(acryloyloxy)-3,3-dimethyl-2-oxobutanoyl)piperidine-2-carbonyl)oxy)-3-(3,4-dimethoxyphenyl)propyl)phenyl)amino-4-oxobutanoic acid (aFKBD).10 (2.50 g), Pd(PPh) (100 mg), and N-methylaniline (1.0 mL) were mixed well in THF (20 mL) at RT for 5 h. The reaction mixture was then diluted with EtOAc (5 mL). The mixture was diluted with HCl (1M, 50 mL x 3) and washed with HCl (1M, 50 mL x 3). The organic phase was dried over NaSO, filtered, and concentrated. The crude product was purified by column chromatography (200-400 mesh), where the by-product was eluted with 2% MeOH in dichloromethane, followed by the desired product with 3% MeOH and 0.05% AcOH in dichloromethane. aFKBD (2.25 g) was collected as an off-white foam. 1 H NMR(500MHz, CDCl3)δ 8.40(s, 1H), 7.62(s, 1H), 7.48(s, 1H), 7.26(t, J=7.5Hz, 1H), 7.01(d, J=7.5Hz, 1H), 6.97(dd, J=16, 7Hz, 1H), 6.86-6.74(m, 1H), 6.74-6.58(m, 2H), 5.85-5.68(m, 2H), 5.39-5.24(m, 1H), 4.29(q, J=11 Hz, 2H), 3.86(s, 3H), 3.84(s, 3H), 3.46(d, J=13Hz, 1H), 3.13(t, J=13Hz, 1H), 2.74(d, J=5.5Hz, 2H) , 2.69(d, J=5.5Hz, 2H), 2.63-2.48(m, 2H), 2.36(d, J=13Hz, 1H), 2.30-2.15(m, 1H), 2.15-1.99(m, 1 H), 1.85(d, J=6Hz, 1H), 1.75(d, J=12Hz, 1H), 1.63(d, J=13Hz, 1H), 1.55-1.38(m, 2H), 1.34(s, 6H). 13C NMR (126 MHz, CDCl3) δ 205.6, 176.8, 170.4, 169.4, 166.4, 166.1, 148.9, 147.3, 145.9, 140.7, 138.5, 133.5, 129.2, 122.1, 121.9, 120.2, 119.5, 117.4, 111.8, 111.4, 76.6, 69.0, 55.9, 55.8, 51.4, 46.8, 44.1, 38.1, 31.6, 31.1, 29.3, 26.2, 25.0, 21.8, 20.9, 18.1. HRMS [M+H]+C36H44O2N11 calculated: 681.3023, observed: 681.3018. FKBD Example 2 2-(3-((R)-1-(((S)-1-(4-(acryloyloxy)-3,3-dimethyl-2-oxobutanoyl)piperidine-2-carbonyl)oxy)-3-(3,4-dimethoxyphenyl)propyl)phenoxy)acetic acid (eFKBD) [ka] Scheme 5. Synthesis of eFKBD

[0244] (R)-1-(3-(2-(tert-butoxy)-2-oxoethoxy)phenyl)-3-(3,4-dimethoxyphenyl)propyl (S)-1-(4-(acryloyloxy)-3,3-dimethyl-2-oxobutanoyl)piperidine-2-carboxylate (6). Alcohol 4 (3.8 g, 1.0 eq.). Its synthesis is described in Liu et al. (2014). Angew. Chem. Int. Ed.(See Angewandte Chemie Internationale Edition) 53:10049-55), carboxylic acid 5 (4.1 g, 1.2 eq.; see FKBD Example 1 for synthesis), and DMAP (134 mg, 0.1 eq.) were dissolved in a mixture of THF (anhydrous, 35 mL) and dichloromethane (anhydrous, 35 mL) in a round-bottom 42 flask under argon protection. EtN (4.7 mL) and benzoyl chloride (2.17 mL, 2.62 g, 1.7 eq.) were added dropwise in sequence via syringe, and the resulting suspension was stirred at room temperature for 2 h. The reaction was monitored by TLC. When complete conversion was achieved, the reaction mixture was diluted with 500 mL of EtOAc and washed with 5% HCl and saturated NaHCO3. The organic phase was washed with brine and dried over Na2SO4. The solvent was then removed and the product was purified by column chromatography (80-200 mesh) using EtOAc / hexane (1 / 10 to 1 / 3). 6 (5.3 g, 69%) was collected as a light yellow foam. 1 H NMR (500MHz, CDCl3)δ 7.26(d, J=8Hz, 1H), 6.97(d, J=8.5Hz, 1H), 6.93-6.89(m, 1H), 6.86-6.81(m, 1H), 6.78(d, J=8.5Hz, 1H), 6.71-6.64(m, 2H), 6.38(dd, J=17, 1.5Hz, 1H), 6.06(dd, J=17, 10.5Hz, 1H), 5.82(dd, J=10.5, 1.5Hz, 1H), 5.78(dd, J=8, 6Hz, 1H), 5.29(d, J=5Hz, 1H), 4.53(s, 2H), 4.36(d, J=11Hz, 1H), 4.27(d, J =11Hz, 1H), 3.86(s, 3H), 3.84(s, 3H), 3.48(d, J=13Hz, 1H), 3.17(td, J=13, 3.0Hz, 1H), 2.67-2.44(m, 2H), 2.37(d, J=14Hz, 1H), 2.32-2.18(m, 1H), 2.14-1.99(m, 1H), 1. 83-1.65(m, 2H), 1.65-1.56(m, 1H), 1.50-1.43(m, 2H), 1.48(s, 9H), 1.35(s, 3H), 1.35(s, 3H). 13C NMR (126 MHz, CDCl3) δ 204.8, 169.4, 167.8, 166.4, 165.4, 158.1, 148.9, 147.3, 141.3, 133.4, 131.2, 129.7, 127.9, 120.2, 119.8, 114.2, 113.2, 111.7, 111.3, 82.3, 76.7, 69.2, 65.7, 55.9, 55.8, 51.4, 46.6, 44.0, 37.9, 31.2, 28.0, 26.4, 25.0, 22.1, 21.6, 21.1. HRMS for [M+H]+C38H49NO11, calculated: 696.3384, observed: 696.3386.

[0245] 2-(3-((R)-1-(((S)-1-(4-(acryloyloxy)-3,3-dimethyl-2-oxobutanoyl)piperidine-2-carbonyl)oxy)-3-(3,4-dimethoxyphenyl)propyl)phenoxy)acetic acid (eFKBD). Compound 6 (5.3 g, 1.0 eq) was dissolved in 60 mL of dichloromethane under Ar protection in a round-bottom flask. TFA (17 mL, 11.4 g, 13 eq.) was then added via syringe in three portions over a period of 3.5 h with stirring at room temperature. The reaction was monitored by TLC. When complete conversion was achieved, the solvent and TFA were removed under reduced pressure. The product was purified by column chromatography (80-200 mesh) using EtOAc / hexane (1 / 5a1 / 1). eFKBD (4.6 g, 96%) was collected as a pale yellow foam. 1H NMR (500MHz, CDCl3)δ 7.28(dd, J=3.5Hz, 3.5Hz, 1H), 6.88(d, J=8.5Hz, 1H), 6.83-6.81(m, 2H), 6.80-6.78(m, 1H), 6.69 -6.67(m, 2H), 6.37(d, J=8.5Hz, 1H), 6.05-6.02(m, 1H), 5.83-5.72(m, 2H), 5.30-5.28(dd, J=10, 5Hz, 1H), 4.67(dd, J=10, 5Hz, 1H), 4.17(dd, J=10, 6Hz, 2H), 3.48-3.45(m, 1H), 3.24-3.22(m, 1H), 2.61-2.55(m, 2H), 2.38(m, 1H), 2.23(m, 1H) , 2.04(m, 1H), 1.79(m, 1H), 1.62(m, 1H), 1.33(m, 1H), 1.30(m, 1H), 1.25(s, 3H), 1.24(s, 3H). 13 C NMR (126 MHz, CDCl3) δ 204.6, 169.2, 166.7, 165.7, 157.9, 149.0, 147.5, 141.7, 131.4, 129.9, 127.9, 120.0, 115.4, 111.8, 111.4, 111.1, 69.3, 65.2, 60.5, 55.9, 51.7, 44.1, 38.0, 31.4, 22.1, 21.1, 14.2. HRMS calculated for [M+H]+C34H42NO11: 640.2758, observed: 640.2761. FKBD Example 3 4-(3-((R)-1-((S)-1-(4-(acryloyloxy)-3,3-dimethyl-2-oxobutanoyl)piperidine-2-carbonyloxy)-3-morpholinopropyl)phenylamino)-4-oxobutanoic acid (Raa1) [ka] Scheme 6. Synthesis of Raa1 FKBD moiety

[0246] 1-(3-Nitrophenyl)prop-2-en-1-one (2). Paraformaldehyde (36 g, 120 mmol) was added to a stirred solution of 1-(3-nitrophenyl)ethanone 1 (20 g, 120 mmol), N-methylanilinium trifluoroacetate (26.8 g, 120 mmol), and TFA (1.4 g, 12 mmol) in THF (300 mL) at room temperature, and the resulting reaction mixture was heated at reflux for 16 h. The solvent was removed in vacuo, and the residue was diluted with water (100 mL) and EA (200 mL). The organic extract was dried over Na2SO4 and concentrated in vacuo to provide compound 2 as a yellow solid (14.2 g, crude), which was used directly in the next step without purification. [M+H] + =178.1.

[0247] 3-Morpholino-1-(3-nitrophenyl)propan-1-one (3). To a solution of 2 (12 g, 33.9 mmol, crude) in DMF (30 mL) was added Morpholine (2.95 g, 33.9 mmol), followed by 4-methylbenzenesulfonic acid (5.83 g, 33.9 mmol). After stirring at room temperature for 5 h, the reaction was quenched with HO (50 mL) and extracted with EA (100 mL × 3). The organic extract was dried over NaSO and concentrated under reduced pressure to give the crude product, which was further purified by column chromatography (SiO, methanol / DCM = 0-10% as eluent) to afford compound 3 (6.6 g, 74%) as a yellow oil. [M+H] + =265.2

[0248] 1-(3-aminophenyl)-3-morpholinopropan-1-one (4). To a solution of 3 (4.2 g, 15.9 mmol) in THF (20 mL) was added 10% Pd / C (wet, 840 mg) at rt. The resulting reaction mixture was hydrogenated with H2(g) at rt for 8 h. The reaction mixture was then filtered and concentrated in vacuo to provide crude compound 4 (3.46 g, crude) as a yellow oil, which was used directly for the next step. [M+H] + =235.1

[0249] tert-Butyl 4-(3-(3-morpholinopropanoyl)phenylamino)-4-oxobutanoate (6). To a solution of 4 (5.05 g, 21.5 mmol) and 4-tert-butoxy-4-oxobutanoic acid 5 (4.86 g, 27.95 mmol) in DMF (20 mL) was added DIPEA (5.55 g, 43 mmol), followed by HATU (10.62 g, 27.95 mmol) at rt. The resulting reaction mixture was stirred at rt for 2 h. The reaction was quenched with HO (50 mL) and extracted with EA (100 mL × 3). The organic extract was dried over NaSO and concentrated in vacuo to give the crude product, which was further purified by column chromatography (SiO, methanol / DCM = 0-5% as eluent) to afford compound 6 (4.3 g, 51%) as a yellow solid. [M+H] + =391.0

[0250] (R)-tert-Butyl 4-(3-(1-hydroxy-3-morpholinopropyl)phenylamino)-4-oxobutanoate (7). To a solution of ketone 6 (4.1 g, 10.5 mmol) in anhydrous THF (40 mL) was added (+)DIPChloride (42 mmol) in heptane (1.7 M, 24.7 mL) at −20° C. The resulting reaction mixture was stirred at −20° C. until complete conversion of 6, which was then quenched with 2,2′-(ethane-1,2-diylbis(oxy))diethanamine (7 g, 47.25 mmol) to form an insoluble complex. After stirring at rt for another 30 min, the suspension was filtered through a pad of Celite and concentrated in vacuo to give the crude product, which was further purified by column (SiO, CHOH / EA=0-5% as eluent) to give compound 7 (1.0 g, 24%) as an off-white solid. [M+H] + =393.0

[0251] (S)-((R)-1-(3-(4-tert-butoxy-4-oxobutanamido)phenyl)-3-morpholinopropyl)1-(4-(acryloyloxy)-3,3-dimethyl-2-oxobutanoyl)piperidine-2-carboxylate (9). A solution of 7 (1.0 g, 2.55 mmol) and 8 (952 mg, 3.06 mmol) in anhydrous DCM (25 mL) was cooled to −20° C., followed by the addition of a solution of DCC (630 mg, 3.06 mmol) in anhydrous DCM (2 mL), followed by the addition of a solution of 4-(dimethylamino)pyridine (DMAP, 31 mg, 0.255 mmol) under an argon atmosphere. The resulting white suspension was stirred at −20° C. for 2 h. The reaction mixture was then filtered, and the filtrate was dried over NaSO and concentrated under reduced pressure to give the crude product, which was further purified by column (SiO, CHOH / DCM=0-5% as eluent) to give compound 9 (1.3 g, 76%) as a white solid. [M+H] + =686.0

[0252] 4-(3-((R)-1-((S)-1-(4-(acryloyloxy)-3,3-dimethyl-2-oxobutanoyl)piperidine-2-carbonyloxy)-3-morpholinopropyl)phenylamino)-4-oxobutanoic acid (RAa-1) To a solution of 9 (1.3 g, 1.9 mmol) in DCM (10 mL) was added TFA (2 mL) at rt. The resulting mixture was stirred at rt for 3 h. The reaction mixture was directly loaded onto a silica gel flash column (CHOH / DCM = 0-5% as eluent) to give RAa-1 (620 mg, 51%) as a white solid. FKBD Example 4 4-(3-((R)-1-((S)-1-(4-(acryloyloxy)-3,3-dimethyl-2-oxobutanoyl)piperidine-2-carbonyloxy)-4-morpholinobutyl)phenylamino)-4-oxobutanoic acid (Raa2) [ka] Scheme 7. Synthesis of Raa2 FKBD Moiety

[0253] 3-(3-Nitrobenzoyl)-dihydrofuran-2(3H)-one (3). To a stirred solution of dihydrofuran-2(3H)-one 2 (6.02 g, 70 mmol) in anhydrous THF (60 mL) was added LiHMDS (1 M in THF, 77 mL, 77 mmol) at -78 °C and stirred for 2 h under an argon atmosphere. Next, a solution of 3-nitrobenzoyl chloride 1 (6.5 g, 35 mmol) in anhydrous THF (10 mL) was added at -78 °C. The resulting reaction mixture was slowly warmed to rt and stirred at rt for 16 h. Saturated NH4Cl aq The reaction was quenched with EA (20 mL) and extracted with EA (100 mL x 3). The organic extract was dried over NaSO and concentrated in vacuo to give compound 3 (8.5 g, crude) as a yellow oil, which was used directly for the next step without purification. [M+H] + =236.1

[0254] A solution of 4-bromo-1-(3-nitrophenyl)butan-1-one (4).3 (25.9 g, 110 mmol, crude) in 40% HBr (150 mL) was heated to 70 °C for 2 h. The reaction mixture was cooled to rt and saturated NaHCO 3aq The pH was adjusted to 5-6 with HCl and extracted with EA (200 mL × 3). The organic extract was dried over NaSO and concentrated in vacuo to give the crude product, which was further purified by column (SiO, EA / PE = 0-10% as eluent) to afford compound 4 (18.5 g, 74% for two steps) as a yellow oil.

[0255] 4-Morpholino-1-(3-nitrophenyl)butan-1-one (5). To a solution of 4 (8.5 g, 31.25 mmol) and Morpholine (2.72 g, 31.25 mmol) in CHCN (100 mL) was added KCO (8.64 g, 62.5 mmol) at rt. The resulting reaction mixture was heated to reflux for 2 h. The reaction mixture was then filtered and concentrated in vacuo to give the crude product, which was further purified by column chromatography (SiO, methanol / DCM = 0-5% as eluent) to give compound 5 (4.6 g, 53%) as a yellow oil. [M+H] + =279.2

[0256] 1-(3-aminophenyl)-4-morpholinobutan-1-one (6). To a solution of 5 (5.9 g, 21.2 mmol) in THF (60 mL) was added 10% Pd / C (wet, 1.18 g) at rt. The resulting reaction mixture was hydrogenated with H2(g) at rt for 10 h. The reaction mixture was filtered and concentrated in vacuo to provide crude compound 6 (4.8 g, crude) as a yellow solid, which was used dirtyly for the next step. [M+H] + =249.0

[0257] To a solution of 6 (4.8 g, 19.35 mmol) and 4-tert-butoxy-4-oxobutanoic acid 7 (4.86 g, 27.95 mmol) in DMF (15 mL) was added DIPEA (5.0 g, 38.7 mmol) at rt, followed by HATU (9.56 g, 25.15 mmol). The resulting reaction mixture was stirred at rt for 2 h. The reaction was quenched with HO (50 mL) and extracted with EA (100 mL × 3). The organic extract was dried over NaSO and concentrated in vacuo to give the crude product, which was further purified by column chromatography (SiO, methanol / DCM = 0-5% as eluent) to give compound 8 (6.6 g, 84%) as a yellow solid. [M+H] + =405.0

[0258] (R)-tert-Butyl 4-(3-(1-hydroxy-4-morpholinobutyl)phenylamino)-4-oxobutanoate (9). To a solution of ketone 8 (5.0 g, 12.4 mmol) in anhydrous THF (20 mL) was added (+)DIPChloride (49.6 mmol) in heptane (1.7 M, 29 mL) at −20° C. The resulting reaction mixture was stirred at −20° C. until complete conversion of 8, and then quenched with 2,2′-(ethane-1,2-diylbis(oxy))diethanolamine (8.3 g, 55.8 mmol) by forming an insoluble complex. After stirring at rt for another 30 min, the suspension was filtered through a pad of Celite and concentrated in vacuo to give the crude product, which was further purified by column (SiO, CHOH / EA=0-5% as eluent) to give compound 9 (2.5 g, 50%) as an off-white solid. [M+H] + =407.3

[0259] (S)-((R)-1-(3-(4-tert-butoxy-4-oxobutanamido)phenyl)-4-morpholinobutyl)1-(4-(acryloyloxy)-3,3-dimethyl-2-oxobutanoyl)piperidine-2-carboxylate (11). A solution of 9 (2.45 g, 6.05 mmol) and 10 (2.29 g, 7.38 mmol) in anhydrous DCM (40 mL) was cooled to −20° C., followed by the addition of a solution of DCC (1.52 g, 7.38 mmol) in anhydrous DCM (5 mL), followed by the addition of a solution of 4-(dimethylamino)pyridine (DMAP, 75 mg, 0.615 mmol) in anhydrous DCM (1 mL) under an argon atmosphere. The resulting white suspension was stirred at −20° C. for 2 h. The reaction mixture was then filtered, and the filtrate was dried over NaSO and concentrated under reduced pressure to give the crude product, which was further purified by column (SiO, CHOH / DCM=0-5% as eluent) to give compound 11 as a white solid (3 g, 69%). [M+H] + =700.0

[0260] 4-(3-((R)-1-((S)-1-(4-(acryloyloxy)-3,3-dimethyl-2-oxobutanoyl)piperidine-2-carbonyloxy)-4-morpholinebutyl)phenylamino)-4-oxobutanoic acid (Raa2). To a solution of 11 (1.0 g, 1.42 mmol) in DCM (10 mL) was added TFA (2 mL) at rt. The resulting mixture was stirred at rt for 2 h. The reaction mixture was directly charged onto a silica gel flash column (CHOH / DCM = 0-5% as eluent) to afford Raa2 (550 mg, 60%) as a white solid. FKBD Example 5 4-(3-((R)-3-(4-((9H-fluoren-9-yl)methoxy)carbonyl)piperazin-1-yl)-1-((S)-1-(4-(acryloyloxy)-3,3-dimethyl-2-oxobutanoyl)piperidine-2-carbonyloxy)propyl)phenylamino)-4-oxobutanoic acid (Raa3) [ka] Scheme 8. Synthesis of Raa3 FKBD moiety

[0261] tert-Butyl 4-(3-(3-nitrophenyl)-3-oxopropyl)piperazine-1-carboxylate (3). To a solution of 1 (10 g, 28.2 mmol, crude) in DMF (20 mL) was added DIPEA (3.64 g, 28.2 mmol), followed by 2 (5.24 g, 28.2 mmol). After stirring at room temperature for 2 h, the reaction was quenched with HO (100 mL) and extracted with EA (100 mL × 3). The organic extract was dried over NaSO and concentrated in vacuo to give the crude product, which was further purified by column chromatography (SiO, methanol / DCM = 0-10% as eluent) to give compound 3 as a yellow oil (6.1 g, 60%). [M+H] + =364.2

[0262] tert-Butyl 4-(3-(3-aminophenyl)-3-oxopropyl)piperazine-1-carboxylate (4). To a solution of 3 (6.1 g, 15.9 mmol) in THF (50 mL) at rt was added 10% Pd / C (wet, 1.22 g). The resulting reaction mixture was hydrogenated with H2(g) at rt for 8 h. The reaction mixture was then filtered and concentrated in vacuo to provide crude compound 4 (5.5 g, crude) as a brown solid, which was used directly for the next step. [M+H] + =334.3

[0263] tert-Butyl 4-(3-(3-(4-tert-butoxy-4-oxobutanamido)phenyl)-3-oxopropyl)piperazine-1-carboxylate (6). To a solution of 4 (5.2 g, 15.6 mmol) and 4-tert-butoxy-4-oxobutanoic acid 5 (3.53 g, 20.27 mmol) in DMF (35 mL) was added DIPEA (5.04 g, 38.99 mmol) at rt, followed by HATU (7.71 g, 20.27 mmol). The resulting reaction mixture was stirred at rt for 4 h. The reaction was quenched with HO (50 mL) and extracted with EA (100 mL × 3). The organic extract was dried over Na2SO4 and concentrated in vacuo to give the crude product, which was further purified by column (SiO2, PE / EA = 0-50% as eluent) to give compound 6 (4.3 g, 56%) as a yellow solid. [M+H] + =490.4

[0264] (R)-tert-Butyl 4-(3-(3-(4-tert-butoxy-4-oxobutanamido)phenyl)-3-hydroxypropyl)piperazine-1-carboxylate (7). To a solution of ketone 6 (3.8 g, 7.76 mmol) in anhydrous THF (30 mL) was added (+)DIPChloride (38.8 mmol) in heptane (1.7 M, 23 mL) at −20 °C. The resulting reaction mixture was stirred at −20 °C until complete conversion of 6 and quenched with 2,2′-(ethane-1,2-diylbis(oxy))diethanamine (6.32 g, 42.68 mmol) by forming an insoluble complex. After stirring at rt for another 30 min, the suspension was filtered through a pad of Celite and concentrated in vacuo to give the crude product, which was further purified by column (SiO, CHOH / EA=0-5% as eluent) to give compound 7 (1.9 g, 51%) as an off-white solid. [M+H] + =492.3

[0265] tert-Butyl 4-((R)-3-((S)-1-(4-(acryloyloxy)-3,3-dimethyl-2-oxobutanoyl)piperidine-2-carbonyloxy)-3-(3-(4-(tert-butoxy-4-oxobutanamido)phenyl)propyl)piperazine-1-carboxylate (9). A solution of 7 (1.03 g, 2.1 mmol) and 8 (784 mg, 2.52 mmol) in anhydrous DCM (20 mL) was cooled to -20 °C, followed by addition of DCC (865 mg, 4.2 mmol) in anhydrous DCM. A solution of 4-(dimethylamino)pyridine (DMAP, 26 mg, 0.21 mmol) was added under an argon atmosphere, followed by the addition of a solution of 4-(dimethylamino)pyridine (DMAP, 26 mg, 0.21 mmol). The resulting white suspension was stirred at −20° C. for 2 hours. The reaction mixture was then filtered, and the filtrate was dried over NaSO and concentrated under reduced pressure to give the crude product, which was further purified by column chromatography (SiO, CHOH / DCM=0-5% as eluent) to give compound 9 (1.2 g, 72%) as a yellow solid. [M+H] + =784.9

[0266] 4-(3-((R)-1-((S)-1-(4-(acryloyloxy)-3,3-dimethyl-2-oxobutanoyl)piperidine-2-carbonyloxy)-3-(piperazin-1-yl)propyl)phenylamino)-4-oxobutanoic acid (10). To a solution of 9 (1.2 g, 1.9 mmol) in DCM (6 mL) was added TFA (3 mL) at rt. The resulting mixture was stirred at rt for 3 h. The reaction mixture was concentrated in vacuo to provide compound 10 (1.1 g, crude) as a yellow solid. [M+H] + =628.9

[0267] 4-(3-((R)-3-(4-(((9H-Fluoren-9-yl)methoxy)carbonyl)piperazin-1-yl)-1-((S)-1-(4-(acryloyloxy)-3,3-dimethyl-2-oxobutanoyl)piperidine-2-carbonyloxy)propyl)phenylamino)-4-oxobutanoic acid (Raa3). To a solution of 10 (1.1 g, 1.74 mmol) in DMF (4 mL) was added NaCO (369 mg, 3.48 mmol) followed by Fmoc Chloride (450 mg, 1.74 mmol) at rt. The resulting reaction mixture was stirred at rt for 30 min. The reaction was quenched with HO (10 mL) and extracted with EA (30 mL × 3). The organic extract was dried over Na2SO4 and concentrated in vacuo to give the crude product, which was further purified by column (SiO2, methanol / DCM = 0-5% as eluent) to afford Raa3 (680 mg, 46%) as a white solid. FKBD Example 6 4-(3-((R)-4-((9H-fluoren-9-yl)methoxy)carbonyl)piperazin-1-yl)-1-((S)-1-(4-(acryloyloxy)-3,3-dimethyl-2-oxobutanoyl)piperidine-2-carbonyloxy)butyl)phenylamino)-4-oxobutanoic acid (Raa4) [ka] Scheme 9. Synthesis of Raa4 FKBD moiety

[0268] tert-Butyl 4-(4-(3-nitrophenyl)-4-oxobutyl)piperazine-1-carboxylate (3). To a solution of 1 (10.5 g, 38.6 mmol) and 2 (7.2 g, 38.6 mmol) in CHCN (100 mL) was added KCO (10.7 g, 77.2 mmol) at rt, and the resulting reaction mixture was heated to reflux for 2 h. The reaction mixture was then filtered and concentrated in vacuo to give the crude product, which was further purified by column chromatography (SiO, methanol / DCM = 0-5% as eluent) to give compound 3 (8.3 g, 57%) as a yellow solid. [M+H] + =378.0

[0269] tert-Butyl 4-(4-(3-aminophenyl)-4-oxobutyl)piperazine-1-carboxylate (4). To a solution of 3 (8.3 g, 22 mmol) in THF (60 mL) was added 10% Pd / C (wet, 1.66 g) at rt. The resulting reaction mixture was hydrogenated with H2(g) at rt for 10 h. The reaction mixture was then filtered and concentrated in vacuo to provide crude compound 4 (7.4 g, crude) as a yellow solid, which was used directly for the next step. [M+H] + =348.3.

[0270] tert-Butyl 4-(3-(4-morpholinobutanoyl)phenylamino)-4-oxobutanoate (6). To a solution of 4 (7.4 g, 21.3 mmol) and 4-tert-butoxy-4-oxobutanoic acid 5 (4.82 g, 27.6 mmol) in DMF (15 mL) was added DIPEA (5.5 g, 42.6 mmol) followed by HATU (10.5 g, 27.69 mmol) at rt. The resulting reaction mixture was stirred at rt for 2 h. The reaction was quenched with HO (50 mL) and extracted with EA (100 mL × 3). The organic extract was dried over NaSO and concentrated in vacuo to give the crude product, which was further purified by column chromatography (SiO, methanol / DCM = 0-5% as eluent) to give compound 6 (8.5 g, 79%) as a yellow solid. [M+H] + =504.0

[0271] (R)-tert-Butyl 4-(4-(3-(4-tert-butoxy-4-oxobutanamido)phenyl)-4-hydroxybutyl)piperazine-1-carboxylate (7). To a solution of ketone 6 (4.5 g, 8.9 mmol) in anhydrous THF (20 mL) was added (+)DIPChloride (35.6 mmol) in heptane (1.7 M, 21 mL) at −20 °C. The resulting reaction mixture was stirred at −20 °C until complete conversion of 6 and then quenched by the addition of 2,2′-(ethane-1,2-diylbis(oxy))diethanamine (5.9 g, 40.0 mmol) to form an insoluble complex. After stirring at rt for another 30 min, the suspension was filtered through a pad of Celite and concentrated in vacuo to give the crude product, which was further purified by column (SiO, methanol / EA=0-5% as eluent) to give compound 7 (2.5 g, 55%) as an off-white solid. [M+H] + =506.0

[0272] tert-Butyl 4-((R)-4-((S)-1-(4-(acryloyloxy)-3,3-dimethyl-2-oxobutanoyl)piperidine-2-carbonyloxy)-4-(3-(4-tert-butoxy-4-oxobutanamido)phenyl)butyl)piperazine-1-carboxylate (9). A solution of 7 (2.3 g, 4.5 mmol) and 8 (1.68 g, 5.4 mmol) in 30 mL of anhydrous DCM was cooled to −20° C., followed by the addition of a solution of DCC (1.11 g, 5.4 mmol) in 5 mL of anhydrous DCM, followed by the addition of a solution of 4-(dimethylamino)pyridine (DMAP, 55 mg, 0.615 mmol) in 1 mL of anhydrous DCM under an argon atmosphere. The resulting white suspension was stirred at −20° C. for 2 h. The reaction mixture was then filtered, and the filtrate was dried over NaSO and concentrated under reduced pressure to give the crude product, which was further purified by column (SiO, methanol / DCM=0-5% as eluent) to give compound 9 (2.9 g, 80%) as a white solid. [M+H] + =799.5

[0273] 4-(3-((R)-1-((S)-1-(4-(acryloyloxy)-3,3-dimethyl-2-oxobutanoyl)piperidine-2-carbonyloxy)-4-(piperazin-1-yl)butyl)phenylamino)-4-oxobutanoic acid (10). To a solution of 9 (2.9 g, 3.6 mmol) in DCM (10 mL) was added TFA (3 mL) at rt. The resulting mixture was stirred at rt for 4 h. The reaction mixture was directly loaded onto a silica gel flash column (CHOH / DCM = 0-5 as eluent) to provide compound 10 (2.6 g, crude) as a yellow solid, which was used directly for the next step. [M+H] + =643.4

[0274] 4-(3-((R)-4-(((9H-Fluoren-9-yl)methoxy)carbonyl)piperazin-1-yl)-1-((S)-1-(4-(acryloyloxy)-3,3-dimethyl-2-oxobutanoyl)piperidine-2-carbonyloxy)butyl)phenylamino)-4-oxobutanoic acid (Raa4). To a solution of 10 (1.2 g, 1.62 mmol) in DMF (2 mL) was added NaCO (343 mg, 3.24 mmol) followed by FmocChloride (419 mg, 1.62 mmol) at rt. The resulting reaction mixture was stirred for 30 min at rt. The reaction was quenched with HO (10 mL) and extracted with EA (30 mL × 3). The organic extract was dried over Na2SO4 and concentrated in vacuo to give the crude product, which was further purified by column (SiO2, methanol / DCM=0-5% as eluent) to afford Raa4 (570 mg, 40%) as a white solid. FKBD Example 7 4-(5-((R)-1-((S)-1-(4-(acryloyloxy)-3,3-dimethyl-2-oxobutanoyl)piperidine-2-carbonyloxy)-3-(3,4-dimethoxyphenyl)propyl)pyridin-3-ylamino)-4-oxobutanoic acid (Raa5) [ka] Scheme 10. Synthesis of Raa5 FKBD moiety

[0275] 1-(5-aminopyridin-3-yl)ethanone (2). To a solution of 1 (13 g, 78.3 mmol) in THF (100 mL) was added 10% Pd / C (wet, 8.0 g) at rt. The resulting reaction mixture was stirred under H2 (g) for 10 h at rt. The reaction mixture was then filtered and concentrated in vacuo to give the crude product, which was further purified by column chromatography (SiO2, methanol / DCM = 0-5% as eluent) to give compound 2 (10 g, 94%) as a yellow solid. [M+H] + =137.0

[0276] (E)-1-(5-aminopyridin-3-yl)-3-(3,4-dimethoxyphenyl)prop-2-en-1-one (4). To a solution of 2 (6.5 g, 47.8 mmol) and 3 (7.9 g, 47.8 mmol) in CHOH (60 mL) was added LiOH. HO (2 g, 47.8 mmol) at 0 °C. The resulting reaction mixture was stirred at rt for 3 h. The solvent was removed in vacuo, and the residue was diluted with DCM and HO. The organic extract was dried over NaSO and concentrated in vacuo to give the crude product, which was further purified by column chromatography (SiO, methanol / DCM = 0-5% as eluent) to give compound 4 (1.8 g, 13%) as a yellow solid. [M+H] + =285.0

[0277] (E)-tert-Butyl 4-(5-(3-(3,4-dimethoxyphenyl)acryloyl)pyridin-3-ylamino)-4-oxobutanoate (6). To a solution of 4 (1.8 g, 6.3 mmol) and 4-tert-butoxy-4-oxobutanoic acid 5 (1.1 g, 6.3 mmol) in DCM (35 mL) was added EtN (12.7 g, 12.6 mmol) followed by T3P (50% in EtOAc, 8.0 g, 12.6 mmol). The resulting reaction mixture was stirred at rt for 1 h. The reaction was quenched with HO (20 mL) and extracted with DCM (40 mL × 2). The organic extract was dried over Na2SO4 and concentrated in vacuo to give the crude product, which was further purified by column (SiO2, methanol / DCM=0-5% as eluent) to give compound 6 (1.88 g, 68%) as a yellow solid. [M+H] + =440.9

[0278] tert-Butyl 4-(5-(3-(3,4-dimethoxyphenyl)propanoyl)pyridin-3-ylamino)-4-oxobutanoate (7). To a solution of 6 (1.88 g, 4.27 mmol) in THF (50 mL) and methanol (5 mL) was added 10% Pd / C (wet, 380 mg) at RT. The resulting reaction mixture was hydrogenated with H2(g) at RT for 4 h. The reaction mixture was then filtered and concentrated in vacuo to give the crude product, which was further purified by column (SiO2, methanol / DCM = 0-5% as eluent) to afford compound 7 (1.34 g, 71%) as a brown solid. [M+H] + =442.9

[0279] (R)-tert-Butyl 4-(5-(3-(3,4-dimethoxyphenyl)-1-hydroxypropyl)pyridin-3-ylamino)-4-oxobutanoate (8). To a solution of ketone 7 (1.34 g, 3.0 mmol) in anhydrous THF (20 mL) was added (+)DIPChloride (12.0 mmol) in heptane (1.7 M, 7.05 mL) at −20° C. The resulting reaction mixture was stirred at −20° C. until complete conversion of 7 and then quenched by forming an insoluble complex with 2,2′-(ethane-1,2-diylbis(oxy))diethanamine (2.0 g, 13.5 mmol). After stirring at rt for another 30 min, the suspension was filtered through a pad of Celite and concentrated in vacuo to give the crude product, which was further purified by column (SiO2, CH3OH / EA = 0-5% as eluent) to give compound 8 (0.99 g, 74%) as a white solid. [M+H] + =445.0

[0280] (S)-((R)-1-(5-(4-tert-butoxy-4-oxobutanamido)pyridin-3-yl)-3-(3,4-dimethoxyphenyl)propyl)1-(4-(acryloyloxy)-3,3-dimethyl-2-oxobutanoyl)piperidine-2-carboxylate (10). A solution of 8 (990 mg, 2.22 mmol) and 9 (827 mg, 2.66 mmol) in anhydrous DCM (20 mL) was cooled to −20° C., followed by the addition of a solution of DCC (548 mg, 2.66 mmol) in anhydrous DCM (2 mL), followed by the addition of a solution of 4-(dimethylamino)pyridine (DMAP, 27 mg, 0.22 mmol) in anhydrous DCM (1 mL) under an argon atmosphere. The resulting white suspension was stirred at −20° C. for 2 h. The reaction mixture was then filtered, and the filtrate was dried over NaSO and concentrated under reduced pressure to give the crude product, which was further purified by column (SiO, CHH / DCM=0-5% as eluent) to give compound 10 (1.3 g, 79%) as a white solid. [M+H] + =738.0

[0281] 4-(5-((R)-1-((S)-1-(4-(acryloyloxy)-3,3-dimethyl-2-oxobutanoyl)piperidine-2-carbonyloxy)-3-(3,4-dimethoxyphenyl)propyl)pyridin-3-ylamino)-4-oxobutanoic acid (Raa5). To a solution of 10 (1.3 g, 1.76 mmol) in DCM (10 mL) was added TFA (5 mL) at rt. The resulting mixture was stirred at rt for 2 h. The reaction mixture was directly charged onto a silica gel flash column (CHOH / DCM = 0-5% as eluent) to afford Raa5 (960 mg, 80%) as a white solid. FKBD Example 8 4-(6-((R)-1-((S)-1-(4-(acryloyloxy)-3,3-dimethyl-2-oxobutanoyl)piperidine-2-carbonyloxy)-3-(3,4-dimethoxyphenyl)propyl)pyridin-2-ylamino)-4-oxobutanoic acid (Raa6) [ka] Scheme 11. Synthesis of Raa6 FKBD moiety

[0282] (E)-1-(6-aminopyridin-2-yl)-3-(3,4-dimethoxyphenyl)prop-2-en-1-one (3). To a solution of 1 (3.75 g, 27.57 mmol) and 2 (4.58 g, 27.57 mmol) in CHOH (40 mL) was added LiOH. HO (1.74 g, 41.35 mmol) at rt. The resulting reaction mixture was stirred at rt for 3 h. The solvent was removed in vacuo, and the residue was diluted with DCM and HO. The organic extract was dried over NaSO and concentrated in vacuo to give the crude product, which was further purified by column chromatography (SiO, methanol / DCM = 0-5% as eluent) to give compound 3 (3.2 g, 41%) as a yellow solid. [M+H] + =285.0

[0283] (E)-tert-Butyl 4-(6-(3-(3,4-dimethoxyphenyl)acryloyl)pyridin-2-ylamino)-4-oxobutanoate (5). To a solution of 3 (3.2 g, 11.26 mmol) and 4-tert-butoxy-4-oxobutanoic acid 4 (2.35 g, 13.5 mmol) in pyridine (10 mL) was added POCl (2.58 g, 16.89 mmol) at 0 °C. The resulting reaction mixture was stirred at 0 °C for 15 min. The reaction was quenched with HO (20 mL) and extracted with EA (30 mL × 3). The organic extract was dried over NaSO and concentrated in vacuo to give the crude product, which was further purified by column chromatography (SiO, methanol / DCM = 0-5% as eluent) to give compound 5 (2.45 g, 49%) as a yellow solid. [M+H] + =440.9

[0284] tert-Butyl 4-(6-(3-(3,4-dimethoxyphenyl)propanoyl)pyridin-2-ylamino)-4-oxobutanoate (6). To a solution of 5 (2.45 g, 5.56 mmol) in THF (30 mL) was added 10% Pd / C (wet, 500 mg) at RT. The resulting reaction mixture was hydrogenated with H2(g) at RT for 4 h. The reaction mixture was then filtered and concentrated in vacuo to give the crude product, which was further purified by column chromatography (SiO2, methanol / DCM = 0-5% as eluent) to provide compound 6 (1.5 g, 61%) as a yellow solid. [M+H] + =443.3

[0285] (R)-tert-Butyl 4-(6-(3-(3,4-dimethoxyphenyl)-1-hydroxypropyl)pyridin-2-ylamino)-4-oxobutanoate (7). To a solution of ketone 6 (1.4 g, 3.16 mmol) in anhydrous DCM (20 mL) was added (+)DIPChloride (12.64 mmol) in heptane (1.7 M, 7.5 mL) at −20° C. The resulting reaction mixture was stirred at −20° C. until complete conversion of 7, and then quenched by the formation of an insoluble complex with 2,2′-(ethane-1,2-diylbis(oxy))diethanolamine (2.1 g, 14.22 mmol). After stirring at rt for another 30 min, the suspension was filtered through a pad of Celite and concentrated in vacuo to give the crude product, which was further purified by column (SiO, CHOH / EA=0-5% as eluent) to give compound 7 (1.0 g, 71%) as a white solid. [M+H] + =445.3

[0286] (S)-((R)-1-(6-(4-tert-butoxy-4-oxobutanamido)pyridin-2-yl)-3-(3,4-dimethoxyphenyl)propyl)1-(4-(acryloyloxy)-3,3-dimethyl-2-oxobutanoyl)piperidine-2-carboxylate (9). A solution of 7 (1.0 g, 2.24 mmol) and 8 (836 mg, 2.69 mmol) in 20 mL of anhydrous DCM was cooled to −20° C., followed by the addition of a solution of DCC (554 mg, 2.69 mmol) in 2 mL of anhydrous DCM, followed by the addition of a solution of 4-(dimethylamino)pyridine (DMAP, 27 mg, 0.22 mmol) in 1 mL of anhydrous DCM under an argon atmosphere. The resulting white suspension was stirred at −20° C. for 2 h. The reaction mixture was then filtered, and the filtrate was dried over NaSO and concentrated under reduced pressure to give the crude product, which was further purified by column (SiO, CHOH / DCM=0-5% as eluent) to give compound 9 (0.38 g, 23%) as a white solid. [M+H] + =738.4

[0287] 4-(6-((R)-1-((S)-1-(4-(acryloyloxy)-3,3-dimethyl-2-oxobutanoyl)piperidine-2-carbonyloxy)-3-(3,4-dimethoxyphenyl)propyl)pyridin-2-ylamino)-4-oxobutanoic acid (Raa6). To a solution of 9 (0.38 g, 1.76 mmol) in DCM (5 mL) was added TFA (2 mL) at rt. The resulting mixture was stirred at rt for 2 h. The reaction mixture was directly charged onto a silica gel flash column (CHOH / DCM = 0-5% as eluent) to give Raa6 (310 mg, 89%) as a white solid. FKBD Example 9 4-((6-((R)-((S)-1-(4-(acryloyloxy)-3,3-dimethyl-2-oxobutanoyl)piperidine-2-carbonyl)oxy)-3-(3,4-dimethoxyphenyl)propyl)pyrazin-2-yl)amino)-4-oxobutanoic acid (Raa7) [ka] Scheme 12. Synthesis of Raa7 FKBD Moiety

[0288] 6-(1-Butoxyvinyl)pyrazin-2-amine (3). To a solution of 1 (16 g, 124 mmol) in ethylene glycol (150 mL) was added Pd(AcO) (0.8 g, 3.7 mmol) and DPPF (4.12 g, 7.4 mmol) at rt. The mixture was degassed with Ar and then 2 and EtN were added sequentially. The reaction mixture was heated to reflux and reacted for 1.5 h. The resulting mixture was poured into water (300 ml) and extracted with DCM (100 ml * 3). The organic phases were combined and washed with brine (100 ml * 3). Filtration and concentration gave 3 (12 g, 50%) as a white solid. [M+H] + =194

[0289] 1-(6-aminopyrazin-2-yl)ethan-1-one (4). To a solution of 3 (12 g, 62 mmol) in DCM (50 ml) was added 5% HCl (20 ml). The reaction mixture was stirred at rt for 0.5 h. The resulting mixture was poured into water (200 ml) and the pH was adjusted to 8-9 with K2CO3 (aq). Extraction with DCM (50 ml * 6) was performed, and the organic phases were combined and concentrated to obtain the crude product. Purification by silica gel chromatography (PE / EA = 20-30% as eluent) gave product 4 (2.9 g, 34%) as a yellow solid. [M+H] + =138

[0290] (E)-1-(5-aminopyridin-2-yl)-3-(3,4-dimethoxyphenyl)prop-2-en-1-one (6). To a solution of 4 (2.9 g, 21 mmol) in MeOH (20 ml), LiOH (1.74 g, 42 mol) and 5 (3.43 g, 21 mmol) were added. The reaction mixture was stirred at 40° C. for 1 h. The resulting mixture was poured into water (200 ml), filtered until no precipitate remained, and the solid cake was washed with water and then with a small amount of MeOH. After drying, the product 6 (3.8 g, 64.5%) was obtained as a yellow solid. [M+H] + =286

[0291] tert-Butyl (E)-4-(6-(3-(3,4-dimethoxyphenyl)acryloyl)pyrazin-2-yl)amino-4-oxobutanoate (8). To a solution of 8 (3.8 g, 133 mmol) and 7 (4.64 g, 266 mmol) in pyridine (100 ml) was added POCl (6.12 g, 400 mmol) at 0 °C. The reaction mixture was stirred at 0 °C for 30 min. The resulting mixture was poured into water (300 ml), extracted with DCM (100 ml * 3), the organic phases were combined, and washed with brine (100 ml * 5). It was dried over Na SO , filtered, and concentrated to obtain the crude product. Purification by silica gel chromatography (MeOH / DCM = 1-2% as eluent) gave the product 8 (5 g, 68%) as a yellow solid. [M+H] + =442

[0292] tert-Butyl 4-((6-(3-(3,4-dimethoxyphenyl)propanoyl)pyrazin-2-yl)amino)-4-oxobutanoate (9). To a solution of 8 (5.0 g, 113 mmol) in THF was added Pd / C (500 mg, 10%), and the reaction mixture was degassed with H2*5 and stirred at rt for 4 h. Filtration and concentration of the filtrate gave the crude product. Purification by silica gel chromatography (MeOH / DCM = 1-2% as eluent) gave product 9 (2.0 g, 40%) as a yellow solid. [M+H] + =444

[0293] tert-Butyl (R) 4-((6-(3,4-dimethoxyphenyl)-1-hydroxyphenyl)pyrazin-2-yl)amino)-4-oxobutanoate (11). To a solution of 9 (2.0 g, 45 mmol) in DCM (50 ml) was added DIPCl (14.5 g, 450 mmol) at −20° C. and degassed with Ar. The reaction mixture was stirred at −20° C. for 5 h. It was quenched with 10 (6.75 g, 455 mmol). The product mixture was directly concentrated, and the brown residue was purified by silica gel chromatography (MeOH / DCM=2-5% as eluent) to give the product 11 (1.0 g, 50%) as a yellow solid. [M+H] + =446

[0294] (R)-1-(6-(4-tert-butoxy)-4-oxobutanamido)pyrazin-2-yl)-3-(3,4-dimethoxyphenyl(S)-1(4-(acryloyloxy)-3,3-dimethyl-2-oxobutanoyl)piperidine-2-carboxylate (13). To a solution of 11 (1.0 g, 22 mmol) in DCM (30 ml) was added 12 (1.05 g, 34 mmol) at −20° C. and degassed with Ar, followed by DCM. CC (0.7 g, 34 mmol) and DMAP (0.03 g, 2.2 mmol) were injected sequentially. The reaction mixture was stirred at -20 °C for 1 h. The solid cake was filtered and washed with DCM (20 ml), and the filtrates were combined and evaporated to obtain the crude product. Purification by silica gel chromatography (MeOH / DCM = 1 to 2% as eluent) gave product 13 (1.8 g, 85%) as a yellow solid. [M+H] + =739

[0295] 4-(6-((R)-(((S)-1-(4-(acryloyloxy)-3,3-dimethyl-2-oxobutanoyl)piperidine-2-carbonyl)oxy)-3-(3,4-dimethoxyphenyl)propyl)pyrazin-2-yl)amino)-4-oxobutanoic acid (Raa7). To a solution of 13 (1.8 g, 24 mmol) in DCM (20 ml) was added TFA (20 ml). The reaction mixture was stirred at rt for 2 h. The product mixture was directly concentrated, and the yellow residue was purified by silica gel chromatography (MeOH / DCM = 1-2% as eluent) to give the product Raa7 (500 mg, 30%) as a pale yellow solid. FKBD Example 10 4-((3-((R)-1-(((S)-4-(4-(acryloyloxy)-3,3-dimethyl-2-oxobutanoyl)morpholine-3-carbonyl)oxy)-3-(3,4-dimethoxyphenyl)propyl)phenyl)amino)-4-oxobutanoic acid (Raa8) [ka] Scheme 13. Synthesis of Raa8 FKBD moiety

[0296] (E)-3-(3,4-Dimethoxyphenyl)-1-(3-nitrophenyl)prop-2-en-1-one (3). To a solution of 3,4-dimethoxybenzaldehyde 1 (60 g, 360 mmol) and 1-(3-nitrophenyl)ethan-1-one 2 (59.6 g, 360 mmol) in MeOH (1100 mL) was added NaOH (15 g) at 0 °C. The resulting solution was stirred at rt for 10 h. The precipitate was collected to give compound 3 as a yellow solid (97 g, 86%). [M+Na] + =336.1

[0297] 1-(3-aminophenyl)-3-(3,4-dimethoxyphenyl)propan-1-one (4). A solution of 3 (32 g, 110 mmol) and 10% Pd / C (10 g) in THF (120 mL) was hydrogenated with H2 at room temperature for 8 hours. The reaction mixture was then filtered and concentrated. The residue was purified by silica gel flash column chromatography (AcOEt / PE 1:3) to give compound 4 as a white solid (24 g, 76%). [M+H] + =286.2

[0298] tert-Butyl 4-((3-(3-(3,4-dimethoxyphenyl)propanoyl)phenyl)amino)-4-oxobutanoate (5). To a solution of 4 (12.0 g, 42 mmol) in DCM (30 mL), 4-tert-butoxy-4-oxobutanoic acid (8.8 g, 50 mmol), DIPEA (13.6 g, 105 mmol), and HATU (19.2 g, 50 mmol) were added. The mixture was stirred at rt for 16 h. The product was purified by silica gel flash column chromatography (AcOEt / PE 1:2) to give compound 5 as a white solid (16 g, 79%). [M+Na] + =464.0

[0299] tert-Butyl (R)-4-((3-(3-(3,4-dimethoxyphenyl)-1-hydroxypropyl)phenyl)amino)-4-oxobutanoate (6). A solution of ketone 5 (11.9 g, 26.9 mmol) in dry THF (120 mL) was treated with a solution of (+)-DIPChloride (135 mmol) in heptane (1.7 M, 79 mL) at −20° C. The resulting mixture was reacted at −20° C. until complete conversion of 5 and then quenched by forming an insoluble complex with 2,2′-(ethylenedioxy)diethylamine (20 g). After stirring for another 30 min at RT, the suspension was filtered through a pad of Celite and concentrated. The crude compound was purified by silica gel flash column chromatography (AcOEt / PE 3:1) to give compound 6 as a pale yellow oil (7.9 g, 66%, ee 97%). [M+Na] + =466.3

[0300] (R)-1-(3-(4-(tert-butoxy)-4-oxobutanamido)phenyl)-3-(3,4-dimethoxyphenyl)propyl (S)-4-(4-(acryloyloxy)-3,3-dimethyl-2-oxobutanoyl)morpholine-3-carboxylate (8). A solution of 6 (2.36 g, 5.32 mmol) and 7 (2 g, 6.38 mmol) in CHCl (10 mL) was cooled to −20 °C, followed by the addition of a solution of DCC (1.65 g, 7.98 mmol) in CHCl (5 mL), followed by the addition of a solution of 4-(dimethylamino)pyridine (DMAP, 65 mg, 0.53 mmol) in CHCl (2 mL) under an argon atmosphere. The resulting white suspension was allowed to stir at −20 °C for 2 h. The reaction mixture was then filtered, evaporated, and the crude compound was purified by silica gel flash column chromatography (AcOEt / PE 2:1) to give compound 8 as a pale yellow oil (2.5 g, 64%). [M+Na] + =761.4

[0301] 4-((R)-1-(((S)-4-(4-(acryloyloxy)-3,3-dimethyl-2-oxobutanoyl)morpholine-3-carbonyl)oxy)-3-(3,4-dimethoxyphenyl)propyl)phenyl)amino)-4-oxobutanoic acid (Raa8). A solution of 8 (2.5 g, 3.45 mmol) in CHCl (12 mL) was treated with a solution of 40% TFA in CHCl (12 mL) at 0 °C. The mixture was allowed to react at room temperature until complete conversion. The reaction mixture was directly charged onto a silica gel flash column (AcOEt / PE / AcOH 1:2:0.5%) to provide Raa8 (815 mg, 38%) as a pale yellow solid. FKBD Example 11 4-((3-((R)-1-(((S)-4-(((9H-fluoren-9-yl)methoxy)carbonyl)-1-(4-(acryloyloxy)-3,3-dimethyl-2-oxobutanoyl)piperazine-2-carbonyl)oxy)-3-(3,4-dimethoxyphenyl)propyl)phenyl)amino)-4-oxobutanoic acid (Raa9) [ka] Scheme 14. Synthesis of Raa9 FKBD Moiety

[0302] 1-((9H-Fluoren-9-yl)methyl)3-((R)-1-(3-(4-(tert-butoxy)-4-oxobutanamido)phenyl)-3-(3,4-dimethoxyphenyl)propyl)(S)-4-(4-(acryloyloxy)-3,3-dimethyl-2-oxobutanoyl)piperazine-1,3-dicarboxylate (3). A solution of 1 (1.35 g, 3.04 mmol) and 2 (1.95 g, 3.65 mmol) in CHCl (10 mL) was cooled to −20 °C before the addition of a solution of DCC (940 mg, 4.56 mmol) in CHCl (5 mL), followed by the addition of a solution of 4-(dimethylamino)pyridine (DMAP, 37 mg, 0.3 mmol) in CHCl (2 mL) under an argon atmosphere. The resulting white suspension was stirred at -20°C for 3 hours. The reaction mixture was then filtered, evaporated, and the crude compound was purified by silica gel flash column chromatography (DCM / MeOH 96:4) to give compound 3 as a white solid (3.0 g, Quant.). [M+Na] + =981.6

[0303] 4-((3)-((R)-1-(((S)-4-((9H-Fluoren-9-yl)methoxy)carbonyl)-1-(4-(acryloyloxy)-3,3-dimethyl-2-oxobutanoyl)piperazine-2-carbonyl)oxy)-3-(3,4-dimethoxyphenyl)propynyl)phenyl)amino)-4-oxobutanoic acid (Raa9). A solution of 3 (1.5 g, 1.56 mmol) in CHCl (12 mL) was treated with a solution of 40% TFA in CHCl (12 mL) at 0 °C. The mixture was allowed to react at room temperature until complete conversion. The reaction mixture was directly charged onto a silica gel flash column (AcOEt / PE / AcOH 1:2:0.5%) to provide Raa9 (1.4 g, 99%) as a white solid. FKBD Example 12 (S)-((R)-1-(3-(4-tert-butoxy-4-oxobutanamido)phenyl)-3-(3,4-dimethoxyphenyl)propyl)1-(4-(acryloyloxy)-3,3-dimethyl-2-oxobutanoyl)-4-methylpiperazine-2-carboxylate (Raa10) [ka] Scheme 15. Synthesis of Raa10 FKBD moiety

[0304] (S)-1-(9H-Fluoren-9-yl)methyl 3-((R)-1-(3-(4-tert-butoxy-4-oxobutanamido)phenyl)-3-(3,4-dimethoxyphenyl)propyl) 4-(4-(acryloyloxy)-3,3-dimethyl-2-oxobutanoyl)piperazine-1,3-dicarboxylate (2). To a solution of 1 (1.3 g, 1.35 mmol) in DMF (5 mL) was added TBAF (3.2 ml, 1.0 M, 3.18 mmol) at 0 °C. The resulting solution was heated at room temperature for 5 h. After this time, the reaction mixture was washed with NaHCO3 (aq. (aq), 50 ml*3) and NaCl (aq., 50 ml*3). The organic phase was concentrated. The reaction mixture was purified on silica in DCM / MEOH=50 / 1 to give 2 (800 mg, 80%) as a colorless oil. [M+H] + =738.4

[0305] (S)-((R)-1-(3-(4-tert-butoxy-4-oxobutanamido)phenyl)-3-(3,4-dimethoxyphenyl)propyl)1-(4-(acryloyloxy)-3,3-dimethyl-2-oxobutanoyl)-4-methylpiperazine-2-carboxylate (Raa10). A solution of 2 (800 mg, 1.08 mmol) in CHOOH (1.6 mL) was treated with aqueous formaldehyde (37% in water, 0.8 ml, 1.3 mmol) and allowed to stir at 50° C. for 1 h. After this time, the reaction mixture was purified with DCM / MeOH=100 / 1 to give 3 (400 mg, 50%) as a colorless oil. [M+H] + =751.9 FKBD Example 13 (S)-4-(3-((R)-1-((S)-1-(4-(acryloyloxy)-3,3-dimethyl-2-oxobutanoyl)piperidine-2-carbonyloxy)-3-(3,4-dimethoxyphenyl)propyl)phenylamino)-3-hydroxy-4-oxobutanoic acid (Raa11) [ka] Scheme 16. Synthesis of Raa11 FKBD moiety.

[0306] tert-Butyl 3-(3-(3,4-dimethoxyphenyl)propanoyl)phenylcarbamate (2). To a solution of 1-(3-aminophenyl)-3-(3,4-dimethoxyphenyl)propan-1-one 1 (8.5 g, 29.79 mmol) in 1,4-dioxane (85 mL) was added (Boc)2O (9.75 g, 44.68 mmol). The resulting solution was heated at 100° C. for 3 h. The solvent was evaporated, and the residue (10.3 g, crude) was used directly for the next step without purification. [M+Na] + =408

[0307] (R)-tert-Butyl 3-(3-(3,4-dimethoxyphenyl)-1-hydroxypropyl)phenylcarbamate (3). A solution of ketone 2 (10 g, crude) in dry THF (200 mL) at -20 °C was treated with a solution of (+)-DIPChloride (1.7 M, 76.2 mL) in heptane at -20 °C. The resulting mixture was reacted at -20 °C until complete conversion of 2, and then quenched by forming an insoluble complex with 2,2'-(ethylenedioxy)diethylamine (23.1 g). After stirring for another 30 min at RT, the suspension was filtered through a pad of Celite and concentrated. The crude compound was purified by silica gel flash column chromatography (AcOEt / PE 1:3) to give compound 3 as a pale yellow oil (8.3 g, 80%). [M+Na] + =410

[0308] (S)-((R)-1-(3-(tert-butoxycarbonylamino)phenyl)-3-(3,4-dimethoxyphenyl)propyl)1-(4-(acryloyloxy)-3,3-dimethyl-2-oxobutanoyl)piperidine-2-carboxylate (5). A solution of 3 (8.3 g, 21.42 mmol) and 4 (8 g, 25.7 mmol) in CHCl (100 mL) was cooled to −20 °C, followed by the addition of a solution of DCC (5.3 g, 25.7 mmol) in CHCl (5 mL), followed by the addition of a solution of 4-(dimethylamino)pyridine (DMAP, 318 mg, 2.6 mmol) in CHCl (2 mL) under an argon atmosphere. The resulting white suspension was allowed to stir at −20 °C for 2 h. The reaction mixture was then filtered, evaporated, and the crude compound was purified by silica gel flash column chromatography (AcOEt / PE 1:3) to give compound 5 as a pale yellow oil (12 g, 83%). [M+Na] + =703.3

[0309] (S)-((R)-1-(3-aminophenyl)-3-(3,4-dimethoxyphenyl)propyl)1-(4-(acryloyloxy)-3,3-dimethyl-2-oxobutanoyl)piperidine-2-carboxylate (6). To a solution of 5 (5 g, 7.34 mmol) in DCM (30 ml) was added TFA (6 ml). The mixture was stirred at 35° C. for 6 h. The solvent was evaporated and the residue (5.0 g, crude) was used directly for the next step without purification. [M+H] + =580.8

[0310] (S)-4-(3-((R)-1-((S)-1-(4-(acryloyloxy)-3,3-dimethyl-2-oxobutanoyl)piperidine-2-carbonyloxy)-3-(3,4-dimethoxyphenyl)propyl)phenylamino)-3-hydroxy-4-oxobutanoic acid (Raa11). To a solution of 6 (1.0 g, crude) in DCM (20 mL) was added 7 (400 mg, 3.4 mmol) and DMAP (25 mg, 0.2 mmol). The mixture was allowed to react at room temperature until complete conversion. The reaction mixture was directly charged onto a silica gel flash column (DCM / MeOH=10:1) to give Raa11 (450 mg, 38%) as a white solid. FKBD Example 14 (S)-4-(3-((R)-1-((S)-(4-(acryloyloxy)-3,3-dimethyl-2-oxobutanoyl)piperidine-2-carbonyloxy)-3-(3,4-dimethoxyphenyl)propyl)phenylamino)-2-hydroxy-4-oxobutanoic acid (Raa12) [ka] Scheme 17. Synthesis of Raa12 FKBD moiety

[0311] The synthesis of 6 is similar to that of Raa11.

[0312] (S)-((R)-1-(3-((S)-4-(allyloxy)-3-hydroxy-4-oxobutanamido)phenyl)-3-(3,4-dimethoxyphenyl)propyl)1-(4-(acryloyloxy)-3,3-dimethyl-2-oxobutanoyl)piperidine-2-carboxylate (8). To a solution of 6 (2 g, 3.44 mmol) in DMF (30 ml) was added 7 (1.2 g, 6.9 mmol), DIPEA (1.33 g, 0.32 mmol), and HATU (1.96 g, 5.16 mmol). The mixture was stirred at rt for 3 h and then diluted with EtOAc. The organic layer was washed with brine, dried over NaSO, and concentrated under reduced pressure. The residue was purified by silica gel column (DCM / MeOH 10:1) to give the product 8 as a yellow oil (800 mg, 32%). [M+H] + =737.

[0313] (S)-4-(3-((R)-1-((S)-1-(4-(acryloyloxy)-3,3-dimethyl-2-oxobutanoyl)piperidine-2-carbonyloxy)-3-(3,4-dimethoxyphenyl)propyl)phenylamino)-2-hydroxy-4-oxobutanoic acid (Raa12). To a solution of 8 (800 mg, 1.09 mmol) in THF (100 mL) was added N-methylaniline (232 mg, 2.17 mmol) and Pd(PPh3)4 (115 mg, 0.1 mmol). The mixture was reacted at room temperature under a N2 atmosphere until complete conversion. The reaction mixture was directly charged onto a silica gel flash column (DCM / MeOH 10:1) to afford Raa12 (120 mg, 16%) as a white solid. FKBD Example 15 (S)-3-(((9H-Fluoren-9-yl)methoxy)carbonylamino)-4-(3-((R)-1-((S)-1-(4-(acryloyloxy)-3,3-dimethyl-2-oxobutanoyl)piperidine-2-carbonyloxy)-3-(3,4-dimethoxyphenyl)propyl)phenylamino)-4-oxobutanoic acid (Raa13) [ka] Scheme 18. Synthesis of Raa13 FKBD Moiety

[0314] (S)-tert-Butyl 3-(((9H-fluoren-9-yl)methoxy)carbonylamino)-4-(3-(3-(3,4-dimethoxyphenyl)propanoyl)phenylamino)-4-oxobutanoate (3). A solution of 1 (4.0 g, 14.03 mmol), 2 (7.0 g, 16.8 mmol) in DCM (150 mL) was treated with DIPEA (8 ml, 42.1 mmol) and HATU (8.0 g, 21.1 mmol) at 0 °C and stirred at room temperature for 15 h. After this time, the reaction mixture was washed with HO and extracted with AcOEt (50 ml * 3). The organic phase was dried over NaSO and concentrated. The residue was purified by silica gel flash column chromatography (AcOEt / PE 1:10) to give compound 3 as a brown oil (9 g, 90%). [M+Na] + =700.9

[0315] (S)-tert-Butyl 3-(((9H-fluoren-9-yl)methoxy)carbonylamino)-4-(3-((R)-3-(3,4-dimethoxyphenyl)-1-hydroxypropyl)phenylamino)-4-oxobutanoate (4). A solution of ketone 3 (4.7 g, 6.9 mmol) in dry THF (130 mL) was treated with a solution of (+)-DIPChloride (27.7 mmol) in heptane (1.7 M, 16.3 mL) at −20° C. The resulting mixture was reacted at −20° C. until complete conversion of 3, then quenched with 2,2′-(ethylenedioxy)diethylamine (2.8 mL) to form an insoluble complex. After stirring for another 30 min at RT, the suspension was filtered through a pad of Celite and concentrated. The crude compound was purified by silica gel flash column chromatography (AcOEt / PE 1:10) to give compound 4 as a pale yellow oil (1.7 g, 40%). [M+Na] + =702.8

[0316] (S)-((R)-1-(3-((S)-2-(((9H-Fluoren-9-yl)methoxy)carbonylamino)-4-tert-butoxy-4 oxobutanamido)phenyl)-3-(3,4-dimethoxyphenyl)propyl)1-(4-(acryloyloxy)-3,3-dimethyl-2-oxobutanoyl)piperidine-2-carboxylate (6). A solution of 4 (1.7 g, 2.5 mmol) and 5 (1.2 g, 3.75 mmol) in CHCl (50 mL) was cooled to −20 °C before the addition of a solution of DCC (0.78 g, 3.75 mmol) in CHCl (5 mL), followed by the addition of a solution of 4-(dimethylamino)pyridine (DMAP, 30 mg, 0.25 mmol) in CHCl (2 mL) under an argon atmosphere. The resulting white suspension was stirred for 2 hours at −20° C. The reaction mixture was then filtered, evaporated, and the crude compound was purified by silica gel flash column chromatography (AcOEt / PE 1:5) to give compound 6 as a pale yellow oil (1.0 g, 50%).

[0317] (S)-3-(((9H-Fluoren-9-yl)methoxy)carbonylamino)-4-(3-((R)-1-((S)-1-(4-(acryloyloxy)-3,3-dimethyl-2-oxobutanoyl)piperidine-2-carbonyloxy)-3-(3,4-dimethoxyphenyl)propynyl)phenylamino)-4-oxobutanoic acid (Raa13). A solution of 6 (1.0 g, 1.02 mmol) in CHCl (10 mL) was treated with a solution of 40% TFA in CHCl (10 mL) at 0 °C. The mixture was allowed to react at room temperature until complete conversion. The reaction mixture was directly charged onto a silica gel flash column (DCM / MeOH = 50 / 1) to give Raa13 (401 mg, 42%) as a white solid. FKBD Example 16 (S)-2-(((9H-Fluoren-9-yl)methoxy)carbonylamino)-4-(3-((R)-1-((S)-1-(4-(acryloyloxy)-3,3-dimethyl-2-oxobutanoyl)piperidine-2-carbonyloxy)-3-(3,4-dimethoxyphenyl)propyl)phenylamino)-4-oxobutanoic acid (Raa14) [ka] Scheme 19. Synthesis of Raa14 FKBD moiety

[0318] (S)-tert-Butyl 2-(((9H-fluoren-9-yl)methoxy)carbonylamino)-4-(3-(3-(3,4-dimethoxyphenyl)propanoyl)phenylamino)-4-oxobutanoate (3). A solution of 1 (4.0 g, 14.03 mmol), 2 (7.0 g, 16.8 mmol) in DCM (150 mL) was treated with DIPEA (8 ml, 42.1 mmol) and HATU (8.0 g, 21.1 mmol) at 0 °C and allowed to stir at room temperature for 15 h. After this time, the reaction mixture was washed with HO and extracted with AcOEt (50 ml * 3). The organic phase was dried over NaSO and concentrated. The residue was purified by silica gel flash column chromatography (AcOEt / PE 1:10) to give compound 3 as a brown oil (9 g, 90%). [M+Na] + =700.9

[0319] (S)-tert-Butyl 2-(((9H-fluoren-9-yl)methoxy)carbonylamino)-4-(3-((S)-3-(3,4-dimethoxyphenyl)-1-hydroxypropyl)phenylamino)-4-oxobutanoate (4). A solution of ketone 3 (4.0 g, 5.9 mmol) in dry THF (80 mL) was treated with a solution of (+)-DIPChloride (23.6 mmol) in heptane (1.7 M, 14.0 mL) at −20° C. The resulting mixture was reacted at −20° C. until complete conversion of 3 and then quenched with 2,2′-(ethylenedioxy)diethylamine (2.8 mL) to form an insoluble complex. After stirring for another 30 min at RT, the suspension was filtered through a pad of Celite and concentrated. The crude compound was purified by silica gel flash column chromatography (AcOEt / PE 1:10) to give compound 4 as a pale yellow oil (2.0 g, 50%). [M+Na] + =702.8

[0320] (S)-((R)-1-(3-((S)-3-(((9H-Fluoren-9-yl)methoxy)carbonylamino)-4-tert-butoxy-4-oxobutanamido)phenyl)-3-(3,4-dimethoxyphenyl)propyl)1-(4-(acryloyloxy)-3,3-dimethyl-2-oxobutanoyl)piperidine-2-carboxylate (6). A solution of 4 (2.0 g, 2.9 mmol) and 5 (1.2 g, 3.82 mmol) in CHCl (50 mL) was cooled to −20 °C before the addition of a solution of DCC (0.91 g, 4.11 mmol) in CHCl (5 mL), followed by the addition of a solution of 4-(dimethylamino)pyridine (DMAP, 35 mg, 0.29 mmol) in CHCl (2 mL) under an argon atmosphere. The resulting white suspension was stirred for 2 hours at −20° C. The reaction mixture was then filtered and evaporated, and the crude compound was purified by silica gel flash column chromatography (AcOEt / PE 1:5) to give compound 6 as a pale yellow oil (1.0 g, 50%).

[0321] (S)-2-(((9H-Fluoren-9-yl)methoxy)carbonylamino)-4-(3-((R)-1-((S)-1-(4-(acryloyloxy)-3,3-dimethyl-2-oxobutanoyl)piperidine-2-carbonyloxy)-3-(3,4-dimethoxyphenyl)propynyl)phenylamino)-4-oxobutanoic acid (Raa14). A solution of 6 (1.0 g, 1.02 mmol) in CHCl (10 mL) was treated with a solution of 40% TFA in CHCl (10 mL) at 0 °C. The mixture was allowed to react at room temperature until complete conversion. The reaction mixture was directly charged onto a silica gel flash column (DCM / MeOH = 50 / 1) to give Raa14 (367 mg, 42%) as a white solid. FKBD Example 17 (2S,3S)-4-((3-((R)-1-(((S)-1-(4-(acryloyloxy)-3,3-dimethyl-2-oxobutanoyl)piperidine-2-carbonyl)oxy)-3-(3,4-dimethoxyphenyl)propyl)phenyl)amino)-2,3-dihydroxy-4-oxobutanoic acid (Raa15) [ka] [ka] Scheme 20. Synthesis of Raa15 FKBD Moiety

[0322] (3R,4S)-3,4-Dihydroxydihydrofuran-2,5-dione (2). To a solution of (2R,3S)-2,3-dihydroxysuccinic acid 1 (10 g, 66.6 mmol) in DCM (100 mL) was added 2,2,2-trifluoroacetic anhydride (27.9 g, 133.2 mmol) at 25 °C. The resulting solution was stirred at room temperature for 12 h. The mixture was concentrated in vacuo. The crude product was washed with petroleum ether (100 mL) to provide 2 (6 g, 68%) as a white solid.

[0323] (2S,3S)-4-((3-((R)-1-(((S)-1-(4-(acryloyloxy)-3,3-dimethyl-2-oxobutanoyl)piperidine-2-carbonyl)oxy)-3-(3,4-dimethoxyphenyl)propyl)phenyl)amino)-2,3-dihydroxy-4-oxobutanoic acid (Raa15). A mixture of 6 (2 g, 3.4 mmol), 2 (0.896 g, 6.8 mmol), and DMAP (80 mg, 0.68 mmol) in THF (60 mL) was stirred at 50° C. for 6 hours. The mixture was filtered and concentrated in vacuo. The resulting residue was purified by prep-HPLC (preparative-HPLC) to provide Raa15 (476 mg, 19%) as a white solid. FKBD Example 18 3-((((9H-Fluoren-9-yl)methoxy)amino)-4-((3-((R)-1-(((S)-1-(4-(acryloyloxy)-3,3-dimethyl-2-oxobutanoyl)piperidine-2-carbonyl)oxy)-3-(3,4-dimethoxyphenyl)propyl)phenyl)amino)-2-hydroxy-4-oxobutanoic acid (Raa16) [ka] Scheme 21. Synthesis of Raa16 FKBD moiety

[0324] 2-((((9H-Fluoren-9-yl)methoxy)carbonyl)amino)-3-hydroxysuccinic acid (2). To a solution of 1 (10 g, 67.1 mmol) in 1,4-dioxane (150 ml) was added 250 ml of 10% NaCO3(aq), and then FmocCl in 1,4-dioxane (150 ml) was added dropwise at 0 °C. The reaction mixture was stirred at 0 °C for 10 min and then warmed to rt and stirred for another 4 h. The resulting mixture was poured into water (500 ml) and extracted three times with EA (200 ml). The aqueous layer was adjusted to pH 2-3 with 2M HCl and then extracted three times with DCM (200 ml). The organic layers were combined, washed three times with brine (200 ml), dried over NaSO, filtered, and concentrated to give product 2 (22 g, 88%) as a white solid. [M+Na] + =394

[0325] 2-((((9H-Fluoren-9-yl)methoxy)carbonyl)amino)-2-(2,2-dimethyl-5-oxo-1,3-dioxolan-4-yl)acetic acid (4). To a solution of 2 (5 g, 13.5 mmol) in EA (50 ml) was added 3 (14 g, 135 mmol) and PTSA (0.46 g 2.7 mmol). The reaction mixture was refluxed for 16 h. The product mixture was concentrated directly, and the brown residue was purified by silica gel chromatography (EA / PE = 10-50% as eluent) to give 4 (3.8 g, 68.6%) as a white solid. [M+Na] + =434

[0326] (1R)-1-(3-(2-((((9H-Fluoren-9-yl)methoxy)carbonyl)amino)-2-(2,2-dimethyl-5-oxo-1,3-dioxolan-4-yl)acetamido)phenyl)-3-(3,4-dimethoxyphenyl(2S)-1-(4-(acryloyloxy)-3,3-dimethyl-2-oxobitanoyl)piperidine-2-carboxylate (6). To a solution of 4 (2.2 g, 5.4 mmol) in DMF (150 ml) was added HATU (3 g, 8 mmol) and DIEA (1.38 g, 10.8 mmol). 5 (2.6 g 4.5 mmol) was finally added. The reaction mixture was stirred at rt for 1 hour. The resulting mixture was poured into water (300 ml), extracted with DCM (100 ml * 3), the organic phases were combined and washed with brine (100 ml * 5). It was dried over Na2SO4, filtered and concentrated to obtain the crude product. It was purified by silica gel chromatography (methanol / DCM = 0-2% as eluent) to give compound 6 (3.7 g, 71%) as a white solid. [M+Na] + =996

[0327] 3-(((9H-Fluoren-9-yl)methoxy)amino)-4-((3-((R)-1-(((S)-1-(4-(acryloyloxy)-3,3-dimethyl-2-oxobutanoyl)piperidine-2-carbonyl)oxy)-3-(3,4-dimethoxyphenyl)propynyl)phenyl)amino)-2-hydroxy-4-oxobutanoic acid (Raa16). To a solution of 6 (3.7 g, 38 mmol) in THF / HO (10 ml / 10 ml) was added THF (40 ml). The reaction mixture was stirred at rt for 1 h. The product mixture was evaporated directly, and the residue was purified by silica gel chromatography (HCOOH / DCM = 0-5% as eluent) to give compound Raa16 (500 mg, 14%) as a pale yellow solid. FKBD Example 19 2-(3-((R)-1-(((S)-1-(4-(acryloyloxy)-3,3-dimethyl-2-oxobutanoyl)piperidine-2-carbonyl)oxy)-3-(3,4,5-trimethoxyphenyl)propyl)phenoxy)acetic acid (Rae1) [ka] Scheme 22. Synthesis of Rae1 FKBD Moiety

[0328] (E)-1-(3-Hydroxyphenyl)-3-(3,4,5-trimethoxyphenyl)prop-2-en-1-one (3). To a solution of 3,4,5-trimethoxybenzaldehyde 1 (5 g, 25.5 mmol) and 3'-hydroxyacetophenone 2 (3.47 g, 25.5 mmol) in EtOH (50 mL) was added a solution of 10% aqueous NaOH (41 mL, 4.1 g, 101.9 mmol) at 0 °C. The resulting solution was heated to 65 °C for 2 h. The solvent was evaporated, and the residue (5.5 g, crude) was used directly for the next step without purification. [M+H] + =314.9.

[0329] 3-(1-Hydroxy-3-(3,4,5-trimethoxyphenyl)propyl)phenol (4). A solution of 3 (5.5 g, crude) and 10% Pd / C (2 g) in THF (40 mL) was hydrogenated with H for 4 h at room temperature. The reaction mixture was then filtered and concentrated to a solid (5.96 g, crude). [M+H-H0] + =300.9

[0330] tert-Butyl 2-(3-(1-hydroxy-3-(3,4,5-trimethoxyphenyl)propyl)phenoxy)acetate (5). A solution of 4 (5.96 g, 18.8 mmol, crude) and K2CO3 (3.12 g, 22.6 mmol) in DMF (30 mL) was treated with tert-butyl bromoacetate (3.68 g, 18.8 mmol) and allowed to stir at room temperature for 5 h. After this time, the reaction mixture was poured into ice, causing a yellow solid to precipitate. The crude product was purified by prep-HPLC to give 5 (4.65 g, 42% (3 steps)) as a yellow solid. [M+Na] + =454.8

[0331] tert-Butyl 2-(3-(3-(3,4,5-trimethoxyphenyl)propanoyl)phenoxy)acetate (6). A solution of 5 (4.65 g, 10.75 mmol) in CHCl (110 mL) was treated with Dess-Martin periodinane (11.4 g, 26.88 mmol) and stirred at room temperature for 3 h before being quenched with a solution of 10% aqueous NaSO. The solution was extracted twice with CHCl. ​​The combined organic layers were washed with saturated NaHCO, brine, dried over NaSO, and concentrated under reduced pressure. The crude compound was purified by silica gel flash column chromatography (AcOEt / PE 1:3) to give compound 6 as a white solid (4.5 g, 97%). [M+Na] + =453.2

[0332] tert-Butyl (R)-2-(3-(1-hydroxy-3-(3,4,5-trimethoxyphenyl)propyl)phenoxy)acetate (7). A solution of ketone 6 (3.98 g, 9.25 mmol) in dry THF (40 mL) was treated with a solution of (+)-DIPChloride (18.5 mmol) in heptane (1.7 M, 10.88 mL) at −20 °C. The resulting mixture was reacted at −20 °C until complete conversion of 6 and then quenched with 2,2′-(ethylenedioxy)diethylamine (2.8 mL) to form an insoluble complex. After stirring at RT for another 30 min, the suspension was filtered through a pad of Celite and concentrated. The crude compound was purified by silica gel flash column chromatography (AcOEt / PE 1:3) to give compound 7 as a pale yellow oil (2.8 g, 70%, ee >99%). [M+Na] + =455.2

[0333] (R)-1-(3-(2-(tert-butoxy)-2-oxoethoxy)phenyl)-3-(3,4,5-trimethoxyphenyl)propyl (S)-1-(4-(acryloyloxy)-3,3-dimethyl-2-oxobutanoyl)piperidine-2-carboxylate (9). A solution of 7 (1.85 g, 4.3 mmol) and 8 (2 g, 6.4 mmol) in CHCl (15 mL) was cooled to −20 °C, followed by the addition of a solution of DCC (1.3 g, 6.4 mmol) in CHCl (5 mL), followed by the addition of a solution of 4-(dimethylamino)pyridine (DMAP, 52.3 mg, 0.43 mmol) in CHCl (2 mL) under an argon atmosphere. The resulting white suspension was allowed to stir at −20 °C for 2 h. The reaction mixture was then filtered, evaporated, and the crude compound was purified by silica gel flash column chromatography (AcOEt / PE 1:2) to give compound 9 as a pale yellow oil (2.5 g, 80%). [M+Na] + =748.4

[0334] 2-(3-((R)-1-(((S)-1-(4-(acryloyloxy)-3,3-dimethyl-2-oxobutanoyl)piperidine-2-carbonyl)oxy)-3-(3,4,5-trimethoxyphenyl)propyl)phenoxy)acetic acid (Rae1). A solution of 9 (2.5 g, 3.44 mmol) in CHCl (11.5 mL) was treated with a solution of 40% TFA in CHCl (11.5 mL) at 0 °C. The mixture was allowed to react at room temperature until complete conversion. The reaction mixture was directly charged onto a silica gel flash column (AcOEt / PE / AcOH 1:2:0.5%) to provide Rae1 (969 mg, 42%) as a white solid. 20 Examples of FKBD 2-(3-((R)-1-(((S)-1-(4-(acryloyloxy)-3,3-dimethyl-2-oxobutanoyl)piperidine-2-carbonyl)oxy)-3-(2,3,4-trimethoxyphenyl)propyl)phenoxy)acetic acid (Rae2) [ka] Scheme 23. Synthesis of Rae2 FKBD Moiety

[0335] (E)-1-(3-Hydroxyphenyl)-3-(2,3,4-trimethoxyphenyl)prop-2-en-1-one (3). To a solution of 2,3,4-trimethoxybenzaldehyde 1 (5 g, 25.5 mmol) and 3'-hydroxyacetophenone 2 (3.47 g, 25.5 mmol) in EtOH (30 mL) was added a solution of 10% aqueous NaOH (41 mL, 4.1 g, 101.9 mmol) at 0 °C. The resulting solution was heated to 65 °C for 3 h. The solvent was evaporated, and the residue was purified by silica gel flash column chromatography (AcOEt / PE 1:5) to give compound 3 as a yellow oil (6.6 g, 83%). [M+H] + =314.9

[0336] 3-(1-Hydroxy-3-(2,3,4-trimethoxyphenyl)propyl)phenol (4). A solution of 3 (6.6 g, 21 mmol) and 10% Pd / C (3 g) in THF (30 mL) was hydrogenated with H for 16 h at room temperature. The reaction mixture was then filtered and concentrated to a colorless oil (8 g, crude). [M+H-H0] + =301.0

[0337] tert-Butyl 2-(3-(1-hydroxy-3-(2,3,4-trimethoxyphenyl)propyl)phenoxy)acetate (5). A solution of 4 (8 g, 25 mmol, crude) and K2CO3 (4.19 g, 30 mmol) in DMF (30 mL) was treated with tert-butyl bromoacetate (5.92 g, 30 mmol) and allowed to stir at room temperature for 6 h. After this time, the reaction mixture was quenched with HO and extracted twice with EtOAc. The combined organic layers were washed with brine, dried over Na2SO4, and concentrated under reduced pressure. The crude compound was purified by silica gel flash column chromatography (AcOEt / PE 1:3) to give compound 6 as a colorless oil (8.2 g, 90% (2 steps)). [M+H-HO-tBu] + =358.8

[0338] tert-Butyl 2-(3-(2,3,4-trimethoxyphenyl)propanoyl)phenoxy)acetate (6). A solution of 5 (5.75 g, 13.29 mmol) in CHCl (30 mL) was treated with Dess-Martin periodinane (11.28 g, 26.59 mmol) and stirred at room temperature for 2 h before quenching with a solution of 10% aqueous NaSO. The solution was extracted twice with CHCl. ​​The combined organic layers were washed with sat. NaHCO, brine, dried over NaSO, and concentrated under reduced pressure. The crude compound was purified by silica gel flash column chromatography (AcOEt / PE 1:3) to give compound 6 as a yellow oil (5 g, 87%).

[0339] tert-Butyl (R)-2-(3-(1-hydroxy-3-(2,3,4-trimethoxyphenyl)propyl)phenoxy)acetate (7). A solution of ketone 6 (5 g, 11.61 mmol) in dry THF (50 mL) was treated with a solution of (+)-DIPChloride (23.23 mmol) in heptane (1.7 M, 13.66 mL) at −20 °C. The resulting mixture was reacted at −20 °C until complete conversion of 6 and then quenched with 2,2′-(ethylenedioxy)diethylamine (3.4 mL) to form an insoluble complex. After stirring for another 30 min at RT, the suspension was filtered through a pad of Celite and concentrated. The crude compound was purified by silica gel flash column chromatography (AcOEt / PE 1:3) to give compound 7 as a pale yellow oil (4 g, 80%, ee 83%). [M+H-H2O-tBu] + =358.9

[0340] (R)-1-(3-(2-(tert-butoxy)-2-oxoethoxy)phenyl)-3-(3,4,5-trimethoxyphenyl)propyl (S)-1-(4-(acryloyloxy)-3,3-dimethyl-2-oxobutanoyl)piperidine-2-carboxylate (9). A solution of 7 (2 g, 4.62 mmol) and 8 (2.16 g, 6.93 mmol) in CHCl (23 mL) was cooled to −20 °C, followed by the addition of a solution of DCC (1.43 g, 6.93 mmol) in CHCl (5 mL), followed by the addition of a solution of 4-(dimethylamino)pyridine (DMAP, 57 mg, 0.46 mmol) in CHCl (2 mL) under an argon atmosphere. The resulting white suspension was allowed to stir at −20 °C for 2 h. The reaction mixture was then filtered, evaporated, and the crude compound was purified by silica gel flash column chromatography (AcOEt / PE 1:2) to give compound 9 as a pale yellow oil (2.13 g, 64%). [M+Na] + =748.4

[0341] 2-(3-((R)-1-(((S)-1-(4-(acryloyloxy)-3,3-dimethyl-2-oxobutanoyl)piperidine-2-carbonyl)oxy)-3-(2,3,4-trimethoxyphenyl)propyl)phenoxy)acetic acid (Rae2). A solution of 9 (2.13 g, 2.93 mmol) in CHCl (12 mL) was treated with a solution of 40% TFA in CHCl (12 mL) at 0 °C. The mixture was allowed to react at room temperature until complete conversion. The reaction mixture was directly charged onto a silica gel flash column (AcOEt / PE / AcOH 1:2:0.5%) to provide Rae2 (508 mg, 25%) as a pale yellow solid. FKBD Example 21 2-(3-((R)-1-(((S)-1-(4-(acryloyloxy)-3,3-dimethyl-2-oxobutanoyl)piperidine-2-carbonyl)oxy)-3-(2,4,5-trimethoxyphenyl)propyl)phenoxy)acetic acid (Rae3) [ka] Scheme 24. Synthesis of Rae3 FKBD Moiety

[0342] (E)-1-(3-Hydroxyphenyl)-3-(2,4,5-trimethoxyphenyl)prop-2-en-1-one (3). To a solution of 2,4,5-trimethoxybenzaldehyde 1 (4.5 g, 22.96 mmol) and 3'-hydroxyacetophenone 2 (3.1 g, 22.96 mmol) in EtOH (50 mL) was added a solution of 10% aqueous KOH (15 mL, 5.1 g, 91.84 mmol) at 0 °C. The resulting solution was heated to 60 °C for 4 h. The solvent was evaporated, and the residue was purified by silica gel flash column chromatography (AcOEt / PE 1:3) to give compound 3 as a yellow oil (5.9 g, 82%). [M+H] + =315.1

[0343] tert-Butyl (E)-2-(3-(3-(2,4,5-trimethoxyphenyl)acryloyl)phenoxy)acetate (4). A solution of 3 (7.4 g, 23.6 mmol) and K2CO3 (3.9 g, 28.3 mmol) in DMF (200 mL) was treated with tert-butyl bromoacetate (5.5 g, 28.3 mmol) and allowed to stir at room temperature for 4 hours. After this time, the reaction mixture was quenched with HO and extracted twice with EtOAc. The combined organic layers were washed with brine, dried over Na2SO4, and concentrated under reduced pressure. The crude compound was purified by silica gel flash column chromatography (AcOEt / PE 1:3) to give compound 4 as a colorless oil (8 g, 80%). [M+H] + =429.3

[0344] tert-Butyl 2-(3-(3-(2,4,5-trimethoxyphenyl)propanoyl)phenoxy)acetate (5). A solution of 4 (8 g, 18.69 mmol) and 10% Pd / C (1 g) in THF (200 mL) was hydrogenated with H2 at room temperature for 8 h. The reaction mixture was then filtered and concentrated. The residue was purified by silica gel flash column chromatography (AcOEt / PE 1:3) to give compound 5 as a colorless oil (6 g, 75%). [M+Na] + =453.2

[0345] tert-Butyl (R)-2-(3-(1-hydroxy-3-(2,4,5-trimethoxyphenyl)propyl)phenoxy)acetate (6). A solution of ketone 5 (6 g, 13.95 mmol) in dry THF (60 mL) was treated with a solution of (+)-DIPChloride (41.86 mmol) in heptane (1.7 M, 24.6 mL) at −20° C. The resulting mixture was reacted at −20° C. until complete conversion of 5, and then quenched with 2,2′-(ethylenedioxy)diethylamine (5.9 mL) to form an insoluble complex. After stirring at RT for another 30 min, the suspension was filtered through a pad of Celite and concentrated. The crude compound was purified by silica gel flash column chromatography (AcOEt / PE 1:1) to give compound 6 as a pale yellow oil (5.5 g, 92%, ee>99%). [M+Na] + =455.2

[0346] (R)-1-(3-(2-(tert-butoxy)-2-oxoethoxy)phenyl)-3-(2,4,5-trimethoxyphenyl)propyl (S)-1-(4-(acryloyloxy)-3,3-dimethyl-2-oxobutanoyl)piperidine-2-carboxylate (8). A solution of 6 (1.85 g, 4.28 mmol) and 7 (2 g, 6.42 mmol) in CHCl (10 mL) was cooled to −20 °C, and then a solution of DCC (1.33 g, 6.42 mmol) in CHCl (5 mL) was added, followed by the addition of a solution of 4-(dimethylamino)pyridine (DMAP, 52 mg, 0.43 mmol) in CHCl (2 mL) under an argon atmosphere. The resulting white suspension was allowed to stir at −20 °C for 2 h. The reaction mixture was then filtered, evaporated, and the crude compound was purified by silica gel flash column chromatography (AcOEt / PE 1:1) to give compound 8 as a pale yellow oil (2.35 g, 76%). [M+Na] + =747.9

[0347] 2-(3-((R)-1-(((S)-1-(4-(acryloyloxy)-3,3-dimethyl-2-oxobutanoyl)piperidine-2-carbonyl)oxy)-3-(2,4,5-trimethoxyphenyl)propyl)phenoxy)acetic acid (Rae3). A solution of 8 (2.35 g, 3.24 mmol) in CHCl (12 mL) was treated with a solution of 40% TFA in CHCl (12 mL) at 0 °C. The mixture was allowed to react at room temperature until complete conversion. The reaction mixture was directly charged onto a silica gel flash column (AcOEt / PE / AcOH 1:2:0.5%) to give Rae3 (815 mg, 37%) as a pale yellow solid. FKBD Example 22 2-(3-((R)-1-(((S)-1-(4-(acryloyloxy)-3,3-dimethyl-2-oxobutanoyl)piperidine-2-carbonyl)oxy)-3-(2,3,5-trimethoxyphenyl)propyl)phenoxy)acetic acid (Rae4) [ka] Scheme 25. Synthesis of Rae4 FKBD moiety

[0348] (E)-1-(3-Hydroxyphenyl)-3-(2,3,5-trimethoxyphenyl)prop-2-en-1-one (3). To a solution of 2,3,5-trimethoxybenzaldehyde 1 (6 g, 30.6 mmol) and 1-(3-hydroxyphenyl)ethan-1-one 2 (4.2 g, 30.6 mmol) in EtOH (50 mL) was added a solution of 10% aqueous NaOH (50 mL, 122.4 mmol) at 0 °C. The resulting solution was stirred at room temperature for 12 h. The solution was adjusted to pH 4 by adding 4 M aqueous HCl dropwise at 0 °C, resulting in the formation of a large yellow solid. The mixture was then filtered, and the solid was washed with water (50 mL) to provide 3 (5.5 g, 57%) as a yellow solid. [M+H] + =315.2.

[0349] tert-Butyl (E)-2-(3-(3-(2,3,5-trimethoxyphenyl)acryloyl)phenoxy)acetate (4). A solution of 3 (5.5 g, 17.4 mmol) and K2CO3 (4.82 g, 34.9 mmol) in DMF (40 mL) was treated with tert-butyl bromoacetate (4.06 g, 20.9 mmol) and allowed to stir at room temperature for 12 h. After this time, the reaction mixture was poured into ice and a yellow solid precipitated. The mixture was filtered and the solid was washed with water (30 mL). The crude product was washed with petroleum ether (50 mL) to give 4 (7 g, 93%) as a yellow solid. [M+H] + =428.8

[0350] tert-Butyl 2-(3-(3-(2,3,5-trimethoxyphenyl)propanoyl)phenoxy)acetate (5). A solution of 4 (7 g, 11.68 mmol) and 10% Pd / C (1 g) in THF (100 mL) was hydrogenated with H2 at room temperature for 4 h. The reaction mixture was then filtered and concentrated. The crude product was purified by column chromatography on silica gel to give 5 (3.5 g, 50%) as a yellow oil. [M+Na] + =452.9.

[0351] tert-Butyl (R)-2-(3-(1-hydroxy-3-(2,3,5-trimethoxyphenyl)propyl)phenoxy)acetate (6). A solution of ketone 5 (3.5 g, 8.14 mmol) in dry THF (30 mL) was treated with (+)-DIPChloride (16.2 mmol) in heptane (1.7 M, 9.5 mL) at 20 °C. The resulting mixture was reacted at -20 °C until complete conversion of 6, and then quenched by forming an insoluble complex with 2,2'-(ethylenedioxy)diethylamine (2.4 g). After stirring for another 30 min at room temperature, the suspension was filtered through a pad of Celite and concentrated. The crude compound was purified by silica gel flash column chromatography (AcOEt / PE 1:4) to give compound 6 (2.2 g, 63%, ee 97% to racemic) as a pale yellow oil. [M+Na] + =454.9

[0352] (R)-1-(3-(2-(tert-butoxy)-2-oxoethoxy)phenyl)-3-(2,3,5-trimethoxyphenyl)propyl (S)-1-(4-(acryloyloxy)-3,3-dimethyl-2-oxobutanoyl)piperidine-2-carboxylate (8). A solution of 6 (2.2 g, 5.09 mmol) and 8 (1.89 g, 6.1 mmol) in CHCl (15 mL) was cooled to −20 °C, followed by the addition of a solution of DCC (1.36 g, 6.6 mmol) in CHCl (5 mL), followed by the addition of a solution of 4-(dimethylamino)pyridine (62 mg, 0.5 mmol) in CHCl (2 mL) under an argon atmosphere. The resulting white suspension was allowed to stir at −20 °C for 2 h. The reaction mixture was then filtered, evaporated, and the crude compound was purified by silica gel flash column chromatography (AcOEt / PE 1:2) to give compound 8 (1.8 g, 49%) as a pale yellow oil. [M+Na] + =748.4

[0353] 2-(3-((R)-1-(((S)-1-(4-(acryloyloxy)-3,3-dimethyl-2-oxobutanoyl)piperidine-2-carbonyl)oxy)-3-(2,3,5-trimethoxyphenyl)propyl)phenoxy)acetic acid (Rae4). A solution of 8 (1.8 g, 2.48 mmol) in CHCl (10 mL) was treated with a solution of 40% TFA in CHCl (10 mL) at 0 °C. The mixture was allowed to react at room temperature until complete conversion. The reaction mixture was directly charged onto a silica gel flash column (AcOEt / PE / AcOH 1:3:0.5%) to give Rae4 (652 mg, 39%) as a slightly yellow solid. FKBD Example 23 2-(3-((R)-1-(((S)-1-(4-(acryloyloxy)-3,3-dimethyl-2-oxobutanoyl)piperidine-2-carbonyl)oxy)-3-(2,3,6-trimethoxyphenyl)propyl)phenoxy)acetic acid (Rae5) [ka] Scheme 26. Synthesis of Rae5 FKBD Moiety

[0354] (E)-1-(3-Hydroxyphenyl)-3-(2,3,6-trimethoxyphenyl)prop-2-en-1-one (3). To a solution of 2,3,6-trimethoxybenzaldehyde 1 (5 g, 25.48 mmol) and 3'-hydroxyacetophenone 2 (3.47 g, 25.48 mmol) in EtOH (40 mL) was added a solution of 40% aqueous KOH (15 mL, 5.7 g, 101.92 mmol) at 0 °C. The resulting solution was reacted at room temperature for 4 h. The solvent was evaporated, and the residue was purified by silica gel flash column chromatography (AcOEt / PE 1:2) to give compound 3 as a yellow oil (4 g, 50%). [M+H] + =315.2

[0355] tert-Butyl (E)-2-(3-(3-(2,3,6-trimethoxyphenyl)acryloyl)phenoxy)acetate (4). A solution of 3 (3.5 g, 11.15 mmol) and K2CO3 (1.85 g, 13.37 mmol) in DMF (60 mL) was treated with tert-butyl bromoacetate (2.6 g, 13.37 mmol) and allowed to stir at room temperature for 4 hours. After this time, the reaction mixture was quenched with HO and extracted twice with EtOAc. The combined organic layers were washed with brine, dried over Na2SO4, and concentrated under reduced pressure. The crude compound was purified by silica gel flash column chromatography (AcOEt / PE 1:4) to give compound 4 as a yellow oil (4.7 g, 98%). [M+H] + =429.0

[0356] tert-Butyl 2-(3-(3-(2,3,6-trimethoxyphenyl)propanoyl)phenoxy)acetate (5). A solution of 4 (4.6 g, 10.75 mmol) and 10% Pd / C (0.5 g) in THF (70 mL) was hydrogenated with H2 at room temperature for 4 h. The reaction mixture was then filtered and concentrated. The residue was purified by silica gel flash column chromatography (AcOEt / PE 1:3) to give compound 5 as a colorless oil (2.9 g, 63%). [M+Na] + =453.3

[0357] tert-Butyl (R)-2-(3-(1-hydroxy-3-(2,3,6-trimethoxyphenyl)propyl)phenoxy)acetate (6). A solution of ketone 5 (2.9 g, 6.7 mmol) in dry THF (30 mL) was treated with a solution of (+)-DIPChloride (13.48 mmol) in heptane (1.7 M, 7.9 mL) at −20° C. The resulting mixture was reacted at −20° C. until complete conversion of 5, and then quenched with 2,2′-(ethylenedioxy)diethylamine (1.96 mL) to form an insoluble complex. After stirring at RT for another 30 min, the suspension was filtered through a pad of Celite and concentrated. The crude compound was purified by silica gel flash column chromatography (AcOEt / PE 1:5) to give compound 6 as a pale yellow oil (2.4 g, 83%, ee>99%). [M+Na] + =454.9

[0358] (R)-1-(3-(2-(tert-butoxy)-2-oxoethoxy)phenyl)-3-(2,3,6-trimethoxyphenyl)propyl (S)-1-(4-(acryloyloxy)-3,3-dimethyl-2-oxobutanoyl)piperidine 2-carboxylate (8). A solution of 6 (1.46 g, 3.45 mmol) and 7 (1.6 g, 5.17 mmol) in CHCl (18 mL) was cooled to −20 °C, followed by the addition of a solution of DCC (1.065 g, 5.17 mmol) in CHCl (5 mL), followed by the addition of a solution of 4-(dimethylamino)pyridine (DMAP, 43 mg, 0.35 mmol) in CHCl (2 mL) under an argon atmosphere. The resulting white suspension was allowed to stir at −20 °C for 2 h. The reaction mixture was then filtered, evaporated, and the crude compound was purified by silica gel flash column chromatography (AcOEt / PE 1:1) to give compound 8 as a pale yellow oil (1.7 g, 68%).

[0359] 2-(3-((R)-1-(((S)-1-(4-(acryloyloxy)-3,3-dimethyl-2-oxobutanoyl)piperidine-2-carbonyl)oxy)-3-(2,3,6-trimethoxyphenyl)propyl)phenoxy)acetic acid (Rae5). A solution of 8 (1.7 g, 2.34 mmol) in CHCl (12 mL) was treated with a solution of 40% TFA in CHCl (12 mL) at 0 °C. The mixture was allowed to react at room temperature until complete conversion. The reaction mixture was directly charged onto a silica gel flash column (AcOEt / PE / AcOH 1:2:0.5%) to give Rae5 (494 mg, 31%) as a pale yellow solid. FKBD Example 24 2-(3-((R)-1-(((S)-1-(4-(acryloyloxy)-3,3-dimethyl-2-oxobutanoyl)piperidine-2-carbonyl)oxy)-3-(2-hydroxy-3,4-dimethoxyphenyl)propyl)phenoxy)acetic acid (Rae9) [ka] Scheme 27. Synthesis of Rae9 FKBD Moiety

[0360] 2-(tert-Butoxy)-3,4-dimethoxybenzaldehyde (2). To a solution of 2-hydroxy-3,4-dimethoxybenzaldehyde 1 (2.77 g, 15.2 mmol) in anhydrous toluene (30 mL) was added 1,1-ditert-butoxy-N,N-dimethylmethanamine 2 (29.1 mL, 122 mmol) under Ar. The mixture was stirred at 80 °C for 6 h, and then the solvent was evaporated. The residue was purified by silica gel flash column chromatography (AcOEt / PE 1:1) to give compound 2 as a yellow solid (2.965 g, 82%). [M+Na] + =261.1

[0361] (E)-3-(2-(tert-butoxy)-3,4-dimethoxyphenyl)-1-(3-hydroxyphenyl)prop-2-en-1-one (4). To a solution of 2 (2.965 g, 12.4 mmol) and 3'-hydroxyacetophenone 3 (2.03 g, 14.9 mmol) in EtOH (50 mL) was added a solution of 40% aqueous KOH (6.98 g, 49.8 mmol) at 0 °C. The resulting solution was stirred at 60 °C for 4 h. The solution was poured into water and acidified to pH 4 with 1 M aqueous HCl, extracted twice with EtOAc. The combined organic layers were washed with brine, dried over Na SO , and concentrated under reduced pressure. The crude compound was purified by silica gel flash column chromatography (AcOEt / PE 1:1) to give compound 4 as a yellow oil (3.2 g, 72%). [M+Na] + =378.9

[0362] tert-Butyl (E)-2-(3-(3-(2-(tert-butoxy)-3,4-dimethoxyphenyl)acryloyl)phenoxy)acetate (5). A solution of 4 (3.2 g, 9 mmol) and K2CO3 (1.49 g, 10.8 mmol) in DMF (30 mL) was treated with tert-butyl bromoacetate (1.58 mL, 10.8 mmol) and allowed to stir at room temperature for 5 hours. After this time, the reaction mixture was quenched with HO and extracted twice with EtOAc. The combined organic layers were washed with brine, dried over Na2SO4, and concentrated under reduced pressure. The crude compound was purified by silica gel flash column chromatography (AcOEt / PE 1:4) to give compound 5 as a yellow oil (4 g, 95%). [M+Na] + =493.3

[0363] tert-Butyl 2-(3-(3-(2-(tert-butoxy)-3,4-dimethoxyphenyl)propanoyl)phenoxy)acetate (6). A solution of 5 (4 g, 8.5 mmol) and 10% Pd / C (0.8 g) in THF (50 mL) was hydrogenated with H2 at room temperature for 3 h. The reaction mixture was then filtered and concentrated. The residue was purified by silica gel flash column chromatography (AcOEt / PE 1:3) to give compound 6 as a colorless oil (2.878 g, 72%). [M+Na] + =495.3

[0364] tert-Butyl (R)-2-(3-(3-(3-(tert-butoxy)-4,5-dimethoxyphenyl)-1-hydroxypropyl)phenoxy)acetate (7). A solution of ketone 6 (2.878 g, 6.1 mmol) in dry THF (30 mL) was treated at −20° C. with a solution of (+)-DIPChloride (24.4 mmol) in heptane (1.7 M, 14.3 mL). The resulting mixture was reacted at −20° C. until complete conversion of 6 and then quenched with 2,2′-(ethylenedioxy)diethylamine (3.6 mL) to form an insoluble complex. After stirring for another 30 min at RT, the suspension was filtered through a pad of Celite and concentrated. The crude compound was purified by silica gel flash column chromatography (AcOEt / PE 1:3) to give compound 7 as a pale yellow oil (2 g, 70%, ee>99%). [M+Na] + =497.0

[0365] (R)-1-(3-(2-(tert-butoxy)-2-oxoethoxy)phenyl)-3-(2-(tert-butoxy)-3,4-dimethoxyphenyl)propyl (S)-1-(4-(acryloyloxy)-3,3-dimethyl-2-oxobutanoyl)piperidine-2-carboxylate (9). A solution of 7 (1.8 g, 3.793 mmol) and 8 (1.77 g, 5.69 mmol) in CHCl (13 mL) was cooled to −20 °C, followed by the addition of a solution of DCC (1.17 g, 5.69 mmol) in CHCl (5 mL), followed by the addition of a solution of 4-(dimethylamino)pyridine (DMAP, 46 mg, 0.379 mmol) in CHCl (2 mL) under an argon atmosphere. The resulting white suspension was allowed to stir at −20 °C for 2 h. The reaction mixture was then filtered, evaporated, and the crude compound was purified by silica gel flash column chromatography (AcOEt / PE 1:3) to give compound 9 as a pale yellow oil (2.5 g, 86%). [M+Na] + =790.4

[0366] 2-(3-((R)-1-(((S)-1-(4-(acryloyloxy)-3,3-dimethyl-2-oxobutanoyl)piperidine-2-carbonyl)oxy)-3-(2-hydroxy-3,4-dimethoxyphenyl)propyl)phenoxy)acetic acid (Rae9): A solution of 9 (2.5 g, 3.26 mmol) in CHCl (12 mL) was treated with a solution of 40% TFA in CHCl (12 mL) at 0 °C. The mixture was allowed to react at room temperature until complete conversion. The reaction mixture was directly charged onto a silica gel flash column (AcOEt / PE / AcOH 1:2:0.5%) to afford Rae9 (636 mg, 30%) as a pale yellow solid. FKBD Example 25 2-(3-((R)-1-(((S)-1-(4-(acryloyloxy)-3,3-dimethyl-2-oxobutanoyl)piperidine-2-carbonyl)oxy)-3-(3-hydroxy-4,5-dimethoxyphenyl)propyl)phenoxy)acetic acid (Rae10) [ka] Scheme 28. Synthesis of Rae10 FKBD Moiety

[0367] 3-(tert-Butoxy)-4,5-dimethoxybenzaldehyde (2). To a solution of 3-hydroxy-4,5-dimethoxybenzaldehyde 1 (2.77 g, 15.2 mmol) in anhydrous toluene (30 mL) was added 1,1-ditert-butoxy-N,N-dimethylmethanamine 2 (29.1 mL, 122 mmol) under Ar. The mixture was stirred at 80 °C for 6 h, and then the solvent was evaporated. The residue was purified by silica gel flash column chromatography (AcOEt / PE 1:1) to give compound 2 as a yellow solid (3.126 g, 86%). [M+H] + =239.0

[0368] (E)-3-(3-(tert-butoxy)-4,5-dimethoxyphenyl)-1-(3-hydroxyphenyl)prop-2-en-1-one (4). To a solution of 2 (3.126 g, 13 mmol) and 3'-hydroxyacetophenone 3 (2.14 g, 15.7 mmol) in EtOH (30 mL) was added a solution of 40% aqueous KOH (7.36 g, 52 mmol) at 0 °C. The resulting solution was stirred at 60 °C for 4 h. The solution was poured into water and acidified to pH 4 with 1 M aqueous HCl, extracted twice with EtOAc. The combined organic layers were washed with brine, dried over Na SO , and concentrated under reduced pressure. The crude compound was purified by silica gel flash column chromatography (AcOEt / PE 1:1) to give compound 4 as a yellow oil (2.489 g, 54%). [M+H] + =357.0

[0369] tert-Butyl (E)-2-(3-(3-(3-(tert-butoxy)-4,5-dimethoxyphenyl)acryloyl)phenoxy)acetate (5). A solution of 4 (2.489 g, 6.98 mmol) and K2CO3 (1.16 g, 8.38 mmol) in DMF (30 mL) was treated with tert-butyl bromoacetate (1.2 mL, 8.38 mmol) and allowed to stir at room temperature for 5 hours. After this time, the reaction mixture was quenched with HO and extracted twice with EtOAc. The combined organic layers were washed with brine, dried over Na2SO4, and concentrated under reduced pressure. The crude compound was purified by silica gel flash column chromatography (AcOEt / PE 1:4) to give compound 5 as a yellow oil (3.1 g, 95%). [M+H] + =471.0

[0370] tert-Butyl 2-(3-(3-(3-(tert-butoxy)-4,5-dimethoxyphenyl)propanoyl)phenoxy)acetate (6). A solution of 5 (3.1 g, 6.59 mmol) and 10% Pd / C (0.5 g) in THF (50 mL) was hydrogenated with H2 at room temperature for 3 h. The reaction mixture was then filtered and concentrated. The residue was purified by silica gel flash column chromatography (AcOEt / PE 1:3) to give compound 6 as a colorless oil (2.88 g, 93%). [M+Na] + =495.3

[0371] tert-Butyl (R)-2-(3-(3-(3-(tert-butoxy)-4,5-dimethoxyphenyl)-1-hydroxypropyl)phenoxy)acetate (7). A solution of ketone 6 (2.868 g, 6.07 mmol) in dry THF (30 mL) was treated at −20° C. with a solution of (+)-DIPChloride (12.1 mmol) in heptane (1.7 M, 7.1 mL). The resulting mixture was reacted at −20° C. until complete conversion of 6 and then quenched with 2,2′-(ethylenedioxy)diethylamine (3.6 mL) to form an insoluble complex. After stirring for another 30 min at RT, the suspension was filtered through a pad of Celite and concentrated. The crude compound was purified by silica gel flash column chromatography (AcOEt / PE 1:3) to give compound 7 as a pale yellow oil (2.03 g, 70%, ee>99% vs. racemate). [M+Na] + =497.3

[0372] (R)-1-(3-(3-(tert-butoxy)-2-oxoethoxy)phenyl)-3-(2-(tert-butoxy)-4,5-dimethoxyphenyl)propyl (S)-1-(4-(acryloyloxy)-3,3-dimethyl-2-oxobutanoyl)piperidine-2-carboxylate (9). A solution of 7 (2.03 g, 4.3 mmol) and 8 (2 g, 6.4 mmol) in CHCl (43 mL) was cooled to −20 °C, followed by the addition of a solution of DCC (1.3 g, 6.4 mmol) in CHCl (5 mL), followed by the addition of a solution of 4-(dimethylamino)pyridine (DMAP, 52.3 mg, 0.43 mmol) in CHCl (2 mL) under an argon atmosphere. The resulting white suspension was allowed to stir at −20 °C for 2 h. The reaction mixture was then filtered, evaporated, and the crude compound was purified by silica gel flash column chromatography (AcOEt / PE 1:3) to give compound 9 as a pale yellow oil (2.5 g, 76%). [M+Na] + =790.3

[0373] 2-(3-((R)-1-(((S)-1-(4-(acryloyloxy)-3,3-dimethyl-2-oxobutanoyl)piperidine-2-carbonyl)oxy)-3-(3-hydroxy-4,5-dimethoxyphenyl)propyl)phenoxy)acetic acid (Rae10). A solution of 9 (2.5 g, 3.26 mmol) in CHCl (12 mL) was treated with a solution of 40% TFA in CHCl (12 mL) at 0 °C. The mixture was allowed to react at room temperature until complete conversion. The reaction mixture was directly charged onto a silica gel flash column (AcOEt / PE / AcOH 1:2:0.5%) to give Rae10 (1.334 g, 62%) as a pale yellow solid. FKBD Example 26 2-(3-((R)-1-(((S)-1-(4-(acryloyloxy)-3,3-dimethyl-2-oxobutanoyl)piperidine-2-carbonyl)oxy)-3-(2-hydroxy-4,5-dimethoxyphenyl)propyl)phenoxy)acetic acid (Rae11) [ka] Scheme 29. Synthesis of Rae11 FKBD Moiety

[0374] 2-(tert-Butoxy)-4,5-dimethoxybenzaldehyde (2). To a solution of 2-hydroxy-4,5-dimethoxybenzaldehyde 1 (3 g, 16.5 mmol) in anhydrous toluene (15 mL) was added 1,1-di-tert-butoxy-N,N-dimethylmethanamine 2 (31.6 mL, 132 mmol) under Ar. The mixture was stirred at 80 °C for 6 h, and then the solvent was evaporated. The residue was purified by silica gel flash column chromatography (AcOEt / PE 1:1) to give compound 2 as a yellow solid (3.875 g, 99%). [M+Na] + =261.2

[0375] (E)-3-(2-(tert-butoxy)-4,5-dimethoxyphenyl)-1-(3-hydroxyphenyl)prop-2-en-1-one (4). To a solution of 2 (3.875 g, 16.3 mmol) and 3'-hydroxyacetophenone 3 (2.436 g, 17.9 mmol) in EtOH (50 mL) was added a solution of 40% aqueous KOH (8.5 mL, 3.65 g, 65.2 mmol) at 0 °C. The resulting solution was stirred at 60 °C for 4 h. The solution was poured into water and acidified to pH 4 with 1 M aqueous HCl and extracted twice with EtOAc. The combined organic layers were washed with brine, dried over Na SO , and concentrated under reduced pressure. The crude compound was purified by silica gel flash column chromatography (AcOEt / PE 1:1) to give compound 4 as a yellow oil (5.2 g, 90%). [M+H] + =357.2

[0376] tert-Butyl (E)-2-(3-(3-(2-(tert-butoxy)-4,5-dimethoxyphenyl)acryloyl)phenoxy)acetate (5). A solution of 4 (5.2 g, 14.59 mmol) and K2CO3 (2.4 g, 17.5 mmol) in DMF (50 mL) was treated with tert-butyl bromoacetate (2.55 mL, 17.5 mmol) and allowed to stir at room temperature for 5 hours. After this time, the reaction mixture was quenched with HO and extracted twice with EtOAc. The combined organic layers were washed with brine, dried over Na2SO4, and concentrated under reduced pressure. The crude compound was purified by silica gel flash column chromatography (AcOEt / PE 1:4) to give compound 5 as a yellow oil (6 g, 88%). [M+H] + =471.0

[0377] tert-Butyl 2-(3-(3-(2-(tert-butoxy)-4,5-dimethoxyphenyl)propanoyl)phenoxy)acetate (6). A solution of 5 (6 g, 12.75 mmol) and 10% Pd / C (1 g) in THF (70 mL) was hydrogenated with H2 at room temperature for 4 h. The reaction mixture was then filtered and concentrated. The residue was purified by silica gel flash column chromatography (AcOEt / PE 1:3) to give compound 6 as a colorless oil (4.5 g, 75%). [M+Na] + =495.3

[0378] tert-Butyl (R)-2-(3-(3-(2-(tert-butoxy)-4,5-dimethoxyphenyl)-1-hydroxypropyl)phenoxy)acetate (7). A solution of ketone 6 (4.5 g, 9.5 mmol) in dry THF (45 mL) was treated at −20° C. with a solution of (+)-DIPChloride (19 mmol) in heptane (1.7 M, 11.2 mL). The resulting mixture was reacted at −20° C. until complete conversion of 6 and then quenched with 2,2′-(ethylenedioxy)diethylamine (2.8 mL) to form an insoluble complex. After stirring for another 30 min at RT, the suspension was filtered through a pad of Celite and concentrated. The crude compound was purified by silica gel flash column chromatography (AcOEt / PE 1:3) to give compound 7 as a pale yellow oil (3.2 g, 70%, ee>99% vs. racemate). [M+Na] + =496.7

[0379] (R)-1-(3-(2-(tert-butoxy)-2-oxoethoxy)phenyl)-3-(2-(tert-butoxy)-4,5-dimethoxyphenyl)propyl (S)-1-(4-(acryloyloxy)-3,3-dimethyl-2-oxobutanoyl)piperidine-2-carboxylate (9). A solution of 7 (1.93 g, 4.07 mmol) and 8 (1.9 g, 6.103 mmol) in CHCl (43 mL) was cooled to −20 °C, followed by the addition of a solution of DCC (1.26 g, 6.103 mmol) in CHCl (5 mL), followed by the addition of a solution of 4-(dimethylamino)pyridine (DMAP, 50 mg, 0.407 mmol) in CHCl (2 mL) under an argon atmosphere. The resulting white suspension was allowed to stir at −20 °C for 2 h. The reaction mixture was then filtered, evaporated, and the crude compound was purified by silica gel flash column chromatography (AcOEt / PE 1:3) to give compound 9 as a pale yellow oil (2.1 g, 67%). [M+Na] + =790.4

[0380] 2-(3-((R)-1-(((S)-1-(4-(acryloyloxy)-3,3-dimethyl-2-oxobutanoyl)piperidine-2-carbonyl)oxy)-3-(2-hydroxy-4,5-dimethoxyphenyl)propyl)phenoxy)acetic acid (Rae11). A solution of 9 (2.1 g, 2.73 mmol) in CHCl (12 mL) was treated with a solution of 40% TFA in CHCl (12 mL) at 0 °C. The mixture was allowed to react at room temperature until complete conversion. The reaction mixture was directly charged onto a silica gel flash column (AcOEt / PE / AcOH 1:2:0.5%) to give Rae11 (638 mg, 23%) as a pale yellow solid. FKBD Example 27 2-(3-((R)-1-(((S)-1-(4-(acryloyloxy)-3,3-dimethyl-2-oxobutanoyl)piperidine-2-carbonyl)oxy)-3-(3-fluoro-4,5-dimethoxyphenyl)propyl)phenoxy)acetic acid (Rae12) [ka] Scheme 30. Synthesis of Rae12 FKBD moiety

[0381] (E)-3-(3-Fluoro-4,5-dimethoxyphenyl)-1-(3-hydroxyphenyl)prop-2-en-1-one (3). To a solution of 3-fluoro-4,5-dimethoxybenzaldehyde 1 (4.5 g, 24.4 mmol) and 1-(3-hydroxyphenyl)ethan-1-one 2 (3.3 g, 24.4 mmol) in EtOH (60 mL) was added a solution of 10% aqueous NaOH (40 mL, 97.6 mmol) at 0 °C. The resulting solution was stirred at room temperature for 12 h. The solution was adjusted to pH 4 with drops of 4 M aqueous HCl at 0 °C, producing a large yellow solid. The mixture was then filtered, and the solid was washed with water (50 mL) to provide 3 (4 g, 54%) as a yellow solid. [M+H] + =303.1

[0382] tert-Butyl (E)-2-(3-(3-(3-fluoro-4,5-dimethoxyphenyl)acryloyl)phenoxy)acetate (4). A solution of 3 (4 g, 13.2 mmol) and K2CO3 (3.65 g, 26.4 mmol) in DMF (30 mL) was treated with tert-butyl bromoacetate (3.08 g, 15.8 mmol) and allowed to stir at room temperature for 5 h. After this time, the reaction mixture was poured into ice and a yellow solid precipitated. The mixture was filtered and the solid was washed with water (30 mL). The crude product was purified by column chromatography on silica gel (AcOEt / PE 1:4) to give 4 (5.2 g, 94%) as a yellow solid. [M+Na] + =438.7

[0383] tert-Butyl 2-(3-(3-(3-fluoro-4,5-dimethoxyphenyl)-1-hydroxypropyl)phenoxy)acetate (5). A solution of 4 (5.2 g, 12.5 mmol) and 10% Pd / C (1 g) in THF (100 mL) was hydrogenated with H2 at room temperature for 2 h. The reaction mixture was then filtered and concentrated. The crude product was purified by column chromatography on silica gel to give 5 (5 g, 96%) as a yellow oil. [M+Na] + =443.2

[0384] tert-Butyl 2-(3-(3-(3-fluoro-4,5-dimethoxyphenyl)propanoyl)phenoxy)acetate (6). A solution of 5 (5 g, 11.9 mmol) in CHCl (100 mL) was treated with Dess-Martin periodinane (15.2 g, 36 mmol) and stirred at room temperature for 2 h before being quenched with a solution of 10% aqueous NaSO. The solution was extracted twice with CHCl. ​​The combined organic layers were washed with sat. NaHCO, brine, dried over NaSO, and concentrated under reduced pressure. The crude compound was purified by silica gel flash column chromatography (AcOEt / PE 1:3) to give compound 6 as a white solid (4 g, 80%). [M+Na] + =441.2

[0385] tert-Butyl (R)-2-(3-(3-(3-fluoro-4,5-dimethoxyphenyl)-1-hydroxypropyl)phenoxy)acetate (7). A solution of ketone 6 (4 g, 9.56 mmol) in dry THF (30 mL) was treated with a solution of (+)-DIPChloride (19.1 mmol) in heptane (1.7 M, 11.2 mL) at −20° C. The resulting mixture was reacted at −20° C. until complete conversion of 6, and then quenched with 2,2′-(ethylenedioxy)diethylamine (2.8 g) by forming an insoluble complex. After stirring for another 30 min at room temperature, the suspension was filtered through a pad of Celite and concentrated. The crude compound was purified by silica gel flash column chromatography (AcOEt / PE 1:4) to give compound 6 (2.2 g, 55%, ee>99%) as a pale yellow oil. [M+Na] + =442.7

[0386] (R)-1-(3-(2-(tert-butoxy)-2-oxoethoxy)phenyl)-3-(3-fluoro-4,5-dimethoxyphenyl)propyl (S)-1-(4-(acryloyloxy)-3,3-dimethyl-2-oxobutanoyl)piperidine-2-carboxylate (9). A solution of 7 (2.2 g, 5.23 mmol) and 8 (1.80 g, 5.76 mmol) in CHCl (20 mL) was cooled to −20 °C, followed by the addition of a solution of DCC (1.4 g, 6.79 mmol) in CHCl (10 mL) under an argon atmosphere, followed by the addition of a solution of 4-(dimethylamino)pyridine (63 mg, 0.52 mmol) in CHCl (2 mL). The resulting white suspension was allowed to stir at −20 °C for 2 h. The reaction mixture was then filtered, evaporated, and the crude compound was purified by silica gel flash column chromatography (AcOEt / PE 1:3) to give compound 9 (1.8 g, 48%) as a pale yellow oil. [M+Na] + =736.4

[0387] 2-(3-((R)-1-(((S)-1-(4-(acryloyloxy)-3,3-dimethyl-2-oxobutanoyl)piperidine-2-carbonyl)oxy)-3-(3-fluoro-4,5-dimethoxyphenyl)propyl)phenoxy)acetic acid (Rae12). A solution of 9 (1.8 g, 2.52 mmol) in CHCl (10 mL) was treated with a solution of 40% TFA in CHCl (10 mL) at 0 °C. The mixture was allowed to react at room temperature until complete conversion. The reaction mixture was directly charged onto a silica gel flash column (AcOEt / PE / AcOH 1:3:0.5%) to provide Rae12 (590 mg, 35%) as a slightly yellow solid. FKBD Example 28 2-(3-((R)-1-(((S)-1-(4-(acryloyloxy)-3,3-dimethyl-2-oxobutanoyl)piperidine-2-carbonyl)oxy)-3-(2-fluoro-4,5-dimethoxyphenyl)propyl)phenoxy)acetic acid (Rae13) [ka] Scheme 31. Synthesis of Rae13 FKBD moiety

[0388] (E)-3-(2-Fluoro-4,5-dimethoxyphenyl)-1-(3-hydroxyphenyl)prop-2-en-1-one (3). To a solution of 2-fluoro-4,5-dimethoxybenzaldehyde 1 (4.5 g, 24.4 mmol) and 1-(3-hydroxyphenyl)ethan-1-one 2 (3.3 g, 24.4 mmol) in EtOH (60 mL) was added a solution of 10% aqueous NaOH (40 mL, 97.6 mmol) at 0 °C. The resulting solution was stirred at 65 °C for 6 h. The solution was adjusted to pH 4 by adding 4 M aqueous HCl dropwise at 0 °C, producing a large yellow solid. The mixture was then filtered, and the solid was washed with water (50 mL) to provide 3 (7 g, 94%) as a yellow solid. [M+H] + =302.8

[0389] 3-(3-(2-Fluoro-4,5-dimethoxyphenyl)-1-hydroxypropyl)phenol (4). A solution of 3 (7 g, 23.1 mmol) and 10% Pd / C (2 g) in THF (150 mL) was hydrogenated with H for 12 h at room temperature. The reaction mixture was then filtered and concentrated. The crude product was purified by column chromatography on silica gel to give 4 (7 g, 98%) as a yellow oil. [M+Na] + =328.8

[0390] tert-Butyl 2-(3-(3-(2-fluoro-4,5-dimethoxyphenyl)-1-hydroxypropyl)phenoxy)acetate (5). A solution of 4 (7 g, 23.1 mmol) and K2CO3 (7 g, 50.6 mmol) in DMF (200 mL) was treated with tert-butyl bromoacetate (6.7 g, 34.5 mmol) and allowed to stir at room temperature for 24 h. After this time, the reaction mixture was poured into ice and a yellow solid precipitated. The mixture was filtered and the solid was washed with water (300 mL). The crude product was purified by column chromatography on silica gel (AcOEt / PE 1:6) to give 5 (8 g, 82%) as a yellow solid. [M+Na] + =443.2

[0391] tert-Butyl 2-(3-(3-(2-fluoro-4,5-dimethoxyphenyl)propanoyl)phenoxy)acetate (6). A solution of 5 (8 g, 19 mmol) in CHCl (100 mL) was treated with Dess-Martin periodinane (16 g, 38 mmol) and stirred at room temperature for 2 hours, then quenched with a solution of 10% aqueous NaSO. The solution was extracted twice with CHCl. ​​The combined organic layers were washed with sat. NaHCO, brine, dried over NaSO, and concentrated under reduced pressure. The crude compound was purified by silica gel flash column chromatography (AcOEt / PE 1:3) to give compound 6 as a yellow solid (6.3 g, 78%). [M+Na] + =440.7

[0392] tert-Butyl (R)-2-(3-(3-(2-fluoro-4,5-dimethoxyphenyl)-1-hydroxypropyl)phenoxy)acetate (7). A solution of ketone 6 (6.3 g, 15.07 mmol) in dry THF (60 mL) was treated with a solution of (+)-DIPChloride (45.2 mmol) in heptane (1.7 M, 26.5 mL) at −20 °C. The resulting mixture was reacted at −20 °C until complete conversion of 6 and then quenched by forming an insoluble complex with 2,2′-(ethylenedioxy)diethylamine (6.6 g). After stirring for another 30 min at room temperature, the suspension was filtered through a pad of Celite and concentrated. The crude compound was purified by silica gel flash column chromatography (AcOEt / PE 1:3) to give compound 7 (4.3 g, 68%, ee >99%) as a pale yellow oil. [M+Na] + =443.2

[0393] (R)-1-(3-(2-(tert-butoxy)-2-oxoethoxy)phenyl)-3-(2-fluoro-4,5-dimethoxyphenyl)propyl (S)-1-(4-(acryloyloxy)-3,3-dimethyl-2-oxobutanoyl)piperidine-2-carboxylate (9). A solution of 7 (1.6 g, 3.81 mmol) and 8 (1.77 g, 5.71 mmol) in CHCl (20 mL) was cooled to −20 °C, followed by the addition of a solution of DCC (1.17 g, 5.71 mmol) in CHCl (10 mL), followed by the addition of a solution of 4-(dimethylamino)pyridine (50 mg, 0.38 mmol) in CHCl (2 mL) under an argon atmosphere. The resulting white suspension was allowed to stir at −20 °C for 2 h. The reaction mixture was then filtered, evaporated, and the crude compound was purified by silica gel flash column chromatography (AcOEt / PE 1:2) to give compound 9 (1.7 g, 62%) as a pale yellow oil. [M+Na] + =736.4

[0394] 2-(3-((R)-1-(((S)-1-(4-(acryloyloxy)-3,3-dimethyl-2-oxobutanoyl)piperidine-2-carbonyl)oxy)-3-(2-fluoro-4,5-dimethoxyphenyl)propyl)phenoxy)acetic acid (Rae13). A solution of 9 (1.7 g, 2.38 mmol) in CHCl (10 mL) was treated with a solution of 40% TFA in CHCl (10 mL) at 0 °C. The mixture was allowed to react at room temperature until complete conversion. The reaction mixture was directly charged onto a silica gel flash column (AcOEt / PE / AcOH 1:3:0.5%) to give Rae13 (520 mg, 33%) as a slightly yellow solid. FKBD Example 29 2-(3-((R)-1-(((S)-1-(4-(acryloyloxy)-3,3-dimethyl-2-oxobutanoyl)piperidine-2-carbonyl)oxy)-3-(2-fluoro-3,4-dimethoxyphenyl)propyl)phenoxy)acetic acid (Rae14) [ka] Scheme 32. Synthesis of Rae14 FKBD Moiety

[0395] (E)-3-(2-Fluoro-3,4-dimethoxyphenyl)-1-(3-hydroxyphenyl)prop-2-en-1-one (3). To a solution of 1 (5.0 g, 27.17 mmol) and 2 (4.10 g, 29.89 mmol) in EtOH (150 mL) at 0 °C was added a solution of 40% aqueous KOH (15.22 g, 108.70 mmol). The resulting solution was heated to 35 °C for 2 h. The solvent was evaporated, and the residue (4.8 g 58%) was used directly for the next step without purification. [M+H] + =303.0

[0396] (E)-tert-Butyl 2-(3-(3-(2-fluoro-3,4-dimethoxyphenyl)acryloyl)phenoxy)acetate (4). A solution of 3 (5.0 g, 16.55 mmol, crude) and K2CO3 (2.74 g, 19.87 mmol) in DMF (40 mL) was treated with tert-butyl bromoacetate (3.9 g, 19.87 mmol) and allowed to stir at room temperature for 5 h. After this time, the reaction mixture was poured into ice and a yellow solid precipitated. The mixture was filtered and the solid was washed with water (30 mL). The crude product was purified by column chromatography on silica gel to give 4 (6.0 g, 80%) as a yellow solid. [M+Na] + =439.2

[0397] tert-Butyl 2-(3-(3-(2-fluoro-3,4-dimethoxyphenyl)propanoyl)phenoxy)acetate (5). A solution of 4 (4.0 g, 9.62 mmol) and 10% Pd / C (1.0 g) in THF (150 mL) was hydrogenated with H2 at room temperature for 4 h. The reaction mixture was then filtered and concentrated. The crude product was purified by column chromatography on silica gel to give 5 (2.8 g, 70%) as a yellow oil. [M+Na] + =440.8

[0398] tert-Butyl (R)-2-(3-(3-(2-fluoro-3,4-dimethoxyphenyl)-1-hydroxypropyl)phenoxy)acetate (6). A solution of ketone 5 (2.8 g, 6.7 mmol) in dry THF (30 mL) was treated with a solution of (+)-DIPChloride (26.8 mmol) in heptane (1.7 M, 15.7 mL) at −20 °C. The resulting mixture was reacted at −20 °C until complete conversion of 6 and then quenched by forming an insoluble complex with 2,2′-(ethylenedioxy)diethylamine (3.96 g). After stirring for another 30 min at room temperature, the suspension was filtered through a pad of Celite and concentrated. The crude compound was purified by silica gel flash column chromatography (AcOEt / PE 1:4) to give compound 6 (1.3 g, 46%, ee >99%) as a pale yellow oil. [M+Na] + =442.7

[0399] (R)-1-(3-(2-(tert-butoxy)-2-oxoethoxy)phenyl)-3-(2-fluoro-3,4-dimethoxyphenyl)propyl (S)-1-(4-(acryloyloxy)-3,3-dimethyl-2-oxobutanoyl)piperidine-2-carboxylate (8). A solution of 6 (1.3 g, 3.09 mmol) and 7 (1.25 g, 4.02 mmol) in CHCl (15 mL) was cooled to −20 °C, followed by the addition of a solution of DCC (0.83 g, 4.02 mmol) in CHCl (5 mL), followed by the addition of a solution of 4-(dimethylamino)pyridine (40 mg, 0.31 mmol) in CHCl (2 mL) under an argon atmosphere. The resulting white suspension was allowed to stir at −20 °C for 2 h. The reaction mixture was then filtered, evaporated, and the crude compound was purified by silica gel flash column chromatography (AcOEt / PE 1:2) to give compound 8 (1.4 g, 63%) as a pale yellow oil. [M+Na] + =736.3

[0400] 2-(3-((R)-1-(((S)-1-(4-(acryloyloxy)-3,3-dimethyl-2-oxobutanoyl)piperidine-2-carbonyl)oxy)-3-(2-fluoro-3,4-dimethoxyphenyl)propyl)phenoxy)acetic acid (Rae14). A solution of 8 (1.4 g, 1.96 mmol) in CHCl (10 mL) was treated with a solution of 40% TFA in CHCl (10 mL) at 0 °C. The mixture was allowed to react at room temperature until complete conversion. The reaction mixture was directly charged onto a silica gel flash column (AcOEt / PE / AcOH 1:3:0.5%) to give Rae14 (585 mg, 45%) as a slightly yellow solid. FKBD Example 30 2-(3-((R)-1-(((S)-4-(((9H-fluoren-9-yl)methoxy)carbonyl)-1-(4-(acryloyloxy)-3,3-dimethyl-2-oxobutanoyl)piperazine-2-carbonyl)oxy)-3-(3,4-dimethoxyphenyl)propyl)phenoxy)acetic acid (Rae16) [ka] Scheme 33. Synthesis of Rae16 FKBD moiety

[0401] (E)-3-(3,4-Dimethoxyphenyl)-1-(3-hydroxyphenyl)prop-2-en-1-one (3). To a solution of 3,4-dimethoxybenzaldehyde 1 (17.6 g, 105.8 mmol) and 3'-hydroxyacetophenone 2 (12 g, 88.2 mmol) in EtOH (160 mL) was added a solution of 40% aqueous KOH (44 mL, 20 g, 352.8 mmol) at 0 °C. The resulting solution was stirred at rt for 2 h, then poured into ice-H2O. The solution was acidified with 1 M HCl solution and extracted with EtOAc. The combined organic layers were dried over Na2SO4 and concentrated under reduced pressure. The residue was recrystallized from EtOAc-PE to give a pale yellow powder (23 g, 92%). [M+H] + =285.2

[0402] 3-(3-(3,4-Dimethoxyphenyl)-1-hydroxypropyl)phenol (4). A solution of 3 (16 g, 56.3 mmol) and 10% Pd / C (1.6 g) in THF (150 mL) was hydrogenated with H for 4 h at room temperature. The reaction mixture was then filtered and concentrated to a solid (16.3 g, quant.). [M+Na] + =311.2

[0403] tert-Butyl 2-(3-(3-(3,4-dimethoxyphenyl)-1-hydroxypropyl)phenoxy)acetate (5). A solution of 4 (16.3 g, 56.53 mmol) and K2CO3 (9.4 g, 67.83 mmol) in DMF (150 mL) was treated with tert-butyl bromoacetate (9.9 mL, 67.83 mmol) and allowed to stir at room temperature for 5 h. After this time, the reaction mixture was poured into ice and a yellow solid precipitated (20 g, 88%). [M+Na] + =424.9.

[0404] tert-Butyl 2-(3-(3-(3,4-dimethoxyphenyl)propanoyl)phenoxy)acetate (6). A solution of 5 (20 g, 49.7 mmol) in CHCl (400 mL) was treated with Dess-Martin periodinane (63 g, 149 mmol) and stirred at room temperature for 3 h before being quenched with a solution of 10% aqueous NaSO. The solution was extracted twice with CHCl. ​​The combined organic layers were washed with sat. NaHCO, brine, dried over NaSO, and concentrated under reduced pressure. The crude compound was purified by silica gel flash column chromatography (AcOEt / PE 1:3) to give compound 6 as a white solid (11 g, 55%). [M+Na] + =423.3.

[0405] tert-Butyl (R)-2-(3-(3-(3,4-dimethoxyphenyl)-1-hydroxypropyl)phenoxy)acetate (7). A solution of ketone 6 (11.156 g, 27.9 mmol) in dry THF (100 mL) was treated with a solution of (+)-DIPChloride (83.6 mmol) in heptane (1.7 M, 49 mL) at −20° C. The resulting mixture was reacted at −20° C. until complete conversion of 6 and then quenched with 2,2′-(ethylenedioxy)diethylamine (11.5 mL) to form an insoluble complex. After stirring at RT for another 30 min, the suspension was filtered through a pad of Celite and concentrated. The crude compound was purified by silica gel flash column chromatography (AcOEt / PE 1:3) to give compound 7 as a pale yellow oil (6.3 g, 58%, ee>99%). [M+Na] + =425.3.

[0406] 1-((9H-Fluoren-9-yl)methyl)3-((R)-1-(3-(2-(tert-butoxy)-2-oxoethoxy)phenyl)-3-(3,4-dimethoxyphenyl)propyl)(S)-4-(4-(acryloyloxy)-3,3-dimethyl-2-oxobutanoyl)piperazine-1,3-dicarboxylate (9). A solution of 7 (1.224 g, 3 mmol) and 8 (2.44 g, 4.56 mmol) in CHCl (10 mL) was cooled to −20 °C before the addition of a solution of DCC (0.94 g, 4.56 mmol) in CHCl (5 mL), followed by the addition of a solution of 4-(dimethylamino)pyridine (DMAP, 37 mg, 0.3 mmol) in CHCl (2 mL) under an argon atmosphere. The resulting white suspension was stirred at -20°C for 2 hours. The reaction mixture was then filtered, evaporated, and the crude compound was purified by silica gel flash column chromatography (AcOEt / PE 1:2) to give compound 9 as a pale yellow oil (1.8g, 70%). [M+Na] + =940.7.

[0407] 2-(3-((R)-1-(((S)-4-(((9H-Fluoren-9-yl)methoxy)carbonyl)-1-(4-(acryloyloxy)-3,3-dimethyl-2-oxobutanoyl)piperazine-2-carbonyl)oxy)-3-(3,4-dimethoxyphenyl)propyl)phenoxy)acetic acid (Rae 16). A solution of 9 (1.8 g, 1.96 mmol) in CHCl (11.5 mL) was treated with a solution of 40% TFA in CHCl (11.5 mL) at 0 °C. The mixture was allowed to react at room temperature until complete conversion. The reaction mixture was directly charged onto a silica gel flash column (AcOEt / PE / AcOH 1:2:0.5%) to provide Rae 16 (964 mg, 57%) as a white solid. FKBD Example 31 2-(3-((R)-1-(((S)-1-(4-(acryloyloxy)-3,3-dimethyl-2-oxobutanoyl)-4-methylpiperazine-2-carbonyl)oxy)-3-(3,4-dimethoxyphenyl)propyl)phenoxy)acetic acid (Rae17) [ka] Scheme 34. Synthesis of Rae17 FKBD Moiety

[0408] (R)-1-(3-(2-(tert-butoxy)-2-oxoethoxy)phenyl)-3-(3,4-dimethoxyphenyl)propyl (S)-1-(4-(acryloyloxy)-3,3-dimethyl-2-oxobutanoyl)piperazine-2-carboxylate (2). To a solution of 1 (1.0 g, 1.09 mmol) in DMF (5 mL) at 0 °C was added TBAF (2.5 ml, 1.0 M, 2.55 mmol). The resulting solution was allowed to warm to room temperature for 5 h. After this time, the reaction mixture was diluted with DCM and washed with saturated aqueous NaHCO3 and brine. The organic layer was concentrated in vacuo, and the residue was purified by silica gel flash column chromatography (DCM / MeOH 50:1) to give compound 2 as a colorless oil (670 mg, 80%). [M+H] + =696.9

[0409] (R)-1-(3-(2-(tert-butoxy)-2-oxoethoxy)phenyl)-3-(3,4-dimethoxyphenyl)propyl (S)-1-(4-(acryloyloxy)-3,3-dimethyl-2-oxobutanoyl)-4-methylpiperazine-2-carboxylate (3). A solution of 2 (670 mg, 0.96 mmol) in CHOOH (1.5 mL) was treated with aqueous formaldehyde (37% in water, 0.77 ml, 1.15 mmol) and allowed to stir at 50° C. for 1 h. After this time, the reaction mixture was purified with DCM / MeOH / AcOH=100 / 1 / 0.5% to give 3 (500 mg, 73%) as a colorless oil. [M+H] + =710.9

[0410] 2-(3-((R)-1-(((S)-1-(4-(acryloyloxy)-3,3-dimethyl-2-oxobutanoyl)-4-methylpiperazine-2-carbonyl)oxy)-3-(3,4-dimethoxyphenyl)propyl)phenoxy)acetic acid (Rae 17). A solution of 9 (0.5 g, 0.7 mmol) in HCOOH (40 mL) was heated to 40 °C for 2 h. The reaction mixture was directly charged to a silica gel flash column (AcOEt / PE / AcOH 1:2:0.5%) to provide Rae 17 (368.7 mg, 80%) as a white solid. FKBD Example 32 2-(3-((R)-1-(((S)-1-(4-(acryloyloxy)-3,3-dimethyl-2-oxobutanoyl)piperidine-2-carbonyl)oxy)-3-(3,4-dimethoxyphenyl)propyl)-4-fluorophenoxy)acetic acid (Rae18) [ka] Scheme 35. Synthesis of Rae18 FKBD moiety

[0411] (E)-3-(3,4-Dimethoxyphenyl)-1-(2-fluoro-5-hydroxyphenyl)prop-2-en-1-one (3). To a solution of 3,4-dimethoxybenzaldehyde 1 (5 g, 30.1 mmol) and 1-(2-fluoro-5-hydroxyphenyl)ethan-1-one 2 (4.6 g, 30.1 mmol) in EtOH (60 mL) was added a solution of 10% aqueous NaOH (50 mL, 120.4 mmol) at 0 °C. The resulting solution was stirred at room temperature for 12 h. The solution was adjusted to pH 4 by adding 4 M aqueous HCl dropwise at 0 °C, resulting in the formation of a large yellow solid. The mixture was then filtered, and the solid was washed with water (50 mL) to provide 3 (9 g, 99%) as a yellow solid. [M+H] + =303.2

[0412] 3-(3,4-Dimethoxyphenyl)-1-(2-fluoro-5-hydroxyphenyl)propan-1-one (4). A solution of 3 (9 g, 29.8 mmol) and 10% Pd / C (2 g) in THF (200 mL) was hydrogenated with H for 12 h at room temperature. The reaction mixture was then filtered and concentrated. The crude product was used for the next step without any further purification. [M+H] + =304.8

[0413] tert-Butyl 2-(3-(3-(3,4-dimethoxyphenyl)propanoyl)-4-fluorophenoxy)acetate (5). A solution of 4 (10 g, 32.8 mmol) and K2CO3 (9 g, 65.6 mmol) in DMF (200 mL) was treated with tert-butyl bromoacetate (7.7 g, 39.3 mmol) and allowed to stir at room temperature for 8 h. After this time, the reaction mixture was poured into ice, precipitating a yellow solid. The mixture was filtered, and the solid was washed with water (300 mL). The crude product was purified by column chromatography on silica gel (AcOEt / PE 1:6) to give 5 (4 g, 32%, 2 steps) as a yellow oil. [M+Na] + =441.0

[0414] tert-Butyl (R)-2-(3-(3-(3,4-dimethoxyphenyl)-1-hydroxypropyl)-4-fluorophenoxy)acetate (6). A solution of ketone 5 (4 g, 9.56 mmol) in dry THF (30 mL) was treated with a solution of (+)-DIPChloride (28.68 mmol) in heptane (1.7 M, 16.8 mL) at −20° C. The resulting mixture was reacted at −20° C. until complete conversion of 6, and then quenched by forming an insoluble complex with 2,2′-(ethylenedioxy)diethylamine (4.2 g). After stirring for another 30 min at room temperature, the suspension was filtered through a pad of Celite and concentrated. The crude compound was purified by silica gel flash column chromatography (AcOEt / PE 1:3) to give compound 6 (2 g, 50%, ee 93%) as a pale yellow oil. [M+Na] + =442.7

[0415] (R)-1-(5-(2-(tert-butoxy)-2-oxoethoxy)-2-fluorophenyl)-3-(3,4-dimethoxyphenyl)propyl (S)-1-(4-(acryloyloxy)-3,3-dimethyl-2-oxobutanoyl)piperidine-2-carboxylate (8). A solution of 6 (2 g, 4.76 mmol) and 7 (2.22 g, 7.14 mmol) in CHCl (20 mL) was cooled to −20 °C, followed by the addition of a solution of DCC (1.47 g, 7.14 mmol) in CHCl (10 mL) under an argon atmosphere, followed by the addition of a solution of 4-(dimethylamino)pyridine (60 mg, 0.47 mmol) in CHCl (2 mL). The resulting white suspension was allowed to stir at −20 °C for 2 h. The reaction mixture was then filtered, evaporated, and the crude compound was purified by silica gel flash column chromatography (AcOEt / PE 1:2) to give compound 8 (1.8 g, 45%) as a pale yellow oil. [M+Na] + =736.3

[0416] 2-(3-((R)-1-(((S)-1-(4-(acryloyloxy)-3,3-dimethyl-2-oxobutanoyl)piperidine-2-carbonyl)oxy)-3-(3,4-dimethoxyphenyl)propyl)-4-fluorophenoxy)acetic acid (Rae18). A solution of 8 (1.7 g, 2.52 mmol) in CHCl (10 mL) was treated with a solution of 40% TFA in CHCl (10 mL) at 0 °C. The mixture was allowed to react at room temperature until complete conversion. The reaction mixture was directly charged onto a silica gel flash column (AcOEt / PE / AcOH 1:3:0.5%) to provide Rae18 (705 mg, 42%) as a white solid. FKBD Example 33 2-(3-((R)-1-(((S)-1-(4-(acryloyloxy)-3,3-dimethyl-2-oxobutanoyl)piperidine-2-carbonyl)oxy)-3-(3,4-dimethoxyphenyl)propyl)-5-fluorophenoxy)acetic acid (Rae-19) [ka] Scheme 36. Synthesis of Rae19 FKBD Moiety

[0417] (E)-3-(3,4-Dimethoxyphenyl)-1-(3-fluoro-5-hydroxyphenyl)prop-2-en-1-one (3). To a solution of 3,4-dimethoxybenzaldehyde 1 (6.391 g, 38.5 mmol) and 1-(3-fluoro-5-hydroxyphenyl)ethan-1-one 2 (5.39 g, 35 mmol) in EtOH (70 mL) was added 40% aqueous KOH (19.6 g, 140 mmol) solution at 0 °C. The resulting solution was reacted at room temperature for 4 h. The yellow solid was filtered to give compound 3 (8.3 g, 78%). [M+H] + =303.0

[0418] tert-Butyl (E)-2-(3-(3-(3,4-dimethoxyphenyl)acryloyl)-5-fluorophenoxy)acetate (4). A solution of 3 (8.3 g, 27.5 mmol) and K2CO3 (4.55 g, 32.9 mmol) in DMF (80 mL) was treated with tert-butyl bromoacetate (6.4 g, 32.9 mmol) and allowed to stir at room temperature for 4 h. After this time, the reaction mixture was quenched with HO and extracted twice with EtOAc. The combined organic layers were concentrated under reduced pressure and used for the next step without purification (11.11 g, 97%). [M+Na] + =439.2

[0419] tert-Butyl 2-(3-(3-(3,4-dimethoxyphenyl)propanoyl)-5-fluorophenoxy)acetate (5). A solution of 4 (11.11 g, 26.7 mmol) and 10% Pd / C (1.11 g) in THF (200 mL) was hydrogenated with H2 at room temperature for 4 h. The reaction mixture was then filtered and concentrated. The residue was purified by silica gel flash column chromatography (AcOEt / PE 1:1) to give compound 5 as a colorless oil (4.2 g, 38%). [M+Na] + =440.7

[0420] tert-Butyl (R)-2-(3-(3-(3,4-dimethoxyphenyl)-1-hydroxypropyl)-5-fluorophenoxy)acetate (6). A solution of ketone 5 (4.2 g, 10 mmol) in dry THF (40 mL) was treated with a solution of (+)-DIPChloride (20 mmol) in heptane (1.7 M, 11.8 mL) at −20 °C. The resulting mixture was reacted at −20 °C until complete conversion of 5 and then quenched with 2,2′-(ethylenedioxy)diethylamine (2.9 mL) to form an insoluble complex. After stirring for another 30 min at RT, the suspension was filtered through a pad of Celite and concentrated. The crude compound was purified by silica gel flash column chromatography (AcOEt / PE 1:5) to give compound 6 as a pale yellow oil (2.94 g, 70%, ee 98% relative to the racemate). [M+Na] + =443.0

[0421] (R)-1-(3-(2-(tert-butoxy)-2-oxoethoxy)-5-fluorophenyl)-3-(3,4-dimethoxyphenyl)propyl (S)-1-(4-(acryloyloxy)-3,3-dimethyl-2-oxobutanoyl)piperidine-2-carboxylate (8). A solution of 6 (1.8 g, 4.28 mmol) and 7 (2 g, 6.42 mmol) in CHCl (18 mL) was cooled to −20 °C, followed by the addition of a solution of DCC (1.33 g, 6.42 mmol) in CHCl (5 mL) under an argon atmosphere, followed by the addition of a solution of 4-(dimethylamino)pyridine (DMAP, 52 mg, 0.43 mmol) in CHCl (2 mL). The resulting white suspension was allowed to stir at −20 °C for 2 h. The reaction mixture was then filtered, evaporated, and the crude compound was purified by silica gel flash column chromatography (AcOEt / PE 1:1) to give compound 8 as a pale yellow oil (2.7 g, 90%). [M+Na] + =735.9

[0422] 2-(3-((R)-1-(((S)-1-(4-(acryloyloxy)-3,3-dimethyl-2-oxobutanoyl)piperidine-2-carbonyl)oxy)-3-(3,4-dimethoxyphenyl)propyl)-5-fluorophenoxy)acetic acid (Rae19). A solution of 8 (2.7 g, 4.11 mmol) in CHCl (12 mL) was treated with a solution of 40% TFA in CHCl (12 mL) at 0 °C. The mixture was allowed to react at room temperature until complete conversion. The reaction mixture was directly charged onto a silica gel flash column (AcOEt / PE / AcOH 1:2:0.5%) to afford Rae 19 (1.094 g, 44%) as a pale yellow solid. FKBD Example 34 2-(5-((R)-1-(((S)-1-(4-(acryloyloxy)-3,3-dimethyl-2-oxobutanoyl)piperidine-2-carbonyl)oxy)-3-(3,4-dimethoxyphenyl)propyl)-2-fluorophenoxy)acetic acid (Rae20) [ka] Scheme 37. Synthesis of Raa20 FKBD Moiety

[0423] (E)-3-(3,4-Dimethoxyphenyl)-1-(4-fluoro-3-hydroxyphenyl)prop-2-en-1-one (3). To a solution of 3,4-dimethoxybenzaldehyde 1 (6.391 g, 38.5 mmol) and 1-(4-fluoro-5-hydroxyphenyl)ethan-1-one 2 (5.39 g, 35 mmol) in EtOH (70 mL) was added 40% aqueous KOH (19.6 g, 140 mmol) at 0 °C. The resulting solution was reacted at room temperature for 4 h. The yellow solid was filtered to give compound 3 (9.368 g, 89%). [M+H] + =303.2

[0424] tert-Butyl (E)-2-(5-(3-(3,4-dimethoxyphenyl)acryloyl)-2-fluorophenoxy)acetate (4). A solution of 3 (9.368 g, 31 mmol) and K2CO3 (5.1 g, 37 mmol) in DMF (90 mL) was treated with tert-butyl bromoacetate (7.2 g, 37 mmol) and allowed to stir at room temperature for 4 h. After this time, the reaction mixture was quenched with HO and extracted twice with EtOAc. The combined organic layers were concentrated under reduced pressure and used for the next step without purification (13 g, quant.). [M+Na] + =438.9

[0425] tert-Butyl 2-(5-(3-(3,4-dimethoxyphenyl)propanoyl)-2-fluorophenoxy)acetate (5). A solution of 4 (13 g, 31.2 mmol) and 10% Pd / C (1.3 g) in THF (200 mL) was hydrogenated with H2 for 4 h at room temperature. The reaction mixture was then filtered and concentrated. The residue was purified by silica gel flash column chromatography (AcOEt / PE 1:1) to give compound 5 as a colorless oil (7 g, 54%). [M+Na] + =441.2

[0426] tert-Butyl (R)-2-(5-(3-(3,4-dimethoxyphenyl)-1-hydroxypropyl)-2-fluorophenoxy)acetate (6). A solution of ketone 5 (7 g, 16.7 mmol) in dry THF (40 mL) was treated with a solution of (+)-DIPChloride (33.5 mmol) in heptane (1.7 M, 19.7 mL) at −20 °C. The resulting mixture was reacted at −20 °C until complete conversion of 5 and then quenched with 2,2′-(ethylenedioxy)diethylamine (4.89 mL) to form an insoluble complex. After stirring for another 30 min at RT, the suspension was filtered through a pad of Celite and concentrated. The crude compound was purified by silica gel flash column chromatography (AcOEt / PE 1:5) to give compound 6 as a pale yellow oil (4.9 g, 71%, ee 96% to racemate). [M+Na]+ =443.3

[0427] (R)-1-(3-(2-(tert-butoxy)-2-oxoethoxy)-4-fluorophenyl)-3-(3,4-dimethoxyphenyl)propyl (S)-1-(4-(acryloyloxy)-3,3-dimethyl-2-oxobutanoyl)piperidine-2-carboxylate (8). A solution of 6 (1.8 g, 4.28 mmol) and 7 (2 g, 6.42 mmol) in CHCl (18 mL) was cooled to −20 °C, followed by the addition of a solution of DCC (1.33 g, 6.42 mmol) in CHCl (5 mL), followed by the addition of a solution of 4-(dimethylamino)pyridine (DMAP, 52 mg, 0.43 mmol) in CHCl (2 mL) under an argon atmosphere. The resulting white suspension was allowed to stir at −20 °C for 2 h. The reaction mixture was then filtered, evaporated, and the crude compound was purified by silica gel flash column chromatography (AcOEt / PE 1:1) to give compound 8 as a pale yellow oil (2 g, 65%). [M+Na] + =736.4

[0428] 2-(5-((R)-1-(((S)-1-(4-(acryloyloxy)-3,3-dimethyl-2-oxobutanoyl)piperidine-2-carbonyl)oxy)-3-(3,4-dimethoxyphenyl)propyl)-2-fluorophenoxy)acetic acid (Rae 20). A solution of 8 (1.8 g, 2.52 mmol) in CHCl (12 mL) was treated with a solution of 40% TFA in CHCl (12 mL) at 0 °C. The mixture was allowed to react at room temperature until complete conversion. The reaction mixture was directly charged onto a silica gel flash column (AcOEt / PE / AcOH 1:2:0.5%) to give Rae 20 (835 g, 50%) as a pale yellow solid. FKBD Example 35 2-(3-((R)-1-(((S)-1-(4-(acryloyloxy)-3,3-dimethyl-2-oxobutanoyl)piperidine-2-carbonyl)oxy)-3-(3,4-dimethoxyphenyl)propyl)-2-fluorophenoxy)acetic acid (Rae21) [ka] Scheme 38. Synthesis of Raa21 FKBD Moiety

[0429] (E)-3-(3,4-Dimethoxyphenyl)-1-(2-fluoro-3-hydroxyphenyl)prop-2-en-1-one (3). To a solution of 3,4-dimethoxybenzaldehyde 1 (9.7 g, 58.4 mmol) and 1-(2-fluoro-3-hydroxyphenyl)ethan-1-one 2 (5.39 g, 35 mmol) in EtOH (70 mL) was added 40% aqueous KOH (19.6 g, 140 mmol) at 0 °C. The resulting solution was reacted at room temperature for 4 h. The yellow solid was filtered to give compound 3 (3.5 g, 30%). [M+H] + =303.0

[0430] tert-Butyl (E)-2-(3-(3-(3,4-dimethoxyphenyl)acryloyl)-2-fluorophenoxy)acetate (4). A solution of 3 (3.5 g, 11.6 mmol) and K2CO3 (1.92 g, 13.9 mmol) in DMF (40 mL) was treated with tert-butyl bromoacetate (2.7 g, 13.9 mmol) and allowed to stir at room temperature for 4 h. After this time, the reaction mixture was quenched with HO and extracted twice with EtOAc. The combined organic layers were concentrated under reduced pressure and used for the next step without purification (3.9 g, 80%). [M+H] + =416.9

[0431] tert-Butyl 2-(3-(3-(3,4-dimethoxyphenyl)propanoyl)-2-fluorophenoxy)acetate (5). A solution of 4 (3.5 g, 8.4 mmol) and 10% Pd / C (350 mg) in THF (50 mL) was hydrogenated with H2 at room temperature for 4 h. The reaction mixture was then filtered and concentrated. The residue was purified by silica gel flash column chromatography (AcOEt / PE 1:1) to give compound 5 as a colorless oil (2.45 g, 70%). [M+Na] + =441.0

[0432] tert-Butyl (R)-2-(3-(3-(3,4-dimethoxyphenyl)-1-hydroxypropyl)-2-fluorophenoxy)acetate (6). A solution of ketone 5 (2.45 g, 5.85 mmol) in dry THF (30 mL) was treated with a solution of (+)-DIPChloride (17.6 mmol) in heptane (1.7 M, 10.3 mL) at −20° C. The resulting mixture wa...

Claims

1. Macrocyclic compounds of formula (XIV): 【Chemistry 1】 or a stereoisomer, solvate or pharma- ceutically acceptable salt thereof [In the ceremony: each n, m, and p is an integer independently selected from 0 to 5; Each R 1 , R 2 , and R 3 H, F, Cl, Br, CF 3 , C.N., N. 3 , -N(R 12 ) 2 , -N(R 12 ) 3 , -CON(R 12 ) 2 , NO 2 , O.H., O.C.H. 3 , methyl, ethyl, propyl, -COOH, -SO 3 H, -PO(OR 12 ) 2 , -OPO(OR 12 ) 2 , -(CH 2 ) q COOH, -O-(CH 2 ) q COOH, -S-(CH 2 ) q COOH, -CO-(CH 2 ) q COOH, -NR 12 -(CH 2 ) q COOH, -(CH 2 ) q SO 3 H, -O-(CH 2 ) q SO 3 H, -S-(CH 2 ) q SO 3 H, -CO-(CH 2 ) q SO 3 H, -NR 12 -(CH 2 ) q SO 3 H, -(CH 2 ) q N(R 12 ) 2 , -O-(CH 2 ) q N(R 12 ) 2 , -S-(CH 2 ) q N(R 12 ) 2 , -CO-(CH 2 ) q N(R 12 ) 2 、-(CH 2 ) q N(R 12 ) 3 、-O-(CH 2 ) q N(R 12 ) 3 、-S-(CH 2 ) q N(R 12 ) 3 、-CO-(CH 2 ) q N(R 12 ) 3 、-NR 12 -(CH 2 ) q N(R 12 ) 3 、-(CH 2 ) q CON(R 12 ) 2 、-O-(CH 2 ) q CON(R 12 ) 2 、-S-(CH 2 ) q CON(R 12 ) 2 、-CO-(CH 2 ) q CON(R 12 ) 2 、-(CH 2 ) q PO(OR 12 ) 2 、-O(CH 2 ) q PO(OR 12 ) 2 、-S(CH 2 ) q PO(OR 12 ) 2 、-CO(CH 2 ) q PO(OR 12 ) 2 、-NR 12 (CH 2 ) q PO(OR 12 ) 2 、-(CH 2 ) q OPO(OR 12 ) 2 , -O(CH 2 ) q OPO(OR 12 ) 2 , -S(CH 2 ) q OPO(OR 12 ) 2 , -CO(CH 2 ) q OPO(OR 12 ) 2 , and -NR 12 (CH 2 ) q OPO(OR 12 ) 2 independently selected from the group consisting of: q is an integer selected from 0 to 5; Each R 4 , R 5 , R 6 , R 7 , R 9 , and R 11 is independently selected from the group consisting of H, methyl, ethyl, propyl, and isopropyl; Each R 8 and R 10 H, halogen, hydroxyl, C 1-20 Alkyl, N 3 , N.H. 2 , NO 2 , C.F. 3 , OCF 3 , O.C.H.F. 2 , COC 1-20 Alkyl, CO 2 C 1-20 alkyl, a 5- or 6-membered ring moiety formed together with R 7 or R 9 , -N(R 12 ) 2 , -N(R 12 ) 3 , -CON(R 12 ) 2 , -COOH, -SO 3 H, -PO(OR 12 ) 2 , -OPO(OR 12 ) 2 , -(CH 2 ) q COOH, -O-(CH 2 ) q COOH, -S-(CH 2 ) q COOH, -CO-(CH 2 ) q COOH, -NR 12 -(CH 2 ) q COOH, -(CH 2 ) q SO 3 H, -O-(CH 2 ) q SO 3 H, -S-(CH 2 ) q SO 3 H, -CO-(CH 2 ) q SO 3 H, -NR 12 -(CH 2 ) q SO 3 H, -(CH 2 ) q N(R 12 ) 2 , -O-(CH 2 ) q N(R 12 ) 2 、-S-(CH 2 ) q N(R 12 ) 2 、-CO-(CH 2 ) q N(R 12 ) 2 、-(CH 2 ) q N(R 12 ) 3 、-O-(CH 2 ) q N(R 12 ) 3 、-S-(CH 2 ) q N(R 12 ) 3 、-CO-(CH 2 ) q N(R 12 ) 3 、-NR 12 -(CH 2 ) q N(R 12 ) 3 、-(CH 2 ) q CON(R 12 ) 2 、-O-(CH 2 ) q CON(R 12 ) 2 、-S-(CH 2 ) q CON(R 12 ) 2 、-CO-(CH 2 ) q CON(R 12 ) 2 、-(CH 2 ) q PO(OR 12 ) 2 、-O(CH 2 ) q PO(OR 12 ) 2 、-S(CH 2 ) q PO(OR 12 ) 2 、-CO(CH 2 ) q PO(OR 12 ) 2 , -NR 12 (CH 2 ) q PO(OR 12 ) 2 , -(CH 2 ) q OPO(OR 12 ) 2 , -O(CH 2 ) q OPO(OR 12 ) 2 , -S(CH 2 ) q OPO(OR 12 ) 2 , -CO(CH 2 ) q OPO(OR 12 ) 2 , and -NR 12 (CH 2 ) q OPO(OR 12 ) 2 selected independently from the group consisting of Each R 12 is independently selected from the group consisting of H, methyl, ethyl, propyl, and isopropyl; However, R 2 , R 3 , R 8 and R 10 At least one of -N(R 12 ) 2 , -N(R 12 ) 3 , -CON(R 12 ) 2 , -COOH, -SO 3 H, -PO(OR 12 ) 2 , -OPO(OR 12 ) 2 , -(CH 2 ) q COOH, -O-(CH 2 ) q COOH, -S-(CH 2 ) q COOH, -CO-(CH 2 ) q COOH, -NR 12 -(CH 2 ) q COOH, -(CH 2 ) q SO 3 H, -O-(CH 2 ) q SO 3 H, -S-(CH 2 ) q SO 3 H, -CO-(CH 2 ) q SO 3 H, -NR 12 -(CH 2 ) q SO 3 H, -(CH 2 ) q N(R 12 ) 2 , -O-(CH 2 ) q N(R 12 ) 2 , -S-(CH 2 ) q N(R 12 ) 2 , -CO-(CH 2 ) q N(R 12 ) 2 , -(CH 2 ) q N(R 12 ) 3 、-O-(CH 2 ) q N(R 12 ) 3 、-S-(CH 2 ) q N(R 12 ) 3 、-CO-(CH 2 ) q N(R 12 ) 3 、-NR 12 -(CH 2 ) q N(R 12 ) 3 、-(CH 2 ) q CON(R 12 ) 2 、-O-(CH 2 ) q CON(R 12 ) 2 、-S-(CH 2 ) q CON(R 12 ) 2 、-CO-(CH 2 ) q CON(R 12 ) 2 、-(CH 2 ) q PO(OR 12 ) 2 、-O(CH 2 ) q PO(OR 12 ) 2 、-S(CH 2 ) q PO(OR 12 ) 2 、-CO(CH 2 ) q PO(OR 12 ) 2 、-NR 12 (CH 2 ) q PO(OR 12 ) 2 、-(CH 2 ) q OPO(OR 12 ) 2 、-O(CH 2 ) q OPO(OR 12 ) 2 , -S(CH 2 ) q OPO(OR 12 ) 2 , -CO(CH 2 ) q OPO(OR 12 ) 2 , and -NR 12 (CH 2 ) q OPO(OR 12 ) 2 are selected from.

2. R 1 is H.

3. R 2 is H.

4. R 3 -O-CH 2 4. The compound of claim 3, wherein said compound is COOH.

5. 5. The compound of claim 4, wherein p is 1.

6. The following structure: 【Chemistry 2】 2. The compound of claim 1, which is compound 1593 having the formula:

7. 13. A pharmaceutical composition comprising an effective amount of a compound of claim 1 and a pharma- ceutically acceptable carrier.

8. 13. A pharmaceutical composition for treating a disease in a subject, comprising administering to the subject an effective amount of a compound according to claim 1.

9. 9. The pharmaceutical composition of claim 8, wherein the disease is selected from acute kidney injury, cerebral ischemia, hepatic ischemia-reperfusion injury, and organ transplant transport solution.

10. The pharmaceutical composition of claim 9, wherein the disease is acute kidney injury.

11. The pharmaceutical composition of claim 8, wherein the compound is administered intravenously.

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