Phenylmaleimide Linker Agents

Abstract

Disclosed are compounds useful as linker-payload compounds, the compounds having the following structure (I) as a stereoisomer, enantiomer or tautomer thereof, or a pharma- ceutically acceptable salt, solvate or prodrug thereof, wherein X 1 , X 2 , X 3 , X 4 , X 5 , R 4a , and R 4b is as defined herein. Additional compounds, conjugates, methods of preparation, pharmaceutical compositions, and methods of treatment related to the conjugates of the compounds of structure (I) and a targeting moiety, or binding fragments thereof, are also provided. JPEG2025513101000528.jpg48154

Description

[Background technology]

[0001] Antibody-drug conjugates (ADCs) are a type of targeted bioconjugate. For the treatment of certain cancers, ADCs combine the targeting characteristics of monoclonal antibodies with the cancer-killing capabilities of cytotoxic drugs, offering several advantages over other non-targeted chemotherapeutic agents. However, challenges associated with the complexity of ADC molecules (specifically, the linker between the targeting peptide and the cytotoxic drug) have significantly hindered the advancement and development of new effective therapies. While the first ADC was approved in 2001, it took nearly a decade for the next generation of ADCs to be approved. Next-generation ADC molecules now contain non-cytotoxic payloads, such as immuno-oncology drugs, to stimulate the innate and adaptive immune systems to attack tumors from within.

[0002] There exists a need for chemical linkers that can enhance the efficacy of targeted therapeutic molecules (e.g., ADCs). The present disclosure fulfills this need and further provides related advantages.

[0003] In the figures, identical reference numbers identify similar elements (e.g., conjugates in Table 2). The sizes and relative positions of elements in the figures are not necessarily drawn to scale. For example, the shapes of various elements and angles are not drawn to scale, and some of these elements are enlarged and positioned to improve visibility in the figures. Furthermore, the particular shapes of the depicted elements are not intended to convey any information regarding the actual shape of the particular elements, but are selected solely for ease of recognition in the drawings. [Brief explanation of the drawings]

[0004] [Figure 1A] 1 shows TNF-α production from human PBMCs induced by conjugate II-12 in a dose-dependent manner in the presence of the HER2-expressing SK-BR-3 tumor cell line. [Figure 1B]1 shows the lack of TNF-α production from human PBMCs induced by its conjugate II-12 in the presence of the non-HER2 expressing MDA-MB-48 cell line. [Figure 2A] Figure 1 shows TNF-α production from human PBMCs induced by conjugates II-15, II-1, II-16, and II-17 in a dose-dependent manner in the presence of Nectin-4-expressing MDA-MB-175-VII tumor cell line. [Figure 2B] 1 shows the lack of TNF-α production from human PBMCs induced by conjugates II-15, II-1, II-16, and II-17 in the presence of HEK-293 cells lacking expression of nectin-4. [Figure 3A] Figure 1 shows TNF-α production from human PBMCs induced by conjugates II-18, II-19, II-19, II-20, and II-21 in a dose-dependent manner in the presence of the Nectin-4-expressing MDA-MB-175-VII tumor cell line. [Figure 3B] Figure 1 shows the lack of TNF-α production from human PBMCs induced by conjugates II-18, II-19, II-19, II-20, and II-21 in the presence of HEK-293 cells lacking expression of nectin-4. [Figure 4A] 1 shows TNF-α production from human PBMCs induced by conjugate II-3 in a dose-dependent manner in the presence of Nectin-4-expressing MDA-MB-175-VII tumor cell line. [Figure 4B] 1 shows the lack of TNF-α production from human PBMCs induced by conjugate II-3 in the presence of HEK-293 cells lacking expression of Nectin-4. [Figure 5A] 1 shows TNF-α production from human PBMCs induced by conjugate II-2 in a dose-dependent manner in the presence of Nectin-4-expressing MDA-MB-175-VII tumor cell line. [Figure 5B] 1 shows the lack of TNF-α production from human PBMCs induced by conjugate II-2 in the presence of HEK-293 cells lacking expression of Nectin-4. DETAILED DESCRIPTION OF THE INVENTION

[0005] The present disclosure provides branched phenylmaleimide compounds (e.g., compounds of structure (I)) for facilitating loading of biologically active molecules (also referred to herein as payloads) onto targeting compounds such as antibodies or ligands. The present disclosure further provides the use of linker compounds to create linker payloads for conjugation to targeting moieties (e.g., antibodies, fusion proteins, ligands). The advantage of using a hydrophilic moiety (e.g., a polar, hydrophilic, charged functional group, etc.) in combination with a phenylmaleimide parallel to the payload topology is that it enhances stability when part of the conjugate and minimizes off-target uptake or activity by masking or masking the payload hydrophobicity compared to when it is connected in series (or linearly) to the payload. Furthermore, such a structure appears to maximize the likelihood that the drug-to-antibody ratio (DAR) will remain nearly fixed while the conjugate is present in the systemic circulation (i.e., conjugate stability is maintained while retaining favorable pharmacokinetic properties). For example, when the thiol present in cysteine ​​is conjugated to the compound of structure (I), a thioether-substituted succinimide is formed.Without wishing to be bound by theory, it is believed that this succinimide is rapidly hydrolyzed due to the aforementioned substituted phenyl group of structure (I), and this hydrolyzed succinimide can no longer be deconjugated via retro-Michael reaction.Therefore, the linker payload is "locked" to the protein, and tends to better maintain its DAR.

[0006] Before describing this disclosure in more detail, it may be useful to provide definitions of certain terms used herein. Additional definitions are set forth throughout this disclosure.

[0007] As used in this specification and claims, the singular forms "a," "an," and "the" include plural referents unless the context clearly dictates otherwise. As used herein, the terms "a" and "an" should be understood to refer to "one or more" of the listed components.

[0008] The use of the alternative (eg, "or") should be understood to mean either one, both, or any combination thereof of the alternatives.

[0009] When used herein in the context of a numerical value, the term "about" refers to a range centered around that numerical value, from 15% less than that numerical value to 15% greater than that numerical value. When used in the context of a range, the term "about" refers to an expanded range, from 15% less than the lowest number recited in the range to 15% greater than the highest number recited in the range. In some embodiments, a given value refers to a range of values ​​(i.e., "about" the given value). For example, in some embodiments, pH 7.4 refers to approximately pH 7.4 (i.e., a range of 6.3 to 8.5).

[0010] Throughout this disclosure, any concentration range, percentage range, ratio range, or integer range should be understood to include any integer value within the recited range, and fractions thereof (such as integer tenths and hundredths), where appropriate, unless otherwise indicated. Also, any numerical range in this disclosure relating to any physical characteristic, such as polymer subunits, size, or thickness, should be understood to include any integer within the recited range, unless otherwise indicated. Throughout this disclosure, numerical ranges include their recited endpoints unless otherwise specified.

[0011] Unless the context otherwise requires, throughout this specification and claims, the word "comprise" and variations thereof, such as "comprises" and "comprising," are to be construed in an open, inclusive sense, i.e., "including but not limited to." As used herein, the terms "include" and "comprise" are used interchangeably.

[0012] The phrase "at least one of," when followed by a list of items or elements, refers to an open-ended set of one or more of the elements in that list, which may, but does not necessarily, include more than one of the elements.

[0013] As used herein, a "variant" protein or polypeptide comprises one or more non-naturally occurring amino acids, one or more amino acid substitutions, one or more amino acid insertions, one or more amino acid deletions, or any combination thereof, which may occur at one or more sites relative to a reference polypeptide of the present disclosure, and the variant protein or polypeptide has substantially similar activity (e.g., enzymatic function, immunogenicity) to the reference polypeptide. A variant protein or polypeptide of the present disclosure may have at least about 70%, 75%, 80%, 85%, 90%, 91%, 92%, 93%, 94%, 95%, 96%, 97%, 98%, 99%, or more sequence identity to the amino acid sequence of a reference polypeptide of the present disclosure as determined by sequence alignment programs and parameters disclosed herein. Variant polypeptides can result, for example, from genetic polymorphism or by human manipulation. Conservative amino acid substitutions are well known and can occur naturally or can be engineered when a protein is produced recombinantly. Amino acid substitutions, deletions, and additions may be introduced into proteins using mutagenesis methods known in the art (see, e.g., Sambrook et al., Molecular Cloning: A Laboratory Manual, 3rd ed., Cold Spring Harbor Laboratory Press, NY, 2001). Oligonucleotide-directed site-specific (or segment-specific) mutagenesis procedures may be used to generate polynucleotides with altered codons that provide the desired substitutions, deletions, or insertions. Alternatively, random or saturation mutagenesis techniques such as alanine scanning mutagenesis, error-prone polymerase chain reaction mutagenesis, or oligonucleotide-directed mutagenesis may be used to prepare polypeptide variants (see, e.g., Sambrook et al., supra).

[0014] In some embodiments, a "binding domain" or "binding region" or "targeting moiety" refers to a protein, polypeptide, oligopeptide, peptide, carbohydrate, nucleic acid, or combination thereof that can specifically bind to a target or targets (e.g., Nectin-4, MSLN, HER2, LRRC15, ASGR1, CD40, or BAFF and / or APRIL). Binding domains include any naturally occurring, synthetic, semi-synthetic, or recombinantly produced binding partner for a biological molecule or another target of interest. Exemplary binding domains of the present disclosure include, for example, Fab', F(ab')2, Fab, Fv, rIgG, scFv, hcAbs (heavy chain antibodies), single domain antibodies, V, and the like. HH , V NAR , sdAb, nanobody, receptor ectodomain or ligand-binding portion thereof, or ligand (e.g., cytokine, chemokine). "Fab" (Fragment Antigen Binding) is the portion of an antibody that binds to an antigen and comprises the variable region and CH1 of the heavy chain linked to the light chain via an interchain disulfide bond. Various assays are known for identifying binding domains of the present disclosure that specifically bind to a particular target, including Western blot, ELISA, and Biacore® analysis. Particularly preferred binding domains comprise immunoglobulin light and heavy chain variable domains (e.g., scFv, Fab), referred to herein as "immunoglobulin binding domains" or "immunoglobulin binding proteins." Immunoglobulin binding domains can be incorporated into various protein scaffolds or structures described herein, such as antibodies or antigen-binding fragments thereof, scFv-Fc fusion proteins, or fusion proteins comprising two or more of such immunoglobulin binding domains.

[0015] Throughout this disclosure, the term "antibody" refers to an immunoglobulin molecule that specifically binds to or is immunologically reactive with an antigen. The portion of an antibody that binds to an antigen may be referred to as an "antigen-binding domain." In certain embodiments, an antibody is an intact antibody comprising at least two heavy (H) chains and two light (L) chains interconnected by disulfide bonds, as well as an antigen-binding portion of an intact antibody that has or retains the ability to bind to a target molecule. Antibodies or antigen-binding fragments thereof of the present disclosure may include, for example, polyclonal antibodies, monoclonal antibodies, and genetically engineered antibodies. Monoclonal antibodies or antigen-binding portions thereof of the present disclosure may be, for example, non-human (e.g., murine, rabbit), chimeric, humanized, or human. In certain embodiments, antibodies of the present disclosure are heteroconjugates, bispecifics, multispecifics, diabodies, triabodies, or tetrabodies. The structure and function of immunoglobulins are reviewed, for example, in Greenfield, Ed., Antibodies: A Laboratory Manual, Chapters 2 and 3 (Cold Spring Harbor Laboratory, Cold Spring Harbor, 2014).

[0016] For example, "V L " and "V H The terms "CL" and "CH" refer to the variable binding regions from the light and heavy chains of an antibody, respectively. The variable binding region is composed of distinct and well-defined subregions known as "complementarity-determining regions" (CDRs) and "framework regions" (FRs). The term "CL" refers to an "immunoglobulin light chain constant region" or "light chain constant region," i.e., the constant region derived from an antibody light chain. The term "CH" refers to an "immunoglobulin heavy chain constant region" or "heavy chain constant region," which can be further divided depending on the antibody isotype into CH1, CH2, and CH3 (IgA, IgD, IgG) or CH1, CH2, CH3, and CH4 domains (IgE, IgM).

[0017] The binding domain and its fusion protein do not significantly bind to other components present in the test sample, but5 M -1 or higher affinity or K a (i.e., the equilibrium association constant of a particular binding interaction has units of 1 / M). Binding domains (or fusion proteins thereof) can be classified as "high affinity" binding domains (or fusion proteins thereof) and "low affinity" binding domains (or fusion proteins thereof). A "high affinity" binding domain has at least 10 8 M -1 , at least 10 9 M -1 , at least 10 10 M -1 , at least 10 11 M -1 , at least 10 12 M -1 , or at least 10 13 M -1 , preferably at least 10 8 M -1 Or at least 10 9 M -1 K a "Low affinity" binding domains refer to those binding domains that have a maximum affinity of 10 8 M -1 , up to 10 7 M -1 , up to 10 6 M -1 , up to 10 5 M -1 Alternatively, affinity is measured in units of M (e.g., 10 -5 M~10 -13 M) the equilibrium dissociation constant (K d The affinity of binding domain polypeptides and fusion proteins according to the present disclosure can be readily determined using conventional techniques (see, e.g., Scatchard et al., Ann. NY Acad. Sci. 51:660, 1949, and U.S. Pat. Nos. 5,283,173, 5,468,614, or equivalents).

[0018] As used herein, a "derivative" refers to a chemically or biologically modified version of a compound that is structurally similar to and (actually or theoretically) derivable from the parent compound. Generally, a "derivative" differs from an "analog" in that the parent compound may be the starting material for generating a "derivative," whereas the parent compound may not necessarily be used as the starting material for generating an "analog." An analog may have different chemical or physical properties than the parent compound. For example, a derivative may be more hydrophilic or have altered reactivity (e.g., a CDR with amino acid changes that alter its affinity for a target) compared to the parent compound.

[0019] As used herein, "identical" or "identity" refers to the similarity between a DNA, RNA, nucleotide, amino acid, or protein sequence and another DNA, RNA, nucleotide, amino acid, or protein sequence. Identity can be expressed in terms of the percentage of sequence identity of a first sequence to a second sequence. The percent sequence identity (%) with respect to a reference DNA sequence can be the percentage of DNA nucleotides in a candidate sequence that are identical to DNA nucleotides in the reference DNA sequence after aligning the sequences. The percent sequence identity (%) with respect to a reference amino acid sequence can be the percentage of amino acid residues in a candidate sequence that are identical to amino acid residues in the reference amino acid sequence after aligning the sequences and introducing gaps, if necessary, to achieve the maximum percent sequence identity (without considering any conservative substitutions as part of the sequence identity). For example, percent sequence identity values ​​for the sequences provided herein can be generated using NCBI BLAST 2.0 software as defined by Altschul et al., "Gapped BLAST and PSI-BLAST: a new generation of protein database search programs," Nucleic Acids Res. 2007, 25, 3389-3402, with parameters set to default values.

[0020] "Oxo" refers to a group having the formula =O.

[0021] "Nitro" refers to a group having the formula -NO2.

[0022] "Thioxo" refers to a group having the formula =S.

[0023] "Cyano" refers to a group having the formula --CN.

[0024] "Amino" refers to a group having the formula -NH2.

[0025] "Hydroxyl" or "hydroxy" refers to a group having the formula --OH.

[0026] "Thiol" refers to a group having the formula --SH.

[0027] "Aldehyde" refers to a group having the formula -C(=O)H.

[0028] "Carboxyl" refers to a functional group of formula -C(=O)OH.

[0029] "Halo" refers to a halogen radical, e.g., -F, -Cl, -Br, -I, and the like.

[0030] "Alkyl" means a group consisting solely of carbon and hydrogen atoms, having 1 to 12 carbon atoms (C1-C 12"Alkyl" refers to a saturated, straight or branched hydrocarbon chain group, such as methyl, ethyl, n-propyl, 1-methylethyl (isopropyl), n-butyl, n-pentyl, 1,1-dimethylethyl (t-butyl), 3-methylhexyl, 2-methylhexyl, etc., having 1 to 8 carbon atoms (C1-C8 alkyl), or 1 to 6 carbon atoms (C1-C6 alkyl), or any value within these ranges (e.g., C4-C6 alkyl), connected to the rest of the molecule by a single bond. The carbon numbers referenced are in relation to the carbon backbone and carbon branches, but do not include carbon atoms belonging to any substituents. Similarly, alkenyl refers to a straight or branched hydrocarbon chain group composed of carbon and hydrogen with at least one carbon-carbon double bond. "Alkynyl" contains at least one carbon-carbon triple bond. Unless stated otherwise in this specification, alkyl, alkenyl, or alkyne groups are optionally substituted.

[0031] "Haloalkyl" refers to an alkyl group, as defined above, that is substituted by one or more halo groups, as defined above, e.g., trifluoromethyl, difluoromethyl, trichloromethyl, 2,2,2-trifluoroethyl, 1,2-difluoroethyl, 3-bromo-2-fluoropropyl, 1,2-dibromoethyl, etc. Unless stated otherwise in the specification, a haloalkyl group is optionally substituted.

[0032] "Alkoxy" means a group of the formula -OR a where R a Contains 1 to 12 carbon atoms (C1 to C 12 and alkyl groups as defined above, containing 1 to 8 carbon atoms (C1-C8 alkoxy), or 1 to 6 carbon atoms (C1-C6 alkoxy), or any value within these ranges. Unless stated otherwise in the specification, alkoxy groups are optionally substituted.

[0033] "Haloalkoxy" refers to an alkoxy group, as defined above, that is substituted by one or more halo groups, as defined above. Unless stated otherwise in the specification, a haloalkoxy group is optionally substituted.

[0034] "Aminyl" is a group of the formula -NR a R b where R a and R b are each independently H or C1-C6 alkyl as defined above. a and R b When both are H, the "aminyl" group is the same as the "amino" group defined above. Unless otherwise specified, the C1-C6 alkyl portion of the aminyl group is optionally substituted.

[0035] "Amindyl" is a compound of the formula -C(=O)NR a R b where R a and R b are each independently H or C1-C6 alkyl as defined above. Alternatively, amine diyl may also be a group or -N(R a )C(=O)R b where R a is either H or C1-C6 alkyl, and R b is C1-C6 alkyl. Unless otherwise specified, the C1-C6 alkyl portion of the aminyl group is optionally substituted.

[0036] "Carbohydrate" refers to a group consisting of carbon, hydrogen, and oxygen atoms. In some embodiments, the carbohydrate has a hydrogen to oxygen ratio of 2:1. In some embodiments, the carbohydrate is C m (H2O) nwhere m and n are integers that may or may not be the same. Exemplary carbohydrates include sugars (e.g., monosaccharides, disaccharides, oligosaccharides, polysaccharides), starch, and cellulose. In some embodiments, the carbohydrate is selected from the group consisting of glucose, fructose, sucrose, ribose, amylose, lactose, galactose, xylose, maltose, isomaltulose, trehlose, sorbitol, mannitol, maltodextrin, raffinose, stachyose, amylose, amylpectin, glycogen, cellulose, hemicellulose, pectin, hydrocolloids, and the like.

[0037] "Haloalkyl" refers to an alkyl group as defined above that is substituted by one or more halo groups. The haloalkyl group is attached to the main chain via an alkyl carbon atom. Unless otherwise specified herein, the haloalkyl group is optionally substituted.

[0038] "Carboxyalkyl" refers to an alkyl group, as defined above, substituted with one or more carboxy groups. The carboxyalkyl group is attached to the main chain via an alkyl carbon atom. Unless otherwise specified herein, the carboxyalkyl group is optionally substituted.

[0039] "Cycloalkyl" or "carbocycle" refers to a stable non-aromatic monocyclic or polycyclic hydrocarbon group, consisting solely of carbon and hydrogen atoms, which may include fused or bridged ring systems, having 3 to 15 carbon atoms, preferably 3 to 10 carbon atoms, saturated or unsaturated, and connected to the remainder of the molecule by a single bond. Monocyclic groups include, for example, cyclopropyl, cyclobutyl, cyclopentyl, cyclohexyl, cycloheptyl, and cyclooctyl. Polycyclic groups include, for example, adamantyl, norbornyl, decalinyl, 7,7-dimethyl-bicyclo[2.2.1]heptanyl, and the like. Unless otherwise specified in the specification, cycloalkyl groups may be optionally substituted.

[0040] "Aryl" refers to any stable monocyclic, bicyclic, or polycyclic carbocyclic ring system of 4 to 12 atoms in each ring, with at least one ring being aromatic. Some examples of aryl include phenyl (sometimes written as "Ph"), naphthyl, tetrahydronaphthyl, indanyl, anthracyl, and biphenyl. When the aryl substituent is bicyclic and one ring is non-aromatic, it is understood that the connection is via the aromatic ring. Aryl can be substituted or unsubstituted.

[0041] The term "heterocycle" or "heterocyclyl" refers to an aromatic or non-aromatic ring system of 5 to 22 atoms, in which 1 to 4 of the ring atoms are heteroatoms selected from oxygen, nitrogen, and sulfur. Thus, a heterocycle can be its heteroaryl, dihydro, or tetrahydro form. Heterocycles include pyrrolidine, tetrihydrofuran, thiolane, indolinyl, 3H-indolyl, azetidine, oxetane, thietane, diazetidine, dioxetane, dithietane, piperidine, tetrahydrofuran, pyran, tetrahydropyran, thiacyclohexane, tetrahydrothiophene, pyridine, pyrimidine, and the like.

[0042] "Heteroaryl" refers to any stable monocyclic, bicyclic, or polycyclic carbocyclic ring system of 4 to 12 atoms in each ring, wherein at least one ring is aromatic and contains 1 to 4 heteroatoms selected from oxygen, nitrogen, and sulfur. Some examples of heteroaryl include acridinyl, quinoxalinyl, pyrazolyl, indolyl, benzotriazolyl, furanyl, thienyl, benzothienyl, benzofuranyl, quinolinyl, isoquinolinyl, oxazolyl, isoxazolyl, pyrazinyl, pyridazinyl, pyridinyl, pyrimidinyl, pyrrolyl, and tetrahydroquinolinyl. Heteroaryl includes N-oxide derivatives of nitrogen-containing heteroaryls.

[0043] "Alkylene" or "alkylene chain" refers to a straight or branched divalent hydrocarbon chain, consisting solely of carbon and hydrogen, containing no unsaturation, having 1 to 12 carbon atoms, e.g., methylene, ethylene, propylene, n-butylene, etc., that connects the rest of the molecule to a radical group. The alkylene chain is connected to the rest of the molecule through a single bond and to the radical group through a single bond. The points of attachment of the alkylene chain to the rest of the molecule and to the radical group can be through one carbon or any two carbons within the chain. Unless stated otherwise in the specification, alkylene is optionally substituted.

[0044] Similarly, "alkenylene" refers to a straight or branched divalent hydrocarbon chain, consisting solely of carbon and hydrogen and containing at least one double carbon-carbon bond, that connects the rest of the molecule to a radical group. The alkenylene chain can contain from 2 to 12 carbon atoms. "Alkynylene" is also a straight or branched divalent hydrocarbon chain, consisting solely of carbon and hydrogen but containing at least one triple carbon-carbon bond, that connects the rest of the molecule to a radical group.

[0045] "Heteroalkylene" refers to an alkylene group, as defined above, containing at least one heteroatom (e.g., Si, N, O, P, or S) within the alkylene chain or at a terminus of the alkylene chain. In some embodiments, the heteroatom is within the alkylene chain (i.e., the heteroalkylene has at least one carbon-[heteroatom] x-carbon bond, where x is 1, 2, or 3. In other embodiments, the heteroatom is at the terminus of the alkylene, thus serving to connect the alkylene to the remainder of the molecule (e.g., M1-HA-M2, where M1 and M2 are part of the molecule, H is the heteroatom, and A is an alkylene). Unless stated otherwise herein, heteroalkylene groups are optionally substituted. Exemplary heteroalkylene groups include ethylene oxide (e.g., polyethylene oxide), propylene oxide, amino acid chains (i.e., short- to medium-length peptides—containing 1-15 amino acids), and alkylene chains (including combinations thereof) connected through various functional groups such as amide, disulfide, phosphate, sulfate, sulfonamide, ester, ether, -S-, carbamate, urea, thiourea, anhydride, and the like. In some embodiments, the heteroalkylene includes polyamino acids having 1-10 amino acids. In some embodiments, the heteroalkylene comprises a polyamino acid having 1 to 5 amino acids.

[0046] "Heteroalkenylene" refers to a heteroalkylene group, as defined above, containing at least one carbon-carbon double bond. "Heteroalkynylene" refers to a heteroalkylene group, as defined above, containing at least one carbon-carbon triple bond.

[0047] A "heteroatom linker" refers to a continuous chain of heteroatoms or heteroatoms that connects a portion of a molecule to a radical group. A heteroatom linker may be polyvalent (e.g., divalent, trivalent, etc.). A heteroatom linker consists solely of non-carbon atoms (e.g., H, O, N, S, Si, P, etc.). Unless otherwise specified herein, a heteroatom linker may be optionally substituted.

[0048] A "cycloalkylene" is a polyvalent (e.g., divalent, trivalent, etc.) cycloalkyl group. Unless stated otherwise in the specification, a cycloalkylene group may be optionally substituted.

[0049] An "arylene" is a polyvalent (e.g., divalent, trivalent, etc.) aryl group. Unless stated otherwise in the specification, an arylene group may be optionally substituted.

[0050] A "heterocyclylene" is a polyvalent (e.g., divalent, trivalent, etc.) heterocyclyl group. Unless stated otherwise in the specification, a heterocyclylene group may be optionally substituted.

[0051] A "heteroarylene" is a polyvalent (e.g., divalent, trivalent, etc.) heteroaryl group. Unless stated otherwise in the specification, a heteroarylene group may be optionally substituted.

[0052] "Linker" refers to a continuous chain of at least one atom, such as carbon, oxygen, silicon, nitrogen, sulfur, phosphorus, and combinations thereof, that connects one part of a molecule to another part of the same molecule or to a different molecule or fragment thereof via a covalent bond (e.g., a single bond, a double bond, or a triple bond). In some embodiments, the linker is an optionally substituted alkylene linker, an optionally substituted alkenylene linker, an optionally substituted alkynylene linker, an optionally substituted heteroalkylene linker, an optionally substituted heteroalkenylene linker, an optionally substituted heteroalkynylene linker, a heteroatom linker, a cycloalkylene linker, an arylene linker, a heterocyclylene linker, a heteroarylene linker, or a combination thereof. Unless otherwise specified herein, the linker may be optionally substituted.

[0053] A "trigger element" refers to a molecular motif that is recognized by a biological molecule (e.g., a protease such as a cathepsin) or is susceptible to chemical reaction in a biological environment (e.g., acid-labile). Typically, enzymatic recognition of the trigger element results in a catalytic cleavage reaction that results in release of the payload from the antibody plus linker(s).

[0054] "Sacrificial element" or "self-immolative spacer" refers to a chemically labile group that facilitates the release or cleavage between molecular moieties (e.g., between an antibody-linker and a payload). A sacrificial element is typically a bivalent linker motif that is compatible with one or more chemical reactions that ultimately result in the cleavage of the covalent bond between the two ends of the group. The reaction then occurs as some kind of stimulus to the system (e.g., a protease catalyzing a reaction during recognition of the trigger element, or a decrease in pH when the antibody-drug conjugate is exposed to the intracellular environment) and then results in the controlled release of the payload. Commonly used sacrificial elements are para-aminobenzyloxycarbonyl (PABC) and aminal. In some embodiments, the sacrificial element is [ka] The structure of the compound is:

[0055] The following scheme illustrates the fragmentation of para-aminobenzyloxycarbonyl (PABC) and release of the payload: [ka] where D represents the unmodified payload.

[0056] In some embodiments, the sacrificial element is [ka] wherein: R 6a , R 6b , R 6c , and R 6d are independently hydrogen, optionally substituted alkyl, optionally substituted aryl, or optionally substituted heteroaryl; or R 6a and R 6c together with the nitrogen and carbon atoms to which they are attached form an azetidinyl, pyrrolodinyl, piperidinyl, or homopiperidinyl, and R 6d is hydrogen, Y1 is -O-, -S-, or -NR 6b -It is.

[0057] In some embodiments, the sacrificial element is [ka] wherein: R 6e , R 6f , R 6g , and R 6h are independently hydrogen, optionally substituted alkyl, optionally substituted aryl, or optionally substituted heteroaryl; or R 6a and R 6c together with the nitrogen and carbon atoms to which they are attached form an azetidinyl, pyrrolodinyl, piperidinyl, or homopiperidinyl, and R 6d is hydrogen, Y 2 is -O-, -S-, or -NR 6f -It is.

[0058] In certain embodiments, the sacrificial element is [ka] wherein: R 10 each occurrence is independently alkyl, alkoxy, or halo; R 11 is hydrogen, alkyl, or -(CH2CH2O) z3 -CH3, R 12 is hydrogen or alkyl, R 13 is hydrogen or alkyl, z1 is 0 or 1, z2 is 0, 1, 2, 3, or 4; z3 is 0, 1, 2, 3, 4, 5, 6, 7, 8, 9, or 10.

[0059] In some embodiments, the sacrificial element is [ka] wherein: R 14a , R 14b , R 14c , R 14d , R 14e , and R 14f are each independently hydrogen, alkyl, hydroxyalkyl, or alkoxyalkyl; z4, z5, z6, and z7 are each independently 1, 2, 3, 4, 5, or 6.

[0060] The above term includes all systems containing effective cleavage elements for endolysosomal processing of antibody-drug conjugates. Exemplary systems include cathepsin-based enzyme peptide sequences such as Val-Cit, Val-Ala, Ala-Ala, and Gly-Gly-Phe-Gly. Also included are trigger elements sensitive to the enzyme b-glucuronidase, which contain glucuronic acid as the trigger element. In some embodiments, the trigger element is directly covalently linked to the sacrificial element. In some embodiments, the sacrificial element is directly covalently linked to the payload, and cleavage irreversibly releases the payload.

[0061] "Non-cleavable element" refers to a linker that does not contain either a trigger element or a sacrificial element and is not cleaved under normal physiological conditions. Depending on the target, many conjugates can be highly effective even without a formal cleavage-release element. Thus, in some embodiments, the payload can be directly connected to the phenylmaleimide via a non-cleavable linker.

[0062] "Heteroalkylene element" refers to a linker comprising one or more heteroalkylenes, as defined above. In some embodiments, heteroalkylene elements can be used to optimize linker-payload characteristics. In some embodiments, the heteroalkylene element(s) increase the linear connectivity between other elements (e.g., charged elements, hydrophilic elements, etc.). In some embodiments, the heteroalkylene element increases the distance between the polar cap or payload of the phenylmaleimide moiety of structure (I). The heteroalkylene elements vary in length, structure, polarity, degree of branching, etc. The aforementioned variables allow the linker-payload, and therefore the antibody-linker-drug conjugate, to have the best overall properties (e.g., solubility, monodispersity, etc.).

[0063] In some embodiments, the heteroalkylene element comprises a linear polyethylene glycol having two ethylene glycol units (i.e., PEG2) and one or more amino acids. In other embodiments, the heteroalkylene element comprises a Gly-Gly didpeptide. In some embodiments, the heteroalkylene element comprises a PEG 1~24 In another embodiment, the heteroalkylene element comprises an amino acid. In another embodiment, the heteroalkylene element comprises PEG, 1~24 In another embodiment, the heteroalkylene element is PEG 1~12 In another embodiment, the heteroalkylene element is PEG 2~6 In some embodiments, PEG x and an amino acid combine to form a heteroalkylene element. Exemplary linear PEG moieties can be found, for example, in PCT Publication No. 2021 / 207701, which is incorporated herein by reference in its entirety.

[0064] "Hydrophilic element" refers to the portion of structure (I) that effectively promotes solubility in aqueous solvents (e.g., water, phosphate buffered saline) throughout the molecule. In effect, the hydrophilic element masks or offsets the hydrophobic portion of the linker-payload, such that the resulting molecule is stable in aqueous environments (e.g., pH 7.4 buffered water). In some embodiments, the hydrophilic element produces a stable and soluble linker-payload with improved overall physicochemical properties. The hydrophilic element provides suitable chemical functionality that allows stable conjugation to proteins. In some embodiments, the incorporation of appropriate hydrophilic element(s) results in the resulting protein conjugate having a more protein-like ADME / DMPK profile.

[0065] In some embodiments, the hydrophilic elements are of medium length (10-14 units or PEG 10~14 In some embodiments, the hydrophilic element comprises a PEG linker of 2-24 units in length (i.e., PEG 2~24 ). In some embodiments, the length and amount of branching included in the hydrophilic element can be adjusted based on other elements present in structure (I). In some embodiments, the hydrophilic element is a C1-C6 alkoxy (e.g., methoxy). In some embodiments, the hydrophilic element includes polysarcosine (PSAR). In one aspect, the hydrophilic element imparts polarity and hydrophilicity to the overall conjugate. In some embodiments, the hydrophilic element is of sufficient size and structural flexibility to mask hydrophobicity with or without the trigger element. In some embodiments, the hydrophilic element is divalent. In some embodiments, the hydrophilic element is a monovalent group. In some embodiments, the hydrophilic element is polyvalent. In some embodiments, the hydrophilic element is branched. In some embodiments, the hydrophilic element is linear.

[0066] Hydrophilic groups (e.g., moieties that constitute hydrophilic elements) are known to those skilled in the art. For example, hydrophilic groups can be found in PCT Publication Nos. 2019 / 217591 and 2018 / 089373, each of which is incorporated herein by reference in its entirety.

[0067] "Polar cap" refers to a group having a structure containing one or more polar or hydrophilic functional groups (e.g., one or more of OH, -NH, -C(=O)OH, -S(O), -OP(O), etc.). In some embodiments, the polar cap is attached to a terminal end of structure (I). In some embodiments, the polar cap is attached to more than one terminal end of structure (I) (e.g., when the polar cap is attached to multiple terminals of a branched heteroalkylene element or a branched hydrophilic element). The polar cap confers hydrophilic properties to the overall compound of structure (I). In some embodiments, the polar cap includes one or more moieties that are negatively charged under physiological conditions (e.g., phosphate, carboxylic acid, sulfate, sulfonic acid, etc.). In some embodiments, the polar cap includes one or more moieties that are positively charged under physiological conditions (e.g., quaternary amines). In some embodiments, the polar cap itself is zwitterionic under physiological conditions, having an overall negative, overall positive, or overall neutral charge. In some embodiments, the polar cap serves to increase both polarity and charge through the incorporation of an amino acid, which may have additional functionality to add further polarity and charge. In some embodiments, the polar cap comprises L-glutamic acid. In some embodiments, the polar cap has two carboxy groups. In some embodiments, the polar cap comprises one or more dibasic acids. In some embodiments, the glutamic acid is further modified to have one or both acids amidated with a polyol-containing amine. In some embodiments, the polar cap comprises an amino acid with a glycoside moiety (e.g., L-serine-beta-D-glucoside, etc.).

[0068] "Stable compound" or "stable structure" is meant to indicate a compound that is sufficiently robust to survive isolation to a useful degree of purity from a reaction mixture, and formulation into an efficacious therapeutic agent.

[0069] "Salts" includes both acid addition salts (e.g., salts formed with inorganic or organic acids such as hydrochloric acid, hydrobromic acid, sulfuric acid, acetic acid, ascorbic acid, etc.) and base addition salts (e.g., salts formed with inorganic or organic bases such as sodium, potassium, amine bases, etc.).

[0070] Crystallization may produce solvates of the compounds described herein (e.g., compounds of structure (I)). Embodiments of the present disclosure include all solvates of the described compounds. As used herein, the term "solvate" refers to an aggregate comprising one or more molecules of a compound of the present disclosure and one or more solvent molecules.

[0071] Embodiments of the disclosed compounds (e.g., compounds of structure (I)), or salts, tautomers, or solvates thereof, may contain one or more stereocenters and thus give rise to enantiomers, diastereomers, and other stereoisomeric forms that may be defined in terms of absolute stereochemistry as (R)- or (S)-, or (D)- or (L)- for amino acids. Embodiments of the disclosure are meant to include all such possible isomers, as well as their racemic and optically pure forms. Optically active (+) and (−), (R)- and (S)-, or (D)- and (L)-isomers may be prepared using chiral synthons or chiral reagents and resolved using conventional techniques, such as chromatography and fractional crystallization. Conventional techniques for the preparation / isolation of individual enantiomers include chiral synthesis from suitable optically pure precursors or resolution of the racemate (or racemate of a salt or derivative) using, for example, chiral high-pressure liquid chromatography (HPLC). When a compound described herein contains an olefinic double bond or other feature which gives rise to geometric asymmetry, unless otherwise specified, the compound is intended to include both E and Z geometric isomers, as well as all tautomeric forms.

[0072] "Stereoisomer" refers to a compound composed of the same atoms joined by the same bonds, but having different three-dimensional structures that are not interchangeable. The present disclosure contemplates various stereoisomers and mixtures thereof, and includes "enantiomers," which refers to two stereoisomers whose molecules are non-superimposable mirror images of one another.

[0073] "Tautomer" refers to a proton shift from one atom of a molecule to another atom of the same molecule. The present disclosure includes tautomers of any of the above-mentioned compounds. Various tautomeric forms of compounds can be easily derived by one skilled in the art.

[0074] The chemical naming protocols and structure diagrams used herein are a modification of the IUPAC naming system, using ACD / Name version 9.07 software and / or ChemDraw Ultra version 11.0 software naming program (CambridgeSoft). Common names familiar to those skilled in the art are also used.

[0075] For ease of illustration, various compounds of structure (I) or conjugates of structure (II) with sulfur- or phosphorus-containing moieties (e.g., sulfonates, phosphates, etc.) may be depicted in an anionic state (e.g., -S(O)3 or -OP(O)3). One of skill in the art will readily appreciate that charge is pH dependent and that uncharged (e.g., protonated form(s) or salt form(s), e.g., sodium or other cations) forms are also within the scope of embodiments of the present disclosure.

[0076] Linker Compounds In certain embodiments, the present disclosure provides branched phenylmaleimide compounds (e.g., compounds of structure (I)) that allow for the formation of a covalent bond between a linker-payload and a protein. As indicated above, certain embodiments of the present disclosure provide compounds useful as linkers between a payload (including fragments thereof) and a targeting peptide (including fragments thereof, e.g., antibodies).

[0077] Thus, some embodiments include a compound having the following structure (I): [ka] During the ceremony, X 1 , X 2 , X 3 , X 4 and X 5 One of them is CL 1 -R 1 and X 1 , X 2 , X 3 , X 4 and X 5Another one of them is CL 2 -R 2 and X 1 , X 2 , X 3 , X 4 and X 5 The remaining three of these are independently N, CR 3 , or CL 3 -R 3a and R 1 , R 2 , and R 3a each independently comprises one or more moieties selected from an amino acid element, a charged element, a heteroalkylene element, a hydrophilic element, a trigger element, a sacrificial element, a polar cap, a payload, and combinations thereof; However, R 1 and R 2 at least one of which contains a payload; R 3 each occurrence independently represents hydrogen, deuterium, alkyl, haloalkyl, halo, alkoxy, haloalkoxy, amino, aminyl, amidyl, aldehyde, hydroxyl, cyano, nitro, thiol, carboxy, carboxyalkyl, alkyl-S(O)3H, alkyl-OP(O)3H, alkyl-P(O)3H, -O-carboxyalkyl, -O-alkyl-S(O)3H, -O-alkyl-OP(O)3H, -O-alkyl-P selected from the group consisting of (O)H, -S(O)H, -OP(O)H, -P(O)H, alkyl-OP(O)-alkyl, alkyl-P(O)-alkyl, -O-alkyl-S(O)-alkyl, -O-alkyl-OP(O)-alkyl, -O-alkyl-P(O)-alkyl, -S(O)-alkyl, -OP(O)-alkyl, -P(O)-alkyl, sulfamide, sulfinimide, and carbohydrate; R 4a and R 4b each independently represents hydrogen, deuterium, halo, or -SR 4c and R 4c Substituted or unsubstituted C6-C 10 aryl or substituted or unsubstituted 5- to 12-membered heteroaryl; L 1 , L 2 , and L 3 are each independently a linker comprising an optionally substituted alkylene, optionally substituted alkenylene, optionally substituted alkynylene, optionally substituted heteroalkylene, optionally substituted heteroalkenylene, optionally substituted heteroalkynylene, a heteroatom linker, an optionally substituted cycloalkylene, an optionally substituted arylene, an optionally substituted heterocyclylene, an optionally substituted heteroarylene, or a combination thereof; Provided are the compounds as stereoisomers, enantiomers or tautomers thereof, or mixtures thereof, or pharmaceutically acceptable salts, solvates or prodrugs thereof.

[0078] A particular embodiment is a compound having the following structure (I): [ka] During the ceremony, X 1 , X 2 , X 3 , X 4 and X 5 One of them is CL 1 -R 1 and X 1 , X 2 , X 3 , X 4 and X 5 Another one of them is CL 2 -R 2 and X 1 , X 2 , X 3 , X 4 and X 5 The remaining three of these are independently N, CR 3 , or CL 3 -R 3a and R 1 , R 2 , and R 3aeach independently comprises one or more moieties selected from an amino acid element, a charged element, a heteroalkylene element, a hydrophilic element, a trigger element, a sacrificial element, a polar cap, a payload, and combinations thereof; However, R 1 and R 2 at least one of which contains a payload; R 3 each occurrence independently represents hydrogen, deuterium, alkyl, haloalkyl, halo, alkoxy, haloalkoxy, amino, aminyl, amidyl, aldehyde, hydroxyl, cyano, nitro, thiol, carboxy, carboxyalkyl, alkyl-S(O)3H, alkyl-OP(O)3H, alkyl-P(O)3H, -O-carboxyalkyl, -O-alkyl-S(O)3H, -O-alkyl-OP(O)3H, -O-alkyl-P selected from the group consisting of (O)H, -S(O)H, -OP(O)H, -P(O)H, alkyl-OP(O)-alkyl, alkyl-P(O)-alkyl, -O-alkyl-S(O)-alkyl, -O-alkyl-OP(O)-alkyl, -O-alkyl-P(O)-alkyl, -S(O)-alkyl, -OP(O)-alkyl, -P(O)-alkyl, sulfamide, sulfinimide, and carbohydrate; R 4a and R 4b each independently represents hydrogen, deuterium, halo, or -SR 4c and R 4c Substituted or unsubstituted C6-C 10 aryl or substituted or unsubstituted 5- to 12-membered heteroaryl; L 1 , L 2 , and L 3are each independently a direct bond or a linker comprising an optionally substituted alkylene, optionally substituted alkenylene, optionally substituted alkynylene, optionally substituted heteroalkylene, optionally substituted heteroalkenylene, optionally substituted heteroalkynylene, a heteroatom linker, optionally substituted cycloalkylene, optionally substituted arylene, optionally substituted heterocyclylene, optionally substituted heteroarylene, or a combination thereof; Provided are the compounds as stereoisomers, enantiomers or tautomers thereof, or mixtures thereof, or pharmaceutically acceptable salts, solvates or prodrugs thereof.

[0079] In some embodiments, R 4a and R 4b and R are both hydrogen. 4a is a halo or R 4b is halo. In some embodiments, R 4a , R 4b , or both [ka] The structure of the compound is:

[0080] In some embodiments, R 1 , R 2 , and / or R 3a comprises elements selected from amino acid elements, charged elements, heteroalkylene elements, hydrophilic elements, trigger elements, sacrificial elements, polar caps, payloads, and combinations thereof. It is understood that these elements can be connected in any order and can be connected linearly or via branched connections. In some embodiments, R 1 R 2 , and / or R 3a includes multiple occurrences of an element (eg, two or more heteroalkylene elements, two or more hydrophilic elements, two or more polar caps, etc.).

[0081] In some embodiments, R 1 R 2 , or R 3a contains a branch point as part of the amino acid element (e.g., lysine), with additional elements connected via the epsilon amine of the lysine and other additional elements linked to the amino acid element via one or more peptide bonds to the alpha carbon of the lysine. A similar motif can be utilized with the amino acid element glutamic acid. In some embodiments, the amino acid element is [ka] It contains one of the following structures:

[0082] In some embodiments, the compound of structure (I) is [ka] [ka] It contains one of the following structures:

[0083] In certain embodiments, R 1 teeth, [ka] wherein: L 1a is an amino acid element, L 1b is the charging element, L 1c is a heteroalkylene element, L 1d is the hydrophilic element, L 1e is the trigger element, L 1f But it is a sacrificial element, L 1a , L 1b , L 1c , L 1d , L 1e , and L 1fOne or more occurrences of, optionally, L 1a , L 1b , L 1c , L 1d , L 1e , and L 1f to form one or more rings, each occurrence of n1, n2, n3, n4, n5, and n6 is independently an integer from 0 to 3, provided that n1+n2+n3+n4+n5+n6=1 or greater; n7 is 1, 2, 3, 4, 5, or 6; R 1a But, L 1a , L 1b , L 1c , L 1d , L 1e , or L 1f wherein the payload is optionally substituted with a polar cap.

[0084] In some embodiments, n7 is 1, 2, or 3. In some embodiments, n7 is 1 or 2. In some embodiments, n7 is 1.

[0085] In some embodiments, R 1 teeth, [ka] wherein: L 1a is an amino acid element, L 1b is the charging element, L 1c is a heteroalkylene element, L 1d is the hydrophilic element, L 1e is the trigger element, L 1f But it is a sacrificial element, L 1a , L 1b , L 1c , L 1d , L 1e, and L 1f One or more occurrences of, optionally, L 1a , L 1b , L 1c , L 1d , L 1e , and L 1f to form one or more rings, each occurrence of n1, n2, n3, n4, n5, and n6 is independently an integer from 0 to 3, provided that n1+n2+n3+n4+n5+n6=1 or greater; n7 is 1, 2, 3, 4, 5, or 6; R 1a is the payload optionally substituted with a polar cap.

[0086] In more embodiments, R 2 teeth, [ka] wherein: L 2a is an amino acid element, L 2b is the charging element, L 2c is a heteroalkylene element, L 2d is the hydrophilic element, L 2e is the trigger element, each occurrence of m1, m2, m3, m4, and m5 is independently an integer from 0 to 3, provided that m1+m2+m3+m4+m5=1 or greater; m6 is 1, 2, 3, 4, or 5; R 2a is hydrogen, alkyl, payload, or polar cap.

[0087] In some embodiments, m6 is 1, 2, or 3. In some embodiments, m6 is 1 or 2. In some embodiments, m6 is 1.

[0088] In even more embodiments, R3a teeth, [ka] wherein: L 3a is an amino acid element, L 3b is the charging element, L 3c is a heteroalkylene element, L 3d is the hydrophilic element, L 3e is the trigger element, each occurrence of p1, p2, p3, p4, and p5 is independently an integer from 0 to 3, provided that p1+p2+p3+p4+p5=1 or greater; p6 is 1, 2, 3, 4, or 5; R 3b is hydrogen, alkyl, or a polar cap.

[0089] In some embodiments, p6 is 1, 2, or 3. In some embodiments, p6 is 1 or 2. In some embodiments, p6 is 1.

[0090] In certain embodiments, n7 is 1, and n1-n6 are each 1. In some embodiments, n7 is 1, n1-n4 are each 0, n5 is 1, and n6 is 1. In certain embodiments, n7 is 1, n1 is 0, n2 is 0, n3 is 1, n4 is 0, n5 is 1, and n6 is 1. In certain embodiments, n7 is 1, n1 is 1, n2 is 0, n3 is 1, n4 is 0, n5 is 1, and n6 is 1. In certain embodiments, n7 is 1, n1 is 1, n2 is 1, n3 is 1, n4 is 0, n5 is 1, and n6 is 1. In certain embodiments, n7 is 1, n1 is 1, n2 is 1, n3 is 1, n4 is 0, n5 is 1, and n6 is 1. In certain embodiments, n7 is 1, n1 is 1, n2 is 1, n3 is 1, n4 is 1, n5 is 1, and n6 is 1. In certain embodiments, n7 is 1, n1 is 1, n2 is 0, n3 is 1, n4 is 0, n5 is 1, and n6 is 1. In certain embodiments, n7 is 2. In certain embodiments, n7 is 3.

[0091] In some embodiments, m6 is 1, and each of m1-m5 is 1. In some embodiments, m6 is 1, m1 is 1, m2 is 0, m3 is 0, m4 is 1, and m5 is 0. In some embodiments, m6 is 1, m1 is 1, m2 is 1, m3 is 0, m4 is 1, and m5 is 0. In some embodiments, m6 is 1, m1 is 1, m2 is 0, m3 is 1, m4 is 1, and m5 is 0. In some embodiments, m6 is 1, m1 is 1, m2 is 0, m3 is 0, m4 is 1, and m5 is 1. In some embodiments, m6 is 2. In certain embodiments, m6 is 3.

[0092] In some embodiments, p6 is 1, and each of p1 through p5 is 1. In particular embodiments, p6 is 1, p1 is 1, p2 is 0, p3 is 0, p4 is 1, and p5 is 0. In some embodiments, p6 is 1, p1 is 1, p2 is 1, p3 is 0, p4 is 1, and p5 is 0. In some embodiments, p6 is 1, p1 is 1, p2 is 0, p3 is 1, p4 is 1, and p5 is 0. In some embodiments, p6 is 1, p1 is 1, p2 is 0, p3 is 0, p4 is 1, and p5 is 1. In some embodiments, p6 is 2, at least one occurrence of p1 is 1, p2 is 1, p3 is 0, p4 is 1, and p5 is 0. In some embodiments, p6 is 2. In certain embodiments, p6 is 3.

[0093] In some embodiments, the amino acid element comprises one or more amino acids selected from the group consisting of glycine, alanine, serine, threonine, cysteine, valine, leucine, isoleucine, methionine, proline, phenylalanine, tyrosine, tryptophan, aspartic acid, glutamic acid, asparagine, glutamine, histidine, lysine, arginine, sarconesine, and beta-alanine.

[0094] In certain embodiments, the amino acid element is selected from the group consisting of glycine, sarcosine, beta-alanine, and glutamic acid.

[0095] In some embodiments, the amino acid element comprises a dipeptide, tripeptide, tetrapeptide, or pentapeptide.

[0096] In more embodiments, the amino acid element is: [ka] wherein: R 5aEach occurrence of is independently hydrogen, alkyl, hydroxyalkyl, or alkoxyalkyl.

[0097] In some embodiments, the charged element comprises a moiety that has a negative charge at pH 7.4 (i.e., in the range of 6.3 to 8.5). In certain embodiments, the charged element comprises a moiety that has a positive charge at pH 7.4 (i.e., in the range of 6.3 to 8.5).

[0098] In some embodiments, the charged element comprises one or more charged amino acids, one or more carboxylic acids, one or more sulfonic acids, one or more sulfonamides, one or more sulfates, one or more phosphates, one or more quaternary amines, one or more sulfamides, one or more sulfinimides, or a combination thereof.

[0099] In certain embodiments, the charged amino acid is aspartic acid, glutamic acid, histidine, lysine, or arginine.

[0100] In some embodiments, R 1 , R 2 , or R 3a comprises a non-cleavable linker (e.g., a linker, or segment thereof, that does not include a trigger element or a sacrificial element).

[0101] In some embodiments, R 1 , R 2 , or R 3a teeth, [ka] wherein: R 5b , R 5c R 5d , and R 5eis independently selected from the group consisting of hydrogen, deuterium, alkyl, haloalkyl, halo, alkoxy, haloalkoxy, amino, hydroxyl, cyano, nitro, thiol, carboxyalkyl, alkyl-S(O)H, alkyl-OP(O)H, alkyl-P(O)H, -O-carboxyalkyl, -O-alkyl-S(O)H, -O-alkyl-OP(O)H, -O-alkyl-P(O)H, -S(O)H, -OP(O)H, -P(O)H, alkyl-OP(O)3-alkyl, alkyl-P(O)3-alkyl, -O-alkyl-S(O)3-alkyl, -O-alkyl-OP(O)3-alkyl, -O-alkyl-P(O)3-alkyl, -S(O)3-alkyl, -OP(O)3-alkyl, -P(O)3-alkyl, sulfamide, sulfinimide; R 5f each occurrence of is independently hydrogen, alkyl, hydroxyalkyl, or alkoxyalkyl; R 9 each occurrence of is independently hydrogen or alkyl; each occurrence of q2 is independently an integer from 1 to 25; Each occurrence of q3 is independently an integer from 5 to 15.

[0102] In some embodiments, the hydrophilic component comprises polyethylene glycol, polysarcosine, cyclodextrin, c-glycoside, or a combination thereof. [ka] wherein: R 5b , R 5c R 5d , and R 5eis independently selected from the group consisting of hydrogen, deuterium, alkyl, haloalkyl, halo, alkoxy, haloalkoxy, amino, hydroxyl, cyano, nitro, thiol, carboxyalkyl, alkyl-S(O)H, alkyl-OP(O)H, alkyl-P(O)H, -O-carboxyalkyl, -O-alkyl-S(O)H, -O-alkyl-OP(O)H, -O-alkyl-P(O)H, -S(O)H, -OP(O)H, and -P(O)H; R 5g each occurrence of is independently hydrogen, alkyl, hydroxyalkyl, or alkoxyalkyl; Each occurrence of q4 is independently an integer from 1 to 24.

[0103] In some embodiments, the hydrophilic element is [ka] It contains one of the following structures:

[0104] In some embodiments, the hydrophilic element is [ka] [ka] It contains one of the following structures:

[0105] In some embodiments, the hydrophilic element is [ka] The structure of the compound is:

[0106] In some embodiments, the hydrophilic element has the following structure: [ka]

[0107] In some embodiments, the hydrophilic element is [ka] The structure of the compound is:

[0108] In some embodiments, the hydrophilic element is [ka] The structure of the compound is:

[0109] In some embodiments, the hydrophilic component comprises polysarcosine. In some embodiments, the hydrophilic component comprises: [ka] It is a polysarcosine containing the structure:

[0110] In some embodiments, the hydrophilic element is [ka] The polysarcosine has one of the following structures:

[0111] In some embodiments, the hydrophilic element has a molecular weight greater than 150 g / mol, greater than 200 g / mol, greater than 300 g / mol, greater than 400 g / mol, greater than 500 g / mol, greater than 600 g / mol, greater than 700 g / mol, greater than 800 g / mol, greater than 900 g / mol, or greater than 1000 g / mol. In some embodiments, the hydrophilic element has a molecular weight less than 150 g / mol, less than 200 g / mol, less than 300 g / mol, less than 400 g / mol, less than 500 g / mol, less than 600 g / mol, less than 700 g / mol, less than 800 g / mol, less than 900 g / mol, or less than 1000 g / mol.

[0112] In certain embodiments, L 1 is alkylene. In some embodiments, L 1 is C1-C6 alkylene. In certain embodiments, L2 is alkylene. In some embodiments, L 2 is C1-C6 alkylene. In certain embodiments, L 3 is alkylene. In some embodiments, L 3 is C1-C6 alkylene.

[0113] In certain embodiments, L 1 is heteroalkylene. In some embodiments, L 1 is C1-C6 heteroalkylene (i.e., containing 1 to 6 carbon atoms and one or more heteroatoms). 2 is heteroalkylene. In some embodiments, L 2 is C1-C6 heteroalkylene. 3 is heteroalkylene. In some embodiments, L 3 is C1-C6 heteroalkylene.

[0114] In more embodiments, L 1 , L 2 , or L 3 is a C1-C6 heteroalkylene containing heteroatoms selected from O and N. In some embodiments, L 3 is a direct bond.

[0115] In more embodiments, L 1 , L 2 , or L 3 teeth, [ka] The structure of the compound is:

[0116] In some embodiments, the trigger element comprises a dipeptide, tripeptide, tetrapeptide, pentapeptide, glucuronide, disulfide, phosphate, diphosphate, triphosphate, hydrazone, or a combination thereof. In some other embodiments, the trigger element comprises beta-glucuronic acid. In certain embodiments, the trigger element comprises a dipeptide, tripeptide, tetrapeptide, or pentapeptide. In some embodiments, the trigger element comprises two or more amino acids selected from the group consisting of valine, citrulline, alanine, glycine, phenylalanine, lysine, or a combination thereof. In certain embodiments, the trigger element comprises a sequence of amino acids selected from the group consisting of valine-citrulline, valine-alanine, glycine-glycine-phenylalanine-glycine, and combinations thereof. In some embodiments, the trigger element comprises [ka] The compound includes one of the following structures (including combinations thereof):

[0117] In some embodiments, the triggering element has a molecular weight greater than 150 g / mol, greater than 200 g / mol, greater than 300 g / mol, greater than 400 g / mol, greater than 500 g / mol, greater than 600 g / mol, greater than 700 g / mol, greater than 800 g / mol, greater than 900 g / mol, or greater than 1000 g / mol. In some embodiments, the triggering element has a molecular weight less than 150 g / mol, less than 200 g / mol, less than 300 g / mol, less than 400 g / mol, less than 500 g / mol, less than 600 g / mol, less than 700 g / mol, less than 800 g / mol, less than 900 g / mol, or less than 1000 g / mol.

[0118] In some embodiments, the trigger element is: [ka] It has the following structure.

[0119] In some embodiments, the trigger element is specifically cleaved by an enzyme. For example, the trigger element can be cleaved by a lysosomal enzyme. The trigger element can be peptide-based or can include a peptide region that can act as a substrate for the enzyme. Peptide-based trigger elements can be more stable in plasma and extracellular environments than chemically labile linkers.

[0120] Exemplary disulfide-containing trigger elements are: [ka] may include the structure where D is a payload and R is independently selected at each occurrence from, for example, hydrogen or C1-C6 alkyl. Increasing steric hindrance adjacent to the disulfide bond can increase the stability of the linker. The above structure can have increased in vivo stability when one or more R groups are selected from lower alkyl, such as methyl.

[0121] The peptide bond can have good serum stability because lysosomal protease activity in blood can be very low due to endogenous inhibitors, and the pH value of blood is unfavorably high compared to lysosomes. The release of the payload from the conjugate of structure (II) can occur due to the action of lysosomal proteases, such as cathepsin and plasmin. These proteases can be present at high levels in certain tumor tissues. The trigger element can be cleavable by a lysosomal enzyme. The lysosomal enzyme can be, for example, cathepsin B, β-glucuronidase, or β-galactosidase.

[0122] The cleavable peptide of the trigger element can be selected from tetrapeptides such as Gly-Phe-Leu-Gly, Ala-Leu-Ala-Leu, tripeptides such as Glu-Val-Cit, or dipeptides such as Val-Cit, Val-Ala, Ala-Ala, and Phe-Lys. Dipeptides may have lower hydrophobicity compared to longer peptides.

[0123] In some embodiments, the trigger element can be a single amino acid residue, hi some embodiments, the trigger element comprises an Asn (e.g., legumain-cleavable).

[0124] The enzymatically cleavable trigger element is combined with a sacrificial element to provide additional spatial separation between the payload and the enzymatic cleavage site. Direct attachment of the payload to the peptide trigger element can result in proteolytic release of the payload or amino acid adducts of the payload, thereby impairing its activity. The use of a sacrificial element can enable release of a fully active, chemically unmodified payload upon amide bond hydrolysis.

[0125] The trigger element may contain a chemically labile group, such as a hydrazone group and / or a disulfide group. Trigger elements containing a chemically labile group or multiple chemically labile groups can take advantage of the different properties between plasma and some cytoplasmic compartments. The intracellular conditions that can facilitate payload release for hydrazone-containing trigger elements may be the acidic environment of endosomes and lysosomes, while disulfide-containing trigger elements may be reduced in the cytoplasm, which may contain high thiol concentrations, such as glutathione. The plasma stability of trigger elements containing chemically labile groups can be increased by introducing steric hindrance using substituents near the chemically labile group.

[0126] Acid-labile groups such as hydrazones can remain intact during systemic circulation in the neutral pH environment of blood (pH 7.3-7.5), and can undergo hydrolysis and release the payload when the conjugate of structure (II) is internalized into the mildly acidic endosomal (pH 5.0-6.5) and lysosomal (pH 4.5-5.0) compartments of cells. This pH-dependent release mechanism may be associated with nonspecific release of the payload. To increase the stability of the hydrazone group of the trigger element, the trigger element can be altered by chemical modification, e.g., substitution, allowing tuning to achieve more efficient release within the lysosome with minimal loss in the circulation.

[0127] In some embodiments, the trigger element is: [ka] wherein R is selected from C1-C6 alkyl, aryl, and —O—C1-C6 alkyl.

[0128] The hydrazone-containing trigger element may contain additional cleavage sites, such as additional acid-labile cleavage sites and / or enzymatically labile cleavage sites (e.g., disulfides). Exemplary conjugates and compounds containing hydrazone-containing trigger elements include, for example, conjugates having the following structures: [ka] wherein R is selected from C1-C6 alkyl, aryl, and —O—C1-C6 alkyl.

[0129] Other acid-labile groups that can be included in the trigger element include cis-aconityl-containing linkers. Cis-aconityl chemistry uses a carboxylic acid juxtaposed to the amide bond to accelerate amide hydrolysis under acidic conditions.

[0130] The trigger element may also contain a disulfide group. The disulfide may be thermodynamically stable at physiological pH and may release the payload upon internalization of the conjugate of structure (II) into cells, where the cytosol may provide a significantly more reducing environment compared to the extracellular environment. Cleavage of the disulfide bond may require the presence of a cytoplasmic thiol cofactor, such as (reduced) glutathione (GSH), such that the disulfide-containing trigger element is reasonably stable in the circulation and can selectively release the payload in the cytoplasm. The intracellular enzyme protein disulfide isomerase, or a similar enzyme capable of cleaving disulfide bonds, may also contribute to preferential cleavage of disulfide bonds within cells. GSH may be present in cells at a concentration range of 0.5 to 10 mM, compared to the significantly lower concentrations of GSH at approximately 5 μM or cysteine, the most abundant low-molecular-weight thiol in the circulation. Tumor cells, where irregular blood flow can cause hypoxia, can result in increased activity of reductase enzymes, thus further increasing glutathione concentrations. The in vivo stability of disulfide-containing trigger elements can be improved by chemical modification of the trigger element, for example, by the use of steric hindrance adjacent to the disulfide bond.

[0131] The trigger element can also be a β-glucuronic acid-based linker. The easy release of the payload can be achieved by cleavage of the β-glucuronide glycosidic bond by the lysosomal enzyme β-glucuronidase. This enzyme can be abundant in lysosomes and can be overexpressed in some tumor types, but the extracellular enzyme activity can be low. β-glucuronic acid-based linkers can be used to avoid the tendency of the conjugate to undergo aggregation due to the hydrophilic nature of β-glucuronide. In some embodiments, the trigger element comprises β-glucuronic acid.

[0132] The following scheme illustrates the release of a payload (D) from a conjugate of structure (II) containing a β-glucuronic acid-based trigger element. [ka]

[0133] A variety of cleavable β-glucuronic acid-based linkers have been described that are useful for linking drugs such as auristatins, camptothecin analogs, doxorubicin analogs, CBI minor groove binders, and psymberin to antibodies. Accordingly, these β-glucuronic acid-based trigger elements are used in conjugates of structure (II). In some embodiments, the trigger element comprises a β-galactoside-based linker. β-galactosides are abundant in lysosomes but have low extracellular enzymatic activity.

[0134] The trigger element may comprise one or more peptides. In some embodiments, the peptides may be selected to contain natural amino acids, unnatural amino acids, or any combination thereof. In some embodiments, the peptides may be tripeptides or dipeptides. In certain embodiments, the dipeptide comprises an L-amino acid, such as Val-Cit; Cit-Val; Ala-Ala; Ala-Cit; Cit-Ala; Asn-Cit; Cit-Asn; Cit-Cit; Val-Glu; Glu-Val; Ser-Cit; Cit-Ser; Lys-Cit; Cit-Lys; Asp-Cit; Cit-Asp; Ala-Val; Val-Ala; Phe-Lys; Lys-Phe; Val-Lys; Lys-Val; Ala-Lys; Lys-Ala; Phe-Cit; Cit-Phe; Leu-Cit; Cit-Leu; Ile-Cit; Cit-Ile; Phe-Arg; Arg-Phe; Cit-Trp; and Trp-Cit, or a salt thereof.

[0135] Trigger elements and sacrificial elements are known in the art, for example, in International Application No. PCT / US2021 / 054296, which trigger elements and sacrificial elements are incorporated herein by reference in their entirety.

[0136] One sacrificial element can be a bifunctional para-aminobenzyl alcohol group that can link to a trigger element via its amino group to form an amide bond, while an amine-containing payload can be attached to the benzyl hydroxyl group of the para-aminobenzyl alcohol via a carbamate functionality (to give a p-amidobenzyl carbamate). The resulting pro-compound can be activated upon protease-mediated cleavage, resulting in a 1,6-elimination reaction that releases the unmodified payload and the remainder of the antibody linker.

[0137] In some embodiments, the sacrificial element comprises para-aminobenzyloxycarbonyl, aminal, hydrazine, disulfide, amide, ester, hydrazine, phosphotriester, diester, β-glucuronide, double bond, triple bond, ethyl bond, ketone, diol, cyano, nitro, quaternary amine, or a combination thereof. In certain embodiments, the sacrificial element comprises para-methoxybenzyl, dialkyldialkoxysilane, diaryldialkoxysilane, orthoester, acetal, optionally substituted β-thiopropionate, ketal, phosphoramidate, hydrazone, vinyl ether, imine, aconityl, trityl, polyketal, bis-arylhydrazone, diazobenzene, bivinaldiol, pyrophosphate diester, or a combination thereof.

[0138] In certain embodiments, the sacrificial element is [ka] It contains one of the following structures:

[0139] In some embodiments, the sacrificial element is [ka] wherein: R 6a , R 6b , R 6c , and R 6dare independently hydrogen, optionally substituted alkyl, optionally substituted aryl, or optionally substituted heteroaryl; or R 6a and R 6c together with the nitrogen and carbon atoms to which they are attached form an azetidinyl, pyrrolodinyl, piperidinyl, or homopiperidinyl, and R 6d is hydrogen, Y 1 is -O-, -S-, or -NR 6b -It is.

[0140] In certain embodiments, the sacrificial element is [ka] wherein: R 6e , R 6f , R 6g , and R 6h are independently hydrogen, optionally substituted alkyl, optionally substituted aryl, or optionally substituted heteroaryl; or R 6a and R 6c together with the nitrogen and carbon atoms to which they are attached form an azetidinyl, pyrrolodinyl, piperidinyl, or homopiperidinyl, and R 6d is hydrogen, Y 2 is -O-, -S-, or -NR 6f -It is.

[0141] In some embodiments, the sacrificial element is [ka] wherein: R 10 each occurrence is independently alkyl, alkoxy, or halo; R 11 is hydrogen, alkyl, or -(CH2CH2O) z3 -CH3, R12 is hydrogen or alkyl, R 13 is hydrogen or alkyl, z1 is 0 or 1, z2 is 0, 1, 2, 3, or 4; z3 is 0, 1, 2, 3, 4, 5, 6, 7, 8, 9, or 10; or In some embodiments, the sacrificial element is [ka] wherein: R 14a , R 14b , R 14c , R 14d , R 14e , and R 14f are each independently hydrogen, alkyl, hydroxyalkyl, or alkoxyalkyl; z4, z5, z6, and z7 are each independently 1, 2, 3, 4, 5, or 6. In some embodiments, the sacrificial element is [ka] wherein: z8 and z9 are each independently 1, 2, 3, 4, 5, or 6; or In certain embodiments, the sacrificial element is [ka] wherein: R 15 is independently H, methyl, ethyl, isopropyl, tert-butyl, or phenyl; Y 3 is O or CH2, q5 is an integer ranging from 1 to 5.

[0142] In some embodiments, the sacrificial element has a molecular weight greater than 150 g / mol, greater than 200 g / mol, greater than 300 g / mol, greater than 400 g / mol, greater than 500 g / mol, greater than 600 g / mol, greater than 700 g / mol, greater than 800 g / mol, greater than 900 g / mol, or greater than 1000 g / mol. In some embodiments, the sacrificial element has a molecular weight less than 150 g / mol, less than 200 g / mol, less than 300 g / mol, less than 400 g / mol, less than 500 g / mol, less than 600 g / mol, less than 700 g / mol, less than 800 g / mol, less than 900 g / mol, or less than 1000 g / mol.

[0143] In some embodiments, the sacrificial element and the trigger element together comprise: [ka] having the structure where the trigger element is designated "peptide" and contains 1 to 10 amino acids, and JPEG2025513101000045.jpg5136 represents the attachment point to the payload. In some embodiments, the peptide comprises Val-Cit or Val-Ala. Heterocyclic variants of this sacrificial element (e.g., pyridinyl, pyrimidinyl, etc.) can also be used.

[0144] In some embodiments, the sacrificial element contains a phenol group covalently bonded to the rest of the molecule via the phenolic oxygen. One such sacrificial element relies on a methodology in which a diamino-ethane "Space Link" is used in conjunction with a conventional "PABO"-based sacrificial element to deliver the phenol.

[0145] In some embodiments, the trigger element may include a non-cleavable moiety or segment: polyethylene glycol (PEG) and related polymers can be included with cleavable groups such as disulfides, hydrazones, or dipeptides to form sacrificial groups and / or trigger elements.

[0146] Other degradable bonds that can be included in the sacrificial element can include esters. Esters can be formed by reacting PEG carboxylic acids or activated PEG carboxylic acids with alcohol groups on the payload. Such ester groups can be hydrolyzed under physiological conditions to release the payload. Other hydrolyzable bonds can include carbonate bonds, imine bonds resulting from the reaction of amines and aldehydes, phosphate ester bonds formed by reacting alcohols with phosphate groups, acetal bonds that are the reaction product of aldehydes and alcohols, orthoester bonds that are the reaction product of formates and alcohols, and oligonucleotide bonds formed by phosphoramidite groups that comprise the termini of polymers and 5' hydroxyl groups of oligonucleotides.

[0147] In some embodiments, the trigger element, the sacrificial group, and the payload are all: [ka] having the structure During the ceremony,

number

[0148] In some embodiments, the phenyl moiety of PABC is substituted with one or more substituents. In some embodiments, the substituents are: [ka] The structure of the compound is: In some embodiments, the sacrificial group is [ka] It contains one of the following structures:

[0149] In some embodiments, the trigger element, victim element, and payload together have one of the following structures, where the payload is represented by a letter: [ka]

[0150] In some embodiments, the sacrificial element is [ka] It has the following structure.

[0151] The above structure shows a substitution pattern of 1, 3, 4 on the phenyl ring of the sacrificial element. In some embodiments, the substitution pattern can be 1, 2, 4 (i.e., 1 is the bond to the payload, 2 is the bond to the remainder of the molecule, and 4 is the bond to the carbohydrate) or 1, 3, 5 (i.e., 1 is the bond to the payload, 3 is the bond to the remainder of the molecule, and 4 is the bond to the carbohydrate).

[0152] Although cleavable linkers (e.g., linkers with trigger or sacrificial elements) may offer certain advantages, the linker does not need to be cleavable. For non-cleavable linkers, payload release may not depend on the different properties between plasma and some cytoplasmic compartments. Payload release may occur after internalization of the conjugate of structure (II) via antigen-mediated endocytosis and delivery to the lysosomal compartment, where the targeting moiety (or its binding fragment) can be degraded to the amino acid level by intracellular proteolytic degradation. This process can release the payload or payload derivative. The payload or payload derivative may be more hydrophilic and less membrane permeable, which can result in fewer bystander effects and nonspecific toxicity compared to conjugates with cleavable linkers. Conjugates with non-cleavable linkers may have higher stability in the circulation than conjugates with cleavable linkers. The non-cleavable linker may comprise an alkylene chain, or may be, for example, a polymer based on a polyalkylene glycol polymer, an amide polymer, or may comprise segments of an alkylene chain, a polyalkylene glycol, and / or an amide polymer. The linker may contain a polyethylene glycol segment having 1 to 6 ethylene glycol units. In some embodiments, -L 1 -R 1 or L 2 -R 2 comprises a linker that is not cleavable in vivo.

[0153] In some embodiments, the trigger element and the sacrificial element together comprise: [ka] [ka] It contains one of the following structures:

[0154] In some embodiments, the heteroalkylene element comprises polyethylene glycol or polypropylene glycol. In some embodiments, the heteroalkylene element comprises [ka] wherein: R 5b , R 5c R 5d , and R 5e is independently selected from the group consisting of hydrogen, deuterium, alkyl, haloalkyl, halo, alkoxy, haloalkoxy, amino, hydroxyl, cyano, nitro, thiol, carboxyalkyl, alkyl-S(O)H, alkyl-OP(O)H, alkyl-P(O)H, -O-carboxyalkyl, -O-alkyl-S(O)H, -O-alkyl-OP(O)H, -O-alkyl-P(O)H, -S(O)H, -OP(O)H, and -P(O)H; Each occurrence of q1 is independently an integer from 1 to 24.

[0155] In some embodiments, R 5b , R 5c R 5d , and R 5e are all hydrogen.

[0156] In some embodiments, the polar cap comprises one or more charged amino acids, one or more polyols, or a combination thereof. In certain embodiments, the polar cap comprises a diol, triol, tetraol, or a combination thereof. In some embodiments, the polar cap comprises glycerol, trimethylolpropane, pentaerythritol, maltitol, sorbitol, xylitol, erythritol, isomalt, or a combination thereof. In certain embodiments, the polar cap comprises one or more natural amino acids. In some embodiments, the polar cap comprises one or more unnatural amino acids. In some embodiments, the polar cap comprises one or more unnatural amino acids and one or more natural amino acids. In certain embodiments, the polar cap comprises serine, threonine, cysteine, proline, asparagine, glutamine, lysine, arginine, histidine, aspartate, glutamate, 4-hydroxyproline, 5-hydroxylysine, homoserine, homocysteine, ornithine, beta-alanine, statin, or gamma-aminobutyric acid. In certain embodiments, the polar cap comprises aspartic acid, serine, glutamic acid, serine-beta-glucose, or a combination thereof.

[0157] In some embodiments, the polar cap is [ka] It contains one of the following structures:

[0158] In more embodiments, the polar cap is [ka] The compound includes one of the following structures (including combinations thereof):

[0159] In more embodiments, L 1 , L 2 , or L 3 is alkylene, alkylene-L a -, alkenylene, alkenylene-L a-, alkynylene, alkynylene-L a -, -L a -, -L a -Alkylene-L a -, -L a -Alkenylene-L a -, -L a -Alkynylene-L a -, and combinations thereof, wherein each alkylene, alkenylene, and alkynylene is optionally substituted; and L a Each occurrence of may independently represent -O-, -S-, -N(R 7 )-, -C(O)-, -C(S)-, -C(O)O-, -OC(O)-, -OC(O)O-, -C(O)N(R 7 )-, -N(R 7 )C(O)-, -C(O)N(R 7 )C(O)-, -C(O)N(R 7 )C(O)N(R 7 ), -N(R 7 )C(O)N(R 7 )-, -N(R 7 )C(O)O-, -OC(O)N(R 7 )-, -C(NR 7 )-, -N(R 7 )C(NR 7 )-, -C(NR 7 )N(R 7 )-, -N(R 7 )C(NR 7 )N(R 7 )-, -S(O)2-, -OS(O)-, -S(O)O-, -S(O), -OS(O)2-, -S(O)2O, -N(R 7 )S(O)2-, -S(O)2N(R 7 )-, -N(R 7 )S(O)-, -S(O)N(R 7 )-, -N(R 7 )S(O)2N(R 7 )- and -N(R 7 )S(O)N(R 7 )-, and R 7 is independently, at each occurrence, hydrogen, —NH2, —C(O)OCH2C6H5; and C 1~10 Alkyl, C2~10 Alkenyl, C 2~10 Alkynyl, C 3~12 cycloalkyl, and 3- to 12-membered heterocycle, each of which is selected from halo, hydroxyl, cyano, nitro, amino, oxo, thioxo, —C(O)OCH2C6H5, —NHC(O)OCH2C6H5, C 1~10 Alkyl, C 1~10 Haloalkyl, C 1~10 Alkoxy, C 2~10 Alkenyl, C 2~10 Alkynyl, C 3~12 It is optionally substituted with one or more substituents independently selected from cycloalkyl, and 3- to 12-membered heterocyclyl.

[0160] In some embodiments, each L 1 , L 2 , or L 3 is alkyl, alkenyl, alkynyl, halo, hydroxyl, cyano, -OR 8 , -SR 8 , amino, aminyl, amido, cycloalkyl, aryl, heterocyclyl, heteroaryl, cycloclkylalkyl, arylalkyl, heterocyclylalkyl, heteroarylalkyl, -C(O)R 8 , -C(O)N(R 8 )2, -N(R 8 )C(O)R 8 , -C(O)OR 8 , -OC(O)R 8 , -S(O)R 8 , -S(O)2R 8 , -P(O)(OR 8 )2, -OP(O)(OR8)2, nitro, oxo, thioxo, =N(R 8 ), or cyano; and R 8 is hydrogen, alkyl, alkenyl, alkynyl, cycloalkyl, aryl, heterocyclyl, heteroaryl, cycloclkylalkyl, arylalkyl, heterocyclylalkyl, or heteroarylalkyl.

[0161] In some embodiments, L 1 , L 2 , or L 3 is, independently, [ka] wherein: R a is hydrogen or alkyl, L b each occurrence of is independently a direct bond, an optionally substituted alkylene linker, an optionally substituted heteroalkylene linker, a heteroatom linker, or a combination thereof; c each occurrence of is independently an optionally substituted alkylene linker, with the proviso that L 1 , L 2 , or L 3 At least one of the [ka] It has the following structure.

[0162] In some embodiments, L 1 and L 2 is, independently, [ka] wherein: R a is hydrogen or alkyl, L b is independently a direct bond, an optionally substituted alkylene linker, an optionally substituted heteroalkylene linker, a heteroatom linker, or a combination thereof; L c each occurrence of is independently an optionally substituted alkylene linker; However, L 1 or L 2 At least one of the [ka] It has the following structure.

[0163] In more embodiments, L 2 teeth, [ka] It has the following structure.

[0164] In some embodiments, L c is unsubstituted. In some embodiments, L c is C1-C6 alkylene. In some more embodiments, L c is C2-C4 alkylene. In some embodiments, L c is a straight chain C1-C6 alkylene. c is a straight chain unsubstituted C1-C6 alkylene. c is a straight-chain unsubstituted C2-C4 alkylene.

[0165] In more embodiments, L 1 , L 2 , and L 3 are each independently [ka] wherein: *but, [ka] shows a direct bond to a substitutable position on the phenyl group in the structure:

[0166] In some embodiments, L 1 or L 2 teeth, [ka] wherein: *but, [ka] shows a direct bond to a substitutable position on the phenyl group in the structure:

[0167] In some embodiments, L c is substituted with one or more substituents selected from the group consisting of halo, haloalkyl, alkoxy, cyano, nitro, carboxy, sulfonamido, sulfonic acid, or combinations thereof.

[0168] In some embodiments, the compound has the following structure (Ia) or (Ib): [ka] It has one of the following.

[0169] In some embodiments, the compound has the following structure (Ic') or (Ic"): [ka] It has one of the following.

[0170] In certain embodiments, X 2 , X 3 , or both are CH, or CF. In some embodiments, X 1 , X 5 , or both are CR 3 and R 3 is H or halo. In some embodiments, X 1 and X 5 But both are CR 3 and R 3 is H or halo. In some embodiments, X 1 is CR 3 and R 3 is halo. In certain embodiments, X 5 is CR 3 and R 3 is halo. In some embodiments, halo is fluoro. In some embodiments, X 1is CF. In some embodiments, X 1 is CH. In some embodiments, X 5 is CH. In some embodiments, X 5 is CF. In some embodiments, X 3 is CR 3 and R 3 is H. In some embodiments, X 3 is CR 3 and R 3 is halo (e.g., fluoro).

[0171] In some embodiments, the compound has the following structure (Ia-1), (Ia-2), (Ia-3), (Ia-4), (Ia-5), or (Ia-6): [ka] wherein: L b is independently a direct bond, an optionally substituted alkylene linker, an optionally substituted heteroalkylene linker, a heteroatom linker, or a combination thereof; q6 is 0, 1, or 2;

[0172] In some embodiments, the compound has the following structure: (Ia-1), (Ia-2), (Ia-3), (Ia-4), (Ia-5), (Ia-6), (Ia-7), or (Ia-8): [ka] wherein: L b is independently a direct bond, an optionally substituted alkylene linker, an optionally substituted heteroalkylene linker, a heteroatom linker, or a combination thereof; q6 is 0, 1, or 2.

[0173] In some embodiments, q6 is 1 and L bis gly-gly.

[0174] In certain embodiments, the compound has the following structure (Ic-1), (Ic-2), (Ic-3), or (Ic-4): [ka] wherein: L b is independently a direct bond, an optionally substituted alkylene linker, an optionally substituted heteroalkylene linker, a heteroatom linker, or a combination thereof; q7 is 1, 2, or 3.

[0175] In some embodiments, q7 is 2.

[0176] In some embodiments, the compound has the following structure (Id), (Ie), (If), or (Ig): [ka] wherein L b is independently a direct bond, an optionally substituted alkylene linker, an optionally substituted heteroalkylene linker, a heteroatom linker, or a combination thereof; q8 is 0, 1, or 2; q9 is 0, 1, or 2.

[0177] In some embodiments, q9 is 0 and q8 is 1.

[0178] In some embodiments, the compound has the following structure (Ih) or (Ii): [ka] wherein: L bEach occurrence of is independently a direct bond, an optionally substituted alkylene linker, an optionally substituted heteroalkylene linker, a heteroatom linker, or a combination thereof.

[0179] In some embodiments, L b is a direct bond, an optionally substituted alkylene linker, or an optionally substituted heteroalkylene linker.

[0180] In some embodiments, L b is a direct bond, or [ka] wherein: R b Each occurrence of is independently hydrogen, alkyl, hydroxyalkyl, or alkoxyalkyl.

[0181] In some embodiments, R b Each occurrence of is -CH3.

[0182] In some embodiments, L 1 or L 2 teeth, [ka] wherein: ** is X 1 , X 2 , X 3 , X 4 or X 5 The bond to

[0183] In certain embodiments, X 2 CL 1 -R 1 and X 3 CL 2 -R 2 In some embodiments, X 3 CL 1 -R 1 and X2 CL 2 -R 2 In some embodiments, X 1 , X 4 , and X 5 All are CR 3 In some embodiments, X 1 , X 4 , and X 5 are all CH.

[0184] In some embodiments, the compound is [ka] [ka] wherein: R 1b is a chemotherapeutic agent, cytotoxic agent, or myeloid cell agonist; R 2b but, [ka] and having one of the structures: L 1g but, [ka] [ka] The structure of the compound is:

[0185] In some embodiments, the compound is [ka] [ka] [ka] wherein: R 1bis a chemotherapeutic agent, cytotoxic agent, or myeloid cell agonist; R 2b but, [ka] and having one of the structures: L 1g but, [ka] [ka] The structure of the compound is:

[0186] In some embodiments, the compound is [ka] The structure of the compound is:

[0187] In some embodiments, the compound is [ka] [ka] The structure of the compound is:

[0188] In some embodiments, the compound is [ka] [ka] [ka] [ka] [ka] [ka] [ka] The structure of the compound is:

[0189] In some embodiments, the compound is [ka] and having one of the structures: This may be as a stereoisomer, enantiomer or tautomer thereof, or a mixture thereof, or as a pharmaceutically acceptable salt, solvate or prodrug thereof.

[0190] In some embodiments, the compound is [ka] [ka] [ka] [ka] [ka] [ka] [ka] [ka] as its stereoisomer, enantiomer or tautomer, or a mixture thereof, or a pharmaceutically acceptable salt, solvate or prodrug thereof.

[0191] In some embodiments, the payload is a chemotherapeutic agent, a cytotoxic agent, or a myeloid cell agonist.

[0192] In some embodiments, the payload is a chemotherapeutic agent selected from the group consisting of camptothecin, paclitaxel, doxorubicin, vinblastine, dacarbazine, irinotecan, topotecan, ciratecan, cositecan, exatecan, lurtotecan, SN-38, Dxd (i.e., CAS number 1599440-33-1), gimatecan, belotecan, and rubitecan.

[0193] In certain embodiments, the payload is a cytotoxic agent selected from the group consisting of calicheamicin, anthramycin, abeimicin, ticamycin, DC-81, mazethramycin, neothramycin A, neothramycin B, polothramycin prothracarcin, sivanomycin, sibiromycin, tomamycin, auristatin F, monomethyl auristatin F, auristatin E, monomethyl auristatin E, dolastatin, monomethyl dolastatin, mertansine, and emtansine.

[0194] In certain embodiments, the payload is a STING agonist, a ligand for TLR2, a ligand for TLR3, a ligand for TLR4, a ligand for TLR5, a ligand for TLR6, a ligand for TLR7, a ligand for TLR8, a ligand for TLR9, a ligand for TLR10, a ligand for a nucleotide oligomerization domain (NOD), a ligand for a RIG-I-like receptor (RLR), a ligand for a C-type lectin receptor (CLR), a ligand for a cytoplasmic DNA sensor (CDS), and an inflammasome. and a myeloid cell agonist selected from the group consisting of ligands of inducers, preferably the myeloid cell agonist is a myeloid cell agonist selected from the group consisting of sergantolimod, motolimod, resiquimod, 3M-051, 3M-052, MCT-465, IMO-4200, VTX-763, VTX-1463, RG7854, ADU-S100, MK-1454, MK-2118, BMS-986301, GSK3745417, SB-11285, and IMSA-101.

[0195] In some embodiments, the payload is an alkylating agent, an antimetabolite, a microtubule inhibitor, a topoisomerase inhibitor, a myeloid agonist, a glucocorticoid receptor agonist, or a cytotoxic antibiotic. In certain embodiments, the payload is a nitrogen mustard, a nitrosourea, a tetrazine, an aziridine, cisplatin or a cisplatin derivative, or a non-classical alkylating agent.

[0196] In certain embodiments, the payload is mechlorethamine, cyclophosphamide, melphalan, chlorambucil, ifosfamide, busulfan, N-nitroso-N-methylurea (MNU), carmustine (BCNU), lomustine (CCNU), semustine (MeCCNU), fotemustine, streptozotocin, dacarbazine, mitozolomide, temozolomide, thiotepa, mitomycin, diaziquone (AZQ), cisplatin, carboplatin, oxaliplatin, procarbazine, or hexamethylmelamine. In some embodiments, the payload is an antifolate, a fluoropyrimidine, a deoxynucleoside analog, or a thiopurine.

[0197] In certain embodiments, the payload is methotrexate, pemetrexed, fluorouracil, capecitabine, cytarabine, gemcitabine, decitabine, azacitidine, fludarabine, nelarabine, cladribine, clofarabine, pentostatin, thioguanine, and mercaptopurine. In some embodiments, the payload is an auristatin, a vinca alkaloid, or a taxane. In certain embodiments, the payload is auristatin F, auristatin E, vincristine, vinblastine, vinorelbine, vindesine, vinflunine, paclitaxel, docetaxel, etoposide, or teniposide. In further embodiments, the payload is a hydrocortisone (e.g., prednisone, fluocinolone), an acetonide (e.g., budesonide, fluocinonide), or a methasone type (e.g., dexamethasone, betamethasone, fluticasone).

[0198] In some embodiments, the payload has a molecular weight greater than 150 g / mol, greater than 200 g / mol, greater than 300 g / mol, greater than 400 g / mol, greater than 500 g / mol, greater than 600 g / mol, greater than 700 g / mol, greater than 800 g / mol, greater than 900 g / mol, or greater than 1000 g / mol. In some embodiments, the payload has a molecular weight less than 150 g / mol, less than 200 g / mol, less than 300 g / mol, less than 400 g / mol, less than 500 g / mol, less than 600 g / mol, less than 700 g / mol, less than 800 g / mol, less than 900 g / mol, or less than 1000 g / mol.

[0199] Payload moieties are known to those skilled in the art. For example, payloads can be found in PCT Publication Nos. 2021 / 207701, 2019 / 217591, 2018 / 089373, 2019 / 136487, and U.S. Patent No. 11,179,473, and the references cited therein, each of which is incorporated by reference in its entirety.

[0200] In some embodiments, the payload is a STING agonist having the following structure (III): [ka] During the ceremony, L 1 But X 1 a first end covalently bonded by a single bond to X 2 and a second end covalently attached by a single bond to L; 1 is C2-C8 alkylene, C2-C8 alkenylene, C2-C8 alkynylene, C2-C8 heteroalkylene, C2-C8 heteroalkenylene, C2-C8 heteroalkynylene, or -(L 1a ) x -L 1b -(L 1c ) y -, each of which is optionally substituted with one or more R; L1a and L 1c are each independently a C1-C6 alkylene, a C1-C6 alkenylene, a C2-C6 alkynylene, a C1-C6 heteroalkylene, a C2-C6 heteroalkenylene, or a C2-C6 heteroalkynylene, each of which is optionally substituted with one or more R; L 1b But C6~C 10 arylene, C-C cycloalkylene, 3- to 6-membered heterocyclene, or 5- to 6-membered heteroarylene, each of which is optionally substituted with one or more R; x and y are each independently 0 or 1; R, in each occurrence, independently represents R*, D, -OR a , -N(R a )2, -NR a C(=O)R b , -C(=O)N(R a )2, -CO2R a , -OC(=O)R b , -OC(=O)N(R a )2, -OC(=O)N(R a )2, -SR a , -S(O)2N(R a )2, -S(O)2R a , C1-C6 alkyl, C1-C6 alkoxy, C6-C 10 aryl, C3-C6 cycloalkyl, 3- to 6-membered heterocyclyl, or 5- to 6-membered heteroaryl, each of which is C1-C6 alkyl, C1-C6 alkoxy, C6-C 10 Aryl, C3-C6 cycloalkyl, 3- to 6-membered heterocyclyl, and 5- to 6-membered heteroaryl are independently selected from D, -OR a , -N(R a )2, -NR a C(=O)R b , -C(=O)N(R a )2, -CO2R a , -OC(=O)R b , -OC(=O)N(R a )2, -OC(=O)N(R a )2, -SR a , -S(=O)2N(R a)2 or -S(O)2R a and optionally substituted with one or more substituents selected from X 1 and X 2 each independently represents O or NR a and (i)W 1 and W 2 But independently, OR c or SR c and Y 1 and Y 2 are N, R 1 is H, CN, OH, Cl, Br, N3, C1-C6 alkyl or C1-C6 alkoxy, R 2 is H, CN, OH, F, Cl, Br, N3, C1-C6 alkyl or C1-C6 alkoxy, (ii)W 1 and W 2 But independently, OR c , S.R. c or BHR c and Y 1 and Y 2 at least one of Y is CH, CF or CCl; 1 and Y 2 the other is CH, CF, CCl or N; R 1 and R 2 are each independently H, CN, OH, F, Cl, Br, N3, C1-C6 alkyl, or C1-C6 alkoxy; (iii)W 1 and W 2 But independently, SR c or BHR c and Y 1 and Y 2 are N, R 1 is H, CN, Cl, Br, N3, C1-C6 alkyl or C1-C6 alkoxy, R 2is H, CN, OH, F, Cl, Br, N3, C1-C6 alkyl or C1-C6 alkoxy, R 3 and R 4 are each independently H or -N(R a )2, R a is, in each occurrence, independently H, C1-C6 alkyl, C2-C6 alkenyl, C1-C6 aminylalkyl, C2-C6 aminylalkenyl, C3-C6 cycloalkyl, C3-C6 aminylcycloalkyl, C3-C6 cycloalky-C1-C6 alkyl, C3-C6 aminylcycloalky-C1-C6 alkyl, 3- to 6-membered heterocyclyl, 3- to 6-membered aminylheterocyclyl, 3- to 6-membered heterocyclyl-C1-C6 alkyl or 3- to 6-membered aminylheterocyclyl-C1-C6 alkyl; and each substitutable R a is optionally substituted with R*; R b is, in each occurrence, independently C1-C6 alkyl, C2-C6 alkenyl, C1-C6 aminylalkyl, C2-C6 aminylalkenyl, C3-C6 cycloalkyl, C3-C6 aminylcycloalkyl, C3-C6 cycloalky-C1-C6 alkyl, C3-C6 aminylcycloalky-C1-C6 alkyl, 3- to 6-membered heterocyclyl, 3- to 6-membered aminylheterocyclyl, 3- to 6-membered heterocyclyl-C1-C6 alkyl or 3- to 6-membered aminylheterocyclyl-C1-C6 alkyl; and each R b is optionally substituted with R*; R c is, in each occurrence, independently H, C1-C6 alkyl, -CH2OC(=O)R d or -R e R f and each substitutable R c is optionally substituted with R*; R d is, in each occurrence, independently C1-C6 alkyl; and each R d is optionally substituted with R*; R e is, in each occurrence, independently absent, C1-C6 alkylene, or C1-C6 heteroalkylene, and each alkylene and heteroalkyl is selected from one or more R g and optionally substituted with R f In each occurrence, independently, H, C1~C 20 Alkyl C1-C 20 Heteroalkyl-OC(O)OC1~C 20 alkyl, cycloalkyl, heterocyclyl, aryl, or heteroaryl, each of which is selected from the group consisting of one or more R h and optionally substituted with R g In each occurrence, independently, R*, halo, -CN, C1~C 20 Alkyl, -OR i , oxo, cycloalkyl, heterocyclyl, aryl, or heteroaryl, each of which may be selected from the group consisting of one or more R j and optionally substituted with R h In each occurrence, independently, R*, C1~C 20 Alkyl, C1-C 20 Heteroalkyl, -C(=O)C1-C 20 Alkyl, -OC(=O)C1~C 20 Alkyl, -C(=O)OC1~C 20 Alkyl, -OC(=O)OC1~C 20 Alkyl, -C(=O)N(R k )C1~C 20 Alkyl, -N(R k )C(=O)C1~C 20 Alkyl, -OC(=O)N(R k )C1~C 20 Alkyl, -C(=O)C1~C 20 Heteroalkyl, -OC(=O)C1-C 20 Heteroalkyl, -C(=O)OC1~C 20Heteroalkyl, -OC(=O)OC1~C 20 Heteroalkyl, -C(=O)N(R k )C1~C 20 Heteroalkyl, -N(R k )C(=O)C1~C 20 Heteroalkyl, -OC(=O)N(R k )C1~C 20 Heteroalkyl, -Oaryl, -Oheteroaryl, -C(=O)aryl, -C(=O)heteroaryl, -OC(=O)aryl, -C(=O)Oaryl, -OC(=O)heteroaryl, -C(=O)Oheteroaryl, -C(=O)N(R k ) aryl, -C(=O)N(R k ) heteroaryl, -N(R k )C(=O)aryl, -N(R k )C(=O)heteroaryl, -OC(=O)N(R k ) aryl, -OC(=O)N(R k ) heteroaryl, -N(R k )C(=O)Oaryl, -N(R k )C(=O)O heteroaryl, -S(=O)2N(R k ) aryl, -S(=O)2N(R k ) heteroaryl or -N(R k )-*Peptide-L 2 where * is the C-terminus of the peptide, and L 2 However, it does not exist or -C1~C 12 Alkyl, -C1-C 12 Heteroalkyl, -C(=O)C1-C 12 Alkyl or -C(=O)C1-C 12 heteroalkyl, and each alkyl, heteroalkyl, aryl, and heteroaryl is selected from one or more R j and optionally substituted with R i In each occurrence, independently, H, C1~C 20 Alkyl, C1-C 20Heteroalkyl is a cycloalkyl, heterocyclyl, aryl, or heteroaryl, and each alkyl, cycloalkyl, heterocyclyl, aryl, and heteroaryl is selected from the group consisting of one or more R j and optionally substituted with R j In each occurrence, independently, R*, C1~C 20 Alkyl, -OC1~C 20 Alkyl, C1-C 20 heteroalkyl, halo, -CN, -OH, oxo, aryl, heteroaryl, -Oaryl or -Oheteroaryl; R k is, in each occurrence, independently H or C1-C 20 is alkyl, R* has a first end covalently bonded to the remainder of Structure (I) or Structure (II) by a single bond, and L 1 , X 1 , X 2 , R 3 , R 4 , W 1 or W 2 and a second end covalently attached by a single bond to at least one of

[0201] In other embodiments, the STING agonist has the following structure (IIIA): [ka] In yet another embodiment, the STING agonist is a compound having the following structure (IIIA'): [ka]

[0202] In other embodiments, the present disclosure provides a compound having activity as a STING agonist and having the following structure (IV): [ka] or a pharmaceutically acceptable salt, tautomer, prodrug, or stereoisomer thereof, wherein: L 1 But X 1 a first end covalently bonded by a single bond to X 2 and a second end covalently attached by a single bond to L; 1 is C2-C8 alkylene, C2-C8 alkenylene, C2-C8 alkynylene, C2-C8 heteroalkylene, C2-C8 heteroalkenylene, C2-C8 heteroalkynylene, or -(L 1a ) x -L 1b -(L 1c ) y -, each of which is optionally substituted with one or more R; L 1a and L 1c are each independently a C1-C6 alkylene, a C1-C6 alkenylene, a C2-C6 alkynylene, a C1-C6 heteroalkylene, a C2-C6 heteroalkenylene, or a C2-C6 heteroalkynylene, each of which is optionally substituted with one or more R; L 1b But C6~C 10 arylene, C-C cycloalkylene, 3- to 6-membered heterocyclene, or 5- to 6-membered heteroarylene, each of which is optionally substituted with one or more R; x and y are each independently 0 or 1; R, in each occurrence, independently represents R*, D, -OR a , -N(R a )2, -NR a C(=O)R b , -C(=O)N(R a )2, -CO2R a , -OC(=O)R b , -OC(=O)N(R a )2, -OC(=O)N(R a )2, -SR a , -S(O)2N(R a )2, -S(O)2R a , C1-C6 alkyl, C1-C6 alkoxy, C6-C 10aryl, C3-C6 cycloalkyl, 3- to 6-membered heterocyclyl, or 5- to 6-membered heteroaryl, each of which is C1-C6 alkyl, C1-C6 alkoxy, C6-C 10 Aryl, C3-C6 cycloalkyl, 3- to 6-membered heterocyclyl, and 5- to 6-membered heteroaryl are independently selected from D, -OR a , -N(R a )2, -NR a C(=O)R b , -C(=O)N(R a )2, -CO2R a , -OC(=O)R b , -OC(=O)N(R a )2, -OC(=O)N(R a )2, -SR a , -S(=O)2N(R a )2 or -S(O)2R a and optionally substituted with one or more substituents selected from W 1 and W 2 However, each independently, OR c , S.R. c or BHR c and X 1 and X 2 each independently represents O or NR a and (i)Y 1 and Y 2 are N, R 5 is H, CN, OH, F, Cl, Br, N3, C1-C6 alkyl or C1-C6 alkoxy, R 6 is H, CN, F, Cl, Br, N3, C1-C6 alkyl or C1-C6 alkoxy, or (ii) Y 1 and Y 2 at least one of Y is CH, CF or CCl; 1 and Y 2 the other is CH, CF, CCl or N; R 5 and R 6are each independently H, CN, OH, F, Cl, Br, N3, C1-C6 alkyl or C1-C6 alkoxy; R 3 and R 4 are each independently H or -N(R a )2, R a is, in each occurrence, independently H, C1-C6 alkyl, C2-C6 alkenyl, C1-C6 aminylalkyl, C2-C6 aminylalkenyl, C3-C6 cycloalkyl, C3-C6 aminylcycloalkyl, C3-C6 cycloalky-C1-C6 alkyl, C3-C6 aminylcycloalky-C1-C6 alkyl, 3- to 6-membered heterocyclyl, 3- to 6-membered aminylheterocyclyl, 3- to 6-membered heterocyclyl-C1-C6 alkyl or 3- to 6-membered aminylheterocyclyl-C1-C6 alkyl; and each substitutable R a is optionally substituted with R*; R b is, in each occurrence, independently C1-C6 alkyl, C2-C6 alkenyl, C1-C6 aminylalkyl, C2-C6 aminylalkenyl, C3-C6 cycloalkyl, C3-C6 aminylcycloalkyl, C3-C6 cycloalky-C1-C6 alkyl, C3-C6 aminylcycloalky-C1-C6 alkyl, 3- to 6-membered heterocyclyl, 3- to 6-membered aminylheterocyclyl, 3- to 6-membered heterocyclyl-C1-C6 alkyl or 3- to 6-membered aminylheterocyclyl-C1-C6 alkyl; and each R b is optionally substituted with R*; R c is, in each occurrence, independently H, C1-C6 alkyl, -CH2OC(=O)R d or -R e R f and each substitutable R c is optionally substituted with R*; R d is, in each occurrence, independently C1-C6 alkyl; and each R dis optionally substituted with R*; R e is, in each occurrence, independently absent, C1-C6 alkylene, or C1-C6 heteroalkylene, and each alkylene and heteroalkyl is selected from one or more R g and optionally substituted with R f In each occurrence, independently, H, C1~C 20 Alkyl C1-C 20 Heteroalkyl-OC(O)OC1~C 20 alkyl, cycloalkyl, heterocyclyl, aryl, or heteroaryl, each of which is selected from the group consisting of one or more R h and optionally substituted with R g In each occurrence, independently, R*, halo, -CN, C1~C 20 Alkyl, -OR i , oxo, cycloalkyl, heterocyclyl, aryl, or heteroaryl, each of which may be selected from the group consisting of one or more R j and optionally substituted with R h In each occurrence, independently, R*, C1~C 20 Alkyl, C1-C 20 Heteroalkyl, -C(=O)C1-C 20 Alkyl, -OC(=O)C1~C 20 Alkyl, -C(=O)OC1~C 20 Alkyl, -OC(=O)OC1~C 20 Alkyl, -C(=O)N(R k )C1~C 20 Alkyl, -N(R k )C(=O)C1~C 20 Alkyl, -OC(=O)N(R k )C1~C 20 Alkyl, -C(=O)C1~C 20 Heteroalkyl, -OC(=O)C1-C 20Heteroalkyl, -C(=O)OC1~C 20 Heteroalkyl, -OC(=O)OC1~C 20 Heteroalkyl, -C(=O)N(R k )C1~C 20 Heteroalkyl, -N(R k )C(=O)C1~C 20 Heteroalkyl, -OC(=O)N(R k )C1~C 20 Heteroalkyl, -Oaryl, -Oheteroaryl, -C(=O)aryl, -C(=O)heteroaryl, -OC(=O)aryl, -C(=O)Oaryl, -OC(=O)heteroaryl, -C(=O)Oheteroaryl, -C(=O)N(R k ) aryl, -C(=O)N(R k ) heteroaryl, -N(R k )C(=O)aryl, -N(R k )C(=O)heteroaryl, -OC(=O)N(R k ) aryl, -OC(=O)N(R k ) heteroaryl, -N(R k )C(=O)Oaryl, -N(R k )C(=O)O heteroaryl, -S(=O)2N(R k ) aryl, -S(=O)2N(R k ) heteroaryl or -N(R k )-*Peptide-L 2 where * is the C-terminus of the peptide, and L 2 However, it does not exist, and -C1~C 12 Alkyl, -C1-C 12 Heteroalkyl, -C(=O)C1-C 12 Alkyl or -C(=O)C1-C 12 heteroalkyl, and each alkyl, heteroalkyl, aryl, and heteroaryl is selected from one or more R j and optionally substituted by R i In each occurrence, independently, H, C1~C 20 Alkyl, C1-C 20Heteroalkyl is a cycloalkyl, heterocyclyl, aryl, or heteroaryl, and each alkyl, cycloalkyl, heterocyclyl, aryl, and heteroaryl is selected from the group consisting of one or more R j and optionally substituted with R j In each occurrence, independently, R*, C1~C 20 Alkyl, -OC1~C 20 Alkyl, C1-C 20 heteroalkyl, halo, -CN, -OH, oxo, aryl, heteroaryl, -Oaryl or -Oheteroaryl; R k is, in each occurrence, independently H or C1-C 20 is alkyl, R* has a first end covalently bonded to the remainder of Structure (I) or Structure (II) by a single bond, and L 1 , X 1 , X 2 , R 3 , R 4 , W 1 or W 2 and a second end covalently attached by a single bond to at least one of

[0203] In other embodiments, the STING agonist of the present disclosure has the following structure (IVA): [ka]

[0204] In other embodiments, the STING agonist has the following structure (IVA'): [ka]

[0205] In some embodiments, R 1a teeth, [ka] The structure of the compound is:

[0206] In some embodiments, R 1a teeth, [ka] [ka] The structure of the compound is:

[0207] In some embodiments, R 1a teeth, [ka] [ka] [ka] The structure of the compound is:

[0208] In some embodiments, the payload or R 1a teeth, [ka] wherein: R' is hydrogen, or [ka] wherein: R a ' is H or C 1~6 is alkyl, R b ', but C 1~6 Alkyl or C 1~6 is an alkoxy, R c 'But, H, C 1~6 Alkyl, -CHOH, or C 1~6 is an alkoxy, R d ' is H or C 1~6is alkyl, or R e ' is H or C 1~6 It is alkyl.

[0209] In some embodiments, the payload has the following structure: [ka]

[0210] In some embodiments, the payload has the following structure: [ka]

[0211] In some embodiments, the payload has the following structure: [ka]

[0212] In a particular embodiment, the payload has the following structure: [ka]

[0213] In some embodiments, the payload has the following structure: [ka]

[0214] In some embodiments, the payload has the following structure: [ka]

[0215] In some embodiments, the payload has the following structure: [ka]

[0216] In a particular embodiment, the payload has the following structure: [ka]

[0217] In some embodiments, the payload has the following structure: [ka]

[0218] In a particular embodiment, the payload has the following structure: [ka]

[0219] In some embodiments, the payload has the following structure: [ka]

[0220] In a particular embodiment, the payload has the following structure: [ka]

[0221] In a particular embodiment, the payload has the following structure: [ka]

[0222] In some embodiments, the payload has the following structure: [ka]

[0223] In a particular embodiment, the payload has the following structure: [ka]

[0224] In some embodiments, the payload has the following structure: [ka]

[0225] In a particular embodiment, the payload has the following structure: [ka]

[0226] Certain embodiments provide a compound having a structure selected from Table 1 (below), or a pharmaceutically acceptable salt, solvate, or prodrug thereof, as a stereoisomer, enantiomer, or tautomer thereof, or a mixture thereof. [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 Table 1-16 Table 1-17 Table 1-18 Table 1-19 Table 1-20 Table 1-21 Table 1-22 Table 1-23 Table 1-24 Table 1-25 [Table 1-26] [Table 1-27] [Table 1-28] [Table 1-29] [Table 1-30] [Table 1-31] [Table 1-32] [Table 1-33] [Table 1-34]

[0227] ‡ Characterization data for compounds I-35-55 and I-57-62 are as follows: Compound I-35 MS m / z[MH] - (ESI):1977.90; 1H NMR(300MHz,メタノール-d4)δ 0.79(d,J=7.1Hz,3H),0.89(s,3H),1.08(t,J=7.1Hz,2H),1.13-1.37( m,7H),1.47(s,4H),1.62-1.76(m,3H),1.73-1.89(m,1H),2.11(d,J=12 .7Hz,3H),2.24(t,J=7.2Hz,3H),2.31-2.45(m,2H),2.49-2.62(m,2H) ,2.78-3.16(m,4H),3.28(d,J=5.4Hz,2H),3.40-3.48(m,2H),3.48-3.5 8(m,36H),3.56-3.74(m,6H),3.79(d,J=11.7Hz,3H),3.98(s,2H),4.1 5-4.30(m,2H),4.30-4.45(m,2H),4.55-4.77(m,4H),4.95(d,J=4.3Hz, 1H),5.39(s,1H),5.51(s,1H),6.13-6.24(m,2H),6.88(s,1H),7.01(t ,J=8.9Hz,1H),7.07-7.25(m,7H),7.23-7.32(m,1H),7.32-7.49(m,2H)

[0228] Compound I-36 MS m / z[MH] - (ESI): 1834.50; 1H NMR(300MHz,メタノール-d4)δ:0.75-1.12(m,3H),1.20-1.43(m,1H),1 .51-1.68(m,4H),1.70-1.84(m,2H),1.85-2.06(m,2H),2.05-2.4 2(m,6H),2.45-2.61(m,4H),2.69(d,J=31.9Hz,1H),3.08(s,2H),3.41(t,J=5.7Hz,2H),3.53-3.59(m,2H),3.59-3.66(m,43H),3.74 (t,J=6.2Hz,2H),3.90(d,J=21.1Hz,4H),4.11(d,J=4.1Hz,2H),4.23(dd,J=7.9Hz,4.2Hz,1H),4.32(d,J=9.7Hz,1H),4.92-5.16(m ,4H),5.34-5.54(m,1H),5.60(s,1H),5.90-6.03(m,1H),6.26-6.63(m,3H),6.82-7.46(m,6H),7.56-7.66(m,1H),7.73-7.88(m,1H)

[0229] Compound I-37 MS m / z[MH] - (ESI): 1801.60; 1 H NMR(300MHz,メタノール-d4)δ:0.98(s,3H),1.27-1.34(m,6H),1.57(s,4H),1.80-1.91(m,4H),2.10-2.20(m,2H) ,2.37-2.39(m,4H),2.63-2.70(m,5H),3.13-3.20(m,2H),3.37-3.40(m,3H),3.52-3.59(m,6H),3.61-3.62(m ,37H),3.72-3.74(m,2H),3.92(s,3H),4.09(s,2H),4.17-4.23(m,3H),4.26-4.43(m,2H),4.66-4.75(m,4H) ,5.03-5.04(m,1H),5.50-5.58(m,2H),6.31-6.34(m,2H),6.97(s,2H),7.10-7.20(m,2H),7.32-7.47(m,6H).

[0230] Compound I-38 MS m / z[MH]- (ESI): 977.30; 1 H NMR(300MHz,メタノール-d4)0.99(s,3H),1.56-1.60(m,4H),1.76-1.78(m,2H),1.91(d,J=13. 4Hz,1H),2.31-2.34(m,3H),2.49-2.53(m,2H),2.84-2.93(m,3H),3.41-3.44(m,2H),3.80 (s,3H),4.02-4.04(m,2H),4.30-4.33(m,1H),4.86-4.99(m,1H),5.02-5.05(m,2H),5.50 -5.59(m,2H),6.30-6.34(m,3H),6.81-6.93(m,3H),7.01-7.21(m,3H),7.33-7.38(m,3H).

[0231] Compound I-39 MS m / z[MH] - (ESI): 949.10; 1 H NMR(300MHz,メタノール-d4)δ:0.99(s,3H),1.20-1.30(m,1H),1.57(s,4H),1.70-1.78(m,2 H),1.90-2.00(m,1H),2.15-2.35(m,3H),2.60-2.70(m,1H),3.60-3.66(m,2H),3.97-3. 99(m,3H),4.13-4.15(m,2H),4.30-4.35(m,1H),5.01-5.05(m,2H),5.50-5.59(m,2H),6 .30-6.34(m,3H),6.52-6.60(m,1H),7.16-7.34(m,6H),7.61(s,1H),7.89-7.92(m,1H).

[0232] Compound I-40 MS m / z[MH] - (ESI): 1121.35; 1H NMR(300MHz,メタノール-d4)δ:0.98(s,3H),1.48-1.62(m,4H),1.72-1.88(m,3H),2.12-2.27(m,1H) ,2.28-2.39(m,2H),2.46-2.79(m,3H),2.81-2.92(m,2H),2.93-3.03(m,1H),3.14-3.26(m,1H), 3.69-3.99(m,9H),4.21-4.54(m,3H),4.64-4.82(m,3H),4.99-5.09(m,1H),5.38-5.70(m,2H), 6.19-6.36(m,3H),6.43-6.59(m,1H),6.89-7.02(m,1H),7.03-7.16(m,2H),7.17-7.42(m,10H).

[0233] Compound I-41 MS m / z[MH]-(ESI):945.30;1H NMR(400MHz,メタノール-d4)δ:0.99(s,3H),1.27-1.29(m,3H),1.35-1.36(m,3H),1.56(s,4H),1.78-1.81(m,3 H),2.15-2.25(m,1H),2.30-2.34(m,2H),2.50-2.54(m,2H),2.60-2.70(m,1H),2.91-2.99(m,2H),3.81(s, 3H),4.21-4.28(m,3H),4.39-4.44(m,1H),4.68-4.77(m,3H),5.04-5.05(m,1H),5.50-5.58(m,2H),6.30-6 .33(m,2H),6.79-6.81(m,1H),6.89-6.94(m,3H),7.11-7.18(m,1H),7.20-7.29(m,2H),7.31-7.39(m,3H).

[0234] Compound I-42 MS m / z[M / 2-H] - (ESI): 1050.05; 1H NMR(300MHz,メタノール-d4)δ:0.71-0.93(m,4H),1.08(t,J=7.1Hz,4H),1.17-1.31(m,1H),1.47(s,4H),1.67(s,3H),1. 76-1.95(m,2H),2.00-2.17(m,2H),2.19-2.33(m,4H),2.32-2.55(m,6H),3.04(t,J=7.3Hz,2H),3.25-3.39(m,6H),3 .47-3.57(m,42H),3.59-3.72(m,4H),3.82(s,2H),4.00(s,2H),4.15-4.37(m,4H),4.63(s,2H),4.76-5.07(m,6H), 5.41(q,J=23.0,21.6Hz,3H),6.24(d,J=19.0Hz,3H),6.80(s,2H),6.88-7.09(m,3H),7.11-7.39(m,7H),8.09(s,1H)

[0235] Compound I-43 MS m / z[MH] - (ESI): 1105.40; 1 H NMR(300MHz,メタノール-d4)δ:0.88-0.98(m,6H),0.99-1.02(m,1H),1. 03-1.19(m,1H),1.27(s,1H),1.39-1.44(m,2H),1.47(s,3H),1.51- 1.64(m,4H),1.66-1.73(m,1H),1.81-1.89(m,3H),2.02-2.33(m,4H),2.31-2.39(m,1H),2.46-2.53(m,2H),2.58-2.69(m,3H),2.91-3. 09(m,3H),3.11-3.19(m,1H),3.21-3.29(m,1H),3.71-2.91(m,6H),3.92-4.11(m,2H),4.39-4.41(m,1H),4.48-4.58(m,2H),4.61-4.6 6(m,1H),4.69-4.79(m,2H),5.99(s,1H),6.12-6.28(m,1H),6.76-6.88(m,1H),6.89-6.99(m,3H),7.11-7.31(m,6H),7.32-7.44(m,1H) Compound I-44 [MH]- (ESI): 1193.40. 1 H NMR(300MHz,メタノール-d4)δ 0.98(s,3H),1.57(s,4H),1.64-1.90(m,3H),1.99-2.23(m,3H),2.24-2.4(m,2H),2.42-2.82(m,5H),2.86 -3.08(m,3H),3.20(dd,J=13.9Hz,5.8Hz,1H),3.68-3.95(m,6H),4.02(t,J=6.1Hz,2H),4.24-4.38(m,1H) ,4.39-4.52(m,2H),4.62(d,J=28.1Hz,1H),4.70-4.81(m,2H),5.04(d,J=4.4Hz,1H),5.37-5.71(m,2H),6 .16-6.35(m,2H),6.81(dd,J=8.0Hz,1.9Hz,1H),6.88-6.98(m,3H),7.03-7.17(m,1H),7.14-7.44(m,10H).

[0236] Compound I-45 MS m / z[MH] - (ESI): 1816.80; 1H NMR(400MHz,メタノール-d4)δ:0.79-0.88(m,6H),0.89-0.92(m,1H),0.94-1.04 (m,1H),1.13-1.24(m,5H),1.26-1.34(m,3H),1.37(s,3H),1.42-1.51(m,4H ),1.58-1.62(m,1H),1.69-1.71(m,2H),1.91-2.19(m,5H),2.21-2.29(m,1 H),2.46-2.61(m,4H),2.79-2.88(m,15H),2.89-2.99(m,13H),3.11-3.18(m ,1H),3.66-3.81(m,6H),3.91-4.01(m,4H),4.02-4.18(m,7H),4.21-4.24( m,3H),4.26-4.32(m,5H),4.39-4.48(m,2H),4.49-4.58(m,3H),4.61-4.68( m,3H),4.71-4.76(m,2H),5.89(s,1H),5.99-6.22(m,1H),6.61-6.78(m,1H) ,6.79-6.98(m,3H),7.01-7.18(m,2H),7.19-7.24(m,4H),7.26-7.38(m,1H)

[0237] Compound I-46 [MH] - (ESI): 1904.75; 1 H NMR(300MHz,メタノール-d4)δ 0.98(s,4H),1.29(s,1H),1.56(s,4H),1.79(d,J=13.8Hz,3H),2.01-2.24(m, 4H),2.33(s,2H),2.52-2.78(m,5H),2.81-3.24(m,33H),3.61-3.90(m,6H),3. 90-4.23(m,12H),4.24-4.53(m,11H),4.66-4.81(m,3H),5.03(d,J=4.4Hz,1H ),5.60(s,2H),6.28(d,J=11.7Hz,2H),6.74-7.16(m,5H),7.16-7.42(m,10H).

[0238] Compound I-47 MS m / z[M+H] + (ESI): 1019.38

[0239] Compound I-48 MS m / z[M+H] + (ESI): 1728.05; 1 H NMR(400MHz,メタノール-d4) δ:0.86-0.90(m,1H),1.02(s,3H),1.22-1.40(m,6H),1.56(s,3H),1.56-1.70(m,1H),1.75-1 .9(m,3H),2.05-2.40(m,3H),2.25-2.45(m,2H),2.50-2.61(m,2H),2.62-2.75(m,2H),2.90- 3.15(m,33H),3.95-4.55(m,24H),4.61-4.82(m,3H),5.05-5.11(m,1H),5.40-5.70(m,2H),6 .25-6.40(m,2H),6.70-6.82(m,1H),6.85-7.00(m,3H),7.05-7.28(m,3H),7.31-7.5(m,3H).

[0240] Compound I-49 MS m / z[M+H] + (ESI): 2008.9; 11H NMR (400 MHz, DMSO-d6) δ 9.97 - 9.80 (m, 1H), 8.70 - 8.62 (m, 1H), 8.27 - 8.16 (m, 2H), 7.92 - 7.85 (m, 2H), 7.64 - 7.16 (m, 12H), 7.14 - 7.08 (m, 2H), 6.29 (dd, J = 1.6, 10.4 Hz, 1H), 6.12 (s, 1H), 5.72 - 5.47 (m, 2H), 5.05 - 4.89 (m, 3H), 4.86 - 4.57 (m, 3H), 4.38 - 4.24 (m, 2H), 4.19 - 4.17 (m, 2H), 4.12 (t, J = 7.6 Hz, 1H), 3.92 - 3.86 (m, 2H), 3.74 - 3.69 (m, 2H), 3.60 - 3.54 (m, 3H), 3.49 - 3.44 (m, 44H), 3.38 (t, J = 5.6 Hz, 3H), 3.31 - 3.27 (m, 1H), 3.22 - 3.19 (m, 2H), 3.01 - 2.91 (m, 2H), 2.38 - 2.26 (m, 4H), 2.20 - 2.09 (m, 1H), 1.99 - 1.87 (m, 3H), 1.78 - 1.64 (m, 4H), 1.47 (s, 3H), 1.27 (d, J = 7.2 Hz, 3H), 1.06 (t, J = 6.8 Hz, 3H), 0.87 - 0.74 (m, 9H).

[0241] Compound I-50 MS m / z [M-H] - (ESI): 1670.0; 1 1H NMR (400 MHz, DMSO-d6) δ 8.75 - 8.57 (m, 1H), 8.39 - 7.95 (m, 1H), 7.50 - 7.19 (m, 6H), 7.18 - 6.97 (m, 2H), 6.28 (d, J = 10.4 Hz, 1H), 6.12 (s, 1H), 5.75 - 5.50 (m, 2H), 4.93 (d, J = 4.0 Hz, 1H), 4.74 - 4.50 (m, 3H), 4.43 - 3.86 (m, 23H), 3.05 - 2.59 (m, 32H), 2.60 - 2.54 (m, 1H), 2.43 (s, 2H), 2.35 - 2.27 (m, 1H), 2.23 - 2.10 (m, 1H), 2.04 - 1.91 (m, 1H), 1.80 - 1.61 (m, 3H), 1.58 - 1.38 (m, 4H), 1.29 - 1.09 (m, 6H), 0.85 (s, 3H)

[0242] Compound I-51 MS m / z[M+H] + (ESI): 993.4; 1 H NMR(DMSO-d6,400MHz)δ 7.73(d,J=1.6Hz,1H),7.46-7.38(m,3H),7.31-7.19(m,4H),7.12(s,2H),6.27(dd,J=1.6,10.0Hz,1H),6.12(s,1 H),5.74-5.52(m,2H),5.05(dd,J=9.6,18.8Hz,1H),4.94(d,J=4.0Hz,1H),4.74(dd,J=8.4,18.8Hz,1H),4.19(d, J=8.8Hz,1H),3.91(q,J=6.0Hz,2H),3.25(t,J=5.6Hz,2H),3.13(t,J=8.0Hz,2H),2.45-2.38(m,2H),2.31-2.26( m,1H),2.20-2.12(m,1H),2.02-1.92(m,1H),1.82(d,J=14.0Hz,1H),1.77-1.66(m,2H),1.47(s,4H),0.88(s,3H)

[0243] Compound I-52 MS m / z[MH] - (ESI): 1701.5896; 1 H NMR(DMSO-d6,400MHz)δ 7.46-7.38(m,2H),7.31-7.14(m,6H),7.08-7.01(m,2H),6.26(d,J=9.6Hz,1H),6.13 (s,1H),5.70-5.48(m,2H),5.07-4.92(m,2H),4.73(dd,J=8.8Hz,18.0Hz,1H),4.32-3 .91(m,25H),3.00-2.74(m,34H),2.43(t,J=7.6Hz,2H),2.25-2.15(m,1H),2.04(d,J =14.0Hz,1H),1.89(d,J=14.0Hz,1H),1.72(d,J=8.0Hz,2H),1.50(s,4H),0.92(s,3H)

[0244] Compound I-53 MS m / z[MH] - (ESI):1846.1; 11H NMR (400 MHz, DMSO-d6) δ 8.58 (s, 1H), 8.39 - 8.28 (m, 1H), 8.21 - 8.03 (m, 2H), 8.20 - 8.03 (m, 2H), 7.33 - 7.07 (m, 11H), 6.25 (d, J = 9.6 Hz, 1H), 6.11 (s, 1H), 5.75 - 5.53 (m, 2H), 4.94 (d, J = 4.8 Hz, 1H), 4.72 - 4.56 (m, 3H), 4.50 - 3.85 (m, 22H), 3.78 - 3.65 (m, 6H), 3.11 - 2.68 (m, 30H), 2.54 - 2.52 (m, 1H), 2.27 - 2.25 (m, 1H), 2.23 - 2.10 (m, 2H), 1.99 (d, J = 12.8 Hz, 1H), 1.82 - 1.63 (m, 3H), 1.48 (s, 4H), 0.86 (s, 3H).

[0245] Compound I-54 MS m / z [M-H] - (ESI): 1678.1; 1 1H NMR (400 MHz, DMSO-d6) δ 7.31 (d, J = 10.0 Hz, 1H), 7.22 - 7.13 (m, 1H), 7.05 (s, 2H), 6.85 (d, J = 10.4 Hz, 1H), 6.78 - 6.70 (m, 1H), 6.21 - 6.11 (m, 1H), 6.05 - 5.96 (m, 1H), 5.92 (s, 1H), 5.32 - 5.22 (m, 1H), 4.91 (d, J = 6.8 Hz, 1H), 4.71 (d, J = 3.6 Hz, 1H), 4.65 - 4.40 (m, 3H), 4.38 - 3.96 (m, 24H), 3.01 - 2.66 (m, 36H), 2.45 - 2.26 (m, 4H), 2.09 - 1.90 (m, 6H), 1.82 - 1.64 (m, 6H), 1.61 - 1.47 (m, 3H), 1.35 (s, 3H), 1.27 - 1.11 (m, 7H), 1.05 - 0.88 (m, 2H), 0.79 (s, 3H).

[0246] Compound I-55 MS m / z [M+Na] + (ESI): 1665.8; 11H NMR (400 MHz, DMSO-d6) δ 8.72 - 8.59 (m, 1H), 7.45 - 7.35 (m, 1H), 7.34 - 7.10 (m, 7H), 7.02 (s, 2H), 6.31 - 6.23 (m, 1H), 6.13 (s, 1H), 5.69 - 5.60 (m, 1H), 5.57 - 5.52 (m, 1H), 4.89 (s, 1H), 4.63 - 4.50 (m, 4H), 4.32 - 4.15 (m, 12H), 2.98 - 2.67 (m, 35H), 2.44 - 2.28 (m, 5H), 2.18 - 2.05 (m, 1H), 1.73 - 1.63 (m, 3H), 1.46 - 1.43 (m, 4H), 0.82 (s, 3H)

[0247] Compound I-57 MS m / z [M+H] + (ESI): 1969.9; 1 1H NMR (400 MHz, DMSO-d6) δ 8.16 (s, 1H), 7.52 - 6.93 (m, 11H), 6.29 (dd, J = 1.6, 10.0 Hz, 1H), 6.12 (s, 1H), 5.75 - 5.47 (m, 2H), 5.09 - 4.90 (m, 3H), 4.89 - 4.59 (m, 4H), 4.44 - 3.77 (m, 22H), 3.39 - 3.25 (m, 7H), 3.03 - 2.63 (m, 31H), 2.62 - 2.53 (m, 4H), 2.44 - 2.26 (m, 4H), 2.23 - 2.12 (m, 1H), 2.02 - 1.97 (m, 1H), 1.77 - 1.65 (m, 3H), 1.52 - 1.45 (m, 4H), 1.15 - 1.02 (m, 3H), 0.86 (s, 3H)

[0248] Compound I-58 MS m / z [M+H] + (ESI): 1672.0; 11H NMR (400 MHz, DMSO-d6) δ 8.76 - 8.59 (m, 1H), 7.33 - 7.24 (m, 1H), 7.22 - 7.15 (m, 1H), 7.11 (s, 2H), 6.88 (brd, J = 8.8 Hz, 1H), 6.76 (brdd, J = 1.6, 7.6 Hz, 1H), 6.22 (dd, J = 1.6, 10.0 Hz, 1H), 6.01 (s, 1H), 4.74 (brs, 1H), 4.69 - 4.42 (m, 4H), 4.38 - 4.14 (m, 13H), 4.13 - 3.86 (m, 12H), 3.06 - 2.67 (m, 32H), 2.61 (dt, J = 5.2, 13.2 Hz, 1H), 2.47 - 2.37 (m, 3H), 2.36 - 2.21 (m, 2H), 2.07 - 1.64 (m, 13H), 1.55 (brd, J = 8.4 Hz, 2H), 1.46 (s, 3H), 1.36 - 1.25 (m, 1H), 1.25 - 1.19 (m, 3H), 1.19 - 1.12 (m, 3H), 0.81 (s, 3H).

[0249] Compound I-59 MS m / z [M+Na] + (ESI): 1675.9; 1 1H NMR (400 MHz, DMSO-d6) δ 8.75 - 8.57 (m, 1H), 7.30 (brd, J = 10.0 Hz, 1H), 7.24 - 7.14 (m, 1H), 7.08 (s, 2H), 6.87 (brd, J = 9.2 Hz, 1H), 6.75 (brdd, J = 1.6, 7.6 Hz, 1H), 6.20 - 6.10 (m, 1H), 5.91 (s, 1H), 4.72 (brd, J = 3.6 Hz, 1H), 4.67 - 4.39 (m, 4H), 4.37 - 4.14 (m, 13H), 4.12 - 3.82 (m, 12H), 3.06 - 2.62 (m, 33H), 2.47 - 2.37 (m, 3H), 2.34 - 2.19 (m, 2H), 2.13 - 1.66 (m, 13H), 1.65 - 1.43 (m, 3H), 1.35 (s, 3H), 1.25 - 1.18 (m, 3H), 1.18 - 1.10 (m, 3H), 1.01 - 0.82 (m, 2H), 0.79 (s, 3H).

[0250] Compound I-60 MS m / z [M+Na] +(ESI): 1722.0; 1 1H NMR (400 MHz, DMSO-d6) δ 8.74 - 8.60 (m, 1H), 7.57 (s, 1H), 7.32 (d, J = 9.6 Hz, 1H), 7.18 (dd, J = 3.2, 10.4 Hz, 1H), 7.12 (s, 2H), 6.97 - 6.86 (m, 2H), 6.80 - 6.71 (m, 1H), 6.21 - 6.14 (m, 1H), 5.96 (d, J = 18.8 Hz, 2H), 5.73 (s, 1H), 4.87 (d, J = 1.6 Hz, 1H), 4.71 - 4.50 (m, 4H), 4.31 (s, 6H), 4.22 (d, J = 14.4 Hz, 9H), 2.94 - 2.75 (m, 35H), 2.43 - 2.38 (m, 3H), 2.34 - 2.29 (m, 2H), 2.06 (s, 5H), 1.98 - 1.88 (m, 3H), 1.82 - 1.68 (m, 4H), 1.65 (s, 3H), 1.38 (s, 4H), 1.24 - 1.11 (m, 9H), 0.85 (s, 4H).

[0251] Compound I-61 MS m / z [M-H] - (ESI): 1724.1; 1 1H NMR (400 MHz, DMSO-d6) δ 7.51 - 7.43 (m, 1H), 7.30 (d, J = 10.0 Hz, 1H), 7.21 - 7.12 (m, 1H), 7.06 (s, 2H), 6.95 (d, J = 6.0 Hz, 1H), 6.90 (d, J = 16.0 Hz, 1H), 6.84 (d, J = 10.0 Hz, 1H), 6.76 - 6.65 (m, 1H), 6.14 (d, J = 9.6 Hz, 1H), 5.91 (s, 1H), 5.87 - 5.76 (m, 1H), 5.12 (d, J = 6.4 Hz, 1H), 4.84 - 4.72 (m, 1H), 4.64 - 4.48 (m, 3H), 4.31 - 4.24 (m, 5H), 4.24 - 4.13 (m, 9H), 4.11 - 3.92 (m, 11H), 2.97 - 2.71 (m, 34H), 2.44 - 2.36 (m, 2H), 2.34 - 2.23 (m, 2H), 2.11 - 1.86 (m, 5H), 1.72 (s, 2H), 1.64 - 1.52 (m, 3H), 1.35 (s, 3H), 1.22 - 1.13 (m, 6H), 1.04 - 0.91 (m, 2H), 0.80 (s, 3H).

[0252] Compound I-62 MS m / z[MH] - (ESI):1696.1; 1 H NMR(400MHz,DMSO-d6)δ 7.33-7.27(m,1H),7.24-7.17(m,1H),7.10(s,2H),6.88(d,J=9.6Hz,1H),6.83-6.72(m,1H),6.28-6.20(m ,1H),6.09-6.00(m,2H),5.27-5.17(m,1H),4.99-4.89(m,1H),4.82-4.74(m,1H),4.67-4.44(m,3H),4.32( s,4H),4.28-4.15(m,10H),4.14-3.90(m,12H),3.03-2.72(m,34H),2.6-2.58(m,1H),2.45-2.29(m,4H),2 .07-1.89(m,6H),1.88-1.51(m,9H),1.50-1.46(m,3H),1.41-1.29(m,1H),1.27-1.15(m,6H),0.82(s,3H).

[0253] Conjugates The compounds of the present disclosure (e.g., compounds of structure (I)) are useful, in part, because they can be attached to a targeting moiety or targeting fragment thereof (e.g., an antibody). Such attachment may be achieved by reducing the disulfide bonds of the protein with a suitable reagent (e.g., TCEP) and coupling with a compound of structure (I) under appropriate conditions.

[0254] Thus, additional embodiments include the following structure (II): [ka] A conjugate having the formula: A is a targeting moiety or a binding fragment thereof; L 4 but, [ka] and having one of the structures: *** indicates the connection point to A, g is an integer from 1 to 20; X 1 , X 2 , X 3 , X 4 and X 5 One of them is CL 1 -R 1 and X 1 , X 2 , X 3 , X 4 and X 5 Another one of them is CL 2 -R 2 and X 1 , X 2 , X 3 , X 4 and X 5 The remaining three of these are independently N, CR 3 , or CL 3 -R 3a and R 1 , R 2 , and R 3a each independently comprises one or more moieties selected from an amino acid element, a charged element, a heteroalkylene element, a hydrophilic element, a trigger element, a sacrificial element, a polar cap, a payload, and combinations thereof; However, R 1 and R 2 at least one of which contains a payload; R 3each occurrence independently represents hydrogen, deuterium, alkyl, haloalkyl, halo, alkoxy, haloalkoxy, amino, aminyl, amidyl, aldehyde, hydroxyl, cyano, nitro, thiol, carboxy, carboxyalkyl, alkyl-S(O)3H, alkyl-OP(O)3H, alkyl-P(O)3H, -O-carboxyalkyl, -O-alkyl-S(O)3H, -O-alkyl-OP(O)3H, -O-alkyl-P selected from the group consisting of (O)H, -S(O)H, -OP(O)H, -P(O)H, alkyl-OP(O)-alkyl, alkyl-P(O)-alkyl, -O-alkyl-S(O)-alkyl, -O-alkyl-OP(O)-alkyl, -O-alkyl-P(O)-alkyl, -S(O)-alkyl, -OP(O)-alkyl, -P(O)-alkyl, sulfamide, sulfinimide, and carbohydrate; R 4a is hydrogen, deuterium, halo, or -SR 4c and R 4c Substituted or unsubstituted C6-C 10 aryl or substituted or unsubstituted 5- to 12-membered heteroaryl; L 1 , L 2 , and L 3 is each independently a linker comprising an optionally substituted alkylene, optionally substituted alkenylene, optionally substituted alkynylene, optionally substituted heteroalkylene, optionally substituted heteroalkenylene, optionally substituted heteroalkynylene, a heteroatom linker, an optionally substituted cycloalkylene, an optionally substituted arylene, an optionally substituted heterocyclylene, an optionally substituted heteroarylene, or a combination thereof, or a conjugate thereof as a stereoisomer, enantiomer, or tautomer, or a mixture thereof, or a pharmaceutically acceptable salt, solvate, or prodrug thereof.

[0255] Additional embodiments include the following structure (II): [ka] A conjugate having the formula: A is a targeting moiety or a binding fragment thereof; L 4 but, [ka] and having one of the structures: *** indicates the connection point to A, g is an integer from 1 to 20; X 1 , X 2 , X 3 , X 4 and X 5 One of them is CL 1 -R 1 and X 1 , X 2 , X 3 , X 4 and X 5 Another one of them is CL 2 -R 2 and X 1 , X 2 , X 3 , X 4 and X 5 The remaining three of these are independently N, CR 3 , or CL 3 -R 3a and R 1 , R 2 , and R 3a each independently comprises one or more moieties selected from an amino acid element, a charged element, a heteroalkylene element, a hydrophilic element, a trigger element, a sacrificial element, a polar cap, a payload, and combinations thereof; However, R 1 and R 2 at least one of which contains a payload; R 3each occurrence independently represents hydrogen, deuterium, alkyl, haloalkyl, halo, alkoxy, haloalkoxy, amino, aminyl, amidyl, aldehyde, hydroxyl, cyano, nitro, thiol, carboxy, carboxyalkyl, alkyl-S(O)3H, alkyl-OP(O)3H, alkyl-P(O)3H, -O-carboxyalkyl, -O-alkyl-S(O)3H, -O-alkyl-OP(O)3H, -O-alkyl-P selected from the group consisting of (O)H, -S(O)H, -OP(O)H, -P(O)H, alkyl-OP(O)-alkyl, alkyl-P(O)-alkyl, -O-alkyl-S(O)-alkyl, -O-alkyl-OP(O)-alkyl, -O-alkyl-P(O)-alkyl, -S(O)-alkyl, -OP(O)-alkyl, -P(O)-alkyl, sulfamide, sulfinimide, and carbohydrate; R 4a is hydrogen, deuterium, halo, or -SR 4c and R 4c Substituted or unsubstituted C6-C 10 aryl or substituted or unsubstituted 5- to 12-membered heteroaryl; L 1 , L 2 , and L 3 is each independently a direct bond or a linker comprising an optionally substituted alkylene, optionally substituted alkenylene, optionally substituted alkynylene, optionally substituted heteroalkylene, optionally substituted heteroalkenylene, optionally substituted heteroalkynylene, a heteroatom linker, an optionally substituted cycloalkylene, optionally substituted arylene, optionally substituted heterocyclylene, optionally substituted heteroarylene, or a combination thereof, or a pharmaceutically acceptable salt, solvate, or prodrug thereof.

[0256] In some embodiments, R 1 teeth, [ka] wherein: L 1a is an amino acid element, L 1b is the charging element, L 1c is a heteroalkylene element, L 1d is the hydrophilic element, L 1e is the trigger element, L 1f But it is a sacrificial element, L 1a , L 1b , L 1c , L 1d , L 1e , and L 1f One or more occurrences of, optionally, L 1a , L 1b , L 1c , L 1d , L 1e , and L 1f to form one or more rings, each occurrence of n1, n2, n3, n4, n5, and n6 is independently an integer from 0 to 3, provided that n1+n2+n3+n4+n5+n6=1 or greater; n7 is 1, 2, 3, 4, 5, or 6; R 1a But, L 1a , L 1b , L 1c , L 1d , L 1e , or L 1f wherein the payload is optionally substituted with a polar cap.

[0257] In certain embodiments, R 1 teeth, [ka] wherein: L 1a is an amino acid element, L 1b is the charging element, L 1c is a heteroalkylene element, L 1d is the hydrophilic element, L 1e is the trigger element, L 1f But it is a sacrificial element, L 1a , L 1b , L 1c , L 1d , L 1e , and L 1f One or more occurrences of, optionally, L 1a , L 1b , L 1c , L 1d , L 1e , and L 1f to form one or more rings, each occurrence of n1, n2, n3, n4, n5, and n6 is independently an integer from 0 to 3, provided that n1+n2+n3+n4+n5+n6=1 or greater; n7 is 1, 2, 3, 4, 5, or 6; R 1a is the payload optionally substituted with a polar cap.

[0258] In some embodiments, R 2 teeth, [ka] wherein: L 2a is an amino acid element, L 2b is the charging element, L 2c is a heteroalkylene element, L 2d is the hydrophilic element, L 2e is the trigger element, each occurrence of m1, m2, m3, m4, and m5 is independently an integer from 0 to 3, provided that m1+m2+m3+m4+m5=1 or greater; m6 is 1, 2, or 3; R 2a is hydrogen, alkyl, payload, or polar cap.

[0259] In certain embodiments, R 3a teeth, [ka] wherein: L 3a is an amino acid element, L 3b is the charging element, L 3c is a heteroalkylene element, L 3d is the hydrophilic element, L 3e is the trigger element, each occurrence of p1, p2, p3, p4, and p5 is independently an integer from 0 to 3, provided that p1+p2+p3+p4+p5=1 or greater; p6 is 1, 2, or 3; R 3b is hydrogen, alkyl, or a polar cap.

[0260] In some embodiments, n7 is 1, and n1-n6 are each 1. In particular embodiments, n7 is 1, n1-n4 are each 0, n5 is 1, and n6 is 1. In some embodiments, n7 is 1, n1 is 0, n2 is 0, n3 is 1, n4 is 0, n5 is 1, and n6 is 1.

[0261] In some embodiments, m6 is 1, and m1-m5 are each 1. In particular embodiments, m6 is 1, m1 is 1, m2 is 0, m3 is 0, m4 is 1, and m5 is 0.

[0262] In certain embodiments, p6 is 1 and p1-p5 are each 1. In some embodiments, p6 is 1, p1 is 1, p2 is 0, p3 is 0, p4 is 1, and p5 is 0.

[0263] In some embodiments, the amino acid element comprises one or more amino acids selected from the group consisting of glycine, alanine, serine, threonine, cysteine, valine, leucine, isoleucine, methionine, proline, phenylalanine, tyrosine, tryptophan, aspartic acid, glutamic acid, asparagine, glutamine, histidine, lysine, arginine, sarcosine, and beta-alanine. In more embodiments, the amino acid element comprises one or more amino acids selected from the group consisting of glycine, sarcosine, beta-alanine, and glutamic acid.

[0264] In some embodiments, the amino acid element comprises a dipeptide, a tripeptide, a tetrapeptide, or a pentapeptide. [ka] wherein: R 5a Each occurrence of is independently hydrogen, alkyl, hydroxyalkyl, or alkoxyalkyl.

[0265] In some embodiments, the charged element comprises a moiety that has a negative charge at pH 7.4 (i.e., in the range of 6.3 to 8.5). In some embodiments, the charged element comprises a moiety that has a positive charge at pH 7.4 (i.e., in the range of 6.3 to 8.5).

[0266] In some embodiments, the charged element comprises one or more charged amino acids, one or more carboxylic acids, one or more sulfonic acids, one or more sulfonamides, one or more sulfates, one or more phosphates, one or more quaternary amines, one or more sulfamides, one or more sulfinimides, or a combination thereof.

[0267] In some embodiments, the charged amino acid is aspartic acid, glutamic acid, histidine, lysine, or arginine.

[0268] In some embodiments, the heteroalkylene element comprises polyethylene glycol or polypropylene glycol. In some embodiments, the heteroalkylene element comprises [ka] wherein: R 5b , R 5c R 5d , and R 5e is independently selected from the group consisting of hydrogen, deuterium, alkyl, haloalkyl, halo, alkoxy, haloalkoxy, amino, hydroxyl, cyano, nitro, thiol, carboxyalkyl, alkyl-S(O)H, alkyl-OP(O)H, alkyl-P(O)H, -O-carboxyalkyl, -O-alkyl-S(O)H, -O-alkyl-OP(O)H, -O-alkyl-P(O)H, -S(O)H, -OP(O)H, and -P(O)H; Each occurrence of q1 is independently an integer from 1 to 24.

[0269] In some embodiments, q1 is 8, 10, 12, or 14.

[0270] In some embodiments, R 1 , R 2 , or R 3a comprises a non-cleavable linker.

[0271] In more embodiments, R 1 , R 2 , or R 3a teeth, [ka] wherein: R5b , R 5c R 5d , and R 5e is independently selected from the group consisting of hydrogen, deuterium, alkyl, haloalkyl, halo, alkoxy, haloalkoxy, amino, hydroxyl, cyano, nitro, thiol, carboxyalkyl, alkyl-S(O)H, alkyl-OP(O)H, alkyl-P(O)H, -O-carboxyalkyl, -O-alkyl-S(O)H, -O-alkyl-OP(O)H, -O-alkyl-P(O)H, -S(O)H, -OP(O)H, -P(O)H, alkyl-OP(O)3-alkyl, alkyl-P(O)3-alkyl, -O-alkyl-S(O)3-alkyl, -O-alkyl-OP(O)3-alkyl, -O-alkyl-P(O)3-alkyl, -S(O)3-alkyl, -OP(O)3-alkyl, -P(O)3-alkyl, sulfamide, sulfinimide; R 5f each occurrence of is independently hydrogen, alkyl, hydroxyalkyl, or alkoxyalkyl; R 9 each occurrence of is independently hydrogen or alkyl; each occurrence of q2 is independently an integer from 1 to 25; Each occurrence of q3 is independently an integer from 5 to 15.

[0272] In some embodiments, q2 is 8, 10, 12, or 14. In some embodiments, q3 is 6, 8, 10, 12, or 14.

[0273] In some embodiments, the hydrophilic component comprises polyethylene glycol, polysarcosine, cyclodextrin, c-glycoside, or a combination thereof. [ka] wherein: R 5b , R 5c R 5d , and R5e is independently selected from the group consisting of hydrogen, deuterium, alkyl, haloalkyl, halo, alkoxy, haloalkoxy, amino, hydroxyl, cyano, nitro, thiol, carboxyalkyl, alkyl-S(O)H, alkyl-OP(O)H, alkyl-P(O)H, -O-carboxyalkyl, -O-alkyl-S(O)H, -O-alkyl-OP(O)H, -O-alkyl-P(O)H, -S(O)H, -OP(O)H, and -P(O)H; R 5g each occurrence of is independently hydrogen, alkyl, hydroxyalkyl, or alkoxyalkyl; Each occurrence of q4 is independently an integer from 1 to 24.

[0274] In some embodiments, R 5g is -CH3 in each occurrence.

[0275] In some embodiments, the trigger element comprises a dipeptide, tripeptide, tetrapeptide, pentapeptide, glucuronide, disulfide, phosphate, diphosphate, triphosphate, hydrazone, or a combination thereof. In some embodiments, the trigger element comprises beta-glucuronic acid. In certain embodiments, the trigger element comprises a dipeptide, tripeptide, tetrapeptide, or pentapeptide. In certain embodiments, the trigger element comprises two or more amino acids selected from the group consisting of valine, citrulline, alanine, glycine, phenylalanine, lysine, or a combination thereof. In some embodiments, the trigger element comprises a sequence of amino acids selected from the group consisting of valine-citrulline, valine-alanine, glycine-glycine-phenylalanine-glycine, and combinations thereof. In certain embodiments, the trigger element comprises [ka] The compound includes one of the following structures (including combinations thereof):

[0276] In some embodiments, the sacrificial element comprises para-aminobenzyloxycarbonyl, aminal, hydrazine, disulfide, amide, ester, hydrazine, phosphotriester, diester, β-glucuronide, double bond, triple bond, ether bond, ketone, diol, cyano, nitro, quaternary amine, or a combination thereof. In certain embodiments, the sacrificial element comprises para-methoxybenzyl, dialkyldialkoxysilane, diaryldialkoxysilane, orthoester, acetal, optionally substituted β-thiopropionate, ketal, phosphoramidate, hydrazone, vinyl ether, imine, aconityl, trityl, polyketal, bis-arylhydrazone, diazobenzene, bivinaldiol, pyrophosphate diester, or a combination thereof.

[0277] In some embodiments, the sacrificial element is [ka] It contains one of the following structures:

[0278] In certain embodiments, the sacrificial element is [ka] wherein: R 6a , R 6b , R 6c , and R 6d are independently hydrogen, optionally substituted alkyl, optionally substituted aryl, or optionally substituted heteroaryl; or R 6a and R 6c together with the nitrogen and carbon atoms to which they are attached form an azetidinyl, pyrrolodinyl, piperidinyl, or homopiperidinyl, and R 6d is hydrogen, Y 1 is -O-, -S-, or -NR 6b -It is.

[0279] In some embodiments, the sacrificial element is [ka] wherein: R 6e , R 6f , R 6g , and R 6h are independently hydrogen, optionally substituted alkyl, optionally substituted aryl, or optionally substituted heteroaryl; or R 6a and R 6c together with the nitrogen and carbon atoms to which they are attached form an azetidinyl, pyrrolodinyl, piperidinyl, or homopiperidinyl, and R 6d is hydrogen, Y 2 is -O-, -S-, or -NR 6f -It is.

[0280] In some embodiments, the sacrificial element is [ka] wherein: R 10 each occurrence is independently alkyl, alkoxy, or halo; R 11 is hydrogen, alkyl, or -(CH2CH2O) z3 -CH3, R 12 is hydrogen or alkyl, R 13 is hydrogen or alkyl, z1 is 0 or 1, z2 is 0, 1, 2, 3, or 4; z3 is 0, 1, 2, 3, 4, 5, 6, 7, 8, 9, or 10.

[0281] In some embodiments, the sacrificial element is [ka] wherein: R 14a , R 14b , R 14c , R 14d , R 14e , and R 14f are each independently hydrogen, alkyl, hydroxyalkyl, or alkoxyalkyl; z4, z5, z6, and z7 are each independently 1, 2, 3, 4, 5, or 6.

[0282] In certain embodiments, the sacrificial element is [ka] wherein: z8 and z9 are each independently 1, 2, 3, 4, 5, or 6; or In some embodiments, the sacrificial element is [ka] wherein: R 15 is independently H, methyl, ethyl, isopropyl, tert-butyl, or phenyl; Y 3 is O or CH2, q5 is an integer ranging from 1 to 5.

[0283] In some embodiments, R 4a is hydrogen,

number

number

[0284] In some embodiments, the trigger element and the sacrificial element together comprise: [ka] [ka] It contains one of the following structures:

[0285] In some embodiments, the polar cap comprises one or more charged amino acids, one or more polyols, or a combination thereof. In certain embodiments, the polar cap comprises a diol, triol, tetraol, or a combination thereof. In some embodiments, the polar cap comprises glycerol, trimethylolpropane, pentaerythritol, maltitol, sorbitol, xylitol, erythritol, isomalt, or a combination thereof. In certain embodiments, the polar cap comprises one or more natural amino acids. In some embodiments, the polar cap comprises one or more unnatural amino acids. In certain embodiments, the polar cap comprises one or more unnatural amino acids and one or more natural amino acids. In some embodiments, the polar cap comprises serine, threonine, cysteine, proline, asparagine, glutamine, lysine, arginine, histidine, aspartate, glutamate, 4-hydroxyproline, 5-hydroxylysine, homoserine, homocysteine, ornithine, beta-alanine, statin, or gamma-aminobutyric acid. In certain embodiments, the polar cap comprises aspartic acid, serine, glutamic acid, serine-beta-glucose, or a combination thereof.

[0286] In some embodiments, the polar cap is [ka] The compound has one of the following structures (including combinations thereof):

[0287] In some embodiments, the payload is a chemotherapeutic agent, a cytotoxic agent, or a myeloid cell agonist.

[0288] In some embodiments, the payload is a chemotherapeutic agent selected from the group consisting of camptothecin, paclitaxel, doxorubicin, vinblastine, dacarbazine, irinotecan, topotecan, ciratecan, cositecan, exatecan, lurtotecan, SN-38, Dxd, gimatecan, belotecan, and rubitecan.

[0289] In yet other embodiments, the payload is a cytotoxic agent selected from the group consisting of calicheamicin, anthramycin, abeimicin, ticamycin, DC-81, mazethramycin, neothramycin A, neothramycin B, polothramycin prothracarcin, sivanomycin, sibiromycin, tomamycin, auristatin F, monomethyl auristatin F, auristatin E, monomethyl auristatin E, dolastatin, monomethyl dolastatin, mertansine, and emtansine.

[0290] In some embodiments, the payload is a STING agonist, a ligand for TLR2, a ligand for TLR3, a ligand for TLR4, a ligand for TLR5, a ligand for TLR6, a ligand for TLR7, a ligand for TLR8, a ligand for TLR9, a ligand for TLR10, a ligand for a nucleotide oligomerization domain (NOD), a ligand for a RIG-I-like receptor (RLR), a ligand for a C-type lectin receptor (CLR), a ligand for a cytoplasmic DNA sensor (CDS), and an inflammasome. a myeloid cell agonist selected from the group consisting of ligands of myeloid cell inducers, preferably the myeloid cell agonist is selected from sergantolimod, motolimod, resiquimod, 3M-051, 3M-052, MCT-465, IMO-4200, VTX-763, VTX-1463, RG7854, ADU-S100, MK-1454, MK-2118, BMS-986301, GSK3745417, SB-11285, and IMSA-101.

[0291] In some embodiments, L 1 , L 2 , or L 3 is alkylene, alkylene-La -, alkenylene, alkenylene-L a -, alkynylene, alkynylene-L a -, -L a -, -L a -Alkylene-L a -, -L a -Alkenylene-L a -, -L a -Alkynylene-L a -, ... L a Each occurrence of is independently -O-, -S-, -N(R 7 )-, -C(O)-, -C(S)-, -C(O)O-, -OC(O)-, -OC(O)O-, -C(O)N(R 7 )-, -N(R 7 )C(O)-, -C(O)N(R 7 )C(O)-, -C(O)N(R 7 )C(O)N(R 7 ), -N(R 7 )C(O)N(R 7 )-, -N(R 7 )C(O)O-, -OC(O)N(R 7 )-, -C(NR 7 )-, -N(R 7 )C(NR 7 )-, -C(NR 7 )N(R 7 )-, -N(R 7 )C(NR 7 )N(R 7 )-, -S(O)2-, -OS(O)-, -S(O)O-, -S(O), -OS(O)2-, -S(O)2O, -N(R 7 )S(O)2-, -S(O)2N(R 7 )-, -N(R 7 )S(O)-, -S(O)N(R 7 )-, -N(R 7 )S(O)2N(R 7 )- and -N(R 7 )S(O)N(R 7 )-selected from R7 are independently, in each occurrence, hydrogen, —NH2, —C(O)OCH2C6H5; and C 1~10 Alkyl, C 2~10 Alkenyl, C 2~10 Alkynyl, C 3~12 cycloalkyl, and 3- to 12-membered heterocycle, each of which is selected from halo, hydroxyl, cyano, nitro, amino, oxo, thioxo, —C(O)OCH2C6H5, —NHC(O)OCH2C6H5, C 1~10 Alkyl, C 1~10 Haloalkyl, C 1~10 Alkoxy, C 2~10 Alkenyl, C 2~10 Alkynyl, C 3~12 It is optionally substituted with one or more substituents independently selected from cycloalkyl, and 3- to 12-membered heterocyclyl.

[0292] In some embodiments, each L 1 , L 2 , or L 3 is alkyl, alkenyl, alkynyl, halo, hydroxyl, cyano, -OR 8 , -SR 8 , amino, aminyl, amido, cycloalkyl, aryl, heterocyclyl, heteroaryl, cycloclkylalkyl, arylalkyl, heterocyclylalkyl, heteroarylalkyl, -C(O)R 8 , -C(O)N(R 8 )2, -N(R 8 )C(O)R 8 , -C(O)OR 8 , -OC(O)R 8 , -S(O)R 8 , -S(O)2R 8 , -P(O)(OR 8 )2, -OP(O)(OR 8 )2, nitro, oxo, thioxo, =N(R 8 ), or cyano; R 8is hydrogen, alkyl, alkenyl, alkynyl, cycloalkyl, aryl, heterocyclyl, heteroaryl, cycloclkylalkyl, arylalkyl, heterocyclylalkyl, or heteroarylalkyl.

[0293] In some embodiments, L 1 , L 2 , or L 3 is, independently, [ka] wherein: R a is hydrogen or alkyl, L b is independently a direct bond, an optionally substituted alkylene linker, an optionally substituted heteroalkylene linker, a heteroatom linker, or a combination thereof; L c each occurrence of is independently an optionally substituted alkylene linker; However, L 1 , L 2 , or L 3 At least one of the [ka] It has the following structure.

[0294] In some embodiments, L 1 and L 2 is, independently, [ka] wherein: R a is hydrogen or alkyl, L b is independently a direct bond, an optionally substituted alkylene linker, an optionally substituted heteroalkylene linker, a heteroatom linker, or a combination thereof; L c each occurrence of is independently an optionally substituted alkylene linker; However, L 1 or L 2 At least one of the [ka] It has the following structure.

[0295] In some embodiments, L 2 teeth, [ka] It has the following structure.

[0296] In some embodiments, L 1 , L 2 , and L 3 are each independently [ka] wherein: *but, [ka] shows a direct bond to a substitutable position on the phenyl group in the structure:

[0297] In some embodiments, L 1 or L 2 teeth, [ka] wherein: *but, [ka] shows a direct bond to a substitutable position on the phenyl group in the structure:

[0298] In some embodiments, L c is C1-C6 alkylene.

[0299] In some embodiments, L c is substituted with one or more substituents selected from the group consisting of halo, haloalkyl, alkoxy, cyano, nitro, carboxy, sulfonamido, sulfonic acid, or combinations thereof. c is non-substituted.

[0300] In some embodiments, the conjugate has the following structure (IIa) or (IIb): [ka] It has one of the following.

[0301] In some embodiments, the conjugate has the following structure (IIc') or (IIc"): [ka] It has one of the following.

[0302] In certain embodiments, X 2 , X 3 , or both are CH, or CF. In some embodiments, X 1 , X 5 , or both are CR 3 and R 3 is H or halo. In some embodiments, X 1 and X 5 But both are CR 3 and R 3 is H or halo. In some embodiments, X 1 is CR 3 and R 3 is halo. In certain embodiments, X 5 is CR 3 and R 3 is halo. In some embodiments, halo is fluoro. In some embodiments, X 1 is CF. In some embodiments, X 1is CH. In some embodiments, X 5 is CH. In some embodiments, X 5 is CF. In some embodiments, X 3 is CR 3 and R 3 is H. In some embodiments, X 3 is CR 3 and R 3 is halo (e.g., fluoro).

[0303] In some embodiments, the conjugate has the following structure (IIa-1), (IIa-2), (IIa-3), (IIa-4), (IIa-5), or (IIa-6): [ka] wherein: L b is independently a direct bond, an optionally substituted alkylene linker, an optionally substituted heteroalkylene linker, a heteroatom linker, or a combination thereof; q6 is 0, 1, or 2.

[0304] In some embodiments, the conjugate has the following structure (IIa-1), (IIa-2), (IIa-3), (IIa-4), (IIa-5), (IIa-6), (IIa-7), or (IIa-8): [ka] wherein: L b is independently a direct bond, an optionally substituted alkylene linker, an optionally substituted heteroalkylene linker, a heteroatom linker, or a combination thereof; q6 is 0, 1, or 2.

[0305] In certain embodiments, the conjugate has the following structure (IIc-1), (IIc-2), (IIc-3), or (IIc-4): [ka] wherein: L b is independently a direct bond, an optionally substituted alkylene linker, an optionally substituted heteroalkylene linker, a heteroatom linker, or a combination thereof; q7 is 1, 2, or 3.

[0306] In some embodiments, the conjugate has the following structure (IId), (IIe), (IIf), or (IIg): [ka] wherein L b is independently a direct bond, an optionally substituted alkylene linker, an optionally substituted heteroalkylene linker, a heteroatom linker, or a combination thereof; q8 is 0, 1, or 2; q9 is 0, 1, or 2.

[0307] In some embodiments, the conjugate has the following structure (IIh) or (IIi): [ka] wherein: L b is independently a direct bond, an optionally substituted alkylene linker, an optionally substituted heteroalkylene linker, a heteroatom linker, or a combination thereof.

[0308] In some embodiments, L bis a direct bond, an optionally substituted alkylene linker, or an optionally substituted heteroalkylene linker.

[0309] In certain embodiments, L b is a direct bond, or [ka] wherein: R b Each occurrence of is independently hydrogen, alkyl, hydroxyalkyl, or alkoxyalkyl.

[0310] In some embodiments, the conjugate is [ka] [ka] [ka] wherein: R 1b is a chemotherapeutic agent, cytotoxic agent, or myeloid cell agonist; R 2b but, [ka] and having one of the structures: L 1g but, [ka] [ka] The structure of the compound is:

[0311] In some embodiments, the conjugate is [ka] [ka] [ka] wherein: R 1b is a chemotherapeutic agent, cytotoxic agent, or myeloid cell agonist; R 2b but, [ka] and having one of the structures: L 1g but, [ka] [ka] The structure of the compound is:

[0312] In certain embodiments, the conjugate is [ka] The structure of the compound is:

[0313] In some embodiments, the conjugate is [ka] [ka] The structure of the compound is:

[0314] In some embodiments, the conjugate is [ka] [ka] [ka] [ka] [ka] [ka] The structure of the compound is:

[0315] In some embodiments, the conjugate is [ka] The structure of the compound is:

[0316] In certain embodiments, the conjugate is [ka] [ka] [ka] [ka] [ka] [ka] [ka] The structure of the compound is:

[0317] In certain embodiments, R 1a teeth, [ka] The structure of the compound is:

[0318] In some embodiments, R 1a teeth, [ka] [ka] as its stereoisomer, enantiomer or tautomer, or a mixture thereof, or a pharmaceutically acceptable salt, solvate or prodrug thereof.

[0319] In some embodiments, R 1a teeth, [ka] [ka] [ka] The structure of the compound is:

[0320] In some embodiments, the payload or R 1a teeth, [ka] wherein: R' is hydrogen, or [ka] wherein: R a ' is H or C 1~6 is alkyl, R b ', C 1~6 Alkyl or C 1~6 is an alkoxy; R c 'But, H, C 1~6 Alkyl, -CHOH, or C 1~6 is an alkoxy; R d ' is H or C 1~6 is alkyl, or R e ' is H or C 1~6 It is alkyl.

[0321] In certain embodiments, L 1 or L 2 teeth, [ka] wherein: ** is X 1 , X 2 , X 3 , X 4 or X 5 The bond to

[0322] In some embodiments, X 2 CL 1 -R 1 and X 3 CL 2 -R 2 In some embodiments, X 3 CL 1 -R 1 and X 2 CL 2 -R 2 is.

[0323] In some embodiments, L 4 teeth, [ka] It has the following structure.

[0324] In some embodiments, L 4 teeth, [ka] It has the following structure.

[0325] In certain embodiments, L 4 teeth, [ka] It has the following structure.

[0326] In some embodiments, the targeting moiety is an anti-CD40 antibody, an antibody selected from an anti-LRRC15 antibody, an anti-CTSK antibody, an anti-ADAM12 antibody, an anti-ITGA11 antibody, an anti-FAP antibody, an anti-NOX4 antibody, an anti-SGCD antibody, an anti-SYNDIG1 antibody, an anti-CDH11 antibody, an anti-PLPP4 antibody, an anti-SLC24A2 antibody, an anti-PDGFRB antibody, an anti-THY1 antibody, an anti-ANTXR1 antibody, an anti-GAS1 antibody, an anti-CALHM5 antibody, an anti-SDC1 antibody, an anti-HER2 antibody, an anti-TROP2 antibody, an anti-MSLN antibody, an anti-Nectin 4 antibody, an anti-ASGR1 antibody, and an anti-MUC16 antibody.

[0327] In some embodiments, the antibody is a full-length antibody. In certain embodiments, the antibody is an antigen-binding fragment. In some embodiments, the antibody is a humanized antibody. In some embodiments, g is 1, 2, 3, 4, 5, 6, 7, 8, 9, or 10.

[0328] Certain embodiments provide conjugates having one of the structures in Table 2, wherein A is a targeting moiety or a binding fragment thereof, as a stereoisomer, enantiomer, or tautomer thereof, or a mixture thereof, or a pharmaceutically acceptable salt, solvate, or prodrug thereof. [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 Table 2-16 Table 2-17 Table 2-18 Table 2-19 Table 2-20 Table 2-21 Table 2-22 Table 2-23 Table 2-24 Table 2-25 Table 2-26 Table 2-27 Table 2-28 Table 2-29 Table 2-30 Table 2-31 Table 2-32 Table 2-33 Table 2-34 Table 2-35 Table 2-36 Table 2-37 Table 2-38 Table 2-39

[0329] In some embodiments, the targeting moiety is an antibody. Antibodies can be of any class, for example, IgA, IgD, IgE, IgG, and IgM. Some of these classes can be further subdivided into isotypes, for example, IgG1, IgG2, IgG3, IgG4, IgA1, and IgA2. The heavy chain constant regions (Fc) corresponding to different classes of immunoglobulins can be α, δ, ε, γ, or μ. The light chain can be either kappa (κ) or lambda (λ) based on the amino acid sequence of the constant domain. The antibody construct can also include antibody fragments or recombinant forms thereof, including single-chain variable fragments (scFv).

[0330] In certain embodiments, the antibody construct or targeting moiety may comprise an antigen-binding antibody fragment. The antibody fragment may include (i) a monovalent fragment consisting of a Fab fragment, a VL domain, a VH domain, a CL domain, and a CH1 domain; (ii) an F(ab')2 fragment, a bivalent fragment comprising two Fab fragments linked by a disulfide bridge at the hinge region; and (iii) an Fv fragment consisting of the VL and VH domains of a single antibody arm. The two domains, VL and VH, of the Fv fragment may be encoded by separate genes, but they may be linked by a synthetic linker produced as a single protein chain in which the VL and VH domains pair to form a monovalent molecule.

[0331] In other embodiments, the antibody construct or targeting moiety may contain, for example, two, three, four, five, six, seven, eight, nine, ten, or more antigen-binding domains. The antibody construct or targeting moiety may contain two antigen-binding domains, each capable of recognizing the same antigen. The antibody construct or targeting moiety may contain two antigen-binding domains, each capable of recognizing a different antigen. In some embodiments, the antibody construct or targeting moiety may comprise an Fc-fusion protein. In further embodiments, the antibody construct comprises an antigen-binding domain and an Fc region or domain. In still further embodiments, the antibody is a chimeric antibody, a humanized antibody, or a human antibody.

[0332] As used throughout this disclosure, "human antibody" may include, for example, antibodies having the amino acid sequence of a human immunoglobulin and may include antibodies isolated from a human immunoglobulin library or from animals transgenic for one or more human immunoglobulins, which typically do not express endogenous immunoglobulins. Human antibodies can be produced using transgenic mice that are incapable of expressing functional endogenous immunoglobulins but can express human immunoglobulin genes. Fully human antibodies that recognize a selected epitope can be generated using guided selection. In this approach, a selected non-human monoclonal antibody, e.g., a murine antibody, is used to guide the selection of a fully human antibody that recognizes the same epitope.

[0333] The antibodies or antigen-binding fragments thereof described herein may be derivatized or otherwise modified. For example, derivatized antibodies may be modified by glycosylation, acetylation, pegylation, phosphorylation, amidation, derivatization with known protecting / blocking groups, proteolytic cleavage, etc.

[0334] The antibody, antibody construct, or targeting moiety may be a derivatized antibody, e.g., the derivatized antibody may be modified by glycosylation, acetylation, pegylation, phosphorylation, amidation, derivatization with known protecting / blocking groups, proteolytic cleavage, or linkage to a cellular ligand or other protein.

[0335] The antibody construct may comprise a light chain of an amino acid sequence that has at least 1, 2, 3, 4, 5, 6, 7, 8, 9, or 10 modifications of the amino acid sequence compared to the native or original amino acid sequence, and in some embodiments has no more than 40, 35, 30, 25, 20, 15, or 10 modifications of the amino acid sequence. The antibody construct may comprise a heavy chain of an amino acid sequence that has at least 1, 2, 3, 4, 5, 6, 7, 8, 9, or 10 modifications of the amino acid sequence compared to the native or original amino acid sequence, and in some embodiments has no more than 40, 35, 30, 25, 20, 15, or 10 modifications of the amino acid sequence.

[0336] In any of the embodiments disclosed herein, the antigen-binding domain may specifically bind to a tumor antigen. In some embodiments, the tumor antigen is ASGR2, LRRC15, mesothelin (MSLN), HER2, CEA, TROP2, EPHA2, p-cadherin, UPK1B, FOLH1, LYPD3, or PVRL4 (Nectin-4). The antigen-binding domain may specifically bind to a molecule on an antigen-presenting cell (APC). In certain embodiments, the antigen-binding domain specifically binds to a human tumor antigen.

[0337] In some embodiments, the antibody (or antigen-binding domain thereof) specifically binds to ASGR1, CTLA4 (also known as CD152), PD-1 (or CD279), PD-L1 (or CD274), TNFR2 (or TNFRSF1B), OX40 (or TNFRSF4), CD27, IL2RA, TNFRSF18, LAG-3, GARP, 4-1BB, ICOS, CD70, PDGFRβ, CD73, CD38, integrin αvβ3, CD248, FAP, integrin αv, or integrin αvβ6.

[0338] In some related embodiments, the targeting moiety is an antibody (e.g., pertuzumab, brentuximab, gemtuzumab, trastuzumab, inotuzumab, polatuzumab, enfortumab, trastuzumab, sacituzumab, belantamab, or moxetumomab).

[0339] In some embodiments, the targeting moiety is an anti-CD40 antibody, an antibody selected from an anti-LRRC15 antibody, an anti-CTSK antibody, an anti-ADAM12 antibody, an anti-ITGA11 antibody, an anti-FAP antibody, an anti-NOX4 antibody, an anti-SGCD antibody, an anti-SYNDIG1 antibody, an anti-CDH11 antibody, an anti-PLPP4 antibody, an anti-SLC24A2 antibody, an anti-PDGFRB antibody, an anti-THY1 antibody, an anti-ANTXR1 antibody, an anti-GAS1 antibody, an anti-CALHM5 antibody, an anti-SDC1 antibody, an anti-HER2 antibody, an anti-TROP2 antibody, an anti-MSLN antibody, an anti-Nectin 4 antibody, an anti-ASGR1 antibody, an anti-Nectin 4 antibody, and an anti-MUC16 antibody.

[0340] In some embodiments, the targeting moiety is an anti-MSR1 antibody (e.g., as disclosed in PCT Publication No. 2019 / 217591, which is incorporated herein by reference).

[0341] Other exemplary antibodies within the scope of the present disclosure will be apparent to those skilled in the art. For example, exemplary targeting moieties (e.g., antibodies) can be found in PCT Publication Nos. 2019 / 217591, 2018 / 089373, PCT / US2021 / 054296, or U.S. Patent No. 11,179,473.

[0342] The antibody construct can be conjugated to a linker via cysteine-based biological conjugation. The antibody construct can be exchanged into an appropriate buffer, such as phosphate, borate, PBS, histidine, or tris-acetate, at a concentration of about 2 mg / mL to about 10 mg / mL, using an appropriate number of equivalents of a reducing agent, such as dithiothreitol (DTT) or tris(2-carboxyethyl)phosphine (TCEP). The resulting solution can be stirred for an appropriate time and temperature to achieve the desired reduction. The compound of structure (I) can be added as a solution with stirring. Depending on the physical properties of the compound of structure (I), a cosolvent can be introduced before adding the compound of structure (I) to promote solubility. The reaction can be stirred at room temperature for about 1 hour to about 12 hours, depending on the observed reactivity. The progress of the reaction can be monitored by liquid chromatography-mass spectrometry (LC-MS). Once the reaction is deemed complete, the remaining free compound of structure (I) may be removed by applicable methods, and the conjugate of structure (II) may be exchanged into a desired formulation buffer. Such conjugates can be synthesized starting from a targeting moiety (e.g., an antibody or mAb) and a compound of structure (I), e.g., 7 equivalents, using the conditions described in the conjugation reaction scheme below. The monomer content and drug-antibody ratio can be determined by methods described herein and known in the art.

[0343] Conjugation Reaction Scheme [ka] Pharmaceutical Compositions The compositions and methods described herein may be useful as pharmaceutical compositions for administration to a subject in need thereof. The pharmaceutical composition may include at least a composition described herein and one or more pharmaceutically acceptable carriers, diluents, excipients, stabilizers, dispersing agents, suspending agents, and / or thickening agents. The composition may include a conjugate of structure (II). In some embodiments, the targeting moiety is an anti-LRRC15 antibody. In some embodiments, the targeting moiety is an anti-ASGR1 antibody. The pharmaceutical composition may include a conjugate of structure (II) and one or more of a buffer, an antibiotic, a steroid, a carbohydrate, a drug (e.g., a chemotherapy drug), radiation, a polypeptide, a chelating agent, an adjuvant, and / or a preservative.

[0344] Pharmaceutical compositions can be formulated using one or more physiologically acceptable carriers, including excipients and adjuvants. Formulations can be modified depending on the selected route of administration. Pharmaceutical compositions containing conjugates can be prepared, for example, by lyophilizing, mixing, dissolving, emulsifying, encapsulating, or encapsulating the conjugate. Pharmaceutical compositions can also contain the conjugate in free base form or in pharmaceutically acceptable salt form.

[0345] Methods for formulating the conjugates may include formulating any of the conjugates with one or more inert pharmaceutically acceptable excipients or carriers to form solid, semi-solid, or liquid compositions. Solid compositions may include, for example, powders, tablets, dispersible granules, and capsules, and in some embodiments, the solid compositions further contain non-toxic auxiliary substances, such as wetting or emulsifying agents, pH buffering agents, and other pharmaceutically acceptable additives. Alternatively, the conjugates of structure (II) may be lyophilized or in powder form for reconstitution with a suitable vehicle, for example, sterile, pyrogen-free water, before use.

[0346] Pharmaceutical compositions of the conjugates of structure (II) may include at least one active ingredient (e.g., a compound, salt, or conjugate and another drug). The active ingredient may be encapsulated, for example, in microcapsules prepared by droplet formation techniques or interfacial polymerization (e.g., hydroxymethylcellulose or gelatin microcapsules and poly-(methyl methacrylate) microcapsules, respectively), in colloidal drug delivery systems (e.g., liposomes, albumin microspheres, microemulsions, nanoparticles, and nanocapsules), or in macroemulsions.

[0347] In many cases, pharmaceutical compositions may contain more than one active compound necessary for the particular indication being treated. The active compounds may have complementary activities that do not adversely affect each other. For example, the composition may further contain a chemotherapeutic agent, a cytotoxic agent, a cytokine, a growth inhibitor, an antihormonal agent, an antiangiogenic agent, and / or a cardioprotective agent. Such molecules may be present in combination in amounts effective for the intended purpose.

[0348] The compositions and formulations may be sterilized. Sterilization may be achieved by filtration by sterile filtration.

[0349] The composition may be formulated for administration as an injection. Non-limiting examples of injection preparations may include sterile suspensions, solutions, or emulsions in oily or aqueous vehicles. Suitable oily vehicles may include lipophilic solvents, or vehicles such as fatty oils or synthetic fatty acid esters, or liposomes. Aqueous injection suspensions may contain substances that increase the viscosity of the suspension. The suspension may also contain suitable stabilizers. The injection may be formulated for bolus injection or continuous infusion. Alternatively, the composition may be lyophilized or in powder form for reconstitution with a suitable vehicle, such as sterile pyrogen-free water, before use.

[0350] As used throughout this disclosure, the phrases "parenteral administration" and "administered parenterally" refer to modes of administration other than enteral and topical administration, usually by injection, and include, but are not limited to, intravenous, intramuscular, intra-arterial, intrathecal, intracapsular, intraorbital, intracardiac, intradermal, intraperitoneal, transtracheal, subcutaneous, subcuticular, intra-articular, subcapsular, subarachnoid, intraspinal and intrapleural injection and infusion.

[0351] The phrase "pharmaceutically acceptable" is used herein to refer to compounds, materials, compositions, and / or dosage forms that are, within the scope of sound medical judgment, suitable for use in contact with the tissues of human beings and animals without undue toxicity, irritation, allergic response, or other problem or complication, commensurate with a reasonable benefit / risk ratio.

[0352] As used throughout this disclosure, the phrase "pharmaceutically acceptable excipient" or "pharmaceutically acceptable carrier" refers to a pharmaceutically acceptable material, composition, or vehicle, such as a liquid or solid filler, diluent, excipient, solvent, or encapsulating material. Each carrier must be "acceptable" in the sense of being compatible with the other ingredients of the formulation and not harmful to the patient. Some examples of materials that can serve as pharmaceutically acceptable carriers include: (a) sugars, such as lactose, glucose, and sucrose; (b) starches, such as corn starch and potato starch; (c) cellulose and its derivatives, such as sodium carboxymethylcellulose, ethylcellulose, and cellulose acetate; (d) powdered tragacanth; (e) malt; (f) gelatin; (g) talc; (h) excipients, such as cocoa butter and suppository wax; and (i) oils, such as peanut oil, cottonseed oil, safflower oil, sesame oil, and olive oil. , corn oil and soybean oil, (j) glycols, such as propylene glycol, (k) polyols, such as glycerin, sorbitol, mannitol and polyethylene glycol, (l) esters, such as ethyl oleate and ethyl laurate, (m) agar, (n) buffers, such as magnesium hydroxide and aluminum hydroxide, (o) alginic acid, (p) pyrogen-free water, (q) isotonic saline, (r) Ringer's solution, (s) ethyl alcohol, (t) phosphate buffer solution, and (u) other non-toxic compatible substances used in pharmaceutical formulations.

[0353] The term "salt" or "pharmaceutically acceptable salt" refers to salts derived from a variety of organic and inorganic counterions known in the art. Pharmaceutically acceptable acid addition salts can be formed with inorganic and organic acids. Inorganic acids from which salts can be derived include, for example, hydrochloric acid, hydrobromic acid, sulfuric acid, nitric acid, phosphoric acid, and the like. Organic acids from which salts can be derived include, for example, acetic acid, propionic acid, glycolic acid, pyruvic acid, oxalic acid, maleic acid, malonic acid, succinic acid, fumaric acid, tartaric acid, citric acid, benzoic acid, cinnamic acid, mandelic acid, methanesulfonic acid, ethanesulfonic acid, p-toluenesulfonic acid, salicylic acid, and the like. Pharmaceutically acceptable base addition salts can be formed with inorganic and organic bases. Inorganic bases from which salts can be derived include, for example, sodium, potassium, lithium, ammonium, calcium, magnesium, iron, zinc, copper, manganese, aluminum, and the like. Organic bases from which salts can be derived include, for example, primary, secondary, and tertiary amines, substituted amines including naturally occurring substituted amines, cyclic amines, basic ion exchange resins, and the like, specifically, for example, isopropylamine, trimethylamine, diethylamine, triethylamine, tripropylamine, and ethanolamine. In some embodiments, pharmaceutically acceptable base addition salts can be selected from ammonium, potassium, sodium, calcium, and magnesium salts.

[0354] For parenteral administration, the compound, salt, or conjugate may be formulated in a unit-dosage injectable form (e.g., solution, suspension, emulsion) in association with a pharmaceutically acceptable parenteral vehicle. Such vehicles may be essentially non-toxic and non-therapeutic. Vehicles may be water, saline, Ringer's solution, dextrose solution, and 5% human serum albumin. Non-aqueous vehicles such as fixed oils and ethyl oleate may also be used. Liposomes may be used as carriers. Vehicles may contain minor amounts of additives, such as substances that enhance isotonicity and chemical stability (e.g., buffers and preservatives).

[0355] Sustained-release preparations may also be prepared. Examples of sustained-release preparations may include semipermeable matrices of solid hydrophobic polymers that can contain the compound, salt, or conjugate, and these matrices may be in the form of shaped articles (e.g., films or microcapsules). Examples of sustained-release matrices may include polyesters, hydrogels (e.g., poly(2-hydroxyethyl-methacrylate) or poly(vinyl alcohol)), polylactides, copolymers of L-glutamic acid and gamma-ethyl-L-glutamate, non-degradable ethylene-vinyl acetate, degradable lactic acid-glycolic acid copolymers, such as LUPRON DEPO™ (i.e., injectable microspheres composed of lactic acid-glycolic acid copolymer and leuprolide acetate), and poly-D-(-)-3-hydroxybutyric acid.

[0356] Pharmaceutical formulations can be prepared for storage by mixing the conjugate of structure (II) with pharmaceutically acceptable carriers, excipients, and / or stabilizers. The formulation can be a lyophilized formulation or an aqueous solution. Acceptable carriers, excipients, and / or stabilizers can be non-toxic to recipients at the dosages and concentrations used. Acceptable carriers, excipients, and / or stabilizers can include buffers such as phosphate, citrate, and other organic acids; antioxidants including ascorbic acid and methionine; preservatives; proteins such as polypeptides, serum albumin, or gelatin; hydrophilic polymers; amino acids; monosaccharides, disaccharides, and other carbohydrates including glucose, mannose, or dextrins; chelating agents such as EDTA; sugars such as sucrose, mannitol, trehalose, or sorbitol; salt-forming counterions such as sodium; metal complexes; and / or non-ionic surfactants; or polyethylene glycol.

[0357] Pharmaceutical formulations of the conjugates may have an average payload-antibody construct ratio ("DAR") selected from about 1 to about 20, about 1 to about 10, about 1 to about 5, about 1 to about 2, or about 1 to about 1. In some embodiments, the average DAR of the formulation is about 2 to about 8, about 3 to about 8, about 3 to about 7, about 3 to about 5, or about 2. In some embodiments, the pharmaceutical formulation has an average DAR of about 3, about 3.5, about 4, about 4.5, or about 5.

[0358] Treatment method The conjugates of the present disclosure are useful for treating diseases (i.e., conjugates of structure (II)). These conjugates disclosed herein provide a targeted approach to drug delivery strategies. Accordingly, certain embodiments provide a method of treating a disease (or a symptom thereof) comprising administering to a mammal (e.g., a human) in need thereof a therapeutically effective amount of a conjugate of structure (II) or a pharmaceutical composition comprising the same.

[0359] Treatment and / or treating refers to any indication of success in treating or alleviating a disease or condition. Treating may include, for example, reducing, delaying, or alleviating the severity of one or more symptoms of a disease or condition, or reducing the frequency with which a symptom of a disease, defect, disorder, or adverse condition is experienced by a patient. Treatment can be used herein to refer to a method that results in some level of treatment or alleviation of a disease or condition, and can contemplate a range of results directed toward that goal, including, but not limited to, complete prevention of the condition.

[0360] Prevention, preventing, etc., refers to the prevention of a disease or condition, e.g., tumor formation, in a patient. For example, if an individual at risk of developing a tumor or other form of cancer is treated with the methods of the present disclosure and does not later develop the tumor or other form of cancer, the disease has been prevented in that individual, at least for a certain period of time. Preventing can also refer to preventing the recurrence of a disease or condition in a patient previously treated for the disease or condition, e.g., by preventing recurrence.

[0361] A therapeutically effective amount (also referred to as an effective amount) may be the amount of a composition comprising a conjugate of structure (II) sufficient to provide a beneficial effect or reduce otherwise harmful and unfavorable events in an individual to whom the composition is administered. A therapeutically effective dose may be a dose that produces one or more desired or desirable (e.g., beneficial) effects for which it is administered, such administration occurring one or more times over a given period of time. The exact dose may depend on the purpose of the treatment and may be ascertainable by one of ordinary skill in the art using known techniques and the teachings provided herein.

[0362] Conjugates that can be used in therapy can be formulated and dosed in a manner consistent with good medical practice, taking into account the disease or condition being treated, the condition of the individual patient, the site of delivery of the composition, the method of administration, and other factors known to physicians. The compositions can be prepared according to the preparation instructions provided herein.

[0363] The pharmaceutical compositions can be used in the methods described herein and can be administered to a subject in need thereof using techniques known to those of skill in the art, which may be suitable as therapy for a disease or condition affecting the subject. Those of skill in the art will understand that the amount, duration, and frequency of administration of a pharmaceutical composition to a subject in need thereof will depend on several factors, including, for example, the health of the subject, the patient's particular disease or condition, the grade or level of the patient's particular disease or condition, additional treatments the subject is receiving or has received, etc.

[0364] The conjugates, compositions, and methods of the present disclosure are useful as single agents for the treatment or prevention of diseases such as infections, autoimmune disorders (e.g., multiple sclerosis), inflammation (including autoinflammatory disorders), and cancer. Alternatively, the conjugates, compositions, and methods of the present disclosure may be used in combination therapy with a second therapeutic agent to treat or prevent diseases such as infections, autoimmune disorders (e.g., multiple sclerosis), inflammation (including autoinflammatory disorders), and cancer.

[0365] Some embodiments provide a pharmaceutical composition comprising a conjugate of structure (II) and a pharmaceutically acceptable excipient.

[0366] Another embodiment provides a method for treating cancer, comprising administering an effective amount of a conjugate of structure (II) or a pharmaceutical composition thereof to a subject in need thereof. Yet another embodiment provides a method for treating infection, inflammation, an autoimmune disorder, or a combination thereof, comprising administering an effective amount of a conjugate of structure (II) or a pharmaceutical composition thereof to a subject in need thereof. In some of the foregoing embodiments, the effective amount of the conjugate is administered intravenously, subcutaneously, or intratumorally.

[0367] Antigens targeted by the conjugate of structure (II) may be derived from the following specific conditions and / or families of conditions, including cancers such as cancer-associated fibroblasts (CAF), brain cancer, skin cancer, lymphoma, sarcoma, lung cancer, liver cancer, leukemia, uterine cancer, breast cancer (including triple-negative breast cancer), ovarian cancer, cervical cancer, uterine cancer, bladder cancer, stomach cancer, esophageal cancer, kidney cancer, angiosarcoma, bone cancer, blood cancer, testicular cancer, prostate cancer, stomach cancer, colon cancer, intestinal cancer, pancreatic cancer, head and neck cancer (including head and neck squamous cell carcinoma), mesothelioma, melanoma, and other types of cancer, as well as precancerous conditions such as hyperplasia.

[0368] Further non-limiting examples of cancers include acute lymphoblastic leukemia (ALL); acute myeloid leukemia (AML); adrenocortical carcinoma; astrocytoma, childhood cerebellar or cerebral cancer; basal cell carcinoma; bone tumors, osteosarcoma / malignant fibrous histiocytoma; brain cancer; brain tumors (e.g., cerebellar astrocytoma, malignant glioma, ependymoma, medulloblastoma, visual pathway and hypothalamic glioma); brain stem glioma; breast cancer (including triple-negative breast cancer); bronchial adenoma / carcinomatous; Burkitt's lymphoma; cerebellar astrocytoma; cervical cancer; bile duct carcinoma; chondrosarcoma; chronic lymphocytic leukemia; chronic myeloid leukemia; chronic Myeloproliferative disorders; colon cancer; cutaneous T-cell lymphoma; endometrial cancer; epithelial cancer; epithelial cancer; eye cancer, e.g., intraocular melanoma and retinoblastoma; gallbladder cancer; gastric cancer; glioma; hairy cell leukemia; head and neck cancer (including head and neck squamous cell carcinoma); heart cancer; hepatocellular (liver) cancer; Hodgkin's lymphoma; hypopharyngeal cancer; islet cell carcinoma (endocrine pancreas); Kaposi's sarcoma; kidney cancer (renal cell carcinoma); laryngeal cancer; leukemia, e.g., acute lymphoblastic, acute myeloid, chronic lymphocytic, chronic myeloid, and hairy cell; lip and oral cavity cancer; liposarcoma; lung cancer, e.g., non-small cell and small cell ;Lymphomas, e.g., AIDS-related, Burkitt's;Lymphoma, cutaneous T-cell, Hodgkin's and non-Hodgkin's, macroglobulinemia, malignant fibrous histiocytoma of bone / osteosarcoma;Melanoma (including cutaneous melanoma);Merkel cell carcinoma;Mesothelioma;Multiple myeloma / plasma cell neoplasms;Mycosis fungoides;Myelodysplastic syndromes;Myelodysplastic / myeloproliferative disorders;Chronic myeloproliferative disorders;Nasal cavity and paranasal sinus cancer;Nasopharyngeal carcinoma;Neuroblastoma;Oligodendroglioma;Oral cancer;Osteosarcoma / malignant fibrous histiocytoma of bone;Ovarian cancer;Pancreatic cancer;Parathyroid cancer;Pharyngeal cancer;Phaeochromocytoma;Pituitary adenoma;Plasma cell neoplasms biological; pleuropulmonary blastoma; prostate cancer; rectal cancer; kidney cancer (renal carcinoma); transitional cell carcinoma of the renal pelvis and ureter; rhabdomyosarcoma; salivary gland cancer; Ewing family of tumors; Kaposi's sarcoma; soft tissue sarcoma; uterine sarcoma; Sezary syndrome; skin cancer (non-melanoma); skin cancer; small intestine cancer; soft tissue sarcoma; squamous cell carcinoma; metastatic cervical carcinoma of occult primary; gastric cancer; testicular cancer; pharyngeal cancer; thymoma and thymic carcinoma; thymoma; thyroid cancer; pediatric thyroid cancer; uterine cancer; vaginal cancer; Waldenstrom's macroglobulinemia; Wilms' tumor, or any combination thereof.

[0369] Additional therapeutic agents that can be combined with the conjugates of the present disclosure can be found in Goodman and Gilman's "The Pharmacological Basis of Therapeutics," Tenth Edition edited by Hardman, Limbird and Gilman, or the Physician's Desk Reference, both of which are incorporated herein by reference in their entireties.

[0370] The conjugates of structure (II) described herein can be used in combination with the drugs disclosed herein or other suitable drugs, depending on the condition being treated. Thus, in some embodiments, one or more conjugates of the present disclosure are co-administered with other drugs described herein. When used in combination therapy, the conjugates described herein are administered simultaneously with the second drug or separately. This combined administration can include simultaneous administration of the two drugs in the same dosage form, simultaneous administration in separate dosage forms, and separate administration. That is, the conjugates described herein and any of the drugs described above can be formulated together in the same dosage form and administered simultaneously. Alternatively, the conjugates of the present disclosure and any of the drugs described above can be administered simultaneously, with both drugs being present in separate formulations. In another alternative, the conjugates of the present disclosure can be administered immediately after any of the drugs described above, or vice versa. In some embodiments of separate administration protocols, the conjugates of the present disclosure and any of the drugs described above are administered within minutes, hours, or days of each other.

[0371] In some embodiments of the present disclosure, including those in which the conjugates and pharmaceutical compositions of the present disclosure are intended to treat or prevent inflammation (including autoinflammatory disorders) or autoimmune disorders, the second therapeutic agent comprises an anti-inflammatory composition, a steroid composition, a nonsteroidal anti-inflammatory drug (NSAID) composition, a cyclooxygenase (COX) enzyme (e.g., a COX1 inhibitor and / or a COX2 inhibitor) composition, or a regulatory T cell antagonist composition.

[0372] In some embodiments of the present disclosure, including those in which the conjugates and pharmaceutical compositions of the present disclosure are intended to treat or prevent cancer, the second therapeutic agent comprises one or more of a second conjugate or pharmaceutical composition of the present disclosure, a chemotherapy composition, radiation therapy, an immunoconjugate composition having specificity for LRRC15, HER2, or MSLN, an immunoconjugate composition having specificity for an antigen other than LRRC15, HER2, or MSLN, or an engineered T cell composition (e.g., a CAR-T composition and / or a TCR-T composition).

[0373] In some embodiments, the methods of treatment provided herein include administering to a subject an additional therapeutic agent, such as an anti-cancer agent or an anti-fibrotic agent, hi some embodiments, the additional therapeutic agent is an anti-cancer agent selected from a chemotherapeutic agent, a cytotoxic agent, a cytokine, a growth inhibitory agent, an anti-hormonal agent, an anti-angiogenic agent, and / or a cardioprotective agent.

[0374] Examples of chemotherapeutic agents contemplated as additional therapeutic agents include alkylating agents, such as nitrogen mustards (e.g., mechlorethamine, cyclophosphamide, ifosfamide (IFEX®), melphalan (Alkeran®), and chlorambucil); bifunctional chemotherapy agents (e.g., bendamustine®); nitrosoureas (e.g., carmustine (BCNU, BiCNU®, polifeprosan 20 implant (Gliadel®)), lomustine (CCNU), and semustine (methyl- CCNU); ethylenimines and methyl-melamines (e.g., triethylenemelamine (TEM), triethylenethiophosphoramide (thiotepa), and hexamethylmelamine (HMM, altretamine)); alkylsulfonates (e.g., busulfan (Myleran®), busulfan injection (Busulfex®)); antimetabolites, such as folic acid analogs (e.g., methotrexate (Folex®), trimetrexate, and pemetrexed (multitargeted antifolates)) and capecitabine (Xeloda®) pyrimidine analogs (e.g., 5-fluorouracil (5-FU, Adrucil®, Efudex®), fluorodeoxyuridine, tezacitabine, gemcitabine, cytosine arabinoside (AraC, cytarabine (Cytosar-U®); cytarabine liposome injection (DepoCyt®)), 5-azacytidine, and 2,2'-difluorodeoxycytidine); purine derivatives (e.g., 6-mercaptopurine (Purinethol®), 6-thioguanine, azathioprine, 2 '-deoxycoformycin (pentostatin), erythrohydroxynonyladenine (EHNA), fludarabine phosphate (Fludara®), 2-chlorodeoxyadenosine (cladribine, 2-CdA)); type I topoisomerase inhibitors, such as camptothecin (CPT), topotecan (Hycamptin®), and irinotecan (Camptosar®); natural products, such as epipodophyllotoxins (e.g., etoposide (Vepesid®) and teniposide (Vumon®));Vinca alkaloids (e.g., vinblastine (Velban®), vincristine (Oncovin®), and vinorelbine (Navelbine®)); antitumor antibiotics, such as actinomycin D (dactinomycin, Cosmegan®), doxorubicin hydrochloride (Adriamycin®, Rubex®), mitoxantrone (Novantrone®), and bleomycin sulfate (Blenoxane®); radiosensitizers, such as platinum coordination complexes, such as cisplatin (Platinol®), carboplatin (Paraplatin®), and oxaliplatin (Eloxatin®); substituted ureas, such as hydroxyurea (Hydrea®); microtubule inhibitors, such as paclitaxel (Taxol®) and docetaxel (Taxotere®); immunosuppressants, such as cyclohexyl 1-hydroxybenzoates (C1H2O4), cyclohexyl 1-hydroxybenzoates (C1H2O5), cyclohexyl 1-hydroxybenzoates (C1H2O6), cyclohexyl 1-hydroxybenzoates (C1H2O7), cyclohexyl 1-hydroxybenzoates (C1H2O8), cyclohexyl 1-hydroxybenzoates (C1H2O9 ... inhibitors, such as cyclophosphamide (Cytoxan® or Neosar®); hormonal compounds, such as anastrozole (Arimidex®), exemestane (Aromasin®), letrozole (Femara®), fulvestrant (Faslodex®), and bicalutamide (Casodex®)) and tamoxifen citrate (Nolvadex®); anti-inflammatory agents, such as dexamethasone; anti-androgen compounds, such as For example, flutamide (Eulexin®); anthracycline compounds, such as idarubicin (Idamycin®, Zavedos®) and epirubicin (Ellence®); bioreductive anticancer drugs, such as tirapazamine (Tirazone®); serine / threonine kinase inhibitors, such as the CDK4 / 6 inhibitors abemaciclib (Verzenio®), palbociclib (Ibrance®), and ribociclib (Kisqali®);and methylhydrazine derivatives, such as N-methylhydrazine (MIH) and procarbazine;

[0375] Preparation method The examples and preparations provided below further describe and illustrate the compounds of the present disclosure, as well as methods for preparing such compounds and their conjugates. It should be understood that the scope of the present disclosure is not limited in any way by the scope of the following examples and preparations. In the following examples, and throughout this specification and claims, unless otherwise stated, molecules and moieties with a single stereocenter exist as racemic mixtures. Unless otherwise stated, these molecules and moieties with two or more stereocenters exist as racemic mixtures of diastereomers. Single enantiomers / diastereomers can be obtained by methods known to those skilled in the art.

[0376] It will also be understood by those skilled in the art that in the processes described below, functional groups of intermediate compounds may need to be protected by suitable protecting groups. Such functional groups include hydroxy, amino, mercapto, and carboxylic acid. Suitable protecting groups for hydroxy include trialkylsilyl or diarylalkylsilyl (e.g., t-butyldimethylsilyl, t-butyldiphenylsilyl, or trimethylsilyl), tetrahydropyranyl, benzyl, and the like. Suitable protecting groups for amino, amidino, and guanidino include t-butoxycarbonyl, benzyloxycarbonyl, and the like. Suitable protecting groups for mercapto include -C(O)-R" (where R" is alkyl, aryl, or arylalkyl), p-methoxybenzyl, trityl, and the like. Suitable protecting groups for carboxylic acid include alkyl, aryl, or arylalkyl esters. Protecting groups can be added or removed according to standard techniques known to those skilled in the art and as described herein. The use of protecting groups is discussed in Green, TW and PGM Wutz, Protective Groups in Organic Synthesis (1999), 3 rdEd., Wiley. As one skilled in the art will appreciate, the protecting group may be a polymer resin such as a Wang resin, a Rink resin, or a 2-chlorotrityl-chloride resin.

[0377] Additionally, all compounds of the present disclosure that exist in free base or acid form can be converted to their salts by treatment with an appropriate inorganic or organic base or acid by methods known to those skilled in the art. Salts of compounds of the present disclosure can be converted to their free base or acid form by standard techniques.

[0378] The following reaction schemes and examples show exemplary methods of making the compounds of the present disclosure. It is understood that one of ordinary skill in the art may be able to make these compounds by analogous methods or by combining other methods known to one of ordinary skill in the art. It is also understood that one of ordinary skill in the art would be able to make other compounds of structure (I) not specifically exemplified below in a manner similar to those described below by using appropriate starting components and modifying the synthetic parameters as necessary. In general, the starting components may be obtained from sources such as Sigma Aldrich, Lancaster Synthesis, Inc., Maybridge, Matrix Scientific, TCI, and Fluorochem USA, or may be synthesized according to sources known to one of ordinary skill in the art (e.g., see Advanced Organic Chemistry: Reactions, Mechanisms, and Structure, 5 thedition (Wiley, December 2000)), or may be prepared as described in the present disclosure. The compound 4-((S)-2-((S)-2-amino-3-methylbutanamido)-5-ureidopentanamido)benzyl 3-(2-amino-4-(dipropylcarbamoyl)-3H-benzo[b]azepine-8-carboxamido)-7,8-dihydro-1,6-naphthyridine-6(5H)-carboxylate can be synthesized according to U.S. Pat. No. 10,239,862, the synthetic procedures for this compound being incorporated herein by reference.

[0379] General conjugation scheme [ka] The above general reaction scheme is illustrative of a conjugation reaction between a compound of structure (I) and its targeting moiety or binding fragment, where mAb-SH represents a targeting moiety or binding fragment thereof (e.g., a monoclonal antibody) having a free thiol (—SH), and X 1 , X 2 , X 2 , X 4 , X 5 , R 4a , and R 4b is as defined herein. Targeting moieties or binding fragments thereof having a free thiol can be prepared and conjugated to compounds of structure (I) using methods known in the art and described herein (see, e.g., Conjugation Example 1). After the initial conjugation reaction, the resulting conjugate of structure (II) can undergo an irreversible hydrolysis reaction to form an alternative conjugate of structure (II) as shown.

[0380] Reaction Scheme 1 [ka] [ka] [ka]

[0381] Intermediate Example 1 Preparation of tert-butyl 2-[[2-(benzyloxycarbonylamino)acetyl]amino]acetate [ka] To a solution of 2-(benzyloxycarbonylamino)acetic acid (5.50 g, 26.3 mmol, 1.0 equiv.) in DMF (100 mL), triethylamine (TEA; 7.98 g, 78.9 mmol, 11.0 mL, 3.0 equiv.), HATU (12.0 g, 31.5 mmol, 1.2 equiv.), and tert-butyl 2-aminoacetate (3.45 g, 26.3 mmol, 1.0 equiv.) were added and stirred at 25 °C for 2 h. The reaction mixture was quenched at 0 °C by adding HO (150 mL) and then extracted with ethyl acetate (100 mL × 3). The combined organic layers were washed with HO (80 mL × 3), then with brine (100 mL), dried over NaSO, filtered, and concentrated under reduced pressure to give a residue. The residue was purified by silica gel chromatography (column height: 250 mm, diameter: 100 mm, 100-200 mesh silica gel, ethyl acetate / methanol=1 / 1) to give tert-butyl 2-[[2-(benzyloxycarbonylamino)acetyl]amino]acetate (8 g, 24.82 mmol, 94.40% yield) as a yellow oil. 1 H NMR(CDCl3,400MHz)δ 7.40-7.31(m,5H),5.14(s,2H),3.98-3.89(m,4H),1.47(s,9H)

[0382] Intermediate Example 2 Preparation of tert-butyl 2-[(2-aminoacetyl)amino]acetate [ka] To a solution of tert-butyl 2-[[2-(benzyloxycarbonylamino)acetyl]amino]acetate (8.80 g, 27.3 mmol, 1.0 equiv.) in methanol (150 mL) was added Pd / C (10%, 2.5 g) under a N2 atmosphere. The suspension was degassed and purged with H2 for 3 minutes. The mixture was stirred under H2 (50 Psi) at 25 °C for 2 hours. The reaction mixture was filtered and concentrated under reduced pressure to give a residue. The compound tert-butyl 2-[(2-aminoacetyl)amino]acetate (4.2 g, 22.31 mmol, 81.74% yield) was obtained as a white solid. 1 H NMR(CDCl3,400MHz)δ 7.67(s,1H),3.96(d,J=5.6Hz,2H),3.39(s,2H),1.75(s,2H),1.46(s,9H)

[0383] Intermediate Example 3 Preparation of methyl (E)-3-(4,4,5,5-tetramethyl-1,3,2-dioxaborolan-2-yl)prop-2-enoate [ka] To a solution of CuCl (1.06 g, 10.7 mmol, 256 μL, 0.03 equiv.) and Xantphos (6.19 g, 10.7 mmol, 0.03 equiv.) in THF (450 mL) was added t-BuONa (2.06 g, 21.4 mmol, 0.06 equiv.) at 0 °C. The reaction solution was stirred at 0 °C for 1 h, and then a solution of Pin2B2 (90.6 g, 357 mmol, 1.0 equiv.) in THF (150 mL) was added. The reaction solution was stirred at 20 °C for 1 h under a nitrogen atmosphere. Methyl prop-2-ynoate (30.0 g, 357 mmol, 29.7 mL, 1.0 equiv.) and methanol (22.9 g, 714 mmol, 28.9 mL, 2.0 equiv.) were added to the above reaction solution. The reaction mixture was then stirred at 20 °C for an additional 12 h. The resulting mixture was poured into ice water (w / w = 1 / 1) (300 mL) and stirred for 5 min. The aqueous phase was extracted with ethyl acetate (200 mL × 3). The combined organic phases were washed with brine (50 mL × 2), dried over anhydrous Na2SO4, filtered, and concentrated under high vacuum. The residue was purified by silica gel chromatography (column height: 250 mm, diameter: 100 mm, 100-200 mesh silica gel, petroleum ether / ethyl acetate = 1 / 0, 3 / 1) to give methyl (E)-3-(4,4,5,5-tetramethyl-1,3,2-dioxaborolan-2-yl)prop-2-enoate (70 g, 330 mmol, 92.51% yield) as a colorless oil. 1 H NMR (400MHz, CDCl3) δ 6.83-6.75 (m, 1H), 6.68-6.59 (m, 1H), 3.77 (s, 3H), 1.29 (s, 12H).

[0384] Intermediate Example 4 Preparation of tert-butyl 2-bromo-5-nitrobenzoate [ka] To a mixture of 2-bromo-5-nitrobenzoic acid (9.70 g, 39.4 mmol, 1.0 equiv) in DCM (100 mL) was added DCC (8.95 g, 43.4 mmol, 8.77 mL, 1.1 equiv) and DMAP (2.41 g, 19.7 mmol, 0.5 equiv) at 0 °C. The mixture was stirred at 0 °C for 10 min, then t-BuOH (4.38 g, 59.1 mmol, 5.66 mL, 1.5 equiv) was added and stirred at 20 °C for 12 h. The mixture was poured into ice water (w / w = 1 / 1) (50 mL) and stirred for 10 min. The aqueous phase was extracted with DCM (50 mL × 2). The combined organic phase was washed with brine (10 mL × 2), dried over anhydrous Na2SO4, filtered, and concentrated under high vacuum. The residue was purified by silica gel chromatography (column height: 250 mm, diameter: 100 mm, 100-200 mesh silica gel, petroleum ether / ethyl acetate = 1 / 0, 5 / 1) to give tert-butyl 2-bromo-5-nitro-benzoate (9.2 g, 30.5 mmol, yield 77.23%) as a white solid. 1 H NMR (400MHz, CDCl3) δ 8.52 (d, J = 2.8 Hz, 1H), 8.13 (dd, J = 2.8, 8.8 Hz, 1H), 7.83 (d, J = 8.8 Hz, 1H), 1.65 (s, 9H).

[0385] Intermediate Example 5 Preparation of tert-butyl 2-[(E)-3-methoxy-3-oxo-prop-1-enyl]-5-nitro-benzoate [ka] A mixture of methyl (E)-3-(4,4,5,5-tetramethyl-1,3,2-dioxaborolan-2-yl)prop-2-enoate (11.2 g, 53.0 mmol, 2.5 equiv), tert-butyl 2-bromo-5-nitrobenzoate (6.40 g, 21.2 mmol, 1.0 equiv), KPO (6.74 g, 31.8 mmol, 1.5 equiv), s-Phos (870 mg, 2.12 mmol, 0.1 equiv), Pd(dba) (970 mg, 1.06 mmol, 0.05 equiv) in dioxane (120 mL) and HO (25 mL) was degassed and purged with N three times, then the mixture was stirred under N at 90 °C for 12 h. The mixture was poured into ice water (w / w = 1 / 1) (30 mL) and stirred for 10 minutes. The aqueous phase was extracted with ethyl acetate (50 mL × 3). The combined organic phases were washed with brine (10 mL × 3), dried over anhydrous Na2SO4, filtered, and concentrated under high vacuum. The residue was purified by silica gel chromatography (column height: 250 mm, diameter: 100 mm, 100-200 mesh silica gel, petroleum ether / ethyl acetate = 1 / 0, 3 / 1) to give tert-butyl 2-[(E)-3-methoxy-3-oxo-prop-1-enyl]-5-nitro-benzoate (6 g, 19.5 mmol, 92.17% yield) as a yellow solid. 1 H NMR(400MHz,CDCl3)δ 8.74(d,J=2.4Hz,1H),8.41(d,J=15.6Hz,1H),8.34(dd,J=2.4,8.4Hz,1H) ,7.72(d,J=8.4Hz,1H),6.38(d,J=15.6Hz,1H),3.85(s,3H),1.65(s,9H).

[0386] Intermediate Example 6 Preparation of tert-butyl 5-amino-2-(3-methoxy-3-oxo-propyl)benzoate [ka] A mixture of tert-butyl 2-[(E)-3-methoxy-3-oxoprop-1-enyl]-5-nitrobenzoate (4.00 g, 13.0 mmol, 1.0 equiv.), Pd / C (400 mg, 13.0 mmol, 10% purity, 1.0 equiv.) in methanol (50 mL) was degassed and purged with H (26.2 mg, 13.0 mmol) three times, then the mixture was stirred under an H atmosphere at 25° C. for 4 h. The mixture was filtered and concentrated under high vacuum to give tert-butyl 5-amino-2-(3-methoxy-3-oxopropyl)benzoate (3.50 g, 12.5 mmol, 96.26% yield) as a yellow oil. 1 H NMR(400MHz,CDCl3)δ 7.14(d,J=2.4Hz,1H),7.04(d,J=8.0Hz,1H),6.73(dd,J=2.4,8.0Hz,1H),3.67(s,3H),3.12(t,J=8.0Hz,2H),2.61(t,J=8.0Hz,2H),1.59(s,9H).

[0387] Intermediate Example 7 Preparation of 5-amino-2-(3-methoxy-3-oxo-propyl)benzoic acid [ka] To a solution of tert-butyl 5-amino-2-(3-methoxy-3-oxo-propyl)benzoate (3.50 g, 12.5 mmol, 1.0 equiv) in ethyl acetate (10 mL) was added HCl / ethyl acetate (4 M, 50 mL, 16.0 equiv), followed by stirring for 12 hours at 25° C. The mixture was concentrated under high vacuum to afford 5-amino-2-(3-methoxy-3-oxo-propyl)benzoic acid (3.20 g, 12.3 mmol, 98.35% yield, HCl) as a white solid. 1 H NMR(400MHz,DMSO-d6)δ 7.69(d,J=2.0Hz,1H),7.39-7.31(m,2H),3.57(s,3H),3.15(t,J=7.6Hz,2H),2.59(t,J=7.6Hz,2H).

[0388] Intermediate Example 8 Preparation of methyl 3-[4-amino-2-[[2-[(2-TERT-butoxy-2-oxo-ethyl)amino]-2-oxo-ethyl]carbamoyl]phenyl]propanoate [ka] To a solution of 5-amino-2-(3-methoxy-3-oxopropyl)benzoic acid (3.20 g, 12.3 mmol, 1.0 equiv., HCl) in DMF (40 mL), NMM (3.74 g, 37.0 mmol, 4.06 mL, 3.0 equiv.), HOBt (833 mg, 6.16 mmol, 0.5 equiv.), EDCI (4.72 g, 24.7 mmol, 2.0 equiv.), and tert-butyl 2-[(2-aminoacetyl)amino]acetate (2.78 g, 14.8 mmol, 1.2 equiv.) were added at 0 °C and stirred at 20 °C for 1 h. The mixture was poured into ice water (w / w = 1 / 1) (40 mL) and stirred for 10 min. The aqueous phase was extracted with ethyl acetate (50 mL × 3). The combined organic phase was washed with brine (20 mL × 2), dried over anhydrous NaSO, filtered, and concentrated under high vacuum. The residue was purified by silica gel chromatography (column height: 250 mm, diameter: 100 mm, 100-200 mesh silica gel, petroleum ether / ethyl acetate = 1 / 0, 0 / 1) to give methyl 3-[4-amino-2-[[2-[(2-tert-butoxy-2-oxo-ethyl)amino]-2-oxo-ethyl]carbamoyl]phenyl]propanoate (3.8 g, 9.66 mmol, 78.38% yield) as a yellow oil. 1 H NMR(400MHz,CDCl3)δ 7.04(d,J=8.0Hz,2H),6.79-6.66(m,3H),4.15(d,J=5.6Hz,2H),3.97(d,J=5.2 Hz,2H),3.61(s,3H),2.99(t,J=7.2Hz,2H),2.68(t,J=7.2Hz,2H),1.47(s,9H).

[0389] Intermediate Example 9 Preparation of 2-[[2-[[5-amino-2-(3-methoxy-3-oxo-propyl)benzoyl]amino]acetyl]amino]acetic acid [ka] To a solution of methyl 3-[4-amino-2-[[2-[(2-tert-butoxy-2-oxo-ethyl)amino]-2-oxo-ethyl]carbamoyl]phenyl]propanoate (0.60 g, 1.53 mmol, 1.0 equiv) in ethyl acetate (5 mL) was added HCl / ethyl acetate (4 M, 10 mL, 26.2 equiv), followed by stirring at 20° C. for 12 h. The mixture was concentrated under high vacuum to give 2-[[2-[[5-amino-2-(3-methoxy-3-oxo-propyl)benzoyl]amino]acetyl]amino]acetic acid (550 mg, 1.47 mmol, 96.48% yield, HCl) as a white solid. 1 H NMR(400MHz,DMSO-d6)δ 8.57(t,J=6.0Hz,1H),8.25(t,J=6.4Hz,1H),7.28(d,J=8.0Hz,1H),7.20-7.10(m,2H),3.89(d, J=6.0Hz,2H),3.81(d,J=6.0Hz,2H),3.57(s,3H),2.90(t,J=8.0Hz,2H),2.60(t,J=8.0Hz,2H).

[0390] Intermediate Example 10 Preparation of Intermediate 1A [ka] 2-[[2-[[5-amino-2-(3-methoxy-3-oxo-propyl)benzoyl]amino]acetyl]amino]acetic acid (320 mg, 856 μmol, 1.0 equiv., HCl), tert-butyl A mixture of 3-[2-[2-[2-[2-[2-[2-[2-[2-[2-[2-[2-[2-[2-[2-(2-aminoethoxy)ethoxy]ethoxy]ethoxy]ethoxy]ethoxy]ethoxy]ethoxy]ethoxy]ethoxy]ethoxy]ethoxy]ethoxy]ethoxy]ethoxy]ethoxy]propanoate (576 mg, 856 μmol, 1.0 equiv), EDCI (328 mg, 1.71 mmol, 2.0 equiv), HOBt (57.8 mg, 428 μmol, 0.5 equiv) and 4-methylmorpholine (259 mg, 2.57 mmol, 282 μL, 3.0 equiv) in DMF (5 mL) was stirred at 25° C. for 2 h. The reaction mixture was quenched at 0° C. by adding HO (10 mL) and then extracted with DCM / i-PrOH (v:v=3:1, 10 mL×5). The combined organic layers were concentrated under reduced pressure to give a residue. The desired product (950 mg, crude) was obtained as a yellow oil. [M+H] + (ESI):994.10.

[0391] Intermediate Example 11 Preparation of intermediate 2A [ka] To a solution of intermediate 1a (950 mg, 956 μmol, 1.0 equiv.) in ethyl acetate (5 mL) was added HCl / ethyl acetate (15 mL, 4 M). The mixture was stirred at 25° C. for 2 hours. The reaction mixture was concentrated under reduced pressure to give the desired product (890 mg, crude) as a yellow solid.

[0392] In a separate reaction, to a solution of intermediate 1a (850 mg, 956 μmol, 1.0 equiv.) in dioxane (5.0 mL) was added hydrogen chloride (gas) / ethyl acetate (15 mL, 4 mol / L). The mixture was stirred at 25° C. for 2 hours. The reaction mixture was concentrated under reduced pressure to give intermediate 2a (800 mg, crude) as a yellow solid. [M+H] + (ESI):936.20.

[0393] Intermediate Example 12 Preparation of Intermediate 3A [ka] A mixture of Intermediate 2 (880 mg, 903 μmol, 1.0 equiv., HCl), di-tert-butyl (2S)-2-aminopentanedioate (588 mg, 1.99 mmol, 2.2 equiv., HCl), EDCI (346 mg, 1.81 mmol, 2.0 equiv.), HOBt (61.0 mg, 451 μmol, 0.5 equiv.), and 4-methylmorpholine (274 mg, 2.71 mmol, 298 μL, 3.0 equiv.) in DMF (10 mL) was stirred at 25° C. for 2 h. The reaction mixture was quenched at 0° C. by adding HO (20 mL) and then extracted with DCM / i-PrOH (v:v=3:1, 20 mL×5). The combined organic layers were concentrated under reduced pressure to give a residue. The desired product (900 mg, 763 μmol, 84.49% yield) was obtained as a yellow oil.

[0394] In another reaction, a mixture of Intermediate 2a (800 mg, 0.903 mmol, 1.0 equiv.), di-tert-butyl (2S)-2-aminopentanedioate (588 mg, 1.99 mmol, 2.2 equiv., HCl), 1-ethyl-3(3-dimethylpropylamine)carbodiimide (346 mg, 1.81 mmol, 2.0 equiv.), 1-hydroxybenzotriazole (61.0 mg, 0.451 mmol, 0.5 equiv.), and 4-methylmorpholine (274 mg, 2.71 mmol, 3.0 equiv.) in N,N-dimethylformamide (10.0 mL) was stirred at 25° C. for 2 hours. The reaction mixture was quenched at 0° C. by adding water (20 mL) and then extracted with methylene chloride / isopropyl alcohol (v:v=3:1, 50 mL×5). The combined organic layers were concentrated under reduced pressure to give a residue. Column, C 18 Silica gel; mobile phase, acetonitrile in water (0.1% trifluoroacetic acid / formic acid), gradient 10% to 70% over 20 min; detector, UV 254 nm. Obtained 900 mg (89%) of intermediate 3a as a yellow oil. [M+H] + (ESI):1179.05

[0395] Intermediate Example 13 Preparation of Intermediate 4A [ka] To a solution of intermediate 3a (800 mg, 679 μmol, 1.0 equiv) in methanol (2.5 mL) and HO (2.5 mL), NaOH (136 mg, 3.39 mmol, 5.0 equiv) was added and then stirred at 20 °C for 1 h. The mixture was concentrated under high vacuum to remove methanol, then diluted with HO (20 mL), and the pH of the aqueous phase was adjusted to approximately 7 with HCl (2 M) and extracted with DCM / i-PrOH (v:v = 3:1, 15 mL × 5). The combined organic layers were dried over anhydrous NaSO, filtered, and concentrated under reduced pressure to give a residue that afforded the desired product (700 mg, crude) as a red oil.

[0396] In a separate reaction, sodium hydroxide (136 mg, 3.39 mmol, 5.0 equiv.) was added to a solution of intermediate 3a (800 mg, 0.68 mmol, 1.0 equiv.) in methanol (8.0 mL) and water (2.5 mL), followed by stirring at 0° C. for 2 hours. The mixture was concentrated under high vacuum to remove methanol, then diluted with water (20 mL). The pH of the aqueous phase was adjusted to approximately 7 with hydrochloric acid (2 mol / L) and extracted with methylene chloride / isopropyl alcohol (v / v=3:1, 30 mL×5). The combined organic layers were dried over sodium sulfate, filtered, and concentrated under reduced pressure to give a residue that afforded intermediate 4a (700 mg, crude) as a red oil. [M+H] + (ESI):1165.10.

[0397] Intermediate Example 14 Preparation of Intermediate 5A [ka] To a solution of intermediate 4a (250 mg, 214 μmol, 1.0 equiv.) and methyl 2,5-dioxopyrrole-1-carboxylate (49.9 mg, 322 μmol, 49.9 μL, 1.5 equiv.) in DCM (3 mL), TEA (65.1 mg, 644 μmol, 89.6 μL, 3.0 equiv.) was added and stirred at 50° C. for 4 h. The pH of the mixture was adjusted to approximately 6 with TFA, diluted with HO (10 mL), and extracted with DCM:i-PrOH (v:v=3:1, 10 mL×5). The combined organic layer was concentrated under reduced pressure to give a residue. The desired product (275 mg, crude) was obtained as a colorless oil.

[0398] In another reaction, trimethylamine (182.2 mg, 1.80 mmol, 3.0 equiv.) was added to a solution of intermediate 4a (700 mg, 0.60 mmol, 1.0 equiv.) and methyl 2,5-dioxopyrrole-1-carboxylate (139.8 mg, 0.92 mmol, 1.5 equiv.) in dichloromethane (7.0 mL), followed by stirring at 50° C. for 4 hours. The pH of the mixture was adjusted to approximately 6 with trifluoroacetic acid, diluted by the addition of water (10 mL), and extracted with methylene chloride:isopropyl alcohol (v / v=3:1, 30 mL×5). The combined organic layer was concentrated under reduced pressure to give a residue. Column, C 18 Silica gel; mobile phase, acetonitrile in water (0.1% trifluoroacetic acid / formic acid), gradient 10% to 70% in 20 min; detector, UV 254 nm. 306 mg (41%) of intermediate 5a was obtained as a colorless oil. [M+H] + (ESI):1245.32.

[0399] Intermediate Example 15 Preparation of Intermediate 6A [ka] A mixture of intermediate 5a (270 mg, 216 μmol, 1.0 equiv.) and EDCI (166 mg, 867 μmol, 4.0 equiv.) in DCM (2 mL) and DMA (2 mL) was stirred at 25 °C for 2 h. The reaction mixture was concentrated under reduced pressure to remove DCM. The residue was purified by preparative HPLC (column: Phenomenex Luna 80 30 mm 3 μm; mobile phase: [water (TFA)-acetonitrile]; B%: 35% to 70%, 8 min). The eluate was removed by lyophilization. The desired product (90 mg, 64.6 μmol, 29.79% yield) was obtained as a colorless oil. 1 H NMR (DMSO-d6, 400 MHz) δ 7.94-7.84(m,1H),7.50(d,J=8.4Hz,1H),7.43-7.34(m,2H),7.18(s,2H),4.1 3(dd,J=5.2,9.1Hz,1H),3.89(s,2H),3.70(s,2H),3.60-3.58(m,3H),3.51-3. 47(m,44H),3.39(t,J=6.0Hz,2H),3.22-3.17(m,2H),3.14(s,3H),2.42-2.32( m, 2H), 2.27-2.19 (m, 2H), 1.95-1.83 (m, 1H), 1.77-1.65 (m, 1H), 1.38 (s, 18H).

[0400] In a separate reaction, a mixture of 2,3,5,6-tetrafluorophenol (108 mg, 0.65 mmol, 3.0 equiv.), Intermediate 5a (270 mg, 216 μmol, 1.0 equiv.), and 1-ethyl-3(3-dimethylpropylamine)carbodiimide (166 mg, 867 μmol, 4.0 equiv.) in N,N-dimethylformamide (3.0 mL) was stirred at 25° C. for 2 h. The reaction mixture was concentrated under reduced pressure. The crude product was purified by flash preparative HPLC with the following conditions (IntelFlash-1): Column, C 18 Silica gel; mobile phase, water and acetonitrile (10.0% acetonitrile up to 100.0% in 20 min); detector, UV 254 nm. 90 mg (30%) of intermediate 6a was obtained as a colorless oil. [M+H] + (ESI):1394.45.

[0401] Additionally, intermediate 6a can be deprotected according to the following reaction scheme and protocol. [ka] Under an inert atmosphere of nitrogen, trifluoroacetic acid (0.25 mL) was added to a solution of intermediate 6a (90 mg, 0.07 mmol, 1.0 equiv) in dichloromethane (1.0 mL) at 0° C. The resulting solution was stirred at 0° C. for 4 hours, concentrated under high vacuum, and concentrated under reduced pressure to give 90 mg (crude) of intermediate 6a′ as a yellow oil. [M+H] + (ESI):1279.95.

[0402] Intermediate Example 16 Preparation of Intermediate 7A [ka] To a solution of [4-[[(2S)-2-[[(2S)-2-amino-3-methylbutanoyl]amino]-5-ureidopentanoyl]amino]phenyl]methyl 3-[[2-amino-4-(dipropylcarbamoyl)-3H-1-benzazepine-8-carbonyl]amino]-7,8-dihydro-5H-1,6-naphthyridine-6-carboxylate (20.0 mg, 23.0 μmol, 1.0 equiv.) in DMF (0.5 mL) was added diisopropylethylamine (DIEA) (8.95 mg, 69.2 μmol, 12.0 μL, 3.0 equiv.) and Intermediate 6a (32.1 mg, 23.0 μmol, 1.0 equiv.). The mixture was stirred at 25° C. for 1 hour. The pH of the mixture was adjusted to about 6 with trifluoroacetic acid (TFA) at 0° C., diluted by adding HO (5 mL), and extracted with dichloromethane (DCM):isopropyl alcohol (i-PrOH) (v:v=3:1, 5 mL×5). The combined organic layer was concentrated under reduced pressure to give a residue. The desired product (55 mg, crude) was obtained as a yellow oil.

[0403] Intermediate Example 17 Synthesis of (19S,22R,23R,23AR,25R,27AR,29R,210R,210AR,212S,214AR,37R,E)-23,210-difluoro-25,212-dimercapto-23,23A,27A,29,210,210A,214,214A-octahydro-19H,22H,27H,37H-4,9-diaza-1(9,6)-purina-3(7,4)-pyrrolo[2,3-D]pyrimidina-2(2,9)-difuro[3,2-D:3',2'-J][1,3,7,9]tetraoxa[2,8]diphosphacyclododecine acyclononaphan-6-ene 25,212-dioxide [ka] [ka] [ka] [ka] [ka] Intermediate 1b (alternatively, in its ammonium salt form) can be obtained using the reaction conditions and synthesis scheme shown above. The reaction methods, reagents, purification techniques, etc. are known in the art. For example, certain embodiments of methods for preparing compounds of the present disclosure can be found in PCT Publication Nos. 2018 / 152453 and / or 2018 / 152450, the entire contents of which are incorporated herein by reference.

[0404] (2R,3R,4R,5R)-5-(4-amino-7H-pyrrolo[2,3-d]pyrimidin-7-yl)-4-fluoro-2-(hydroxymethyl)tetrahydrofuran-3-ol was protected using Bz-Cl and pyridine to give N-(7-((2R,3R,4R,5R)-3-fluoro-4-hydroxy-5-(hydroxymethyl)tetrahydrofuran-2-yl)-7H-pyrrolo[2,3-d]pyrimidin-4-yl)benzamide, which was then protected using TBS-Cl and imidazole in DMF. After cooling to 0° C., the reaction mixture was warmed to 40° C. The resulting product is then reacted with N-(9-((2R,3R,4R,5R)-5-((bis(4-methoxyphenyl)(phenyl)methoxy)methyl)-4-((tert-butyldimethylsilyl)oxy)-3-fluorotetrahydrofuran-2-yl)-9H-purin-6-yl)-N-((E)-4-hydroxybut-2-en-1-yl)benzamide using DIAD and triphenylphosphine in THF, the reaction was cooled to 0° C., and the reaction mixture was allowed to warm to room temperature over time. The free OH group of the resulting product is then converted to a phosphonate under basic conditions using the reagent shown. The DMT protecting group of the resulting product is then removed with dichloroacetic acid, water, and aqueous sodium bicarbonate. The deprotected product is then reacted to form an intramolecular bond with the phosphonate group as shown. A sulfur functionality is then added to the resulting product using sulfur and trimethylamine. The TBS protecting group is then removed using the reagent shown. The resulting product is then functionalized with a 2-nitrobenzyl group. The resulting product is then converted to a phosphoramidite in dichloromethane (diluted with acetonitrile). The phosphoramidite product then forms an intramolecular bond via the free OH group as shown. The coupled product is then converted to a sulfur-containing product. The resulting product is reacted with benzenethiol, trimethylamine, and dioxane. The resulting product is then deprotected to give the desired compound, which is used as the starting material in Reaction Scheme 2 below.

[0405] Reaction Scheme 2 [ka] [ka] [ka]

[0406] Intermediate Example 18 Preparation of intermediate 2B [ka] To a solution of intermediate 1b (100 mg, 134 μmol, 1 equiv.) in DCM (6 mL) were added pyridine (318 mg, 4.02 mmol, 325 μL, 30 equiv.) and 2-[tert-butoxycarbonyl(methyl)amino]ethyl carbonochloridate (542 mg, 2.28 mmol, 17 equiv.). The mixture was stirred at 25° C. for 1 h. The mixture was concentrated under high vacuum. The residue was purified by preparative HPLC (column: C 18-4 Purification by HPLC (150 × 30 mm × 5 μm; mobile phase: [A: 0.1 M water (TFA) B: acetonitrile]; B%: 15%–45%, 20 min) afforded the desired product (50 mg, 47.7 μmol, 35.57% yield, TEA) as a white solid.

[0407] Intermediate Example 19 Preparation of Intermediate 3B [ka] To a solution of intermediate 2b (100 mg, 95.4 μmol, 1.0 equiv., TEA) in acetonitrile (0.5 mL) and HO (0.5 mL) was added TFA (544 mg, 4.77 mmol, 353 μL, 50 equiv.). The mixture was stirred at 50° C. for 1 h. The mixture was concentrated under high vacuum to give the desired product (84 mg, 87.4 μmol, 91.63% yield, TFA) as a white solid.

[0408] Intermediate Example 20 Preparation of Intermediate 5B [ka] [ka] To a solution of intermediate 3b (40 mg, 41.6 μmol, 1 equiv., TFA) in DMF (0.2 mL) was added DIEA (16.1 mg, 125 μmol, 21.8 μL, 3.0 equiv.) and intermediate 4b (40.3 mg, 62.5 μmol, 1.5 equiv.). The mixture was stirred at 0° C. for 1 h. The pH of the mixture was adjusted to approximately 6 with TFA and then concentrated under high vacuum. The residue was diluted with HO (5 mL), and the aqueous phase was extracted with ethyl acetate (5 mL) to remove by-products. The aqueous phase was lyophilized to give the desired product (50 mg, 37.0 μmol, 88.81% yield) as a white solid.

[0409] Intermediate Example 21 Preparation of Intermediate 6B [ka] To a solution of intermediate 5b (50 mg, 37.0 μmol, 1.0 equiv) in acetonitrile (0.5 mL) and HO (0.5 mL) was added TFA (211 mg, 1.85 mmol, 137 μL, 50 equiv). The mixture was stirred at 50° C. for 2 h. The mixture was concentrated under high vacuum. The residue was purified by preparative HPLC (column: C 18-4 Purification by HPLC (150 30 mm 5 μm; mobile phase: [A: 0.1 M water (TFA) B: acetonitrile]; B%: 10% to 40%, 20 min) afforded the desired product (20 mg, 14.6 μmol, 39.59% yield, TFA) as a white solid.

[0410] Intermediate Example 22 Preparation of Intermediate 8B [ka] To a solution of intermediate 6b (9.00 mg, 6.59 μmol, 1.0 equiv, TFA) in DMF (0.1 mL) was added DIEA (2.55 mg, 19.8 μmol, 3.44 μL, 3.0 equiv) and intermediate 7b (9.18 mg, 6.59 μmol, 1 equiv) at 0° C. The mixture was stirred at 20° C. for 1 h. The pH of the mixture was adjusted to approximately 6 with TFA at 0° C. The mixture was concentrated under high vacuum to give the desired product (15 mg crude) as a colorless oil.

[0411] Intermediate Example 23 Synthesis of 4-[[TERT-butyl(dimethyl)silyl]oxymethyl]aniline [ka]

[0412] To a solution of (4-aminophenyl)methanol (5.00 g, 40.6 mmol, 1.0 equiv) in DCM (50 mL) was added imidazole (4.15 g, 60.9 mmol, 1.5 equiv) and TBSCl (7.34 g, 48.7 mmol, 5.97 mL, 1.2 equiv) at 0 °C, followed by stirring at 25 °C for 1 h. The reaction mixture was quenched at 0 °C by adding 100 mL of HO and then extracted with DCM (30 mL × 3). The combined organic layers were washed with 50 mL of brine, dried over NaSO, filtered, and concentrated under reduced pressure to give a residue. The residue was purified by flash silica gel chromatography (Biotage®, 80 g SepaFlash® Silica Flash Column, elution with a 50–70% ethyl acetate / petroleum ether gradient at 80 mL / min). 4-[[tert-Butyl(dimethyl)silyl]oxymethyl]aniline (9.6 g, 40.44 mmol, 99.60% yield) was obtained as a colorless oil. 1 H NMR(400MHz,CDCl3)δ 7.13(d,J=8.0Hz,2H),6.67(d,J=8.0Hz,2H),4.64(s,2H),0.94(s,9H),0.10(s,6H);LC / MS[M+H]238.2(calculated value);LC / MS[M+H]238.1(observed value).

[0413] Intermediate Example 24 Synthesis of tert-butyl N-[(1S)-2-[4-[[Tert-butyl(dimethyl)silyl]oxymethyl]anilino]-1-methyl-2-oxo-ethyl]carbamate [ka]

[0414] To a solution of 4-[[tert-butyl(dimethyl)silyl]oxymethyl]aniline (9.34 g, 39.3 mmol, 1 equiv.) and (2S)-2-(tert-butoxycarbonylamino)propanoic acid (8.93 g, 47.2 mmol, 1.2 equiv.) in DCM (50 mL) and methanol (50 mL) was added EEDQ (29.2 g, 118 mmol, 3.0 equiv.), followed by stirring at 25 °C for 1 h. The reaction mixture was quenched at 0 °C by adding 100 mL of HO and then extracted with ethyl acetate (50 mL × 3). The combined organic layers were washed with 50 mL of brine, dried over NaSO, filtered, and concentrated under reduced pressure to give a residue. The residue was purified by flash silica gel chromatography (Biotage®, 120 g SepaFlash® Silica Flash Column, elution with a 15-30% ethyl acetate / petroleum ether gradient at 80 mL / min) to give tert-butyl N-[(1S)-2-[4-[[tert-butyl(dimethyl)silyl]oxymethyl]anilino]-1-methyl-2-oxo-ethyl]carbamate (25 g, crude) as an orange oil.

[0415] Intermediate Example 25 Synthesis of tert-butyl N-[(2S)-2-[bis(tert-butoxycarbonyl)amino]propanoyl]-N-[4-[[TERT-butyl(dimethyl)silyl]oxymethyl]phenyl]carbamate [ka] To a solution of tert-butyl N-[(1S)-2-[4-[[tert-butyl(dimethyl)silyl]oxymethyl]anilino]-1-methyl-2-oxo-ethyl]carbamate (10.0 g, 24.5 mmol, 1.0 equiv.) in DCM (100 mL) was added DMAP (2.99 g, 24.5 mmol, 1.0 equiv.), DIEA (9.49 g, 73.4 mmol, 12.8 mL, 3.0 equiv.), and BocO (16.0 g, 73.4 mmol, 16.9 mL, 3.0 equiv.). The mixture was stirred at 40 °C for 24 h. The reaction mixture was quenched at 0 °C by adding 100 mL of HO and then extracted with DCM (50 mL × 2). The combined organic layers were washed with 100 mL of brine, dried over Na2SO4, filtered, and concentrated under reduced pressure to give a residue. The residue was purified by flash silica gel chromatography (Biotage®, 120 g SepaFlash® Silica Flash Column, eluting with a 10-20% ethyl acetate / petroleum ether gradient at 80 mL / min). Tert-butyl N-[(2S)-2-[bis(tert-butoxycarbonyl)amino]propanoyl]-N-[4-[[tert-butyl(dimethyl)silyl]oxymethyl]phenyl]carbamate (4.70 g, 7.72 mmol, 31.54% yield) was obtained as a colorless oil. 1 H NMR(CDCl3,400MHz)δ 7.31(d,J=8.4Hz,2H),7.11(d,J=8.4Hz,2H),5.48-5.43(m,1H),4.75(s,2H),1.55-1.50(m,27H),1. 02(d,J=6.4Hz,3H),0.94(s,9H),0.08(s,6H);LC / MS[M+Na]631.35(calculated value);LC / MS[M+Na]631.3(observed value).

[0416] Intermediate Example 26 Synthesis of tert-butyl N-[(2S)-2-[bis(tert-butoxycarbonyl)amino]propanoyl]-N-[4-(hydroxymethyl)phenyl]carbamate [ka] To a solution of tert-butyl N-[(2S)-2-[bis(tert-butoxycarbonyl)amino]propanoyl]-N-[4-[[tert-butyl(dimethyl)silyl]oxymethyl]phenyl]carbamate (4.68 g, 7.69 mmol, 1.0 equiv.) in THF (50 mL) was added TBAF (1 M, 15.4 mL, 2.0 equiv.), followed by stirring at 25 °C for 1 h. The reaction mixture was quenched at 0 °C by adding HO (100 mL) and then extracted with ethyl acetate (50 mL × 3). The combined organic layers were washed with 50 mL of brine, dried over NaSO, filtered, and concentrated under reduced pressure to give a residue. The residue was purified by flash silica gel chromatography (Biotage®, 40 g SepaFlash® Silica Flash Column, elution with a 50–70% ethyl acetate / petroleum ether gradient at 60 mL / min). Tert-butyl N-[(2S)-2-[bis(tert-butoxycarbonyl)amino]propanoyl]-N-[4-(hydroxymethyl)phenyl]carbamate (2.10 g, 4.25 mmol, 55.24% yield) was obtained as a colorless oil. 1 H NMR(400MHz,CDCl3)δ 7.36(d,J=8.4Hz,2H),7.15(d,J=8.4Hz,2H),5.49-5.44(m,1H),4.70(d,J=2.8Hz, 2H),1.53(s,21H),1.35(s,9H);LC / MS[M+H]517.2(calculated value);LC / MS[M+H]517.2(observed value).

[0417] Intermediate Example 27 Synthesis of [4-[[(2S)-2-[bis(Tert-butoxycarbonyl)amino]propanoyl]-Tert-butoxycarbonyl-amino]phenyl]methyl(4-nitrophenyl)carbonate [ka] To a solution of tert-butyl N-[(2S)-2-[bis(tert-butoxycarbonyl)amino]propanoyl]-N-[4-(hydroxymethyl)phenyl]carbamate (1.00 g, 2.02 mmol, 1.0 equiv.) in DCM (15 mL) was added DIEA (784 mg, 6.07 mmol, 1.06 mL, 3.0 equiv.) and bis(4-nitrophenyl)carbonate (1.23 g, 4.04 mmol, 2.0 equiv.), followed by stirring at 25 °C for 1 h. The reaction mixture was quenched at 0 °C by adding HO (30 mL) and then extracted with DCM (10 mL × 3). The combined organic layers were washed with 20 mL of brine, dried over NaSO, filtered, and concentrated under reduced pressure to give a residue. The residue was purified by flash silica gel chromatography (Biotage®, 20 g SepaFlash® Silica Flash Column, eluting with a 10-30% ethyl acetate / petroleum ether gradient at 40 mL / min). [4-[[(2S)-2-[bis(tert-butoxycarbonyl)amino]propanoyl]-tert-butoxycarbonyl-amino]phenyl]methyl (4-nitrophenyl)carbonate (1.71 g, crude) was obtained as a colorless oil. LC / MS [M+Na] 682.3 (calculated); LC / MS [M+Na] 682.2 (observed).

[0418] Intermediate Example 28 Synthesis of tert-butyl N-[(2S)-2-[bis(tert-butoxycarbonyl)amino]propanoyl]-N-[4-(ethylcarbamoyloxymethyl)phenyl]carbamate [ka] To a solution of ethanamine (315 mg, 3.87 mmol, 457 μL, 1.5 equiv, HCl) in DMF (15 mL) was added DIEA (1.67 g, 12.9 mmol, 2.24 mL, 5.0 equiv) and [4-[[(2S)-2-[bis(tert-butoxycarbonyl)amino]propanoyl]-tert-butoxycarbonyl-amino]phenyl]methyl(4-nitrophenyl)carbonate (1.70 g, 2.58 mmol, 1.0 equiv) at 0° C. The mixture was stirred at 25° C. for 1 h. The reaction mixture was quenched at 0° C. by adding HO (20 mL) and then extracted with ethyl acetate (20 mL × 3). The combined organic layers were washed with 20 mL of brine, dried over NaSO, filtered, and concentrated under reduced pressure to give a residue. The residue was purified by flash silica gel chromatography (Biotage®, 20 g SepaFlash® Silica Flash Column, elution with a 30-50% ethyl acetate / petroleum ether gradient at 40 mL / min). Tert-butyl N-[(2S)-2-[bis(tert-butoxycarbonyl)amino]propanoyl]-N-[4-(ethylcarbamoyloxymethyl)phenyl]carbamate (1.14 g, 2.02 mmol, 78.21% yield) was obtained as a white solid. LC / MS [M+Na] 588.3 (calculated); LC / MS [M+Na] 588.3 (observed).

[0419] Intermediate Example 29 Synthesis of tert-butyl N-[(2S)-2-[bis(tert-butoxycarbonyl)amino]propanoyl]-N-[4-[[chloromethyl(ethyl)carbamoyl]oxymethyl]phenyl]carbamate [ka] To a solution of tert-butyl N-[(2S)-2-[bis(tert-butoxycarbonyl)amino]propanoyl]-N-[4-(ethylcarbamoyloxymethyl)phenyl]carbamate (1.00 g, 1.77 mmol, 1.0 equiv.) in toluene (10 mL) was added paraformaldehyde (3.15 g, 2.65 mmol, 1.5 equiv.) and 1-chloro-N,N,2-trimethyl-prop-1-en-1-amine (1.18 g, 8.84 mmol, 1.17 mL, 5.0 equiv.). The mixture was stirred at 80° C. for 1 hour. The reaction mixture was filtered and concentrated under reduced pressure to give a residue. The compound tert-butyl N-[(2S)-2-[bis(tert-butoxycarbonyl)amino]propanoyl]-N-[4-[[chloromethyl(ethyl)carbamoyl]oxymethyl]phenyl]carbamate (2 g, crude) was obtained as a colorless oil.

[0420] Intermediate Example 30 Synthesis of tert-butyl N-[(2S)-2-[bis(tert-butoxycarbonyl)amino]propanoyl]-N-[4-[[[2-[(1S,2S,4R,6R,8S,9S,11S,12R,13S,19S)-12,19-difluoro-6-(3-fluorophenyl)-11-hydroxy-9,13-dimethyl-16-oxo-5,7-dioxapentacyclo[10.8.0.02,9.04,8.013,18]icosa-14,17-dien-8-yl]-2-oxo-ethoxy]methyl-ethyl-carbamoyl]oxymethyl]phenyl]carbamate [ka] To a solution of tert-butyl N-[(2S)-2-[bis(tert-butoxycarbonyl)amino]propanoyl]-N-[4-[[chloromethyl(ethyl)carbamoyl]oxymethyl]phenyl]carbamate (1.18 g, 1.93 mmol, 2.0 equiv.) in DCM (15 mL) was added DIEA (371 mg, 2.87 mmol, 0.5 mL, 2.98 equiv.) and (1S,2S,4R,6R,8S,9S,11S, (12R,13S,19S)-12,19-Difluoro-6-(3-fluorophenyl)-11-hydroxy-8-(2-hydroxyacetyl)-9,13-dimethyl-5,7-dioxapentacyclo[10.8.0.02,9.04,8.013,18]icosa-14,17-dien-16-one (500 mg, 964.28 μmol, 1.0 equiv.) was added, followed by stirring at 25°C for 12 h. The combined organic layers were washed with 20 mL of brine, dried over Na2SO4, filtered, and concentrated under reduced pressure to give a residue. The residue was purified by flash silica gel chromatography (Biotage®, 20 g SepaFlash® Silica Flash Column, elution with a 50-70% ethyl acetate / petroleum ether gradient at 45 mL / min). Tert-butyl N-[(2S)-2-[bis(tert-butoxycarbonyl)amino]propanoyl]-N-[4-[[[2-[(1S,2S,4R,6R,8S,9S,11S,12R,13S,19S)-12,19-difluoro-6-(3-fluorophenyl)-11-hydroxy-9,13-dimethyl-16-oxo-5,7-dioxapentacyclo[10.8.0.02,9.04,8.013,18]icosa-14,17-dien-8-yl]-2-oxo-ethoxy]methyl-ethyl-carbamoyl]oxymethyl]phenyl]carbamate (658 mg, 600.26 μmol, 62.25% yield) was obtained as a colorless oil. LC / MS [M+Na] 1118.5 (calculated); LC / MS [M+Na] 1118.3 (observed).

[0421] Intermediate Example 31 Synthesis of [4-[[(2S)-2-aminopropanoyl]amino]phenyl]methyl N-[[2-[(1S,2S,4R,6R,8S,9S,11S,12R,13S,19S)-12,19-difluoro-6-(3-fluorophenyl)-11-hydroxy-9,13-dimethyl-16-oxo-5,7-dioxapentacyclo[10.8.0.02,9.04,8.013,18]icosa-14,17-dien-8-yl]-2-oxo-ethoxy]methyl]-N-ethyl-carbamate [ka] tert-Butyl N-[(2S)-2-[bis(tert-butoxycarbonyl)amino]propanoyl]-N-[4-[[[2-[(1S,2S,4R,6R,8S,9S,11S,12R,13S,19S)-12,19-difluoro-6-(3-fluorophenyl)-11-hydroxy-9,13-dimethyl-16-oxo-5,7-dioxapentacyclo[3.0]heptyl] To a solution of [10.8.0.02,9.04,8.013,18]icosa-14,17-dien-8-yl]-2-oxo-ethoxy]methyl-ethyl-carbamoyl]oxymethyl]phenyl]carbamate (0.96 g, 875 μmol, 1.0 equiv.) in toluene (15 mL) was added SiO2 (10.0 g, 166 mmol, 190 equiv.). The mixture was stirred at 120 °C for 1 h. The reaction mixture was filtered, the SiO2 was washed with methanol (20 mL × 2), and the filtrate was concentrated under reduced pressure to give a residue. [4-[[(2S)-2-aminopropanoyl]amino]phenyl]methyl N-[[2-[(1S,2S,4R,6R,8S,9S,11S,12R,13S,19S)-12,19-difluoro-6-(3-fluorophenyl)-11-hydroxy-9,13-dimethyl-16-oxo-5,7-dioxapentacyclo[10.8.0.02,9.04,8.013,18]icosa-14,17-dien-8-yl]-2-oxo-ethoxy]methyl]-N-ethyl-carbamate (500 mg, 628.27 μmol, 71.74% yield) was obtained as a colorless oil. LC / MS [M+H] 796.3 (calculated); LC / MS [M+H] 796.4 (observed).

[0422] Intermediate Example 32 Synthesis of [4-[[(2S)-2-[[(2S)-2-amino-3-methyl-butanoyl]amino]propanoyl]amino]phenyl]methyl N-[[2-[(1S,2S,4R,6R,8S,9S,11S,12R,13S,19S)-12,1 9-difluoro-6-(3-fluorophenyl)-11-hydroxy-9,13-dimethyl-16-oxo-5,7-dioxapentacyclo[10.8.0.02,9.04,8.013,18]icosa-14,17-dien-8-yl]-2-oxo-ethoxy]methyl]-N-ethyl-carbamate [ka] [4-[[(2S)-2-Aminopropanoyl]amino]phenyl]methyl N-[[2-[(1S,2S,4R,6R,8S,9S,11S,12R,13S,19S)-12,19-difluoro-6-(3-fluorophenyl)-11-hydroxy-9,13-dimethyl-16-oxo-5,7-dioxapentacyclo[10.8.0.02,9.04,8.013,18]icosa-14,17-dien-8-yl]-2-oxo-ethoxy]methyl]-N-ethyl- To a solution of bamate (200 mg, 251 μmol, 1.0 equiv.) in THF (2 mL), DIEA (97.4 mg, 754 μmol, 131 μL, 3.0 equiv.) and (2,5-dioxopyrrolidin-1-yl)(2S)-2-(9H-fluoren-9-ylmethoxycarbonylamino)-3-methylbutanoate (164 mg, 377 μmol, 1.5 equiv.) were added and stirred at 25°C for 1 h. Piperidine (86.2 mg, 1.01 mmol, 0.10 mL, 5.66 equiv.) was added. The mixture was stirred at 25°C for an additional 1 h. The mixture was purified by preparative HPLC (column: Phenomenex Luna 80 × 30 mm × 3 μm; mobile phase: [water (TFA)-ACN]; B%: 30% to 65%, 8 min). [4-[[(2S)-2-[[(2S)-2-amino-3-methyl-butanoyl]amino]propanoyl]amino]phenyl]methyl N-[[2-[(1S,2S,4R,6R,8S,9S,11S,12R,13S,19S)-12,1 9-difluoro-6-(3-fluorophenyl)-11-hydroxy-9,13-dimethyl-16-oxo-5,7-dioxapentacyclo[10.8.0.02,9.04,8.013,18]icosa-14,17-dien-8-yl]-2-oxo-ethoxy]methyl]-N-ethyl-carbamate (100 mg, 44.69 μmol, 24.97% yield, 40% purity) was obtained as a pale yellow solid. LC / MS [M+H] 895.4 (calculated); LC / MS [M+H] 895.5 (observed).

[0423] Intermediate Example 33 Synthesis of tert-butyl(((9H-fluoren-9-yl)methoxy)carbonyl)-L-alanyl-L-alanylglycinate [ka] Under an inert atmosphere of nitrogen, to a stirred solution of (2S)-2-[(2S)-2-{[(9H-fluoren-9-ylmethoxy)carbonyl]amino}propanamido]propanoic acid (1.0 g, 2.6 mmol, 1.0 equiv.) in N,N-dimethylformamide (5.0 mL) was added tert-butyl 2-aminoacetate (343.0 mg, 2.6 mmol, 1.0 equiv.), N,N-diisopropylethylamine (675.9 mg, 5.2 mmol, 2.0 equiv.), and benzotriazol-1-yl-oxytripyrrolidinophosphonium hexafluorophosphate (530.0 mg, 3.9 mmol, 1.5 equiv.) at 0° C. The resulting solution was stirred at 25° C. for 4 hours and diluted with water (100 mL). The resulting mixture was extracted with dichloromethane (3 x 100 mL), and the organic layer was washed with saturated sodium chloride solution (3 x 100 mL), dried over anhydrous sodium sulfate, filtered, and concentrated under reduced pressure. The crude product was purified by flash preparative HPLC with the following conditions (IntelFlash-1): Column, C 18 Silica gel; mobile phase, water and acetonitrile (10.0% acetonitrile up to 100.0% in 20 min); detector, UV 254 nm. Obtained 1.0 g (77%) of tert-butyl (((9H-fluoren-9-yl)methoxy)carbonyl)-L-alanyl-L-alanylglycinate as a white solid. MS m / z [M+H] + (ESI):496.23.

[0424] Intermediate Example 34 Synthesis of (((9H-fluoren-9-yl)methoxy)carbonyl)-L-alanyl-L-alanylglycine [ka] Under an inert atmosphere of nitrogen, trifluoroacetic acid (10.0 mL) was added to a solution of tert-butyl (((9H-fluoren-9-yl)methoxy)carbonyl)-L-alanyl-L-alanylglycinate (1.0 g, 2.0 mmol, 1.0 equiv.) in dichloromethane (20.0 mL) at 25° C. The resulting solution was stirred at 25° C. for 5 hours and concentrated under high vacuum. The crude product was purified by flash preparative HPLC with the following conditions (IntelFlash-1). Column, C 18 Silica gel; mobile phase, water and acetonitrile (10.0% acetonitrile up to 100.0% in 20 min); detector, UV 254 nm. Obtained 800 mg (90%) of (((9H-fluoren-9-yl)methoxy)carbonyl)-L-alanyl-L-alanylglycine as a yellow oil. MS m / z [M−H] - (ESI):438.17.

[0425] Intermediate Example 35 (5S,8S)-1-(9H-fluoren-9-yl)-5,8-dimethyl-3,6,9-trioxo-2-oxa-4,7,10-triazaundecan-11-yl acetate [ka] To a stirred solution of (((9H-fluoren-9-yl)methoxy)carbonyl)-L-alanyl-L-alanylglycine (800.0 mg, 1.8 mmol, 1.0 equiv.) and cupric acetate monohydrate (33.0 mg, 0.2 mmol, 0.1 equiv.) in N,N-dimethylformamide (10.0 mL) at room temperature under an air atmosphere, acetic acid (164.0 mg, 2.7 mmol, 1.5 equiv.) and lead tetraacetate (4.0 g, 9.0 mmol, 5.0 equiv.) were added. The resulting mixture was stirred at 60°C for an additional 5 hours. The resulting solution was cooled to 25°C and diluted with water (100 mL). The resulting mixture was extracted with dichloromethane (3 × 100 mL), and the organic layer was washed with saturated sodium chloride solution (3 × 100 mL), dried over anhydrous sodium sulfate, filtered, and concentrated under reduced pressure. The crude product was purified by flash preparative HPLC with the following conditions (IntelFlash-1): Column, C18 Silica gel; mobile phase, water and acetonitrile (10.0% acetonitrile up to 100.0% in 20 min); detector, UV 254 nm. 425 mg (51%) of (5S,8S)-1-(9H-fluoren-9-yl)-5,8-dimethyl-3,6,9-trioxo-2-oxa-4,7,10-triazaundecan-11-yl acetate was obtained as a white solid. MS m / z [M+Na] + (ESI):476.17.

[0426] Intermediate Example 36 Synthesis of (5S,8S)-1-(9H-fluoren-9-yl)-5,8-dimethyl-3,6,9-trioxo-2-oxa-4,7,10-triazaundecan-11-yl acetate [ka] (5S,8S)-1-(9H-fluoren-9-yl)-5,8-dimethyl-3,6,9-trioxo-2-oxa-4,7,10-triazaundecan-11-yl acetate (425.0 mg, 0.94 mmol, 1.0 equiv.) and (2S,6aS,6bR,7S,8aS,8bS,10R,11aR,12aS,12bS)-2,6b-difluoro-10-(3-fluorophenyl)-7-hydroxy-8b-(2-hydroxyacetyl)-6a,8a-dimethyl-1,2,6a,6b,7,8,8a,8b,11a,1 To a stirred solution of 2,12a,12b-dodecahydro-4H-naphtho[2',1':4,5]indeno[1,2-d][1,3]dioxol-4-one (243.0 mg, 0.5 mmol, 0.5 equiv.) in dichloromethane (5.0 mL) was added p-toluenesulfonic acid (32.2 mg, 0.2 mmol, 0.20 equiv.) in portions at 0°C under air. The resulting mixture was stirred at 25°C for an additional 12 hours. The crude product was purified under the following conditions (column, C 18Purification was performed by reverse-phase flash on silica gel, mobile phase: acetonitrile in water (10 mmol / L ammonium bicarbonate), gradient 10% to 100% in 20 min, detector: UV 254 nm. 200 mg (47%) of (9H-fluoren-9-yl)methyl ((S)-1-(((S)-1-(((2-((2S,6aS,6bR,7S,8aS,8bS,10R,11aR,12aS,12bS)-2,6b-difluoro-10-(3-fluorophenyl)-7-hydroxy-6a,8a-dimethyl-4-oxo-1,2,4,6 a,6b,7,8,8a,11a,12,12a,12b-dodecahydro-8bH-naphtho[2',1':4,5]indeno[1,2-d][1,3]dioxol-8b-yl)-2-oxoethoxy)methyl)amino)-1-oxopropan-2-yl)amino)-1-oxopropan-2-yl)carbamate was obtained as a white solid. MS m / z [M+H] + (ESI):912.36.

[0427] Intermediate Example 37 Synthesis of (5S,8S)-1-(9H-fluoren-9-yl)-5,8-dimethyl-3,6,9-trioxo-2-oxa-4,7,10-triazaundecan-11-yl acetate [ka] (9H-Fluoren-9-yl)methyl((S)-1-(((S)-1-(((2-((2S,6aS,6bR,7S,8aS,8bS,10R,11aR,12aS,12bS)-2,6b-difluoro-10-(3-fluorophenyl)-7-hydroxy-6a,8a-dimethyl-4-oxo-1,2,4,6a,6b,7,8,8a,11a,12,12a,12b-dodecahydro-8bH-na) To a stirred solution of 200 mg (0.2 mmol, 1.0 equiv.) of 1-oxopropan-2-yl-2-oxoethoxymethylamino)-1-oxopropan-2-ylamino)-1-oxopropan-2-ylcarbamate (200 mg, 0.2 mmol, 1.0 equiv.) in N,N-dimethylformamide (2.0 mL) was added morpholine (0.3 mL) at 25°C under an air atmosphere. The resulting mixture was stirred at 25°C for an additional hour. The crude product was purified under the following conditions (column, C 18 Purification was performed by reverse-phase flash on silica gel, mobile phase: acetonitrile in water (0.1% trifluoroacetic acid), gradient 10% to 100% in 20 min, detector: UV 254 nm. Obtained 100 mg (66%) of (S)-2-amino-N-((S)-1-(((2-((2S,6aS,6bR,7S,8aS,8bS,10R,11aR,12aS,12bS)-2,6b-difluoro-10-(3-fluorophenyl)-7-hydroxy-6a,8a-dimethyl-4-oxo-1,2,4,6a,6b,7,8,8a,11a,12,12a,12b-dodecahydro-8bH-naphtho[2',1':4,5]indeno[1,2-d][1,3]dioxol-8b-yl)-2-oxoethoxy)methyl)amino)-1-oxopropan-2-yl)propanamide as a white solid. MS m / z [M+H] + (ESI):690.29.

[0428] Intermediate Example 38 Synthesis of tert-butyl (((9H-fluoren-9-yl)methoxy)carbonyl)-L-phenylalanylglycinate [ka] Under an inert atmosphere of nitrogen, a solution of (2S)-2-{[(9H-fluoren-9-ylmethoxy)carbonyl]amino}-3-phenylpropanoic acid (5.0 g, 12.9 mmol, 1.0 equiv.) in N,N-dimethylformamide (50.0 mL) was added with tert-butyl 2-aminoacetate hydrochloride (2.6 g, 15.5 mmol, 1.2 equiv.), benzotriazol-1-yl-oxytripyrrolidinophosphonium hexafluorophosphate (10.1 g, 19.4 mmol, 1.5 equiv.), 1-hydroxybenzotriazole (2.6 g, 19.4 mmol, 1.5 equiv.), and N,N-diisopropylethylamine (4.9 g, 38.7 mmol, 3.0 equiv.) at 0° C. The resulting solution was stirred at 25° C. for 4 hours and then diluted with water (1000 mL). The resulting mixture was extracted with dichloromethane (3 x 1000 mL), and the organic layer was washed with saturated sodium chloride solution (3 x 1000 mL), dried over anhydrous sodium sulfate, filtered, and concentrated under reduced pressure. The crude product was purified by flash preparative HPLC with the following conditions (IntelFlash-1): Column, C 18 Silica gel; mobile phase, water and acetonitrile (10.0% acetonitrile up to 100.0% in 20 min); detector, UV 254 nm. 5.2 g (80%) of tert-butyl (((9H-fluoren-9-yl)methoxy)carbonyl)-L-phenylalanylglycinate was obtained as a white solid. MS m / z [M+H] + (ESI):501.23.

[0429] Intermediate Example 39 Synthesis of tert-butyl L-phenylalanylglycinate [ka] Under an inert atmosphere of nitrogen, tert-butyl (((9H-fluoren-9-yl)methoxy)carbonyl)-L-phenylalanylglycinate (5.2 g, 10.4 mmol, 1.0 equiv.) was added to a solution of morpholine / N,N-dimethylformamide (7.0 mL / 49.0 mL). The resulting solution was stirred at 25° C. for 1 hour and diluted with water (1000 mL). The resulting mixture was extracted with dichloromethane (3×1000 mL), and the organic layer was washed with saturated sodium chloride solution (3×1000 mL), dried over anhydrous sodium sulfate, filtered, and concentrated under reduced pressure. The crude product was purified by flash preparative HPLC with the following conditions (IntelFlash-1): Column, C 18 Silica gel; mobile phase, water and acetonitrile (10.0% acetonitrile up to 100.0% in 20 min); detector, UV 254 nm. Obtained 2.7 g (93%) of tert-butyl L-phenylalanylglycinate as a white solid. MS m / z [M+H] + (ESI):279.16.

[0430] Intermediate Example 40 Synthesis of tert-butyl(((9H-fluoren-9-yl)methoxy)carbonyl)glycylglycyl-L-phenylalanylglycinate [ka] Under an inert atmosphere of nitrogen, to a solution of tert-butyl L-phenylalanylglycinate (2.7 g, 9.7 mmol, 1.0 equiv.) in N,N-dimethylformamide (30.0 mL) was added (2-{[(9H-fluoren-9-ylmethoxy)carbonyl]amino}acetamido)acetic acid (3.4 g, 9.7 mmol, 1.0 equiv.), benzotriazol-1-yl-oxytripyrrolidinophosphonium hexafluorophosphate (7.6 g, 14.6 mmol, 1.5 equiv.), 1-hydroxybenzotriazole (1.9 g, 14.6 mmol, 1.5 equiv.), and N,N-diisopropylethylamine (3.8 g, 29.1 mmol, 3.0 equiv.) at 0° C. The resulting solution was stirred at 25° C. for 4 hours and diluted with water (500 mL). The resulting mixture was extracted with dichloromethane (3 x 500 mL), and the organic layer was washed with saturated sodium chloride solution (3 x 500 mL), dried over anhydrous sodium sulfate, filtered, and concentrated under reduced pressure. The crude product was purified by flash preparative HPLC with the following conditions (IntelFlash-1): Column, C 18 Silica gel; mobile phase, water and acetonitrile (10.0% acetonitrile up to 100.0% in 20 min); detector, UV 254 nm. 3.8 g (64%) of tert-butyl (((9H-fluoren-9-yl)methoxy)carbonyl)glycylglycyl-L-phenylalanylglycinate was obtained as a white solid. MS m / z [M+H] + (ESI):615.27.

[0431] Intermediate Example 40 Synthesis of tert-butyl(((9H-fluoren-9-yl)methoxy)carbonyl)glycylglycyl-L-phenylalanylglycylglycinate [ka] Under an inert atmosphere of nitrogen, trifluoroacetic acid (10.0 mL) was added to a solution of tert-butyl (((9H-fluoren-9-yl)methoxy)carbonyl)glycylglycyl-L-phenylalanylglycinate (3.8 g, 6.2 mmol, 1.0 equiv) in dichloromethane (40.0 mL) at 0° C. The resulting solution was stirred at 25° C. for 5 hours and concentrated under high vacuum. To this solution in N,N-dimethylformamide (30.0 mL) was added tert-butyl 2-aminoacetate hydrochloride (1.0 g, 6.2 mmol, 1.0 equiv.), benzotriazol-1-yl-oxytripyrrolidinophosphonium hexafluorophosphate (4.8 g, 9.3 mmol, 1.5 equiv.), 1-hydroxybenzotriazole (1.3 g, 9.3 mmol, 1.5 equiv.), and N,N-diisopropylethylamine (2.4 g, 18.6 mmol, 3.0 equiv.) at 0 °C. The resulting solution was stirred at 25 °C for 4 h and diluted with water (300 mL). The resulting mixture was extracted with dichloromethane (3 × 500 mL), and the organic layer was washed with saturated sodium chloride solution (3 × 500 mL), dried over anhydrous sodium sulfate, filtered, and concentrated under reduced pressure. The crude product was purified by flash preparative HPLC with the following conditions (IntelFlash-1): Column, C 18 Silica gel; mobile phase, water and acetonitrile (10.0% acetonitrile up to 100.0% in 20 min); detector, UV 254 nm. 2.6 g (64%) of tert-butyl (((9H-fluoren-9-yl)methoxy)carbonyl)glycylglycyl-L-phenylalanylglycylglycinate was obtained as a white solid. MS m / z [M+H] + (ESI):672.30; 1 H NMR(300MHz,DMSO-d6) δ:1.39(s,9H),2.75-2.82(m,1H),3.03-3.09(m,1H),3.56-3.80(m,8H),4.19-4.31(m,3H),4.49-4.57(m,1H),7.15-7.24 (m,7H),7.25-7.44(m,2H),7.57-7.61(m,1H),7.69-7.72(m,2H),7.88-7.99(m,2H),8.01-8.15(m,3H),8.31-8.35(m,1H).

[0432] Intermediate Example 40 Synthesis of (((9H-fluoren-9-yl)methoxy)carbonyl)glycylglycyl-L-phenylalanylglycylglycine [ka] Under an inert atmosphere of nitrogen, trifluoroacetic acid (15.0 mL) was added to a solution of tert-butyl (((9H-fluoren-9-yl)methoxy)carbonyl)glycylglycyl-L-phenylalanylglycylglycinate (2.6 g, 3.9 mmol, 1.0 equiv.) in dichloromethane (30.0 mL) at 0° C. The resulting solution was stirred at 25° C. for 5 hours and concentrated under high vacuum. The crude product was purified by flash preparative HPLC with the following conditions (IntelFlash-1). Column, C 18 Silica gel; mobile phase, water (containing 0.1% trifluoroacetic acid) and acetonitrile (10.0% acetonitrile up to 100.0% in 20 min); detector, UV 254 nm. 2.1 g (88%) of (((9H-fluoren-9-yl)methoxy)carbonyl)glycylglycyl-L-phenylalanylglycylglycine was obtained as a white solid. MS m / z [M+H] + (ESI):616.20.

[0433] Intermediate Example 41 Synthesis of (S)-11-benzyl-1-(9H-fluoren-9-yl)-3,6,9,12,15-pentaoxo-2-oxa-4,7,10,13,16-pentaazaheptadecan-17-yl acetate [ka] Under an inert atmosphere of nitrogen, cupric acetate anhydride (254 mg, 1.4 mmol, 0.4 equiv.), acetic acid (468 mg, 7.8 mmol, 2.3 equiv.), and lead tetrachloride (1.8 g, 4.1 mmol, 1.2 equiv.) were added to a solution of (((9H-fluoren-9-yl)methoxy)carbonyl)glycylglycyl-L-phenylalanylglycylglycine (2.1 g, 3.4 mmol, 1.0 equiv.) in N,N-dimethylformamide (30.0 mL) at 25° C. The resulting mixture was stirred at 60° C. for 5 hours. The mixture was cooled to 25° C., diluted with water (300 mL), extracted with dichloromethane (3×500 mL), and the organic layer was washed with saturated sodium chloride solution (3×500 mL), dried over anhydrous sodium sulfate, filtered, and concentrated under reduced pressure. The crude product was purified by flash preparative HPLC with the following conditions (IntelFlash-1): Column, C 18 Silica gel; mobile phase, water and acetonitrile (10.0% acetonitrile up to 100.0% in 20 min); detector, UV 254 nm. 870 mg (41%) of (S)-11-benzyl-1-(9H-fluoren-9-yl)-3,6,9,12,15-pentaoxo-2-oxa-4,7,10,13,16-pentaazaheptadecan-17-yl acetate was obtained as a white solid. MS m / z [M+H] + (ESI):630.20.

[0434] Intermediate Example 42 Synthesis of (9H-fluoren-9-yl)methyl ((S)-10-benzyl-1-((2S,6AS,6BR,7S,8AS,8BS,10R,11AR,12AS,12BS)-2,6B-difluoro-10-(3-fluorophenyl)-7-hydroxy-6A,8A-dimethyl-4-oxo-1,2,4,6A,6B,7,8,8A,11A,12,12A,12B-dodecahydro-8BH-naphtho[2',1':4,5]indeno[1,2-D][1,3]dioxol-8B-yl)-1,6,9,12,15-pentaoxo-3-oxa-5,8,11,14-tetraazahexadecan-16-yl)carbamate [ka] To a solution of (S)-11-benzyl-1-(9H-fluoren-9-yl)-3,6,9,12,15-pentaoxo-2-oxa-4,7,10,13,16-pentaazaheptadecan-17-yl (870.0 mg, 1.38 mmol, 3.0 equiv) in dichloromethane (8.0 mL) was added (2S,6aS,6bR,7S,8aS,8bS,10R,11aR,12aS,12bS)-2,6b-difluoro-10-(3-fluorophenyl)-7-hydroxybenzoate. C1-8b-(2-hydroxyacetyl)-6a,8a-dimethyl-1,2,6a,6b,7,8,8a,8b,11a,12,12a,12b-dodecahydro-4H-naphtho[2',1':4,5]indeno[1,2-d][1,3]dioxol-4-one (238.8 mg, 0.46 mmol, 1.0 equiv.) was added under a nitrogen atmosphere at 25 °C, followed by p-toluenesulfonic acid (31.7 mg, 0.18 mmol, 0.4 equiv.) at 25 °C. The resulting mixture was stirred at 40 °C for 12 h. The mixture was cooled to 25 °C, diluted with water (300 mL), extracted with dichloromethane (3 × 500 mL), and the organic layer was washed with saturated sodium chloride solution (3 × 500 mL), dried over anhydrous sodium sulfate, filtered, and concentrated under reduced pressure. The crude product was purified by flash preparative HPLC with the following conditions (IntelFlash-1): Column, C 18 Silica gel; mobile phase, water and acetonitrile (10.0% acetonitrile up to 100.0% in 20 min); detector, UV 254 nm. 150.0 mg (30%) of (9H-fluoren-9-yl)methyl ((S)-10-benzyl-1-((2S,6aS,6bR,7S,8aS,8bS,10R,11aR,12aS,12bS)-2,6b-difluoro-10-(3-fluorophenyl)-7-hydroxy-6a,8a-dimethyl-4-oxo-1,2,4,6a ,6b,7,8,8a,11a,12,12a,12b-dodecahydro-8bH-naphtho[2',1':4,5]indeno[1,2-d][1,3]dioxol-8b-yl)-1,6,9,12,15-pentaoxo-3-oxa-5,8,11,14-tetraazahexadecan-16-yl)carbamate was obtained as a white solid. MS m / z [M+H] +(ESI):1088.42.

[0435] Intermediate Example 43 Synthesis of (S)-2-(2-(2-aminoacetamido)acetamido)-N-(2-(((2-((2S,6AS,6BR,7S,8AS,8BS,10R,11AR,12AS,12BS)-2,6B-difluoro-10-(3-fluorophenyl)-7-hydroxy-6A,8A-dimethyl-4-oxo-1,2,4,6A,6B,7,8,8A,11A,12,12A,12B-dodecahydro-8BH-naphtho[2',1':4,5]indeno[1,2-D][1,3]dioxol-8B-yl)-2-oxoethoxy)methyl)amino)-2-oxoethyl)-3-phenylpropanamide [ka] Under an inert atmosphere of nitrogen, (9H-fluoren-9-yl)methyl ((S)-10-benzyl-1-((2S,6aS,6bR,7S,8aS,8bS,10R,11aR,12aS,12bS)-2,6b-difluoro-10-(3-fluorophenyl)-7-hydroxy-6a,8a-dimethyl-4-oxo-1,2,4,6a,6b,7,8,8a,11a,12,12a,12b-dodecahydro-8bH-naphtho[ A solution of 2',1':4,5]indeno[1,2-d][1,3]dioxol-8b-yl)-1,6,9,12,15-pentaoxo-3-oxa-5,8,11,14-tetraazahexadecan-16-yl)carbamate (150.0 mg, 0.14 mmol, 1.0 equiv.) in morpholine / N,N-dimethylformamide (0.1 mL / 0.7 mL) was stirred at 25 °C for 1 hour and diluted with water (10 mL). The resulting mixture was extracted with dichloromethane (3 × 30 mL), and the organic layer was washed with saturated sodium chloride solution (3 × 30 mL), dried over anhydrous sodium sulfate, filtered, and concentrated under reduced pressure. The crude product was purified by flash preparative HPLC with the following conditions (IntelFlash-1): Column, C 18Silica gel; mobile phase, water and acetonitrile (10.0% acetonitrile up to 100.0% in 20 minutes); detector, UV 254 nm. Obtained 100.6 mg (84%) of (S)-2-(2-(2-aminoacetamido)acetamido)-N-(2-(((2-((2S,6aS,6bR,7S,8aS,8bS,10R,11aR,12aS,12bS)-2,6b-difluoro-10-(3-fluorophenyl)-7-hydroxy-6a,8a-dimethyl-4-oxo-1,2,4,6a,6b,7,8,8a,11a,12,12a,12b-dodecahydro-8bH-naphtho[2',1':4,5]indeno[1,2-d][1,3]dioxol-8b-yl)-2-oxoethoxy)methyl)amino)-2-oxoethyl)-3-phenylpropanamide as a white solid. MS m / z[M+H] + (ESI):866.35.

[0436] Intermediate Example 44 Synthesis of (2S,6AS,6BR,7S,8AS,8BS,10R,11AR,12AS,12BS)-2,6B-difluoro-10-(3-fluorophenyl)-7-hydroxy-8B-(2-hydroxyacetyl)-6A,8A-dimethyl-1,2,6A,6B,7,8,8A,8B,11A,12,12A,12B-dodecahydro-4H-naphtho[2',1':4,5]indeno[1,2-d][1,3]dioxol-4-one [ka] Under an inert atmosphere of nitrogen, magnesium sulfate (2.0 g, 16.9 mmol, 3.5 equiv.) and 3-fluorobenzaldehyde (903.0 mg, 7.28 mmol, 1.5 equiv.) were added to a solution of fluocinolone (2.0 g, 4.85 mmol, 1.0 equiv.) in acetonitrile (20.0 mL), followed by trifluoromethanesulfonic acid (2.2 g, 14.6 mmol, 3.0 equiv.) at 0 °C. The resulting solution was stirred at 0 °C for 3 h, and the pH was adjusted to 7 with saturated sodium bicarbonate solution (1.0 mL). The mixture was diluted with 300 mL of water and extracted with 3 × 500 mL of dichloromethane. The organic layers were combined, washed with 3 × 500 mL of saturated sodium chloride solution, dried over anhydrous sodium sulfate, filtered, and concentrated under reduced pressure. The crude product (>1 g) was purified by achiral SFC using the following conditions: Column: Torus 2-PIC Column 4.6 x 100 mm, 5 μm; Mobile phase: isopropyl alcohol (1% 2 mol / L NH3-methanol) and CO2; Detector: UV 254 nm. 800 mg (32%) of (2S,6aS,6bR,7S,8aS,8bS,10R,11aR,12aS,12bS)-2,6b-difluoro-10-(3-fluorophenyl)-7-hydroxy-8b-(2-hydroxyacetyl)-6a,8a-dimethyl-1,2,6a,6b,7,8,8a,8b,11a,12,12a,12b-dodecahydro-4H-naphtho[2',1':4,5]indeno[1,2-d][1,3]dioxol-4-one was obtained as a white solid. MS m / z [M+H] + (ESI):519.20; 1 H NMR (400MHz, methanol-d4) δ:0.99(s,3H),1.28-1.65(m,4H),1.71-1.83(m,3H),2.24-2.39(m,3H),2.66-2.74(m,1H),4.31-4.36(m,2H),4.65(d,J=19 .6Hz,1H),5.08-5.09(m,1H),5.49-5.62(m,2H),6.32-6.35(m,2H),7.09-7.18(m,2H),7.27-7.33(m,2H),7.38-7.41(m,1H).

[0437] Intermediate Example 45 Synthesis of (9H-fluoren-9-yl)methyl (2-((((2-((2S,6AS,6BR,7S,8AS,8BS,10R,11AR,12AS,12BS)-2,6B-difluoro-10-(3-fluorophenyl)-7-hydroxy-6A,8A-dimethyl-4-oxo-1,2,4,6A,6B,7,8,8A,11A,12,12A,12B-dodecahydro-8BH-naphtho[2',1':4,5]indeno[1,2-D][1,3]dioxol-8B-yl)-2-oxoethoxy)(hydroxy)phosphoryl)oxy)(hydroxy)phosphoryl)oxy)ethyl)carbamate [ka] Under an inert nitrogen atmosphere, trimethylamine (91.0 mg, 0.90 mmol, 1.2 equiv.) and 1,1'-carbonyldiimidazole (183.0 mg, 1.13 mmol, 1.5 equiv.) were added to a solution of (9H-fluoren-9-yl)methyl(2-(phosphonooxy)ethyl)carbamate (327.0 mg, 0.90 mmol, 1.5 equiv.) in N,N-dimethylformamide (5.0 mL). The resulting mixture was stirred at 25°C for 30 minutes. To this mixture was added 2-((2S,6aS,6bR,7S,8aS,8bS,10R,11aR,12aS,12bS)-2,6b-difluoro-10-(3-fluorophenyl)-7-hydroxy-6a,8a-dimethyl-4-oxo-1,2,4,6a,6b,7,8,8a,11a,12,12a,12b-dodecahydro-8bH-naphtho[2′,1′:4,5]indeno[1,2-d][1,3]dioxol-8b-yl)-2-oxoethyl dihydrogen phosphate (450.0 mg, 0.75 mmol, 1.0 equiv.) and zinc chloride (816.0 mg, 6.0 mmol, 8.0 equiv.). The resulting mixture was stirred at 25° C. overnight, diluted with methanol (10.0 mL), and filtered. The filtrate was concentrated under reduced pressure. The crude product was purified by flash column with the following conditions: 18Silica gel; mobile phase, water (containing 5 mmol / L ammonium hydroxide) and acetonitrile (10.0% acetonitrile up to 100.0% in 20 minutes); detector, UV 254 nm. 310 mg of (9H-fluoren-9-yl)methyl (2-((((2-((2S,6aS,6bR,7S,8aS,8bS,10R,11aR,12aS,12bS)-2,6b-difluoro-10-(3-fluorophenyl)-7-hydroxy-6a,8a-dimethyl-4-oxo-1,2,4,6a,6b,7,8,8a,11a,12,12a,12b-dodecahydro-8bH-naphtho[2',1':4,5]indeno[1,2-d][1,3]dioxol-8b-yl)-2-oxoethoxy)(hydroxy)phosphoryl)oxy)(hydroxy)phosphoryl)oxy)ethyl)carbamate was obtained as a white solid. MS m / z [MH] - (ESI):942.20.

[0438] Intermediate Example 46 Synthesis of (2-((((2-((2S,6AS,6BR,7S,8AS,8BS,10R,11AR,12AS,12BS)-2,6B-difluoro-10-(3-fluorophenyl)-7-hydroxy-6A,8A-dimethyl-4-oxo-1,2,4,6A,6B,7,8,8A,11A,12,12A,12B-dodecahydro-8BH-naphtho[2',1':4,5]indeno[1,2-D][1,3]dioxol-8B-yl)-2-oxoethoxy)(hydroxy)phosphoryl)oxy)(hydroxy)phosphoryl)oxy)ethyl)carbamate [ka] Under an inert atmosphere of nitrogen, (9H-fluoren-9-yl)methyl (2-((((2-((2S,6aS,6bR,7S,8aS,8bS,10R,11aR,12aS,12bS)-2,6b-difluoro-10-(3-fluorophenyl)-7-hydroxy-6a,8a-dimethyl-4-oxo-1,2,4,6a,6b,7,8,8a,11a,12,12a,12b-dodecahydro-8 To a solution of bH-naphtho[2',1':4,5]indeno[1,2-d][1,3]dioxol-8b-yl)-2-oxoethoxy)(hydroxy)phosphoryl)oxy)(hydroxy)phosphoryl)oxy)ethyl)carbamate (310.0 mg, 0.33 mmol, 1.0 equiv.) in dichloromethane (5.0 mL) was added piperidine (196.0 mg, 2.31 mmol, 7.0 equiv.). The resulting solution was stirred at 25°C for 3 hours and concentrated under reduced pressure. The crude product was purified by flash column with the following conditions: 18 Silica gel; mobile phase, water (containing 5 mmol / L ammonium hydroxide) and acetonitrile (5.0% acetonitrile up to 100.0% in 20 min); detector, UV 254 nm. The eluate was concentrated under reduced pressure. Under an inert atmosphere of nitrogen, to a solution of the desired product (ammonium salt) in water (5.0 mL) was added Dowex 50w x 8 (200.0 mg). The resulting solution was stirred at 25 °C for an additional 1 h and filtered. The filtrate was concentrated under reduced pressure. 115 mg (21% over two steps) of the desired product was obtained as a white solid. MS m / z [M−H] - (ESI):720.20.

[0439] Intermediate Example 47 Synthesis of benzyl (E)-3-(4,4,5,5-tetramethyl-1,3,2-dioxaborolan-2-yl)acrylate [ka] To a stirred solution of copper(I) chloride (0.09 g, 0.93 mmol, 0.03 equiv.) and 4,5-bis(diphenylphosphino)-9,9-dimethylxanthene (0.54 g, 0.93 mmol, 0.03 equiv.) in tetrahydrofuran (50 mL), sodium tert-butoxide (0.18 g, 1.87 mmol, 0.06 equiv.) was added at 0° C. The reaction solution was stirred at 0° C. for 1 hour. To the above mixture, 4,4,5,5-tetramethyl-2-(4,4,5,5-tetramethyl-1,3,2-dioxaborolan-2-yl)-1,3,2-dioxaborolane (15.85 g, 62.43 mmol, 2.0 equiv.) was added in several portions at room temperature. The resulting mixture was stirred at room temperature for an additional 1 hour. To the above mixture, benzyl prop-2-ynoate (5.0 g, 31.22 mmol, 1.0 equiv.) and methanol (1.5 g, 46.82 mmol, 1.5 equiv.) were added portionwise at room temperature. The resulting mixture was stirred for an additional 14 h at 25 °C. The aqueous layer was extracted with methylene chloride (3 × 200 mL). The residue was purified by silica gel chromatography (column height: 250 mm, diameter: 100 mm, 100-200 mesh silica gel, petroleum ether / ethyl acetate = 1 / 0, 3 / 1). 5.0 g (55%) of the desired product was obtained as a colorless liquid.

[0440] Intermediate Example 48 Synthesis of tert-butyl 4-(2-bromo-5-nitrophenoxy)butanoate [ka] To a stirred solution of 2-bromo-5-nitrophenol (5.0 g, 22.9 mmol, 1.0 equiv.) and tert-butyl 4-bromobutanoate (7.7 g, 34.40 mmol, 1.5 equiv.) in N,N-dimethylformamide (100.0 mL) was added potassium carbonate (9.5 g, 68.8 mmol, 3 equiv.) in portions at 25° C. under a nitrogen atmosphere. The resulting mixture was stirred at 25° C. under a nitrogen atmosphere for 3 hours and diluted with water (500 mL). The resulting mixture was extracted with dichloromethane (3×500 mL), and the organic layer was washed with saturated sodium chloride solution (3×500 mL), dried over anhydrous sodium sulfate, filtered, and concentrated under reduced pressure. The crude product was purified by flash preparative HPLC with the following conditions (IntelFlash-1): Column, C 18 Silica gel; mobile phase, water and acetonitrile (10.0% acetonitrile up to 100.0% in 20 min); detector, UV 254 nm. Obtained 5 g (60%) of tert-butyl 4-(2-bromo-5-nitrophenoxy)butanoate as a yellow solid.

[0441] Intermediate Example 49 Synthesis of tert-butyl (E)-4-(2-(3-(benzyloxy)-3-oxoprop-1-en-1-yl)-5-nitrophenoxy)butanoate [ka] tert-Butyl 4-(2-bromo-5-nitrophenoxy)butanoate (5.0 g, 13.88 mmol, 1.0 equiv.), benzyl (2E)-3-(4,4,5,5-tetramethyl-1,3,2-dioxaborolan-2-yl)prop-2-enoate (4.8 g, 16.65 mmol, 1.2 equiv.), and potassium phosphate tribasic (5.89 g, 27.76 mmol, 2.0 equiv.) in an N,N-dimethyl phosphate solution. To a stirred solution of methyl phthalamide (50.0 mL), 2-dicyclohexylphosphino-2',6'-dimethoxy-1,1'-biphenyl (0.28 g, 0.69 mmol, 0.05 equiv.) and tris(dibenzylideneacetone)dipalladium (0.64 g, 0.69 mmol, 0.05 equiv.) were added portionwise at room temperature under a nitrogen atmosphere, and the reaction solution was stirred at 90°C under a nitrogen atmosphere for an additional 12 hours. The resulting mixture was cooled to 25°C and diluted with water (500 mL). The resulting mixture was extracted with dichloromethane (3 x 500 mL), and the organic layer was washed with saturated sodium chloride solution (3 x 500 mL), dried over anhydrous sodium sulfate, filtered, and concentrated under reduced pressure. The crude product was purified by flash preparative HPLC with the following conditions (IntelFlash-1): Column, C 18 Silica gel; mobile phase, water and acetonitrile (10.0% acetonitrile up to 100.0% in 20 min); detector, UV 254 nm. 3 g (49%) of tert-butyl (E)-4-(2-(3-(benzyloxy)-3-oxoprop-1-en-1-yl)-5-nitrophenoxy)butanoate was obtained as a yellow oil.

[0442] Intermediate Example 50 Synthesis of 3-(4-amino-2-(4-(tert-butoxy)-4-oxobutoxy)phenyl)propanoic acid [ka] To a solution of tert-butyl (E)-4-(2-(3-(benzyloxy)-3-oxoprop-1-en-1-yl)-5-nitrophenoxy)butanoate (3.0 g, 6.80 mmol, 1.0 equiv) in methanol (30.0 mL) was added palladium on carbon (1.0 g). The flask was evacuated and flushed with hydrogen five times. The resulting solution was stirred at 25 °C for 3 h. The solid was filtered off and washed with methanol (3 × 10 mL). The resulting mixture was concentrated under reduced pressure, and the crude product was used directly in the next step without further purification. 2.5 g (81%) of 3-(4-amino-2-(4-(tert-butoxy)-4-oxobutoxy)phenyl)propanoic acid was obtained as a white solid.

[0443] Intermediate Example 51 Synthesis of 3-(2-(4-(TERT-butoxy)-4-oxobutoxy)-4-(2,5-dioxo-2,5-dihydro-1H-pyrrol-1-yl)phenyl)propanoic acid [ka] To a solution of 3-(4-amino-2-(4-(tert-butoxy)-4-oxobutoxy)phenyl)propanoic acid (2.5 g, 7.73 mmol, 1.0 equiv.) and methyl 2,5-dioxopyrrole-1-carboxylate (1.80 g, 11.59 mmol, 1.5 equiv.) in dichloromethane (25.0 mL) was added trimethylamine (1.56 g, 15.46 mmol, 2.0 equiv.) at room temperature under a nitrogen atmosphere. The resulting mixture was stirred at 50° C. for 4 hours. The resulting mixture was cooled to 25° C. and concentrated under reduced pressure to afford 1.5 g (48%) of 3-(2-(4-(tert-butoxy)-4-oxobutoxy)-4-(2,5-dioxo-2,5-dihydro-1H-pyrrol-1-yl)phenyl)propanoic acid as a yellow oil. MS m / z[MH] - (ESI):402.16.

[0444] Intermediate Example 52 Synthesis of tert-butyl 4-(5-(2,5-dioxo-2,5-dihydro-1H-pyrrol-1-yl)-2-(3-oxo-3-(2,3,5,6-tetrafluorophenoxy)propyl)phenoxy)butanoate [ka] A solution of 3-(2-(4-(tert-butoxy)-4-oxobutoxy)-4-(2,5-dioxo-2,5-dihydro-1H-pyrrol-1-yl)phenyl)propanoic acid (1.5 g, 3.71 mmol, 1 equiv.) in N,N-dimethylformamide (15 mL) was treated with 2,3,5,6-tetrafluorophenol (1.85 g, 11.15 mmol, 3 equiv.) under a nitrogen atmosphere at room temperature, followed by the addition of 1-ethyl-3(3-dimethylpropylamine)carbodiimide (2.14 g, 11.15 mmol, 3 equiv.) at 25° C. The resulting mixture was stirred at 25° C. under a nitrogen atmosphere for 2 hours. The residue was purified by reverse-phase flash chromatography with the following conditions: Column, C 18 Silica gel; mobile phase, acetonitrile (0.1% formic acid) in water, 10% to 100% gradient over 10 min; detector, UV 254 nm. The resulting mixture was concentrated under reduced pressure. 800 mg (39%) of tert-butyl 4-(5-(2,5-dioxo-2,5-dihydro-1H-pyrrol-1-yl)-2-(3-oxo-3-(2,3,5,6-tetrafluorophenoxy)propyl)phenoxy)butanoate was obtained as a colorless oil.

[0445] Intermediate Example 53 Synthesis of 4-(5-(2,5-dioxo-2,5-dihydro-1H-pyrrol-1-yl)-2-(3-oxo-3-(2,3,5,6-tetrafluorophenoxy)propyl)phenoxy)butanoic acid [ka] Under an inert atmosphere of nitrogen, trifluoroacetic acid (2.0 mL) was added to a solution of tert-butyl 4-(5-(2,5-dioxo-2,5-dihydro-1H-pyrrol-1-yl)-2-(3-oxo-3-(2,3,5,6-tetrafluorophenoxy)propyl)phenoxy)butanoate (800 mg, 1.45 mmol, 1 equiv.) in dichloromethane (6.0 mL) at 25° C. The resulting mixture was stirred at 25° C. for 2 hours. The resulting mixture was concentrated under reduced pressure to afford 700 mg (97%) of 4-(5-(2,5-dioxo-2,5-dihydro-1H-pyrrol-1-yl)-2-(3-oxo-3-(2,3,5,6-tetrafluorophenoxy)propyl)phenoxy)butanoic acid as a yellow oil.

[0446] Intermediate Example 54 Synthesis of (2,3,5,6-tetrafluorophenyl) 3-[2-chlorocarbonyl-4-(2,5-dioxopyrrol-1-yl)phenyl]propanoate [ka] To a solution of 5-(2,5-dioxopyrrol-1-yl)-2-[3-oxo-3-(2,3,5,6-tetrafluorophenoxy)propyl]benzoic acid (1.80 g, 4.12 mmol, 1.0 equiv.) in DCM (36 mL) was added 1-chloro-N,N,2-trimethyl-prop-1-en-1-amine (1.10 g, 8.23 ​​mmol, 1.09 mL, 2.0 equiv.). The mixture was stirred at 25 °C for 1 hour. The reaction mixture was concentrated under reduced pressure to remove the solvent. This was used directly in the next step without further purification. The compound (2,3,5,6-tetrafluorophenyl) 3-[2-chlorocarbonyl-4-(2,5-dioxopyrrol-1-yl)phenyl]propanoate (1.88 g, 4.13 mmol, 100% yield) was obtained as a yellow oil.

[0447] Intermediate Example 55 Synthesis of 2-[[2-[[2-[[2-[[2-[[2-[[2-[[2-[[2-[[2-[[2-[[2-[[2-[[5-(2,5-dioxopyrrol-1-yl)-2-[3-oxo-3-(2,3,5,6-tetrafluorophenoxy)propyl]benzoyl]-methyl-amino]acetyl]-methyl-amino]acetyl]-methyl-amino]acetyl]-methyl-amino]acetyl]-methyl-amino]acetyl]-methyl-amino]acetyl]-methyl-amino]acetyl]-methyl-amino]acetyl]-methyl-amino]acetyl]-methyl-amino]acetic acid [ka] To a solution of 2-[methyl-[2-[methyl-[2-[methyl-[2-[methyl-[2-[methyl-[2-[methyl-[2-[methyl-[2-[methyl-[2-[methyl-[2-[methyl-[2-(methylamino)acetyl]amino]acetyl]amino]acetyl]amino]acetyl]amino]-acetyl]amino]acetyl]amino]acetyl]amino]acetyl]amino]acetic acid (3.01 g, 4.13 mmol, 1.0 equiv) in DCM (36 mL) was added DIEA (1.60 g, 12.4 mmol, 2.16 mL, 3.0 equiv) at 0° C. After 10 minutes, a solution of (2,3,5,6-tetrafluorophenyl) 3-[2-chlorocarbonyl-4-(2,5-dioxopyrrol-1-yl)phenyl]propanoate (1.88 g, 4.13 mmol, 1.0 equiv.) in DCM (18 mL) was added and then stirred at 0° C. for 1 hour. The pH of the resulting mixture was adjusted to approximately 6 with TFA at 0° C., diluted with MeCN (2 mL), and concentrated under reduced pressure to remove DCM. The residue was purified by preparative HPLC (column: Phenomenex Luna C 18(250 × 70 mm, 15 μm; mobile phase: [water (TFA)-ACN]; B%: 25%–55%, 20 min) to give 2-[[2-[[2-[[2-[[2-[[2-[[2-[[2-[[2-[[2-[[2-[[5-(2,5-dioxopyrrol-1-yl)-2-[3-oxo-3-(2,3,5,6-tetrafluorophenoxy)propyl]benzoyl]-methyl-amino]acetyl]-methyl-amino]acetyl]-methyl-amino]acetyl]-methyl-amino]acetyl]-methyl-amino]acetyl]-methyl-amino]acetyl]-methyl-amino]acetyl]-methyl-amino]acetyl]-methyl-amino]acetic acid (360 mg, 314 μmol, 7.60% yield) as a white solid. LC / MS [M+H] 1148.4 (calculated); LC / MS [M+H] 1148.5 (observed).

[0448] Intermediate Example 56 Synthesis of (2S,3S,4S,5R,6S)-2-(methoxycarbonyl)-6-(2-nitro-4-((((4-nitrophenoxy)carbonyl)oxy)methyl)phenoxy)tetrahydro-2H-pyran-3,4,5-tolyltriacetate [ka] To a stirred solution of methyl (2S,3S,4S,5R,6S)-3,4,5-tris(acetyloxy)-6-[4-(hydroxymethyl)-2-nitrophenoxy]oxane-2-carboxylate (5.0 g, 10.3 mmol, 1.0 equiv.) and diisopropylethylamine (4.0 g, 31.0 mmol, 3.0 equiv.) in N,N-dimethylformamide (50.0 mL) was added bis(4-nitrophenyl)carbonate (4.7 g, 15.5 mmol, 1.5 equiv.) at room temperature under air. The resulting solution was stirred at 25 °C for 16 h and diluted with water (500 mL). The resulting mixture was extracted with dichloromethane (3 × 500 mL), and the organic layer was washed with saturated sodium chloride solution (3 × 500 mL), dried over anhydrous sodium sulfate, filtered, and concentrated under reduced pressure. The crude product was purified by flash preparative HPLC with the following conditions (IntelFlash-1): Column, C 18 Silica gel; mobile phase, water and acetonitrile (10.0% acetonitrile up to 100.0% in 20 min); detector, UV 254 nm. 5.5 g (82%) of (2S,3S,4S,5R,6S)-2-(methoxycarbonyl)-6-(2-nitro-4-((((4-nitrophenoxy)carbonyl)oxy)methyl)phenoxy)tetrahydro-2H-pyran-3,4,5-tolyl triacetate was obtained as a white solid. MS m / z [M+NH4] + (ESI):668.12.

[0449] Intermediate Example 57 Synthesis of (2S,3R,4S,5S,6S)-2-(4-(((ethylcarbamoyl)oxy)methyl)-2-nitrophenoxy)-6-(methoxycarbonyl)tetrahydro-2H-pyran-3,4,5-tolyltriacetate [ka] To a stirred solution of (2S,3S,4S,5R,6S)-2-(methoxycarbonyl)-6-(2-nitro-4-((((4-nitrophenoxy)carbonyl)oxy)methyl)phenoxy)tetrahydro-2H-pyran-3,4,5-tolyltriacetate (5.5 g, 8.5 mmol, 1.0 equiv.) and ethylamine (1.4 g, 16.9 mmol, 2.0 equiv.) in N,N-dimethylformamide (50.0 mL) was added diisopropylethylamine (3.3 g, 25.4 mmol, 3.0 equiv.) under air at 25° C. The resulting solution was stirred at 25° C. for 2 hours and diluted with water (500 mL). The resulting mixture was extracted with dichloromethane (3×500 mL), and the organic layer was washed with saturated sodium chloride solution (3×500 mL), dried over anhydrous sodium sulfate, filtered, and concentrated under reduced pressure. The crude product was purified by flash preparative HPLC with the following conditions (IntelFlash-1): Column, C 18 Silica gel; mobile phase, water and acetonitrile (10.0% acetonitrile up to 100.0% in 20 min); detector, UV 254 nm. 3.5 g (75%) of (2S,3R,4S,5S,6S)-2-(4-(((ethylcarbamoyl)oxy)methyl)-2-nitrophenoxy)-6-(methoxycarbonyl)tetrahydro-2H-pyran-3,4,5-tolyl triacetate was obtained as a white solid. MS m / z [M+NH4] + (ESI):574.15.

[0450] Intermediate Example 58 Synthesis of (2S,3R,4S,5S,6S)-2-(4-((((chloromethyl)(ethyl)carbamoyl)oxy)methyl)-2-nitrophenoxy)-6-(methoxycarbonyl)tetrahydro-2H-pyran-3,4,5-tolyltriacetate [ka] To a stirred solution of (2S,3R,4S,5S,6S)-2-(4-(((ethylcarbamoyl)oxy)methyl)-2-nitrophenoxy)-6-(methoxycarbonyl)tetrahydro-2H-pyran-3,4,5-tolyltriacetate (3.5 g, 6.3 mmol, 1.0 equiv.) and paraformaldehyde (1.7 g, 18.9 mmol, 3.0 equiv.) in dichloromethane (35.0 mL) was added chlorotrimethylsilane (2.1 g, 18.9 mmol, 3.0 equiv.) under air at 25° C. The resulting mixture was stirred at 25° C. for an additional 2 hours. The resulting mixture was concentrated under reduced pressure to give (2S,3R,4S,5S,6S)-2-(4-((((chloromethyl)(ethyl)carbamoyl)oxy)methyl)-2-nitrophenoxy)-6-(methoxycarbonyl)tetrahydro-2H-pyran-3,4,5-tolyltriacetate (3.0 g, 79%) as a white solid.

[0451] Intermediate Example 59 Synthesis of (2S,3R,4S,5S,6S)-2-(4-((((2-((2S,6AS,6BR,7S,8AS,8BS,10R,11AR,12AS,12BS)-2,6B-difluoro-10-(3-fluorophenyl)-7-hydroxy-6A,8A-dimethyl-4-oxo-1,2,4,6A,6B,7,8,8A,11A,12,12A,12B-dodecahydro-8BH-naphtho[2',1':4,5]indeno[1,2-D][1,3]dioxol-8B-yl)-2-oxoethoxy)methyl)(ethyl)carbamoyl)oxy)methyl)-2-nitrophenoxy)-6-(methoxycarbonyl)tetrahydro-2H-pyran-3,4,5-tolyltriacetate [ka] To a stirred solution of (2S,3R,4S,5S,6S)-2-(4-((((chloromethyl)(ethyl)carbamoyl)oxy)methyl)-2-nitrophenoxy)-6-(methoxycarbonyl)tetrahydro-2H-pyran-3,4,5-tolyltriacetate (3.0 g, 5.0 mmol, 2 equiv.) and diisopropylethylamine (51.28 mg, 0.397 mmol, 3 equiv.) in N,N-dimethylformamide (20.0 mL) was added (2S,6aS,6bR,7S,8a (S,8bS,10R,11aR,12aS,12bS)-2,6b-difluoro-10-(3-fluorophenyl)-7-hydroxy-8b-(2-hydroxyacetyl)-6a,8a-dimethyl-1,2,6a,6b,7,8,8a,8b,11a,12,12a,12b-dodecahydro-4H-naphtho[2',1':4,5]indeno[1,2-d][1,3]dioxol-4-one (1.3 g, 2.5 mmol, 1.0 equiv.) was added under air at 25 °C. The resulting solution was stirred at 25 °C for 2 h and diluted with water (300 mL). The resulting mixture was extracted with dichloromethane (3 × 300 mL), and the organic layer was washed with saturated sodium chloride solution (3 × 300 mL), dried over anhydrous sodium sulfate, filtered, and concentrated under reduced pressure. The crude product was purified by flash preparative HPLC with the following conditions (IntelFlash-1): Column, C 18Silica gel; mobile phase, water and acetonitrile (10.0% acetonitrile up to 100.0% in 20 min); detector, UV 254 nm. 1.1 g (41%) of (2S,3R,4S,5S,6S)-2-(4-((((2-((2S,6aS,6bR,7S,8aS,8bS,10R,11aR,12aS,12bS)-2,6b-difluoro-10-(3-fluorophenyl)-7-hydroxy-6a,8a-dimethyl-4-oxo-1,2,4,6a,6b,7,8,8a,11a,12,1 2a,12b-dodecahydro-8bH-naphtho[2',1':4,5]indeno[1,2-d][1,3]dioxol-8b-yl)-2-oxoethoxy)methyl)(ethyl)carbamoyl)oxy)methyl)-2-nitrophenoxy)-6-(methoxycarbonyl)tetrahydro-2H-pyran-3,4,5-tolyl triacetate was obtained as a white solid. MS m / z [M+H] + (ESI):1087.35.

[0452] Intermediate Example 60 Synthesis of (2S,3R,4S,5S,6S)-2-(2-amino-4-((((2-((2S,6AS,6BR,7S,8AS,8BS,10R,11AR,12AS,12BS)-2,6B-difluoro-10-(3-fluorophenyl)-7-hydroxy-6A,8A-dimethyl-4-oxo-1,2,4,6A,6B,7,8,8A,11A,12,12A,12B-dodecahydro-8BH-naphtho[2',1':4,5]indeno[1,2-D][1,3]dioxol-8B-yl)-2-oxoethoxy)methyl)(ethyl)carbamoyl)oxy)methyl)phenoxy)-6-(methoxycarbonyl)tetrahydro-2H-pyran-3,4,5-tolyltriacetate [ka] Methyl (2S,3R,4S,5S,6S)-2-(4-((((2-((2S,6aS,6bR,7S,8aS,8bS,10R,11aR,12aS,12bS)-2,6b-difluoro-10-(3-fluorophenyl)-7-hydroxy-6a,8a-dimethyl-4-oxo-1,2,4,6a,6b,7,8,8a,11a,12,12a,12b-dodecahydro-8bH-naphtho[2',1':4,5]indeno[1,2-d][1,3]dioxane To a stirred solution of (ethyl-8b-yl)-2-oxoethoxy)methyl)(ethyl)carbamoyl)oxy)methyl)-2-nitrophenoxy)-6-(methoxycarbonyl)tetrahydro-2H-pyran-3,4,5-tolyltriacetate (1.1 g, 1.0 mmol, 1.0 equiv.) and ammonium chloride (536.8 mg, 10.1 mmol, 10.0 equiv.) in methanol (11.0 mL) was added iron powder (1.1 g, 20.0 mmol, 20.0 equiv.) at 25° C. The resulting mixture was stirred at 70° C. for 5 hours. The solid was filtered off and washed with methanol (3×10 mL). The resulting mixture was concentrated under reduced pressure. The crude product was purified by flash preparative HPLC with the following conditions (IntelFlash-1): Column, C 18 Silica gel; mobile phase, water and acetonitrile (10.0% acetonitrile up to 100.0% in 20 min); detector, UV 254 nm. 810 mg (76%) of (2S,3R,4S,5S,6S)-2-(2-amino-4-((((2-((2S,6aS,6bR,7S,8aS,8bS,10R,11aR,12aS,12bS)-2,6b-difluoro-10-(3-fluorophenyl)-7-hydroxy-6a,8a-dimethyl-4-oxo-1,2,4,6a,6b,7,8,8a,11 a,12,12a,12b-dodecahydro-8bH-naphtho[2',1':4,5]indeno[1,2-d][1,3]dioxol-8b-yl)-2-oxoethoxy)methyl)(ethyl)carbamoyl)oxy)methyl)phenoxy)-6-(methoxycarbonyl)tetrahydro-2H-pyran-3,4,5-tolyl triacetate was obtained as a white solid. MS m / z [M+H] + (ESI):1057.37.

[0453] Intermediate Example 61 (2S,3R,4S,5S,6S)-2-(2-(3-((((9H-fluoren-9-yl)methoxy)carbonyl)amino)propanamido)-4-((((2-((2S,6AS,6BR,7S,8AS,8BS,10R,11AR,12AS,12BS)-2,6B-difluoro-10-(3-fluorophenyl)-7-hydroxy-6A,8A-dimethyl-4-oxo-1, Synthesis of 2,4,6A,6B,7,8,8A,11A,12,12A,12B-dodecahydro-8BH-naphtho[2',1':4,5]indeno[1,2-D][1,3]dioxol-8B-yl)-2-oxoethoxy)methyl)(ethyl)carbamoyl)oxy)methyl)phenoxy)-6-(methoxycarbonyl)tetrahydro-2H-pyran-3,4,5-tolyltriacetate [ka] (2S,3R,4S,5S,6S)-2-(2-amino-4-((((2-((2S,6aS,6bR,7S,8aS,8bS,10R,11aR,12aS,12bS)-2,6b-difluoro-10-(3-fluorophenyl)-7-hydroxy-6a,8a-dimethyl-4-oxo-1,2,4,6a,6b,7,8,8a,11a,12,12a,12b-do To a stirred solution of decahydro-8bH-naphtho[2',1':4,5]indeno[1,2-d][1,3]dioxol-8b-yl)-2-oxoethoxy)methyl)(ethyl)carbamoyl)oxy)methyl)phenoxy)-6-(methoxycarbonyl)tetrahydro-2H-pyran-3,4,5-tolyltriacetate (810.0 mg, 0.7 mmol, 1.0 equiv) and ethyl 2-ethoxy-2H-quinoline-1-carboxylate (378.9 mg, 1.5 mmol, 2.0 equiv) in dichloromethane (10.0 mL) was added 3-{[(9H-fluoren-9-ylmethoxy)carbonyl]amino}propanoic acid (357.8 mg, 1.2 mmol, 1.5 equiv) at 25 °C. The resulting solution was stirred at 25°C for 3 hours and diluted with water (200 mL). The resulting mixture was extracted with dichloromethane (3 x 300 mL), and the organic layer was washed with saturated sodium chloride solution (3 x 300 mL), dried over anhydrous sodium sulfate, filtered, and concentrated under reduced pressure. The crude product was purified by flash preparative HPLC with the following conditions (IntelFlash-1): Column, C 18Silica gel; mobile phase, water (10 mmol / L ammonium bicarbonate) and acetonitrile (10.0% acetonitrile up to 100.0% in 20 min); detector, UV 254 nm. 600.0 mg (58%) of (2S,3R,4S,5S,6S)-2-(2-(3-((((9H-fluoren-9-yl)methoxy)carbonyl)amino)propanamide)-4-(((((2-((2S,6aS,6bR,7S,8aS,8bS,10R,11aR,12aS,12bS)-2,6b-difluoro-10-(3-fluorophenyl)-7-hydroxy-6a,8a-dimethyl-4-oxo -1,2,4,6a,6b,7,8,8a,11a,12,12a,12b-dodecahydro-8bH-naphtho[2',1':4,5]indeno[1,2-d][1,3]dioxol-8b-yl)-2-oxoethoxy)methyl)(ethyl)carbamoyl)oxy)methyl)phenoxy)-6-(methoxycarbonyl)tetrahydro-2H-pyran-3,4,5-tolyl triacetate was obtained as a white solid. MS m / z [M+H] + (ESI):1050.48.

[0454] Intermediate Example 62 Synthesis of (2S,3S,4S,5R,6S)-6-(2-(3-aminopropanamido)-4-((((2-((2S,6AS,6BR,7S,8AS,8BS,10R,11AR,12AS,12BS)-2,6B-difluoro-10-(3-fluorophenyl)-7-hydroxy-6A,8A-dimethyl-4-oxo-1,2,4,6A,6B,7,8,8A,11A,12,12A,12B-dodecahydro-8BH-naphtho[2',1':4,5]indeno[1,2-D][1,3]dioxol-8B-yl)-2-oxoethoxy)methyl)(ethyl)carbamoyl)oxy)methyl)phenoxy)-3,4,5-trihydroxytetrahydro-2H-pyran-2-carboxylic acid [ka] Under an inert atmosphere of nitrogen, (2S,3R,4S,5S,6S)-2-(2-(3-((((9H-fluoren-9-yl)methoxy)carbonyl)amino)propanamido)-4-((((2-((2S,6aS,6bR,7S,8aS,8bS,10R,11aR,12aS,12bS)-2,6b-difluoro-10-(3-fluorophenyl)-7-hydroxy-6a,8a-dimethyl-4-oxo-1,2,4,6a,6b,7,8,8a,11a,12,12a,12b-dodecahydro- To a stirred solution of 8bH-naphtho[2',1':4,5]indeno[1,2-d][1,3]dioxol-8b-yl)-2-oxoethoxy)methyl)(ethyl)carbamoyl)oxy)methyl)phenoxy)-6-(methoxycarbonyl)tetrahydro-2H-pyran-3,4,5-tolyl triacetate (600.0 mg, 0.4 mmol, 1.0 equiv.) in methanol (10.0 mL) and water (5.0 mL) was added lithium hydroxide (31.9 mg, 1.2 mmol, 3.0 equiv.) at 0°C. The resulting solution was stirred at 25°C for 12 hours and the pH was adjusted to 7 with hydrochloric acid (1 mol / L). The crude product was purified by flash preparative HPLC with the following conditions (IntelFlash-1): Column, C 18 Silica gel; mobile phase, water and acetonitrile (10.0% acetonitrile up to 100.0% in 20 min); detector, UV 254 nm. 180 mg (41%) of (2S,3S,4S,5R,6S)-6-(2-(3-aminopropanamido)-4-((((2-((2S,6aS,6bR,7S,8aS,8bS,10R,11aR,12aS,12bS)-2,6b-difluoro-10-(3-fluorophenyl)-7-hydroxy-6a,8a-dimethyl-4-oxo-1,2,4,6a,6b ,7,8,8a,11a,12,12a,12b-dodecahydro-8bH-naphtho[2',1':4,5]indeno[1,2-d][1,3]dioxol-8b-yl)-2-oxoethoxy)methyl)(ethyl)carbamoyl)oxy)methyl)phenoxy)-3,4,5-trihydroxytetrahydro-2H-pyran-2-carboxylic acid was obtained as a white solid. MS m / z [M+H] + (ESI):988.36.

[0455] Intermediate Example 63 Synthesis of methyl (E)-3-(4,4,5,5-tetramethyl-1,3,2-dioxaborolan-2-yl)acrylate [ka] To a solution of copper(I) chloride (1.06 g, 10.7 mmol, 0.03 equiv.) and 4,5-bis(diphenylphosphino)-9,9-dimethylxanthene (6.19 g, 10.7 mmol, 0.03 equiv.) in tetrahydrofuran (450.0 mL) was added sodium tert-butoxide (2.06 g, 21.4 mmol, 0.06 equiv.) at 0°C. The reaction solution was stirred at 0°C for 1 hour, and then a solution of bis(pinacolato)diboron (90.6 g, 357 mmol, 1.0 equiv.) in tetrahydrofuran (150.0 mL) was added. The reaction solution was stirred at 20°C for 1 hour under a nitrogen atmosphere. Methyl prop-2-ynoate (30.0 g, 357.0 mmol, 1.0 equiv.) and methanol (22.9 g, 714 mmol, 2.0 equiv.) were added to the above reaction solution. The reaction solution was then stirred at 20° C. for an additional 12 hours. The resulting mixture was poured into ice water (w / w=1 / 1) (300 mL) and stirred for 5 minutes. The aqueous phase was extracted with ethyl acetate (200 mL×3). The combined organic phase was washed with brine (50 mL×2), dried over anhydrous sodium sulfate, filtered, and concentrated under high vacuum. The residue was purified by silica gel chromatography (column height: 250 mm, diameter: 100 mm, 100-200 mesh silica gel, petroleum ether / ethyl acetate = 1 / 0, 3 / 1) to give methyl (E)-3-(4,4,5,5-tetramethyl-1,3,2-dioxaborolan-2-yl)acrylate (70 g, 92%) as a colorless oil. 1 H NMR (400MHz, CDCl3) δ 6.83-6.75 (m, 1H), 6.68-6.59 (m, 1H), 3.77 (s, 3H), 1.29 (s, 12H).

[0456] Intermediate Example 64 Synthesis of tert-butyl((benzyloxy)carbonyl)glycyl [ka] To a solution of 2-(benzyloxycarbonylamino)acetic acid (5.50 g, 26.3 mmol, 1.0 equiv.) in N,N-dimethylformamide (100.0 mL) was added triethylamine (7.98 g, 78.9 mmol, 3.0 equiv.), 2-(7-azabenzotriazol-1-yl)-N,N,N′,N′-tetramethyluronium hexafluorophosphate (12.0 g, 31.5 mmol, 1.2 equiv.), and tert-butyl 2-aminoacetate (3.45 g, 26.3 mmol, 1.0 equiv.), followed by stirring at 25° C. for 2 hours. The reaction mixture was quenched at 0° C. by adding water (150 mL) and then extracted with ethyl acetate (100 mL×3). The combined organic layers were washed with water (80 mL × 3), then brine (100 mL), dried over sodium sulfate, filtered, and concentrated under reduced pressure to give a residue. The residue was purified by silica gel chromatography (column height: 250 mm, diameter: 100 mm, 100-200 mesh silica gel, ethyl acetate / methanol = 1 / 1) to give tert-butyl ((benzyloxy)carbonyl)glycylglycinate (7.7 g, 91%) as a yellow oil. 1 H NMR (CDCl3, 400MHz) δ 7.40-7.31 (m, 5H), 5.14 (s, 2H), 3.98-3.89 (m, 4H), 1.47 (s, 9H).

[0457] Intermediate Example 65 Synthesis of [ka] To a solution of tert-butyl ((benzyloxy)carbonyl)glycylglycinate (8.8 g, 27.3 mmol, 1.0 equiv) in methanol (150 mL) was added palladium on carbon (10%, 2.5 g) under a nitrogen atmosphere. The suspension was degassed and purged with hydrogen for 3 minutes. The mixture was stirred under hydrogen (50 Psi) at 25° C. for 2 hours. The reaction mixture was filtered and concentrated under reduced pressure to give a residue. tert-Butyl glycylglycinate (4.2 g, 81%) was obtained as a white solid. MS m / z [M+H] + (ESI):189.12; 1 H NMR(CDCl3,400MHz)δ 7.67(s,1H),3.96(d,J=5.6Hz,2H),3.39(s,2H),1.75(s,2H),1.46(s,9H).

[0458] Intermediate Example 66 Synthesis of tert-butyl 2-bromo-5-nitrobenzoate [ka] To a mixture of 2-bromo-5-nitrobenzoic acid (9.7 g, 39.4 mmol, 1.0 equiv.) in dichloromethane (100 mL), dicyclohexylcarbodiimide (8.95 g, 43.4 mmol, 1.1 equiv.) and 4-dimethylaminopyridine (2.41 g, 19.7 mmol, 0.5 equiv.) were added at 0° C. The mixture was stirred at 0° C. for 10 minutes, and then tert-butanol (4.4 g, 59.1 mmol, 1.5 equiv.) was added and stirred at 20° C. for 12 hours. The mixture was poured into ice water (w / w=1 / 1) (50 mL) and stirred for 10 minutes. The aqueous phase was extracted with dichloromethane (500 mL × 3). The combined organic phase was washed with brine (100 mL × 3), dried over anhydrous sodium sulfate, filtered, and concentrated under high vacuum. The residue was purified by silica gel chromatography (column height: 250 mm, diameter: 100 mm, 100-200 mesh silica gel, petroleum ether / ethyl acetate = 1 / 0, 5 / 1) to give tert-butyl 2-bromo-5-nitrobenzoate (9.2 g, 77%) as a white solid. 1H NMR (400MHz, CDCl3) δ 8.52 (d, J = 2.8 Hz, 1H), 8.13 (dd, J = 2.8, 8.8 Hz, 1H), 7.83 (d, J = 8.8 Hz, 1H), 1.65 (s, 9H).

[0459] Intermediate Example 67 Synthesis of tert-butyl (E)-2-(3-methoxy-3-oxoprop-1-en-1-yl)-5-nitrobenzoate [ka] Methyl (E)-3-(4,4,5,5-tetramethyl-1,3,2-dioxaborolan-2-yl)acrylate (11.2 g, 53.0 mmol, 2.5 equiv.), tert-butyl A mixture of 2-bromo-5-nitrobenzoate (6.40 g, 21.2 mmol, 1.0 equiv.), tribasic potassium phosphate (6.74 g, 31.8 mmol, 1.5 equiv.), 2-dicyclohexylphosphino-2',6'-dimethoxy-1,1'-biphenyl (870 mg, 2.12 mmol, 0.1 equiv.), tris(dibenzylideneacetone)dipalladium (970 mg, 1.06 mmol, 0.05 equiv.) in dioxane (120 mL) and water (25 mL) was degassed and purged with nitrogen three times. The mixture was then stirred at 90 °C under a nitrogen atmosphere for 12 h. The mixture was poured into ice water (w / w = 1 / 1) (30 mL) and stirred for 10 min. The aqueous phase was extracted with ethyl acetate (500 mL × 3). The combined organic phase was washed with brine (100 mL × 3), dried over anhydrous sodium sulfate, filtered, and concentrated under high vacuum. The residue was purified by silica gel chromatography (column height: 250 mm, diameter: 100 mm, 100-200 mesh silica gel, petroleum ether / ethyl acetate = 1 / 0, 3 / 1) to give tert-butyl (E)-2-(3-methoxy-3-oxoprop-1-en-1-yl)-5-nitrobenzoate (6 g, 92%) as a yellow solid. 1H NMR(400MHz,CDCl3)δ 8.74(d,J=2.4Hz,1H),8.41(d,J=15.6Hz,1H),8.34(dd,J=2.4,8.4Hz,1H) ,7.72(d,J=8.4Hz,1H),6.38(d,J=15.6Hz,1H),3.85(s,3H),1.65(s,9H).

[0460] Intermediate Example 68 Synthesis of tert-butyl 5-amino-2-(3-methoxy-3-oxopropyl)benzoate [ka] A mixture of tert-butyl (E)-2-(3-methoxy-3-oxoprop-1-en-1-yl)-5-nitrobenzoate (4.0 g, 13.0 mmol, 1.0 equiv.), palladium on carbon (10%, 400.0 mg) in methanol (50.0 mL) was degassed and purged with hydrogen three times, then the mixture was stirred under a hydrogen atmosphere at 25° C. for 3 hours. The mixture was filtered and concentrated under high vacuum to give tert-butyl 5-amino-2-(3-methoxy-3-oxopropyl)benzoate (3.50 g, 96%) as a yellow oil. [M+H] + (ESI):280.30; 1 H NMR(400MHz,CDCl3)δ 7.14(d,J=2.4Hz,1H),7.04(d,J=8.0Hz,1H),6.73(dd,J=2.4,8.0Hz,1H),3.67(s,3H),3.12(t,J=8.0Hz,2H),2.61(t,J=8.0Hz,2H),1.59(s,9H).

[0461] Intermediate Example 69 Synthesis of 5-amino-2-(3-methoxy-3-oxopropyl)benzoic acid [ka] To a solution of tert-butyl 5-amino-2-(3-methoxy-3-oxopropyl)benzoate (3.5 g, 12.5 mmol, 1.0 equiv.) in ethyl acetate (10.0 mL) was added hydrogen chloride (gas) / ethyl acetate (4 mol / L, 50 mL, 16.0 equiv.), followed by stirring at 25° C. for 12 hours. The mixture was concentrated under high vacuum to give 5-amino-2-(3-methoxy-3-oxopropyl)benzoic acid (3.20 g, 98%) as a white solid. [M+H] + (ESI):224.23; 1 H NMR(400MHz,DMSO-d6)δ 7.69(d,J=2.0Hz,1H),7.39-7.31(m,2H),3.57(s,3H),3.15(t,J=7.6Hz,2H),2.59(t,J=7.6Hz,2H).

[0462] Intermediate Example 70 Synthesis of methyl 3-(4-amino-2-((2-((2-(Tert-butoxy)-2-oxoethyl)amino)-2-oxoethyl)carbamoyl)phenyl)propanoate [ka] To a solution of 5-amino-2-(3-methoxy-3-oxopropyl)benzoic acid (3.20 g, 12.3 mmol, 1.0 equiv.) in N,N-dimethylformamide (40 mL), 4-methylmorpholine (3.74 g, 37.0 mmol, 3.0 equiv.), 1-hydroxybenzotriazole (833 mg, 6.16 mmol, 0.5 equiv.), N-(3-dimethylaminopropyl)-N′-ethylcarbodiimide hydrochloride (4.72 g, 24.7 mmol, 2.0 equiv.), and tert-butyl 2-[(2-aminoacetyl)amino]acetate (2.78 g, 14.8 mmol, 1.2 equiv.) were added at 0° C. and stirred at 20° C. for 2 h. The mixture was poured into ice water (w / w=1 / 1) (40 mL) and stirred for 10 min. The aqueous phase was extracted with ethyl acetate (500 mL × 3). The combined organic phases were washed with brine (200 mL × 3), dried over anhydrous sodium sulfate, filtered, and concentrated under high vacuum. The residue was purified by silica gel chromatography (column height: 250 mm, diameter: 100 mm, 100-200 mesh silica gel, petroleum ether / ethyl acetate = 1 / 0, 0 / 1) to give methyl 3-(4-amino-2-((2-((2-(tert-butoxy)-2-oxoethyl)amino)-2-oxoethyl)carbamoyl)phenyl)propanoate (3.8 g, 78%) as a yellow oil. [M+H] + (ESI):394.35; 1 H NMR(400MHz,CDCl3)δ 7.04(d,J=8.0Hz,2H),6.79-6.66(m,3H),4.15(d,J=5.6Hz,2H),3.97(d,J=5.2 Hz,2H),3.61(s,3H),2.99(t,J=7.2Hz,2H),2.68(t,J=7.2Hz,2H),1.47(s,9H).

[0463] Intermediate Example 71 Synthesis of (5-amino-2-(3-methoxy-3-oxopropyl)benzoyl)glycylglycine [ka] To a solution of methyl 3-(4-amino-2-((2-((2-(tert-butoxy)-2-oxoethyl)amino)-2-oxoethyl)carbamoyl)phenyl)propanoate (0.60 g, 1.53 mmol, 1.0 equiv.) in ethyl acetate (5.0 mL) was added hydrogen chloride (gas) / ethyl acetate (4 mol / L, 10 mL, 26.2 equiv.), followed by stirring at 25°C for 12 hours. The mixture was concentrated under high vacuum to give (5-amino-2-(3-methoxy-3-oxopropyl)benzoyl)glycylglycine (550 mg, 96% yield) as a white solid. [M+H] + (ESI):338.14; 1 H NMR(400MHz,DMSO-d6)δ 8.57(t,J=6.0Hz,1H),8.25(t,J=6.4Hz,1H),7.28(d,J=8.0Hz,1H),7.20-7.10(m,2H),3.89(d, J=6.0Hz,2H),3.81(d,J=6.0Hz,2H),3.57(s,3H),2.90(t,J=8.0Hz,2H),2.60(t,J=8.0Hz,2H).

[0464] Intermediate Example 72 Synthesis of tert-butyl 1-(5-amino-2-(3-methoxy-3-oxopropyl)phenyl)-1,4,7-trioxo-11,14,17,20,23,26,29,32,35,38,41,44-dodecaoxa-2,5,8-triazaheptatetracontan-47-oate [ka] (5-amino-2-(3-methoxy-3-oxopropyl)benzoyl)glycylglycine (320.0 mg, 0.86 mmol, 1.0 equiv.), tert-butyl A mixture of 3-[2-[2-[2-[2-[2-[2-[2-[2-[2-[2-[2-[2-[2-[2-(2-aminoethoxy)ethoxy]ethoxy]ethoxy]ethoxy]ethoxy]ethoxy]ethoxy]ethoxy]ethoxy]ethoxy]ethoxy]ethoxy]propanoate (576.0 mg, 0.86 mmol, 1.0 equiv), N-(3-dimethylaminopropyl)-N'-ethylcarbodiimide hydrochloride (328.0 mg, 1.71 mmol, 2.0 equiv), 1-hydroxybenzotriazole (57.8 mg, 0.43 mmol, 0.5 equiv), and 4-methylmorpholine (259.0 mg, 2.57 mmol, 3.0 equiv) in N,N-dimethylformamide (5.0 mL) was stirred at 25°C for 2 hours. The reaction mixture was quenched at 0° C. by adding water (10 mL) and then extracted with methylene chloride / isopropyl alcohol (v:v=3:1, 100 mL×5). The combined organic layer was concentrated under reduced pressure to give a residue. Column, C 18 Silica gel; mobile phase, acetonitrile in water (0.1% trifluoroacetic acid / formic acid), 10% to 70% gradient over 20 minutes; detector, UV 254 nm. Obtained 950 mg (90%) of tert-butyl 1-(5-amino-2-(3-methoxy-3-oxopropyl)phenyl)-1,4,7-trioxo-11,14,17,20,23,26,29,32,35,38,41,44-dodecaoxa-2,5,8-triazaheptatetracontan-47-oate as a yellow oil. [M+H] + (ESI):994.10.

[0465] Intermediate Example 73 Synthesis of (S)-3-(4-amino-2-((47-(TERT-butoxycarbonyl)-52,52-dimethyl-2,5,45,50-tetraoxo-9,12,15,18,21,24,27,30,33,36,39,42,51-tridecaoxa-3,6,46-triazatripentacontyl)carbamoyl)phenyl)propanoic acid [ka] [ka] To a solution of tert-butyl 1-(5-amino-2-(3-methoxy-3-oxopropyl)phenyl)-1,4,7-trioxo-11,14,17,20,23,26,29,32,35,38,41,44-dodecaoxa-2,5,8-triazaheptatetracontan-47-oate (850 mg, 956 μmol, 1.0 equiv.) in dioxane (5.0 mL) was added hydrogen chloride (gas) / ethyl acetate (15 mL, 4 mol / L). The mixture was stirred at 25° C. for 2 hours. The reaction mixture was concentrated under reduced pressure. 1-(5-amino-2-(3-methoxy-3-oxopropyl)phenyl)-1,4,7-trioxo-11,14,17,20,23,26,29,32,35,38,41,44-dodecaoxa-2,5,8-triazaheptatetracontan-47-oic acid (800 mg, crude) was obtained as a yellow solid. [M+H] + (ESI):936.20.

[0466] 1-(5-amino-2-(3-methoxy-3-oxopropyl)phenyl)-1,4,7-trioxo-11,14,17,20,23,26,29,32,35,38,41,44-dodecaoxa-2,5,8-triazaheptatetracontan-47-oic acid (800 mg, 0.903 mmol, 1.0 equiv.), di-tert-butyl (2S)-2-aminopentanedioate (588 mg, 1.99 mmol, 2.2 equiv., HCl), 1-ethyl-3(3-dimethylpropylamine A mixture of carbodiimide (346 mg, 1.81 mmol, 2.0 equiv.), 1-hydroxybenzotriazole (61.0 mg, 0.451 mmol, 0.5 equiv.), and 4-methylmorpholine (274 mg, 2.71 mmol, 3.0 equiv.) in N,N-dimethylformamide (10.0 mL) was stirred at 25° C. for 2 hours. The reaction mixture was quenched at 0° C. by adding water (20 mL) and then extracted with methylene chloride / isopropyl alcohol (v:v=3:1, 50 mL×5). The combined organic layer was concentrated under reduced pressure to give a residue. Column, C18 Silica gel; mobile phase, acetonitrile in water (0.1% trifluoroacetic acid / formic acid), 10% to 70% gradient over 20 min; detector, UV 254 nm. 900 mg (89%) of di-tert-butyl(1-(2-(5-amino-2-(3-methoxy-3-oxopropyl)benzamido)acetamido)-2-oxo-6,9,12,15,18,21,24,27,30,33,36,39-dodecaoxa-3-azadotetracontan-42-oyl)-L-glutamate was obtained as a yellow oil. [M+H] + (ESI):1179.05.

[0467] To a solution of di-tert-butyl(1-(2-(5-amino-2-(3-methoxy-3-oxopropyl)benzamido)acetamido)-2-oxo-6,9,12,15,18,21,24,27,30,33,36,39-dodecaoxa-3-azadotetracontan-42-oyl)-L-glutamate (800 mg, 0.68 mmol, 1.0 equiv) in methanol (8.0 mL) and water (2.5 mL) was added sodium hydroxide (136 mg, 3.39 mmol, 5.0 equiv), followed by stirring at 0° C. for 2 hours. The mixture was concentrated under high vacuum to remove methanol, then diluted with water (20 mL). The pH of the aqueous phase was adjusted to approximately 7 with hydrochloric acid (2 mol / L) and extracted with methylene chloride / isopropyl alcohol (v / v=3:1, 30 mL×5). The combined organic layers were dried over sodium sulfate, filtered, and concentrated under reduced pressure to give a residue, which afforded (S)-3-(4-amino-2-((47-(tert-butoxycarbonyl)-52,52-dimethyl-2,5,45,50-tetraoxo-9,12,15,18,21,24,27,30,33,36,39,42,51-tridecaoxa-3,6,46-triazatripentacontyl)carbamoyl)phenyl)propanoic acid (700 mg, crude) as a red oil. [M+H] + (ESI):1165.10.

[0468] Intermediate Example 74 Synthesis of di-tert-butyl(1-(2-(5-(2,5-dioxo-2,5-dihydro-1H-pyrrol-1-yl)-2-(3-oxo-3-(2,3,5,6-tetrafluorophenoxy)propyl)benzamido)acetamido)-2-oxo-6,9,12,15,18,21,24,27,30,33,36,39-dodecaoxa-3-azadotetracontan-42-oyl)-L-glutamate [ka] [ka] To a solution of (S)-3-(4-amino-2-((47-(tert-butoxycarbonyl)-52,52-dimethyl-2,5,45,50-tetraoxo-9,12,15,18,21,24,27,30,33,36,39,42,51-tridecaoxa-3,6,46-triazatripentacontyl)carbamoyl)phenyl)propanoic acid (700 mg, 0.60 mmol, 1.0 equiv) and methyl 2,5-dioxopyrrole-1-carboxylate (139.8 mg, 0.92 mmol, 1.5 equiv) in dichloromethane (7.0 mL) was added trimethylamine (182.2 mg, 1.80 mmol, 3.0 equiv), followed by stirring at 50° C. for 4 hours. The pH of the mixture was adjusted to about 6 with trifluoroacetic acid, diluted by adding water (10 mL), and extracted with methylene chloride:isopropyl alcohol (v / v=3:1, 30 mL×5). The combined organic layer was concentrated under reduced pressure to give a residue. Column, C 18 Silica gel; mobile phase, acetonitrile in water (0.1% trifluoroacetic acid / formic acid), 10% to 70% gradient in 20 min; detector, UV 254 nm. Obtained 306 mg (41%) of (S)-3-(2-((47-(tert-butoxycarbonyl)-52,52-dimethyl-2,5,45,50-tetraoxo-9,12,15,18,21,24,27,30,33,36,39,42,51-tridecaoxa-3,6,46-triazatripentacontyl)carbamoyl)-4-(2,5-dioxo-2,5-dihydro-1H-pyrrol-1-yl)phenyl)propanoic acid as a colorless oil. [M+H]+ (ESI):1245.32.

[0469] 2,3,5,6-Tetrafluorophenol (108 mg, 0.65 mmol, 3.0 equiv.), (S)-3-(2-((47-(tert-butoxycarbonyl)-52,52-dimethyl-2,5,45,50-tetraoxo-9,12,15,18,21,24,27,30,33,36,39,42,51-tridecaoxa-3,6,46-triazatripentacone) A mixture of (2,5-dioxo-2,5-dihydro-1H-pyrrol-1-yl)phenyl)-4-(2,5-dioxo-2,5-dihydro-1H-pyrrol-1-yl)phenyl)propanoic acid (270 mg, 216 μmol, 1.0 equiv.) and 1-ethyl-3(3-dimethylpropylamine)carbodiimide (166 mg, 867 μmol, 4.0 equiv.) in N,N-dimethylformamide (3.0 mL) was stirred at 25° C. for 2 hours. The reaction mixture was concentrated under reduced pressure. The crude product was purified by flash preparative HPLC with the following conditions (IntelFlash-1). Column: C 18 Silica gel; mobile phase, water and acetonitrile (10.0% acetonitrile up to 100.0% in 20 min); detector, UV 254 nm. 90 mg (30%) of di-tert-butyl(1-(2-(5-(2,5-dioxo-2,5-dihydro-1H-pyrrol-1-yl)-2-(3-oxo-3-(2,3,5,6-tetrafluorophenoxy)propyl)benzamido)acetamido)-2-oxo-6,9,12,15,18,21,24,27,30,33,36,39-dodecaoxa-3-azadotetracontan-42-oyl)-L-glutamate was obtained as a colorless oil. [M+H] + (ESI):1394.45; 1H NMR (DMSO-d6, 400 MHz) δ 7.94-7.84(m,1H),7.50(d,J=8.4Hz,1H),7.43-7.34(m,2H),7.18(s,2H),4.1 3(dd,J=5.2,9.1Hz,1H),3.89(s,2H),3.70(s,2H),3.60-3.58(m,3H),3.51-3. 47(m,44H),3.39(t,J=6.0Hz,2H),3.22-3.17(m,2H),3.14(s,3H),2.42-2.32( m, 2H), 2.27-2.19 (m, 2H), 1.95-1.83 (m, 1H), 1.77-1.65 (m, 1H), 1.38 (s, 18H).

[0470] Intermediate Example 75 Synthesis of (1-(2-(5-(2,5-dioxo-2,5-dihydro-1H-pyrrol-1-yl)-2-(3-oxo-3-(2,3,5,6-tetrafluorophenoxy)propyl)benzamido)acetamido)-2-oxo-6,9,12,15,18,21,24,27,30,33,36,39-dodecaoxa-3-azadotetracontan-42-oyl)-L-glutamic acid [ka] Under an inert atmosphere of nitrogen, to a solution of di-tert-butyl (1-(2-(5-(2,5-dioxo-2,5-dihydro-1H-pyrrol-1-yl)-2-(3-oxo-3-(2,3,5,6-tetrafluorophenoxy)propyl)benzamido)acetamido)-2-oxo-6,9,12,15,18,21,24,27,30,33,36,39-dodecaoxa-3-azadotetracontan-42-oyl)-L-glutamate (90 mg, 0.07 mmol, 1.0 equiv) in dichloromethane (1.0 mL) was added trifluoroacetic acid (0.25 mL) at 0 °C. The resulting solution was stirred at 0 °C for 4 h, concentrated under high vacuum, and concentrated under reduced pressure. 90 mg (crude) of (1-(2-(5-(2,5-dioxo-2,5-dihydro-1H-pyrrol-1-yl)-2-(3-oxo-3-(2,3,5,6-tetrafluorophenoxy)propyl)benzamido)acetamido)-2-oxo-6,9,12,15,18,21,24,27,30,33,36,39-dodecaoxa-3-azadotetracontan-42-oyl)-L-glutamic acid was obtained as a yellow oil. [M+H] + (ESI):1279.95.

[0471] Synthesis Example 1 Preparation of Compound I-15 [ka] To a solution of intermediate 7 (55.0 mg, 26.2 μmol, 1.0 equiv) in DCM (1 mL) was added TFA (1.54 g, 13.5 mmol, 1.00 mL, 513 equiv). The mixture was stirred at 25 °C for 2 h. The reaction mixture was concentrated under high vacuum to give a residue, which was purified by preparative HPLC (column: Phenomenex Luna 80 × 30 mm × 3 μm; mobile phase: [water (TFA)-acetonitrile]; B%: 15% to 45%, 8 min). The eluent was removed by lyophilization. The desired product (8.50 mg, 4.04 μmol, 15.3% yield, 99.4% purity, TFA) was obtained as a white solid. 1H NMR(D2O,400MHz)δ 9.05(s,1H),8.31(s,1H),7.91-7.76(m,2H),7.63(d,J=8.6Hz,1H),7.46-7.16(m,7H),7.09(s,1H),6.93(s,2H),5.14(s,2H),4.43 -4.34(m,1H),4.32-4.24(m,1H),4.17-4.02(m,2H),3.97-3.79(m,5H),3.72(t,J=6.0Hz,2H),3.68-3.55(m,46H),3.53(t,J=6.0Hz ,2H),3.48-3.30(m,8H),3.16-3.11(m,2H),3.08-2.97(m,4H),2.66-2.61(m,2H),2.58-2.48(m,2H),2.42(t,J=7.2Hz,2H),2.21-2 .07(m,1H),1.98-1.85(m,2H),1.85-1.75(m,1H),1.73-1.56(m,5H),1.54-1.34(m,2H),0.89(t,J=7.2Hz,3H),0.79(t,J=6.0Hz,9H) HPLC:99.45%(220nm), 100.00%(254nm) MS (ESI): Mass Calculation Value. 95 H 134 N 16 O 30 について1978.95, m / z measured value 1979.9641[M+H] +

[0472] Synthesis Example 2 Modulation of Compound I-13

change

[0473] Synthesis Example 3 Modulation of compound I-49

change

[0474] The compounds in the following table were prepared in a manner similar to the methods and protocols of Synthesis Example 3 and the methods and protocols preceding it (see also, for example, Intermediate Examples 25-32), using the appropriate compounds as starting materials. [Table 3-1] [Table 3-2]

[0475] Synthesis Example 4 Synthesis of Compound I-44 [ka] A solution of 4-(5-(2,5-dioxo-2,5-dihydro-1H-pyrrol-1-yl)-2-(3-oxo-3-(2,3,5,6-tetrafluorophenoxy)propyl)phenoxy)butanoic acid (120.0 mg, 0.20 mmol, 1.0 equiv) in N,N-dimethylformamide (2.0 mL) was treated with N,N-diisopropylethylamine (52.3 mg, 0.40 mmol, 2.0 equiv) at room temperature. To the above mixture, (S)-2-(2-(2-aminoacetamido)acetamido)-N-(2-(((2-((2S,6aS,6bR,7S,8aS,8bS,10R,11aR,12aS,12bS)-2,6b-difluoro-10-(3-fluorophenyl)-7-hydroxy-6a,8a-dimethyl-4-oxo-1,2,4,6a,6b, 7,8,8a,11a,12,12a,12b-dodecahydro-8bH-naphtho[2',1':4,5]indeno[1,2-d][1,3]dioxol-8b-yl)-2-oxoethoxy)methyl)amino)-2-oxoethyl)-3-phenylpropanamide (192.6 mg, 0.22 mmol, 1.1 equivalents) was added at 25°C. The resulting mixture was stirred at 25°C for an additional 5 hours. The crude product was purified by preparative HPLC using the following conditions: acetonitrile / water containing 0.05% formic acid. 64 mg (26%) of compound I-44 was obtained as a white solid. [MH] - (ESI):1193.40. 1H NMR(300MHz, methanol-d4)δ 0.98(s,3H),1.57(s,4H),1.64-1.90(m,3H),1.99-2.23(m,3H),2.24-2.4(m,2H),2.42-2.82(m,5H),2.86 -3.08(m,3H),3.20(dd,J=13.9Hz,5.8Hz,1H),3.68-3.95(m,6H),4.02(t,J=6.1Hz,2H),4.24-4.38(m,1H) ,4.39-4.52(m,2H),4.62(d,J=28.1Hz,1H),4.70-4.81(m,2H),5.04(d,J=4.4Hz,1H),5.37-5.71(m,2H),6 .16-6.35(m,2H),6.81(dd,J=8.0Hz,1.9Hz,1H),6.88-6.98(m,3H),7.03-7.17(m,1H),7.14-7.44(m,10H).

[0476] The compounds in the following table were prepared in a manner similar to the methods and protocols of Synthetic Example 4 and those preceding it, using the appropriate compounds as starting materials. [Table 4]

[0477] Synthesis Example 5 Synthesis of compound I-46 [ka] Compound I-46 was synthesized using the conditions and materials according to the reaction scheme shown above. [MH]-(ESI):1904.75;1H NMR (300MHz, methanol-d4) δ0.98(s,4H),1.29(s,1H),1.56(s,4H),1.79(d,J=13.8Hz, 3H),2.01-2.24(m,4H),2.33(s,2H),2.52-2.78(m,5H),2.81-3.24(m,33H),3.61-3. 90(m,6H),3.90-4.23(m,12H),4.24-4.53(m,11H),4.66-4.81(m,3H),5.03(d,J=4.4 Hz, 1H), 5.60 (s, 2H), 6.28 (d, J=11.7Hz, 2H), 6.74-7.16 (m, 5H), 7.16-7.42 (m, 10H).

[0478] The compounds in the following table were prepared in a manner similar to the methods and protocols of Synthetic Example 5 and those preceding it, using the appropriate compounds as starting materials. [Table 5]

[0479] Synthesis Example 6 Synthesis of compound I-47 [ka] 4-(5-(2,5-dioxo-2,5-dihydro-1H-pyrrol-1-yl)-2-(3-oxo-3-(2,3,5,6-tetrafluorophenoxy)propyl)phenoxy)butanoic acid (200.0 mg, 0.40 mmol, 1.0 equiv.) and (S)-2-amino-N-((S)-1-(((2-((2S,6aS,6bR,7S,8aS,8bS,10R,11aR,12aS,12bS)-2,6b-difluoro-10-(3-fluorophenyl)-7-hydroxy-6a,8a-dimethyl-4-oxo- To a stirred solution of 1,2,4,6a,6b,7,8,8a,11a,12,12a,12b-dodecahydro-8bH-naphtho[2',1':4,5]indeno[1,2-d][1,3]dioxol-8b-yl)-2-oxoethoxy)methyl)amino)-1-oxopropan-2-yl)propanamide (417.7 mg, 0.60 mmol, 1.5 equiv.) in N,N-dimethylformamide (5.0 mL) was added N,N-diisopropylethylamine (156.5 mg, 1.20 mmol, 3.0 equiv.) dropwise at 25°C. The resulting mixture was stirred at 25°C for an additional 5 hours. The crude product was purified by flash chromatography with the following conditions (IntelFlash-1). Column, C 18 Silica gel; mobile phase, water (containing 5 mmol / L formic acid) and acetonitrile (10.0% acetonitrile up to 100.0% in 20 min); detector, UV 254 nm. 150 mg (36%) of compound I-47 was obtained as a white solid. MS m / z [M+H] + (ESI):1019.38.

[0480] The compounds in the following table were prepared in a manner similar to the methods and protocols of Synthesis Example 6 and the methods and protocols preceding it (see also, for example, Intermediate Examples 33-37), using the appropriate compounds as starting materials. [Table 6]

[0481] Synthesis Example 7 Synthesis of compound I-48 [ka] To a stirred solution of compound I-47 (100.0 mg, 0.098 mmol, 1.0 equiv.) and HATU (2-(3H-[1,2,3]triazolo[4,5-b]pyridin-3-yl)-1,1,3,3-tetramethylisouronium) (55.97 mg, 0.147 mmol, 1.5 equiv.) in N,N-dimethylformamide (2.0 mL) was added N,N-diisopropylethylamine (38.05 mg, 0.294 mmol, 3.0 equiv.) and (5,8,11,14,17,20,23,26,29-nonamethyl-4,7,10,13,16,19,22,25,28-nonaoxo-2,5,8,11,14,17,20,23,26,29-decaazahentriacontan-31-oic acid) (143.04 mg, 0.196 mmol, 2.0 equiv.) were added dropwise under air at 25°C. The resulting mixture was stirred at 25°C for an additional 3 hours. The crude product was purified by flash chromatography with the following conditions (IntelFlash-1). Column, C 18 Silica gel; mobile phase, water and acetonitrile (10.0% acetonitrile up to 100.0% in 20 min); detector, UV 254 nm. The resulting mixture was lyophilized in a cool, dry place. 20 mg (10%) of compound I-48 was obtained as a white solid. MS m / z [M+H] + (ESI):1728.05; 1 H NMR (400MHz, methanol-d4) δ:0.86-0.90(m,1H),1.02(s,3H),1.22-1.40(m,6H),1.56(s,3H),1.56-1.70(m,1H),1.75-1 .9(m,3H),2.05-2.40(m,3H),2.25-2.45(m,2H),2.50-2.61(m,2H),2.62-2.75(m,2H),2.90- 3.15(m,33H),3.95-4.55(m,24H),4.61-4.82(m,3H),5.05-5.11(m,1H),5.40-5.70(m,2H),6 .25-6.40(m,2H),6.70-6.82(m,1H),6.85-7.00(m,3H),7.05-7.28(m,3H),7.31-7.5(m,3H).

[0482] The compounds in the following table were prepared in a manner similar to the methods and protocols of Synthetic Example 7 and those preceding it, using the appropriate compounds as starting materials. [Table 7-1] [Table 7-2] [Table 7-3]

[0483] Synthesis Example 8 Synthesis of I-40 [ka] Under an inert atmosphere of nitrogen, (S)-2-(2-(2-aminoacetamido)acetamido)-N-(2-(((2-((2S,6aS,6bR,7S,8aS,8bS,10R,11aR,12aS,12bS)-2,6b-difluoro-10-(3-fluorophenyl)-7-hydroxy-6a,8a-dimethyl-4-oxo-1,2,4,6a,6b,7,8,8a,11a,12,12a,12b-dodecahydro-8bH- To a stirred mixture of naphtho[2',1':4,5]indeno[1,2-d][1,3]dioxol-8b-yl)-2-oxoethoxy)methyl)amino)-2-oxoethyl)-3-phenylpropanamide in N,N-dimethylformamide was added N,N-diisopropylethylamine and 3-(4-(2,5-dioxo-2,5-dihydro-1H-pyrrol-1-yl)-2-methoxyphenyl)propanoic acid at 0° C. The resulting mixture was stirred at 25° C. for 3 hours. The crude product was purified by preparative HPLC. MS m / z[MH] - (ESI):1121.35; 1H NMR (300MHz, methanol-d4) δ:0.98(s,3H),1.48-1.62(m,4H),1.72-1.88(m,3H),2.12-2.27(m,1H),2.28-2.39(m,2H ),2.46-2.79(m,3H),2.81-2.92(m,2H),2.93-3.03...

Claims

1. A compound having the following structure (Ia), 【Chemistry 1】 During the ceremony, X １ , X ２ and X ５ One of them is either CH or CF, independently of each other. R １ and R ２ However, each is independent of the following: i) An amino acid element comprising one or more amino acids selected from the group consisting of glycine, alanine, serine, threonine, cysteine, valine, leucine, isoleucine, methionine, proline, phenylalanine, tyrosine, tryptophan, aspartic acid, glutamic acid, asparagine, glutamine, histidine, lysine, arginine, sarconsine, and beta-alanine, ii) A charged element comprising one or more charged amino acids, one or more carboxylic acids, one or more sulfonic acids, one or more sulfonamides, one or more sulfates, one or more phosphates, one or more quaternary amines, one or more sulfamides, one or more sulfinimides, or a combination thereof, at pH 7.4 iii) Below: 【Chemistry 2】 A heteroalkylene element containing one of the structures, In the formula, each occurrence of R 5b, R 5c, R 5d, and R 5e is independently selected from the group consisting of hydrogen, deuterium, alkyl, haloalkyl, halo, alkoxy, haloalkoxy, amino, hydroxyl, cyano, nitro, thiol, carboxyalkyl, alkyl-S(O)3H, alkyl-O-P(O)3H, alkyl-P(O)3H, -O-carboxyalkyl, -O-alkyl-S(O)3H, -O-alkyl-O-P(O)3H, -O-alkyl-P(O)3H, -S(O)3H, -OP(O)3H, and -P(O)3H; each occurrence of q1 is independently an integer from 1 to 24, and the stereoisomer is a heteroalkylene element. iv) Hydrophilic elements comprising polyethylene glycol, polysarcosine, cyclodextrin, c-glycoside, or combinations thereof, v) A trigger element comprising a phosphate, diphosphate, triphosphate, glucuronide, or one or more amino acids selected from the group consisting of valine, citrulline, alanine, glycine, phenylalanine, lysine, asparagine, or combinations thereof. vi) An immolative element, including para-aminobenzyloxycarbonyl, aminal, hydrazine, disulfide, amide, ester, phosphotriester, diester, β-glucuronide, double bond, triple bond, ether bond, ketone, diol, cyano, nitro, quaternary amine, or combination thereof. vii) A polar cap containing -OH, -NH₂, -C(=O)OH, -S(O)₃H, or -OP(O)₃ viiii) A payload that is a glucocorticoid receptor agonist, chemotherapeutic agent, cytotoxic agent, or myeloid cell agonist, It includes one or more parts selected from, However, R １ and R ２ At least one of them contains a payload, R ４ａ and R ４ｂ are each independently hydrogen, deuterium, halo, or -S-R ４ｃ where R ４ｃ is substituted or unsubstituted C ６ to C １０ aryl or substituted or unsubstituted 5- to 12-membered heteroaryl, and L １ , and L ２ However, each is independent of the following: 【Transformation 3】 It has a structure, In the formula, * indicates a direct bond to the ring carbon shown in structural formula (Ia); Ra a is hydrogen or alkyl; each occurrence of L b is independently a direct bond, an optionally substituted alkylene linker, an optionally substituted heteroalkylene linker, a heteroatom linker, or a combination thereof; each occurrence of L c is independently an optionally substituted alkylene linker. Compounds, or their stereoisomers, enantiomers, or tautomers, mixtures thereof, or pharmaceutically acceptable salts or solvates thereof.

2. a) The amino acid element comprises one or more amino acids selected from the group consisting of glycine, sarcosine, beta-alanine, and glutamic acid; b) The amino acid element comprises a dipeptide, tripeptide, tetrapeptide, or pentapeptide; c) The amino acid elements are as follows: 【Chemistry 4】 It has one of the structures, in the formula, R ５ａ Each occurrence is independently hydrogen, alkyl, hydroxyalkyl, or alkoxyalkyl; d) The charged element includes a portion that has a negative charge at pH 7.4; e) The charged element includes a portion that has a positive charge at pH 7.4; or, f) The charged element comprises one or more charged amino acids, one or more carboxylic acids, one or more sulfonic acids, one or more sulfonamides, one or more sulfates, one or more phosphates, one or more quaternary amines, one or more sulfamides, one or more sulfinimides, or a combination thereof, wherein the charged amino acid is aspartic acid, glutamic acid, histidine, lysine, or arginine. A compound as described in claim 1, or a stereoisomer, enantiomer, or tautomer thereof, a mixture thereof, or a pharmaceutically acceptable salt or solvate thereof.

3. R １ or R ２ However, the following: 【Transformation 5】 It further includes one of the structures of, in the formula, R ５ｂ , R ５ｃ R ５ｄ , and R ５ｅ Each occurrence is independently hydrogen, deuterium, alkyl, haloalkyl, halo, alkoxy, haloalkoxy, amino, hydroxyl, cyano, nitro, thiol, carboxyalkyl, alkyl-S(O) ３ H, alkyl-O-P(O) ３ H, alkyl-P(O) ３ H,-O-carboxyalkyl,-O-alkyl-S(O) ３ H,-O-alkyl-O-P(O) ３ H,-O-alkyl-P(O) ３ H, -S(O) ３ H, -OP(O) ３ H, -P(O) ３ H, alkyl-O-P(O) ３ -Alkyl, alkyl-P(O) ３ -alkyl, -O-alkyl-S(O) ３ -alkyl, -O-alkyl-O-P(O) ３ -alkyl, -O-alkyl-P(O) ３ -alkyl, -S(O) ３ -Alkyl, -OP(O) ３ -alkyl, -P(O) ３ - Selected from the group consisting of alkyl, sulfamide, and sulfinimide, R ５ｆ Each occurrence is independently hydrogen, alkyl, hydroxyalkyl, or alkoxyalkyl. R ９ Each occurrence is independently hydrogen or alkyl. Each occurrence of q2 is an integer between 1 and 25, Each occurrence of q3 is an integer between 5 and 15, independently. A compound as described in claim 1, or a stereoisomer, enantiomer, or tautomer thereof, a mixture thereof, or a pharmaceutically acceptable salt or solvate thereof.

4. The hydrophilic element is as follows: 【Transformation 6】 It includes one of the structures, in the formula, R ５ｂ , R ５ｃ R ５ｄ , and R ５ｅ Each occurrence is independently hydrogen, deuterium, alkyl, haloalkyl, halo, alkoxy, haloalkoxy, amino, hydroxyl, cyano, nitro, thiol, carboxyalkyl, alkyl-S(O) ３ H, alkyl-O-P(O) ３ H, alkyl-P(O) ３ H,-O-carboxyalkyl,-O-alkyl-S(O) ３ H,-O-alkyl-O-P(O) ３ H,-O-alkyl-P(O) ３ H, -S(O) ３ H, -OP(O) ３ H, and -P(O) ３ Selected from the group consisting of H, R ５ｇ Each occurrence is independently hydrogen, alkyl, hydroxyalkyl, or alkoxyalkyl. Each occurrence of q4 is an integer between 1 and 24, independently. A compound as described in claim 1, or a stereoisomer, enantiomer, or tautomer thereof, a mixture thereof, or a pharmaceutically acceptable salt or solvate thereof.

5. The trigger element is a) Asparagine, b) Dipeptides, tripeptides, tetrapeptides, pentapeptides, glucuronides, disulfides, phosphates, diphosphates, triphosphates, hydrazones, or combinations thereof c) Beta-glucuronic acid, d) A sequence of amino acids selected from the group consisting of valine-citrulline, valine-alanine, glycine-glycine-phenylalanine-glycine, and combinations thereof. e) 【Transformation 7】 One of the structures (including combinations thereof), f) The sacrificial element includes paramethoxybenzyl, dialkyldialkoxysilane, diaryldialkoxysilane, orthoester, acetal, optionally substituted β-thiopropionate, ketal, phosphoramide, hydrazone, vinyl ether, imine, aconityl, trityl, polyketal, bis-arylhydrazone, diazobenzene, vivinal diol, pyrophosphate diester, or combinations thereof, or stereoisomers, enantiomers or tautomers thereof, mixtures thereof, or pharmaceutically acceptable salts, solvates or prodrugs thereof. g) The sacrificial elements are as follows: 【Transformation 8】 Includes one of the structures, h) The sacrificial elements are as follows: 【Chemistry 9】 Structure [In the formula, R ６ａ , R ６ｂ , R ６ｃ , and R ６ｄ However, independently, these are hydrogen, optionally substituted alkyl, optionally substituted aryl, or optionally substituted heteroaryl, or R ６ａ and R ６ｃ However, together with the nitrogen and carbon atoms to which they are bonded, they form azetidinyl, pyrrolodinyl, piperidinyl, or homopiperidinyl, R ６ｄ However, it is hydrogen, Y １ However, -O-, -S-, or -NR ６ｂ -is] including, i) The sacrificial elements are as follows: 【Chemistry 10】 Structure [In the formula, R ６ｅ , R ６ｆ , R ６ｇ , and R ６ｈ However, independently, these are hydrogen, optionally substituted alkyl, optionally substituted aryl, or optionally substituted heteroaryl, or R ６ａ and R ６ｃ However, together with the nitrogen and carbon atoms to which they are bonded, they form azetidinyl, pyrrolodinyl, piperidinyl, or homopiperidinyl, R ６ｄ However, it is hydrogen, Y ２ However, -O-, -S-, or -NR ６ｆ -is] including, j) The aforementioned sacrificial elements are as follows: 【Chemistry 11】 Structure [In the formula, R １０ Each occurrence is independently alkyl, alkoxy, or halo. R １１ However, hydrogen, alkyl, or -(CH ２ CH ２ O) ｚ３ -CH ３ And, R １２ However, it is hydrogen or alkyl, R １３ However, it is hydrogen or alkyl, z1 is 0 or 1, z2 is 0, 1, 2, 3, or 4, z3 is 0, 1, 2, 3, 4, 5, 6, 7, 8, 9, or 10. including, or k) The sacrificial elements are as follows: 【Chemistry 12】 One of the structures [in the formula, R １４ａ , R １４ｂ , R １４ｃ , R １４ｄ , R １４ｅ , and R １４ｆ However, each is independently hydrogen, alkyl, hydroxyalkyl, or alkoxyalkyl, [Includes z4, z5, z6, and z7 being independently 1, 2, 3, 4, 5, or 6] l) The sacrificial elements are as follows: 【Chemistry 13】 One of the structures [in the formula, z8 and z9 are independently 1, 2, 3, 4, 5, or 6. including, or m) The sacrificial elements are as follows: 【Chemistry 14】 One of the structures [in the formula, R １５ Each of the occurrences is independently H, methyl, ethyl, isopropyl, tert-butyl, or phenyl. Y ３ is O or CH ２ and [Includes cases where q5 is an integer between 1 and 5] A compound as described in claim 1, or a stereoisomer, enantiomer, or tautomer thereof, a mixture thereof, or a pharmaceutically acceptable salt or solvate thereof.

6. a) R ４ａ and R ４ｂ However, both are hydrogen; or, b) The trigger element and the sacrificial element are both as follows: 【Chemistry 15-1】 【Chemistry 15-2】 Includes one of the structures, A compound as described in claim 1, or a stereoisomer, enantiomer, or tautomer thereof, a mixture thereof, or a pharmaceutically acceptable salt or solvate thereof.

7. The aforementioned polarity cap, a) comprising one or more charged amino acids, one or more polyols, or a combination thereof b) Diols, triols, tetraols, or combinations thereof c) Glycerol, trimethylolpropane, pentaerythritol, maltitol, sorbitol, xylitol, erythritol, isomalt, or combinations thereof, d) Contains one or more natural amino acids, e) Containing one or more non-natural amino acids, f) Containing one or more non-natural amino acids and one or more natural amino acids, g) containing serine, threonine, cysteine, proline, asparagine, glutamine, lysine, arginine, histidine, aspartate, glutamate, 4-hydroxyproline, 5-hydroxylysine, homoserine, homocysteine, ornithine, beta-alanine, statin, or gamma-aminobutyric acid, or h) containing aspartic acid, serine, glutamic acid, serine-beta-glucose, or a combination thereof i) Below: 【Chemistry 16】 Having one of the structures (including combinations thereof), A compound as described in claim 1, or a stereoisomer, enantiomer, or tautomer thereof, a mixture thereof, or a pharmaceutically acceptable salt or solvate thereof.

8. The aforementioned payload, a) A chemotherapy agent selected from the group consisting of camptothecin, paclitaxel, doxorubicin, vinblastine, dacarbazine, irinotecan, topotecan, siratecan, cocitecan, exatecan, lulutotecan, SN-38, Dxd, gimatecan, berotecan, and rubitecan. b) A cytotoxic agent selected from the group consisting of calicheamicin, anthramycin, abeimycin, thikamycin, DC-81, mazetramycin, neotramycin A, neotramycin B, polotoramycin proslacalcin, cibanomycin, cibilomycin, tomamycin, auristatin F, monomethyl auristatin F, auristatin E, monomethyl auristatin E, drastatin, monomethyl drastatin, meltansine, and emtansine, or c) STING agonists, TLR2 ligands, TLR3 ligands, TLR4 ligands, TLR5 ligands, TLR6 ligands, TLR7 ligands, TLR8 ligands, TLR9 ligands, TLR10 ligands, nucleotide oligomerization domain (NOD) ligands, RIG-I-like receptor (RLR) ligands, C-type lectin receptor (CLR) ligands, cytoplasmic DNA sensor (CDS) ligands, and inflammasome-inducible factor ligands A myeloid cell agonist selected from the group consisting of, preferably, the myeloid cell agonist is a myeloid cell agonist selected from the group consisting of, selgantrimod, motlimod, resikimod, 3M-051, 3M-052, MCT-465, IMO-4200, VTX-763, VTX-1463, RG7854, ADU-S100, MK-1454, MK-2118, BMS-986301, GSK3745417, SB-11285, and IMSA-101. A compound as described in claim 1, or a stereoisomer, enantiomer, or tautomer thereof, a mixture thereof, or a pharmaceutically acceptable salt or solvate thereof.

9. L ｃ but, a) C １ ~C ６ It is alkylene; or b) Substituted with one or more substituents selected from the group consisting of halo, haloalkyl, alkoxy, cyano, nitro, carboxy, sulfonamide, sulfonic acid, or combinations thereof, A compound as described in claim 1, or a stereoisomer, enantiomer, or tautomer thereof, a mixture thereof, or a pharmaceutically acceptable salt or solvate thereof.

10. a) X １ is C-F and X ５ is C-H; or b) X １ However, C-H and X ５ However, it is C-F. A compound as described in claim 1, or a stereoisomer, enantiomer, or tautomer thereof, a mixture thereof, or a pharmaceutically acceptable salt or solvate thereof.

11. a) The compound has the following structures (Ia-1), (Ia-2), (Ia-3), (Ia-4), (Ia-5), (Ia-6), (Ia-7), or (Ia-8) 【Chemistry 17】 It possesses one of these, and in the formula, L ｂ Each occurrence is, independently, a direct bond, an optionally substituted alkylene linker, an optionally substituted heteroalkylene linker, a heteroatom linker, or a combination thereof, and q6 is 0, 1, or 2; b) The compound has the following structures (Id), (Ie), (If), or (Ig) [Chemistry 18] It has, in the formula, L ｂ Each occurrence is independently a direct bond, an optionally substituted alkylene linker, an optionally substituted heteroalkylene linker, a heteroatom linker, or a combination thereof. q8 is 0, 1, or 2, and, q9 is 0, 1, or 2; or, c) The compound has the following structure (Ih) or (Ii) 【Chemistry 19】 It possesses one of these, and in the formula, L ｂ Each occurrence is independently a direct bond, an optionally substituted alkylene linker, an optionally substituted heteroalkylene linker, a heteroatom linker, or a combination thereof. A compound as described in claim 1, or a stereoisomer, enantiomer, or tautomer thereof, a mixture thereof, or a pharmaceutically acceptable salt or solvate thereof.

12. a) L ｂ However, it is directly bonded, optionally substituted alkylene linkers, or optionally substituted heteroalkylene linkers; or, b) L ｂ However, it is either a direct link or the following: 【Chemistry 20】 It has one of the structures, in the formula, R ｂ Each occurrence is independently hydrogen, alkyl, hydroxyalkyl, or alkoxyalkyl. A compound as described in claim 1, or a stereoisomer, enantiomer, or tautomer thereof, a mixture thereof, or a pharmaceutically acceptable salt or solvate thereof.

13. a) The compound is as follows: 【Chemistry 21-1】 【Chemistry 21-2】 【Chemistry 21-3】 It has one of the structures, in the formula, R １ｂ However, it is a chemotherapy drug, a cytotoxic agent, or a myeloid cell agonist. R ２ｂ However, the following: 【Chemistry 22】 It has one of the structures, L １ｇ However, the following: 【Chemistry 23-1】 【Chemistry 23-2】 Having one of the structures of; or, b) The compound is as follows: 【Chemistry 24-1】 【Chemistry 24-2】 【Chemistry 24-3】 【Chemistry 24-4】 【Chemistry 24-5】 【Chemistry 24-6】 【Chemistry 24-7】 【Chemistry 24-8】 Having one of the structures, A compound as described in claim 1, or a stereoisomer, enantiomer, or tautomer thereof, a mixture thereof, or a pharmaceutically acceptable salt or solvate thereof.

14. a) R １ａ However, the following: 【Chemistry 25-1】 【Chemistry 25-2】 【Chemistry 25-3】 It has one of the following structures; b) R １ａ However, the following: 【Chemistry 26】 It has one of the structures, in the formula, R' is either hydrogen or one of the following: 【Chemistry 27】 It has one of the structures, in the formula, R ａ 'but H or C １～６ It is alkyl, R ｂ 'But, C １～６ Alkyl or C １～６ It is an alkoxy, R ｃ 'But, H, C １～６ Alkyl, -CH ２ OH, or C １～６ It is an alkoxy, R ｄ 'but H or C １～６ It is alkyl, or R ｅ 'but H or C １～６ It is alkyl. The compound described in claim 13, or its stereoisomers, enantiomers, or tautomers, mixtures thereof, or pharmaceutically acceptable salts or solvates thereof.

15. The following: 【Chemistry 28-1】 【Chemistry 28-2】 【Chemistry 28-3】 【Chemistry 28-4】 【Chemistry 28-5】 【Chemistry 28-6】 【Chemistry 28-7】 【Chemistry 28-8】 [Chemistry 28-9] 【Chemistry 28-10】 【Chemistry 28-11】 【Chemistry 28-12】 【Chemistry 28-13】 [Chemistry 28-14] 【Chemistry 28-15】 [Chemistry 28-16] 【Chemistry 28-17】 【Chemistry 28-18】 [Chemistry 28-19] 【Chemistry 28-20】 【Chemistry 28-21】 A compound having one of the structures of, or a stereoisomer, enantiomer, or tautomer thereof, a mixture thereof, or a pharmaceutically acceptable salt or solvate thereof.

16. A conjugate having the following structure (IIa), 【Chemistry 29】 (IIa) During the ceremony, A is the target portion or its binding fragment, L ４ However, the following: 【Transformation 30】 It has one of the structures, *** indicates the connection point to A, g is an integer between 1 and 20. X １ , X ２ and X ５ One of them is either CH or CF, independently of each other. R １ and R ２ However, each is independent of the following: i) An amino acid element comprising one or more amino acids selected from the group consisting of glycine, alanine, serine, threonine, cysteine, valine, leucine, isoleucine, methionine, proline, phenylalanine, tyrosine, tryptophan, aspartic acid, glutamic acid, asparagine, glutamine, histidine, lysine, arginine, sarconsine, and beta-alanine, ii) A charged element comprising one or more charged amino acids, one or more carboxylic acids, one or more sulfonic acids, one or more sulfonamides, one or more sulfates, one or more phosphates, one or more quaternary amines, one or more sulfamides, one or more sulfinimides, or a combination thereof, at pH 7.4 iii) Below: 【Chemistry 31】 A heteroalkylene element containing one of the structures, In the formula, each occurrence of R 5b, R 5c, R 5d, and R 5e is independently selected from the group consisting of hydrogen, deuterium, alkyl, haloalkyl, halo, alkoxy, haloalkoxy, amino, hydroxyl, cyano, nitro, thiol, carboxyalkyl, alkyl-S(O)3H, alkyl-O-P(O)3H, alkyl-P(O)3H, -O-carboxyalkyl, -O-alkyl-S(O)3H, -O-alkyl-O-P(O)3H, -O-alkyl-P(O)3H, -S(O)3H, -OP(O)3H, and -P(O)3H; each occurrence of q1 is independently an integer from 1 to 24, and the stereoisomer is a heteroalkylene element. iv) Hydrophilic elements comprising polyethylene glycol, polysarcosine, cyclodextrin, c-glycoside, or combinations thereof, v) A trigger element comprising a phosphate, diphosphate, triphosphate, glucuronide, or one or more amino acids selected from the group consisting of valine, citrulline, alanine, glycine, phenylalanine, lysine, asparagine, or combinations thereof. vi) Sacrificial elements including para-aminobenzyloxycarbonyl, aminal, hydrazine, disulfide, amide, ester, phosphotriester, diester, β-glucuronide, double bond, triple bond, ether bond, ketone, diol, cyano, nitro, quaternary amine, or combinations thereof, vii) A polar cap containing -OH, -NH₂, -C(=O)OH, -S(O)₃H, or -OP(O)₃ viiii) A payload that is a glucocorticoid receptor agonist, chemotherapeutic agent, cytotoxic agent, or myeloid cell agonist, It includes one or more parts selected from, However, R １ and R ２ At least one of them contains a payload, R ４ａ However, hydrogen, deuterium, halo, or -S-R ４ｃ And R ４ｃ However, substitution or non-substitution C ６ ~C １０ It is an aryl or a substituted or unsubstituted 5- to 12-membered heteroaryl, and, L １ and L ２ but, Each independently, as follows: 【Chemistry 32】 It has a structure, In the formula, * indicates a direct bond to the ring carbon shown in structural formula (Ia); Ra a is hydrogen or alkyl; each occurrence of L b is independently a direct bond, an optionally substituted alkylene linker, an optionally substituted heteroalkylene linker, a heteroatom linker, or a combination thereof; each occurrence of L c is independently an optionally substituted alkylene linker. Compounds, or their stereoisomers, enantiomers, or tautomers, mixtures thereof, or pharmaceutically acceptable salts or solvates thereof.

17. A pharmaceutical composition comprising the conjugate described in claim 16 and a pharmaceutically acceptable excipient.

18. A method for treating cancer, infection, inflammation, autoimmune disorder, or a combination thereof, the method comprising administering an effective amount of the conjugate according to claim 16 or the pharmaceutical composition according to claim 17 to a subject in need thereof.

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