RAS inhibitors

Abstract

This invention provides a macrocyclic compound capable of inhibiting the Ras protein, and a pharmaceutical composition thereof. [Solution] Examples of compounds relating to this disclosure are compounds having the structure of formula I, or pharmaceutically acceptable salts thereof. JPEG2026048800000188.jpg70170

Description

[Background technology]

[0001] The vast majority of small molecule drugs act by binding to functionally important pockets on target proteins, thereby regulating the activity of those proteins. For example, cholesterol-lowering drugs known as statins bind to the enzymatic active site of HMG-CoA reductase, thus preventing the enzyme from engaging with its substrate. The fact that many such drug / target interaction pairs are known can be misleading to some extent, leading people to believe that, given a reasonable amount of time, effort, and resources, small molecule modulators can be discovered for most, if not all, proteins. This is far from the truth. Current estimates suggest that only about 10% of all human proteins are targetable by small molecules. Non-patent document 1. The other 90% are currently considered refractory or refractory to the small molecule drug discovery described above. Such targets are commonly referred to as "undruggable." These undruggable targets represent a vast and large, untapped reservoir of clinically important human proteins. Therefore, there is considerable interest in discovering novel molecular modalities that can control the function of such undruggable targets.

[0002] The literature has well established that Ras proteins (K-Ras, H-Ras, and N-Ras) play essential roles in various human cancers and are therefore appropriate targets for anti-cancer therapies. In fact, mutations in the Ras protein account for approximately 30% of all human cancers in the United States, many of which are lethal. Dysregulation of the Ras protein due to mutation activation, overexpression, or upstream expression is common in human tumors, and mutation activation in Ras is frequently found in human cancers. For example, activation of a mutation at codon 12 in the Ras protein functions by inhibiting both the GTPase-activating protein (GAP)-dependent and intrinsic hydrolysis rates of GTP, significantly distorting the population of Ras mutant proteins into an "on" (GTP-bound) state (Ras(ON)), leading to oncogenic MAPK signaling. In particular, Ras exhibits picomolar affinity for GTP, allowing Ras to be activated even in the presence of low concentrations of this nucleotide. Mutations in codon 13 of Ras (e.g., G13D) and codon 61 (e.g., Q61K) also contribute to oncogenic activity in some cancers.

[0003] Despite extensive drug discovery efforts against Ras in recent decades, only drugs targeting the K-Ras G12C variant have been approved (sotracib). Further efforts are needed to identify additional drugs for cancers driven by other Ras mutations. [Prior art documents] [Non-patent literature]

[0004] [Non-Patent Document 1] Bojadzic and Buchwald,Curr Top Med Chem 18:674-699(2019) [Overview of the project]

[0005] Ras inhibitors are provided herein. The approach described herein requires the formation of a high-affinity triplicate or conjugate between a synthetic ligand and two intracellular proteins that do not interact under normal physiological conditions: the target protein (e.g., Ras) and a cytosolic chaperone (presenter protein) that is widely expressed intracellularly (e.g., cyclophyllin A). More specifically, in some embodiments, the Ras inhibitors described herein include a novel binding pocket cytosol in Ras, which drives the formation of a high-affinity triplicate or conjugate between the Ras protein and the widely expressed cytosolic chaperone cyclophyllin A (CYPA). While not theoretically bound, one way in which the inhibitory effect on Ras is affected by the compounds and complexes or conjugates of the present invention is the formation of steric occlusion of the interaction site between Ras and downstream effector molecules such as RAF and PI3K, which the inventors believe is necessary for the growth of oncogenic signals.

[0006] Therefore, in some embodiments, the present invention relates to a compound of structural formula I, or a pharmaceutically acceptable salt thereof:

[0007] [ka]

[0008] [In the formula, A is an optionally substituted 3-6 member heterocycloalkylene, an optionally substituted 3-6 member cycloalkylene, an optionally substituted 6 member arylene, or an optionally substituted 5-10 member heteroarylene.] X 1 , X 2 , and X 3 Each of these is independently selected from CH2, CF2, C=O, or O. m is either 1 or 2. n is either 0 or 1. R 1is hydrogen, optionally substituted C1-C6 heteroalkyl, or optionally substituted 3-10 member heterocycloalkyl. R 2 is an arbitrarily substituted C1-C6 alkyl, and R 3 This is an optionally substituted C1-C6 alkyl group, an optionally substituted 3-6 member cycloalkyl group, or an optionally substituted heterocycloalkyl group. Each hydrogen atom is independently an arbitrarily isotopically enriched deuterium.

[0009] In some embodiments, the present invention is characterized by a compound of structural formula II, or a pharmaceutically acceptable salt thereof:

[0010] [ka]

[0011] [In the formula, A is an optionally substituted 3-6 member heterocycloalkylene, an optionally substituted 3-6 member cycloalkylene, an optionally substituted 6 member arylene, or an optionally substituted 5-10 member heteroarylene.] R 2 is an arbitrarily substituted C1-C6 alkyl, and R 3 This is an optionally substituted C1-C6 alkyl group, an optionally substituted 3-6 member cycloalkyl group, or an optionally substituted heterocycloalkyl group. Each hydrogen atom is independently an arbitrarily isotopically enriched deuterium.

[0012] In some embodiments, the present invention is characterized by a compound of structural formula V, or a pharmaceutically acceptable salt thereof:

[0013] [ka]

[0014] [wherein, A is optionally substituted 3- to 6-membered heterocycloalkylene, optionally substituted 3- to 6-membered cycloalkylene, optionally substituted 6-membered arylene, or optionally substituted 5- to 10-membered heteroarylene, R 2 is optionally substituted C1-C6 alkyl, and R 3 is optionally substituted C_{1}-C_{6} alkyl, optionally substituted 3- to 6-membered cycloalkyl, or optionally substituted heterocycloalkyl, each hydrogen is independently optionally isotope-enriched deuterium].

[0015] In some embodiments, the present invention features a compound of Structural Formula VI, or a pharmaceutically acceptable salt thereof:

[0016]

Chemical Formula

[0017] [wherein, A is optionally substituted 3- to 6-membered heterocycloalkylene, optionally substituted 3- to 6-membered cycloalkylene, optionally substituted 6-membered arylene, or optionally substituted 5- to 10-membered heteroarylene, R 2 is optionally substituted C1-C6 alkyl, and R 3 is optionally substituted C_{1}-C_{6} alkyl, optionally substituted 3- to 6-membered cycloalkyl, or optionally substituted heterocycloalkyl, each hydrogen is independently optionally isotope-enriched deuterium].

[0018] In some embodiments, the present invention features a compound of Structural Formula VII, or a pharmaceutically acceptable salt thereof:

[0019]

Chemical Formula

[0020] [In the formula, A is an optionally substituted 3-6 member heterocycloalkylene, an optionally substituted 3-6 member cycloalkylene, an optionally substituted 6 member arylene, or an optionally substituted 5-10 member heteroarylene.] R 2 is an arbitrarily substituted C1-C6 alkyl, and R 3 This is an optionally substituted C1-C6 alkyl group, an optionally substituted 3-6 member cycloalkyl group, or an optionally substituted heterocycloalkyl group. Each hydrogen atom is independently an arbitrarily isotopically enriched deuterium.

[0021] In some embodiments, the present invention also features compounds selected from Table 1 or Table 2, or pharmaceutically acceptable salts thereof. Pharmaceutical compositions comprising compounds of formula I, formula II, formula V, formula VI, or formula VII, or pharmaceutically acceptable salts thereof, and pharmaceutically acceptable excipients are also provided. Pharmaceutical compositions comprising compounds of Table 1 or Table 2, or pharmaceutically acceptable salts thereof, and pharmaceutically acceptable excipients are also provided.

[0022] The present invention also provides a method for treating cancer in a subject requiring cancer treatment, wherein the method comprises administering to the subject a therapeutically effective amount of the compound of the present invention, or a pharmaceutically acceptable salt thereof.

[0023] In some embodiments, the present invention provides a method for treating a Ras protein-related disease in a subject requiring treatment for the Ras protein-related disease, the method comprising administering to the subject a therapeutically effective amount of the compound of the present invention or a pharmaceutically acceptable salt thereof.

[0024] A method for inhibiting the Ras protein in cells is provided, further comprising contacting the cells with an effective amount of the compound of the present invention or a pharmaceutically acceptable salt thereof.

[0025] Any limitations discussed in relation to one embodiment of the present invention can be specifically conceived to apply to any other embodiment of the present invention. Furthermore, any compound or composition of the present invention can be used in any manner of the present invention, and any compound or composition of the present invention can be produced or utilized using any manner of the present invention.

[0026] Definitions and Chemical Terms In this application, unless otherwise clearly indicated by context, (i) the term “one (a)” means “one or more”; (ii) is used to mean “and / or” unless it is explicitly indicated that it means only alternative expressions or that such alternative expressions are mutually exclusive; however, the present invention supports definitions that refer only to alternative expressions and to “and / or”; (iii) the terms “comprising” and “including” are understood to encompass itemized components or processes, whether presented by themselves or together with one or more additional components or processes; and (iv) where a scope is indicated, it includes endpoints.

[0027] As used herein, the term “approximately” is used to indicate that a value includes the standard deviation of the error of the device or method used to determine the value. In certain embodiments, unless otherwise stated or evident from the content (for example, if such a number may exceed 100% of the possible values), the term “approximately” refers to a range of values ​​that fall within 25%, 20%, 19%, 18%, 17%, 16%, 15%, 14%, 13%, 12%, 11%, 10%, 9%, 8%, 7%, 6%, 5%, 4%, 3%, 2%, and 1% or less in either direction (above or below) the stated value.

[0028] As used herein, the term "adjacent" in the context of describing adjacent atoms means divalent atoms directly bonded by a covalent bond. As used herein, “compounds of the present invention” and similar terms mean, whether expressly stated or not, compounds of any one of formulas I to VII or their subformulas, as well as the compounds of Table 1 or Table 2, in addition to their salts (e.g., pharmaceutically acceptable salts), solvates, hydrates, stereoisomers (including atropisomers), and tautomers, as described herein, the Ras inhibitors.

[0029] The term "wild-type" refers to an entity possessing a structure or activity found in nature in a "normal" state or context (as opposed to variants, diseases, or changes). Those skilled in the art will understand that wild-type genes and polypeptides often exist in multiple different forms (e.g., alleles).

[0030] Those skilled in the art will understand that certain compounds described herein may exist in one or more different isomeric forms (e.g., stereoisomers, geometric isomers, atropisomers, tautomers) or isotopic forms (e.g., hydrogen substituted with deuterium, in which one or more atoms are substituted with different isotopes of that atom). Unless otherwise specified or made clear from the context, the structures described may be understood to represent any such isomeric or isotopic forms, individually or in combination.

[0031] The compounds described herein may be asymmetric (for example, having one or more stereocenters). All stereoisomers, such as enantiomers and diastereomers, are intended unless otherwise specified. Compounds of the present invention containing asymmetrically substituted carbon atoms can be isolated in optically active forms or as racemates. Methods for preparing optically active forms from optically active starting materials, such as by resolution of racemic mixtures or stereoselective synthesis, are known in the art. Many geometric isomers, such as olefins and C=N double bonds, can also be present in the compounds described herein, and all such stable isomers are intended in the present invention. Cis and trans geometric isomers of the compounds of the present invention are described and can be isolated as mixtures of isomers or as separated isomers.

[0032] In some embodiments, one or more compounds described herein may exist in different tautomerized forms. As will be apparent from the context, unless explicitly excluded, references to such compounds encompass all such tautomerized forms. In some embodiments, the tautomerized form results from the exchange of a single bond with an adjacent double bond and the accompanying transfer of a proton. In certain embodiments, the tautomerized form may be a prototropic tautomer, which is a protonated state of an isomer having the same empirical formula and total charge as the reference form. Examples of moieties having a prototropic tautomerized form include ketone-enol pairs, amide-imoid acid pairs, lactam-lactim pairs, amide-imoid acid pairs, enamine-imine pairs, and cyclic forms in which a proton can occupy two or more positions in a heterocyclic system, such as 1H- and 3H-imidazoles, 1H-, 2H-, and 4H-1,2,4-triazoles, 1H- and 2H-isoindoles, and 1H- and 2H-pyrazoles. In some embodiments, the tautomer morphs may be in equilibrium or sterically fixed to one morph by appropriate substitution. In certain embodiments, the tautomer morphs arise from acetal interconversion.

[0033] Unless otherwise specified, the structures shown herein also mean that they include compounds that differ only in the presence of one or more isotopic enriched atoms. Exemplary isotopes that can be incorporated into the compounds of the present invention include: 2 H, 3 H, 11 C, 13 C, 14 C, 13 N, 15 N, 15 O, 17 O, 18 O, 32 P, 33 P, 35 S, 18 F, 36 Cl, 123 I, and 125 Examples of isotopes include hydrogen, carbon, nitrogen, oxygen, phosphorus, sulfur, fluorine, chlorine, and iodine, such as I. Isotope-labeled compounds (e.g., 3 H and 14 Compounds labeled with 1C can be useful in compound or substrate tissue partitioning assays. Tritium labeling (i.e., 3 H), and carbon-14 (i.e., 14 C) Isotopes can be useful due to their ease of preparation and detection. Furthermore, heavier isotopes, such as deuterium (i.e., 2 Substitution with H) etc. can lead to greater metabolic stability, which may result in certain therapeutic benefits (e.g., longer in vivo half-life or lower required dose). In some embodiments, one or more hydrogen atoms are 2 H or 3 Replaced by H, or one or more carbon atoms 13 C or 14 It is replaced by carbon-rich carbon. 15 O, 13 N, 11 C and 18Positron-emitting isotopes such as fluorine are useful in positron emission tomography (PET) studies to examine substrate receptor occupancy. The preparation of isotopically labeled compounds is known to those skilled in the art. For example, isotopically labeled compounds can generally be prepared by replacing unlabeled reagents with isotopically labeled reagents, following a procedure similar to the procedure disclosed for the compounds of the present invention as described herein.

[0034] As used interchangeably in this specification, “deuterium substitution,” “deuterated,” or “deuterium enrichment” refers to deuterium (D or 2 H) means a compound or part thereof of the present invention having a level. In certain embodiments, the composition of the present invention has a minimum isotopic enrichment factor of at least 5 (0.075% deuterium incorporated), for example, at least 10 (0.15% deuterium incorporated). In other embodiments, the composition has an isotopic enrichment factor of at least 50 (0.75% deuterium incorporated), at least 500 (7.5% deuterium incorporated), at least 2000 (30% deuterium incorporated), at least 3000 (45% deuterium incorporated), at least 4000 (60% deuterium incorporated), at least 4500 (67.5% deuterium incorporated), at least 5000 (75% deuterium incorporated), at least 5500 (82.5% deuterium incorporated), at least 6000 (90% deuterium incorporated), or at least 6600 (99% deuterium incorporated).

[0035] Non-limiting examples of a compound of the present invention that may contain one or more deuterium substitutions (where any position "R" is deuterium (D)) include:

[0036] [ka]

[0037] Examples include:

[0038] [ka]

[0039] Furthermore, similar R 1 One example is the deuteration of the mold portion, R 1 The definition of is found herein (for example, in any one of the compounds of formulas I to VII). Deuteration of a portion of a crosslinking group (for example, an optionally substituted aziridine moiety) in the compounds of the present invention is also contemplated, where the crosslinking group is as defined herein (for example, general formulas I to VII, as well as their subformulas, in addition to specific examples of W described herein, e.g.

[0040] [ka]

[0041] (See also).

[0042] [ka]

[0043] Deuteration at any available position in the A portion of the compounds of the formulas described herein is also intended. Furthermore, deuterium substitution is also,

[0044] [ka]

[0045] The compounds of the present invention can be formed at the linker position of the compounds of the formulas described herein. In further embodiments, silylation substitutions, such as those at the linker position as follows, are also considered:

[0046] [ka]

[0047] An example of additional silylation is,

[0048] [ka]

[0049] parts such as and similar R 1 Silylation of the mold portion is one example, R 1 The definition is found herein (for example, in any one of the compounds of formulas I to VII). As is known from the prior art, many chemical components can be used in various different solid forms, such as amorphous or crystalline forms (e.g., polymorphs, hydrates, solvates). In some embodiments, the compounds of the present invention can be used in any such form, including any solid form. In some embodiments, the compounds described or explained herein can be provided or used in hydrate or solvate form.

[0050] In various parts of this specification, substituents of the compounds of the present invention are disclosed in groups or ranges. It is specifically intended that the present invention includes each individual partial combination of members of such groups and ranges. For example, the term “C1-C6 alkyl” is specifically intended to disclose methyl, ethyl, C3 alkyl, C4 alkyl, C5 alkyl, and C6 alkyl individually. Furthermore, if a compound includes multiple positions where substituents are disclosed in groups or ranges, unless otherwise specified, the present invention is intended to extend to individual compounds containing each and all individual element subcombinations at each position, as well as to groups of compounds (e.g., genera and sub-genera).

[0051] The term “optionally substituted X” (e.g., optionally substituted alkyl) is intended to be equivalent to “X, where X is optionally substituted” (e.g., “alkyl, where the alkyl is optionally substituted”). It is not intended to mean that the feature “X” (e.g., alkyl) itself is optional. Certain compounds of interest, as described herein, may contain one or more “optionally substituted” moieties. Typically, the term “substituted” means that one or more hydrogens of a specified moiety are substituted by a preferred substituent, e.g., one of the substituents or groups described herein, whether preceded by the term “optionally”. Unless otherwise specified, an “optionally substituted” group may have preferred substituents at each preferred position of the group. Furthermore, if two or more positions in any given structure can be substituted by two or more substituents selected from a particular group, the substituents may be identical or different at each position. For example, in the term “optionally substituted C1-C6 alkyl-C2-C9 heteroaryl”, the alkyl moiety, the heteroaryl moiety, or both may be optionally substituted. The substituent combinations conceived in the present invention are preferably obtained by forming stable or chemically suitable compounds. The term “stable,” as used herein, means a compound that remains substantially unchanged when subjected to conditions anticipating the production, detection, and, in certain embodiments, their recovery, purification, and use for one or more purposes disclosed herein.

[0052] Preferred monovalent substituents on the replaceable carbon atoms of the "optionally substituted" group are, independently, deuterium, halogen, -(CH2)O-4R°, -(CH2)O-4OR°, -O(CH2)O-4R°, -O-(CH2)O-4C(O)OR°, -(CH2)O-4CH(OR°)2, -(CH2)O-4SR°, -(CH2)O-4Ph[may be substituted with R°], -(CH2)O-4O(CH2)O-1Ph[may be substituted with R°], -CH=CHPh[may be substituted with R°], and -(CH2)O-4O(CH2)O-1-pyridyl[may be substituted with R°]. ], 4-8 member saturated or unsaturated heterocycloalkyl (e.g., pyridyl), 3-8 member saturated or unsaturated cycloalkyl (e.g., cyclopropyl, cyclobutyl, or cyclopentyl), -NO2, -CN, -N3, -(CH2)O-4N(R°)2, -(CH2)O-4N(R°)C(O)R°, -N(R°)C(S)R°, -(CH2)O-4N(R °)C(O)NR°2, -N(R°)C(S)NR°2, -(CH2)0-4N(R°)C(O)OR°, -N(R°)N(R°)C(O)R°, -N(R°)N(R°)C(O)N R°2, -N(R°)N(R°)C(O)OR°, -(CH2)0-4C(O)R°, -C(S)R°, -(CH2)0-4C(O)OR°, -(CH2)0-4-C(O)-N(R o )2,-(CH2)O-4-C(O)-N(R o )-S(O)2-R o, -C(NCN)NR°2, -(CH2)0-4C(O)SR°, -(CH2)0-4C(O)OSiR°3, -(CH2)0-4OC(O)R°, -OC(O)(CH2)0-4SR°, -SC(S)SR°, -(CH2)0-4 SC(O)R°, -(CH2)0-4C(O)NR°2, -C(S)NR°2, -C(S)SR°, -(CH2)0-4OC(O)NR°2, -C(O)N(OR°)R°, -C(O)C(O)R°, -C(O)CH2C(O)R° , -C(NOR°)R°, -(CH2)0-4SSR°, -(CH2)0-4S(O)2R°, -(CH2)0-4S(O)2OR°, -(CH2)0-4OS(O)2R°, -S(O)2NR°2, -(CH2)0-4S(O)R °, -N(R°)S(O)2NR°2, -N(R°)S(O)2R°, -N(OR°)R°, -C(NOR°)NR°2, -C(NH)NR°2, -P(O)2R°, -P(O)R°2, -P(O)(OR°)2, -OP(O)R°2 -OP(O)(OR°)2, -OP(O)(OR°)R°, -SiR°3, -(C1-C4 linear or branched alkylene)ON(R°)2, or -(C1-C4 linear or branched alkylene)C(O)ON(R°)2 [wherein each R° may be substituted as defined below, independently from hydrogen, -C1-C6 aliphatic, -CH2Ph, -O(CH2)O-1Ph, -CH2-(5-6 membered heteroaryl ring), or nitrogen, oxygen, or sulfur] It can be a 3-6 member saturated, partially unsaturated, or aryl ring having 0-4 heteroatoms independently selected, or, notwithstanding the above definition, two independently existing R° can, together with the atom(s) between them, form a 3-12 member saturated, partially unsaturated, or aryl mono- or bicyclic ring having 0-4 heteroatoms independently selected from nitrogen, oxygen, or sulfur, which may be substituted as defined below.

[0053] Suitable monovalent substituents on R° (or the ring formed by using two independently existing R° atoms together with the atoms in between) are, independently, halogens, -(CH2)O-2R ● , -(HaroR● ), -(CH2)O-2OH, -(CH2)O-2OR ● ,-(CH2)0-2CH(OR ● )2, -O(HaroR ● ), -CN, -N3, -(CH2)0-2C(O)R ● , -(CH2)0-2C(O)OH, -(CH2)0-2C(O)OR ● ,-(CH2)0-2SR ● , -(CH2)0-2SH, -(CH2)0-2NH2, -(CH2)0-2NHR ● ,-(CH2)0-2NR ● 2, -NO2, -SiR ● 3. -OSiR ● 3. -C(O)SR ● -(C1-4 linear or branched alkylene)C(O)OR ● , or -SSR ● It can be, and in the formula, each R ● The atoms are either unsubstituted, or, where the prefix "halo" is located, substituted by only one or more halogens, and are independently selected from C1-C4 aliphatic, -CH2Ph, -O(CH2)0-1Ph, or 5-6 membered saturated, partially unsaturated, or aryl rings having 0-4 heteroatoms independently selected from nitrogen, oxygen, or sulfur. Preferred divalent substituents on the saturated carbon atom of R° include =O and =S.

[0054] Suitable divalent substituents on the saturated carbon atom of the "arbitrarily substituted" group are: =O, =S, =NNR * 2. =NNHC(O)R * ,=NNHC(O)OR * ,=NNHS(O)2R * ,=NR * 、=NOR * , -O(C(R * 2))2-3O-, or -S(C(R * 2))2-3S- are listed, and in the formula, R exists independently in each case. *is selected from hydrogen, a C1-C6 aliphatic which can be substituted as defined below, or an unsubstituted 5- to 6-membered, saturated, partially unsaturated, or aryl ring having 0 to 4 heteroatoms independently selected from nitrogen, oxygen, or sulfur. Suitable divalent substituents bonded to vicinal substitutable carbons of the "optionally substituted" group include -O(CR * 2)2-3O-, where each independently occurring R * is selected from hydrogen, a C1-C6 aliphatic which can be substituted as defined below, or an unsubstituted 5- to 6-membered, saturated, partially unsaturated, or aryl ring having 0 to 4 heteroatoms independently selected from nitrogen, oxygen, or sulfur.

[0055] R * Suitable substituents on the aliphatic group of are -R ● , (haloR ● ), -OH, -OR ● , -O(haloR ● ), -CN, -C(O)OH, -C(O)OR ● , -NH2, -NHR ● , -NR ● 2, or -NO2, where each R ● is unsubstituted or, where "halo" precedes, is substituted only by one or more halogens and is independently a C1-C4 aliphatic, -CH2Ph, -O(CH2)0-1Ph, or a 5- to 6-membered, saturated, partially unsaturated, or aryl ring having 0 to 4 heteroatoms independently selected from nitrogen, oxygen, or sulfur.

[0056] Suitable substituents on the substitutable nitrogen of the "optionally substituted" group include -R † , -NR † 2, -C(O)R † , -C(O)OR † , -C(O)C(O)R † , -C(O)CH2C(O)R † , -S(O)2R † , -S(O)2NR † , -C(S)NR †2, -C(NH)NR † 2, or -N(R † )S(O)2R † These are listed, and in the formula, each R † R is independently a saturated, partially unsaturated, or aryl ring having 0 to 4 heteroatoms independently selected from hydrogen, C1-C6 aliphatic, unsubstituted -OPh, or nitrogen, oxygen, or sulfur, or two independently existing R rings notwithstanding the above definitions. † These atoms, along with the atoms (or multiple atoms) interposed between them, form unsubstituted 3-12 member saturated, partially unsaturated, or aryl mono- or bicyclic rings containing 0-4 heteroatoms independently selected from nitrogen, oxygen, or sulfur.

[0057] R † Suitable substituents on the aliphatic group are, independently, halogens, -R ● ,-(HaroR ● ), -OH, -OR ● ,-O(HaroR ● ), -CN, -C(O)OH, -C(O)OR ● -NH2, -NHR ● , -NR ● 2, or -NO2, where each R ● R is either unsubstituted, or, where it is preceded by "halo", substituted by only one or more halogens, and is independently a C1-C4 aliphatic, -CH2Ph, -O(CH2)0-1Ph, or a 5-6 membered, saturated, partially unsaturated, or aryl ring having 0-4 heteroatoms independently selected from nitrogen, oxygen, or sulfur. † Suitable divalent substituents on the saturated carbon atom include =O and =S.

[0058] As used herein, the term "acetyl" means the group -C(O)CH3. As used herein, the term "alkoxy" means -O-C1-C 20 Alkyl grouping means that the alkoxy group is bonded to the rest of the compound via an oxygen atom.

[0059] As used herein, the term “alkyl” means a saturated, linear, or branched monovalent hydrocarbon group containing 1 to 20 (e.g., 1 to 10, or 1 to 6) carbon atoms. In some embodiments, the alkyl group is unbranched (i.e., linear), and in some embodiments, the alkyl group is branched. Alkyl groups are exemplified by, but are not limited to, methyl, ethyl, n- and isopropyl, n-, sec-, iso and tert-butyl, and neopentyl.

[0060] As used herein, the term "alkylene" refers to a saturated divalent hydrocarbon group derived from a straight-chain or branched-chain saturated hydrocarbon by removing two hydrogen atoms, exemplified by methylene, ethylene, isopropylene, etc. x -C y "Alkylene" refers to an alkylene group having x to y carbon atoms. Exemplary values ​​for x are 1, 2, 3, 4, 5, and 6, and exemplary values ​​for y are 2, 3, 4, 5, 6, 7, 8, 9, 10, 12, 14, 16, 18, or 20 (e.g., C1-C6, C1-C 10 , C2-C 20 , C2-C6, C2-C 10 , or C2-C 20 Alkylene). In some embodiments, the alkylene may be further substituted with one, two, three, or four substituents as defined herein.

[0061] As used herein, the term “alkenyl” means, unless otherwise specified, a monovalent linear or branched group of 2 to 20 carbon atoms (e.g., 2 to 6 or 2 to 10 carbon atoms) containing one or more carbon-carbon double bonds, exemplified by ethenyl, 1-propenyl, 2-propenyl, 2-methyl-1-propenyl, 1-butenyl, and 2-butenyl. Alkenyls include both cis and trans isomers. As used herein, the term “alkenylene” means, unless otherwise specified, a divalent linear or branched group of 2 to 20 carbon atoms (e.g., 2 to 6 or 2 to 10 carbon atoms) containing one or more carbon-carbon double bonds.

[0062] As used herein, the term "alkynyl" refers to a monovalent linear or branched group consisting of 2 to 20 carbon atoms (e.g., 2 to 4, 2 to 6, or 2 to 10 carbon atoms) containing a carbon-carbon triple bond, as exemplified by ethynyl and 1-propynyl.

[0063] As used herein, the term "amino" means -N(R † )2, for example, represents -NH2 and -N(CH3)2. As used herein, the term "aminoalkyl" refers to an alkyl moiety in which one or more carbon atoms are substituted with one or more amino moieties.

[0064] As used herein, the term “amino acid” means a molecule having a side chain, an amino group, and an acidic group (e.g., -CO2H or -SO3H), and an amino acid is bonded to a parent molecule by a side chain, an amino group, or an acidic group (e.g., a side chain). As used herein, the term “amino acid” in its broadest sense means any compound or substance that can be incorporated into a polypeptide chain, for example, by the formation of one or more peptide bonds. In some embodiments, an amino acid has the general structure H2N-C(H)(R)-COOH. In some embodiments, an amino acid is a naturally occurring amino acid. In some embodiments, an amino acid is a synthetic amino acid, in some embodiments, an amino acid is a D-amino acid, and in some embodiments, an amino acid is an L-amino acid. “Standard amino acid” means any of the 20 standard L-amino acids commonly found in naturally occurring peptides. Examples of amino acids include alanine, arginine, asparagine, aspartic acid, cysteine, glutamic acid, glutamine, glycine, histidine, optionally substituted hydroxylnorvaline, isoleucine, leucine, lysine, methionine, norvaline, ornithine, phenylalanine, proline, pyrrolicine, selenocysteine, serine, taurine, threonine, tryptophan, tyrosine, and valine.

[0065] As used herein, the term "aryl" refers to a monovalent monocyclic, bicyclic, or polycyclic ring system formed by carbon atoms, wherein the ring bonded to the pendant group is aromatic. Examples of aryl groups include phenyl, naphthyl, phenantrenyl, and anthracenyl. The aryl ring may be bonded to its pendant group by any heteroatom or carbocyclic atom that results in a stable structure, and unless otherwise specified, any of the ring atoms may be substituted.

[0066] As used herein, the term "C0" represents a bond. For example, part of the term -N(C(O)-(C0-C5alkylene-H)- includes -N(C(O)-(C0alkylene-H)-, which is also represented by -N(C(O)-H)-.

[0067] As used herein, the terms “carbocyclic” and “carbocyclyl” refer to a monovalent, optionally substituted, 3- to 12-membered monocyclic, bicyclic, or tricyclic structure, which may be optionally bridged, condensed, or spirocyclic, where all rings are formed of carbon atoms and at least one ring is non-aromatic. Examples of carbocyclic structures include cycloalkyl, cycloalkenyl, and cycloalkynyl groups. Examples of carbocyclyl groups include cyclohexyl, cyclohexenyl, cyclooctinyl, 1,2-dihydronaphthyl, 1,2,3,4-tetrahydronaphthyl, fluorenyl, indenyl, indanyl, and dekalinyl. The carbocyclic ring can be bonded to its pendant group at any ring atom that results in a stable structure, and any of the ring atoms may be optionally substituted unless otherwise specified.

[0068] As used herein, the term "carbonyl" refers to a C(O) group, which can also be represented as C=O. As used herein, the term "carboxyl" means -CO2H, (C=O)(OH), COOH, or C(O)OH, or the corresponding aprotonated group.

[0069] As used herein, the term "cyano" refers to the -CN group. As used herein, the term "cycloalkyl" means a monovalent saturated cyclic hydrocarbon group, which may be crosslinked, condensed, or a spirocyclic group having 3 to 8 carbon atoms, unless otherwise specified, and is exemplified by cyclopropyl, cyclobutyl, cyclopentyl, cyclohexyl, cycloheptyl, and cycloheptyl.

[0070] As used herein, the term "cycloalkenyl" means a monovalent non-aromatic saturated cyclic hydrocarbon group, which, unless otherwise specified, may be crosslinked, condensed, or a spirocyclic group having 3 to 8 carbon atoms and containing one or more carbon-carbon double bonds.

[0071] As used herein, the term “diastereomer” means a stereoisomer that is not a mirror image of another and cannot be superimposed on another. As used herein, “enantiomer” means each individual optically active form of the compound of the present invention having at least 80% (i.e., at least 90% of one enantiomer and up to 10% of the other enantiomer), preferably at least 90%, and more preferably at least 98% optical purity or enantiomer excess (measured by methods standard in the art).

[0072] As used herein, the term "haloacetyl" means an acetyl group in which at least one hydrogen atom is substituted with a halogen. As used herein, the term "haloalkyl" refers to an alkyl moiety in which one or more carbon atoms are substituted with one or more identical or different halogen moieties.

[0073] As used herein, the term "halogen" refers to a halogen selected from bromine, chlorine, iodine, or fluorine. As used herein, the term “heteroalkyl” refers to an “alkyl” group (as defined herein) in which at least one carbon atom is replaced by a heteroatom (e.g., an O, N, or S atom). The heteroatom may appear in the middle or at the ends of the radical.

[0074] As used herein, the term “heteroaryl” refers to a monovalent monocyclic or polycyclic cyclic structure containing at least one complete aromatic ring. That is, these contain 4n+2 π electrons within the monocyclic or polycyclic ring system and contain at least one ring heteroatom selected from N, O, or S within the aromatic ring. Exemplary unsubstituted heteroaryl groups have 1 to 12 (e.g., 1 to 11, 1 to 10, 1 to 9, 2 to 12, 2 to 11, 2 to 10, or 2 to 9) carbon atoms. The term “heteroaryl” also includes bicyclic, tricyclic, and tetracyclic groups in which any of the above heteroaromatic rings are fused to one or more aryl or carbocyclic rings, such as a phenyl ring or a cyclohexane ring. Examples of heteroaryl groups include, but are not limited to, pyridyl, pyrazolyl, benzoxazolyl, benzimidazolyl, benzothiazolyl, imidazolyl, thiazolyl, quinolinyl, tetrahydroquinolinyl, and 4-azaindylol. The heteroaryl ring can be bonded to its pendant group at any ring atom that results in a stable structure, and any of the ring atoms can be optionally substituted unless otherwise specified. In some embodiments, the heteroaryl is substituted with 1, 2, 3, or 4 substituents.

[0075] As used herein, the term "heterocycloalkyl" refers to a monovalent monocyclic, bicyclic, or polycyclic ring system, which may be bridged, condensed, or spirocyclic, in which at least one ring is non-aromatic, and the non-aromatic ring contains 1, 2, 3, or 4 heteroatoms independently selected from the group consisting of nitrogen, oxygen, and sulfur. Five-membered rings have 0 to 2 double bonds, and six- and seven-membered rings have 0 to 3 double bonds. Exemplary unsubstituted heterocycloalkyl groups have 1 to 12 (e.g., 1 to 11, 1 to 10, 1 to 9, 2 to 12, 2 to 11, 2 to 10, or 2 to 9) carbon atoms. The term "heterocycloalkyl" also refers to heterocyclic compounds having a bridged polycyclic structure in which one or more carbons or heteroatoms bridge two non-adjacent elements of a monocyclic ring, e.g., a quinuclidinyl group. The term "heterocycloalkyl" includes bicyclic, tricyclic, and tetracyclic groups in which any of the above heterocyclic rings is fused to one or more aromatic, carbocyclic, heteroaromatic, or heterocyclic rings, such as an aryl ring, cyclohexane ring, cyclohexene ring, cyclopentane ring, cyclopentene ring, pyridine ring, or pyrrolidine ring. Examples of heterocycloalkyl groups are pyrrolidinyl, piperidinyl, 1,2,3,4-tetrahydroquinolinyl, decahydroquinolinyl, dihydropyrrolopyridine, and decahydronaphthilidinyl. Heterocycloalkyl rings can be bonded to their pendant group at any ring atom that results in a stable structure, and any of the ring atoms can be substituted unless otherwise specified.

[0076] As used herein, the term "hydroxy" refers to the -OH group. As used herein, the term "hydroxyalkyl" refers to an alkyl moiety in which one or more carbon atoms are substituted with one or more -OH moieties.

[0077] As used herein, “isomer” means any tautomer, stereoisomer, atropisomer, enantiomer, or diastereomer of any compound of the present invention. Compounds of the present invention may have one or more chiral centers or double bonds and are therefore recognized to exist as stereoisomers, e.g., double bond isomers (i.e., geometric E / Z isomers), or diastereomers (e.g., enantiomers (i.e., (+) or (-), or cis / trans isomers)). In accordance with the present invention, the chemical structures described herein, and therefore the compounds of the present invention, have all corresponding stereoisomers, i.e., stereoisomerically pure forms (e.g., geometrically pure, enantiomerically pure, or diastereally pure), as well as mirror images. This includes both enantiomers and stereoisomers (e.g., racemic compounds). Enantiomers and stereoisomers of the compounds of the present invention can usually be broken down into their constituent enantiomers or stereoisomers by well-known methods, such as chiral phase gas chromatography, chiral phase high-performance liquid chromatography, crystallization of the compound as a chiral salt complex, or crystallization of the compound in a chiral solvent. Enantiomers and stereoisomers can also be obtained from stereoisomerically or enantiomerically pure intermediates, reagents, and catalysts by well-known asymmetric synthesis methods.

[0078] As used herein, the "monovalent organic moiety" is less than 500 kDa. In some embodiments, the "monovalent organic moiety" is less than 400 kDa. In some embodiments, the "monovalent organic moiety" is less than 300 kDa. In some embodiments, the "monovalent organic moiety" is less than 200 kDa. In some embodiments, the "monovalent organic moiety" is less than 100 kDa. In some embodiments, the "monovalent organic moiety" is less than 50 kDa. In some embodiments, the "monovalent organic moiety" is less than 25 kDa. In some embodiments, the "monovalent organic moiety" is less than 20 kDa. In some embodiments, the "monovalent organic moiety" is less than 15 kDa. In some embodiments, the "monovalent organic moiety" is less than 10 kDa. In some embodiments, the "monovalent organic moiety" is less than 1 kDa. In some embodiments, the "monovalent organic moiety" is less than 500 g / mol. In some embodiments, the "monovalent organic moiety" is in the range of 500 g / mol to 500 kDa.

[0079] As used herein, the term “stereoisomer” means all possible different isomeric and structural forms that a compound may have (for example, any compound of any formula described herein), in particular all possible stereochemical and structural isomeric forms of the basic molecular structure, including atropisomers, all diastereomers, enantiomers, or conformational isomers. Some of the compounds of the present invention may exist in different tautomers, all of which are included in the scope of the present invention.

[0080] As used herein, the term "sulfonyl" refers to the -S(O)2- group. As used herein, the term "thiocarbonyl" means a -C(S)- group. Those skilled in the art will understand that certain compounds described herein can be provided or utilized in any of various forms, such as salt form, protected form, prodrug form, ester form, isomer form (e.g., optical or structural isomer), isotopic form, etc. In some embodiments, reference to a particular compound may relate to a particular form of that compound. In some embodiments, reference to a particular compound may relate to the compound in any form. In some embodiments, for example, a preparation of a single stereoisomer of a compound may be considered a different form of the compound from a racemic mixture of the compound; a particular salt of a compound may be considered a different form from another salt form of the compound; a preparation containing a structural isomer of a double bond ((Z) or (E)) may be considered a different form from one containing other structural isomers of the double bond ((E) or (Z)); and a preparation in which one or more atoms are isotopes different from those present in the reference preparation may be considered a different form. [Brief explanation of the drawing]

[0081] [Figure 1] The present invention has revealed that Compound A and Compound B, representative covalent KRAS G12D inhibitors, exhibited potent and sustained RAS pathway modulation in a human pancreatic adenocarcinoma HPAC KRASG12D / wt mouse xenograft model. The RAS / ERK signaling pathway modulation was evaluated by measuring human DUSP6 mRNA levels in a qPCR assay. Both Compound A and Compound B suppressed DUSP6 mRNA levels in HPAC xenograft tumors within 4 hours of administration, indicating potent RAS pathway modulation. [Figure 2]Compounds A and B exhibited strong tumor crosslinking activity up to 4 hours and up to 24 hours, which was consistent with significant DUSP6 suppression. Tumor samples collected from the assay in Figure 1 were homogenized for protein extraction. The protein lysates were subjected to Western blotting using Ras Rabbit mAb (Abcam ab108602) and β-actin mAb (CST-4967). Tumor samples in which the compounds were covalently bound to the KRAS G12D protein showed the appearance of bands with larger molecular weights (MW) (crosslinked KRAS G12D bands). [Figure 3A] In HPAC CDX mouse xenograft models containing heterozygous KRASG12D, administration of both compound A and compound B as monotherapy at 100 mg / kg po per day resulted in total tumor regression (defined as >10% tumor regression from baseline) at the end of treatment (day 28 after the start of treatment). [Figure 3B] The study showed that neither compound A (100 mg / kg po qd) nor compound B (100 mg / kg po qd) resulted in weight loss, indicating that both compounds at 100 mg / kg are well-tolerated in the human pancreatic adenocarcinoma HPAC KRASG12D / wt mouse xenograft model. [Figure 3C] In a human pancreatic adenocarcinoma HPAC KRASG12D / wt mouse xenograft model, it was revealed that in the compound A (100 mg / kg po qd) group and the compound B (100 mg / kg po qd) group, 8 out of 10 tumors and 9 out of 10 tumors, respectively, achieved complete regression (complete regression defined as >85% tumor regression from baseline) on day 28. [Modes for carrying out the invention]

[0082] compound Ras inhibitors are provided herein. The approach described herein requires the formation of a high-affinity triplicate or conjugate between a synthetic ligand and two intracellular proteins that do not interact under normal physiological conditions: the target protein (e.g., Ras) and a cytosolic chaperone (presenter protein) that is widely expressed intracellularly (e.g., cyclophyllin A). More specifically, in some embodiments, the Ras inhibitors described herein include a novel binding pocket cytosol in Ras, which drives the formation of a high-affinity triplicate or conjugate between the Ras protein and the widely expressed cytosolic chaperone cyclophyllin A (CYPA). While not theoretically bound, the inventors believe that one way in which the inhibitory effect on Ras is affected by the compounds and complexes or conjugates of the present invention is the formation of steric occlusion of the interaction site between Ras and downstream effector molecules such as RAFs, which is necessary for the growth of oncogenic signals.

[0083] While not bound by theory, the inventors hypothesize that both covalent and non-covalent interactions of the compounds of the present invention with Ras and chaperone proteins (e.g., cyclophyllin A) can contribute to the inhibition of Ras activity. In some embodiments, the compounds of the present invention form covalent adducts with side chains of the Ras protein (e.g., the -CH2-COOH or -CH2-COO- side chain of aspartic acid at position 12 or 13 of a mutant Ras protein). Covalent adducts may also be formed with other side chains of Ras. Furthermore, or alternatively, non-covalent interactions may be at play: for example, van der Waals interactions, hydrophobic interactions, hydrophilic interactions, and hydrogen bonding interactions, and combinations thereof, may contribute to the ability of the compounds of the present invention to form complexes and act as RAS inhibitors. Therefore, various Ras proteins can be inhibited by the compounds of the present invention (for example, K-Ras, N-Ras, H-Ras at positions 12, 13, and 61, and their variants, e.g., G12C, G12D, G12V, G12S, G13C, G13D, and Q61L, as well as others described herein).

[0084] Methods for measuring covalent adduct formation are known in the art. Another method for measuring covalent adduct formation is to perform a "crosslinking" assay, as described below: Note - The following protocol describes a procedure for monitoring the crosslinking of K-Ras G12D(GMP-PNP) to the compound of the present invention. This protocol can also be performed by substituting other Ras proteins or nucleotides.

[0085] The purpose of this biochemical assay is to measure the ability of the test compound to covalently label the nucleotide-loaded K-Ras isoform. The stock solution of 75 mM NaCl, 1 mM MgCl2, 5 μM cyclophyllin A, 2 μM test compound, and 5 μM GMP-PNP-loaded K-Ras(1-169)G12D was diluted 10-fold in assay buffer containing 12.5 mM HEPES (pH 7.4) to obtain a final concentration of 0.5 μM. The final sample volume was 100 μL.

[0086] After incubating the sample at 25°C for up to 24 hours (multiple hours are possible), quench the sample by adding 10 μL of 5% formic acid. The quenched sample is centrifuged in a benchtop centrifuge at 15,000 rpm for 15 minutes, and a 10 μL aliquot is injected into a reverse-phase C4 column. The sample is then eluted into a mass spectrometer while increasing the acetonitrile gradient in the mobile phase. Raw data can be analyzed using Waters MassLynx MS software, and binding ratios are calculated from the reverse-convolution protein peaks for labeled and unlabeled K-Ras.

[0087] Therefore, compounds having the structure of formula 0, or pharmaceutically acceptable salts thereof, are provided herein:

[0088] [ka]

[0089] [In the formula, A is an optionally substituted 3-6 member heterocycloalkylene, an optionally substituted 3-6 member cycloalkylene, an optionally substituted 6 member arylene, or an optionally substituted 5-10 member heteroarylene.] X 1 , X 2 , and X 3 Each of these is independently selected from CH2, CF2, C=O, or O. m is either 1 or 2. n is either 0 or 1. R 1is hydrogen, optionally substituted C1-C6 heteroalkyl, or optionally substituted 3-10 member heterocycloalkyl. R 2 is an arbitrarily substituted C1-C6 alkyl, and R 3 This is an optionally substituted C1-C6 alkyl group, an optionally substituted 3-6 member cycloalkyl group, or an optionally substituted heterocycloalkyl group. Each hydrogen atom is independently an arbitrarily isotopically enriched deuterium.

[0090] In some embodiments, the compounds of the present invention have the structure of formula I, or a pharmaceutically acceptable salt thereof:

[0091] [ka]

[0092] [In the formula, A is an optionally substituted 3-6 member heterocycloalkylene, an optionally substituted 3-6 member cycloalkylene, an optionally substituted 6 member arylene, or an optionally substituted 5-10 member heteroarylene.] X 1 , X 2 , and X 3 Each of these is independently selected from CH2, CF2, C=O, or O. m is either 1 or 2. n is either 0 or 1. R 1 is hydrogen, optionally substituted C1-C6 heteroalkyl, or optionally substituted 3-10 member heterocycloalkyl. R 2 is an arbitrarily substituted C1-C6 alkyl, and R 3 This is an optionally substituted C1-C6 alkyl group, an optionally substituted 3-6 member cycloalkyl group, or an optionally substituted heterocycloalkyl group. Each hydrogen atom is independently an arbitrarily isotopically enriched deuterium.

[0093] In some embodiments, the compounds of the present invention have the structures of formulas Ia, Ib, and Ic, or pharmaceutically acceptable salts thereof:

[0094] [ka]

[0095] [In the formula, each D represents hydrogen having a deuterium isotope enrichment factor of at least 5.] In some embodiments, the compounds of the present invention have the structure of formula II, or a pharmaceutically acceptable salt thereof:

[0096] [ka]

[0097] [In the formula, A is an optionally substituted 3-6 member heterocycloalkylene, an optionally substituted 3-6 member cycloalkylene, an optionally substituted 6 member arylene, or an optionally substituted 5-10 member heteroarylene.] R 2 is an arbitrarily substituted C1-C6 alkyl, and R 3 This is an optionally substituted C1-C6 alkyl group, an optionally substituted 3-6 member cycloalkyl group, or an optionally substituted heterocycloalkyl group. Each hydrogen atom is independently an arbitrarily isotopically enriched deuterium.

[0098] In some embodiments, the compounds of the present invention have the structure of formula V, or a pharmaceutically acceptable salt thereof:

[0099] [ka]

[0100] [In the formula, A is an optionally substituted 3-6 member heterocycloalkylene, an optionally substituted 3-6 member cycloalkylene, an optionally substituted 6 member arylene, or an optionally substituted 5-10 member heteroarylene.] R 2 is an arbitrarily substituted C1-C6 alkyl, and R 3 This is an optionally substituted C1-C6 alkyl group, an optionally substituted 3-6 member cycloalkyl group, or an optionally substituted heterocycloalkyl group. Each hydrogen atom is independently an arbitrarily isotopically enriched deuterium.

[0101] In some embodiments, the compounds of the present invention have the structure of formula VI, or a pharmaceutically acceptable salt thereof:

[0102] [ka]

[0103] [In the formula, A is an optionally substituted 3-6 member heterocycloalkylene, an optionally substituted 3-6 member cycloalkylene, an optionally substituted 6 member arylene, or an optionally substituted 5-10 member heteroarylene.] R 2 is an arbitrarily substituted C1-C6 alkyl, and R 3 This is an optionally substituted C1-C6 alkyl group, an optionally substituted 3-6 member cycloalkyl group, or an optionally substituted heterocycloalkyl group. Each hydrogen atom is independently an arbitrarily isotopically enriched deuterium.

[0104] In some embodiments, the compounds of the present invention have the structure of formula VII, or a pharmaceutically acceptable salt thereof:

[0105] [ka]

[0106] [In the formula, A is an optionally substituted 3-6 member heterocycloalkylene, an optionally substituted 3-6 member cycloalkylene, an optionally substituted 6 member arylene, or an optionally substituted 5-10 member heteroarylene.] R 2 is an arbitrarily substituted C1-C6 alkyl, and R 3 This is an optionally substituted C1-C6 alkyl group, an optionally substituted 3-6 member cycloalkyl group, or an optionally substituted heterocycloalkyl group. Each hydrogen atom is independently an arbitrarily isotopically enriched deuterium.

[0107] In some embodiments, the compounds of the present invention have the structures of formulas Va, Vb, and Vc, or pharmaceutically acceptable salts thereof:

[0108] [ka]

[0109] [In the formula, each D represents hydrogen having a deuterium isotope enrichment factor of at least 5.] In some embodiments, the compounds of the present invention have the structures of formulas Vd, Ve, Vf, or pharmaceutically acceptable salts thereof:

[0110] [ka]

[0111] [In the formula, each D represents hydrogen having a deuterium isotope enrichment factor of at least 5.] In some embodiments, A is an optionally substituted thiazole-diyl, optionally substituted oxazole-diyl, optionally substituted morpholine-diyl, optionally substituted pyrrolidine-diyl, optionally substituted piperidine-diyl, or optionally substituted phenylene. In some embodiments, A is an optionally substituted thiazole-diyl or optionally substituted morpholine-diyl. In some embodiments of the compounds of the present invention, A is an optionally substituted 5- to 10-membered heteroarylene. In some embodiments, A is

[0112] [ka]

[0113] In some embodiments, A is

[0114] [ka]

[0115] That is the case. In some embodiments of the compounds of the present invention, A is optionally substituted phenylene. In some embodiments, A is

[0116] [ka]

[0117] In some embodiments, A is

[0118] [ka]

[0119] That is the case. In some embodiments of the compounds of the present invention, A is an optionally substituted 3- to 6-membered heterocycloalkylene. In some embodiments, A is an optionally substituted 6-membered heterocycloalkylene. In some embodiments, A is selected from the following, or their stereoisomers:

[0120] [ka]

[0121] In some embodiments, A is selected from the following, or their stereoisomers:

[0122] [ka]

[0123] In some embodiments of the compounds of the present invention, R 1 R is a hydrogen atom or an optionally substituted 3- to 10-membered heterocycloalkyl group. In some embodiments of the compounds of the present invention, R 1 R is an optionally substituted 3- to 10-membered heterocycloalkyl group. In some embodiments of the compounds of the present invention, R 1 teeth,

[0124] [ka]

[0125] That is the case. In some embodiments of the compounds of the present invention, R 1 teeth,

[0126] [ka]

[0127] In the formula, each D represents hydrogen having a deuterium isotope enrichment factor of at least 5. In some embodiments of the compounds of the present invention, R 2 teeth,

[0128] [ka]

[0129] That is the case. In some embodiments of the compounds of the present invention, R 2 teeth,

[0130] [ka]

[0131] In the formula, each D represents hydrogen having a deuterium isotope enrichment factor of at least 5. In some embodiments of the compounds of the present invention, R 3 R is an optionally substituted C1-C6 alkyl or an optionally substituted 3-6 member cycloalkyl. In some embodiments of the compounds of the present invention, R 3 is an optionally substituted C1-C6 alkyl group. In some embodiments, R 3 teeth,

[0132] [ka]

[0133] In some embodiments, R 3 teeth,

[0134] [ka]

[0135] In some embodiments, R 3 teeth,

[0136] [ka]

[0137] In the formula, each D represents hydrogen having a deuterium isotope enrichment factor of at least 5. In some embodiments of the compounds of the present invention, R 3 R is an optionally substituted 3- to 6-membered cycloalkyl group. In some embodiments, R 3 teeth,

[0138] [ka]

[0139] In some embodiments, R 3 teeth,

[0140] [ka]

[0141] That is the case. In some embodiments of the compounds of the present invention, R 2 teeth,

[0142] [ka]

[0143] And R 3 teeth,

[0144] [ka]

[0145] And A is,

[0146] [ka]

[0147] That is the case. In some embodiments, R 2 teeth,

[0148] [ka]

[0149] And R 3 teeth,

[0150] [ka]

[0151] And A is,

[0152] [ka]

[0153] That is the case. In some embodiments of the compounds of the present invention, m is 1. In some embodiments, n is 1. In some embodiments, X 1 This is CH2. In some embodiments, X 2 This is CH2. In some embodiments, X 3 is CH2. In some embodiments, m is 1, n is 1, and X 1 , X 2 , and X 3 Each of these is CH2.

[0154] In some embodiments, the compounds of the present invention are selected from Table 1, or are pharmaceutically acceptable salts or stereoisomers thereof. In some embodiments, the compounds of the present invention are selected from Table 1, or are pharmaceutically acceptable salts or atropisomers thereof.

[0155] [Table 1-1]

[0156] [Table 1-2]

[0157] Table 1-3

[0158] Table 1-4

[0159] Table 1-5

[0160] Table 1-6

[0161] Table 1-7

[0162] Table 1-8

[0163] Table 1-9

[0164] Table 1-10

[0165] Table 1-11

[0166] Table 1-12

[0167] [Table 1-13]

[0168] [Table 1-14]

[0169] [Table 1-15]

[0170] [Table 1-16]

[0171] [Table 1-17]

[0172] In some embodiments, compounds from Table 2 or pharmaceutically acceptable salts thereof are provided. In some embodiments, the compounds of the present invention are selected from Table 2, or are pharmaceutically acceptable salts or atrop isomers thereof.

[0173] [Table 2-1]

[0174] [Table 2-2]

[0175] [Table 2-3]

[0176] [Table 2-4]

[0177] [Table 2-5]

[0178] [Table 2-6]

[0179] [Table 2-7]

[0180] [Table 2-8]

[0181] In some embodiments, the compound of the present invention is a compound selected from Table 3, or a pharmaceutically acceptable salt or stereoisomer thereof. In some embodiments, the compound of the present invention is a compound selected from Table 3, or a pharmaceutically acceptable salt or atropisomer thereof.

[0182] In some embodiments, the compounds of the present invention are not compounds selected from Table 3. In some embodiments, the compounds of the present invention are not compounds selected from Table 3, or are not pharmaceutically acceptable salts or stereoisomers thereof. In some embodiments, the compounds of the present invention are not compounds selected from Table 3, or are not pharmaceutically acceptable salts or atropisomers thereof.

[0183] [Table 3-1]

[0184] [Table 3-2]

[0185] [Table 3-3]

[0186] In some embodiments, the compound of the present invention comprises a crosslinking group (e.g., an optionally substituted aziridine moiety) bound to an organic moiety that is a Ras binding moiety, and when the compound comes into contact with a Ras protein, the compound binds to the Ras protein to form a conjugate. For example, the crosslinking group (e.g., an optionally substituted aziridine moiety) of the compound binds to, for example, an amino acid of the Ras protein, crosslinking to form a conjugate. In some embodiments, the Ras binding moiety is a K-Ras binding moiety. In some embodiments, the K-Ras binding moiety binds to a residue in the K-Ras Switch-II binding pocket of the K-Ras protein. In some embodiments, the Ras binding moiety is an H-Ras binding moiety that binds to a residue in the H-Ras Switch-II binding pocket of the H-Ras protein. In some embodiments, the Ras binding moiety is an N-Ras binding moiety that binds to a residue in the N-Ras Switch-II binding pocket of the N-Ras protein. The Ras binding moiety typically has a molecular weight of less than 1200 Da. For example, see Johnson et al., 292:12981-12993 (2017), which is incorporated herein by reference to the Ras protein domain.

[0187] In some embodiments, the compounds of the present invention are, or act as, prodrugs for administration to, for example, cells or other targets requiring administration. In another embodiment, the present invention provides a pharmaceutical composition comprising a compound of the present invention or a pharmaceutically acceptable salt thereof and a pharmaceutically acceptable excipient.

[0188] Further providing are conjugates containing the structure of formula III, or salts thereof: MP 1 Formula III [In the formula, P 1 is a monovalent organic part, and M has the structure of equation IV:

[0189] [ka]

[0190] [In the formula, A is an optionally substituted 3-6 member heterocycloalkylene, an optionally substituted 3-6 member cycloalkylene, an optionally substituted 6 member arylene, or an optionally substituted 5-10 member heteroarylene.] X 1 , X 2 , and X 3 Each of these is independently selected from CH2, CF2, C=O, or O. m is either 1 or 2. n is either 0 or 1. R 1 is hydrogen, optionally substituted C1-C6 heteroalkyl, or optionally substituted 3-10 member heterocycloalkyl. R 2 is an arbitrarily substituted C1-C6 alkyl, and R 3 This is an optionally substituted C1-C6 alkyl group, an optionally substituted 3-6 member cycloalkyl group, or an optionally substituted heterocycloalkyl group. Each hydrogen in Equation IV is independently an arbitrarily isotopically enriched deuterium.

[0191] In some embodiments of the conjugate of the present invention, the monovalent organic moiety is a protein. In some embodiments, the protein is a Ras protein. In some embodiments, the Ras protein is K-Ras G12D or K-Ras G13D. In some embodiments of the conjugate of the present invention, M is bound to an amino acid residue of the monovalent organic moiety.

[0192] In some embodiments, the compounds of the present invention exhibit improved oral bioavailability (%F) compared to those known in the art. Methods for measuring oral bioavailability are known in the art, and one such method is provided below: Oral bioavailability can be measured in BALB / c mice. After intravenous (IV) bolus administration and oral enteral (PO) administration of the test compound, approximately 30 μL of whole blood sample is collected at predetermined time points in a tube containing K2EDTA. For some compounds, the blood sample is centrifuged at 4600 rpm at 4°C for approximately 5 minutes, and the plasma sample is stored at -80°C before bioanalysis. Either the blood or plasma sample is extracted by protein precipitation and analyzed by tandem mass spectrometry (UPLC MS / MS) using an AB Sciex Triple Quad 6500+ mass spectrometer in conjunction with an Acquity UPLC system, for example, using electrospray positive ionization.

[0193] All PK parameters can be derived from time-series blood (or plasma) concentration data obtained by non-compartmental analysis using WinNonlin. Bioavailability (both F% and %F) is estimated using the following formula:

[0194]

number

[0195] AUC inf,PO This is the area under blood (or plasma) concentration over time from zero time after PO administration to infinity. AUC inf,IV This is the area under blood (or plasma) concentration over time from zero time after IV administration to infinity.

[0196] Dose IV This is the total dose administered intravenously. Dose PO This is the total dose administered by PO. Generally, an F% (or %F) value greater than 10% is preferred.

[0197] Of the 19 compounds listed in Table 1 described herein that were tested for oral bioavailability, all but three had a %F greater than 10%. Furthermore, all but three cross-linked the K-Ras G12D residue by more than 60% over 6 hours in the biochemical cross-linking assay described herein. In 13 of the tested compounds, the %F greater than 10% and cross-linked the K-Ras G12D residue by more than 60%. While not bound by theory, the inventors argue that the N-methylaziridine moiety of the compounds described herein is the main cause of this unexpected activity.

[0198] In some embodiments, the compounds of the present invention are more selective to one or more specific Ras variants than other Ras variants or the wild type compared to those known in the art. Methods for measuring such selectivity are known in the art, such as Ras-Raf binding assays, and such protocols are provided herein: The purpose of this biochemical assay is to measure the ability of the test compound to facilitate the formation of a ternary complex between the nucleotide load Ras isoform and cyclophyllin A, and the resulting ternary complex is BRAF. RBD It disrupts binding to constructs and inhibits Ras signaling via RAF effectors.

[0199] In an assay buffer containing 25 mM HEPES (pH 7.3), 0.002% Tween20, 0.1% BSA, 100 mM NaCl and 5 mM MgCl2, untagged cyclophyllin A, His6-K-Ras-GMPPNP (or other Ras variants), and GST-BRAF were added. RBDThese compounds were combined in a 384-well assay plate at final concentrations of 25 μM, 12.5 nM, and 50 nM, respectively. The compounds were present in the plate wells as a 10-point 3-fold dilution series, starting at a final concentration of 30 μM. After incubation at 25°C for 3 hours, the mixture of anti-His Eu-W1024 and anti-GST allophycocyanin was added to the assay sample wells at final concentrations of 10 nM and 50 nM, respectively, and the reaction was incubated for a further 1.5 hours. The TR-FRET signal was read using a microplate reader (excitation 320 nm, fluorescence 665 / 615 nm). Compounds that promote the disruption of the Ras:RAF complex were identified as those that induced a decrease in the TR-FRET ratio compared to the DMSO control well. Therefore, in some embodiments, the compounds of the present invention are KRAS G12D The compounds of the present invention exhibit greater selectivity for other Ras variants or the wild type than for other Ras variants or the wild type.

[0200] In some embodiments, the compounds of the present invention are potent against one or more specific Ras variants compared to other Ras variants or wild-types compared to those known in the art. Methods for measuring such potency are known in the art, such as the pERK assay, and the protocol is provided in the following examples: Thus, in some embodiments, the compounds of the present invention are potent against KRAS G12D The compounds of the present invention exhibit higher efficacy with respect to those known in the art. The compounds of the present invention may also exhibit higher efficacy with respect to other RAS variants disclosed herein, or combinations thereof.

[0201] In some embodiments, the compounds of the present invention exhibit greater adverse effects on cell viability with respect to one or more specific Ras variants compared to other Ras variants or wild-type variants, compared to those known in the art. Methods for measuring cell viability are known in the art, such as the CellTiter-Glo® Cell Viability Assay, and the assay is described herein: Note: The following protocol describes the procedure for monitoring the cell viability of K-Ras mutant cancer cell lines corresponding to the compounds of the present invention. Other RAS isoforms can be used, but the number of cells seeded will vary depending on the cell line used.

[0202] The purpose of this cell assay is to measure the effect of the test compound on the proliferation of human Ras cancer cell lines (e.g., NCI-H358 (K-Ras G12C), AsPC-1 (K-Ras G12D), and Capan-1 (K-Ras G12V)) over a 5-day treatment period by quantifying the amount of ATP present at the endpoint using CellTiter-Glo® 2.0 reagent (Promega).

[0203] Cells are seeded at a rate of 250 cells / well in 40 μL of growth medium in a 384-well assay plate and incubated overnight in a humid atmosphere at 37°C with 5% CO2. On assay day, a 10 mM stock solution of the test compound is first diluted with 100% DMSO to make a 3 mM solution. A well-mixed solution of the compound (15 μL) is transferred to the next well containing 30 μL of 100% DMSO, and this process is repeated until nine 3-fold serial dilutions of the compound are prepared (starting assay concentration of 10 μM). The test compound (132.5 nL) is directly dispensed into the assay plate containing the cells. The plate is shaken at 300 rpm for 15 seconds, centrifuged, and incubated in a humid atmosphere at 37°C with 5% CO2 for 5 days. On day 5, the assay plate and its contents are allowed to equilibrate to room temperature for approximately 30 minutes. Add CellTiter-Glo® 2.0 reagent (25 μL), mix the plate contents with an orbital shaker for 2 minutes, and then incubate at room temperature for 10 minutes. Measure luminescence using PerkinElmer Enspire. Normalize the data by: (Sample signal / Mean DMSO) * 100. Fit the data using 4-parameter logistic fit. Therefore, in some embodiments, the compounds of the present invention contain KRAS G12D With respect to this, a greater reduction in cell viability is shown compared to those known in the art. The compounds of the present invention may also be shown to have a greater reduction in cell viability compared to other RAS variants or combinations thereof disclosed herein.

[0204] In some embodiments, the compounds of the present invention may exhibit greater metabolic stability, permeability, or solubility, or a combination thereof, compared to those known in the art. The compounds of the present invention may exhibit improved acid stability, such as in simulated gastric juice stability assays. Methods for measuring such properties are known in the art. The compounds of the present invention may exhibit better Ras crosslinking than compounds known in the art. Methods for measuring Ras crosslinking are provided herein. In some embodiments, the compounds of the present invention may exhibit improvements in any or a combination thereof, compared to those known in the art: selectivity, potency, cell viability, metabolic stability, acid stability, crosslinking, permeability, or solubility.

[0205] A method for treating cancer in a subject requiring cancer treatment is provided, the method comprising administering to the subject a therapeutically effective amount of the compound of the present invention or a pharmaceutically acceptable salt thereof. The cancer may be, for example, pancreatic cancer, colorectal cancer, non-small cell lung cancer, acute myeloid leukemia, multiple myeloma, thyroid cancer, myelodysplastic syndrome, or small squamous cell carcinoma or follicular carcinoma. In some embodiments, the cancer includes a Ras mutation such as K-Ras G12D or K-Ras G13D. Other Ras mutations are described herein.

[0206] A method for treating a Ras protein-related disorder in a subject requiring treatment for the disorder, further comprising administering to the subject a therapeutically effective amount of the compound of the present invention or a pharmaceutically acceptable salt thereof.

[0207] A method for inhibiting the Ras protein in cells is provided, further comprising contacting the cells with an effective amount of the compound of the present invention or a pharmaceutically acceptable salt thereof. For example, the Ras protein may be K-Ras G12D or K-Ras The Ras protein is G13D. Other Ras proteins are described herein. The cells may be cancer cells such as pancreatic cancer cells, colorectal cancer cells, non-small cell lung cancer cells, acute myeloid leukemia cells, multiple myeloma cells, thyroid cancer cells, myelodysplastic syndrome cells, or small squamous cell carcinoma follicular carcinoma cells. Other types of cancer are described herein. The cells may be in vivo or in vitro.

[0208] With respect to the compounds of the present invention, one stereoisomer may exhibit better inhibition than another. For example, one atropisomer may exhibit inhibition, while other atropisomers may exhibit little to no inhibition.

[0209] In some embodiments, the methods or uses described herein further include administering an additional anticancer therapy. In some embodiments, the additional anticancer therapy is an EGFR inhibitor, a second Ras inhibitor, a SHP2 inhibitor, a SOS1 inhibitor, a Raf inhibitor, a MEK inhibitor, an ERK inhibitor, a PI3K inhibitor, a PTEN inhibitor, an AKT inhibitor, an mTORC1 inhibitor, a BRAF inhibitor, a PD-L1 inhibitor, a PD-1 inhibitor, a CDK4 / 6 inhibitor, a HER2 inhibitor, or a combination thereof. In some embodiments, the additional anticancer therapy is an SHP2 inhibitor. Other additional anticancer therapies are described herein.

[0210] Synthesis method The compounds described herein may be prepared from commercially available starting materials or synthesized using known organic, inorganic, or enzymatic processes.

[0211] The compounds of the present invention can be prepared by several methods well known to those skilled in the art of organic synthesis. For example, the compounds of the present invention can be synthesized using the methods shown in the following scheme, along with synthetic methods known in the field of organic synthesis chemistry, or variations thereof as understood by those skilled in the art. These methods include, but are not limited to, the methods described in the following scheme.

[0212] The compounds in Table 1 of this specification were prepared using the methods disclosed herein, or prepared using the methods disclosed herein in combination with the knowledge of those skilled in the art. The compounds in Table 2 can be prepared using the methods disclosed herein, or prepared using the methods disclosed herein in combination with the knowledge of those skilled in the art.

[0213] Scheme 1. General synthesis of aziridine-containing macrocyclic compounds

[0214] [ka]

[0215] As shown in Scheme 1, this type of compound can be prepared by reacting a suitable amine (1) with a protected amine (2)-containing carboxylic acid in the presence of a standard amide coupling reagent to obtain 3, and then deprotecting the amine to produce 4. The final compound (6) is obtained by coupling with aziridine carboxylate (5) in the presence of a standard amide coupling reagent.

[0216] Pharmaceutical composition and method of use Pharmaceutical composition and administration method The compounds related to the present invention are RAS inhibitors and are useful in cancer treatment. Accordingly, one embodiment of the present invention provides a pharmaceutical composition containing the compound of the present invention or a pharmaceutically acceptable salt thereof, and a pharmaceutically acceptable excipient, as well as a method for preparing such a composition using the compound of the present invention.

[0217] In the present invention, the term "pharmaceutical composition" means a compound such as the compound of the present invention, or a pharmaceutically acceptable salt thereof, formulated with a pharmaceutically acceptable excipient.

[0218] In some embodiments, the compound is present in the pharmaceutical composition in a unit dose appropriate for administration in a treatment regimen that exhibits a statistically significant probability of achieving a predetermined therapeutic effect when administered to the relevant population. In some embodiments, the pharmaceutical composition can be specifically formulated for administration in solid or liquid form, including: oral administration, e.g., oral tablets (aqueous or non-aqueous or suspension), tablets, e.g., cheek, sublingual, and those targeted for intracellular absorption, pills, powders, granules, and pastes for application to the tongue; parenteral administration, e.g., by subcutaneous, intramuscular, intravenous, or epidural injection as a sterile solution or suspension, or as a sustained-release formulation; topical application, e.g., as a cream, ointment, or sustained-release patch, or as a spray applied to the skin, lungs, or oral cavity; e.g., as a pessary, cream, or foam suitable for vaginal or rectal, sublingual, intraocular, transdermal, or transnasal, lung, and other mucosal surfaces.

[0219] As used herein, “pharmaceutically acceptable excipient” means any inert component having properties that are toxic and non-inflammatory within the subject matter (e.g., a vehicle capable of suspending or dissolving an active compound). Typical excipients include, for example, anti-adhesives, antioxidants, binders, coatings, compression aids, disintegrants, dyes (colorants), emollients, emulsifiers, fillers (diluents), film-forming or coating agents, flavorings, fragrances, lubricants (flow enhancers), preservatives, printing inks, adsorbents, suspending or dispersing agents, sweeteners, or hydration water. Examples of excipients include, but are not limited to, optionally substituted butylated hydroxytoluene (BHT), calcium carbonate, dibasic calcium phosphate, calcium stearate, croscarmellose, cross-linked polyvinylpyrrolidone, citric acid, crospovidone, cysteine, ethylcellulose, gelatin, optionally substituted hydroxypropylcellulose, optionally substituted hydroxypropylmethylcellulose, lactose, magnesium stearate, maltitol, mannitol, methionine, methylcellulose, methylparaben, microcrystalline cellulose, polyethylene glycol, polyvinylpyrrolidone, povidone, pregelatinized starch, propylparaben, retinyl palmitate, shellac, silicon dioxide, sodium carboxymethylcellulose, sodium citrate, sodium starch glycolate, sorbitol, starch (corn), stearic acid, sucrose, talc, titanium dioxide, vitamin A, vitamin E, vitamin C, and xylitol. Those skilled in the art are familiar with a wide variety of agents and materials useful as excipients. For example, for example, Ansel, et al., Ansel's Pharmaceutical Dosage Forms and Drug Delivery Systems.Philadelphia: Lippincott, Williams & Wilkins, 2004; Gennaro, et al., Remington: The Science and Practice of Pharmacy. Philadelphia: Lippincott, Williams & Wilkins, 2000; and Rowe, Handbook of Pharmaceutical Excipients. Chicago, Pharmaceutical See Press, 2005. In some embodiments, the composition comprises at least two different pharmaceutically acceptable excipients.

[0220] The compounds described herein may be provided or used in salt form, e.g., pharmaceutically acceptable salt form, whether or not expressly stated otherwise. As used herein, the term “pharmaceutically acceptable salt” means these salts of the compounds described herein that are suitable for use in contact with human and other animal tissues without causing excessive toxicity, irritation, allergic reactions, etc., within the normal range of reasonable medical judgment, and that are balanced by a reasonable benefit / risk ratio. pharmaceutically acceptable salts are well known in the art. For example, pharmaceutically acceptable salts are described in Berge et al., J. Pharmaceutical Sciences 66:1-19, 1977 and in Pharmaceutical Salts: Properties, Selection, and Use, (Eds. PHStahl). The salts can be prepared in situ during the final isolation and purification of the compounds described herein, or separately by reacting the free base group with a suitable organic acid.

[0221] The compounds of the present invention may have ionic groups so that they can be prepared as pharmaceutically acceptable salts. These salts may be acid addition salts with inorganic or organic acids, or, in the case of the acidic form of the compounds of the present invention, the salts may be prepared from inorganic or organic bases. In some embodiments, the compounds are prepared or used as pharmaceutically acceptable salts prepared as addition products of pharmaceutically acceptable acids or bases. Suitable pharmaceutically acceptable acids and bases are well known in the art for forming acid addition salts, such as hydrochloric acid, sulfuric acid, hydrobromic acid, acetic acid, lactic acid, or tartaric acid, and for forming basic salts, such as potassium hydroxide, sodium hydroxide, ammonium hydroxide, caffeine, and various amines. Methods for preparing suitable salts are well established in the art.

[0222] Typical acid addition salts include acetate, adipine, alginate, ascorbate, aspartate, benzenesulfonate, benzoate, bisulfate, borate, butyrate, camphorate, camphorsulfonate, citrate, cyclopentanepropionate, digluconate, dodecyl sulfate, ethanesulfonate, fumarate, glucoheptonate, glycerophosphate, hemisulfate, heptanoate, hexanoate, hydrobromide, hydrochloride, hydroiodide, and 2-(optionally substituted)hydroxylethane. Examples include sulfonates, lactobionates, lactates, laurates, lauryl sulfates, malates, maleates, malons, methanesulfonates, 2-naphthalenesulfonates, nicotinates, nitrates, oleates, oxalates, palmitates, pamoates, pectins, persulfates, 3-phenylpropionates, phosphates, picrates, pivalates, propions, stearates, succinates, sulfates, tartrates, thiocyanates, toluenesulfonates, undecanoates, and valersates. Representative alkali or alkaline earth metal salts include sodium, lithium, potassium, calcium, and magnesium, as well as non-toxic ammonium, quaternary ammonium, and amine cations (including, but not limited to, ammonium, tetramethylammonium, tetraethylammonium, methylamine, dimethylamine, trimethylamine, triethylamine, and ethylamine).

[0223] As used herein, the term “subject” means any element in the animal kingdom. In some embodiments, “subject” means a human at any stage of development. In some embodiments, “subject” means a human patient. In some embodiments, “subject” means a non-human animal. In some embodiments, the non-human animal is a mammal (e.g., rodents, mice, rats, rabbits, monkeys, dogs, cats, sheep, cattle, primates, or pigs). In some embodiments, subjects may be mammals, birds, reptiles, amphibians, fish, or insects, but are not limited to these. In some embodiments, subjects may be transgenic animals, genetically modified animals, or clones.

[0224] As used herein, the term “dosage form” means a physically distinct unit of a compound (e.g., the compound of the present invention) for administration to a subject. Each unit contains a predetermined amount of the compound. In some embodiments, such an amount is a unit dose (or its entire fraction) appropriate for administration according to an administration regimen (i.e., using a therapeutic administration regimen) that has been measured to correlate with a desired or beneficial outcome when administered to the relevant population. Those skilled in the art will understand that the total amount of a therapeutic composition or compound administered to a particular subject may be determined by one or more attending physicians and may involve administration of multiple dosage forms.

[0225] As used herein, the term “dosage regimen” means a set of unit doses (usually two or more) administered individually to a subject, usually separated by a period of time. In some embodiments, a given therapeutic compound (e.g., a compound of the present invention) has a recommended dosage regimen, which may have one or more doses. In some embodiments, the dosage regimen comprises multiple doses, each separated from the others by a period of time equal in length, and in some embodiments, the dosage regimen comprises multiple doses and at least two different period of time separating the individual doses. In some embodiments, all doses in the dosage regimen are identical unit doses. In some embodiments, different doses in the dosage regimen are different amounts. In some embodiments, the dosage regimen comprises one or more further doses, a first dose at a first dose, followed by a second dose different from the first dose. In some embodiments, the dosage regimen comprises one or more further doses, a first dose at a first dose, followed by a second dose that is the same as the first dose. In some embodiments, the administration regimen correlates with a desired or beneficial outcome when administered across relevant populations (i.e., it is a therapeutic administration regimen).

[0226] A "treatment regimen" refers to a dosing regimen across relevant populations that correlates with a desired or beneficial therapeutic outcome. The term “treatment” (and additionally, “to treat” or “to treat”) means, in its broadest sense, any administration of a substance (e.g., a compound of the present invention) that partially or completely remits, alleviates, reduces or inhibits a particular disease, disorder or condition; partially or completely delays the onset of a particular disease, disorder or condition; partially or completely reduces the severity of a particular disease, disorder or condition; or partially or completely reduces the occurrence of one or more symptoms, features, or causes of a particular disease, disorder or condition. In some embodiments, such treatment may be administered to subjects who show no signs of the related disease, disorder or condition, or to subjects who show only the initial signs of the disease, disorder or condition. Alternatively, or in addition, in some embodiments, such treatment may be administered to subjects showing established signs of one or more of the related diseases, disorders or conditions. In some embodiments, treatment may be administered to subjects diagnosed with suffering from the related disease, disorder or condition. In some embodiments, the treatment may be in subjects known to have one or more susceptibility factors that are statistically correlated with an increased risk of progression of the associated disease, disorder, or condition.

[0227] The term “therapeutic dose” means a quantity sufficient to treat a disease, disorder, or condition when administered to a population suffering from or suspected of having a disease, disorder, or condition, according to a therapeutic administration regimen. In some embodiments, a therapeutic dose is a quantity that reduces the onset or severity of one or more symptoms of the disease, disorder, or condition, or delays the onset of one or more symptoms of the disease, disorder, or condition. Those skilled in the art will understand that the term “therapeutic dose” does not actually require that successful treatment be achieved in a particular individual. Rather, a therapeutic dose can be a quantity that, when administered to subjects requiring such treatment, produces a specific desired pharmacological response in a significant number of subjects. It is specifically understood that a particular subject may actually be “refractory” to the “therapeutic dose.” In some embodiments, a reference to a therapeutic dose may refer to a quantity measured in one or more specific tissues (e.g., tissues affected by the disease, disorder, or condition) or fluids (e.g., blood, saliva, serum, sweat, tears, urine). Those skilled in the art will understand that in some embodiments, a therapeutically effective amount can be formulated or administered as a single dose. In some embodiments, a therapeutically effective amount can be formulated or administered in multiple doses, for example, as part of an administration regimen.

[0228] The compounds of the present invention, or pharmaceutically acceptable salts thereof, can be formulated as pharmaceutical or veterinary compositions for use as a treatment for a target. Depending on the target to be treated, the method of administration, and the type of treatment desired, e.g., prevention, prophylaxis, or treatment, the compounds, or pharmaceutically acceptable salts thereof, are formulated in a manner that matches these parameters. A summary of such techniques can be found in Remington: The Science and Practice of Pharmacy, 21 stThis can be found in Edition, Lippincott Williams & Wilkins, (2005); and Encyclopedia of Pharmaceutical Technology, eds. J. Swarbrick and JCBoylan, 1988–1999, Marcel Dekker, New York, each of which is incorporated herein by reference.

[0229] Each composition can be prepared according to conventional mixing, granulation, or coating methods, and the pharmaceutical composition may contain about 0.1% to about 99%, about 5% to about 90%, or about 1% to about 20% (by weight or by volume) of the compound of the present invention or a pharmaceutically acceptable salt thereof. In some embodiments, the compounds described herein or a pharmaceutically acceptable salt thereof may be present in total in an amount of 1 to 95% of the total weight of the composition, such as the pharmaceutical composition.

[0230] The composition can be provided in dosage forms suitable for intra-articular, oral, parenteral (e.g., intravenous, intramuscular), rectal, cutaneous, subcutaneous, topical, transdermal, sublingual, transnasal, intravaginal, intrabladderal, intraurethral, ​​intrathecal, epidural, transaural, or intraocular administration, or for injection, inhalation, or direct contact with the nasal, genitourinary, reproductive, or oral mucosa. Accordingly, the pharmaceutical composition may be in the form of, for example, tablets, capsules, pills, powders, granules, suspensions, emulsions, solutions, gels including hydrogels, pastes, ointments, creams, plasters, oral medications, infiltration delivery devices, suppositories, enemas, injections, implants, sprays, preparations suitable for iontophoresis delivery, or aerosols. The composition can be formulated according to conventional pharmaceutical regulations.

[0231] As used herein, the term “administration” means the administration of a composition (e.g., a compound, or a preparation containing a compound as described herein) to a subject or system. Administration to an animal subject (e.g., a human) may be by any suitable route. For example, in some embodiments, administration may be bronchial (including bronchial infusion), cheek, enteral, interdermal, intra-arterial, intradermal, intragastric, intramedullary, intramuscular, transnasal, intraperitoneal, intrathecal, intravenous, intraventricular, mucosal, nasal, oral, rectal, subcutaneous, sublingual, topical, intratracheal (including intratracheal infusion), transdermal, vaginal, or intravitreous.

[0232] Formulations can be prepared in a manner suitable for systemic administration or local or local administration. Systemic formulations may be designed for injection (e.g., intramuscular, intravenous, or subcutaneous injection), or they may be prepared for transdermal, transmucosal, or oral administration. Formulations generally contain diluents, and optionally adjuvants, buffers, and preservatives. The compound, or a pharmaceutically acceptable salt thereof, may also be administered in a liposome composition or as a microemulsion.

[0233] For injection, formulations can be prepared in conventional forms, such as a solution or suspension, or as a solid suitable for solution or suspension in liquid before injection, or as an emulsion. Suitable excipients include, for example, water, saline, dextrose, and glycerol. Such compositions may also contain certain amounts of non-toxic auxiliary substances (e.g., wetting agents or emulsifiers), pH buffers, such as sodium acetate and sorbitan monolaurate.

[0234] Various sustained-release systems for pharmaceuticals have also been devised. See, for example, U.S. Patent No. 5,624,677. Systemic administration can also be achieved through relatively non-invasive methods such as suppositories, transdermal patches, transmucosal delivery, and intranasal administration. Oral administration is also suitable for the compounds of the present invention or pharmaceutically acceptable salts thereof. Preferred forms, as understood in the art, include syrups, capsules, and tablets.

[0235] Each compound described herein, or a pharmaceutically acceptable salt thereof, can be formulated in various ways known in the art. For example, the first and second agents of a combination therapy can be formulated together or individually. Other modalities of combination therapy are described herein.

[0236] Individually formulated or separately manufactured drugs can be packaged together as a kit. Non-limiting examples include, but are not limited to, kits containing two pills, pills and powder, suppositories and liquid in a vial, or two topical creams. A kit may include any components that assist in administering a unit dose to a subject, such as vials for reconstituting powder forms, syringes for injection, customized IV delivery systems, or inhalers. In addition, a unit dose kit may include instructions for preparing or administering the composition. A kit may be manufactured as a single-use unit dose for a particular subject, or as a multi-use unit dose for a specific subject (where the efficacy of individual compounds or their pharmaceutically acceptable salts changes at a constant concentration or as treatment progresses), or a kit may contain multi-use doses (bulk packaging) suitable for administration to multiple subjects. The components of a kit can be assembled into cartons, blister packs, bottles, tubes, etc.

[0237] Formulations for oral use include tablets containing the active ingredient(s) in a mixture with pharmaceutically acceptable, non-toxic excipients. These excipients may include, for example, inert diluents or fillers (e.g., sucrose, sorbitol, sugars, mannitol, microcrystalline cellulose, starch containing potato starch, calcium carbonate, sodium chloride, lactose, calcium phosphate, calcium sulfate, or sodium phosphate); granulators and disintegrants (e.g., cellulose derivatives containing microcrystalline cellulose, starch containing potato starch, croscarmellose sodium, alginate, or arginine); binders (e.g., sucrose, glucose, sorbitol, acacia, arginine, sodium alginate, gelatin, starch, pregelatinized starch, microcrystalline cellulose, aluminum magnesium silicate, sodium carboxymethylcellulose, methylcellulose, optionally substituted hydroxypropylmethylcellulose, ethylcellulose, polyvinylpyrrolidone, or polyethylene glycol); as well as smoothers, lubricants, and anti-adhesives (e.g., magnesium stearate, zinc stearate, stearic acid, silica, hydrogenated vegetable oil, or talc). Other pharmaceutically acceptable excipients may include colorants, flavorings, plasticizers, wetting agents, and buffering agents.

[0238] Two or more compounds can be mixed or fractionated in a tablet, capsule, or other vehicle. In one example, the first compound is contained inside the tablet, the second compound is on the outside, and a substantial portion of the second compound is released before the first compound.

[0239] Formulations for oral use may be provided as chewable tablets, or as rigid gelatin capsules in which the active ingredient is mixed with an inert solid diluent (e.g., potato starch, lactose, microcrystalline cellulose, calcium carbonate, calcium phosphate, or kaolin), or as flexible gelatin capsules in which the active ingredient can be mixed with water or an oil culture medium, e.g., peanut oil, liquid paraffin, or olive oil. Powders, granules, and pellets can be prepared using the above-mentioned components below tablets and capsules by conventional methods, for example, using a mixer, fluidized bed apparatus, or spray dryer.

[0240] Dissolution or diffusion-controlled release can be achieved by appropriate coatings for tablet, capsule, pellet, or granule formation of the compound, or by incorporating the compound, or a pharmaceutically acceptable salt thereof, into a suitable matrix. Sustained-release coatings may include the coating materials described above, or one or more of the following: shellac, beeswax, glycowax, castor wax, carnauba wax, stearyl alcohol, glyceryl monostearate, glyceryl distearate, glycerol palmitostearate, ethylcellulose, acrylic resin, dl-polylactic acid, cellulose acetate / butyrate, polyvinyl chloride, polyvinyl acetate, vinylpyrrolidone, polyethylene, polymethacrylate, methyl methacrylate, 2-(optionally substituted) hydroxyl methacrylate, methacrylate hydrogel, 1,3-butylene glycol, ethylene glycol methacrylate, or polyethylene glycol. In sustained-release matrix formulations, examples of matrix materials include hydrated methylcellulose, carnauba wax and stearyl alcohol, Carbopol 934, silicone, glyceryl tristearate, methyl methacrylate, polyvinyl chloride, polyethylene, or halogenated fluorocarbons.

[0241] Liquid forms in which the compounds of the present invention, or pharmaceutically acceptable salts thereof, and compositions may be incorporated for oral administration include aqueous solutions, suitably flavored syrups, aqueous or oily suspensions, and emulsions flavored with edible oils such as cottonseed oil, sesame oil, coconut oil, or peanut oil, as well as elixirs and similar pharmaceutical vehicles.

[0242] In general, when administered to humans, the oral dose of any of the compounds of the present invention, or any of its pharmaceutically acceptable salts, depends on the properties of the compound and can be quickly determined by those skilled in the art. The dose may be, for example, about 0.001 mg to about 2000 mg / day, about 1 mg to about 1000 mg / day, about 5 mg to about 500 mg / day, about 100 mg to about 1500 mg / day, about 500 mg to about 1500 mg / day, about 500 mg to about 2000 mg / day, or any of these range variables. In some embodiments, the daily dose range for oral administration may be, for example, within the range of about 0.001 mg to about 2000 mg / kg human body weight, in single or divided doses. On the other hand, in some cases, it may be necessary to use dosages outside these limits.

[0243] In some embodiments, the pharmaceutical composition may further contain additional compounds having antiproliferative activity. Depending on the method of administration, the compounds, or pharmaceutically acceptable salts thereof, are formulated into a suitable composition that allows for easy delivery. Each compound, or pharmaceutically acceptable salt thereof, of the combination therapy can be formulated in various ways known in the art. For example, the first and second agents of the combination therapy can be formulated together or individually. Preferably, the first and second agents are formulated together for simultaneous or near-simultaneous administration.

[0244] It will be understood that the compounds and pharmaceutical compositions of the present invention can be formulated and used in combination therapy, that is, the compounds and pharmaceutical compositions can be formulated or administered simultaneously with, before, or after one or more other desired therapeutic agents or medical procedures. In specific combinations of treatments (therapeutic agents or procedures) using combination regimens, the suitability of the desired therapeutic agent or procedure and the desired therapeutic effect to be achieved will be taken into consideration. Furthermore, it will be understood that the treatments used may achieve the desired effect for the same disease, or different effects (e.g., control of any adverse effects).

[0245] As described herein, each drug in combination therapy may be administered independently once to four times a day for a period of one to one year, and may also be administered throughout the patient's lifetime. Chronic, long-term administration may be indicated.

[0246] How to use In some embodiments, the present invention discloses methods for treating diseases or disorders characterized by ectopic Ras activity caused by Ras variants. In some embodiments, the disease or disorder is cancer.

[0247] Accordingly, the present invention also provides a method for treating cancer in a subject requiring treatment for cancer, wherein the method comprises administering to the subject a therapeutically effective amount of the compound of the present invention, or a pharmaceutically acceptable salt thereof, or a pharmaceutical composition containing such compound or salt. In some embodiments, the cancer is colorectal cancer, non-small cell lung cancer, small cell lung cancer, pancreatic cancer, appendiceal cancer, melanoma, acute myeloid leukemia, small intestine cancer, ampulla cancer, germ cell carcinoma, cervical cancer, cancer of unknown primary origin, endometrial cancer, esophageal cancer, GI neuroendocrine cancer, ovarian cancer, sex cord-stromal tumor cancer, hepatobiliary cancer, or bladder cancer. In some embodiments, the cancer is appendiceal cancer, endometrial cancer, or melanoma. The present invention also provides a method for treating Ras protein-related disorders in a subject requiring treatment for Ras protein-related disorders, wherein the method comprises administering to the subject a therapeutically effective amount of the compound of the present invention, or a pharmaceutically acceptable salt thereof, or a pharmaceutical composition containing such compound or salt.

[0248] In some embodiments, the compounds of the present invention, or pharmaceutically acceptable salts thereof, pharmaceutical compositions containing such compounds or salts, and the methods provided herein can be used to treat a wide variety of cancers, including, for example, lung cancer, prostate cancer, breast cancer, brain cancer, skin cancer, cervical cancer, and testicular cancer. More specifically, cancers treatable by the compounds or salts thereof, pharmaceutical compositions containing such compounds or salts, and the methods of the present invention include, but are not limited to, astrocyte, breast, cervix, colorectal, uterine, esophageal, gastric, head and neck, hepatocyte, larynx, lung, pharyngeal, ovarian, prostate, and thyroid cancer and sarcoma. Other cancers include, for example: Cardiac malignancies, for example, non-epithelial malignancies (angiosarcoma, fibrosarcoma, rhabdomyosarcoma, liposarcoma), myxoma, rhabdomyomas, fibromas, lipomas, and teratomas; Lung cancers, for example, bronchogenic carcinomas (squamous cell carcinomas, anaplastic small cell carcinomas, anaplastic large cell carcinomas, adenocarcinomas), alveolar (bronchial) carcinomas, bronchial adenomas, non-epithelial malignancies, lymphomas, chondrotoxic hamartomas, mesotheliomas; The gastrointestinal tract, for example, the esophagus (squamous cell carcinoma, adenocarcinoma, leiomyosarcoma, lymphoma), the stomach (epithelial malignant tumor, lymphoma, leiomyosarcoma), the pancreas (ductal adenocarcinoma, islet cell tumor, glucagon-producing tumor, gastrin-producing tumor, carcinoid tumor, VIP-producing tumor), the small intestine (adenocarcinoma, lymphoma, carcinoid tumor, Kaposi's sarcoma, leiomyoma, hemangioma, lipoma, neurofibroma, fibroma), and the large intestine (adenocarcinoma, tubular adenoma, chorioadenoma, hamartoma, leiomyoma); The urogenital organs, for example, the kidneys (adenocarcinoma, Wilms' tumor, (nephroblastoma), lymphoma, leukemia), bladder and urethra (squamous cell carcinoma, transitional cell carcinoma, adenocarcinoma), prostate (adenocarcinoma, non-epithelial malignant tumor), testicles (seminal carcinoma, teratoma, embryonic carcinoma, teratocarcinoma, choriocarcinoma, non-epithelial malignant tumor, stromal cell carcinoma, fibroma, fibroadenoma, adenoid tumor, lipoma); Liver, for example, liver cancer (hepatocellular carcinoma), cholangiocarcinoma, hepatoblastoma, angiosarcoma, hepatocellular adenoma, hemangioma; Biliary tract cancer, for example, gallbladder cancer, ampulla cancer, bile duct cancer; Bone, for example, osteogenic sarcoma (osteosarcoma), fibrosarcoma, malignant fibrous histiocytoma, chondrosarcoma, Ewing's sarcoma, malignant lymphoma (reticulosarcoma), multiple myeloma, malignant giant cell tumor chordoma, osteochondroma (osteochondroma), benign chondroma, chondroblastoma, chondromyxofibroma, osteoid osteoma, and giant cell tumor; Nervous system, for example, skull (osteoma, hemangioma, granuloma, xanthomas, degenerative osteitis), meninges (meningioma, meningiosarcoma, gliomas), brain (astrocytoma, medulloblastoma, glioma, ependymoma, germ cell tumor (pineal glandoma), glioblastoma multiforme, oligodendroglioma, schwannoma, retinoblastoma, congenital tumors), spinal neurofibroma, neurofibromatosis type 1, meningioma, glioma, non-epithelial malignant tumors); Gynecology, for example, the uterus (endometrial cancer, uterine cancer, endometrial cancer of the uterine body), the cervix (cervical cancer, pre-tumor cervical dysplasia), the ovaries (ovarian cancer (serous cystadenocarcinoma, mucinous cystadenocarcinoma, unclassifiable cancer), granulosa-theca cell tumor, Sertoli-Leydig cell tumor, undifferentiated germ cell tumor, malignant teratoma), the vulva (squamous cell carcinoma, carcinoma in situ, adenocarcinoma, fibrosarcoma, melanoma), the vagina (clear cell carcinoma, squamous cell carcinoma, staphyloid sarcoma (embryonic rhabdomyosarcoma), the fallopian tubes (epithelial malignant tumors); Hematological conditions, e.g., blood (myeloid leukemia (acute and chronic), acute lymphoblastic leukemia, chronic lymphocytic leukemia); myeloproliferative disorders (e.g., myelofibrosis and myeloproliferative neoplasms); multiple myeloma; myelodysplastic syndromes, Hodgkin's disease, non-Hodgkin lymphoma (malignant lymphoma); Skin conditions such as malignant melanoma, basal cell carcinoma, squamous cell carcinoma, Kaposi's sarcoma, dysplastic moles, lipomas, hemangiomas, dermatofibromas, keloids, and psoriasis; Adrenal glands, for example, neuroblastoma.

[0249] In some embodiments, the Ras protein is wild-type (Ras WT Therefore, in some embodiments, the compounds of the present invention are Ras WT (For example, K-Ras WT H-Ras WT , or N-Ras WT It is used in treatment methods for patients with cancer including ). In some embodiments, the Ras protein is used for Ras amplification (e.g., K-Ras amp Therefore, in some embodiments, the compounds of the present invention are Ras amp (K-Ras amp H-Ras amp , or N-Ras amp It is used in methods for treating patients with cancer including ). In some embodiments, the cancer includes Ras mutations such as the Ras mutations described herein. In some embodiments, the mutations are selected from the following: (a) The following K-Ras variants: G12D, G12V, G12C, G13D, G12R, G12A, Q61H, G12S, A146T, G13C, Q61L, Q61R, K117N, A146V, G12F, Q61K, L19F, Q22K, V14I, A59T, A146P, G13R, G12L, or G13V, and combinations thereof; (b) The following H-Ras variants: Q61R, G13R, Q61K, G12S, Q61L, G12D, G13V, G13D, G12C, K117N, A59T, G12V, G13C, Q61H, G13S, A18V, D119N, G13N, A146T, A66T, G12A, A146V, G12N, or G12R, and combinations thereof; and (c) The following N-Ras variants: Q61R, Q61K, G12D, Q61L, Q61H, G13R, G13D, G12S, G12C, G12V, G12A, G13V, G12R, P185S, G13C, A146T, G60E, Q61P, A59D, E132K, E49K, T50I, A146V, or A59T, and combinations thereof; Or any combination of the above. In some embodiments, the compound may inhibit both K-Ras G12C and K-Ras G12D. In some embodiments, the compound may inhibit both K-Ras G12D and K-Ras G13D. In some embodiments, the compound may inhibit both K-Ras G12V and K-Ras G12S. In some embodiments, the compound of the present invention inhibits Ras in addition to one or more additional Ras mutations. WT Inhibits (e.g., K, H, or N-Ras) WT (and K-Ras G12D). In some embodiments, the compounds of the present invention are Ras in addition to one or more additional Ras mutations. amp Inhibits (e.g., K, H, or N-Ras) amp (And K-Ras G12D).

[0250] Methods for detecting Ras mutations are known in the art. Such methods include, but are not limited to, direct sequencing and the use of highly sensitive diagnostic assays (using the CE-IVD mark) including TheraScreen PCR; AmoyDx; PNAClamp; RealQuality; EntroGen; LightMix; StripAssay; Hybcell plexA; Devyser; Surveyor; Cobas; and TheraScreen Pyro, which are incorporated herein by reference, for example. See also, for example, WO2020 / 106640.

[0251] In some embodiments, the cancer is non-small cell lung cancer, and the Ras mutation includes a K-Ras mutation such as K-Ras G12D. In some embodiments, the cancer is colorectal cancer, and the Ras mutation includes a K-Ras mutation such as K-Ras G12D. In some embodiments, the cancer is pancreatic cancer, and the Ras mutation includes a K-Ras mutation such as K-Ras G12D. In some embodiments, the cancer is pancreatic cancer, and the Ras mutation includes an N-Ras mutation such as N-Ras G12D. Unless otherwise specified, in any of the foregoing, the compound is also Ras WT (For example, K-, H-, or N-Ras) WT ), or Ras amp (For example, K-, H-, or N-Ras) amp ) may be inhibited.

[0252] A method for inhibiting the Ras protein in cells is also provided, wherein the method comprises contacting the cells with an effective amount of the compound of the present invention or a pharmaceutically acceptable salt thereof. The compound or a pharmaceutically acceptable salt thereof can inhibit two or more types of Ras protein in cells. A method for inhibiting RAF-Ras binding is also provided, wherein the method comprises contacting the cells with an effective amount of the compound of the present invention or a pharmaceutically acceptable salt thereof. The cells may be cancer cells. The cancer cells may be of any type of cancer described herein. The cells may be in vivo or in vitro.

[0253] Combination therapy The methods of the present invention may include compounds of the present invention used alone or in combination with one or more additional therapies (e.g., non-pharmacological treatments or therapeutic agents). The dosage of one or more additional therapies (e.g., non-pharmacological treatments or therapeutic agents) may be reduced from the standard dosage when administered alone. For example, the dosage may be determined empirically from the combination and permutation of drugs or estimated by isoborographic analysis (e.g., Black et al.). al., Neurology 65:S3-S6(2005)).

[0254] The compounds of the present invention may be administered before, after, or concurrently with one or more additional therapies. When combined, the dosage of the compounds of the present invention and the dosage of one or more additional therapies (e.g., non-pharmacological treatments or therapeutic agents) provide a therapeutic effect (e.g., a synergistic or additive therapeutic effect). The compounds of the present invention and additional therapies, such as anticancer agents, may be administered together or separately in a single pharmaceutical composition, and if administered separately, they may be administered simultaneously or sequentially. Such sequential administration may have short or long intervals between doses.

[0255] In some embodiments, additional therapy involves the administration of side effect limiting agents (e.g., agents intended to reduce the occurrence or severity of side effects of the treatment). For example, in some embodiments, the compounds of the present invention may also be used in combination with therapeutic agents for treating nausea. Examples of agents that can be used to treat nausea include dronabinol, granisetron, metoclopramide, ondansetron, and prochlorperazine, or pharmaceutically acceptable salts thereof.

[0256] In some embodiments, one or more additional therapies include non-pharmacological treatment (e.g., surgery or radiotherapy). In some embodiments, one or more additional therapies include therapeutic agents (e.g., compounds or biological agents that are anti-angiogenic agents, signaling inhibitors, antiproliferative agents, glycolysis inhibitors, or autophagy inhibitors). In some embodiments, one or more additional therapies include non-pharmacological treatment (e.g., surgery or radiotherapy) and therapeutic agents (e.g., compounds or biological agents that are anti-angiogenic agents, signaling inhibitors, antiproliferative agents, glycolysis inhibitors, or autophagy inhibitors). In other embodiments, one or more additional therapies include two therapeutic agents. In yet another embodiment, one or more additional therapies include three therapeutic agents. In some embodiments, one or more additional therapies include four or more therapeutic agents.

[0257] In this section on combination therapies, all references for the listed drugs, or their pharmaceutically acceptable salts, solvates, isomers (e.g., stereoisomers), prodrugs, or tautomers, whether explicitly stated or not, are incorporated by reference.

[0258] Non-pharmacological therapy Examples of non-pharmacological treatments include, but are not limited to, radiation therapy, cryotherapy, hyperthermia, surgery (e.g., surgical excision of tumor tissue), and T-cell adoptive transfer (ACT) therapy.

[0259] In some embodiments, the compounds of the present invention can be used as adjuvant therapy after surgery. In some embodiments, the compounds of the present invention can be used as preoperative adjuvant therapy before surgery.

[0260] Radiotherapy may be used in subjects (e.g., mammals (e.g., humans)) to inhibit abnormal cell proliferation or to treat hyperproliferative disorders such as cancer. Techniques for administering radiotherapy are known in the art. Radiotherapy can be administered by one or a combination of several methods, including, but not limited to, external beam therapy, internal radiotherapy, implantable radiation, stereotactic radiosurgery, total body radiotherapy, radiotherapy, and permanent or transient interstitial near-brightening therapy. As used herein, the term “near-brightening therapy” refers to radiotherapy delivered by spatially confined radioactive material inserted into the body at or near a tumor or other proliferative tissue disease site. The term is intended to include, but is not limited to, exposure to radioisotopes (e.g., At-211, I-131, I-125, Y-90, Re-186, Re-188, Sm-153, Bi-212, P-32, and radioisotopes of Lu). Suitable radiation sources for use as cell modifiers in the present invention include both solids and liquids. In non-limiting examples, the radiation source may be radionuclides such as I-125, I-131, Yb-169, Ir-192 as solid sources, I-125 as a solid source, or other radionuclides that emit photons, beta particles, gamma rays, or other therapeutic rays. The radioactive material may also be a fluid prepared from any solution of the radionuclide(s), e.g., a solution of I-125 or I-131, or the radioactive fluid may be produced using a suitable fluid slurry containing small particles of a solid radionuclide such as Au-198 or Y-90. Furthermore, the radionuclide(s) may be embodied in gels or radioactive microspheres.

[0261] In some embodiments, the compounds of the present invention can make abnormal cells more sensitive to radiotherapy aimed at killing or inhibiting the proliferation of such cells. Therefore, the present invention further relates to a method for sensitizing abnormal cells in mammals to radiotherapy, the method comprising administering to a mammal an amount of the compound of the present invention effective in sensitizing the abnormal cells to radiotherapy. The amount of the compound in this method can be determined according to the means for determining an effective amount of such compound described herein. In some embodiments, the compounds of the present invention may be used as adjuvant therapy after radiotherapy or as neoadjuvant therapy before radiotherapy.

[0262] In some embodiments, the non-pharmacological treatment is T cell adoptive transfer (ACT) therapy. In some embodiments, the T cells are activated T cells. The T cells can be modified to express a chimeric antigen receptor (CAR). CAR-modified T (CAR-T) cells can be generated by any method known in the art. For example, CAR-T cells can be generated by introducing a suitable expression vector encoding a CAR into T cells. The T cell source is obtained from a subject before proliferation and genetic modification of the T cells. T cells can be obtained from several sources, including peripheral blood mononuclear cells, bone marrow, lymph node tissue, umbilical cord blood, thymic tissue, tissue from infection sites, ascites, pleural fluid, spleen tissue, and tumors. In certain embodiments of the present invention, any number of T cell lines available in the art may be used. In some embodiments, the T cells are autologous T cells. Regardless of whether the T cells are genetically modified to express a desired protein (e.g., CAR), T cells are generally, for example, U.S. Patent Nos. 6,352,694, 6,534,055, 6,905,680, 6,692,964, 5,858,358, 6,887,466, 6,905,681, and It can be activated and propagated using the methods described in Nos. 7,144,575, 7,067,318, 7,172,869, 7,232,566, 7,175,843, 7,572,631, 5,883,223, 6,905,874, 6,797,514, and 6,867,041.

[0263] Therapeutic drugs The therapeutic agent may be a compound used to treat cancer or related conditions. The compounds of the present invention can be used in combination with a second, third, or fourth therapeutic agent, or more therapeutic agents. The compounds of the present invention can be used in combination with one or more therapeutic agents, along with one or more non-pharmacological therapies.

[0264] For example, the therapeutic agent may be a steroid. Steroids are known in the art. Therefore, in some embodiments, one or more additional therapies include steroids. Preferred steroids include 21-acetoxypregnenolone, alclomethasone, algestone, amcinonide, beclomethasone, betamethasone, budesonide, chloroprednisone, clobetasol, crocortol, cloprednol, corticosterone, cortisone, cortibazole, deflazacort, desonide, dexoxymethasone, dexamethasone, diflorasone, diflucortol, difprednate, enoxolone, fluazacort, fluchloronide, flumethasone, flunisolide, fluocinolone acetonide, fluocinonide, flucortin butyl, flucortolone, fluorometholone, fluperolone acetate, flupredniden acetate, fluprednisolone, and flulandrenolide. Examples include, but are not limited to, fluticasone propionate, formocortal, halcinonide, halobetazole propionate, halomethasone, hydrocortisone, loteprednol etavonate, mazipredone, medrisone, meprednisone, methylprednisolone, mometasone furoate, paramethasone, prednicarbate, prednisolone, prednisolone 25-diethylaminoacetate, sodium prednisolone phosphate, prednisone, prednival, prednylidene, rimexolone, thixocortol, triamcinolone, triamcinolone acetonide, triamcinolone benetonide, triamcinolone hexaacetonide, and their salts or derivatives.

[0265] Further examples of therapeutic agents that may be used in combination therapy with the compounds of the present invention include the compounds described in the following patents: U.S. Patent Nos. 6,258,812, 6,630,500, 6,515,004, 6,713,485, 5,521,184, 5,770,599, 5,747,498, 5,990,141, 6,235,764, and 8,623,885, as well as International Patent Application No. WO0 Issues 1 / 37820, WO01 / 32651, WO02 / 68406, WO02 / 66470, WO02 / 55501, WO04 / 05279, WO04 / 07481, WO04 / 07458, WO04 / 09784, WO02 / 59110, WO99 / 45009, WO00 / 59509, WO99 / 61422, WO00 / 12089, and WO00 / 02871.

[0266] The therapeutic agent may be a biological agent used to treat cancer or related conditions (e.g., cytokines (e.g., interferons or interleukins such as IL-2)). Biological agents are known in the art. In some embodiments, the biological agent is a biological agent of the immunoglobulin system, e.g., monoclonal antibodies (e.g., humanized antibodies, fully human antibodies, Fc fusion proteins, or functional fragments thereof) that inflict pain on a target to stimulate an anti-cancer response or antagonize antigens important to cancer. Antibody-drug conjugates are also included.

[0267] The therapeutic agent may be a T-cell checkpoint inhibitor. Such checkpoint inhibitors are known in the art. In one embodiment, the checkpoint inhibitor is an inhibitory antibody (e.g., a monospecific antibody such as a monoclonal antibody). The antibody may be, for example, a humanized or fully human antibody. In some embodiments, the checkpoint inhibitor is a fusion protein, e.g., an Fc receptor fusion protein. In some embodiments, the checkpoint inhibitor is a drug such as an antibody that interacts with a checkpoint protein. In some embodiments, the checkpoint inhibitor is a drug such as an antibody that interacts with a ligand of a checkpoint protein. In some embodiments, the checkpoint inhibitor is an inhibitor (e.g., an inhibitory antibody or small molecule inhibitor) of CTLA-4 (e.g., an anti-CTLA-4 antibody or fusion protein). In some embodiments, the checkpoint inhibitor is a PD-1 inhibitor or antagonist (e.g., an inhibitory antibody or small molecule inhibitor). In some embodiments, the checkpoint inhibitor is a PD-L1 inhibitor or antagonist (e.g., an inhibitory antibody or small molecule inhibitor). In some embodiments, the checkpoint inhibitor is an inhibitor or antagonist (e.g., an inhibitory antibody or Fc fusion or small molecule inhibitor) of PDL-L2 (e.g., a PDL-L2 / Ig fusion protein). In some embodiments, the checkpoint inhibitor is an inhibitor or antagonist (e.g., an inhibitory antibody or small molecule inhibitor) of B7-H3, B7-H4, BTLA, HVEM, TIM3, GAL9, LAG3, VISTA, KIR, 2B4, CD160, CGEN-15049, CHK1, CHK2, A2aR, B-7 family ligands, or combinations thereof.In some embodiments, the checkpoint inhibitor is pembrolizumab, nivolumab, PDR001 (NVS), REGN2810 (Sanofi / Regeneron), PD-L1 antibody, e.g., avelumab, durvalumab, atezolizumab, pizilizumab, JNJ-63723283 (JNJ), BGB-A317 (BeiGene & Celgene), or Preusser, M. et al. Checkpoint inhibitors disclosed in al. (2015) Nat. Rev. Neurol. include, but are not limited to, ipilimumab, tremelimumab, nivolumab, pembrolizumab, AMP224, AMP514 / MEDI0680, BMS936559, MEDl4736, MPDL3280A, MSB0010718C, BMS986016, IMP321, lirirumab, IPH2101, 1-7F9, and KW-6002.

[0268] The therapeutic agent may be an anti-TIGIT antibody, such as MBSA43, BMS-986207, MK-7684, COM902, AB154, MTIG7192A, or OMP-313M32 (ethigirimab). Other anti-TIGIT antibodies are known in the art.

[0269] The therapeutic agent may be a drug that treats cancer or related conditions (e.g., cytotoxic agents, non-peptide small molecules, or other compounds useful in treating cancer or related conditions, collectively referred to as "anticancer agents"). Anticancer agents may be, for example, chemotherapeutic agents or targeted therapy agents. Such agents are known in the art.

[0270] Anticancer agents include mitotic inhibitors, insertive antibiotics, growth factor inhibitors, cell cycle inhibitors, enzymes, topoisomerase inhibitors, biological response modifiers, alkylating agents, antimetabolites, folic acid analogs, pyrimidine analogs, purine analogs and related inhibitors, vinca alkaloids, epipodophyllotoxin, antibiotics, L-asparaginase, topoisomerase inhibitors, interferons, platinum-coordination complexes, anthracendione-substituted ureas, methylhydrazine derivatives, corticosteroids, progestins, estrogens, antiestrogens, androgens, and gonadotropin-releasing hormone analogs. Further anticancer agents include leucovorin (LV), irenotecan, oxaliplatin, capecitabine, paclitaxel, and doxetaxel. In some embodiments, one or more additional therapies comprise two or more anticancer agents. Two or more anticancer drugs can be used in a cocktail, either administered in combination or individually. Preferred administration regimens for combination anticancer drugs are known in the art and are described, for example, in Saltz et al., Proc.Am.Soc.Clin.Oncol.18:233a(1999) and Douillard et al., Lancet 355(9209):1041-1047(2000).

[0271] Other non-exclusive examples of anticancer drugs include Gleevec® (imatinib mesylate); Kyprolis® (carfilzomib); Velcade® (bortezomib); Casodex (bicalutamide); Iressa® (gefitinib); alkylating agents, e.g., thiotepa and cyclophosphamide; alkyl sulfonates, e.g., busulfan, improsulfan, and pigosulfan; aziridines, e.g., be Nzodopa, carbocone, metsuredopa, and uredopa; ethyleneimines and methylamelamines, e.g., altoretamine, triethylenemelamine, triethylenephosphoramide, triethylenethiophosphoramide, and trimethylolomelamine; acetogenins (especially bratacin and bratacinone); camptothecin (e.g., synthetic analog topotecan); briostatin; callistatin; CC-1065 (e.g., its adzeresin, karzeresin, and Beizelesin synthetic analogs); cryptophycin (specifically, cryptophycin 1 and cryptophycin 8); dorastatin; duocalmycin (e.g., synthetic analogs KW-2189 and CB1-TM1); eryuterobin; pancratistatin; sarcodictiin A; spongistatin; nitrogen mustard, e.g., chlorambucil, chlornafadin, colophosphamide, estramustine, ifosfamide, mechloretamine, mechloretamine Minoxide hydrochloride, melphalan, nobenbitin, fenestrine, prednimustine, trophosphamide, and uracil mustard; nitrosoureas, e.g., camulstine, chlorozotosine, fotemustine, lomustine, nimustine, and ranimustine; antibiotics, e.g., engine antibiotics (e.g., calicheamicin, e.g., calicheamicin gamma II and calicheamicin omega II (see, e.g., Agnew, Chem. Intl. Ed Engl. 33:183-186 (1994)); dynemicins such as dynemicin A; bisphosphonates such as clodronate; esperamicin;Neocardinostatin chromophore and related pigment protein enediin antibiotic chromophore, acrasinomycin, actinomycin, autoramycin, azaserin, bleomycin, kactinomycin, calicheamicin, carabicin, kaminomycin, carminomycin, cardinophilin, chromomycin, dactinomycin, daunorubicin, detrubicin, 6-diazo-5-oxo-L-norleucine, adriamycin (doxorubicin), morpholino-doxorubicin, cyanomorpholino-doxorubicin, 2-pyrrolino - Doxorubicin, deoxydoxorubicin, epirubicin, esorubicin, idarubicin, marcelomycin, mitomycin C and other mitomycins, mycophenolic acid, nogaramycin, olibomycin, peplomycin, potophyllomycin, puromycin, keramycin, rhodorubicin, streptonigrin, streptozocin, tubercidine, ubenimex, dinostatin, zorubicin; antimetabolites such as methotrexate and 5-fluorouracil (5-FU); folic acid analogs such as denopterin, pteropterin, and trimethrexate; f Purine analogs such as rudarabine, 6-mercaptopurine, thiamiprine, and thioguanine; pyrimidine analogs such as ancitabine, azacitidine, 6-azauridine, carmofur, cytarabine, dideoxyuridine, doxifluridine, enocitabine, and phloxuridine; androgens such as carsterone, dromostanolone propionate, epithiostanol, mepitiostane, and testolactone; anti-adrenal agents such as aminoglutethimide, mitotane, and trilostane; folic acid infusions such as fluphosphate; acegraton; aldofamide glycoside; aminolevulinic acid Acids; Enyluracil; Amsacrine; Bestrabusil; Bisanthren; Edatrexate; Defofamine; Demecolsin; Diadiquan; Elfomithine; Erliptinium acetate; Epotilon B and other Epotilons; Etoglucid; Gallium nitrate; Hydroxyurea; Lentinan; Mytansinoids such as Ronidynin, Mytansin and Ansamitosin; Mitoguazone; Mitoxantrone; Mopidamole; Nitracrine; Pentostatin; Fenamet; Pirarubicin; Rosoxantrone; Podophyllic acid;2-Ethylhydrazide; Procarbazine; PSK® Polysaccharide Complex (JHS Natural Products, Eugene, OR); Lazoxane; Rhizoxin; Schizophyllan; Spirogermanium; Tenuazonic Acid; Triadicone; 2,2',2''-Trichlorotriethylamine; Trichothecenes such as T-2 toxin, Veraculine A, Loridine A, and Anguidine; Urethane; Vindesine; Dacarbazine; Mannomustine; Mitobronitol; Mitractol; Pipobroman; Gacytosine; Arabinoside ("Ara-C"); Cyclophosphamide; Thiotepa; Taxoids, e.g., Taxol® (paclitaxel), Abraxane® (chromophore-free, albumin-modified nanoparticle formulation of paclitaxel), and Taxotere® (doxetaxel); Chlorambucil; Tamoxifen (Nolvadex®); Raloxifene; Aromatase inhibitory 4(5)-imidazole; 4-hydroxytamoxifen; Trioxyfen; Keoxyfen; LY 117018; Onapristone; Toremifene (Fareston®); Flutamide, Niltamide, Bicalutamide, Leuprolide, Goserelin; Chlorambucil; Gemzar® Gemcitabine; 6-Thiogunine; Mercaptopurine; Platinum analogs such as cisplatin, oxaliplatin, and carboplatin; Vinblastine; Platinum; Etoposide (VP-16); Ifosfamide; Mitoxantrone; Vincristine; Navelbine® (Vinorelbine); Novantrone; Teniposide; Edatrexate; Daunomycin; Aminopterin; Ibandronate; Irinotecan (e.g., CPT-11); Topoisomerase inhibitors RFS Examples include difluoromethylornithine (DMFO); retinoids such as retinoic acid; esperamicin; capecitabine (e.g., Xeloda®); and any pharmaceutically acceptable salt of any of the above.

[0272] Non-exclusive examples of additional anticancer drugs include trastuzumab (Herceptin®), bevacizumab (Avastin®), cetuximab (Erbitux®), rituximab (Rituxan®), Taxol®, Arimidex®, ABVD, Avisin, avagovomab, acridine carboxamide, adecatumumab, 17-N-allylamino-17-demethoxygeldanamycin, alfarazine, arbocidib, 3-aminopyridine-2-carboxaldehyde thiosemicarbazone, amonafide, anthracendione, anti-CD22 immunotoxin, antitumor drugs (e.g., cell cycle nonspecific antitumor drugs, and other antitumor drugs described herein), antitumor herbs, apadiquon, atiprimod, azathioprine, berotecan, bendamustine, and BIBW. 2992, Bilicodal, Brostarisin, Briostatin, Butionine sulfoximine, CBV (chemotherapy), Calyculine, Dichloroacetate, Discordamorid, Elsamitolu, Enocitabine, Eribulin, Exatecan, Exislind, Ferginol, Forodesine, Phosfestrol, ICE chemotherapy regimen, IT-101, Imexone, Imiquimod, Indocarbazole, Ilofluben, Lanikidal, Lalotaxel, Lenalidomide, Lucanton, Lulutotecan, Maphosfamide, Mitozolomide, Examples include napoxidine, nedaplatin, olaparib, ortataxel, PAC-1, pawpaw, pixantrone, proteasome inhibitors, rebeccamycin, reximod, rubitecan, SN-38, salinosporamide A, sapacitabine, stanford V, swainsonin, talaporfin, talikidal, tegafur-uracil, temodal, tesetaxel, triplatin tetranitrate, tris(2-chloroethyl)amine, troxacitabine, uramustine, bajimezan, vinflunin, ZD6126, and zoskidal.

[0273] Further non-exclusive examples of anticancer drugs include vinca alkaloids (e.g., vinblastine, vincristine, and vinorelbine), epidipodophyllotoxins (e.g., etoposide and teniposide), antibiotics (e.g., dactinomycin (actinomycin D), daunorubicin, and idarubicin), anthracyclines, mitoxantrone, bleomycin, plicamycin (mitramycin), mitomycin, and enzymes (e.g., enzymes that systemically metabolize L-asparagine to synthesize its own asparagine). L-asparaginase, which deprives cells of the ability to proliferate, antiplatelet agents, antiproliferative / antimitotic alkylating agents such as nitrogen mustard (e.g., mechloretamine, cyclophosphamide and its analogs, melphalan, and chlorambucil), ethyleneimine and methylmelamine (e.g., hexamethylmelamine and thiotepa), CDK inhibitors (e.g., CDK4 / 6 inhibitors such as abemaciclib, ribociclib, and palbociclib), sericiclib, UCN-01, P1446A-05, PD- Antiproliferative / antimitotic metabolites and related inhibitors (e.g., Mercadora, 0332991, Dynacyclib, P27-00, AT-7519, RGB286638, and SCH727965), alkyl sulfonates (e.g., busulfan), nitrosoureas (e.g., carmustine (BCNU) and its analogues, as well as streptozocin), trazeneth-dacarbadinine (DTIC), folic acid analogues, pyrimidine analogues (e.g., fluorouracil, floxuridine, and cytarabine), purine analogues, etc. Putopurine, thioguanine, pentostatin, and 2-chlorodeoxyadenosine), aromatase inhibitors (e.g., anastrozole, exemestane, and letrozole), and platinum-coordinated complexes (e.g., cisplatin and carboplatin), procarbazine, hydroxyurea, mitotane, aminoglutethimide, histone deacetylase (HDAC) inhibitors (e.g., trichostatin, sodium butyrate, apicidan, suberoylanilide hydroamic acid, vorinostat, belinostat, LBH589, romidepsin, ACY-1215, and panobinostat), mTOR inhibitors (e.g., vistocertib, temsirolimus, everolimus, ridafololimus, and sirolimus), KSP(Eg5) inhibitors (e.g., Array 520), DNA binding agents (e.g., Zalypsis®), PI3K inhibitors such as PI3K delta inhibitors (e.g., GS-1101 and TGR-1202), PI3K delta and gamma inhibitors (e.g., CAL-130), copanlisib, alpelisib, and idelalisib; multikinase inhibitors (e.g., TG02 and sorafenib), hormones (e.g., estrogen), and hormone agonists such as leutinizing hormone-releasing hormone (LHRH) agonists (e.g., goserelin, leuprolide, and triptorelin), BAFF neutralizing antibodies (e.g., LY2127399), IKK inhibitors, p38MAPK inhibitors, anti-IL-6 (e.g., CNT0328), telomerase inhibitors (e.g., GRN 163L), aurora kinase inhibitors (e.g., MLN8237), cell surface monoclonal antibodies (e.g., anti-CD38 (HUMAX-CD38)), anti-CS1 (e.g., elotuzumab), HSP90 inhibitors (e.g., 17AAG and KOS953), P13K / Akt inhibitors (e.g., perifosine), Akt inhibitors (e.g., GSK-2141795), PKC inhibitors (e.g., enzastaurin), FTIs (e.g., Za Examples include natural products such as rnestra (trademark), anti-CD138 (e.g., BT062), Torcl / 2-specific kinase inhibitors (e.g., INK128), ER / UPR targeting agents (e.g., MKC-3946), cFMS inhibitors (e.g., ARRY-382), JAK1 / 2 inhibitors (e.g., CYT387), PARP inhibitors (e.g., olaparib and veliparib (ABT-888)), and BCL-2 antagonists.

[0274] In some embodiments, the anticancer agent is selected from mechloretamine, camptothecin, ifosfamide, tamoxifen, raloxifene, gemcitabine, Navelbine®, sorafenib, or any analogue or derivative variant of the foregoing.

[0275] In some embodiments, the anticancer agent is a HER2 inhibitor. HER2 inhibitors are known in the art. Non-limiting examples of HER2 inhibitors include monoclonal antibodies, e.g., trastuzumab (Herceptin®) and pertuzumab (Perjeta®); small molecule tyrosine kinase inhibitors, e.g., gefitinib (Iressa®), erlotinib (Tarceva®), pyritinib, CP-654577, CP-724714, canertinib (CI 1033), HKI-272, lapatinib (GW-572016; Tykerb®), PKI-166, AEE788, BMS-599626, HKI-357, BIBW 2992, ARRY-334543, and JNJ-26483327.

[0276] In some embodiments, the anticancer agent is an ALK inhibitor. ALK inhibitors are known in the art. Non-limiting examples of ALK inhibitors include ceritinib, TAE-684 (NVP-TAE694), PF02341066 (crizotinib or 1066), alectinib, brigatinib, entrectinib, ensartinib (X-396), lorlatinib, ASP3026, CEP-37440, 4SC-203, TL-398, PLB1003, TSR-011, CT-707, TPX-0005, and AP26113. Additional examples of ALK kinase inhibitors are described in Examples 3-39 of WO05016894.

[0277] In some embodiments, the anticancer agent is an inhibitor of a downstream member of the receptor tyrosine kinase (RTK) / growth factor receptor (e.g., SHP2 inhibitors (e.g., SHP099, TNO155, RMC-4550, RMC-4630, JAB-3068, JAB-3312, RLY-1971, ERAS-601, SHP3809, PF-07284892, or BBP-398, or a pharmaceutically acceptable salt, solvate, isomer (e.g., stereoisomer), prodrug, or tautomer)). These include SOS1 inhibitors (e.g., BI-1701963, BI-3406, SDR5, BAY-293, or RMC-5845, or their pharmaceutically acceptable salts, solvates, isomers (e.g., stereoisomers), prodrugs, or tautomers), Raf inhibitors, MEK inhibitors, ERK inhibitors, PI3K inhibitors, PTEN inhibitors, AKT inhibitors, or mTOR inhibitors (e.g., mTORC1 inhibitors or mTORC2 inhibitors). In some embodiments, the anticancer agent is JAB-3312.

[0278] In some embodiments, the anticancer agent is an SOS1 inhibitor. SOS1 inhibitors are known in the art. In some embodiments, the SOS1 inhibitor is WO 2022146698, WO 2022081912, WO 2022058344, WO 2022026465, WO 2022017519, WO 2021173524, WO 2021130731, WO 2021127429, WO 2021092115, WO 2021105960, WO 2021074227, WO 2020180768, WO 2020180770, WO 2020173935, WO 2020146470, WO 2019201848, WO The compounds disclosed in 2019122129, WO 2018172250, and WO 2018115380, or selected from their pharmaceutically acceptable salts, solvates, isomers (e.g., stereoisomers), prodrugs, or tautomers. In some embodiments, the compounds of the present invention are K-Ras It is used in combination with SOS1 inhibitors to treat G13D cancer.

[0279] In some embodiments, the anticancer agent is a further Ras inhibitor or Ras vaccine, or another treatment designed to directly or indirectly reduce the oncogenic activity of Ras. Such agents are known in the art. In some embodiments, the anticancer agent is a further Ras inhibitor. In some embodiments, the Ras inhibitor targets Ras in its active or GTP-bound state. In some embodiments, the Ras inhibitor targets Ras in its inactive or GDP-bound state. In some embodiments, the Ras inhibitor is, for example, an inhibitor of K-Ras G12C, such as AMG 510 (sotrasib), MRTX1257, MRTX849 (adaglasib), JNJ-74699157, LY3499446, ARS-1620, ARS-853, BPI-421286, LY3537982, JDQ443, AZ4625, JAB-21822, JAB-21000, IBI351, ERAS-3490, RMC-6291, or GDC-6036, or a pharmaceutically acceptable salt, solvate, isomer (e.g., stereoisomer), prodrug, or tautomer thereof. In some embodiments, the Ras inhibitor is a K-Ras G12D inhibitor, e.g., MRTX1133 or JAB-22000, or a pharmaceutically acceptable salt, solvate, isomer (e.g., stereoisomer), prodrug, or tautomer thereof. In some embodiments, the Ras inhibitor is a K-Ras G12V inhibitor, e.g., JAB-23000, or a pharmaceutically acceptable salt, solvate, isomer (e.g., stereoisomer), prodrug, or tautomer thereof. In some embodiments, the Ras inhibitor is RMC-6236, or a pharmaceutically acceptable salt, solvate, isomer (e.g., stereoisomer), prodrug, or tautomer thereof. In some embodiments, the Ras inhibitor is: WO 2021091982, WO 2021091967, WO 2021091956, or WORas(ON) inhibitors disclosed in 2020132597 (the entirety of which is incorporated herein by reference), or selected from pharmaceutically acceptable salts, solvates, isomers (e.g., stereoisomers), prodrugs, or tautomers thereof. Other examples of Ras inhibitors are known in the art, for example: WO 20220133038, WO 2022133345, WO 2022132200, WO 2022119748, WO 2022109485, WO 2022109487, WO 2022066805, WO 2021190467, WO 2021185233, WO 2021180181, WO 2021175199, 2021173923, WO 2021169990, WO 2021169963, WO 2021168193, WO 2021158071, WO 2021155716, WO 2021152149, WO 2021150613, WO 2021147967, WO 2021147965, WO 2021143693, WO 2021142252, WO 2021141628, WO 2021139748, WO 2021139678, WO 2021129824, WO 2021129820, WO 2021127404, WO 2021126816, WO 2021126799, WO 2021124222, WO 2021121371, WO 2021121367, WO 2021121330, WO 2020050890, WO 2020047192, WO 2020035031, WO 2020028706, WO 2019241157, WO 2019232419, WO 2019217691, WO 2019217307, WO 2019215203, WO 2019213526, WO 2019213516, WO 2019155399, WO 2019150305, WO 2019110751, WO 2019099524, WO 2019051291, WO 2018218070, WO 2018217651, WO 2018218071, WO 2018218069, WO 2018206539, WO 2018143315, WO 2018140600, WO 2018140599, WO 2018140598,WO2018140514, WO 2018140513, WO 2018140512, WO 2018119183, WO 2018112420, WO 2018068017, WO 2018064510, WO 2017201161, WO 2017172979, WO 2017100546, WO 2017087528, WO 2017058807, WO 2017058805, WO 2017058728, WO 2017058902, WO 2017058792, WO 2017058768, WO Those found in 2017058915, WO 2017015562, WO 2016168540, WO 2016164675, WO 2016049568, WO 2016049524, WO 2015054572, WO 2014152588, WO 2014143659, and WO 2013155223 (all of which are incorporated herein by reference), or their pharmaceutically acceptable salts, solvates, isomers (e.g., stereoisomers), prodrugs, or tautomers.

[0280] In some embodiments, therapeutic agents that can be combined with the compounds of the present invention are inhibitors of the MAP kinase (MAPK) pathway (or "MAPK inhibitors"). Such agents are known in the art. Examples of MAPK inhibitors include, but are not limited to, one or more MAPK inhibitors described in Cancers (Basel) 2015 Sep;7(3):1758-1784. For example, MAPK inhibitors include trametinib, binimetinib, selumetinib, cobimetinib, LErafAON (NeoPharm), and ISIS. One or more of the following may be selected: 5132, vemurafenib, pimacertib, TAK733, RO4987655 (CH4987655), CI-1040, PD-0325901, CH5126766, MAP855, AZD6244, refametinib (RDEA 119 / BAY 86-9766), GDC-0973 / XL581, AZD8330 (ARRY-424704 / ARRY-704), RO5126766 (Roche, PLoS One. 2014 Nov 25;9(11)), and GSK1120212 (or JTP-74057, Clin Cancer Res. 2011 Mar 1;17(5):989-1000). MAPK inhibitors may include PLX8394, LXH254, GDC-5573, and LY3009120.

[0281] In some embodiments, the anticancer agent is a disruptor or inhibitor of the RAS-RAF-ERK, PI3K-AKT-TOR, or PI3K-AKT signaling pathway. Such agents are known in the art. Examples of PI3K / AKT inhibitors include, but are not limited to, one or more PI3K / AKT inhibitors described in Cancers (Basel) 2015 Sep;7(3):1758-1784. For example, the PI3K / AKT inhibitor may be selected from one or more of NVP-BEZ235, BGT226, XL765 / SAR245409, SF1126, GDC-0980, PI-103, PF-04691502, PKI-587, and GSK2126458.

[0282] In some embodiments, the anticancer agent is a PD-1 or PD-L1 antagonist. Such agents are known in the art. In some embodiments, additional therapeutic agents include ALK inhibitors, HER2 inhibitors, EGFR inhibitors, IGF-1R inhibitors, MEK inhibitors, PI3K inhibitors, AKT inhibitors, TOR inhibitors, MCL-1 inhibitors, BCL-2 inhibitors, SHP2 inhibitors, proteasome inhibitors, and immunotherapies. In some embodiments, additional therapeutic agents include FGFR inhibitors, PARP inhibitors, BET inhibitors, PRMT5i inhibitors, MAT2A inhibitors, VEGF inhibitors, and HDAC inhibitors. In some embodiments, the therapeutic agent may be a pan-RTK inhibitor, such as afatinib.

[0283] IGF-1R inhibitors are known in the art, and examples of IGF-1R inhibitors include lincitinib or its pharmaceutically acceptable salts. EGFR inhibitors are known in the art and include, but are not limited to, small molecule antagonists, antibody inhibitors, or specific antisense nucleotides or siRNAs. Useful antibody inhibitors of EGFR include cetuximab (Erbitux®), panitumumab (Vectibix®), zaltumumab, nimotuzumab, and matuzumab. Further antibody-based EGFR inhibitors include any anti-EGFR antibody or antibody fragment that can partially or completely block EGFR activation by its native ligand. Non-limiting examples of antibody-based EGFR inhibitors include Modjtahedi et al., Br.J. Cancer 1993, 67:247-253; Teramoto et al., Cancer Examples include those described in 1996,77:639-645; Goldstein et al., Clin. Cancer Res. 1995,1:1311-1318; Huang et al., 1999, Cancer Res. 15:59(8):1935-40; and Yang et al., Cancer Res. 1999,59:1236-1243. EGFR inhibitors may be monoclonal antibodies such as Mab E7.6.3 (Yang, 1999 above) or Mab C225 (ATCC accession number HB-8508), or antibodies or antibody fragments having binding specificity thereto.

[0284] Small molecule antagonists of EGFR include gefitinib (Iressa®), erlotinib (Tarceva®), and lapatinib (TykerB®). See, for example, Yan et al., Pharmacogenetics and Pharmacogenomics In Oncology Therapeutic Antibody Development, BioTechniques 2005, 39(4):565-8, and Paez et al., EGFR Mutations In Lung Cancer Correlation With Clinical Response To Gefitinib Therapy, Science 2004, 304(5676):1497-500. In some embodiments, the EGFR inhibitor is osimertinib (Tagrisso®). Further examples of the limited scope of small molecule EGFR inhibitors include any of the EGFR inhibitors described in the following patent publications, and any pharmaceutically acceptable salts of such EGFR inhibitors: EP0520722;EP0566226;WO96 / 33980;U.S. Patent No. 5,747,498;WO96 / 30347;EP0787772;WO97 / 30034;WO97 / 30044;WO97 / 38994;WO97 / 49688;EP837063;WO98 / 02434;WO97 / 38983;WO95 / 19774;WO95 / 19970;WO97 / 13771;WO98 / 02437;WO9 8 / 02438;WO97 / 32881;DE19629652;WO98 / 33798;WO97 / 32880;WO97 / 32880;EP68202 7;WO97 / 02266;WO97 / 27199;WO98 / 07726;WO97 / 34895;WO96 / 31510;WO98 / 14449;WO 98 / 14450; WO98 / 14451; WO95 / 09847; WO97 / 19065; WO98 / 17662; U.S. Patent No. 5,789,427; U.S. Patent No. 5,650,415; U.S. Patent No. 5,656,643; WO99 / 35146; WO99 / 35132; WO99 / 07701; and O92 / 20642.Additional non-limiting examples of small molecule EGFR inhibitors include any of the EGFR inhibitors described in Traxler et al., Exp. Opin.Ther. Patents 1998, 8(12):1599-1625. In some embodiments, EGFR inhibitors are ERBB inhibitors. In humans, the ERBB family includes HER1 (EGFR, ERBB1), HER2 (NEU, ERBB2), HER3 (ERBB3), and HER (ERBB4).

[0285] MEK inhibitors are known in the art and include, but are not limited to, pimacertib, selumetinib, cobimetinib (Cotellic®), trametinib (Mekinist®), and binimetinib (Mektovi®). In some embodiments, the MEK inhibitor targets a MEK mutation that is a class I MEK1 mutation selected from D67N, P124L, P124S, and L177V. In some embodiments, the MEK mutation is a class II MEK1 mutation selected from ΔE51-Q58, ΔF53-Q58, E203K, L177M, C121S, F53L, K57E, Q56P, and K57N.

[0286] PI3K inhibitors are known in the art, and include wartmannin, a 17-hydroxywartmannin analog described in WO06 / 044453, 4-[2-(1H-indazole-4-yl)-6-[[4-(methylsulfonyl)piperazine-1-yl]methyl]thieno[3,2-d]pyrimidine-4-yl]morpholine (also known as pictilisib or GDC-0941, described in WO09 / 036082 and WO09 / 055730), 2-methyl-2-[4-[3-methyl-2-oxo-8-(quinoline-3-yl)-2,3-dihydroimidazo[4,5-c]quinoline-1-yl]phenyl]propionitrile (BEZ 235 or NVP-BEZ Also known as 235 and described in WO06 / 122806), (S)-1-(4-((2-(2-aminopyrimidine-5-yl)-7-methyl-4-morpholinothieno[3,2-d]pyrimidine-6-yl)methyl)piperazin-1-yl)-2-hydroxypropan-1-one (described in WO08 / 070740), LY294002 (2-(4-morpholinyl)-8-phenyl-4H-1-benzopyran-4-one (available from Axon Medchem), PI 103 hydrochloride (3-[4-(4-morpholinylpyrido[3',2':4,5]fl[3,2-d]pyrimidine-2-yl]phenol hydrochloride (available from Axon Medchem), PIK 75(2-methyl-5-nitro-2-[(6-bromoimidazo[1,2-a]pyridine-3-yl)methylene]-1-methylhydrazide-benzenesulfonic acid, monohydrochloride) (available from Axon Medchem), PIK 90(N-(7,8-dimethoxy-2,3-dihydroimidazo[1,2-c]quinazolin-5-yl)-nicotinamide (available from Axon Medchem), AS-252424(5-[1-[5-(4-fluoro-2-hydroxyphenyl)-furan-2-yl]-meth-(Z)-ylidene]-thiazolidined-2,4-dione (available from Axon Medchem), TGX-221(7-methyl-2-(4-morpholinyl)-9-[1-(phenylamino)ethyl]-4H-pyrido-[1,2-a] Pyrinidine-4-one (available from Axon Medchem), XL-765, and XL-147 are examples, but are not limited to these. Other PI3K inhibitors include demethoxypyridine, perifosine, CAL101, PX-866, BEZ235, SF1126, INK1117, IPI-145, BKM120, XL147, XL765, Palomid, Examples include 529, GSK1059615, ZSTK474, PWT33597, IC87114, TGI 00-115, CAL263, PI-103, GNE-477, CUDC-907, and AEZS-136.

[0287] AKT inhibitors are known in the art, and examples of AKT inhibitors include Akt-1-1 (inhibits Akt1) (Barnett et al., Biochem.J.2005,385(Pt.2):399-408); Akt-1-1,2 (inhibits Ak1 and 2) (Barnett et al., Biochem.J.2005,385(Pt.2):399-408); API-59CJ-Ome (e.g., Jin et al., Br.J.Cancer 2004,91:1808-12); 1-H-imidazo[4,5-c]pyridinyl compounds (e.g., WO05 / 011700); indole-3-carbinol and its derivatives (e.g., U.S. Patent No. 6,656,963; Sarkar and Li J Nutr.2004,134(12)). Suppl):3493S-3498S); Perifosine (e.g., interferes with Akt membrane localization; Dasmahapatra et al. Clin. Cancer Res. 2004, 10(15):5242-52); Phosphatidylinositol ether lipid analogs (e.g., Gills and Dennis Expert.Opin.Investig.Drugs 2004,13:787-97); as well as trisilibine (TCN or API-2 or NCI discriminant: NSC 154020; Yang et al., Cancer Res. 2004,64:4394-9), but not limited to these.

[0288] mTOR inhibitors are known in the art, and these include ATP-competitive mTORC1 / mTORC2 inhibitors, such as PI-103, PP242, PP30; and Torin. 1; FKBP12 enhancers; 4H-1-benzopyran-4-one derivatives; and rapamycin (also known as sirolimus) and its derivatives, e.g., temsirolimus (Torisel®); everolimus (Afinitor®, WO94 / 09010); ridafololimus (also known as deforolimus or AP23573); rapalogs, e.g., those disclosed in WO98 / 02441 and WO01 / 14387, e.g., AP23464 and AP23841; 40-(2-hydroxyethyl)rapamycin; 40-[3-hydroxy(hydroxymethyl)methylpropanoate]-rapamycin (also known as CC1779); 40-epi-(tetrazolite)-rapamycin (also referred to as ABT578); 32-deoxo Rapamycin; 16-pentinyloxy-32(S)-dihydrolapanisin; derivatives disclosed in WO05 / 005434; U.S. Patents 5,258,389, 5,118,677, 5,118,678, 5,100,883, 5,151,413, 5,120,842, and 5,256,790, as well as WO94 / 090101, WO92 / 051 Examples include, but are not limited to, derivatives disclosed in 79, WO93 / 111130, WO94 / 02136, WO94 / 02485, WO95 / 14023, WO94 / 02136, WO95 / 16691, WO96 / 41807, WO96 / 41807, and WO2018 / 204416, as well as phosphorus-containing rapamycin derivatives (e.g., WO05 / 016252). In some embodiments, the mTOR inhibitor is a disteric inhibitor (see e.g., WO2018204416, WO2019212990, and WO2019212991), e.g., RMC-5552, having the following structure.

[0289] [ka]

[0290] BRAF inhibitors that can be used in combination with the compounds of the present invention are known in the art, and include, for example, vemurafenib, dabrafenib, and encorafenib. BRAF may include class 3 BRAF mutations. In some embodiments, class 3 BRAF mutations are selected from one or more of the following amino acid substitutions in human BRAF: D287H; P367R; V459L; G466V; G466E; G466A; S467L; G469E; N581S; N581I; D594N; D594G; D594A; D594H; F595L; G596D; G596R; and A762E.

[0291] MCL-1 inhibitors are known in the art and include, but are not limited to, AMG-176, MIK665, and S63845. The myeloid cell leukemia-1 (MCL-1) protein is one of the major anti-apoptotic members of the B-cell lymphoma-2 (BCL-2) protein family. Overexpression of MCL-1 is closely associated with tumor progression and resistance to targeted therapies, including BCL-2 inhibitors such as ABT-263, as well as to conventional chemotherapy.

[0292] In some embodiments, additional therapeutic agents are SHP2 inhibitors. SHP2 is known in the art. SHP2 is a non-receptor protein tyrosine phosphatase encoded by the PTPN11 gene and contributes to several cellular functions, including proliferation, differentiation, maintenance of the cell cycle, and migration. SHP2 has two N-terminal Src homology domains (N-SH2 and C-SH2), a catalytic domain (PTP), and a C-terminal tail. The two SH2 domains regulate the intracellular localization and functional regulation of SHP2. The molecule exists in an inactive, self-inhibitory conformation stabilized by a binding network involving residues from both the N-SH2 and PTP domains. For example, stimulation by cytokines or growth factors acting via receptor tyrosine kinases (RTKs) leads to exposure of the catalytic site, resulting in enzymatic activation of SHP2.

[0293] SHP2 is involved in signaling via the RAS mitogen-activated protein kinase (MAPK), JAK-STAT, or phosphoinositol 3-kinase-AKT pathways. Mutations in the PTPN11 gene and subsequent SHP2 mutations have been identified in several human developmental disorders, including Noonan syndrome and Leopard syndrome, as well as in human cancers such as juvenile myelomonocytic leukemia, neuroblastoma, melanoma, acute myeloid leukemia, breast cancer, lung cancer, and colon cancer. Some of these mutations destabilize the autoinhibitory conformation of SHP2, promoting SHP2 autoactivation or enhanced growth factor-driven activation. Therefore, SHP2 is a very attractive target for the development of novel therapies for the treatment of various diseases, including cancer. SHP2 inhibitors (e.g., RMC-4550 or SHP099) combined with RAS pathway inhibitors (e.g., MEK inhibitors) have been shown to inhibit the growth of several cancer cell lines (e.g., pancreatic cancer, lung cancer, ovarian cancer, and breast cancer) in vitro. Therefore, combination therapy using SHP2 inhibitors and RAS pathway inhibitors can be a common strategy for preventing tumor resistance in a wide range of malignancies.

[0294] Non-limiting examples of such SHP2 inhibitors are known in the art, including Chen et al. Mol Pharmacol. 2006, 70, 562; Sarver et al., J.Med.Chem. 2017, 62, 1793; Xie et al., J.Med.Chem. 2017, 60, 113734; and Igbe et al., Oncotarget, 2017, 8, 113734; as well as PCT applications incorporated herein by reference: WO 2022135568, WO 2021176072, WO 2021171261, WO 2021149817, WO 2021148010, WO 2021147879, WO 2021143823, WO 2021143701, WO 2021143680, WO 2021121397, WO 2021119525, WO 2021115286, WO 2021110796, WO 2021088945, WO 2021073439, WO 2021061706, WO 2021061515, WO 2021043077, WO 2021033153, WO 2021028362, WO 2021033153, WO 2021028362, WO 2021018287, WO 2020259679, WO 2020249079, WO 2020210384, WO 2020201991, WO 2020181283, WO 2020177653, WO 2020165734, WO 2020165733, WO 2020165732, WO 2020156243, WO 2020156242, WO 2020108590, WO 2020104635, WO 2020094104, WO 2020094018, WO 2020081848, WO 2020073949, WO 2020073945, WO 2020072656, WO 2020065453, WO 2020065452, WO 2020063760, WO 2020061103, WO 2020061101, WO 2020033828, WO 2020033286, WO 2020022323, WO 2019233810, WO 2019213318, WO 2019183367, WO 2019183364, WO 2019182960, WO 2019167000, WO 2019165073, WO 2019158019, WO 2019152454, WO 2019051469, WO 2019051084, WO 2018218133, WO 2018172984, WO 2018160731, WO 2018136265, WO 2018136264, WO 2018130928, WO 2018129402, WO 2018081091, WO 2018057884, WO 2018013597, WO 2017216706, WO 2017211303, WO 2017210134, WO 2017156397, WO 2017100279, WO 2017079723, WO 2017078499, WO 2016203406, WO 2016203405, WO 2016203404, WO 2016196591, WO 2016191328, WO 2015107495, WO 2015107494, WO 2015107493, WO 2014176488, WO Examples include US 2014113584, US 20210085677, US 10858359, US 10934302, US 10954243, US 10988466, US 11001561, US 11033547, US 11034705, or US 11044675, or their pharmaceutically acceptable salts, solvates, isomers (e.g., stereoisomers), prodrugs, or tautomers.

[0295] In some embodiments, the SHP2 inhibitor binds to the active site. In some embodiments, the SHP2 inhibitor is a mixed-type irreversible inhibitor. In some embodiments, the SHP2 inhibitor binds to an allosteric site, for example, a non-covalent allosteric inhibitor. In some embodiments, the SHP2 inhibitor is a covalent SHP2 inhibitor, such as an inhibitor that targets a cysteine ​​residue (C333) outside the active site of the phosphatase. In some embodiments, the SHP2 inhibitor is a reversible inhibitor. In some embodiments, the SHP2 inhibitor is an irreversible inhibitor. In some embodiments, the SHP2 inhibitor is SHP099. In some embodiments, the SHP2 inhibitor is TNO155 having the following structure.

[0296] [ka]

[0297] Alternatively, it may be a pharmaceutically acceptable salt, solvate, isomer (e.g., stereoisomer), prodrug, or tautomer thereof. In some embodiments, the SHP2 inhibitor is RMC-4550 having the following structure

[0298] [ka]

[0299] Alternatively, it may be a pharmaceutically acceptable salt, solvate, isomer (e.g., stereoisomer), prodrug, or tautomer thereof. In some embodiments, the SHP2 inhibitor is RMC-4630 having the following structure

[0300] [ka]

[0301] Alternatively, it may be a pharmaceutically acceptable salt, solvate, isomer (e.g., stereoisomer), prodrug, or tautomer thereof. In some embodiments, the SHP2 inhibitor is JAB-3068 having the following structure.

[0302] [ka]

[0303] Alternatively, it may be a pharmaceutically acceptable salt, solvate, isomer (e.g., stereoisomer), prodrug, or tautomer thereof. In some embodiments, the SHP2 inhibitor is JAB-3312. In some embodiments, the SHP2 inhibitor is the following compound

[0304] [ka]

[0305] Alternatively, it may be a pharmaceutically acceptable salt, solvate, isomer (e.g., stereoisomer), prodrug, or tautomer thereof. In some embodiments, the SHP2 inhibitor is RLY-1971 having the following structure.

[0306] [ka]

[0307] Alternatively, it may be a pharmaceutically acceptable salt, solvate, isomer (e.g., stereoisomer), prodrug, or tautomer. In some embodiments, the SHP2 inhibitor is ERAS-601, or a pharmaceutically acceptable salt, solvate, isomer (e.g., stereoisomer), prodrug, or tautomer. In some embodiments, the SHP2 inhibitor is BBP-398, or a pharmaceutically acceptable salt, solvate, isomer (e.g., stereoisomer), prodrug, or tautomer. In some embodiments, the SHP2 inhibitor is SH3809. In some embodiments, the SHP2 inhibitor is PF-07284892, or a pharmaceutically acceptable salt, solvate, isomer (e.g., stereoisomer), prodrug, or tautomer.

[0308] In some embodiments, additional therapeutic agents are selected from the group consisting of MEK inhibitors, HER2 inhibitors, SHP2 inhibitors, CDK4 / 6 inhibitors, mTOR inhibitors, SOS1 inhibitors, and PD-L1 inhibitors. See, for example, Hallin et al., Cancer Discovery, DOI:10.1158 / 2159-8290 (October 28, 2019) and Canon et al., Nature, 575:217 (2019). In some embodiments, the Ras inhibitor of the present invention is used in combination with a MEK inhibitor and an SOS1 inhibitor. In some embodiments, the Ras inhibitor of the present invention is used in combination with a PD-L1 inhibitor and an SOS1 inhibitor. In some embodiments, the Ras inhibitor of the present invention is used in combination with a PD-L1 inhibitor and an SHP2 inhibitor. In some embodiments, the Ras inhibitor of the present invention is used in combination with a MEK inhibitor and an SHP2 inhibitor. In some embodiments, the Ras inhibitor of the present invention is used in combination with an SHP2 inhibitor and a Ras inhibitor that inhibits multiple Ras isoforms and / or variants (e.g., RMC-6236). In some embodiments, the cancer is colorectal cancer, and the treatment comprises administering the Ras inhibitor of the present invention in combination with a second or third therapeutic agent (e.g., an SHP2 inhibitor and a Ras inhibitor that inhibits multiple Ras isoforms and / or variants). In some embodiments, the cancer is cholangiocarcinoma, and the treatment comprises administering the Ras inhibitor of the present invention, sorafenib, and a chemotherapeutic agent. In some embodiments, the cancer is gastric cancer, and the treatment comprises administering the Ras inhibitor of the present invention and an FGFR inhibitor (e.g., FGFR2i or FGFR4i). In some embodiments, the Ras inhibitor of the present invention is used in combination with immunotherapy and optionally with a chemotherapeutic agent.

[0309] Proteasome inhibitors are known in the art and include, but are not limited to, carfilzomib (Kyprolis®), bortezomib (Velcade®), and oprozomib.

[0310] Immunotherapy includes, but is not limited to, monoclonal antibodies, immunomodulatory imides (IMiDs), GITR agonists, genetically modified T cells (e.g., CAR-T cells), bispecific antibodies (e.g., BiTEs), and anti-PD-1, anti-PD-L1, anti-CTLA4, anti-LAG1, and anti-OX40 agents. Other immunotherapies are known in the art.

[0311] Immunomodulators (IMiDs) are a class of immunomodulatory drugs (drugs that modulate the immune response) that contain an imide group. The IMiD class includes thalidomide and its analogues (lenalidomide, pomalidomide, and apremilast).

[0312] Exemplary anti-PD-1 antibodies and their uses are described in Goldberg et al., Blood 2007, 110(1):186-192; Thompson et al., Clin. Cancer Res. 2007, 13(6):1757-1761; and WO06 / 121168 A1), and further described elsewhere in this specification.

[0313] FGFR inhibitors are known in the art and include FGFR2 inhibitors and FGFR4 inhibitors, such as pemigatinib and erdafitinib. See, for example, Cancers (Basel), 2021 Jun;13(12)2968.

[0314] BET inhibitors are known in the art, such as romidepsin, panobinostat, and bellinostat. See, for example, British J. Cancer 124:1478 (2021).

[0315] PRMT5i inhibitors are known in the art, such as PF-0693999, PJ-68, and MRTX1719. See, for example, Biomed.Pharmacotherapy 144:112252 (2021).

[0316] MAT2A inhibitors are known in the art, such as AG-270 and IDE397. See, for example, Exp Opin Ther Patents (2022) DOI:10.1080 / 13543776.2022.2119127.

[0317] GITR agonists are known in the art, and GITR agonists include GITR fusion proteins and anti-GITR antibodies (e.g., bivalent anti-GITR antibodies), for example, GITR fusion proteins described in U.S. Patent No. 6,111,090, No. 8,586,023, WO2010 / 003118, and WO2011 / 090754, or for example, U.S. Patent No. 7,025,962, EP1947183, U.S. Patent No. 7,812,135, No. 8,388, Examples of anti-GITR antibodies include, but are not limited to, those described in No. 967, No. 8,591,886, No. 7,618,632, EP1866339, and WO2011 / 028683, WO2013 / 039954, WO05 / 007190, WO07 / 133822, WO05 / 055808, WO99 / 40196, WO01 / 03720, WO99 / 20758, WO06 / 083289, WO05 / 115451, and WO2011 / 051726.

[0318] Another example of a therapeutic agent that may be used in combination with the compounds of the present invention is an anti-angiogenic agent. Anti-angiogenic agents are known in the art and include, but are not limited to, chemical compositions, antibodies, antigen-binding domains, radionuclides, and combinations and conjugates thereof, which are synthetically prepared in vitro. Anti-angiogenic agents may be agonists, antagonists, allosteric modulators, toxins, or, more generally, may act to inhibit or stimulate their targets (e.g., by activating or inhibiting receptors or enzymes), thereby promoting cell death or halting cell proliferation. In some embodiments, one or more additional therapies include an anti-angiogenic agent.

[0319] Anti-angiogenic agents may include MMP-2 (matrix-metalloproteinase 2) inhibitors, MMP-9 (matrix-metalloproteinase 9) inhibitors, and COX-II (cyclooxygenase 11) inhibitors. Non-limited examples of anti-angiogenic agents include rapamycin, temsirolimus (CCI-779), everolimus (RAD001), sorafenib, sunitinib, and bevacizumab. Examples of useful COX-II inhibitors include arecoxib, valdecoxib, and rofecoxib. Examples of useful matrix metalloproteinase inhibitors include WO96 / 33172, WO96 / 27583, WO98 / 07697, WO98 / 03516, WO98 / 34918, WO98 / 34915, WO98 / 33768, WO98 / 30566, WO90 / 05719, WO99 / 52910, and WO99 / 528. As described in 89, WO99 / 29667, WO99007675, EP0606046, EP0780386, EP1786785, EP1181017, EP0818442, EP1004578, and US20090012085, and U.S. Patents 5,863,949 and 5,861,510. Preferred MMP-2 and MMP-9 inhibitors are those with little to no activity to inhibit MMP-1. More preferred are those that selectively inhibit MMP-2 or AMP-9 compared to other matrix metalloproteinases (i.e., MAP-1, MMP-3, MMP-4, MMP-5, MMP-6, MMP-7, MMP-8, MMP-10, MMP-11, MMP-12, and MMP-13). Some specific examples of MMP inhibitors are AG-3340, RO 32-3555, and RS13-0830.

[0320] Further exemplary anti-angiogenic agents include KDR (kinase domain receptor) inhibitors (e.g., antibodies and antigen-binding regions that specifically bind to kinase domain receptors), anti-VEGF agents (e.g., VEGF (e.g., bevacizumab), or antibodies or antigen-binding regions that specifically bind to their soluble VEGF receptors or ligand-binding regions), e.g., VEGF-TRAP®, and anti-VEGF receptor agents (e.g., antibodies or antigen-binding regions that specifically bind to them), VEGF inhibitors, EGFR inhibitors (e.g., antibodies or antigen-binding regions that specifically bind to them), e.g., Vectibix® (panitumumab), erlotinib (Tarceva®), anti-Ang1 and anti-Ang2 agents (e.g., antibodies or antigen-binding regions that specifically bind to them or their receptors, e.g., Tie2 / Tek), and anti-Tie2 kinase inhibitors (e.g., antibodies or antigen-binding regions that specifically bind to them). Other anti-angiogenic agents include Canas, IL-8, B-FGF, Tek antagonists (US2003 / 0162712, US6,413,932), anti-Tweak agents (e.g., antibodies or antigen-binding domains that specifically bind, or soluble Tweak receptor antagonists, see US6,727,225), ADAM distointegrin domains that antagonize the binding of integrins to their ligands (US2002 / 0042368), and anti-eph receptors or anti-ephrin antibodies or antigen-binding domains that specifically bind. Examples include (U.S. Patents No. 5,981,245, No. 5,728,813, No. 5,969,110, No. 6,596,852, No. 6,232,447, No. 6,057,124, and their respective patent family members), anti-PDGF-BB antagonists (e.g., antibodies or antigen-binding regions that specifically bind to them), antibodies or antigen-binding regions that specifically bind to PDGF-BB ligands, and PDGFR kinase inhibitors (e.g., antibodies or antigen-binding regions that specifically bind to them).Additional anti-angiogenic agents include SD-7784 (Pfizer, USA), cilentide (Merck KGaA, Germany, EPO0770622), pegaptanib octasodium (Gilead Sciences, USA), alpha-statin (BioActa, UK), M-PGA (Celgene, USA, US5712291), ilomastat (Arriva, USA, US5892112), emaxanib (Pfizer, USA, US5792783), batalanib (Novartis, Switzerland), 2-methoxyestradiol (EntreMed, USA), TLC ELL-12 (Elan, Ireland), anecoltab acetate (Alcon, USA), and alpha-D148. Mab (Amgen, USA), CEP-7055 (Cephalon, USA), anti-Vn Mab (Crucell, Netherlands), DAC anti-angiogenic agent (ConjuChem, Canada), angiocidin (InKine). Pharmaceuticals, USA), KM-2550 (Kyowa Hakko, Japan), SU-0879 (Pfizer, USA), CGP-79787 (Novartis, Switzerland, EP0970070), ARGENT technology (Ariad, USA), YIGSR-Stealth (Johnson & Johnson, USA), Fibrinogen-E fragment (BioActa, UK), Angiogenesis inhibitor (Trigen, UK), TBC-1635 (Encysive Pharmaceuticals, USA), SC-236 (Pfizer, USA), ABT-567 (Abbott, USA), Metastatin (EntreMed, USA), Maspin (Sosei, Japan), 2-Methoxyestradiol (Oncology Sciences Corporation, USA), ER-68203-00 (IV AX, USA), BeneFin (Lane Labs, USA), Tz-93 (Tsumura, Japan), TAN-1120 (Takeda, Japan), FR-111142 (Fujisawa, Japan, JP02233610), Platelet Factor IV (RepliGen, USA, EP407122), Vascular Endothelial Growth Factor Antagonist (Borean, Denmark), Bevacizumab (pINN) (Genentech, USA), Angiogenesis Inhibitor (SUGEN, USA), XL 784 (Exelixis, USA), XL 647 (Exelixis, USA), MAb, Alpha-5 Beta-3 Integrin, Second Generation (Applied Molecular Evolution, USA and MedImmune, USA), Enzastaurin Hydrochloride (Lilly, USA), CEP 7055 (Cephalon, USA and Sanofi-Synthelabo, France), BC 1 (Genoa Institute of Cancer Research, Italy), rBPI 21 and BPI-derived anti-angiogenic agents (XOMA, USA), PI 88 (Progen, Australia), Silenditide (Merck KGaA, Germany, Munich Technical University, Germany, Scripps Clinic and Research Foundation)USA), AVE 8062 (Ajinomoto, Japan), AS 1404 (Cancer Research Laboratory, New Zealand), SG, 292 (Telios, USA), Endostatin (Boston Children's Hospital, USA), ATN 161 (Attenuon, USA), 2-Methoxyestradiol (Boston Children's Hospital, USA), ZD 6474 (AstraZeneca, UK), ZD 6126 (Angiogene Pharmaceuticals, UK), PPI 2458 (Praecis, USA), AZD 9935 (AstraZeneca, UK), AZD 2171 (AstraZeneca, UK), Vatalanib (pINN) (Novartis, Switzerland and Schering AG, Germany), Tissue Factor Pathway Inhibitor (EntreMed, USA), Pegaptanib (Pinn) (Gilead Sciences, USA), Xantrizole (Yonsei University, South Korea), Vaccine, Gene-based VEGF-2 (Scripps Clinic and Research) Foundation, USA), SPV5.2 (Supratek, Canada), SDX 103 (University of California at San Diego, USA), PX 478 (ProlX, USA), Metastatin (EntreMed, USA), Troponin I (Harvard University, USA), SU 6668 (SUGEN, USA), OXI 4503 (OXiGENE, USA), o-Guanidine (Dimensional Pharmaceuticals, USA), Motupolamine C (British Columbia University, Canada), CDP 791 (Celltech) Group, UK), atiprimod (pINN) (GlaxoSmithKline, UK), E 7820 (Eisai, Japan), CYC 381 (Harvard University, USA), AE 941 (Aeterna, Canada), vaccines, angiogenic agents (EntreMed, USA), urokinase plasminogen activator inhibitors (Dendreon, USA), ogluphanide (pINN) (Melmotte, USA), HIF-alfa inhibitors (Xenova, UK), CEP 5214 (Cephalon, USA), BAY RES 2622 (Bayer, Germany), angiocidin (InKine, USA), A6 (Angstrom, USA), KR 31372 (Korea Research Institute of Chemical Technology, South Korea), GW 2286 (GlaxoSmithKline, UK), EHT 0101 (ExonHit, France), CP 868596 (Pfizer, USA), CP 564959 (OSI, USA), CP 547632 (Pfizer, USA), 786034 (GlaxoSmithKline, UK), KRN 633 (Kirin Brewery, Japan), drug delivery system, intraocular, 2-methoxyestradiol, Anguinex (Maastricht University, Netherlands and Minnesota University, USA), ABT 510 (Abbott, USA), AAL 993 (Novartis, Switzerland), VEGI (ProteomTech, USA), tumor necrosis factor-alpha inhibitor, SU 11248 (Pfizer, USA and SUGEN, USA), ABT 518 (Abbott, USA), YH16 (Yantai Rongchang, China), S-3APG (Boston Children's Hospital, USA and EntreMed, USA), MAb, KDR (ImClone Systems, USA), MAb, alpha 5 beta (Protein Design, USA), KDR kinase inhibitor (Celltech Group, UK and Johnson & Johnson,(USA), GFB 116 (South Florida University, USA and Yale University, USA), CS 706 (Sankyo, Japan), Combretastatin A4 Prodrug (Arizona State University, USA), Chondroitinase AC (IBEX, Canada), BAY RES 2690 (Bayer, Germany), AGM 1470 (Harvard University, USA, Takeda, Japan, and TAP, USA), AG 13925 (Agouron, USA), Tetrathiomolybdate (University of Michigan, USA), GCS 100 (Wayne State University, USA), CV 247 (Ivy Medical, UK), CKD 732 (Chong Kun Dang, South Korea), Ilsogladine (Nippon Shinyaku, Japan), RG 13577 (Aventis, France), WX 360 (Wilex, Germany), Squalamine (Genaera, USA), RPI 4610 (Sirna, USA), Heparanase inhibitor (InSight, Israel), KL 3106 (Kolon, South Korea), Honokiol (Emory University, USA), ZK CDK (Schering AG, Germany), ZK Angio (Schering AG, Germany), ZK 229561 (Novartis, Switzerland and Schering AG, Germany), XMP 300 (XOMA, USA), VGA 1102 (Taisho, Japan), VE-Cadherin-2 antagonist (ImClone Systems, USA), Vasostatin (National Institutes of Health, USA), Flk-1 (ImClone Systems, USA), TZ 93 (Tsumura, Japan), Tamstatin (Beth Israel Hospital, USA), truncated soluble FLT 1 (vascular endothelial growth factor receptor 1) (Merck & Co, USA), Tie-2 ligand (Regeneron, USA, Examples include ), and thrombospongin 1 inhibitors (Allegheny Health, Education and Research Foundation, USA).

[0321] Further examples of therapeutic agents that may be used in combination with the compounds of the present invention include agents (e.g., antibodies, antigen-binding domains, or soluble receptors) that specifically bind to and inhibit the activity of growth factors, such as antagonists of hepatocyte growth factor (HGF, also known as scatter factor), as well as antibodies or antigen-binding domains that specifically bind to the receptor c-Met. Such agents are known in the art.

[0322] Another example of a therapeutic agent that may be used in combination with the compounds of the present invention is an autophagy inhibitor. Autophagy inhibitors are known in the art and include, but are not limited to, chloroquine, 3-methyladenine, hydroxychloroquine (Plaquenil®), bafilomycin A1, 5-amino-4-imidazole carboxamidriboside (AICAR), okadaic acid, autophagy-suppressing algal toxins that inhibit type 2A or type 1 protein phosphatases, cAMP analogs, and drugs that increase cAMP levels, such as adenosine, LY204002, N6-mercaptopurine riboside, and vinblastine. Furthermore, antisense or siRNAs that inhibit the expression of proteins, including but not limited to ATG5 (which is involved in autophagy), may also be used. In some embodiments, one or more additional therapies include an autophagy inhibitor.

[0323] Another example of therapeutic agents that can be used in combination with the compounds of the present invention is antineoplastic agents, which are known in the art. In some embodiments, one or more additional therapies include antineoplastic agents. Non-limiting examples of antineoplastic agents include acemannan, acralubicin, aldesleukin, alemtuzumab, alitretinoin, altretamine, amiphostine, aminolevulinic acid, amrubicin, amsacrine, anagrelide, anastrozole, ancer, ancestim, algravin, arsenic trioxide, BAM-002 (Novelos), bexarotene, bicalutamide, bromodeoxyuridine, capecitabine, cermoloukin, cetrorelix, cladribine, clotrimazole, cytarabine ocphosphate, and DA 3030 (Dong-A), daclizumab, denileukin difutox, deslorerin, dexrazoxane, dilazep, docetaxel, docosanol, doxelcalciferol, doxifluridine, doxorubicin, bromocriptine, carmustine, cytarabine, fluorouracil, HIT diclofenac, interferon alpha, daunorubicin, doxorubicin, tretinoin, ederfosine, edrecolomab eflornithine, emiteflu, epirubicin, epoetin beta, etoposide phosphate, exemestane, exislind, fadrozol, filgrastim, finasteride, fludarabine phosphate, formestan, fotemustine, gallium nitrate, gemcitabine, gemtuzumab zogamicin, gimeracil / oteracil / Gafour combination, glycopine, goserelin, heptaplatin, human chorionic gonadotropin, human fetal alpha-fetoprotein, ibandronic acid, idarubicin (imiquimod), interferon alpha, interferon alpha, natural type, interferon alpha-2, interferon alpha-2a, interferon alpha-2b, interferon alpha-NI, interferon alpha-n3, interferon alpha-con-1, interferon alpha, natural type, interferon beta, interferon beta-la, interferon beta-lb, interferon gamma, natural type interferon gamma-la, interferon gamma-lb, interleukin-1 beta, iobenguan,Irinotecan, Ilsogladine, Lanreotide, LC9018 (Yakult), Leflunomide, Lenograstim, Lentinan sulfate, Letrozole, Leukocyte alpha interferon, Leuprorelin, Levamizole + Fluorouracil, Rialozol, Lovaplatin, Ronidamin, Lovastatin, Masopropyl, Melalsoprole, Metoclopramide, Mifepristone, Miltefosine, Millimostim, Mispaired double-stranded RNA, Mitoguazone, Mitractol, Mitoxantrone, Morglamostim, Nafarelin, Naloxone + Pentazocine, Naltgrastim, Nedaplatin, Niltamide, Noscapine, Novel Erythropoiesis-Promoting Protein, NSC631570 Octreotide, Oprelbequin, Osateron, Oxaliplatin, Paclitaxel, Pamidronic Acid, Pegaspargase, Peginterferon Alpha-2b, Pentosan, Sodium Polysulfate, Pentostatin, Picibanil, Pirarubicin, Rabbit Antithymocyte Polyclonal Antibody, Polyethylene Glycol Interferon Alpha-2a, Porfimer Sodium, Raloxifene, Larcitrexed, Rasbrien Bodymen Rasburiembodiment, rhenium etidronate Re186, RII retinamide, rituximab, romultide, samarium (153Sm) lexidonam, salglamostim, schizophyllan, sobuzoxane, sonelmin, strontium-89 chloride, suramin, tasonelmin, tazarotene, tegafur, temoporfin, temozolomide, teniposide, tetrachlorodecaoxide, thalidomide, thymalfacin, thyroid-stimulating hormone alpha, topotecan, toremifene, tositumomab-iodine-131, trastuzumab, treosulf Avan, tretinoin, trilostane, trimethrexate, triptorelin, tumor necrosis factor alpha, natural type, ubenimex, bladder cancer vaccine, Maruyama vaccine, melanoma solubilizing solution vaccine, barrubicin, verteporfin, vinorelbine, bilirulysine, dinostatin stimulamer or zoledronic acid; Abarelix; AE941 (Aeterna), ambamustin, antisense oligonucleotide, bcl-2 (Genta), APC8015 (Dendreon), decitabine, dexaaminoglutethimide, diazicon, EL532 (Elan),EM800 (Endorecherche), eniluracil, etanidazole, fenretinide, filgrastim SD01 (Amgen), fulvestrant, gallocitabine, gastrin-17 immunogene, HLA-B7 gene therapy (Vical), granulocyte-macrophage colony-stimulating factor, histamine dihydrochloride, ibritumomab tiuxetan, ilomastat, IM862 (Cytran), interleukin-2, iproxyfen, LDI200 (Milkhaus), religistim, lintuzumab, CA125 MAb (Biomira), cancer MAb (Japan Pharmaceutical Development), HER-2 and Fc MAb (Medarex), idiotype 105AD7 MAb (CRC Technology), idiotype CEA MAb (Trilex), LYM-1-iodine-131MAb (Techniclone), polymorphoemic mucin-yttrium-90MAb (Antisoma), marimast, menogalil, mitumomab, motexafine, gadolinium, MX6 (Galderma), nelarabine, noratexed, P30 protein, pegvisomant, pemetrexed, porphyromycin, prinomast, RL0903 (Shire), rubitecan, satoraplatin, sodium phenylacetate, sparphosic acid, SRL172 (SR Pharma), SU5416 (SUGEN), TA077 (Tanabe), tetrathiomolybdate, saliblastin, thrombopoietin, tin ethylethiopurine, tirapazamine, cancer vaccine (Biomira), melanoma vaccine (New York University), melanoma vaccine (Sloan Kettering Examples include the melanoma tumor lysis product vaccine (New York Medical College), the viral melanoma cell solubilization vaccine (Royal Newcastle Hospital), or Valspodar.

[0324] Additional examples of therapeutic agents that may be used in combination with the compounds of the present invention include ipilimumab (Yervoy®); tremelimumab; galiximab; nivolumab, also known as BMS-936558 (Opdivo®); pembrolizumab (Keytruda®); avelumab (Bavencio®); AMP224; BMS-936559; MPDL3280A, also known as RG7446; MEDI-570; AMG557; MGA271; IMP321; BMS-663513; PF-05082566; CDX-1127; anti-OX40 (Providence Health Services); huMAbOX40L; Atasicept; CP-870893; Lucatumumab; Dasetuzumab; Muromonab-CD3; Ipirumumab; MEDI4736 (Imfinzi (registered trademark)); MSB0010718C; AMP 224; Adalimumab (Humira®); Adtrastuzumab emtansine (Kadcyla®); Aflibercept (Eylea®); Alemtuzumab (Campath®); Basiliximab (Simulect®); Belimumab (Benlysta®); Basiliximab (Simulect®); Belimumab (Benlysta®); Brentuximab vedotin (Adcetris®); Canakinumab (Ilaris®); Certolizumab pegol (Cimzia®); Daclizumab (Zenapax®); Daratumumab (Darzale x(registered trademark)); denosumab (Prolia(registered trademark)); eculizumab (Soliris(registered trademark)); efalizumab (Raptiva(registered trademark)); gemtuzumab ozogamicin (Mylotarg(registered trademark)); golimumab (Simponi(registered trademark)); ibritumomab tiuxetan (Zevalin(registered trademark)); infliximab (Remicade(registered trademark)); motabizumab (Numax(registered trademark)); natalizumab (Tysabri(registered trademark)); obinutuzumab (Gazyva(registered trademark)); ofatumumab (Arzerra(registered trademark)); omalizumab (Xolair(registered trademark)); palivizumab (Synagis(registered trademark));Pertuzumab (Perjeta®); Pertuzumab (Perjeta®); Ranibizumab (Lucentis®); Laxibakumab (Abthrax®); Tocilizumab (Actemra®); Tositumomab; Tositumomab-i-131; Tositumomab and Tositumomab-i-131 (Bexxar®); Ustekinumab (Stelara®); AMG 102; AMG; Examples include the 386; AMG 479; AMG 655; AMG 706; AMG 745; and AMG 951.

[0325] The compounds described herein can be used in combination with other agents disclosed herein or other suitable agents, depending on the condition being treated. Therefore, in some embodiments, one or more compounds of the present invention are administered concurrently with other therapies, such as those described herein. When used in combination therapy, the compounds described herein may be administered concurrently with or separately from a second agent. This combined administration may include concurrent administration of the two agents in the same dosage form, concurrent administration in separate dosage forms, and separate administration. That is, the compounds described herein and any agents described herein may be formulated together in the same dosage form and administered concurrently. Alternatively, the compounds of the present invention and any therapy described herein may be administered concurrently, with both agents existing in separate formulations. In another alternative, the compounds of the present invention may be administered, followed by any therapy described herein, and vice versa. In some embodiments of separate administration protocols, the compounds of the present invention and any therapy described herein are administered at intervals of several minutes, several hours, or several days.

[0326] In some embodiments of any of the methods described herein, the first therapy (e.g., the compound of the present invention) and one or more additional therapies are administered simultaneously or sequentially in any order. The first therapeutic agent is administered immediately before or after one or more additional therapies, or up to 1 hour, 2 hours, 3 hours, 4 hours, 5 hours, 6 hours, 7 hours, 8 hours, 9 hours, 10 hours, 11 hours, 12 hours, 13 hours, 14 hours, 16 hours, 17 hours, 18 hours, 19 hours, 20 hours, 21 hours, 22 hours, 23 hours, 24 hours, or up to 1-7, 1-14, 1-21, or 1-30 days before or after.

[0327] The present invention also features a kit comprising (a) a pharmaceutical composition comprising an agent described herein (e.g., a compound of the present invention), and (b) a package containing instructions for carrying out any of the methods described herein. In some embodiments, the kit comprises (a) a pharmaceutical composition comprising an agent described herein (e.g., a compound of the present invention), (b) one or more additional therapies (e.g., non-pharmacological treatments or therapeutic agents), and (c) a package containing instructions for carrying out any of the methods described herein.

[0328] One aspect of the present invention, intending to treat a disease or associated symptoms using a combination of pharmaceutically active compounds that can be administered separately, further relates to combining separate pharmaceutical compositions in the form of a kit. The kit may comprise two separate pharmaceutical compositions, namely the compounds of the present invention, and one or more additional therapies. The kit may comprise containers for housing the separate compositions, such as divided bottles or divided foil packets. Examples of additional containers include syringes, boxes, and bags. In some embodiments, the kit may include instructions for using the separate components. The kit form is particularly advantageous when the separate components are preferably administered in different dosage forms (e.g., oral and parenteral), administered at different dosing intervals, or when titration of the individual components of the combination is desired by the prescribing healthcare professional.

[0329] Numbered embodiments Embodiment 1. A compound having the structure of formula I, or a pharmaceutically acceptable salt thereof:

[0330] [ka]

[0331] [In the formula, A is an optionally substituted 3-6 member heterocycloalkylene, an optionally substituted 3-6 member cycloalkylene, an optionally substituted 6 member arylene, or an optionally substituted 5-10 member heteroarylene.] X 1 , X 2 , and X 3 Each of these is independently selected from CH2, CHF, CF2, C=O, or O. m is either 1 or 2. n is either 0 or 1. R 1 is hydrogen, optionally substituted C1-C6 heteroalkyl, or optionally substituted 3-10 member heterocycloalkyl. R 2 is an arbitrarily substituted C1-C6 alkyl, and R 3 This is an optionally substituted C1-C6 alkyl, an optionally substituted C1-C6 heteroalkyl, an optionally substituted 3- to 6-membered cycloalkyl, or an optionally substituted heterocycloalkyl. Each hydrogen atom is independently an arbitrarily isotopically enriched deuterium.

[0332] Embodiment 2. A compound having the structure of formula I, or a pharmaceutically acceptable salt thereof:

[0333] [ka]

[0334] [In the formula, A is an optionally substituted 3-6 member heterocycloalkylene, an optionally substituted 3-6 member cycloalkylene, an optionally substituted 6 member arylene, or an optionally substituted 5-10 member heteroarylene.] X 1 , X 2 , and X 3 Each of these is independently selected from CH2, CF2, C=O, or O. m is either 1 or 2. n is either 0 or 1. R 1 is hydrogen, optionally substituted C1-C6 heteroalkyl, or optionally substituted 3-10 member heterocycloalkyl. R 2 is an arbitrarily substituted C1-C6 alkyl, and R 3 This is an optionally substituted C1-C6 alkyl group, an optionally substituted 3-6 member cycloalkyl group, or an optionally substituted heterocycloalkyl group. Each hydrogen atom is independently an arbitrarily isotopically enriched deuterium.

[0335] Embodiment 3. A compound according to Embodiment 1 or 2, or a pharmaceutically acceptable salt thereof, having the structure of any one of formulas Ia, Ib, or Ic:

[0336] [ka]

[0337] [In the formula, each D represents hydrogen having a deuterium isotope enrichment factor of at least 5.] Embodiment 4. R 1 The compound according to any one of Embodiments 1 to 3, or a pharmaceutically acceptable salt thereof, wherein is a hydrogen atom or an optionally substituted 3- to 10-membered heterocycloalkyl group.

[0338] Embodiment 5. R 1The compound according to any one of Embodiments 1 to 4, or a pharmaceutically acceptable salt thereof, wherein is an optionally substituted 3- to 10-membered heterocycloalkyl group. Embodiment 6. R 1 teeth,

[0339] [ka]

[0340] The compound described in Embodiment 5, or a pharmaceutically acceptable salt thereof. Embodiment 7. R 1 teeth,

[0341] [ka]

[0342] The compound according to Embodiment 5, or a pharmaceutically acceptable salt thereof, wherein each D represents hydrogen having a deuterium isotope enrichment factor of at least 5. Embodiment 8. A compound according to any one of Embodiments 1 to 7, or a pharmaceutically acceptable salt thereof, wherein m is 1.

[0343] Embodiment 9. A compound according to any one of Embodiments 1 to 8, or a pharmaceutically acceptable salt thereof, wherein n is 1. Embodiment 10. X 1 , X 2 , and X 3 Each of the compounds described in any one of Embodiments 1 to 9, or a pharmaceutically acceptable salt thereof, wherein each of the compounds is CH2.

[0344] Embodiment 11. The compound described in Embodiment 1, having the structure of Formula II, or a pharmaceutically acceptable salt thereof.

[0345] [ka]

[0346] Embodiment 12. The compound described in Embodiment 1, having the structure of formula V, or a pharmaceutically acceptable salt thereof.

[0347] [ka]

[0348] Embodiment 13. A compound according to Embodiment 12, having the structure of formula Va, formula Vb, or formula Vc, or a pharmaceutically acceptable salt thereof:

[0349] [ka]

[0350] [In the formula, each D represents hydrogen having a deuterium isotope enrichment factor of at least 5.] Embodiment 14. A compound according to Embodiment 12, or a pharmaceutically acceptable salt thereof, having the structure of any one of the following: formula Vd, formula Ve, or formula Vf:

[0351] [ka]

[0352] [In the formula, each D represents hydrogen having a deuterium isotope enrichment factor of at least 5.] Embodiment 15. The compound described in Embodiment 1, having the structure of formula VI, or a pharmaceutically acceptable salt thereof.

[0353] [ka]

[0354] Embodiment 16. The compound described in Embodiment 1, having the structure of formula VII, or a pharmaceutically acceptable salt thereof.

[0355] [ka]

[0356] Embodiment 17. A compound according to any one of Embodiments 1 to 16, or a pharmaceutically acceptable salt thereof, wherein A is optionally substituted thiazole-diyl, optionally substituted oxazole-diyl, optionally substituted morpholine-diyl, optionally substituted pyrrolidine-diyl, optionally substituted piperidine-diyl, or optionally substituted phenylene.

[0357] Embodiment 18. The compound described in Embodiment 17, or a pharmaceutically acceptable salt thereof, wherein A is optionally substituted thiazole-diyl or optionally substituted morpholine-diyl.

[0358] Embodiment 19. A is an optionally substituted 5- to 10-membered heteroarylene, the compound according to any one of Embodiments 1 to 16, or a pharmaceutically acceptable salt thereof. Embodiment 20. A is,

[0359] [ka]

[0360] The compound described in Embodiment 19, or a pharmaceutically acceptable salt thereof. Embodiment 21. A is,

[0361] [ka]

[0362] The compound described in Embodiment 20, or a pharmaceutically acceptable salt thereof. Embodiment 22. A is a compound according to any one of Embodiments 1 to 16, or a pharmaceutically acceptable salt thereof, wherein A is optionally substituted phenylene.

[0363] Embodiment 23. A is,

[0364] [ka]

[0365] The compound described in Embodiment 22, or a pharmaceutically acceptable salt thereof. Embodiment 24. A is,

[0366] [ka]

[0367] The compound described in Embodiment 23, or a pharmaceutically acceptable salt thereof. Embodiment 25. A is an optionally substituted 3- to 6-membered heterocycloalkylene, the compound according to any one of Embodiments 1 to 16, or a pharmaceutically acceptable salt thereof.

[0368] Embodiment 26. The compound described in Embodiment 25, or a pharmaceutically acceptable salt thereof, wherein A is an optionally substituted six-membered heterocycloalkylene. Embodiment 27. A is one of the following, or a compound described in Embodiment 25, selected from these stereoisomers, or a pharmaceutically acceptable salt thereof:

[0369] [ka]

[0370] Embodiment 28. A is one of the following, or a compound described in Embodiment 26, selected from these stereoisomers, or a pharmaceutically acceptable salt thereof:

[0371] [ka]

[0372] Embodiment 29. R 2 teeth,

[0373] [ka]

[0374] The compound described in any one of Embodiments 1 to 28, or a pharmaceutically acceptable salt thereof. Embodiment 30. R 2 teeth,

[0375] [ka]

[0376] The compound according to any one of Embodiments 1 to 28, or a pharmaceutically acceptable salt thereof, wherein each D represents hydrogen having a deuterium isotope enrichment factor of at least 5. Embodiment 31. R 3 The compound according to any one of Embodiments 1 to 30, or a pharmaceutically acceptable salt thereof, wherein is an optionally substituted C1-C6 alkyl or an optionally substituted 3- to 6-membered cycloalkyl.

[0377] Embodiment 32. R 3 The compound according to any one of Embodiments 1 to 31, or a pharmaceutically acceptable salt thereof, wherein is an optionally substituted C1-C6 alkyl. Embodiment 33. R 3 teeth,

[0378] [ka]

[0379] The compound described in Embodiment 32, or a pharmaceutically acceptable salt thereof. Embodiment 34. R 3 teeth,

[0380] [ka]

[0381] The compound described in Embodiment 33, or a pharmaceutically acceptable salt thereof. Embodiment 35. R 3 teeth,

[0382] [ka]

[0383] The compound according to Embodiment 32, or a pharmaceutically acceptable salt thereof, wherein each D represents hydrogen having a deuterium isotope enrichment factor of at least 5. Embodiment 36. R 3 The compound according to any one of Embodiments 1 to 31, or a pharmaceutically acceptable salt thereof, wherein is an optionally substituted 3- to 6-membered cycloalkyl group.

[0384] Embodiment 37. R 3 teeth,

[0385] [ka]

[0386] The compound described in Embodiment 36, or a pharmaceutically acceptable salt thereof. Embodiment 38. R 3 The compound according to Embodiment 36, or a pharmaceutically acceptable salt thereof, wherein is an optionally substituted five-membered cycloalkyl group.

[0387] Embodiment 39. R 3 teeth,

[0388] [ka]

[0389] The compound described in Embodiment 38, or a pharmaceutically acceptable salt thereof. Embodiment 40. R 2 teeth,

[0390] [ka]

[0391] And, R 3 teeth,

[0392] [ka]

[0393] and A is

[0394] [ka]

[0395] The compound described in any one of embodiments 11 to 16, or a pharmaceutically acceptable salt thereof. Embodiment 41.

[0396] R 2 teeth,

[0397] [ka]

[0398] And, R 3 teeth,

[0399] [ka]

[0400] and A is

[0401] [ka]

[0402] The compound according to any one of embodiments 11 to 16. Embodiment 42. The compound is a compound described in any one of Embodiments 1 to 41, other than the compounds listed in Table 3, or a pharmaceutically acceptable salt thereof.

[0403] Embodiment 43. A compound having the structure of a compound described in Table 1 or Table 2, or a pharmaceutically acceptable salt thereof. Embodiment 44. A pharmaceutical composition comprising a compound described in any one of Embodiments 1 to 43, or a pharmaceutically acceptable salt thereof, and a pharmaceutically acceptable excipient.

[0404] Embodiment 45. A method for treating cancer in a subject requiring treatment, comprising administering to the subject a therapeutically effective amount of a compound described in any one of Embodiments 1 to 43, or a pharmaceutically acceptable salt thereof, or a pharmaceutical composition described in Embodiment 44.

[0405] Embodiment 46. The method according to Embodiment 45, wherein the cancer is pancreatic cancer, non-small cell lung cancer, colorectal cancer, or endometrial cancer. Embodiment 47. The method according to Embodiment 45 or 46, wherein the cancer is a Ras mutation.

[0406] Embodiment 48. The method according to Embodiment 47, wherein the Ras mutation is K-Ras G12D or K-Ras G13D. Embodiment 49. A method for treating a Ras protein-related disease in a subject requiring treatment, the method comprising administering to the subject a therapeutically effective amount of a compound described in any one of Embodiments 1 to 43, or a pharmaceutically acceptable salt thereof, or a pharmaceutical composition described in Embodiment 44.

[0407] Embodiment 50. A method for inhibiting Ras protein in cells, comprising contacting the cells with an effective amount of a compound described in any one of Embodiments 1 to 43, or a pharmaceutically acceptable salt thereof, or the pharmaceutical composition described in Embodiment 44.

[0408] Embodiment 51. The method according to Embodiment 49 or 50, wherein the Ras protein is K-Ras G12D or K-Ras G13D. Embodiment 52. The method according to Embodiment 50 or 51, wherein the cells are cancer cells.

[0409] Embodiment 53. The method according to Embodiment 52, wherein the cancer cells are pancreatic cancer cells, non-small cell lung cancer cells, colorectal cancer cells, or endometrial cells. Embodiment 54. The method or use according to any one of Embodiments 45 to 53, further comprising administering an additional anti-cancer treatment.

[0410] Embodiment 55. The method according to Embodiment 54, wherein the additional anticancer treatment described above is an EGFR inhibitor, a second Ras inhibitor, a SHP2 inhibitor, a SOS1 inhibitor, a Raf inhibitor, a MEK inhibitor, an ERK inhibitor, a PI3K inhibitor, a PTEN inhibitor, an AKT inhibitor, an mTORC1 inhibitor, a BRAF inhibitor, a PD-L1 inhibitor, a PD-1 inhibitor, a CDK4 / 6 inhibitor, a HER2 inhibitor, or a combination thereof.

[0411] Embodiment 56. The method according to Embodiment 54 or 55, wherein the additional anti-cancer treatment is an SHP2 inhibitor. Embodiment 57. A conjugate containing the structure of Formula III, or a salt thereof: MP 1 Formula III [In the formula, P 1 is a monovalent organic part, and M has the structure of equation IV:

[0412] [ka]

[0413] [In the formula, A is an optionally substituted 3-6 member heterocycloalkylene, an optionally substituted 3-6 member cycloalkylene, an optionally substituted 6 member arylene, or an optionally substituted 5-10 member heteroarylene.] X 1 , X 2 , and X 3 Each of these is independently selected from CH2, CHF, CF2, C=O, or O. m is either 1 or 2. n is either 0 or 1. R 1 is hydrogen, optionally substituted C1-C6 heteroalkyl, or optionally substituted 3-10 member heterocycloalkyl. R 2 is an arbitrarily substituted C1-C6 alkyl, and R 3 This is an optionally substituted C1-C6 alkyl, an optionally substituted 3-6 member cycloalkyl, an optionally substituted C1-C6 heteroalkyl, or an optionally substituted heterocycloalkyl. Each hydrogen in Equation IV is independently an arbitrarily isotopically enriched deuterium.

[0414] Embodiment 58. The conjugate or salt thereof according to Embodiment 57, wherein A is optionally substituted thiazole-diyl, optionally substituted oxazole-diyl, optionally substituted morpholine-diyl, optionally substituted pyrrolidine-diyl, optionally substituted piperidine-diyl, or optionally substituted phenylene.

[0415] Embodiment 59. R 1 teeth,

[0416] [ka]

[0417] The conjugate or salt thereof according to embodiment 57 or 58. Embodiment 60. m is 1, n is 1, X 1 , X 2 , and X 3 Each of the components is CH2, the conjugate or salt thereof according to any one of embodiments 57 to 59.

[0418] Embodiment 61. The conjugate according to any one of Embodiments 57 to 60, or a salt thereof, wherein the monovalent organic portion is a protein. Embodiment 62. The conjugate or salt thereof according to Embodiment 61, wherein the protein is a Ras protein.

[0419] Embodiment 63. The conjugate or salt thereof according to Embodiment 62, wherein the Ras protein is K-Ras G12D or K-Ras G13D. Embodiment 64. A conjugate according to any one of Embodiments 57 to 63, wherein M is bound to an amino acid residue of the monovalent organic moiety.

[0420] Examples The present invention is further illustrated by the following examples and synthesis examples, which should not be considered to limit the scope or spirit of the invention to the specific procedures described herein. It should be understood that the examples are provided to illustrate specific embodiments and are not intended to impose any limitation on the scope of the invention. It should be further understood that various other embodiments, modifications, and equivalents can be taken, which themselves may be suggested to those skilled in the art without departing from the spirit of the invention or the appended claims.

[0421] chemical synthesis The following examples and definitions used elsewhere in this specification are as follows: JPEG2026048800000113.jpg97170

[0422] Synthesis of intermediates Synthesis of intermediate 1:3-(5-bromo-1-ethyl-2-[2-[(1S)-1-methoxyethyl]pyridine-3-yl]indole-3-yl)-2,2-dimethylpropan-1-ol

[0423] [ka]

[0424] Step 1: Synthesis of 1-(5-bromo-1H-indole-3-yl)-3-((tert-butyldiphenylsilyl)oxy)-2,2-dimethylpropan-1-one Under an N2 atmosphere at 0°C, 3-((tert-butyldiphenylsilyl)oxy)-2,2-dimethylpropanoyl chloride (65 g, 137 mmol, crude) was slowly added to a mixture of DCM (120 mL) with 1 M SnCl4 in DCM (137 mL, 137 mmol). After stirring the mixture at 0°C for 30 minutes, a solution of 5-bromo-1H-indole (26.8 g, 137 mmol) in DCM (40 mL) was added dropwise. After stirring the mixture at 0°C for 45 minutes, it was diluted with siRNA (300 mL), washed with brine (4 × 100 mL), dried over Na2SO4, and filtered. The filtrate was concentrated under reduced pressure, and the residue was purified by silica gel column chromatography to obtain the product (55 g, 75% yield). LCMS(ESI) m / z:[M+Na]C 29 H 32 Calculated value for BrNO2SiNa: 556.1; measured value: 556.3.

[0425] Step 2: Synthesis of 1-(5-bromo-1H-indole-3-yl)-3-((tert-butyldiphenylsilyl)oxy)-2,2-dimethylpropan-1-one Under an N2 atmosphere at 0°C, LiBH4 (6.1 g, 281 mmol) was added to a mixture of 1-(5-bromo-1H-indole-3-yl)-3-((tert-butyldiphenylsilyl)oxy)-2,2-dimethylpropan-1-one (50 g, 93.6 mmol) with THF (100 mL). The mixture was heated to 60°C and stirred for 20 hours. Then, MeOH (10 mL) and siRNA (100 mL) were added. The mixture was washed with brine (50 mL), dried over Na2SO4, filtered, and the filtrate was concentrated under reduced pressure. The residue was diluted with DCM (50 mL), cooled to 10°C, and zirzine (9.5 g, 37.4 mmol) and TsOH·H2O (890 mg, 4.7 mmol) were added. The mixture was stirred at 10°C for 2 hours and filtered. The filtrate was concentrated under reduced pressure, and the residue was purified by silica gel column chromatography to obtain the product (41 g, 84% yield). LCMS(ESI) m / z:[M+H]C 29 H 34 Calculated value for BrNOSi: 519.2; measured value: 520.1.

[0426] Step 3: Synthesis of 5-bromo-3-(3-((tert-butyldiphenylsilyl)oxy)-2,2-dimethylpropyl)-2-iodo-1H-indole 1-(5-bromo-1H-indol-3-yl)-3-((tert-butyldiphenylsilyl)oxy)-2,2-dimethylpropan-1-one (1.5 g, 2.9 mmol) and I2 (731 mg, 2.9 mmol) were mixed with THF (15 mL) and AgOTf (888 mg, 3.5 mmol) at room temperature. The mixture was stirred at room temperature for 2 hours, then diluted with siRNA (200 mL), washed with saturated Na2S2O3 aqueous solution (100 mL), dried over anhydrous Na2SO4, and filtered. The filtrate was concentrated under reduced pressure, and the residue was purified by silica gel column chromatography to obtain the product (900 mg, 72% yield) as a solid.

[0427] Step 4: Synthesis of (1S)-1-(3-bromopyridine-2-yl)ethanol Under an Ar atmosphere at 0°C, (4S,5S)-2-chloro-2-methyl-1-(4-methylbenzenesulfonyl)-4,5-diphenyl-1,3-diaza-2-lutenacyclopentancymene (3.9 g, 6.0 mmol) was added in several portions to a stirred mixture of HCO2H (66.3 g, 1.44 mol) and Et3N (728 g, 7.2 mol). The mixture was heated to 40°C and stirred for 15 minutes, then cooled to room temperature, and 1-(3-bromopyridine-2-yl)ethanone (120 g, 600 mmol) was added in small portions. The mixture was heated to 40°C and stirred for a further 2 hours, after which the solvent was concentrated under reduced pressure. Brine (2 L) was added to the residue, and the mixture was extracted with ELISA (4 × 700 mL), dried over anhydrous Na2SO4, and filtered. The filtrate was concentrated under reduced pressure, and the residue was purified by silica gel column chromatography to obtain the product (100g, 74% yield) as oil. Calculated value for LCMS(ESI) m / z[M+H]C7H8BrNO: 201.98; measured value: 201.9.

[0428] Step 5: Synthesis of 3-bromo-2-[(1S)-1-methoxyethyl]pyridine At 0°C, (1S)-1-(3-bromopyridine-2-yl)ethanol (100g, 495 mmol) was mixed with DMF (1 L) in a stirring mixture. NaH (60% dispersion of oil) (14.25g, 594 mmol) was added in small quantities. The mixture was stirred at 0°C for 1 hour. MeI (140.5g, 990 mmol) was added dropwise at 0°C, and the mixture was warmed to room temperature and stirred for 2 hours. The mixture was cooled to 0°C, and saturated NH4Cl aqueous solution (5 L) was added. The mixture was extracted with ELISA (3 × 1.5 L), dried over anhydrous Na2SO4, and filtered. The filtrate was concentrated under reduced pressure, and the residue was purified by silica gel column chromatography to obtain the product (90g, 75% yield) as oil. LCMS(ESI) m / z[M+H]C8H 10 Calculated value for BrNO: 215.99; Measured value: 215.9.

[0429] Step 6: Synthesis of 2-[(1S)-1-methoxyethyl]-3-(4,4,5,5-tetramethyl-1,3,2-dioxaborolan-2-yl)pyridine Under an Ar atmosphere at room temperature, 3-bromo-2-[(1S)-1-methoxyethyl]pyridine (90 g, 417 mmol) was mixed with toluene (900 mL) in a stirred mixture. Bis(pinacolato)diborone (127 g, 500 mmol), KOAc (81.8 g, 833 mmol), and Pd(dppf)Cl2 (30.5 g, 41.7 mmol) were added. The mixture was heated to 100 °C and stirred for 3 hours. The filtrate was concentrated under reduced pressure, and the residue was purified by Al2O3 column chromatography to obtain the product (100 g, 63% yield) as a semi-solid. LCMS(ESI) m / z[M+H]C 14 H 22 Calculated value for BNO3: 264.17; Measured value: 264.1.

[0430] Step 7: Synthesis of 5-bromo-3-[3-[(tert-butyldiphenylsilyl)oxy]-2,2-dimethylpropyl]-2-[2-[(1S)-1-methoxyethyl]pyridine-3-yl]-1H-indole 5-Bromo-3-[3-[(tert-butyldiphenylsilyl)oxy]-2,2-dimethylpropyl]-2-iodo-1H-indole (140 g, 217 mmol) and 2-[(1S)-1-methoxyethyl]-3-(4,4,5,5-tetramethyl-1,3,2-dioxaborolan-2-yl)pyridine (100 g, 380 mmol) were mixed with dioxane (1.4 L) and added in small quantities to a stirred mixture. K2CO3 (74.8 g, 541 mmol), Pd(dppf)Cl2 (15.9 g, 21.7 mmol), and H2O (280 mL) were added in small quantities at room temperature under an Ar atmosphere. The mixture was heated to 85°C and stirred for 4 hours, then cold water (5 L) was added, and the mixture was extracted with ELISA (3 × 2 L). The combined organic layers were washed with brine (2 × 1 L), dried over anhydrous Na₂SO₄, and filtered. The filtrate was concentrated under reduced pressure, and the residue was purified by silica gel column chromatography to obtain the product (71 g, 45% yield) as a solid. LCMS(ESI) m / z[M+H]C 37 H 43 Calculated value for BrN2O2Si: 655.23; Measured value: 655.1.

[0431] Step 8: Synthesis of 5-bromo-3-[3-[(tert-butyldiphenylsilyl)oxy]-2,2-dimethylpropyl]-1-ethyl-2-[2-[(1S)-1-methoxyethyl]pyridine-3-yl]indole Under an N2 atmosphere at 0°C, 71 g of 5-bromo-3-[3-[(tert-butyldiphenylsilyl)oxy]-2,2-dimethylpropyl]-2-[2-[(1S)-1-methoxyethyl]pyridine-3-yl]-1H-indole (71 g, 108 mmol) was mixed with 0.8 L of DMF and added in fractions of Cs2CO3 (70.6 g, 217 mmol) and EtI (33.8 g, 217 mmol). The mixture was warmed to room temperature and stirred for 16 hours, then 4 L of H2O was added, and the mixture was extracted with siRNA (3 × 1.5 L). The combined organic layer was washed with brine (2 × 1 L), dried over anhydrous Na2SO4, and filtered. The filtrate was concentrated under reduced pressure, and the residue was purified by silica gel column chromatography to obtain the product (66 g, 80% yield) as oil. LCMS(ESI) m / z[M+H]C 39 H 47 Calculated value for BrN2O2Si: 683.26; Measured value: 683.3.

[0432] Step 9: Synthesis of 3-(5-bromo-1-ethyl-2-[2-[(1S)-1-methoxyethyl]pyridine-3-yl]indole-3-yl)-2,2-dimethylpropan-1-ol Under an N2 atmosphere at room temperature, 5-bromo-3-[3-[(tert-butyldiphenylsilyl)oxy]-2,2-dimethylpropyl]-1-ethyl-2-[2-[(1S)-1-methoxyethyl]pyridine-3-yl]indole (66 g, 97 mmol) was added in fractions to a stirred mixture of TBAF (172.6 g, 660 mmol) and THF (660 mL). The mixture was heated to 50 °C and stirred for 16 hours, then cooled and diluted with H2O (5 L), and extracted with ethyl acetate (3 × 1.5 L). The combined organic layers were washed with brine (2 × 1 L), dried over anhydrous Na2SO4, and filtered. After filtration, the filtrate was concentrated under reduced pressure, and the residue was purified by silica gel column chromatography to obtain the product (30 g, 62% yield) as a solid. LCMS(ESI) m / z[M+H]C 23 H 29 Calculated value for BrN2O2: 445.14; Measured value: 445.1.

[0433] Intermediate 2: Alternative synthesis via the Fischer indole pathway.

[0434] [ka]

[0435] Step 1: Synthesis of 5-[2-[(1S)-1-methoxyethyl]pyridine-3-yl]-2,2-dimethyl-5-oxopentanoic acid Under an N2 atmosphere, at -10°C, n-BuLi (2.5M, 333mL, 833 mmol in hexane) was added dropwise to a mixture of i-PrMgCl (2M, in THF, 0.5L) over 15 minutes. After stirring the mixture at -10°C for 30 minutes, a solution of 3-bromo-2-[(1S)-1-methoxyethyl]pyridine (180g, 833 mmol) in THF (0.5L) was added dropwise at -10°C over 30 minutes. The resulting mixture was warmed to -5°C and stirred for 1 hour, after which a solution of 3,3-dimethyloxane-2,6-dione (118g, 833 mmol) in THF (1.2L) was added dropwise at -5°C over 30 minutes. The mixture was heated to 0°C and stirred for 1.5 hours. Then, HCl from pre-pooled 4M dioxane (0.6 L) was added at 0°C to quench the mixture and adjust the pH to approximately 5. The mixture was diluted with H2O (3 L) at 0°C and extracted with RINKAN (3 × 2.5 L). The combined organic layers were dried over anhydrous Na2SO4, filtered, and the filtrate was concentrated under reduced pressure. The residue was purified by silica gel column chromatography to obtain the product (87 g, 34% yield) as a solid. LC-MS (ESI) m / z[M+H]C 15 H 21 Calculated value for NO4: 280.15; Measured value: 280.1.

[0436] Step 2: Synthesis of 3-(5-bromo-2-[2-[(1S)-1-methoxyethyl]pyridine-3-yl]-1H-indole-3-yl)-2,2-dimethylpropanoic acid and ethyl(S)-3-(5-bromo-2-(2-(1-methoxyethyl)pyridine-3-yl)-1H-indole-3-yl)-2,2-dimethylpropanoate Under an N2 atmosphere at room temperature, 78 g of 5-[2-[(1S)-1-methoxyethyl]pyridine-3-yl]-2,2-dimethyl-5-oxopentanoic acid (279 mmol) was mixed with 0.78 L of EtOH, to which 68.7 g of (4-bromophenyl)hydrazine hydrochloride (307 mmol) was added in fractional amounts. The mixture was heated to 85°C and stirred for 2 hours, then cooled to room temperature, after which 69.8 mL of 4 M HCl (in dioxane, 279 mmol) was added dropwise. The mixture was heated to 85°C and stirred for a further 3 hours, then concentrated under reduced pressure, and the residue was dissolved in 0.78 L of TFA. The mixture was heated to 60°C and stirred for 1.5 hours, then concentrated under reduced pressure, and the residue was basicized to approximately pH 5 with saturated NaHCO3, followed by extraction with SiO2 (3 × 1.5 L). The combined organic layers were dried over anhydrous Na2SO4 and filtered. The filtrate was concentrated under reduced pressure, and the residue was purified by silica gel column chromatography to obtain the product (78 g, crude). LCMS(ESI) m / z[M+H]C 21 H 23 Calculated value for BrN2O3: 430.1, and C 23 H 27 Calculated value for BrN2O3: 459.12; Measured value: 431.1 (carboxylic acid) and 459.1.

[0437] Step 3: Synthesis of ethyl 3-(5-bromo-1-ethyl-2-[2-[(1S)-1-methoxyethyl]pyridine-3-yl]indole-3-yl)-2,2-dimethylpropanoate Under an N2 atmosphere at 0°C, 3-(5-bromo-2-[2-[(1S)-1-methoxyethyl]pyridine-3-yl]-1H-indole-3-yl)-2,2-dimethylpropanoic acid and ethyl(S)-3-(5-bromo-2-(2-(1-methoxyethyl)pyridine-3-yl)-1H-indole-3-yl)-2,2-dimethylpropanoate (198 g, 459 mmol) were added in a mixture with DMF (1.8 L) to which Cs2CO3 (449 g, 1.38 mol) was added in fractions. Next, a solution of EtI (215 g, 1.38 mmol) in DMF (200 mL) was added dropwise at 0°C. The mixture was warmed to room temperature, stirred for 4 hours, diluted with brine (5 L), and extracted with ethyl (3 × 2.5 L). The combined organic layers were washed with brine (2 × 1.5 L), dried over anhydrous Na₂SO₄, and filtered. The filtrate was concentrated under reduced pressure, and the residue was purified by silica gel column chromatography to obtain the product (160 g, 57% yield) as a solid. LCMS(ESI) m / z[M+H]C 25 H 31 Calculated value for BrN2O3: 487.17; Measured value: 487.2.

[0438] Step 4: Synthesis of 3-(5-bromo-1-ethyl-2-(2-((S)-1-methoxyethyl)pyridine-3-yl)-1H-indole-3-yl)-2,2-dimethylpropan-1-ol Under an N2 atmosphere at 0°C, 160 g of ethyl 3-(5-bromo-1-ethyl-2-[2-[(1S)-1-methoxyethyl]pyridine-3-yl]indole-3-yl)-2,2-dimethylpropanoate (328 mmol) was mixed with 1.6 L of THF, to which 28.6 g of LiBH4 (1.3 mol) was added. The mixture was heated to 60°C for 16 hours, cooled, and quenched with a pre-pooled (0°C) saturated NH4Cl aqueous solution (5 L). The mixture was extracted with ethyl phosphate (3 × 2 L), the combined organic layers were washed with brine (2 × 1 L), dried over anhydrous sodium 2 SO4, and filtered. The filtrate was concentrated under reduced pressure, and the residue was purified by silica gel column chromatography to obtain two atropisomers of 3-(5-bromo-1-ethyl-2-(2-((S)-1-methoxyethyl)pyridine-3-yl)-1H-indole-3-yl)-2,2-dimethylpropan-1-ol (as a single atropisomer) (60 g, 38% yield, and 40 g, 26% yield), both as solids. LCMS(ESI) m / z[M+H]C 23 H 29 Calculated value for BrN2O2: 445.14; Measured value: 445.2.

[0439] Intermediate 3: (6 3 S,4S)-4-amino-1 1 -ethyl-1 2 -(2-((S)-1-methoxyethyl)pyridine-3-yl)-10,10-dimethyl-6 1 ,6 2 ,6 3 ,6 4 ,6 3 ,6 6 -Hexahydro-1 1 Synthesis of H-8-oxa-1(5,3)-indola-6(1,3)-pyridadina-2(1,3)-benzenacycloundecafane-5,7-dione

[0440] [ka]

[0441] Step 1: Synthesis of methyl(S)-3-(3-bromophenyl)-2-((tert-butoxycarbonyl)amino)propanoate At room temperature, (2S)-3-(3-bromophenyl)-2-[(tert-butoxycarbonyl)amino]propanoic acid (100 g, 290 mmol) was dissolved in DMF (1 L), to which NaHCO3 (48.8 g, 581.1 mmol) and MeI (61.9 g, 435.8 mmol) were added. The reaction mixture was stirred for 16 hours, then quenched with H2O (1 L) and extracted with SiO2 (3 × 1 L). The combined organic layer was washed with brine (3 × 500 mL), dried over Na2SO4, filtered, and concentrated under reduced pressure. The residue was purified by silica gel column chromatography (13% SiO2 / petroleum ether) to obtain the final product (109 g, crude). LCMS (ESI) m / z[M+Na]C 15 H 20 Calculated value for BrNO4: 380.05; Measured value: 380.0.

[0442] Step 2: Synthesis of methyl(S)-2-((tert-butoxycarbonyl)amino)-3-(3-(4,4,5,5-tetramethyl-1,3,2-dioxaborolan-2-yl)phenyl)propanoate Methyl(2S)-3-(3-bromophenyl)-2-[(tert-butoxycarbonyl)amino]propanoate (108 g, 301.5 mmol) and bis(pinacolato)diborone (99.53 g, 391.93 mmol) were mixed with dioxane (3.2 L) and KOAc (73.97 g, 753.70 mmol) and Pd(dppf)Cl2 (22.06 g, 30.15 mmol) were added. The reaction mixture was heated to 90°C for 3 hours, then cooled to room temperature and extracted with SiO2 (2 × 3 L). The combined organic layer was washed with brine (3 × 800 mL), dried over Na2SO4, filtered, and concentrated under reduced pressure. The residue was purified by silica gel column chromatography (5% SiO2 / petroleum ether) to obtain the product (96 g, 78.6% yield). LCMS(ESI) m / z[M+Na]C 21 H 32 Calculated value for BNO6: 428.22; Measured value: 428.1.

[0443] Step 3: Synthesis of methyl(S)-3-(3-(3-(3-acetoxy-2,2-dimethylpropyl)-1H-indole-5-yl)phenyl)-2-((tert-butoxycarbonyl)amino)propanoate Methyl(2S)-2-[(tert-butoxycarbonyl)amino]-3-[3-(4,4,5,5-tetramethyl-1,3,2-dioxaborolan-2-yl)phenyl]propanoate (94 g, 231.9 mmol) and 3-(5-bromo-1H-indole-3-yl)-2,2-dimethylpropyl acetate (75.19 g, 231.93 mmol) were mixed with dioxane (1.5 L) and H2O (300 mL). K2CO3 (64.11 g, 463.85 mmol) and Pd(DtBPF)Cl2 (15.12 g, 23.19 mmol) were added to the mixture. The reaction mixture was heated to 70°C and stirred for 4 hours. The reaction mixture was extracted with RINKAN (2 × 2 L), the combined organic layer was washed with brine (3 × 600 mL), dried over Na₂SO₄, filtered, and concentrated under reduced pressure. The residue was purified by silica gel column chromatography (20% RINKAN / petroleum ether) to obtain the product (130 g, crude). LC-MS (ESI) m / z: [M + H]C 30 H 38 Calculated value for N2O6: 523.28; measured value: 523.1.

[0444] Step 4: Synthesis of methyl(S)-3-(3-(3-(3-acetoxy-2,2-dimethylpropyl)-2-iodo-1H-indole-5-yl)phenyl)-2-((tert-butoxycarbonyl)amino)propanoate At -10°C, methyl(2S)-3-(3-[3-[3-(acetoxy)-2,2-dimethylpropyl]-1H-indole-5-yl]phenyl)-2-[(tert-butoxycarbonyl)amino]propanoate (95.0 g, 181.8 mmol) and iodine (36.91 g, 145.41 mmol) were dissolved in THF (1 L), to which AgOTf (70.0 g, 272.7 mmol) and NaHCO3 (22.9 g, 272.65 mmol) were added. The reaction mixture was stirred for 30 minutes and then quenched at 0°C by adding saturated Na2S2O3 (100 mL). The resulting mixture was extracted with ELISA (3 × 1 L), the combined organic layer was washed with brine (3 × 500 mL), dried over Na2SO4, filtered, and concentrated under reduced pressure. The residue was purified by silica gel column chromatography (50% ₹ / petroleum ether) to obtain the desired product (49.3 g, 41.8% yield). LCMS(ESI) m / z[M+H]C 30 H 37 Calculated value for IN2O6: 649.18; Measured value: 649.1.

[0445] Step 5: Synthesis of (S)-2-((tert-butoxycarbonyl)amino)-3-(3-(3-(3-hydroxy-2,2-dimethylpropyl)-2-iodo-1H-indole-5-yl)phenyl)propanoic acid A solution of methyl(2S)-3-(3-[3-[3-(acetyloxy)-2,2-dimethylpropyl]-2-iodo-1H-indole-5-yl]phenyl)-2-[(tert-butoxycarbonyl)amino]propanoate (60 g, 92.5 mmol) in THF (600 mL) was mixed with an aqueous solution of LiOH·H₂O (19.41 g, 462.5 mmol) (460 mL). The resulting solution was stirred overnight, and the pH was adjusted to 6 with HCl (1 M). The resulting solution was extracted with ELISA (2 × 500 mL), the combined organic layers were washed with brine (2 × 500 mL), dried over Na₂SO₄, filtered, and concentrated under reduced pressure to obtain the product (45 g, 82.1% yield). LCMS(ESI) m / z[M+Na]C 27 H 33Calculated value for IN2O6: 615.13; Measured value: 615.1.

[0446] Step 6: Synthesis of methyl(S)-1-((S)-2-((tert-butoxycarbonyl)amino)-3-(3-(3-(3-hydroxy-2,2-dimethylpropyl)-2-iodo-1H-indole-5-yl)phenyl)propanoyl)hexahydropyridazine-3-carboxylate (2S)-2-[(tert-butoxycarbonyl)amino]-3-[3-[3-(3-hydroxy-2,2-dimethylpropyl)-2-iodo-1H-indole-5-yl]phenyl]propanoic acid (30 g, 50.6 mmol) and methyl(3S)-1,2-diadinane-3-carboxylate (10.9 g, 75.9 mmol) were dissolved in DCM (400 mL), to which NMM (40.97 g, 405.08 mmol), HOBt (2.05 g, 15.19 mmol), and EDCI (19.41 g, 101.27 mmol) were added. The reaction mixture was stirred overnight, washed with saturated NH4Cl aqueous solution (2 × 200 mL) and brine (2 × 200 mL), dried over Na2SO4, filtered, and concentrated under reduced pressure to obtain the product (14 g, 38.5% yield). LCMS(ESI) m / z:[M+H]C 33 H 43 Calculated value for IN4O6: 718.23; measured value: 719.4.

[0447] Step 7: Synthesis of (S)-1-((S)-2-((tert-butoxycarbonyl)amino)-3-(3-(3-(3-hydroxy-2,2-dimethylpropyl)-2-iodo-1H-indole-5-yl)phenyl)propanoyl)hexahydropyridazine-3-carboxylic acid At 0°C, methyl(S)-1-((S)-2-((tert-butoxycarbonyl)amino)-3-(3-(3-(3-hydroxy-2,2-dimethylpropyl)-2-iodo-1H-indole-5-yl)phenyl)propanoyl)hexahydropyridazine-3-carboxylate (92 g, 128.0 mmol) was dissolved in THF (920 mL), to which an aqueous solution of LiOH·H2O (26.86 g, 640.10 mmol) (640 mL) was added. The reaction mixture was stirred for 2 hours, and then concentrated under reduced pressure to obtain the product (90 g, crude). LCMS(ESI) m / z:[M+H]C 32 H 41 Calculated value for IN4O6: 705.22; measured value: 705.1.

[0448] Step 8: tert-butyl((6 3 S,4S)-1 2 -iodo-10,10-dimethyl-5,7-dioxo-6 1 ,6 2 ,6 3 ,6 4 ,6 5 ,6 6 -Hexahydro-1 1 Synthesis of H-8-oxa-1(5,3)-indola-6(1,3)-pyridadina-2(1,3)-benzenacycloundecafan-4-yl)carbamate At 0°C, (3S)-1-[(2S)-2-[(tert-butoxycarbonyl)amino]-3-[3-[3-(3-hydroxy-2,2-dimethylpropyl)-2-iodo-1H-indole-5-yl]phenyl]propanoyl]-1,2-diadinane-3-carboxylic acid (90 g, 127.73 mmol) was dissolved in DCM (10 L), to which HOBt (34.52 g, 255.46 mmol), DIPEA (330.17 g, 2554.62 mmol), and EDCI (367.29 g, 1915.96 mmol) were added. The reaction mixture was stirred for 16 hours and then concentrated under reduced pressure. The mixture was extracted with DCM (2 × 2 L), the combined organic layers were washed with brine (3 × 1 L), dried over Na₂SO₄, filtered, and concentrated under reduced pressure. The residue was purified by silica gel column chromatography (50% siRNA / petroleum ether) to obtain the product (70 g, 79.8% yield). LCMS(ESI) m / z:[M+H]C 32 H 39 Calculated value for IN4O5: 687.21; Measured value: 687.1.

[0449] Step 9: tert-butyl((6 3 S,4S)-10,10-dimethyl-5,7-dioxo-12-(4,4,5,5-tetramethyl-1,3,2-dioxaborolan-2-yl)-6 1 ,6 2 ,6 3 ,6 4 ,6 5 ,6 6 -Hexahydro-1 1 Synthesis of H-8-oxa-1(5,3)-indola-6(1,3)-pyridadina-2(1,3)-benzenacycloundecafan-4-yl)carbamate At room temperature, in a 1 L round-bottom flask, add tert-butyl ((6 3 S,4S)-1 2 -iodo-10,10-dimethyl-5,7-dioxo-6 1 ,6 2 ,6 3 ,6 4 ,6 5 ,6 6 -Hexahydro-1 1H-8-oxa-1(5,3)-indola-6(1,3)-pyridadina-2(1,3)-benzenacycloundecafan-4-yl)carbamate (22.0 g, 32.042 mmol), toluene (300.0 mL), Pd2(dba)3 (3.52 g, 3.845 mmol), S-Phos (3.95 g, 9.613 mmol), and KOAc (9.43 g, 96.127 mmol) were packed into the mixture. At room temperature, 4,4,5,5-tetramethyl-1,3,2-dioxaborolane (26.66 g, 208.275 mmol) was added dropwise with stirring. The resulting solution was stirred at 60°C for 3 hours. The resulting mixture was filtered, and the filtrate was washed with ELISA. The filtrate was concentrated under reduced pressure, and the remaining residue was purified by silica gel column chromatography to obtain the product (22 g, 90% yield) as a solid. LCMS(ESI) m / z:[M+H]C 38 H 51 Calculated value for BN4O7: 687.3; measured value: 687.4.

[0450] Step 10: tert-butyl((6 3 S,4S)-1 2 -(2-((S)-1-methoxyethyl)pyridine-3-yl)-10,10-dimethyl-5,7-dioxo-6 1 ,6 2 ,6 3 ,6 4 ,6 5 ,6 6 -Hexahydro-1 1 Synthesis of H-8-oxa-1(5,3)-indola-6(1,3)-pyridadina-2(1,3)-benzenacycloundecafan-4-yl)carbamate tert-butyl((6 3 S,4S)-10,10-dimethyl-5,7-dioxo-1 2 -(4,4,5,5-tetramethyl-1,3,2-dioxaborolan-2-yl)-6 1 ,6 2 ,6 3 ,6 4 ,6 5 ,6 6 -Hexahydro-1 1A mixture of H-8-oxa-1(5,3)-indola-6(1,3)-pyridadina-2(1,3)-benzenacycloundecafan-4-yl)carbamate (2.0 g, 2.8 mmol), 3-bromo-2-[(1S)-1-methoxyethyl]pyridine (0.60 g, 2.8 mmol), Pd(dppf)Cl2 (0.39 g, 0.5 mmol), and K3PO4 (1.2 g, 6.0 mmol) with dioxane (50 mL) and H2O (10 mL) was heated to 70°C under an N2 atmosphere and stirred for 2 hours. The mixture was diluted with H2O (50 mL) and extracted with ethyl acetate (3 × 50 mL). The combined organic layer was washed with brine (3 × 50 mL), dried over anhydrous Na2SO4, and filtered. The resulting mixture was concentrated under reduced pressure, and the residue was purified by silica gel column chromatography to obtain the desired product (1.5 g, 74% yield). LCMS(ESI) m / z:[M+H]C 40 H 49 Calculated value for N5O6: 695.4; measured value: 696.5.

[0451] Step 11: tert-butyl((6 3 S,4S)-1 1 -ethyl-1 2 -(2-((S)-1-methoxyethyl)pyridine-3-yl)-10,10-dimethyl-5,7-dioxo-6 1 ,6 2 ,6 3 ,6 4 ,6 5 ,6 6 -Hexahydro-1 1 Synthesis of H-8-oxa-1(5,3)-indola-6(1,3)-pyridadina-2(1,3)-benzenacycloundecafan-4-yl)carbamate At 0°C, tert-butyl((6 3 S,4S)-1 2 -(2-((S)-1-methoxyethyl)pyridine-3-yl)-10,10-dimethyl-5,7-dioxo-6 1 ,6 2 ,6 3 ,6 4 ,6 5 ,6 6 -Hexahydro-11 H-8-oxa-1(5,3)-indola-6(1,3)-pyridadina-2(1,3)-benzenacycloundecafan-4-yl)carbamate (20 g, 28.7 mmol) and Cs2CO3 (18.7 g, 57.5 mmol) were mixed in DMF (150 mL) and EtI (13.45 g, 86.22 mmol) in DMF (50 mL). The resulting mixture was stirred overnight at 35°C and then diluted with H2O (500 mL). The mixture was extracted with siRNA (2 × 300 mL), the combined organic layer was washed with brine (3 × 100 mL), dried over Na2SO4, filtered, and concentrated under reduced pressure. The residue was purified by silica gel column chromatography to obtain the product (4.23 g, 18.8% yield) and the atropisomer (5.78 g, 25.7%) as solids. LCMS(ESI) m / z:[M+H]C 42 H 53 Calculated value for N5O6: 724.4; measured value: 724.6.

[0452] Step 12:(6 3 S,4S)-4-amino-1 1 -ethyl-1 2 -(2-((S)-1-methoxyethyl)pyridine-3-yl)-10,10-dimethyl-6 1 ,6 2 ,6 3 ,6 4 ,6 5 ,6 6 -Hexahydro-1 1 Synthesis of H-8-oxa-1(5,3)-indola-6(1,3)-pyridadina-2(1,3)-benzenacycloundecafane-5,7-dione tert-butyl((6 3 S,4S)-1 1 -ethyl-1 2 -(2-((S)-1-methoxyethyl)pyridine-3-yl)-10,10-dimethyl-5,7-dioxo-6 1 ,6 2 ,6 3 ,6 4 ,6 5 ,6 6 -Hexahydro-1 1A mixture of H-8-oxa-1(5,3)-indola-6(1,3)-pyridadina-2(1,3)-benzenacycloundecafan-4-yl)carbamate (1.3 g, 1.7 mmol) with TFA (10 mL) and DCM (20 mL) was stirred at 0°C for 2 hours. The mixture was concentrated under reduced pressure to obtain the product (1.30 g, crude) as a solid. LC-MS (ESI) m / z: [M+H]C 37 H 45 Calculated value for N5O4: 623.3; measured value: 624.4.

[0453] Intermediate 4: tert-butyl((6 3 S,4S,Z)-1 1 -ethyl-1 2 -(2-((S)-1-methoxyethyl)pyridine-3-yl)-10,10-dimethyl-5,7-dioxo-6 1 ,6 2 ,6 3 ,6 4 ,6 5 ,6 6 -Hexahydro-1 1 Synthesis of H-8-oxa-2(4,2)-thiazola-1(5,3)-indola-6(1,3)-pyridadinacycloundecafan-4-yl)carbamate

[0454] [ka]

[0455] Step 1: Synthesis of (S)-3-(4-bromothiazole-2-yl)-2-((tert-butoxycarbonyl)amino)propanoic acid At room temperature, methyl(2S)-3-(4-bromo-1,3-thiazole-2-yl)-2-[(tert-butoxycarbonyl)amino]propanoate (110 g, 301.2 mmol) was dissolved in THF (500 mL) and H2O (200 mL) with LiOH (21.64 g, 903.6 mmol). The resulting solution was stirred for 1 hour and then concentrated under reduced pressure. The resulting residue was adjusted to pH 6 with 1 M HCl and extracted by DCM (3 × 500 mL). The combined organic layers were dried over Na2SO4, filtered, and concentrated under reduced pressure to obtain the desired product (108 g, crude). LCMS(ESI) m / z[M+H]C 11 H 15 Calculated value for BrN2O4S: 351.00; Measured value: 351.0.

[0456] Step 2: Synthesis of methyl(S)-1-((S)-3-(4-bromothiazole-2-yl)-2-((tert-butoxycarbonyl)amino)propanoyl)hexahydropyridazine-3-carboxylate At 0°C, (S)-3-(4-bromothiazole-2-yl)-2-((tert-butoxycarbonyl)amino)propanoic acid (70 g, 199.3 mmol) was dissolved in DCM (500 mL) and then methyl(3S)-1,2-diadinane-3-carboxylate bis(trifluoroacetic acid) (111.28 g, 298.96 mmol), NMM (219.12 mL, 1993.0 mmol), EDCI (76.41 g, 398.6 mmol), and HOBt (5.39 g, 39.89 mmol) were added. The resulting solution was warmed to room temperature and stirred for 1 hour. Next, the reaction mixture was quenched with H2O (500 mL) and extracted with ELISA (3 × 500 mL). The combined organic layer was dried over Na2SO4, filtered, and concentrated under reduced pressure. The residue was purified by silica gel column chromatography (0 → 50% Â / petroleum ether) to obtain the desired product (88.1 g, 93% yield). LCMS(ESI) m / z[M+H]C 17 H 25 Calculated value for BrN4O5S: 477.08; Measured value: 477.1.

[0457] Step 3: Synthesis of (S)-3-(1-ethyl-2-(2-(1-methoxyethyl)pyridine-3-yl)-5-(4,4,5,5-tetramethyl-1,3,2-dioxaborolan-2-yl)-1H-indole-3-yl)-2,2-dimethylpropane-1-ol At room temperature, 60 g, 134.7 mmol of 3-(5-bromo-1-ethyl-2-(2-((S)-1-methoxyethyl)pyridine-3-yl)-1H-indole-3-yl)-2,2-dimethylpropan-1-ol was dissolved in 500 mL of toluene. Bis(pinacolato)diborone (51.31 g, 202.1 mmol), Pd(dppf)Cl2 (9.86 g, 13.48 mmol), and KOAc (26.44 g, 269.4 mmol) were added to this solution. The reaction mixture was then heated to 90°C and stirred for 2 hours. The reaction solution was then cooled to room temperature and concentrated under reduced pressure. The desired product was obtained by purification by silica gel column chromatography (0 → 50% siRNA / petroleum ether) (60.6 g, 94% yield). LCMS(ESI) m / z[M+H]C 29 H 41 Calculated value for BN2O4: 493.32; Measured value: 493.3.

[0458] Step 4: Synthesis of methyl(S)-1-((S)-2-((tert-butoxycarbonyl)amino)-3-(4-(1-ethyl-3-(3-hydroxy-2,2-dimethylpropyl)-2-(2-((S)-1-methoxyethyl)pyridine-3-yl)-1H-indole-5-yl)thiazole-2-yl)propanoyl)hexahydropyridazine-3-carboxylate At room temperature, (S)-3-(1-ethyl-2-(2-(1-methoxyethyl)pyridine-3-yl)-5-(4,4,5,5-tetramethyl-1,3,2-dioxaborolan-2-yl)-1H-indole-3-yl)-2,2-dimethylpropan-1-ol (30 g, 60.9 mmol) is dissolved in toluene (600 mL), dioxane (200 mL), and H2O (200 mL). To the solution, methyl(S)-1-((S)-3-(4-bromothiazole-2-yl)-2-((tert-butoxycarbonyl)amino)propanoyl)hexahydropyridazine-3-carboxylate (43.62 g, 91.4 mmol), K3PO4 (32.23 g, 152.3 mmol), and Pd(dppf)Cl2 (8.91 g, 12.18 mmol) were added. The resulting solution was heated to 70°C and stirred overnight. Next, the reaction mixture was cooled to room temperature and quenched with H2O (200 mL). The resulting mixture was extracted with ELISA (3 × 1000 mL), and the combined organic layer was dried over Na2SO4, filtered, and concentrated under reduced pressure. The residue was purified by silica gel column chromatography (0 → 90% ELISA / petroleum ether) to obtain the desired product (39.7 g, 85% yield). LCMS(ESI) m / z[M+H]C 40 H 54 Calculated value for N6O7S: 763.39; Measured value: 763.3.

[0459] Step 5: Synthesis of (S)-1-((S)-2-((tert-butoxycarbonyl)amino)-3-(4-(1-ethyl-3-(3-hydroxy-2,2-dimethylpropyl)-2-(2-((S)-1-methoxyethyl)pyridine-3-yl)-1H-indole-5-yl)thiazole-2-yl)propanoyl)hexahydropyridazine-3-carboxylic acid At room temperature, methyl(S)-1-((S)-2-((tert-butoxycarbonyl)amino)-3-(4-(1-ethyl-3-(3-hydroxy-2,2-dimethylpropyl)-2-(2-((S)-1-methoxyethyl)pyridine-3-yl)-1H-indole-5-yl)thiazole-2-yl)propanoyl)hexahydropyridazine-3-carboxylate (39.7 g, 52.0 mmol) was dissolved in THF (400 mL) and H2O (100 mL) and LiOH·H2O (3.74 g, 156.2 mmol) was added. The resulting mixture was stirred for 1.5 hours and then concentrated under reduced pressure. The residue was acidified to pH 6 with 1 M HCl and extracted by DCM (3 × 1000 mL). The combined organic layers were dried over Na2SO4, filtered, and concentrated under reduced pressure to obtain the desired product (37.9 g, crude). LC-MS(ESI) m / z[M+H]C 39 H 52 Calculated value for N6O7S: 749.37; Measured value: 749.4.

[0460] Step 6: tert-butyl((6 3 S,4S,Z)-1 1 -ethyl-1 2 -(2-((S)-1-methoxyethyl)pyridine-3-yl)-10,10-dimethyl-5,7-dioxo-6 1 ,6 2 ,6 3 ,6 4 ,6 5 ,6 6 -Hexahydro-1 1 Synthesis of H-8-oxa-2(4,2)-thiazola-1(5,3)-indola-6(1,3)-pyridadinacycloundecafan-4-yl)carbamate At 0°C, EDCI (271.63 g, 1416.9 mmol) was added to a solution of (S)-1-((S)-2-((tert-butoxycarbonyl)amino)-3-(4-(1-ethyl-3-(3-hydroxy-2,2-dimethylpropyl)-2-(2-((S)-1-methoxyethyl)pyridine-3-yl)-1H-indole-5-yl)thiazole-2-yl)propanoyl)hexahydropyridazine-3-carboxylic acid (37.9 g, 50.6 mmol), HOBt (34.19 g, 253.0 mmol), and DIPEA (264.4 mL, 1518 mmol) in DCM (4 L). The resulting mixture was warmed to room temperature and stirred overnight. The reaction mixture was then quenched with H2O and washed with 1 M HCl (4 × 1 L). The organic layer was separated and concentrated under reduced pressure. The residue was purified by silica gel column chromatography (0 → 70% Â / petroleum ether) to obtain the desired product (30 g, 81% yield). LCMS(ESI) m / z[M+H]C 39 H 50 Calculated value for N6O6S: 731.36; Measured value: 731.3.

[0461] Intermediate 5:(6 3 S)-4-amino-1 1 -ethyl-1 2 -(2-((S)-1-methoxyethyl)pyridine-3-yl)-10,10-dimethyl-2 1 ,2 2 ,2 3 ,2 6 ,6 1 ,6 2 ,6 3 ,6 4 ,6 5 ,6 6 - Decahydro-1 1 Synthesis of H-8-oxa-1(5,3)-indola-6(1,3)-pyridadina-2(5,1)-pyridinacyclodecafane-5,7-dione

[0462] [ka]

[0463] Step 1: Synthesis of methyl 2-((tert-butoxycarbonyl)amino)acrylate DIPEA (17 g, 137 mmol) was added to a solution of methyl (tert-butoxycarbonyl)-L-serinate (10 g, 45 mmol) in anhydrous MeCN (150 mL). The mixture was stirred at 45 °C for 2 hours, and the product was obtained in solution. LC-MS (ESI): m / z [M+Na]C9H 15 Calculated value for NO4: 201.1; measured value: 224.1.

[0464] Step 2: Synthesis of methyl 2-(bis(tert-butoxycarbonyl)amino)acrylate At 0°C, methyl 2-((tert-butoxycarbonyl)amino)acrylate (12 g, 60 mmol) was added to a solution of anhydrous MeCN (150 mL) with DMAP (13 g, 90 mmol) and (Boc)2O (26 g, 120 mmol). After stirring the reaction mixture for 6 hours, it was quenched with H2O (100 mL) and extracted by DCM (3 × 200 mL). The combined organic layer was washed with brine (150 mL), dried over anhydrous Na2SO4, filtered, and concentrated under reduced pressure. The residue was purified by silica gel column chromatography to obtain the product (12.5 g, 65% yield) as a solid. LCMS(ESI): m / z [M+Na]C 14 H 23 Calculated value for NO6: 301.2; measured value: 324.1.

[0465] Step 3: Synthesis of methyl 2-(bis(tert-butoxycarbonyl)amino)-3-(5-bromo-3,6-dihydropyridine-1(2H)-yl)propanoate Under an Ar atmosphere, methyl 2-{bis[(tert-butoxy)carbonyl]amino}prop-2-enoate (22 g, 74 mmol) was added to a mixture of 5-bromo-1,2,3,6-tetrahydropyridine (8.0 g, 49 mmol) and MeOH (120 mL). After stirring the mixture for 16 hours, it was concentrated under reduced pressure, and the residue was purified by silica gel column chromatography to obtain the product (12 g, 47% yield) as oil. LCMS(ESI) m / z:[M+H]C19 H 31 Calculated value for BrN2O6: 462.1; measured value: 463.1.

[0466] Step 4: Synthesis of 3-(5-bromo-3,6-dihydropyridine-1(2H)-yl)-2-((tert-butoxycarbonyl)amino)propanoic acid A mixture of methyl 2-(bis(tert-butoxycarbonyl)amino)-3-(5-bromo-3,6-dihydropyridine-1(2H)-yl)propanoate (14 g, 30 mmol) with dioxane (30 mL) and H2O (12 mL) was mixed with LiOH (3.6 g, 151 mmol). The mixture was heated to 35°C and stirred for 12 hours, then 1 M was added. HCl was added to adjust the pH to approximately 3-4. The mixture was extracted using DCM (2 × 300 mL), and the combined organic layer was dried over anhydrous Na₂SO₄ and filtered. The filtrate was concentrated under reduced pressure to obtain the product (10 g, 85% yield) as a solid. LCMS(ESI)m / z [M+H]C 13 H 21 Calculated value for BrN2O4: 348.1; Measured value: 349.0.

[0467] Step 5: Synthesis of methyl(3S)-1-(3-(5-bromo-3,6-dihydropyridine-1(2H)-yl)-2-((tert-butoxycarbonyl)amino)propanoyl)hexahydropyridazine-3-carboxylate Under an Ar atmosphere at 0°C, HATU (13 g, 34 mmol) was added to a mixture of 3-(5-bromo-3,6-dihydropyridine-1(2H)-yl)-2-((tert-butoxycarbonyl)amino)propanoic acid (10 g, 30 mmol), DIPEA (12 g, 93 mmol), and methyl(3S)-1,2-diadinane-3-carboxylate (5.4 g, 37 mmol) with DMF (100 mL). The mixture was stirred at 0°C for 2 hours, then H2O was added and the mixture was extracted with ELISA (2 × 300 mL). The combined organic layer was dried over anhydrous Na2SO4 and filtered. The filtrate was concentrated under reduced pressure, and the residue was purified by reverse-phase chromatography to obtain the product (9.0 g, 55% yield) as a solid. LCMS(ESI)m / z [M+H]C 19 H 31 Calculated value for BrN4O5: 474.1; Measured value: 475.1.

[0468] Step 6: Synthesis of methyl(3S)-1-(2-((tert-butoxycarbonyl)amino)-3-(5-(1-ethyl-3-(3-hydroxy-2,2-dimethylpropyl)-2-(2-((S)-1-methoxyethyl)pyridine-3-yl)-1H-indole-5-yl)-3,6-dihydropyridine-1(2H)-yl)propanoyl)hexahydropyridazine-3-carboxylate Under an Ar atmosphere, methyl(3S)-1-(3-(5-bromo-3,6-dihydropyridine-1(2H)-yl)-2-((tert-butoxycarbonyl)amino)propanoyl)hexahydropyridazine-3-carboxylate (9.0g, 18mmol), K2CO3 (4.5g, 32mmol), Pd(dppf)Cl2·DCM (1.4g, 2mmol), 3-( A mixture of 1-ethyl-2-{2-[(1S)-1-methoxyethyl]pyridine-3-yl}-5-(4,4,5,5-tetramethyl-1,3,2-dioxaborolan-2-yl)indole-3-yl)-2,2-dimethylpropan-1-ol (9.8 g, 20 mmol) with dioxane (90 mL) and H2O (10 mL) was heated to 75°C and stirred for 2 hours. H2O was added, and the mixture was extracted with ELISA (3 × 200 mL). The combined organic layer was dried over Na2SO4, filtered, and the filtrate was concentrated under reduced pressure. The residue was purified by silica gel column chromatography to obtain 5-[2-[(1S)-1-methoxyethyl]pyridine-3-yl]-2,2-dimethyl-5-oxopentanoic acid (4.0 g, 25% yield) as a solid. LCMS(ESI)m / z [M+H]C 42 H 60 Calculated value for N6O7: 760.5; measured value: 761.4.

[0469] Step 7: Synthesis of (3S)-1-(2-((tert-butoxycarbonyl)amino)-3-(5-(1-ethyl-3-(3-hydroxy-2,2-dimethylpropyl)-2-(2-((S)-1-methoxyethyl)pyridine-3-yl)-1H-indole-5-yl)-3,6-dihydropyridine-1(2H)-yl)propanoyl)hexahydropyridazine-3-carboxylic acid At 0°C, methyl(3S)-1-(2-((tert-butoxycarbonyl)amino)-3-(5-(1-ethyl-3-(3-hydroxy-2,2-dimethylpropyl)-2-(2-((S)-1-methoxyethyl)pyridine-3-yl)-1H-indole-5-yl)-3,6-dihydropyridine-1(2H)-yl)propanoyl)hexahydropyridazine-3-carboxylate (4.1 g, 5.0 mmol) was mixed with THF (35 mL) and LiOH (0.60 g, 27 mmol) was added. The mixture was stirred at 0°C for 1.5 hours, then 1 M HCl was added to adjust the pH to approximately 6-7, and the mixture was extracted with ELISA (3 × 200 mL). The combined organic layer was dried over Na₂SO₄ and filtered. The filtrate was concentrated under reduced pressure to obtain the product (3.6 g, 80% yield) as a solid. LCMS(ESI)m / z [M+H]C 41 H 58 Calculated value for N6O7: 746.4; measured value: 747.4.

[0470] Step 8: tert-butyl((6 3 S)-1 1 -ethyl-1 2 -(2-((S)-1-methoxyethyl)pyridine-3-yl)-10,10-dimethyl-5,7-dioxo-2 1 ,2 2 ,2 3 ,2 6 ,6 1 ,6 2 ,6 3 ,6 4 ,6 5 ,6 6 - Decahydro-1 1 Synthesis of H-8-oxa-1(5,3)-indola-6(1,3)-pyridadina-2(5,1)-pyridinacyclodecafan-4-yl)carbamate Under an Ar atmosphere, (3S)-1-(2-((tert-butoxycarbonyl)amino)-3-(5-(1-ethyl-3-(3-hydroxy-2,2-dimethylpropyl)-2-(2-((S)-1-methoxyethyl)pyridine-3-yl)-1H-indole-5-yl)-3,6-dihydropyridine-1(2H)-yl)propanoyl)hexahydropyridazine-3-carboxylic acid (3.6 g, 5.0 mmol) and DIPEA (24 g, 190 mmol) were mixed with DCM (700 mL), to which EDCI·HCl (28 g, 140 mmol) and HOBt (6.5 g, 50 mmol) were added. The mixture was heated to 30°C, stirred at 30°C for 16 hours, and then concentrated under reduced pressure. The residue was diluted with siRNA (200 mL), washed with H₂O (2 × 200 mL) and brine (200 mL), dried over Na₂SO₄, and filtered. The filtrate was concentrated under reduced pressure, and the residue was purified by silica gel column chromatography to obtain the product (1.45 g, 40% yield) as a solid. LCMS(ESI) m / z:[M+H]C 41 H 56 Calculated value for N6O6: 728.4; measured value: 729.4.

[0471] Step 9:(6 3 S)-4-amino-1 1 -ethyl-1 2 -(2-((S)-1-methoxyethyl)pyridine-3-yl)-10,10-dimethyl-2 1 ,2 2 ,2 3 ,2 6 ,6 1 ,6 2 ,6 3 ,6 4 ,6 5 ,6 6 - Decahydro-1 1 Synthesis of H-8-oxa-1(5,3)-indola-6(1,3)-pyridadina-2(5,1)-pyridinacyclodecafane-5,7-dione At 0°C, tert-butyl((6 3 S)-1 1 -ethyl-1 2-(2-((S)-1-methoxyethyl)pyridine-3-yl)-10,10-dimethyl-5,7-dioxo-2 1 ,2 2 ,2 3 ,2 6 ,6 1 ,6 2 ,6 3 ,6 4 ,6 5 ,6 6 - Decahydro-1 1 A mixture of H-8-oxa-1(5,3)-indola-6(1,3)-pyridadina-2(5,1)-pyridinacyclodecafan-4-yl)carbamate (130 mg, 0.20 mmol) and DCM (1.0 mL) was mixed with TFA (0.3 mL). The mixture was heated to room temperature and stirred for 2 hours, then concentrated under reduced pressure to obtain the product, which was used directly in the next step without further purification. LC-MS(ESI) m / z[M+H]C 36 H 48 Calculated value for N6O4: 628.4; measured value: 629.4.

[0472] Intermediate 6: (2 2 S,6 3 S,4S)-4-amino-1 1 -ethyl-1 2 -(2-((S)-1-methoxyethyl)pyridine-3-yl)-10,10-dimethyl-6 1 ,6 2 ,6 3 ,6 4 ,6 5 ,6 6 -Hexahydro-1 1 Synthesis of H-8-oxa-2(4,2)-morpholina-1(5,3)-indola-6(1,3)-pyridadinacycloundecafane-5,7-dione

[0473] [ka]

[0474] Step 1: Synthesis of tert-butyl(2R)-2-formylmorpholine-4-ylformate At room temperature, tert-butyl(2R)-2-(hydroxymethyl)morpholine-4-ylformate (50 g, 230 mmol) was dissolved in ethyl acetate (1 L), to which TEMPO (715 mg, 4.6 mmol) and NaHCO3 (58 g, 690 mmol) were added. After cooling the mixture to -50°C, a solution of TCCA (56 g, 241 mmol) in ethyl acetate (100 mL) was added dropwise over 30 minutes. The reaction mixture was warmed to 5°C for 2 hours, then quenched with 10% Na2S2O3 (200 mL) and stirred for 20 minutes. The resulting mixture was filtered to separate the organic phase. The aqueous phase was extracted with ethyl acetate (2 × 100 mL). The combined organic layers were washed with H2O (100 mL) and brine (100 mL) and dried over anhydrous Na2SO4. The organic layers were concentrated under reduced pressure to obtain the product (50 g, crude) as oil.

[0475] Step 2: Synthesis to obtain tert-butyl(S,Z)-2-(2-(((benzyloxy)carbonyl)amino)-3-methoxy-3-oxopropane-1-en-1-yl)morpholine-4-carboxylate To a solution of tert-butyl(2R)-2-formylmorpholine-4-ylformate (49 g, 153 mmol) and methyl 2-{[(benzyloxy)carbonyl]amino}-2-(dimethoxyphosphoryl)acetate (60 g, 183 mmol) in MeCN (300 mL), tetramethylguanidine (35 g, 306 mmol) was added at 0-10°C. The reaction mixture was stirred at 10°C for 30 minutes and then warmed to room temperature for 2 hours. The reaction mixture was diluted with DCM (200 mL) and washed with 10% citric acid (200 mL) and 10% NaHCO3 aqueous solution (200 mL). The organic phase was concentrated under reduced pressure and purified by silica gel column chromatography to obtain the product (36 g, 90% yield) as a solid. LCMS(ESI): m / z[M+Na]C 21 H 28 The calculated value for N2O4 is 420.2; the measured value is 443.1.

[0476] Step 3: Synthesis of tert-butyl(S)-2-((S)-2-(((benzyloxy)carbonyl)amino)-3-methoxy-3-oxopropyl)morpholine-4-carboxylate To a solution of tert-butyl(S,Z)-2-(2-(((benzyloxy)carbonyl)amino)-3-methoxy-3-oxopropa-1-en-1-yl)morpholine-4-carboxylate (49 g, 0.12 mol) in MeOH (500 mL), (S,S)-Et-DUPHOS-Rh (500 mg, 0.7 mmol) was added. The mixture was stirred at room temperature for 48 hours under an H2 (60 psi) atmosphere. The reaction product was concentrated and purified by silica gel column chromatography to obtain the product (44 g, 90% yield) as a solid. LCMS(ESI): m / z [M+Na]C 21 H 30 The calculated value for N2O7 is 422.2; the measured value is 445.2.

[0477] Step 4: Synthesis of methyl(S)-2-(((benzyloxy)carbonyl)amino)-3-((S)-morpholine-2-yl)propanoate At 15°C, 2.2 g, 5.2 mmol of tert-butyl(S)-2-((S)-2-(((benzyloxy)carbonyl)amino)-3-methoxy-3-oxopropyl)morpholine-4-carboxylate was stirred with 2 mL of HCl, to which 25 mL of HCl / HCl was added. The reaction mixture was stirred at 15°C for 2 hours, then concentrated under reduced pressure to obtain the product (1.51 g, 90% yield) as oil. LCMS(ESI) m / z[M+H]C 16 H 22 Calculated value for N2O5: 322.1; Measured value: 323.2.

[0478] Step 5: Synthesis of (S)-5-bromo-3-(3-((tert-butyldimethylsilyl)oxy)-2,2-dimethylpropyl)-1-ethyl-2-(2-(1-methoxyethyl)pyridine-3-yl)-1H-indole 3-(5-bromo-1-ethyl-2-{2-[(1S)-1-methoxyethyl]pyridine-3-yl}indole-3-yl)-2,2-dimethylpropan-1-ol (100 g, 0.22 mol) and imidazole (30.6 g, 0.45 mol) were added to a solution of TBSCl (50.7 g, 0.34 mol) in a solution of TBSCl (200 mL) at 0°C. The reaction mixture was stirred at room temperature for 2 hours. The resulting solution was washed with H2O (3 × 300 mL) and brine (2 × 200 mL), dried over anhydrous Na2SO4, filtered, and concentrated under reduced pressure. The residue was purified by silica gel column chromatography to obtain the product (138 g, 90% yield) as a solid. LCMS(ESI) m / z:[M+H]C 29 H 43 Calculated value for BrN2O2Si: 558.2; measured value: 559.2.

[0479] Step 6: Synthesis of methyl(2S)-2-{[(benzyloxy)carbonyl]amino}-3-[(2S)-4-(3-{3-[(tert-butyldimethylsilyl)oxy]-2,2-dimethylpropyl}-1-ethyl-2-{2-[(1S)-1-methoxyethyl]pyridine-3-yl}indole-5-yl)morpholine-2-yl]propanoate At 105°C, under an N2 atmosphere, (S)-5-bromo-3-(3-((tert-butyldimethylsilyl)oxy)-2,2-dimethylpropyl)-1-ethyl-2-(2-(1-methoxyethyl)pyridine-3-yl)-1H-indole (50 g, 89.3 mmol) was mixed with dioxane (500 mL) in a stirred solution. S)-Morpholin-2-yl]propanoate (31.7 g, 98.2 mmol), RuPhos (16.7 g, 35.7 mmol), di-μ-chlorobis(2-amino-1,1-biphenyl-2-yl-C,N)dipalladium(II) (2.8 g, 4.4 mmol), and cesium carbonate (96 g, 295 mmol), followed by RuPhos-Pd-G2 (3.5 g, 4.4 mmol). The reaction mixture was stirred at 105°C for 6 hours under an N2 atmosphere. The resulting mixture was filtered, and the filtrate was concentrated under reduced pressure. The residue was purified by preparative TLC chromatography to obtain the desired product (55 g, 73% yield) as a solid. LCMS(ESI) m / z:[M+H]C 45 H 64 Calculated value for N4O7Si: 800.5; Measured value: 801.5.

[0480] Step 7: Synthesis of (2S)-2-{[(benzyloxy)carbonyl]amino}-3-[(2S)-4-(3-{3-[(tert-butyldimethylsilyl)oxy]-2,2-dimethylpropyl}-1-ethyl-2-{2-[(1S)-1-methoxyethyl]pyridine-3-yl}indole-5-yl)morpholine-2-yl]propanoic acid At room temperature, methyl(2S)-2-{[(benzyloxy)carbonyl]amino}-3-[(2S)-4-(3-{3-[(tert-butyldimethylsilyl)oxy]-2,2-dimethylpropyl}-1-ethyl-2-{2-[(1S)-1-methoxyethyl]pyridine-3-yl}indole-5-yl)morpholin-2-yl]propanoate (10 g, 12 mmol) was dissolved in THF (270 mL) to which an aqueous solution of LiOH (1.3 g, 31 mmol) (45 mL) was added. The reaction mixture was stirred at room temperature for 2 hours, and then treated with 1N HCl at 0-5°C to adjust the pH to 4-5. The resulting mixture was extracted with ELISA (2 × 50 mL). The combined organic layers were washed with brine and dried over anhydrous Na₂SO₄. The organic phase was then concentrated under reduced pressure to obtain the product (9.5 g, 97% yield) as a solid. LCMS(ESI) m / z:[M+H]C 44 H 62 Calculated value for N4O7Si: 786.4; measured value: 787.4.

[0481] Step 8: Synthesis of methyl(S)-1-((S)-2-(((benzyloxy)carbonyl)amino)-3-((S)-4-(3-(3-((tert-butyldimethylsilyl)oxy)-2,2-dimethylpropyl)-1-ethyl-2-(2-((S)-1-methoxyethyl)pyridine-3-yl)-1H-indole-5-yl)morpholine-2-yl)propanoyl)hexahydropyridazine-3-carboxylate (2S)-2-{[(benzyloxy)carbonyl]amino}-3-[(2S)-4-(3-{3-[(tert-butyldimethylsilyl)oxy]-2,2-dimethylpropyl}-1-ethyl-2-{2-[(1S)-1-methoxyethyl]pyridine-3-yl}indole-5-yl)morpholin-2-yl]propanoic acid (10 g, 12.7 mmol) was mixed with DMF (150 mL) and methyl(S)-hexahydropyridazine-3-carboxylate (2 g, 14 mmol). The mixture was then cooled to 0°C, and DIPEA (32.8 g, 254 mmol), followed by HATU (9.7 g, 25.4 mmol), was added at 0-5°C. The reaction mixture was stirred at 0-5°C for 1 hour. The resulting mixture was diluted with ELISA (500 mL) and H2O (200 mL). The organic layer was separated and washed with H2O (2 × 100 mL) and brine (100 mL), and dried over anhydrous sodium sulfate. The solution was filtered and concentrated under reduced pressure, and the residue was purified by silica gel column chromatography to obtain the product. LCMS(ESI) m / z:[M+H]C 50 H 72 Calculated value for N6O8Si: 912.5; measured value: 913.4.

[0482] Step 9: Synthesis of methyl(S)-1-((S)-2-(((benzyloxy)carbonyl)amino)-3-((S)-4-(1-ethyl-3-(3-hydroxy-2,2-dimethylpropyl)-2-(2-((S)-1-methoxyethyl)pyridine-3-yl)-1H-indole-5-yl)morpholine-2-yl)propanoyl)hexahydropyridazine-3-carboxylate At room temperature, methyl(S)-1-((S)-2-(((benzyloxy)carbonyl)amino)-3-((S)-4-(3-(3-((tert-butyldimethylsilyl)oxy)-2,2-dimethylpropyl)-1-ethyl-2-(2-((S)-1-methoxyethyl)pyridine-3-yl)-1H-indole-5-yl)morpholine-2-yl)propanoyl)hexahydropyridazine-3-carboxylate (8.5g To a solution of 9 mmol of THF (8 mL), tetrabutylammonium fluoride (1 M, 180 mL, 180 mmol in THF) and AcOH (11 g, 200 mmol) were added. The reaction mixture was stirred at 75°C for 3 hours. The resulting mixture was diluted with RINKAN (150 mL) and washed with H2O (6 × 20 mL). The organic phase was concentrated under reduced pressure to obtain the product (7.4 g, 100% yield) as a solid. LC-MS (ESI) m / z: [M + H]C 44 H 58 Calculated value for N6O8: 799.4; Measured value: 798.4.

[0483] Step 10: Synthesis of (S)-1-((S)-2-(((benzyloxy)carbonyl)amino)-3-((S)-4-(1-ethyl-3-(3-hydroxy-2,2-dimethylpropyl)-2-(2-((S)-1-methoxyethyl)pyridine-3-yl)-1H-indole-5-yl)morpholine-2-yl)propanoyl)hexahydropyridazine-3-carboxylic acid A solution of methyl(S)-1-((S)-2-(((benzyloxy)carbonyl)amino)-3-((S)-4-(1-ethyl-3-(3-hydroxy-2,2-dimethylpropyl)-2-(2-((S)-1-methoxyethyl)pyridine-3-yl)-1H-indole-5-yl)morpholine-2-yl)propanoyl)hexahydropyridazine-3-carboxylate (8 g, 10 mmol) in THF (200 mL) was mixed with an aqueous solution of LiOH (600 mg, 25 mmol) (30 mL). The reaction mixture was stirred at room temperature for 1 hour, then treated with 1N HCl at 0-5°C to adjust the pH to 4-5, and extracted with ELISA (2 × 500 mL). The organic phase was washed with brine and concentrated under reduced pressure to obtain the product (8 g, crude) as a solid. LCMS(ESI) m / z[M+H]C 43 H 56 Calculated value for N6O8: 784.4; measured value: 785.4.

[0484] Step 11: Benzyl ((2 2 S,6 3 S,4S)-1 1 -ethyl-1 2 -(2-((S)-1-methoxyethyl)pyridine-3-yl)-10,10-dimethyl-5,7-dioxo-6 1 ,6 2 ,6 3 ,6 4 ,6 5 ,6 6 -Hexahydro-1 1 Synthesis to obtain H-8-oxa-2(4,2)-morpholina-1(5,3)-indola-6(1,3)-pyridadinacycloundecafan-4-yl)carbamate Under an argon atmosphere at room temperature, (S)-1-((S)-2-(((benzyloxy)carbonyl)amino)-3-((S)-4-(1-ethyl-3-(3-hydroxy-2,2-dimethylpropyl)-2-(2-((S)-1-methoxyethyl)pyridine-3-yl)-1H-indole-5-yl)morpholine-2-yl)propanoyl)hexahydropyridazine-3-carboxylic acid (8 g, 10.2 mmol) and DIPEA (59 g, 459 mmol) were mixed with DCM (800 mL), to which EDCI (88 g, 458 mmol) and HOBt (27.6 g, 204 mmol) were added. The reaction mixture was stirred at room temperature for 16 hours. The resulting mixture was concentrated under reduced pressure, and the residue was purified by silica gel column chromatography to obtain the product (5 g, 66% yield) as a solid. LCMS(ESI) m / z:[M+H]C 43 H 54 The calculated value for N6O7 was 766.4; the measured value was 767.4.

[0485] Step 12:(2 2 S,6 3 S,4S)-4-amino-1 1 -ethyl-1 2 -(2-((S)-1-methoxyethyl)pyridine-3-yl)-10,10-dimethyl-6 1 ,6 2 ,6 3 ,6 4 ,6 5 ,6 6 -Hexahydro-1 1 Synthesis of H-8-oxa-2(4,2)-morpholina-1(5,3)-indola-6(1,3)-pyridadinacycloundecafane-5,7-dione Under an H2 atmosphere, at room temperature, benzyl((2 2 S,63S,4S)-1 1 -ethyl-1 2 -(2-((S)-1-methoxyethyl)pyridine-3-yl)-10,10-dimethyl-5,7-dioxo-6 1 ,6 2 ,6 3 ,6 4 ,6 5 ,6 6-Hexahydro-1 1 To a solution of H-8-oxa-2(4,2)-morpholina-1(5,3)-indola-6(1,3)-pyridadinacycloundecafan-4-yl)carbamate (400 mg, 0.5 mmol) in MeOH (20 mL), Pd / C (200 mg) and ammonium acetate (834 mg, 16 mmol) were added, and the mixture was stirred for 2 hours. The resulting mixture was filtered and concentrated under reduced pressure. The residue was again dissolved in DCM (20 mL) and washed with H2O (5 mL x 2), then concentrated under reduced pressure to obtain the product (320 mg, 97% yield) as a solid. LCMS(ESI) m / z:M+H]C 35 H 48 The calculated value for N6O5 was 632.4; the measured value was 633.3.

[0486] Intermediate 7: tert-butyl((6 3 S,4S,Z)-11-ethyl-1 2 -(2-((S)-1-methoxyethyl)-5-(4-methylpiperazine-1-yl)pyridine-3-yl)-10,10-dimethyl-5,7-dioxo-6 1 ,6 2 ,6 3 ,6 4 ,6 5 ,6 6 -Hexahydro-1 1 Synthesis of H-8-oxa-2(4,2)-thiazola-1(5,3)-indola-6(1,3)-pyridadinacycloundecafan-4-yl)carbamate

[0487] [ka]

[0488] Step 1: Synthesis of (S)-(5-bromo-6-(1-methoxyethyl)pyridine-3-yl)boronic acid Under an argon atmosphere, dtbpy (14.91 g, 55.5 mmol) and chloro(1,5-cyclooctadiene)iridium(I) dimer (7.46 g, 11.1 mmol) were added to a stirred solution of (S)-3-bromo-2-(1-methoxyethyl)pyridine (80.0 g, 370.24 mmol) and bis(pinacolato)diborone (141.03 g, 555.3 mmol) in THF (320 mL). The resulting mixture was stirred at 75°C for 16 hours. The mixture was concentrated under reduced pressure, and the resulting residue was dissolved in siRNA (200 mL) and adjusted to pH 10 with a solution of Na₂CO₃ (40 g) and NaOH (10 g) in H₂O (600 mL). The aqueous layer was extracted with RINKAN (800 mL), and then the aqueous phase was acidified with HCl (6N) to pH 6 to precipitate the desired product (50 g, 52% yield) as a solid. LCMS(ESI) m / z[M+H]C8H 11 Calculated value for BBrNO3: 260.01; Measured value: 260.0.

[0489] Step 2: Synthesis of (S)-3-bromo-5-iodo-2-(1-methoxyethyl)pyridine (S)-(5-bromo-6-(1-methoxyethyl)pyridine-3-yl)boronic acid (23.0 g, 88.5 mmol) was mixed in MeCN (230 mL) with NIS (49.78 g, 221.2 mmol) added at room temperature. The resulting mixture was stirred overnight at 80 °C under an argon atmosphere. The mixture was concentrated under reduced pressure, the residue was dissolved in DCM (2.1 L), and washed with Na2S2O3 (3 × 500 mL). The organic layer was dried over anhydrous Na2SO4 and filtered, and the filtrate was concentrated under reduced pressure. The residue was purified by silica gel column chromatography to obtain the desired product (20 g, 66% yield). Calculated value for LCMS (ESI) m / z [M+H]C8H9BrINO: 341.90; measured value: 341.7.

[0490] Step 3: Synthesis of benzyl(S)-4-(5-bromo-6-(1-methoxyethyl)pyridine-3-yl)piperazine-1-carboxylate In a 3 L three-necked round-bottom flask, purged and maintained under an inert argon atmosphere, 3-bromo-5-iodo-2-[(1S)-1-methoxymethyl]pyridine (147 g, 429.8 mmol), benzylpiperazine-1-carboxylate (94.69 g, 429.8 mmol), Pd(OAc)2 (4.83 g, 21.4 mmol), BINAP (5.35 g, 8.6 mmol), Cs2CO3 (350.14 g, 1074.6 mmol), and toluene (1 L) were added. The resulting solution was stirred overnight in an oil bath at 100°C. The reaction mixture was cooled to room temperature. The resulting mixture was concentrated under reduced pressure. The MeOH / DCM residue was purified by silica gel column chromatography (50% siRNA / hexane) to obtain the product (135 g, 65% yield) as a solid. LCMS(ESI) m / z:[M+H]C 20 H 24 Calculated value for BrN3O3: 433.1; measured value: 434.1.

[0491] Step 4: Synthesis of benzyl(S)-4-(6-(1-methoxyethyl)-5-(4,4,5,5-tetramethyl-1,3,2-dioxaborolan-2-yl)pyridine-3-yl)piperazine-1-carboxylate In a 3 L three-necked round-bottom flask, purged and maintained with an inert argon atmosphere, benzyl 4-[5-bromo-6-[(1S)-1-methoxyethyl]pyridine-3-yl]piperazine-1-carboxylate (135 g, 310.8 mmol), bis(pinacolato)diborone (86.82 g, 341.9 mmol), Pd(dppf)Cl2 (22.74 g, 31.0 mmol), KOAc (76.26 g, 777.5 mmol), and toluene (1 L) were placed. The resulting solution was stirred in an oil bath at 90°C for 2 days. The reaction mixture was cooled to room temperature. The resulting mixture was concentrated under reduced pressure. The residue was purified by neutral alumina column chromatography (30% siRNA / hexane) to obtain the product (167 g, crude) as a solid. LCMS(ESI) m / z:[M+H]C 26 H 36 The calculated value for BN3O5 was 481.3; the measured value was 482.1.

[0492] Step 5: Synthesis of benzyl(S)-4-(5-(5-bromo-3-(3-((tert-butyldiphenylsilyl)oxy)-2,2-dimethylpropyl)-1H-indole-2-yl)-6-(1-methoxyethyl)pyridine-3-yl)piperazine-1-carboxylate In a 3 L three-necked round-bottom flask, which was purged and maintained with an inert argon atmosphere, (S)-4-(6-(1-methoxyethyl)-5-(4,4,5,5-tetramethyl-1,3,2-dioxaborolan-2-yl)pyridine-3-yl)piperazine-1-carboxylate (167 g, 346.9 mmol), 5-bromo-3-[3-[(tert-butyldiphenylsilyl)oxy]-2,2-dimethylpropyl]-2-iodo-1H-indole (224.27 g, 346.9 mmol), Pd(dppf)Cl2 (25.38 g, 34.6 mmol), dioxane (600 mL), H2O (200 mL), K3PO4 (184.09 g, 867.2 mmol), and toluene (200 mL) were added. The resulting solution was stirred overnight in an oil bath at 70°C. The reaction mixture was cooled to room temperature. The resulting mixture was concentrated under reduced pressure. The MeOH / DCM residue was purified by silica gel column chromatography (50% Âxane) to obtain the product (146 g, 48% yield) as a solid. LCMS(ESI) m / z:[M+H]C 49 H 57 Calculated value for BrN4O4Si: 872.3; measured value: 873.3.

[0493] Step 6: Synthesis of benzyl(S)-4-(5-(5-bromo-3-(3-((tert-butyldiphenylsilyl)oxy)-2,2-dimethylpropyl)-1-ethyl-1H-indole-2-yl)-6-(1-methoxyethyl)pyridine-3-yl)piperazine-1-carboxylate Under an N2 atmosphere at 0°C, benzyl(S)-4-(5-(5-bromo-3-(3-((tert-butyldiphenylsilyl)oxy)-2,2-dimethylpropyl)-1H-indole-2-yl)-6-(1-methoxyethyl)pyridine-3-yl)piperazine-1-carboxylate (146 g, 167.0 mmol) and Cs2CO3 (163.28 g, 501.1 mmol) were mixed with DMF (1200 mL) and ethyl iodide (52.11 g, 334.0 mmol) was added in small quantities. The final reaction mixture was stirred at room temperature for 12 hours. The resulting mixture was diluted with ELISA (1 L) and washed with brine (3 × 1.5 L). The organic layer was dried over anhydrous Na2SO4 and filtered. After filtration, the filtrate was concentrated under reduced pressure to obtain the product (143 g, crude) as a solid, which was used directly in the next step without further purification. LCMS(ESI) m / z:[M+H]C 51 H 61 Calculated value for BrN4O4Si: 900.4; measured value: 901.4.

[0494] Step 7: Synthesis of benzyl(S)-4-(5-(5-bromo-1-ethyl-3-(3-hydroxy-2,2-dimethylpropyl)-1H-indole-2-yl)-6-(1-methoxyethyl)pyridine-3-yl)piperazine-1-carboxylate Benzyl(S)-4-(5-(5-bromo-3-(3-((tert-butyldiphenylsilyl)oxy)-2,2-dimethylpropyl)-1-ethyl-1H-indole-2-yl)-6-(1-methoxyethyl)pyridine-3-yl)piperazine-1-carboxylate (143 g, 158.5 mmol) was mixed with DMF (1250 mL) and CsF (72.24 g, 475.5 mmol) was added. The reaction mixture was then stirred at 60 °C for 2 days under an N2 atmosphere. The resulting mixture was diluted with RINKAN (1 L) and washed with brine (3 × 1 L). The organic phase was then concentrated under reduced pressure. The residue was purified by silica gel column chromatography (30% RINKAN / petroleum ether) to obtain two atropisomers A (38 g, 36% yield) and B (34 g, 34% yield), both as solids. LCMS(ESI) m / z:[M+H]C35 H 43 Calculated value for BrN4O4: 663.2; measured value: 662.2.

[0495] Step 8: Synthesis of benzyl(S)-4-(5-(1-ethyl-3-(3-hydroxy-2,2-dimethylpropyl)-5-(4,4,5,5-tetramethyl-1,3,2-dioxaborolan-2-yl)-1H-indole-2-yl)-6-(1-methoxyethyl)pyridine-3-yl)piperazine-1-carboxylate In a 500 mL three-necked round-bottom flask purged and maintained under an inert nitrogen atmosphere, benzyl(S)-4-(5-(5-bromo-1-ethyl-3-(3-hydroxy-2,2-dimethylpropyl)-1H-indole-2-yl)-6-(1-methoxyethyl)pyridine-3-yl)piperazine-1-carboxylate (14 g, 21.1 mmol), bis(pinacolato)diborone (5.89 g, 23.21 mmol), Pd(dppf)Cl2 (1.54 g, 2.1 mmol), KOAc (5.18 g, 52.7 mmol), and toluene (150 mL) were placed. The resulting solution was stirred in an oil bath at 90°C for 5 hours. After the reaction mixture was cooled to room temperature, it was concentrated under reduced pressure. The residue was purified by silica gel column chromatography (30% siRNA / petroleum ether) to obtain the product (12 g, 76% yield) as a solid. LCMS(ESI) m / z:[M+H]C 41 H 55 Calculated value for BN4O6: 710.4; measured value: 711.3.

[0496] Step 9: Synthesis of methyl(S)-1-((S)-3-(4-(2-(5-(4-((benzyloxy)carbonyl)piperazin-1-yl)-2-((S)-1-methoxyethyl)pyridine-3-yl)-1-ethyl-3-(3-hydroxy-2,2-dimethylpropyl)-1H-indole-5-yl)thiazole-2-yl)-2-((tert-butoxycarbonyl)amino)propanoyl)hexahydropyridazine-3-carboxylate In a 250 mL round-bottom flask purged and maintained under an inert argon atmosphere, benzyl(S)-4-(5-(1-ethyl-3-(3-hydroxy-2,2-dimethylpropyl)-5-(4,4,5,5-tetramethyl-1,3,2-dioxaborolan-2-yl)-1H-indole-2-yl)-6-(1-methoxyethyl)pyridine-3-yl)piperazine-1-carboxylate (10.8 g, 15.2 mmol), methyl (3S)-1-[(2S)-3-(4-bromo-1,3-thiazole-2-yl)-2-[(tert-butoxycarbonyl)amino]propanoyl]-1,2-diadinane-3-carboxylate (7.98 g, 16.7 mmol), Pd(dtbpf)Cl2 (0.99 g, 1.52 mmol), K3PO4 (8.06 g, 37.9 mmol), toluene (60 mL), dioxane (20 mL), and H2O (20 mL) were added. The resulting solution was stirred in an oil bath at 70°C for 3 hours. The reaction mixture was cooled to room temperature. The resulting solution was extracted with ELISA (2 × 50 mL) and concentrated under reduced pressure. The residue was purified by silica gel column chromatography (10% ELISA / hexane). The solvent was removed under reduced pressure to obtain the product (8 g, 51% yield) as a solid. LCMS(ESI) m / z[M+H]C 52 H 68 Calculated value for N8O9S: 980.5; Measured value: 980.9.

[0497] Step 10: Synthesis of (S)-1-((S)-3-(4-(2-(5-(4-((benzyloxy)carbonyl)piperazin-1-yl)-2-((S)-1-methoxyethyl)pyridine-3-yl)-1-ethyl-3-(3-hydroxy-2,2-dimethylpropyl)-1H-indole-5-yl)thiazole-2-yl)-2-((tert-butoxycarbonyl)amino)propanoyl)hexahydropyridazine-3-carboxylic acid To a stirred mixture of methyl(S)-1-((S)-3-(4-(2-(5-(4-((benzyloxy)carbonyl)piperazin-1-yl)-2-((S)-1-methoxyethyl)pyridine-3-yl)-1-ethyl-3-(3-hydroxy-2,2-dimethylpropyl)-1H-indole-5-yl)thiazole-2-yl)-2-((tert-butoxycarbonyl)amino)propanoyl)hexahydropyridazine-3-carboxylate (12 g, 12.23 mmol) in THF (100 mL) / H2O (100 mL), LiOH (2.45 g, 61.1 mmol) was added under an N2 atmosphere, and the resulting mixture was stirred at room temperature for 2 hours. The THF was removed under reduced pressure. At 0°C, the pH of the aqueous phase was acidified to 5 with 1N HCl. The aqueous layer was extracted by DCM (3 × 100 mL). The organic phase was concentrated under reduced pressure to obtain the product (10 g, 85% yield) as a solid. LC-MS(ESI) m / z[M+H]C 51 H 66 Calculated value for N8O9S: 966.5; Measured value: 967.0.

[0498] Step 11: Benzyl 4-(5-((6 3 S,4S,Z)-4-((tert-butoxycarbonyl)amino)-1 1 -ethyl-10,10-dimethyl-5,7-dioxo-6 1 ,6 2 ,6 3 ,6 4 ,6 5 ,6 6 -Hexahydro-1 1 H-8-Oxa-2(4,2)-Thiazola-1(5,3)-Indola-6(1,3)-Pyridadinacycloundecaphane-1 2 Synthesis of (-yl)-6-((S)-1-methoxyethyl)pyridine-3-yl)piperazine-1-carboxylate (S)-1-((S)-3-(4-(2-(5-(4-((benzyloxy)carbonyl)piperazin-1-yl)-2-((S)-1-methoxyethyl)pyridine-3-yl)-1-ethyl-3-(3-hydroxy-2,2-dimethylpropyl)-1H-indole-5-yl)thiazole-2-yl)-2-((tert-butoxycarbonyl)amino)propanoyl)hexahydropyridazine-3-carboxylic acid (18 g, 18.61 mmol), MeCN (1.8 L), DIPEA (96.21 g, 744.4 mmol), EDCI (107.03 g, 558.3 ​​mmol), and HOBt (25.15 g, 186.1 mmol) were placed in a 3 L round-bottom flask purged and maintained with an inert nitrogen atmosphere. The obtained solution was stirred at room temperature and then concentrated under reduced pressure. The resulting solution was diluted with DCM (1 L) and washed with 1 M HCl (3 × 1 L) and H2O (3 × 1 L). Next, the organic layer was concentrated under reduced pressure and purified by silica gel column chromatography (50% HCl / hexane) to obtain the product (10.4 g, 55% yield) as a solid. LCMS (ESI) m / z:[M+H]C 51 H 64 Calculated value for N8O8S: 948.5; Measured value: 949.3.

[0499] Step 12: tert-butyl((6 3 S,4S,Z)-11-ethyl-1 2 -(2-((S)-1-methoxyethyl)-5-(piperazin-1-yl)pyridine-3-yl)-10,10-dimethyl-5,7-dioxo-6 1 ,6 2 ,6 3 ,6 4 ,6 5 ,6 6 -Hexahydro-1 1 Synthesis of H-8-oxa-2(4,2)-thiazola-1(5,3)-indola-6(1,3)-pyridadinacycloundecafan-4-yl)carbamate In a 250 mL round-bottom flask purged and maintained under a nitrogen-reducing atmosphere, benzyl 4-(5-((6 3S,4S,Z)-4-((tert-butoxycarbonyl)amino)-1 1 -ethyl-10,10-dimethyl-5,7-dioxo-6 1 ,6 2 ,6 3 ,6 4 ,6 5 ,6 6 -Hexahydro-1 1 H-8-Oxa-2(4,2)-Thiazola-1(5,3)-Indola-6(1,3)-Pyridadinacycloundecaphane-1 2 (-yl)-6-((S)-1-methoxyethyl)pyridine-3-yl)piperazine-1-carboxylate (10.40 g, 10.9 mmol), Pd(OH)2 / C (5 g, 46.9 mmol), and MeOH (100 mL) were added. The resulting solution was stirred at room temperature for 3 hours under a 2 atm H2 atmosphere. The solid was filtered, and the filtrate was washed with MeOH (3 × 100 mL). The combined organic phase was concentrated under reduced pressure to obtain the product (8.5 g, 95% yield) as a solid. LC-MS(ESI) m / z[M+H]C 43 H 58 Calculated value for N8O6S: 814.4; Measured value: 815.3.

[0500] Step 13: tert-butyl((6 3 S,4S,Z)-1 1 -ethyl-1 2 -(2-((S)-1-methoxyethyl)-5-(4-methylpiperazine-1-yl)pyridine-3-yl)-10,10-dimethyl-5,7-dioxo-6 1 ,6 2 ,6 3 ,6 4 ,6 5 ,6 6 -Hexahydro-1 1 Synthesis of H-8-oxa-2(4,2)-thiazola-1(5,3)-indola-6(1,3)-pyridadinacycloundecafan-4-yl)carbamate In a 1000 mL round-bottom flask purged and maintained under a nitrogen-reducing atmosphere, tert-butyl ((6 3 S,4S,Z)-1 1 -ethyl-12 -(2-((S)-1-methoxyethyl)-5-(piperazin-1-yl)pyridine-3-yl)-10,10-dimethyl-5,7-dioxo-6 1 ,6 2 ,6 3 ,6 4 ,6 5 ,6 6 -Hexahydro-1 1 H-8-oxa-2(4,2)-thiazola-1(5,3)-indola-6(1,3)-pyridadinacycloundecafan-4-yl)carbamate (8.5 g, 10.4 mmol), MeOH (100 mL), and AcOH (1.88 g, 31.2 mmol) were added. After stirring the solution for 15 minutes, HCHO (1.88 g, 23.15 mmol, 37% aqueous solution) and NaBH3CN (788 mg, 12.5 mL) were added at room temperature. The resulting solution was stirred for 3 hours. Next, the mixture was quenched with H2O (100 mL) and concentrated under reduced pressure to remove MeOH. The resulting solution was diluted with DCM (300 mL) and washed with H2O (3 × 100 mL). The solution was concentrated under reduced pressure to obtain the product (8.2 g, 90% yield) as a solid. LCMS(ESI) m / z[M+H]C 44 H 60 Calculated value for N8O6S: 828.4; Measured value: 829.3.

[0501] Intermediate 8:(6 3 S,4S,Z)-4-amino-1 1 -ethyl-1 2 -(5-((S)-Hexahydropyridazino[2,1-c][1,4]oxazin-8(1H)-yl)-2-((S)-1-methoxyethyl)pyridine-3-yl)-10,10-dimethyl-6 1 ,6 2 ,6 3 ,6 4 ,6 5 ,6 6 -Hexahydro-1 1 Synthesis of H-8-oxa-2(4,2)-thiazola-1(5,3)-indola-6(1,3)-pyridadinacycloundecafane-5,7-dione

[0502] [ka]

[0503] Step 1: Synthesis of (S)-3-(5-bromo-1-ethyl-2-(2-(1-methoxyethyl)pyridine-3-yl)-1H-indole-3-yl)-2,2-dimethylpropyl acetate At 0°C under an argon atmosphere, (S)-3-(5-bromo-1-ethyl-2-(2-(1-methoxyethyl)pyridine-3-yl)-1H-indol-3-yl)-2,2-dimethylpropan-1-ol (100 g, 224.517 mmol) and Et3N (45.44 g, 449.034 mmol) were mixed in 1 L of DCM, to which DMAP (2.74 g, 22.452 mmol) and Ac2O (27.50 g, 269.420 mmol) were added in small quantities. The resulting mixture was stirred at room temperature for 3 hours. The resulting mixture was concentrated under reduced pressure and then diluted with 1000 mL of ethyl acetate. The resulting mixture was washed with 500 mL of 1 M HCl, then washed with 500 mL of saturated NaHCO3 and brine, and dried over anhydrous Na2SO4. After filtration, the filtrate was concentrated under reduced pressure. The residue was purified by trituration with petroleum ether (500 mL) to obtain the product (93.3 g, 85% yield) as a white solid. LCMS(ESI) m / z[M+H]C 25 H 31 Calculated value for BrN2O3: 487.16; Measured value: 489.2.

[0504] Step 2: Synthesis of (S)-(5-(3-(3-acetoxy-2,2-dimethylpropyl)-5-bromo-1-ethyl-1H-indole-2-yl)-6-(1-methoxyethyl)pyridine-3-yl)boronic acid At room temperature under an argon atmosphere, (S)-3-(5-bromo-1-ethyl-2-(2-(1-methoxyethyl)pyridine-3-yl)-1H-indole-3-yl)-2,2-dimethylpropyl acetate (93.3 g, 191.409 mmol) and B2PIN2 (72.91 g, 287.113 mmol) were mixed in 370 mL of THF. dtbpy (7.71 g, 28.711 mmol) and chloro(1,5-cyclooctadiene)iridium(I) dimer (6.43 g, 9.570 mmol) were added in small quantities. The resulting mixture was stirred overnight at 75°C. The mixture was concentrated under reduced pressure, and the product (190 g, crude) was obtained as oil. LCMS(ESI) m / z:[M+H]C 25 H 32 Calculated value for BBrN2O5: 531.17; Measured value: 533.3.

[0505] Step 3: Synthesis of (S)-3-(5-bromo-1-ethyl-2-(5-iodo-2-(1-methoxyethyl)pyridine-3-yl)-1H-indole-3-yl)-2,2-dimethylpropyl acetate At 0°C under an air atmosphere, 110 g (207.059 mmol) of (S)-(5-(3-(3-acetoxy-2,2-dimethylpropyl)-5-bromo-1-ethyl-1H-indole-2-yl)-6-(1-methoxyethyl)pyridine-3-yl)boronic acid and chloramine-T trihydrate (349.96 g (1242.354 mmol) were added in 550 mL of THF, in small quantities, to a 225 mL aqueous solution of NaI (186.22 g (1242.354 mmol)). The resulting mixture was stirred overnight at 50°C under an argon atmosphere. The resulting mixture was concentrated under reduced pressure and washed with CHCl3 (500 mL). The resulting mixture was filtered, and the filtrate was washed with CHCl3 (3 × 250 mL). The filtrate was extracted with CHCl3 (3 × 500 mL). The combined organic layers were washed with Na2S2O3 (500 mL), then with brine (2 × 200 mL), and dried over anhydrous Na2SO4. After filtration, the filtrate was concentrated under reduced pressure. The residue was purified by silica gel column chromatography (18% siRNA / petroleum ether) to obtain the product (24 g, 18% yield) as a solid. LCMS(ESI) m / z:[M+H]C 25 H 30 Calculated value for BrIN2O3: 613.06; Measured value: 614.7.

[0506] Step 4: Synthesis of 3-(5-bromo-1-ethyl-2-(5-((S)-hexahydropyridazino[2,1-c][1,4]oxazin-8(1H)-yl)-2-((S)-1-methoxyethyl)pyridine-3-yl)-1H-indole-3-yl)-2,2-dimethylpropyl acetate At room temperature under an argon atmosphere, 3-(5-bromo-1-ethyl-2-{5-iodo-2-[(1S)-1-methoxyethyl]pyridine-3-yl}indole-3-yl)-2,2-dimethylpropyl acetate (9 g, 14.674 mmol), (S)-octahydropyrazino[2,1-c][1,4]oxazine (2.469 g, 17.609 mmol), Cs2CO3 (11.953 g, 36.685 mmol), and BINAP (456.9 mg, 0.734 mmol) were stirred, to which Pd(OAc)2 (329.44 mg, 1.467 mmol) was added. The resulting mixture was stirred at 6°C for 100 hours. After filtration, the filtrate was washed with ELISA (100 mL). The filtrate was concentrated under reduced pressure. The residue was purified by preparative TLC (8% MeOH / DCM) to obtain the product (6.9 g, 75% yield) as a solid. LC-MS (ESI) m / z:[M+H]C 32 H 43 Calculated value for BrN4O4: 627.25; measured value: 627.4.

[0507] Step 5: Synthesis of 3-(1-ethyl-2-(5-((S)-hexahydropyridazino[2,1-c][1,4]oxazin-8(1H)-yl)-2-((S)-1-methoxyethyl)pyridine-3-yl)-5-(4,4,5,5-tetramethyl-1,3,2-dioxaborolan-2-yl)-1H-indole-3-yl)-2,2-dimethylpropyl acetate At room temperature under an argon atmosphere, 3-(5-bromo-1-ethyl-2-(5-((S)-hexahydropyridazino[2,1-c][1,4]oxazin-8(1H)-yl)-2-((S)-1-methoxyethyl)pyridine-3-yl)-1H-indole-3-yl)-2,2-dimethylpropyl acetate (3.2 g, 5.115 mmol), KOAc (1.51 g, 15.345 mmol), and 4,4,5,5-tetramethyl-2-(tetramethyl-1,3,2-dioxaborolan-2-yl)-1,3,2-dioxaborolan (2.60 g, 10.230 mmol) were stirred in toluene (48 mL), to which Pd(dppf)Cl2 (0.37 g, 0.512 mmol) was added in small quantities. The resulting mixture was stirred at 90°C for 1.5 hours. The mixture was filtered, and the filtrate was washed with dimethyl acetate (100 mL). The filtrate was concentrated under reduced pressure. The residue was purified by preparative TLC (8% MeOH / DCM) to obtain the product (3.0 g, 88% yield) as a solid. LC-MS (ESI) m / z:[M+H]C 38 H 55 Calculated value for BN4O6: 675.43; Measured value: 675.1.

[0508] Step 6: Synthesis of methyl(S)-1-((S)-3-(4-(3-(3-acetoxy-2,2-dimethylpropyl)-1-ethyl-2-(5-((S)-hexahydropyridazino[2,1-c][1,4]oxazin-8(1H)-yl)-2-((S)-1-methoxyethyl)pyridine-3-yl)-1H-indole-5-yl)thiazole-2-yl)-2-((tert-butoxycarbonyl)amino)propanoyl)hexahydropyridazin-3-carboxylate At room temperature under an argon atmosphere, 3-(1-ethyl-2-(5-((S)-hexahydropyridazino[2,1-c][1,4]oxazine-8(1H)-yl)-2-((S)-1-methoxyethyl)pyridine-3-yl)-5-(4,4,5,5-tetramethyl-1,3,2-dioxaborolan-2-yl)-1H-indole-3-yl)-2,2-dimethylpropyl acetate (5 g, 7.433 mmol) and K3PO4 (4.26 g, 20.067 mmol) were stirred with toluene (54 mL), to which dioxane (17.82 mL, 210.307 mmol) and H2O (17.82 mL) were added. The resulting mixture was stirred at 70°C for 2 hours. The resulting mixture was filtered, and the filtrate was washed with ELISA (100 mL). The filtrate was concentrated under reduced pressure. The resulting mixture was extracted with ethyl acetate (200 mL). The combined organic layers were washed with brine (100 mL) and dried over anhydrous sodium 2SO4. After filtration, the filtrate was concentrated under reduced pressure. The residue was purified by preparative TLC (8% MeOH / DCM) to obtain the product (4.6 g, 66% yield) as a solid. LC-MS (ESI) m / z:[M+H]C 49 H 68 Calculated value for N8O9S: 945.49; Measured value: 945.7.

[0509] Step 7: Synthesis of (S)-1-((S)-2-((tert-butoxycarbonyl)amino)-3-(4-(1-ethyl-2-(5-((S)-hexahydropyridazino[2,1-c][1,4]oxazin-8(1H)-yl)-2-((S)-1-methoxyethyl)pyridine-3-yl)-3-(3-hydroxy-2,2-dimethylpropyl)-1H-indole-5-yl) and azole-2-yl)propanoyl)hexahydropyridazin-3-carboxylic acid At 0°C, methyl(S)-1-((S)-3-(4-(3-(3-acetoxy-2,2-dimethylpropyl)-1-ethyl-2-(5-((S)-hexahydropyridazino[2,1-c][1,4]oxazin-8(1H)-yl)-2-((S)-1-methoxyethyl)pyridine-3-yl)-1H-indole-5-yl)thiazole-2-yl)-2-((tert-butoxycarbonyl)amino)propanoyl)hexahydropyridazin-3-carboxylate (6 g, 6.361 mmol) was mixed with THF (43 mL) and LiOH·H2O (573.92 mg, 13.677 mmol) was added. The resulting mixture was stirred at room temperature for 16 hours. The mixture was acidified to pH 6 with hydrochloric acid. The resulting mixture was extracted with ELISA (3 × 100 mL). The combined organic layers were washed with brine and dried over anhydrous Na2SO4. After filtration, the filtrate was concentrated under reduced pressure to obtain the product (4 g, crude) as a solid. LC-MS(ESI) m / z:[M+H]C 45 H 60 Calculated value for N8O9S: 889.43; Measured value: 889.7.

[0510] Step 8: tert-butyl((6 3 S,4S,Z)-1 1 -ethyl-1 2 -(5-((S)-Hexahydropyridadino[2,1-c][1,4]oxazin-8(1H)-yl)-2-((S)-1-methoxyethyl)pyridine-3-yl)-10,10-dimethyl-5,7-dioxo-61,6 2 ,6 3 ,6 4 ,6 5 ,6 6 -Hexahydro-1 1 Synthesis of H-8-oxa-2(4,2)-thiazola-1(5,3)-indola-6(1,3)-pyridadinacycloundecafan-4-yl)carbamate At 0°C under an argon atmosphere, (S)-1-((S)-2-((tert-butoxycarbonyl)amino)-3-(4-(1-ethyl-2-(5-((S)-hexahydropyridazino[2,1-c][1,4]oxazin-8(1H)-yl)-2-((S)-1-methoxyethyl)pyridine-3-yl)-3-(3-hydroxy-2,2-dimethylpropyl)-1H-yl A mixture of dole-5-yl) and azole-2-yl)propanoyl)hexahydropyridazine-3-carboxylic acid (4 g, 4.51 mmol), HOBt (6.09 g, 45.09 mmol), and DIPEA (23.31 g, 180.36 mmol) in 200 mL of DCM was stirred, and a solution of EDCI (25.93 g, 135.27 mmol) in 200 mL of DCM was added dropwise. The resulting mixture was stirred at room temperature for 16 hours and then concentrated under reduced pressure. The reaction was quenched with H2O at 0°C and extracted with siRNA (500 mL). The combined organic layer was washed with brine and dried over anhydrous Na2SO4. After filtration, the filtrate was concentrated under reduced pressure. The residue was purified by preparative TLC (8% MeOH / DCM) to obtain the product (2.0 g, 52% yield) as a solid. LCMS(ESI) m / z:[M+H]C 46 H 62 Calculated value for N8O7S: 870.4; Measured value: 871.8.

[0511] Step 9:(6 3 S,4S,Z)-4-amino-1 1 -ethyl-1 2 -(5-((S)-Hexahydropyridadino[2,1-c][1,4]oxazin-8(1H)-yl)-2-((S)-1-methoxyethyl)pyridine-3-yl)-10,10-dimethyl-61,6 2 ,6 3 ,6 4 ,6 5 ,6 6 -Hexahydro-1 1 Synthesis of H-8-oxa-2(4,2)-thiazola-1(5,3)-indola-6(1,3)-pyridadinacycloundecafane-5,7-dione At 0°C, under an argon atmosphere, tert-butyl((6 3 S,4S,Z)-11 -ethyl-1 2 -(5-((S)-Hexahydropyridadino[2,1-c][1,4]oxazin-8(1H)-yl)-2-((S)-1-methoxyethyl)pyridine-3-yl)-10,10-dimethyl-5,7-dioxo-61,6 2 ,6 3 ,6 4 ,6 5 ,6 6 -Hexahydro-1 1 To a stirred solution of H-8-oxa-2(4,2)-thiazola-1(5,3)-indola-6(1,3)-pyridadinacycloundecafan-4-yl)carbamate (316 mg, 0.345 mmol) in DCM (3 mL), TFA (1 mL) was added dropwise. The resulting mixture was stirred at room temperature for 2 hours. The mixture was neutralized to pH 8 with saturated NaHCO3. The resulting mixture was extracted with ELISA (3 × 80 mL). The combined organic layers were washed with brine (3 × 40 mL) and dried over anhydrous Na2SO4. After filtration, the filtrate was concentrated under reduced pressure. The crude product mixture was used directly in the next step without further purification. LCMS(ESI) m / z:[M+H]C 41 H 54 Calculated value for N8O5S: 771.4; Measured value: 771.6.

[0512] Intermediate 9:(6 3 S,4S,Z)-4-amino-1 1 -ethyl-1 2 -(2-((S)-1-methoxyethyl)-5-((R)-octahydro-2H-pyrido[1,2-a]pyrazine-2-yl)pyridine-3-yl)-10,10-dimethyl-6 1 ,6 2 ,6 3 ,6 4 ,6 5 ,6 6 -Hexahydro-1 1 Synthesis of H-8-oxa-2(4,2)-thiazola-1(5,3)-indola-6(1,3)-pyridadinacycloundecafane-5,7-dione

[0513] [ka]

[0514] Step 1: Synthesis of 3-(5-bromo-1-ethyl-2-(2-((S)-1-methoxyethyl)-5-((R)-octahydro-2H-pyrido[1,2-a]pyrazine-2-yl)pyridine-3-yl)-1H-indole-3-yl)-2,2-dimethylpropyl acetate At room temperature under an argon atmosphere, 3-(5-bromo-1-ethyl-2-{5-iodo-2-[(1S)-1-methoxyethyl]pyridine-3-yl}indole-3-yl)-2,2-dimethylpropyl acetate (9 g, 14.674 mmol), (R)-octahydro-2H-pyrido[1,2-a]pyrazine (2.469 g, 17.609 mmol), Cs2CO3 (11.9523 g, 36.685 mmol), and BINAP (456.85 mg, 0.734 mmol) were stirred in toluene (63 mL), to which Pd(OAc)2 (329.44 mg, 1.467 mmol) was added in small quantities. The resulting mixture was stirred at 100°C for 6 hours, then filtered, and the filtrate was washed with ELISA (100 mL). The filtrate was concentrated under reduced pressure. The residue was purified by preparative TLC (8% MeOH / DCM) to obtain the product (6 g, 65% yield) as a solid. LC-MS (ESI) m / z:[M+H]C 33 H 45 Calculated value for BrN4O3: 625.28; Measured value: 627.4.

[0515] Step 2: Synthesis of 3-(1-ethyl-2-(2-((S)-1-methoxyethyl)-5-((R)-octahydro-2H-pyrido[1,2-a]pyrazine-2-yl)pyridine-3-yl)-5-(4,4,5,5-tetramethyl-1,3,2-dioxaborolan-2-yl)-1H-indole-3-yl)-2,2-dimethylpropyl acetate At room temperature under an argon atmosphere, 3-(5-bromo-1-ethyl-2-(2-((S)-1-methoxyethyl)-5-((R)-octahydro-2H-pyrido[1,2-a]pyrazine-2-yl)pyridine-3-yl)-1H-indole-3-yl)-2,2-dimethylpropyl acetate (3.2 g, 5.115 mmol), KOAc (1.51 g, 15.345 mmol), and 4,4,5,5-tetramethyl-2-(tetramethyl-1,3,2-dioxaborolan-2-yl)-1,3,2-dioxaborolan (2.60 g, 10.230 mmol) were stirred in toluene (48 mL), to which Pd(dppf)Cl2 (0.37 g, 0.512 mmol) was added in small quantities. The resulting mixture was stirred at 90°C for 1.5 hours. The mixture was filtered, and the filtrate was washed with ELISA (100 mL). The filtrate was concentrated under reduced pressure and purified by preparative TLC (8% MeOH / DCM) to obtain the product (3.1 g, 81% yield) as a solid. LCMS(ESI) m / z:[M+H]C 39 H 57 Calculated value for BN4O5: 673.45; Measured value: 673.4.

[0516] Step 3: Synthesis of methyl(S)-1-((S)-3-(4-(3-(3-acetoxy-2,2-dimethylpropyl)-1-ethyl-2-(2-((S)-1-methoxyethyl)-5-((R)-octahydro-2H-pyrido[1,2-a]pyrazine-2-yl)pyridine-3-yl)-1H-indole-5-yl)thiazole-2-yl)-2-((tert-butoxycarbonyl)amino)propanoyl)hexahydropyridazine-3-carboxylate At room temperature under an argon atmosphere, 3-(1-ethyl-2-(2-((S)-1-methoxyethyl)-5-((R)-octahydro-2H-pyrido[1,2-a]pyrazine-2-yl)pyridine-3-yl)-5-(4,4,5,5-tetramethyl-1,3,2-dioxaborolan-2-yl)-1H-indole-3-yl)-2,2-dimethylpropyl acetate (5g, 7.433 mmol), methyl(S)-1-((S)-3 -(4-bromothiazole-2-yl)-2-((tert-butoxycarbonyl)amino)propanoyl)hexahydropyridazine-3-carboxylate (3.89 g, 8.176 mmol) and K3PO4 (4.26 g, 20.067 mmol) were mixed with toluene (54 mL), dioxane (18 mL), and H2O (18 mL) and Pd(dtbpf)Cl2 (969 mg, 1.486 mmol) was added. The resulting mixture was stirred at 70°C for 2 hours. The mixture was filtered and the filtrate was washed with ethyl acetate (100 mL). The filtrate was concentrated under reduced pressure and the resulting mixture was extracted with ethyl acetate (200 mL). The combined organic layers were washed with brine (100 mL) and dried over anhydrous sodium 2SO4. After filtration, the filtrate was concentrated under reduced pressure. The residue was purified by preparative TLC (8% MeOH / DCM) to obtain the product (6.8 g, 83% yield) as a solid. LC-MS (ESI) m / z:[M+H]C 50 H 70 Calculated value for N8O8S: 943.51; Measured value: 943.4.

[0517] Step 4: Synthesis of (S)-1-((S)-2-((tert-butoxycarbonyl)amino)-3-(4-(1-ethyl-3-(3-hydroxy-2,2-dimethylpropyl)-2-(2-((S)-1-methoxyethyl)-5-((R)-octahydro-2H-pyrido[1,2-a]pyrazine-2-yl)pyridine-3-yl)-1H-indole-5-yl)thiazole-2-yl)propanoyl)hexahydropyridazine-3-carboxylic acid At 0°C under an argon atmosphere, methyl(S)-1-((S)-3-(4-(3-(3-acetoxy-2,2-dimethylpropyl)-1-ethyl-2-(2-((S)-1-methoxyethyl)-5-((R)-octahydro-2H-pyrido[1,2-a]pyrazine-2-yl)pyridine-3-yl)-1H-indole-5-yl)thiazole-2-yl)-2-((tert-butoxycarbonyl)amino)propanoyl)hexahydropyridazine-3-carboxylate (6 g, 6.361 mmol) was stirred in THF (43 mL) and LiOH·H2O (573.92 mg, 13.677 mmol) was added. The resulting mixture was stirred at room temperature for 16 hours. The mixture was acidified to pH 6 with hydrochloric acid. The resulting mixture was extracted with ELISA (3 × 150 mL). The combined organic layers were washed with brine and dried over anhydrous Na2SO4. After filtration, the filtrate was concentrated under reduced pressure to obtain the product (4 g, crude) as a solid. LC-MS(ESI) m / z:[M+H]C 47 H 66 Calculated value for N8O7S: 887.49; Measured value: 887.6.

[0518] Step 5: tert-butyl((6 3 S,4S,Z)-1 1 -ethyl-1 2 -(2-((S)-1-methoxyethyl)-5-((R)-octahydro-2H-pyrido[1,2-a]pyrazine-2-yl)pyridine-3-yl)-10,10-dimethyl-5,7-dioxo-6 1 ,6 2 ,6 3 ,6 4 ,6 5 ,6 6 -Hexahydro-1 1 Synthesis of H-8-oxa-2(4,2)-thiazola-1(5,3)-indola-6(1,3)-pyridadinacycloundecafan-4-yl)carbamate At 0°C under an argon atmosphere, (S)-1-((S)-2-((tert-butoxycarbonyl)amino)-3-(4-(1-ethyl-3-(3-hydroxy-2,2-dimethylpropyl)-2-(2-((S)-1-methoxyethyl)-5-((R)-octahydro-2H-pyrido[1,2-a]pyrazine-2-yl)pyridine-3-yl)-1H-indole-5- A stirring solution of yl)thiazole-2-yl)propanoyl)hexahydropyridazine-3-carboxylic acid (4 g, 4.509 mmol), HOBt (6.09 g, 45.090 mmol), and DIPEA (23.31 g, 180.360 mmol) in DCM (200 mL) was added dropwise to a stirring solution of EDCI (25.93 g, 135.270 mmol) in DCM (200 mL). The resulting mixture was stirred at room temperature for 16 hours. The resulting mixture was concentrated under reduced pressure, quenched with H2O at 0°C, and extracted with siRNA (500 mL). The combined organic layers were washed with brine and dried over anhydrous Na2SO4. After filtration, the filtrate was concentrated under reduced pressure. The residue was purified by preparative TLC (8% MeOH / DCM) to obtain the product (2.0 g, 49% yield) as a solid. LCMS(ESI) m / z:[M+H]C 47 H 64 Calculated value for N8O6S: 869.47; measured value: 869.8.

[0519] Step 6:(6 3 S,4S,Z)-4-amino-1 1 -ethyl-1 2 -(2-((S)-1-methoxyethyl)-5-((R)-octahydro-2H-pyrido[1,2-a]pyrazine-2-yl)pyridine-3-yl)-10,10-dimethyl-6 1 ,6 2 ,6 3 ,6 4 ,6 5 ,6 6 -Hexahydro-1 1 Synthesis of H-8-oxa-2(4,2)-thiazola-1(5,3)-indola-6(1,3)-pyridadinacycloundecafane-5,7-dione At 0°C, tert-butyl((6 3 S,4S,Z)-11 -ethyl-1 2 -(2-((S)-1-methoxyethyl)-5-((R)-octahydro-2H-pyrido[1,2-a]pyrazine-2-yl)pyridine-3-yl)-10,10-dimethyl-5,7-dioxo-6 1 ,6 2 ,6 3 ,6 4 ,6 5 ,6 6 -Hexahydro-1 1 To a stirred solution of H-8-oxa-2(4,2)-thiazola-1(5,3)-indola-6(1,3)-pyridadinacycloundecafan-4-yl)carbamate (900 mg, 1.035 mmol) in DCM (9 mL), TFA (3 mL) was added dropwise. The resulting mixture was stirred at room temperature for 2 hours. The mixture was basicized to pH=8 with saturated NaHCO3 aqueous solution and extracted with ethyl acetate (3 × 100 mL). The combined organic layers were washed with brine (3 × 50 mL) and dried over anhydrous Na2SO4. After filtration, the filtrate was concentrated under reduced pressure to obtain the product (800 mg), which was used directly in the next step without further purification. LCMS(ESI) m / z:[M+H]C 42 H 56 Calculated value for N8O4S: 769.42; measured value: 769.5.

[0520] Intermediate 10:(6 3 S,4S)-4-amino-1 2 -(5-((S)-Hexahydropyridazino[2,1-c][1,4]oxazin-8(1H)-yl)-2-((S)-1-methoxyethyl)pyridine-3-yl)-10,10-dimethyl-1 1 -(2,2,2-trifluoroethyl)-6 1 ,6 2 ,6 3 ,6 4 ,6 5 ,6 6 -Hexahydro-1 1 Synthesis of H-8-oxa-1(5,3)-indola-6(1,3)-pyridadina-2(1,3)-benzenacycloundecafane-5,7-dione

[0521] [ka]

[0522] Step 1: Synthesis of (S)-3-(5-bromo-2-(2-(1-methoxyethyl)pyridine-3-yl)-1-(2,2,2-trifluoroethyl)-1H-indole-3-yl)-2,2-dimethylpropyl acetate At 0°C under an argon atmosphere, (S)-3-(5-bromo-2-(2-(1-methoxyethyl)pyridine-3-yl)-1-(2,2,2-trifluoroethyl)-1H-indole-3-yl)-2,2-dimethylpropan-1-ol (60 g, 0.12 mol) and Et3N (24.33 g, 0.24 mol) were stirred in 600 mL of DCM, to which DMAP (1.46 g, 0.012 mol) and acetic anhydride (14.7 g, 144 mmol) were added dropwise. The resulting mixture was stirred at room temperature for 2 hours. The resulting mixture was concentrated under reduced pressure and washed with HCl (500 mL). The resulting mixture was washed with saturated NaHCO3 aqueous solution (500 mL). The combined organic layers were washed with brine (500 mL) and dried over anhydrous Na2SO4. After filtration, the filtrate was concentrated under reduced pressure to obtain the product (59.6 g, 92% yield) as oil. LCMS(ESI) m / z:[M+H]C 25 H 28 Calculated value for BrF3N2O3: 541.13; Measured value: 543.2.

[0523] Step 2: Synthesis of (S)-(5-(3-(3-acetoxy-2,2-dimethylpropyl)-5-bromo-1-(2,2,2-trifluoroethyl)-1H-indole-2-yl)-6-(1-methoxyethyl)pyridine-3-yl)boronic acid At room temperature under an argon atmosphere, (S)-3-(5-bromo-2-(2-(1-methoxyethyl)pyridine-3-yl)-1-(2,2,2-trifluoroethyl)-1H-indole-3-yl)-2,2-dimethylpropyl acetate (55.1 g, 101.771 mmol) and 4,4,5,5-tetramethyl-2-(tetramethyl-1,3,2-dioxaborolan-2-yl)-1,3,2-dioxaborolan (38.77 g, 152.656 mmol) were stirred with THF (40 mL) to which dtbpy (4.10 g, 15.266 mmol) and chloro(1,5-cyclooctadiene)iridium(I) dimer (3.42 g, 5.089 mmol) were added in fractions. The resulting mixture was stirred at 75°C for 5 hours. The resulting mixture was concentrated under reduced pressure, and the product (102.4 g, crude) was obtained as oil. LCMS(ESI) m / z:[M+H]C 25 H 29 Calculated value for BBrF3N2O5: 585.14; Measured value: 585.2.

[0524] Step 3: (S)-3-(5-bromo-2-(5-iodo-2-(1-methoxyethyl)pyridine-3-yl)-1-(2,2,2-trifluoroethyl)-1H-indole-3-yl)-2,2-dimethylpropyl acetate At 0°C under an argon atmosphere, (S)-(5-(3-(3-acetoxy-2,2-dimethylpropyl)-5-bromo-1-(2,2,2-trifluoroethyl)-1H-indole-2-yl)-6-(1-methoxyethyl)pyridine-3-yl)boronic acid (51.2 g, 87.487 mmol) and sodium chloro[(4-methylbenzene)sulfonyl]azanide (197 g, 699.896 mmol) were stirred in THF (258 mL), to which a solution of NaI (104.91 g, 699.896 mmol) in aqueous solution (129 mL) was added dropwise. The resulting mixture was stirred at 55°C for 16 hours. The resulting mixture was concentrated under reduced pressure and extracted with CH3Cl (2 × 200 mL). The combined organic layers were washed with brine (2 × 20 mL) and dried over anhydrous Na2SO4. After filtration, the filtrate was concentrated under reduced pressure. The residue was purified by silica gel column chromatography (20% siRNA / petroleum ether) to obtain the product (15.3 g, 26% yield) as a solid. LCMS(ESI) m / z:[M+H]C 32 H 40 Calculated value for BrF3N4O4: 666.0; Measured value: 667.3.

[0525] Step 4: Synthesis of methyl(S)-1-((S)-3-(3-(3-(3-acetoxy-2,2-dimethylpropyl)-2-(5-((S)-hexahydropyridazino[2,1-c][1,4]oxazin-8(1H)-yl)-2-((S)-1-methoxyethyl)pyridine-3-yl)-1-(2,2,2-trifluoroethyl)-1H-indole-5-yl)phenyl)-2-((tert-butoxycarbonyl)amino)propanoyl)hexahydropyridazin-3-carboxylate At room temperature under an argon atmosphere, (S)-3-(5-bromo-2-(5-iodo-2-(1-methoxyethyl)pyridine-3-yl)-1-(2,2,2-trifluoroethyl)-1H-indole-3-yl)-2,2-dimethylpropyl acetate (2.70 g, 4.046 mmol) and (S)-octahydropyrazino[2,1-c][1,4]oxazine dihydrochloride (1.044 g, 4.855 mmol) were stirred with toluene (18.9 mL) and Cs2CO3 (5932.38 mg, 18.207 mmol) and BINAP (125.97 mg, 0.202 mmol) were added in small amounts. Pd(OAc)2 (90.84 mg, 0.405 mmol) was added in small amounts to the above mixture. The resulting mixture was stirred for a further 16 hours at 90°C. After cooling the mixture to room temperature, it was filtered, and the filtrate was washed with ethyl acetate (2 × 20 mL). The filtrate was concentrated under reduced pressure. The MeOH / DCM residue was purified by silica gel column chromatography (10% MeOH / DCM) to obtain the product (2.3 g, 83% yield) as a solid. LCMS(ESI) m / z:[M+H]C 32 H 40 Calculated value for BrF3N4O4: 681.23; Measured value: 681.4.

[0526] Step 5: Synthesis of methyl(S)-1-((S)-3-(3-(3-(3-acetoxy-2,2-dimethylpropyl)-2-(5-((S)-hexahydropyridazino[2,1-c][1,4]oxazin-8(1H)-yl)-2-((S)-1-methoxyethyl)pyridine-3-yl)-1-(2,2,2-trifluoroethyl)-1H-indole-5-yl)phenyl)-2-((tert-butoxycarbonyl)amino)propanoyl)hexahydropyridazin-3-carboxylate At room temperature and under an air atmosphere, methyl(S)-1-((S)-3-(3-(3-(3-acetoxy-2,2-dimethylpropyl)-2-(5-((S)-hexahydropyridazino[2,1-c][1,4]oxazin-8(1H)-yl)-2-((S)-1-methoxyethyl)pyridine-3-yl)-1-(2,2,2-trifluoroethyl)-1H-indole-5-yl)phenyl)-2-((tert-butoxycarbonyl)amino)propanoyl)hexahydropyridazin-3-carboxylate (2.33 g, 4.512 mmol) and K3PO4 (1.59 g, 7.490 mmol) were added to a 250 mL three-necked round-bottom flask. At room temperature, Pd(dtbpf)Cl2 (0.29 g, 0.451 mmol) was added in fractions to a stirred mixture of H2O (8.20 mL) and dioxane (8.20 mL) with toluene. The resulting mixture was stirred at 65°C for 3 hours. The resulting mixture was filtered, and the filtrate was washed with ethyl acetate (2 × 100 mL). The filtrate was concentrated under reduced pressure. The resulting mixture was extracted with ethyl acetate (2 × 100 mL). The combined organic layers were washed with brine (2 × 150 mL) and dried over anhydrous Na2SO4. After filtration, the filtrate was concentrated under reduced pressure. The MeOH / DCM residue was purified by silica gel column chromatography (3 → 4% MeOH / DCM) to obtain the product (2.7 g, 90% yield) as a solid. LCMS(ESI) m / z:[M+H]C 52 H 68 Calculated value for F3N7O9: 991.5; Measured value: 992.7.

[0527] Step 6: Synthesis of (S)-1-((S)-2-((tert-butoxycarbonyl)amino)-3-(3-(2-(5-((S)-hexahydropyridazino[2,1-c][1,4]oxazin-8(1H)-yl)-2-((S)-1-methoxyethyl)pyridine-3-yl)-3-(3-hydroxy-2,2-dimethylpropyl)-1-(2,2,2-trifluoroethyl)-1H-indole-5-yl)phenyl)propanoyl)hexahydropyridazin-3-carboxylic acid At room temperature, methyl(S)-1-((S)-3-(3-(3-(3-acetoxy-2,2-dimethylpropyl)-2-(5-((S)-hexahydropyridazino[2,1-c][1,4]oxazine-8(1H)-yl)-2-((S)-1-methoxyethyl)pyridine-3-yl)-1-(2,2,2-trifluoroethyl)-1H-indole-5-yl)phenyl)-2-((tert-butoxycarbonyl)amino)propanoyl)hexahydropyridazin-3-carboxylate (3 g, 3.024 mmol) and THF (30 mL) were added to a 100 mL three-necked round-bottom flask. At 0°C, the mixture was divided into smaller portions and followed by a solution of LiOH (0.30 g, 12.701 mmol) in water (12.7 mL). The resulting mixture was stirred at room temperature for 16 hours. The mixture was acidified to pH 5 with 1N HCl. The resulting mixture was extracted with dimethylammonium phosphate (2 × 100 mL). The combined organic layer was washed with brine (2 × 100 mL) and dried over anhydrous sodium 2SO4. After filtration, the filtrate was concentrated under reduced pressure to obtain the product (2.7 g, crude) as a solid. LC-MS (ESI) m / z: [M+H]C 49 H 64 Calculated value for F3N7O8: 936.48; Measured value: 936.7.

[0528] Step 7: tert-butyl((6 3 S,4S)-12-(5-((S)-Hexahydropyridazino[2,1-c][1,4]oxazin-8(1H)-yl)-2-((S)-1-methoxyethyl)pyridine-3-yl)-10,10-dimethyl-5,7-dioxo-11-(2,2,2-trifluoroethyl)-6 1 ,6 2 ,6 3 ,6 4 ,6 5 ,6 6 -Hexahydro-1 1 Synthesis of H-8-oxa-1(5,3)-indola-6(1,3)-pyridadina-2(1,3)-benzenacycloundecafan-4-yl)carbamate At room temperature, (S)-1-((S)-2-((tert-butoxycarbonyl)amino)-3-(3-(2-(5-((S)-hexahydropyridazino[2,1-c][1,4]oxazin-8(1H)-yl)-2-((S)-1-methoxyethyl)pyridine-3-yl)-3-(3-hydroxy-2,2-dimethylpropyl)-1-(2,2,2-trifluoroethyl)-1H-indole-5-yl)phenyl)propanoyl)hexahydropyridazin-3-carboxylic acid (3.12 g, 3.333 mmol) and DCM (624 mL) were added to a 2 L three-necked round-bottom flask. At 0°C, DIPEA (17.23 g, 133.320 mmol) and HOBt (4.50 g, 33.330 mmol) were added in small portions to the above mixture. The resulting mixture was stirred for a further 30 minutes. At room temperature for 16 hours, EDCI (19.17 g, 99.990 mmol) was added in small portions to the above mixture. The resulting mixture was concentrated under reduced pressure. The reaction product was quenched with H2O at 0°C. The resulting mixture was extracted with ELISA (3 × 100 mL). The combined organic layers were washed with brine (3 × 100 mL) and dried over anhydrous Na2SO4. After filtration, the filtrate was concentrated under reduced pressure. The MeOH / DCM residue was purified by silica gel column chromatography (3 → 4% MeOH / DCM) to obtain the product (3 g, 98% yield) as a solid. LCMS(ESI) m / z:[M+H]C 49 H 62 Calculated value for F3N7O7: 918.47; Measured value: 918.8.

[0529] Step 8:(6 3 S,4S)-4-amino-1 2 -(5-((S)-Hexahydropyridazino[2,1-c][1,4]oxazin-8(1H)-yl)-2-((S)-1-methoxyethyl)pyridine-3-yl)-10,10-dimethyl-11-(2,2,2-trifluoroethyl)-6 1 ,6 2 ,6 3 ,6 4 ,6 5 ,6 6 -Hexahydro-1 1Synthesis of H-8-oxa-1(5,3)-indola-6(1,3)-pyridadina-2(1,3)-benzenacycloundecafane-5,7-dione At 0°C, under an argon atmosphere, tert-butyl((6 3 S,4S)-12-(5-((S)-Hexahydropyridazino[2,1-c][1,4]oxazin-8(1H)-yl)-2-((S)-1-methoxyethyl)pyridine-3-yl)-10,10-dimethyl-5,7-dioxo-1 1 -(2,2,2-trifluoroethyl)-6 1 ,6 2 ,6 3 ,6 4 ,6 5 ,6 6 A stirring solution of -hexahydro-11H-8-oxa-1(5,3)-indola-6(1,3)-pyridadina-2(1,3)-benzenacycloundecafan-4-yl)carbamate (930 mg, 1.013 mmol) in DCM (15 mL) was mixed with TFA (5 mL, 67.315 mmol) dissolved in DCM (5 mL). The resulting mixture was stirred at 0°C for 2 hours. The residue was basicized to pH 8 with saturated NaHCO3 aqueous solution. The resulting mixture was extracted with DCM, and the combined organic layers were washed with brine and dried over anhydrous Na2SO4. After filtration, the filtrate was concentrated under reduced pressure to obtain the product (880 mg, crude) as a solid. LCMS(ESI) m / z:[M+H]C 44 H 54 Calculated value for F3N7O5: 818.42; measured value: 818.6.

[0530] Intermediate 11:(6 3 S,4S)-4-amino-12-(2-((S)-1-methoxyethyl)-5-((R)-octahydro-2H-pyrido[1,2-a]pyrazine-2-yl)pyridine-3-yl)-10,10-dimethyl-1 1 -(2,2,2-trifluoroethyl)-6 1 ,6 2 ,6 3 ,6 4 ,6 5 ,6 6 -Hexahydro-11 Synthesis of H-8-oxa-1(5,3)-indola-6(1,3)-pyridadina-2(1,3)-benzenacycloundecafane-5,7-dione

[0531] [ka]

[0532] Step 1: Synthesis of (S)-(5-(3-(3-acetoxy-2,2-dimethylpropyl)-5-bromo-1-(2,2,2-trifluoroethyl)-1H-indole-2-yl)-6-(1-methoxyethyl)pyridine-3-yl)boronic acid At room temperature, 3-(5-bromo-2-{2-[(1S)-1-methoxyethyl]pyridine-3-yl}-1-(2,2,2-trifluoroethyl)indole-3-yl)-2,2-dimethylpropyl acetate (10 g, 18.470 mmol), 4,4,5,5-tetramethyl-2-(tetramethyl-1,3,2-dioxaborolan-2-yl)-1,3,2-dioxaborolan (8.44 g, 33.25 mmol), and dtbpy (0.89 g, 3.325 mmol) were added to a 100 mL three-necked round-bottom flask. To the above mixture, chloro(1,5-cyclooctadiene)iridium(I) dimer (0.74 g, 1.108 mmol) and THF (40 mL) were added. The resulting mixture was further stirred at 80°C for 16 hours. The resulting mixture was concentrated under reduced pressure. The crude product was used directly in the next step without further purification. LC-MS(ESI) m / z:[M+H]C 25 H 29 Calculated value for BBrF3N2O5: 585.14; Measured value: 585.0.

[0533] Step 2: Synthesis of (S)-3-(5-bromo-2-(5-iodo-2-(1-methoxyethyl)pyridine-3-yl)-1-(2,2,2-trifluoroethyl)-1H-indole-3-yl)-2,2-dimethylpropyl acetate At 0°C under a nitrogen atmosphere, (S)-(5-(3-(3-acetoxy-2,2-dimethylpropyl)-5-bromo-1-(2,2,2-trifluoroethyl)-1H-indole-2-yl)-6-(1-methoxyethyl)pyridine-3-yl)boronic acid (17.9 g, 30.586 mmol) was added dropwise to a stirred solution of (S)-(5acetoxy-2,2-dimethylpropyl)-5-bromo-1-(2,2,2-trifluoroethyl)-1H-indole-2-yl)-6-(5)-(5)-(5)-methoxyethyl)pyridine-3-yl)boronic acid (17.9 g, 30.586 mmol) in THF (89.5 mL) to which sodium chloro[(4-methylbenzene)sulfonyl]azanide (68.93 g, 244.688 mmol) and NaI (36.68 g, 244.688 mmol) in water (44.75 mL) was added dropwise. The resulting mixture was stirred at room temperature for a further 20 minutes, and then heated to 50°C for 16 hours. The resulting mixture was washed After filtration, the filtrate was washed with CHCl3 (3 × 100 mL). The filtrate was extracted with CHCl3 (3 × 200 mL). The combined organic layers were washed with Na2S2O3 (300 mL) and brine (2 × 150 mL) and dried over anhydrous Na2SO4. After filtration, the filtrate was concentrated under reduced pressure. The residue was purified by silica gel column chromatography (16% Â / petroleum ether) to obtain the product (6.6 g, 32% yield) as a solid. LCMS(ESI) m / z:[M+H]C 25 H 27 Calculated value for BrF3IN2O3: 667.03; Measured value: 668.7.

[0534] Step 3: Synthesis of 3-(5-bromo-2-(2-((S)-1-methoxyethyl)-5-((R)-octahydro-2H-pyrido[1,2-a]pyrazine-2-yl)pyridine-3-yl)-1-(2,2,2-trifluoroethyl)-1H-indole-3-yl)-2,2-dimethylpropyl acetate (S)-3-(5-bromo-2-(5-iodo-2-(1-methoxyethyl)pyridine-3-yl)-1-(2,2,2-trifluoroethyl)-1H-indole-3-yl)-2,2-dimethylpropyl acetate (1.4 g, 2.098 mmol) and (R)-octahydro-2H-pyrido[1,2-a]pyrazine (353.04 mg, 2.518 mmol) were mixed with toluene (10 mL) and Cs2CO3 (3076.05 mg, 9.441 mmol), BINAP (65.32 mg, 0.105 mmol), and Pd(OAc)2 (47.10 mg, 0.210 mmol) were added. The resulting mixture was stirred overnight at 90°C under an argon atmosphere. The reaction product was quenched with H2O (100 mL). The resulting mixture was extracted using DCM (3 × 100 mL). The combined organic layer was washed with H2O (3 × 100 mL) and dried over anhydrous Na2SO4. After filtration, the filtrate was concentrated under reduced pressure. The residue was purified by silica gel column chromatography (5% MeOH / DCM) to obtain the product (1 g, 49% yield) as oil. LCMS (ESI) m / z:[M+H]C 33 H 42 Calculated value for BrF3N4O3: 679.25; measured value: 679.5.

[0535] Step 4: Synthesis of methyl(S)-1-((S)-3-(3-(3-(3-acetoxy-2,2-dimethylpropyl)-2-(2-((S)-1-methoxyethyl)-5-((R)-octahydro-2H-pyrido[1,2-a]pyrazine-2-yl)pyridine-3-yl)-1-(2,2,2-trifluoroethyl)-1H-indole-5-yl)phenyl)-2-((tert-butoxycarbonyl)amino)propanoyl)hexahydropyridazine-3-carboxylate 3-(5-bromo-2-(2-((S)-1-methoxyethyl)-5-((R)-octahydro-2H-pyrido[1,2-a]pyrazine-2-yl)pyridine-3-yl)-1-(2,2,2-trifluoroethyl)-1H-indole-3-yl)-2,2-dimethylpropyl acetate (1g, 1.471 mmol) and methyl(S)-1-((S)-2-((tert-butoxycarbonyl)amino)-3-(3-(4,4,5,5-tetramethyl-1 To a stirred mixture of ,3,2-dioxaborolan-2-yl)phenyl)propanoyl)hexahydropyridazine-3-carboxylate (913.62 mg, 1.765 mmol) and toluene (9 mL), dioxane (6 mL), H2O (3 mL), K3PO4 (780.82 mg, 3.678 mmol), and Pd(dtbpf)Cl2 (95.90 mg, 0.147 mmol) were added, and the resulting mixture was stirred for 2 hours at 70°C under a nitrogen atmosphere. The mixture was basicized to pH 8 with saturated NaHCO3 aqueous solution. The resulting mixture was extracted with DCM (3 × 30 mL). The combined organic layers were washed with H2O (3 × 30 mL) and dried over anhydrous Na2SO4. After filtration, the filtrate was concentrated under reduced pressure. The residue was purified by silica gel column chromatography (20% siRNA / petroleum ether) to obtain the product (1.2 g, 74% yield) as a solid. LCMS(ESI) m / z:[M+H]C 53 H 70 Calculated value for F3N7O8: 990.53; Measured value: 990.8.

[0536] Step 5: Synthesis of (S)-1-((S)-2-((tert-butoxycarbonyl)amino)-3-(3-(3-(3-hydroxy-2,2-dimethylpropyl)-2-(2-((S)-1-methoxyethyl)-5-((R)-octahydro-2H-pyrido[1,2-a]pyrazine-2-yl)pyridine-3-yl)-1-(2,2,2-trifluoroethyl)-1H-indole-5-yl)phenyl)propanoyl)hexahydropyridazine-3-carboxylic acid At 0°C, methyl(S)-1-((S)-3-(3-(3-(3-acetoxy-2,2-dimethylpropyl)-2-(2-((S)-1-methoxyethyl)-5-((R)-octahydro-2H-pyrido[1,2-a]pyrazine-2-yl)pyridine-3-yl)-1-(2,2,2-trifluoroethyl)-1H-indole-5-yl)phenyl)-2-((tert-butoxycarbonyl)amino)propanoyl)hexahydropyridazine-3-carboxylate (1.2 g, 1.212 mmol) and LiOH (252 mg, 10.523 mmol) were mixed with THF (6 mL) and H2O (6 mL) was added in fractional amounts. The resulting mixture was stirred overnight at 0°C. The mixture was acidified to pH 7 with 1N HCl (aqueous solution). The aqueous layer was extracted by DCM (3 × 30 mL). The combined organic layer was washed with H2O (30 mL) and dried over anhydrous Na2SO4. After filtration, the filtrate was concentrated under reduced pressure to obtain the product (1.2 g, 84% yield) as a solid. LCMS(ESI) m / z:[M+H]C 50 H 66 Calculated value for F3N7O7: 934.51; Measured value: 935.0.

[0537] Step 6: tert-butyl((6 3 S,4S)-12-(2-((S)-1-methoxyethyl)-5-((R)-octahydro-2H-pyrido[1,2-a]pyrazine-2-yl)pyridine-3-yl)-10,10-dimethyl-5,7-dioxo-1 1 -(2,2,2-trifluoroethyl)-6 1 ,6 2 ,6 3 ,6 4 ,6 5 ,6 6 -Hexahydro-1 1 H-8-oxa-1(5,3)-indola-6(1,3)-pyridadina-2(1,3)-benzenacycloundecafan-4-yl)carbamate At 0°C, (S)-1-((S)-2-((tert-butoxycarbonyl)amino)-3-(3-(3-(3-hydroxy-2,2-dimethylpropyl)-2-(2-((S)-1-methoxyethyl)-5-((R)-octahydro-2H-pyrido[1,2-a]pyrazine-2-yl)pyridine-3-yl)-1-(2,2,2-trifluoroethyl)-1H-indole-5-yl)phenyl)propanoyl)hexahydropyridazine-3-carboxylic acid (1.2 g, 1.285 mmol) and DIPEA (7.83 mL, 44.975 mmol) were stirred with DCM (100 mL), to which HOBt (0.87 g, 6.425 mmol) and EDCI·HCl (5.58 g, 35.980 mmol) were added in fractions. The resulting mixture was stirred overnight at 0°C. The mixture was diluted with DCM (30 mL). The combined organic layer was washed with H2O (3 × 30 mL) and dried over anhydrous Na2SO4. After filtration, the filtrate was concentrated under reduced pressure. The MeOH / DCM residue was purified by silica gel column chromatography (5% MeOH / DCM) to obtain the product (850 mg, 65% yield) as a solid. LCMS(ESI) m / z:[M+H]C 50 H 64 Calculated value for F3N7O6: 916.49; Measured value: 917.0.

[0538] Step 7:(6 3 S,4S)-4-amino-12-(2-((S)-1-methoxyethyl)-5-((R)-octahydro-2H-pyrido[1,2-a]pyrazine-2-yl)pyridine-3-yl)-10,10-dimethyl-1 1 -(2,2,2-trifluoroethyl)-6 1 ,6 2 ,6 3 ,6 4 ,6 5 ,6 6 -Hexahydro-1 1 Synthesis of H-8-oxa-1(5,3)-indola-6(1,3)-pyridadina-2(1,3)-benzenacycloundecafane-5,7-dione At 0°C, tert-butyl((6 3S,4S)-12-(2-((S)-1-methoxyethyl)-5-((R)-octahydro-2H-pyrido[1,2-a]pyrazine-2-yl)pyridine-3-yl)-10,10-dimethyl-5,7-dioxo-1 1 -(2,2,2-trifluoroethyl)-6 1 ,6 2 ,6 3 ,6 4 ,6 5 ,6 6 -Hexahydro-1 1 H-8-oxa-1(5,3)-indola-6(1,3)-pyridadina-2(1,3)-benzenacycloundecafan-4-yl)carbamate (1000 mg, 1.092 mmol) was mixed with DCM (4 mL) and TFA (4 mL) was added. The resulting mixture was stirred for 1 hour at 0°C under a nitrogen atmosphere. The resulting mixture was concentrated under reduced pressure. The residue was basicized to pH 8 with saturated NaHCO3 aqueous solution. The aqueous layer was extracted with DCM (3 × 30 mL). The combined organic layers were dried over anhydrous Na2SO4. After filtration, the filtrate was concentrated under reduced pressure to obtain the product (800 g, 80% yield) as a solid. LCMS(ESI) m / z:[M+H]C 45 H 56 Calculated value for F3N7O4: 816.44; measured value: 816.6.

[0539] Intermediate 12: (6 3 S,4S)-4-amino-1 1 -ethyl-1 2 -(5-((S)-Hexahydropyridazino[2,1-c][1,4]oxazin-8(1H)-yl)-2-((S)-1-methoxyethyl)pyridine-3-yl)-10,10-dimethyl-6 1 ,6 2 ,6 3 ,6 4 ,6 5 ,6 6 -Hexahydro-1 1 Synthesis of H-8-oxa-1(5,3)-indola-6(1,3)-pyridadina-2(1,3)-benzenacycloundecafane-5,7-dione

[0540] [ka]

[0541] Step 1: Synthesis of methyl(S)-1-((S)-3-(3-(3-(3-acetoxy-2,2-dimethylpropyl)-1-ethyl-2-(5-((S)-hexahydropyridazino[2,1-c][1,4]oxazin-8(1H)-yl)-2-((S)-1-methoxyethyl)pyridine-3-yl)-1H-indole-5-yl)phenyl)-2-((tert-butoxycarbonyl)amino)propanoyl)hexahydropyridazin-3-carboxylate At room temperature under an argo...

Claims

1. Structure of formula I: 【Chemistry 1】 A compound having, or a pharmaceutically acceptable salt thereof, In the formula, A is an optionally substituted 3- to 6-membered heterocycloalkylene, an optionally substituted 3- to 6-membered cycloalkylene, an optionally substituted 6-membered arylene, or an optionally substituted 5- to 10-membered heteroarylene. X1, X2, and X3 are each independently selected from CH2, CHF, CF2, C=O, or O. m is either 1 or 2. n is either 0 or 1, R1 is hydrogen, an optionally substituted C1-C6 heteroalkyl, or an optionally substituted 3- to 10-membered heterocycloalkyl. R2 is an optionally substituted C1-C6 alkyl, and R3 is an optionally substituted C1-C6 alkyl, an optionally substituted C1-C6 heteroalkyl, an optionally substituted 3- to 6-membered cycloalkyl, or an optionally substituted heterocycloalkyl. Each hydrogen atom is independently an arbitrarily isotopically enriched deuterium. A compound, or a pharmaceutically acceptable salt thereof.

2. The compound according to claim 1, or a pharmaceutically acceptable salt thereof, wherein R1 is hydrogen or an optionally substituted 3- to 10-membered heterocycloalkyl group.

3. The compound according to claim 2, or a pharmaceutically acceptable salt thereof, wherein R1 is an optionally substituted 3- to 10-membered heterocycloalkyl group.

4. R1 is, 【Chemistry 2】 The compound according to claim 3, or a pharmaceutically acceptable salt thereof.

5. The compound according to claim 1, or a pharmaceutically acceptable salt thereof, wherein m is 1.

6. The compound according to claim 1, or a pharmaceutically acceptable salt thereof, wherein n is 1.

7. The compound according to claim 1, or a pharmaceutically acceptable salt thereof, wherein each of X1, X2, and X3 is CH2.

8. Structure of Formula II: 【Transformation 3】 A compound according to claim 1, or a pharmaceutically acceptable salt thereof, having the above.

9. Structure of formula V: 【Chemistry 4】 A compound according to claim 1, or a pharmaceutically acceptable salt thereof, having the above.

10. Structure of formula VI: 【Transformation 5】 A compound according to claim 1, or a pharmaceutically acceptable salt thereof, having the above.

11. Structure of formula VII: 【Transformation 6】 A compound according to claim 1, or a pharmaceutically acceptable salt thereof, having the above.

12. The compound according to claim 1, or a pharmaceutically acceptable salt thereof, wherein A is optionally substituted thiazole-diyl, optionally substituted oxazole-diyl, optionally substituted morpholine-diyl, optionally substituted pyrrolidine-diyl, optionally substituted piperidine-diyl, or optionally substituted phenylene.

13. The compound according to claim 1, or a pharmaceutically acceptable salt thereof, wherein A is an optionally substituted 5- to 10-membered heteroarylene.

14. A is, 【Transformation 7】 The compound according to claim 13, or a pharmaceutically acceptable salt thereof.

15. The compound according to claim 1, or a pharmaceutically acceptable salt thereof, wherein A is optionally substituted phenylene.

16. A is, 【Transformation 8】 The compound according to claim 15, or a pharmaceutically acceptable salt thereof.

17. The compound according to claim 1, or a pharmaceutically acceptable salt thereof, wherein A is an optionally substituted 3- to 6-membered heterocycloalkylene.

18. A is as follows: 【Chemistry 9】 A compound according to claim 17, or a pharmaceutically acceptable salt thereof, selected from the above.

19. R2 is, 【Chemistry 10】 The compound according to claim 1, or a pharmaceutically acceptable salt thereof.

20. The compound according to claim 1, or a pharmaceutically acceptable salt thereof, wherein R3 is optionally substituted C1-C6 alkyl.

21. R 3 is, 【Chemistry 11】 The compound according to claim 20, or a pharmaceutically acceptable salt thereof.

22. The compound according to claim 1, or a pharmaceutically acceptable salt thereof, wherein R3 is an optionally substituted 3- to 6-membered cycloalkyl group.

23. R 3 is, 【Chemistry 12】 The compound according to claim 22, or a pharmaceutically acceptable salt thereof.

24. The following: 【Chemistry 13-1】 【Chemistry 13-2】 【Chemistry 13-3】 [Chemistry 13-4] 【Chemistry 13-5】 【Chemistry 13-6】 【Chemistry 13-7】 【Chemistry 13-8】 【Chemistry 13-9】 A compound or a pharmaceutically acceptable salt thereof having the structure of a compound selected from the above.

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

26. The pharmaceutical composition according to claim 25 for use in the treatment of cancer.

27. ​​The pharmaceutical composition according to claim 26, wherein the cancer is pancreatic cancer, non-small cell lung cancer, colorectal cancer, or endometrial cancer.

28. The pharmaceutical composition according to claim 26, wherein the cancer comprises a Ras mutation.

29. The pharmaceutical composition according to claim 28, wherein the Ras mutation is K-Ras G12D or K-Ras G13D.

30. The pharmaceutical composition according to claim 25 for use in the treatment of Ras protein-related diseases.

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