Topoisomerase inhibitors, antibody-drug conjugates comprising them, and methods of their use

Abstract

Disclosed herein are novel topoisomerase inhibitors, compounds comprising the inhibitors coupled to linkers, antibody-drug conjugates, and methods of their preparation and use. Particular antibody-drug conjugates have a structure according to formula (V) or (Va) or the substituents of which are defined herein.

Description

[0001] TOPOISOMERASE INHIBITORS, ANTIBODY-DRUG CONJUGATES COMPRISING THEM, AND METHODS OF THEIR USE

[0002] This application claims priority to U.S. provisional patent application no. 63 / 657,480, filed June 7, 2024, the entirety of which is incorporated herein by reference.

[0003] BACKGROUND OF THE DISCEOSURE

[0004] A type of anticancer drug that is generating strong interest is a conjugate, in which an anticancer drug is attached to a targeting agent that binds to a ligand on a cancer cell. The targeting agent, by binding to the ligand, directs the drug to the cancer cell, where it is released by one of several mechanisms, to act on the cancer cell. But, while in systemic circulation, the drug is inactive by virtue of its being attached to the targeting agent.

[0005] A common type of conjugate is an antibody-drug conjugate (“ADC”, also referred to as an immunoconjugate). In an ADC, an anticancer drug (synonymously therapeutic agent, cytotoxin, payload, or warhead) is covalently linked to an antibody whose antigen is a tumor-associated antigen - i. e. , an antigen that is exclusively or predominantly expressed by a cancer cell, to the exclusion of normal tissue cells.

[0006] The moiety covalently linking the antibody and the drug is referred to as the linker. Thus, generally, an ADC has the formula

[0007] Antibody- [Linker -Drug]mwhere m refers to the number of drug molecules attached to the antibody. Typically, m is between 1 and 8.

[0008] The antibody, upon binding to its antigen, delivers the ADC to the cancer site. There, cleavage of the linker or degradation of the antibody releases the drug. Frequently, the ADC is internalized within a cancer cell by endocytosis and release of the drug takes place inside the cancer cell. The drug can be a much more potent (cytotoxic) agent than an ordinary chemotherapy agent because its localized release reduces systemic toxicity. For a review of ADCs, see Fu et al., Signal Transduction and Targeted Therapy, 2022, 7, 93.

[0009] Over ten ADCs have received approval in the US and abroad for hematologic and solid malignancies. The list of target antigens is diverse, including CD33, CD30, CD79b, B-cell maturation agent (BCMA), CD19, PE38, HER2, NECTIN-4, TROP-2, tissue factor, folate receptor 1, and epidermal growth factor receptor. For a review, see Riccardi et al., Front. Pharmacol. 2023 14, 1274088.

[0010] Many drugs have been tried as a payload in an ADCs, representing many different mechanisms of action and structures. Some mechanisms of action mirror those of conventional chemotherapy agents; others are unique to ADCs. Exemplary ADC payloads include microtubule inhibitors / disrupters; DNA intercalator / alkylators, enediynes, pyrrolobenzodiazepines, EG5 inhibitors, RNA polymerase II inhibitors, topoisomerase I inhibitors, and topoisomerase II inhibitors. For a review, see Wang et al., Acta Pharmaceutica Sinica B, 2023, 13 (10), 4025.

[0011] Camptothecin is an anticancer drug that has spawned numerous analogues seeking to improve upon one pharmacological property or another. Some examples are irinotecan, topotecan, SN38, exatecan, and Dxd. For a review, see Venditto et al., Mol. Pharmacol. 2010, 7, 307.

[0012] One family of camptothecin analogues that has been explored entails the expansion of the six-member lactone ring to a seven-member ring. These analogues - e.g., homocamptothecin and BN 80927 - have interesting anticancer drug properties (Lavergne et al. Bioorg. Med. Chem. Lett., 1999, 9, 2599). However, they have not been used as a payload for an ADC.

[0013] Other disclosures involving homocamptothecin-based compounds include CN 117164601; WO 2023 / 207773; CN 116253747; CN 1557814; US 6,339,091; and WO 2924 / 230752.

[0014] BRIEF SUMMARY OF THE DISCLOSURE

[0015] This invention is based on the discovery of new topoisomerase inhibitors that are suitable for conjugation in an ADC. The compounds possess either a primary or secondary hydroxyl or amino group that is reactive and serves as a conjugation handle for attachment to a linker. A primary or secondary hydroxyl or amino group is a preferred conjugation site, as opposed to a tertiary hydroxyl or amino group as they are more reactive and less sterically hindered, although a tertiary hydroxyl or amino group can also be used as a conjugation site.

[0016] Particular compounds of the invention have cell permeability and / or pharmacokinetic properties that render them particularly useful as ADC payloads.

[0017] Accordingly, there is provided a compound of formula (I) or a pharmaceutically acceptable salt thereof, wherein each X is independently O or NH; and

[0018] Y is CH orN; where the open valence of R1that is bonded to -XH is denoted by an * and the other end of R1that is bonded to the aromatic core of the compound of formula (I) is denoted by a wiggly line ( ). Particular embodiments of the invention encompass compounds of formula (I) with the provisos that: when R1is absent, R2is methyl and R3is fluoro, X is not O or NH; and when X is O, R2is methyl and R3is fluoro, R1is not

[0019] In one embodiment, R2is Me and R3is Cl or F. In a preferred embodiment, R2is Me and R3is Cl. In another preferred embodiment, R2is Me and R3is F.

[0020] In another embodiment, there is provided a compound having the structure or a pharmaceutically acceptable salt thereof. We also provide a drug -linker compound of formula (II) wherein R1, R2, R3, and X are as defined hereinabove, L is a linker, and Rbis -NH2, -OH, -CO2H, -SH, maleimido, cyclooctyne, azido (-N3), hydroxylamino (-ONH2) or N-hydroxysuccinimido. We further provide an antibody-drug -linker conjugate of the formula (V): where R1, R2, R3, and X are as defined hereinabove, L’ is a linker, Ab is an antibody, and m is 1, 2, 3, 4, 5, 6, 7 or 8. We also provide a drug -linker compound of formula (Ila):

[0021] R5is H, Cl, F, (CHJ^H. OCCH^^H, or S(CH2)1.3H; R6is H, Cl, F, (CH^H, OCCH^^H, or SCCH^H;

[0022] L is a linker; and

[0023] Rbis -NH2, -OH, -CO2H, -SH, maleimido, cyclooctyne, azido (-N3), hydroxylamino (-ONH2) or N -hydroxy succinimido .

[0024] We also provide an antibody-drug -linker conjugate of formula (Va): wherein R4, R5, and R6are as defined above; L’ is a linker; Ab is an antibody; and m is 1, 2, 3, 4, 5, 6, 7 or 8.

[0025] DETAILED DESCRIPTION OF THE INVENTION

[0026] TOPOISOMERASE INHIBITORS Topoisomerase inhibitors of this invention include compounds of Formula (I), shown above. In one embodiment of the invention, R1is present and the X immediately adjacent to R1is NH. Thus, in this embodiment, there is provided a compound according to formula (F)

[0027] R2is H, Cl, F, (CH^H, O(CH2)!.3H, or SCCH^H;

[0028] R3is H, Cl, F, (CH^H, ©(CH^^H, or SCCH^H; or R2and R3in combination form each X is independently O or NH;

[0029] Y is CH orN; where the open valence of R1that is bonded to -NH2 is denoted by an * and the other end of R1that is bonded to the aromatic core of the compound of formula (I’) is denoted by a wiggly line ( ).

[0030] In another preferred embodiment, R1comprises a cycloaliphatic or heterocycloaliphatic moiety and the X immediately adjacent to R1is O. Thus, in this embodiment, there is provided a compound according to formula (I”)

[0031] R2is H, Cl, F, (CH^H, ©(CH^^H, or SCCH^^H;

[0032] R3is H, Cl, F, (CH^H, ©(CH^^H, or SCCH^^H; or R2and R3in combination form each X is independently O or NH;

[0033] Y is CH orN; where the open valence of R1that is bonded to -NH2 is denoted by an * and the other end of R1that is bonded to the aromatic core of the compound of formula (I’) is denoted by a wiggly line ( ); with the provision that is excluded.

[0034] Preferred compounds of this invention include the following, where R3is Cl or F:

[0035]

[0036] In one embodiment of compounds (A)-(R), R3is Cl. In another embodiment of compounds (A)-(R), R3is

[0037] F.

[0038] Preferred compounds where R3is F include:

[0039]

[0040] CONJUGATION

[0041] The compounds disclosed herein can be delivered to the site of intended action by targeted delivery in a conjugate with a targeting agent. Preferably, the targeting agent is an antibody or antigen binding portion thereof and its cognate antigen is found at the locality of intended action, for example a tumor associated antigen if the site of intended action is a tumor (cancer). Preferably, the tumor associated antigen is uniquely expressed or overexpressed by the cancer cell, compared to a normal cell. The tumor associated antigen can be located on the surface of the cancer cell or secreted by the cancer cell into its environs.

[0042] By binding to a target tissue or cell where its antigen or receptor is located, the antibody directs the conjugate there. Cleavage of the linker at the target tissue or cell releases the drug to exert its effect locally. In this manner, precise delivery of the drug is achieved at the site of intended action, reducing the dosage needed. Also, the drug is normally biologically inactive (or significantly less active) in its conjugated state, thereby reducing off-target effects.

[0043] Thus, a conjugate of this disclosure has the general formula

[0044] Antibody- [Linker -Drug]m

[0045] As reflected by the subscript m, each antibody can conjugate with more than one drug, depending on the number of sites each antibody has available for conjugation and the experimental conditions employed. Those skilled in the art will appreciate that, while each individual antibody is conjugated to an integer number of drug moieties, a preparation of the conjugate may analyze for a non-integer ratio of drug to antibody, reflecting a statistical average. This ratio is referred to as the substitution ratio (“SR”) or the drug -antibody ratio (“DAR”). Typically, m is between 1 and 8, depending on the conjugation chemistry employed and conditions of conjugation

[0046] Generally, an ADC is constructed by first making a drug-linker compound and then conjugating it to an antibody. The conjugation step generally takes place in an aqueous medium. This method requires complementary reactive functional groups on the antibody and drug-linker compound:

[0047] Antibody-Ra+. Rb-[Linker-Drug] Antibody -Rc-[Linker-Drug] where Rais a reactive functional group on the antibody, Rbis a complementary reactive functional group on the drug -linker compound, and Rcis the resultant link formed by the reaction of Raand Rb.

[0048] Chemistry generally usable for the preparation of suitable moieties Rb- [Linker-Drug] is disclosed in Ng et al., US 7,087,600 B2 (2006); Ng etal., US 6,989,452 B2 (2006); Ng etal., US 7,129,261 B2 (2006); Ng etal., WO 02 / 096910 Al; Boyd et al., US 7,691,962 B2; Chen et al., US 7,517,903 B2 (2009); Gangwar et al., US 7,714,016 B2 (2010); Boyd et al., US 2008 / 0279868 Al; Gangwar et al. , US 7,847,105 B2 (2010); Gangwar et al., US 7,968,586 B2 (2011); Sufi et al., US 8,461,117 B2 (2013); and Chen et al., US 8,664,407 B2 (2014); the disclosures of which are incorporated herein by reference.

[0049] Preferably reactive functional group Rbis -NH2, -OH, -CO2H, -SH, maleimido, cyclooctyne, azido (-N3), hydroxylamino (-ONH2) or N-hydroxysuccinimido. Especially preferred functional groups Rbare:

[0050] An -OH group can be esterified with a carboxy group on the antibody, for example, on an aspartic or glutamic acid side chain.

[0051] A -CO2H group can be esterified with a -OH group or amidated with an amino group (for example on a lysine side chain) on the antibody.

[0052] An N-hydroxy succinimide group is functionally an activated carboxyl group and can conveniently be amidated by reaction with an amino group (e.g., from lysine).

[0053] A maleimide group can be conjugated with an -SH group on the antibody (e.g., from cysteine or from the chemical modification of the antibody to introduce a sulfhydryl functionality), in a Michael addition reaction.

[0054] Where an antibody does not have a cysteine -SH available for conjugation, an s-amino group in the side chain of a lysine residue can be reacted with 2-iminothiolane or N-succinimidyl-3-(2-pyridyldithio)- propionate (“SPDP”) to introduce a free thiol (-SH) group - creating a cysteine surrogate, as it were. The thiol group can react with a maleimide or other nucleophile acceptor group to effect conjugation. The mechanism is illustrated below with 2-iminothiolane:

[0055] Lys thiolane

[0056] Typically, athiolation level of two to three thiols per antibody is achieved. For a representative procedure, see Cong et al., US 8,980,824 B2 (2015), the disclosure of which is incorporated herein by reference.

[0057] In a reversed arrangement, an antibody Z can be modified with N-succinimidyl 4-(maleimidomethyl)- cyclohexanecarboxylate (“SMCC”) or its sulfonated variant sulfo-SMCC, both of which are available from Sigma- Aldrich, to introduce a maleimide group thereto. Then, conjugation can be effected with a drug-linker compound having an -SH group on the linker.

[0058] An alternative conjugation method employs copper-free “click chemistry,” in which an azide group adds across a strained cyclooctyne to form an 1,2,3-triazole ring. See, e.g., Agard et al., J. Amer. Chem. Soc. 2004, 126, 15046; Best, Biochemistry 2009, 48, 6571, the disclosures of which are incorporated herein by reference. The azide can be located on the antibody and the cyclooctyne on the drug -linker moiety, or vice-versa. A preferred cyclooctyne group is dibenzocyclooctyne (DIBO). Various reagents having a DIBO group are available from Invitrogen / Molecular Probes, Eugene, Oregon. The reaction below illustrates click chemistry conjugation for the instance in which the DIBO group is attached to the antibody (but the reverse arrangement can also be employed):

[0059] Yet another conjugation technique involves introducing a non-natural amino acid into an antibody, with the non-natural amino acid providing a functionality for conjugation with a reactive functional group in the drug moiety. For instance, the non-natural amino acid p-acetylphenylalanine can be incorporated into an antibody or other polypeptide, as taught in Tian et al., WO 2008 / 030612 A2 (2008). The ketone group in p-acetylphenyalanine can be a conjugation site via the formation of an oxime with a hydroxylamino group on the linker-drug moiety. Alternatively, the non-natural amino acid p-azidophenylalanine can be incorporated into an antibody to provide an azide functional group for conjugation via click chemistry, as discussed above. Non-natural amino acids can also be incorporated into an antibody or other polypeptide using cell-free methods, as taught in Goerke et al., US 2010 / 0093024 Al (2010). The foregoing disclosures are incorporated herein by reference.

[0060] Still another conjugation technique uses the enzyme transglutaminase (preferably bacterial transglutaminase from Streptomyces mobaraensis or BTG), per Jeger et al., Angew. Chem. Int. Ed. 2010, 49, 9995. BTG forms an amide bond between the side chain carboxamide of a glutamine (the amine acceptor) and an alkyleneamino group (the amine donor), which can be, for example, the s-amino group of a lysine or a 5-amino-n-pentyl group. In atypical conjugation reaction, the glutamine residue is located on the antibody, while the alkyleneamino group is located on the linker-drug moiety, as shown below: O — (CH2)2-C-NH— [Linker-Drug]

[0061] The positioning of a glutamine residue on a polypeptide chain has a large effect on its susceptibility to BTG mediated transamidation. None of the glutamine residues on an antibody are normally BTG substrates. However, if the antibody is deglycosylated - the glycosylation site being asparagine 297 (N297) of the heavy chain - nearby glutamine 295 (Q295) is rendered BTG susceptible. An antibody can be deglycosylated enzymatically by treatment with PNGase F (Peptide-N-Glycosidase F). Alternatively, an antibody can be synthesized glycoside free by introducing an N297A mutation in the constant region, to eliminate the N297 glycosylation site. Further, it has been shown that an N297Q substitution not only eliminates glycosylation, but also introduces a second glutamine residue (at position 297) that too is an amine acceptor. Thus, in one embodiment, the antibody is deglycosylated. In another embodiment, the antibody has an N297Q substitution. Those skilled in the art will appreciate that deglycosylation by postsynthesis modification or by introducing an N297A mutation generates two BTG-reactive glutamine residues per antibody (one per heavy chain, at position 295), while an antibody with an N297Q substitution will have four BTG-reactive glutamine residues (two per heavy chain, at positions 295 and 297).

[0062] An antibody can also be rendered susceptible to BTG-mediated conjugation by introducing into it a glutamine containing peptide, or “tag,” as taught, for example, in Pons et al., US 2013 / 0230543 Al (2013) and Rao-Naik et al., WO 2016 / 144608 Al.

[0063] In another approach, the substrate specificity of BTG can be altered by varying its amino acid sequence, such that it becomes capable of reacting with glutamine 295 in an unmodified antibody, as taught in Strop et al. WO 2020 / 123425 and Rao-Naik et al. WO 2017 / 059158.

[0064] Conjugation can also be effected using the enzyme Sortase A, as taught in Levary et al., PLoS One 2011, 6(4), el 8342; Ploegh et al., WO 2010 / 087994 A2 (2010); and Mao et al., WO 2005 / 051976 A2 (2005).

[0065] ANTIBODIES

[0066] Antibodies that can be used in conjugates of this invention include those recognizing the following antigens: mesothelin, prostate specific membrane antigen (PSMA), CD19, CD22, CD30, CD70, B7H3, B7H4 (also known as O8E), protein tyrosine kinase 7 (PTK7), glypican-3, RG1, fucosyl-GMl, CTLA-4, and CD44. The antibody can be animal (e.g., murine), chimeric, humanized, or, preferably, human. The antibody preferably is monoclonal, especially a monoclonal human antibody. The type can be IgGl, IgG2, IgG3, or IgG4.

[0067] An antibody recognizes, or specifically binds to, an antigen when it preferentially recognizes the antigen in a complex mixture of proteins and / or macromolecules. In some embodiments, an antibody comprises an antigen-binding site that specifically binds to a particular epitope. In some such embodiments, the antibody is capable of binding different antigens so long as the different antigens comprise that particular epitope. In some instances, for example, homologous proteins from different species may comprise the same epitope. In some embodiments, an antibody is said to specifically bind an antigen when the dissociation constant (KD) is < 1 pM, < 100 nM, or < 10 nM.

[0068] Particularly preferred antibodies are those whose antigen is selected from the following list: GPC3, PTK7, DLL3, Claudin 6, Claudin 1, Claudin 18.2, ROR1, ROR2, AXL, PSMA, B7H3 (CD276), B7H4, B7H7, LIV-1, MUC 1, CSPG4, 5T4, Trop2, Nectin4, TF, HER3, CEACAM5, Cadherin 17, Cadherin 6, STn, ITGB6 (integrin beta 6), SEZ6, EGFR, mesothelin, CD70, CD74, CDCP1, FRalpha, cMet, GUCY2C, EFNA4, HER2, EphA5, Napi2B, 5T4, uPARAP, CD30, CD33, CD79b, CD22, CD19, CD46, CD56, BCMA, and GPRC5D. Especially preferred antibodies are anti-GPC3 and anti-PTK7 antibodies.In addition to a full-length antibody, an antibody fragment (such as Fab, Fab', F(ab')2, Fd, or Fv) or antibody mimetic, such as an affibody, a domain antibody (dAb), a nanobody, a unibody, a DARPin, an anticalin, a versabody, a duocalin, a lipocalin, or an avimer can be used.

[0069] Any one of several different reactive groups on the antibody can be a conjugation site, including s-amino groups in lysine residues, pendant carbohydrate moieties, carboxylic acid groups on aspartic or glutamic acid side chains, cysteine-cysteine disulfide groups, and cysteine thiol groups. For reviews on antibody reactive groups suitable for conjugation, see, e.g., Garnett, Adv. Drug Delivery Rev. 2001, 53, 171-216 and Dubowchik and Walker, Pharmacology & Therapeutics 1999, 83, 67-123, the disclosures of which are incorporated herein by reference.

[0070] Most antibodies have multiple lysine residues, which can be conjugated via their e-amino groups via amide, urea, thiourea, or carbamate bonds.

[0071] A thiol (-SH) group in the side chain of a cysteine can be used to form a conjugate by several methods. It can be used to form a disulfide bond between it and a thiol group on the linker. Another method is via its Michael addition to a maleimide group on the linker.

[0072] Typically, although antibodies have cysteine residues, they lack free thiol groups because all their cysteines are engaged in intra- or inter-chain disulfide bonds. To generate a free thiol group, a native disulfide group can be reduced. See, e.g., Naito et al. , US 11,584,236. Alternatively, a cysteine having a free -SH group can be introduced by mutating the antibody, substituting a cysteine for another amino acid or inserting one into the polypeptide chain. See, for example, Eigenbrot et al., US 7,521,541 B2 (2009). In yet another approach, a cysteine is added to the C-terminus of the heavy of light chain. See, e.g., Liu et al., US 8,865,875 B2 (2014). The disclosures of the documents cited in this paragraph are incorporated herein by reference.

[0073] LINKERS

[0074] Sometimes the linker desirably contains a self-immolating group adjacent to the drug DXH (where X represent NEE or OH), especially where the linker is designed to be cleaved enzymatically. The drug DXH may interfere sterically or electronically with the action of the enzyme and the use of a selfimmolating group provides some spatial separation from the enzymatic cleavage site and the drug DXH. Thereafter, the self-immolating group spontaneously self-eliminates to release the active drug DXH. (Hence, a self-immolating group is also referred to as a self-eliminating group.)

[0075] Exemplary self-immolating groups are shown in formula (i) through (viii) below, where X is NH or O and D is the residue of a drug molecule DXH.

[0076]

[0077] The self-immolating moiety is the structure between dotted lines a and b (or dotted lines b and c), with adjacent structural features shown to provide context. Cleavage of the bond at dotted line a by an enzyme

[0078] - a peptidase in the instance of structures (i)-(vi) and a P-glucuronidase in the instance of structure (vii) - initiates a self-immolating reaction sequence that results in the cleavage of the bond at dotted line b and the consequent release of DXH. By way of illustration, self-immolating mechanisms for structures (i),

[0079] (iv), (v), (vi), and (vii) are shown below.

[0080]

[0081] In other words, cleavage of a first chemical bond at one part of a self-immolating group initiates an electronic cascade that results in the cleavage of a second chemical bond - the one connecting the selfimmolating group to the drug - at a different part of the self-immolating group, thereby releasing the drug.

[0082] In some instances, self-immolating groups can be used in tandem, as shown by structure (viii). In such case, cleavage at dotted line a triggers self-immolation of the moiety between dotted lines a and b by a 1,6-elimination reaction, followed by self-immolation of the moiety between dotted lines b and c by a cyclization-elimination reaction.

[0083] For additional disclosures regarding self-immolating moieties, see Carl et al., WO 81 / 01145 (1981); Firestone et al., US 6,214,345 Bl (2001); Boyd et al., US 7,691,962 B2; Boyd et al., Ogitani et al., Cancer Sci. 2016, 107, 1039; US 2008 / 0279868 Al; Sufi et al., WO 2008 / 083312 A2; Feng, US 7,375,078 B2; Jeffrey et al., US 8,039,273; and Senter et al., US 2003 / 0096743 Al; the disclosures of which are incorporated by reference.

[0084] Another optional but frequently desirable component of the linker is a polyethylene glycol) (“PEG”) group. Since conjugation typically entails coupling a drug-linker compound to an antibody in an aqueous medium, a PEG group can be used to enhance the aqueous solubility of the former. Also, a PEG group can reduce aggregation of the resulting ADC. The number of repeat units in a PEG group is typically between 2 to 20, preferably 4 and 10. See for example Howard et al., US 2011 / 0256157.

[0085] For conjugates designed to be internalized by a cell, peptide P preferably comprises an amino acid sequence selected for cleavage by an endosomal or lysosomal protease, especially the latter. Non-limiting examples of such proteases include cathepsins B, C, D, H, L and S, especially cathepsin B. Exemplary cathepsin B cleavable peptides include Ala-Ala-Ala, Vai-Ala, Val-Cit, Val-Lys, Lys-Val-Ala, Asp-Val- Ala, Vai-Ala, Lys-Val-Cit, Ala-Val-Cit, Val-Gly, Val-Gln, and Asp-Val-Cit. (Herein, amino acid sequences are writen in the N-to-C direction, as in FtN-AA^AA'-CChH. unless the context clearly indicates otherwise.) See Dubowchik et al., Biorg. Med. Chem. Lett. 1998, 8, 3341; Dubowchik et al., Bioorg. Med. Chem. Lett. 1998, 8, 3347; and Dubowchik et al. , Bioconjugate Chem. 2002, 13, 855; the disclosures of which are incorporated by reference.

[0086] Another enzyme that can be utilized for cleaving peptidyl linkers is legumain, a lysosomal cysteine protease that preferentially cleaves at Ala-Ala-Asn.

[0087] In one embodiment, peptide P is a peptide comprising atwo-amino acid sequence -AA2-AA'-. wherein AA1is lysine, arginine, or citrulline and AA2is phenylalanine, valine, alanine, leucine or isoleucine. In another embodiment, peptide P consists of a sequence of one to three amino acids, such as Ala-Ala-Ala, Val-Cit, Ala-Vai, Val-Ala-Val, Lys-Lys, Ala-Asn-Val, Val-Leu-Lys, Cit-Cit, Val-Lys, Ala-Ala-Asn, Lys, Cit, Ser, and Glu. In a preferred embodiment, P is a two to three amino acid peptide selected from the group consisting of Ala-Ala-Ala, Val-Cit, Ala-Vai, Val-Ala-Val, Lys-Lys, Ala-Asn-Val, Val-Leu-Lys, Cit-Cit, Val-Lys, Ala-Ala-Asn, Lys, Cit, Ser, and Glu.

[0088] In another embodiment, peptide P is a tetrapeptide of the structure Gly-Gly-Phe-Gly-. See, e.g., Ogitani et al., Cancer Sci 2016 107, 1039.

[0089] In another embodiment, the linker is non-cleavable. Release of the drug DXH relies on the metabolism of the antibody, eventually reducing the linker to a small appended moiety that does not interfere with the biological activity of DXH. See, e.g., Erickson et al., Cancer Res. 2006, 66 (8), 4426.

[0090] DRUG-LINKER COMPOUNDS

[0091] In one embodiment, there is provided a drug-linker compound of formula (II): where R1, R2, R3, and X are as defined hereinabove in the context of formula (I), L is a linker, and Rbis -NH2, -OH, -CO2H, -SH, maleimido, cyclooctyne, azido (-N3), hydroxylamino (-ONH2) or N-hydroxysuccinimido. Preferably, Rbis maleimido. Examples of drug -linkers according to formula (II) include: where x is an integer from 2 to 10, inclusive, y (when present) is an integer from 3 to 6, inclusive, Gly is glycyl and Phe is phenylalanyl.

[0092] A preferred embodiment is

[0093] In a preferred embodiment, the drug-linker compound is according to formula (II’), where

[0094] R1, R2, R3, and X are as defined hereinabove in the context of formula (I); Rbis -NH2, -OH, -CO2H, -SH, maleimido, cyclooctyne, azido (-N3), hydroxylamino (-ONH2) or

[0095] N -hydroxy succinimido ;

[0096] T is a self-immolating group; t is 0, 1, or 2;

[0097] Rcis the side chain of an amino acid selected from the group consisting of alanine, arginine, asparagine, aspartic acid, citrulline, cysteine, glutamic acid, glutamine, glycine, histidine, isoleucine, leucine, lysine, methionine, norleucine, norvaline, ornithine, phenyalanine, proline, serine, threonine, tryptophan, tyrosine, and valine (preferably lysine, arginine, citrulline, phenylalanine, valine, alanine, leucine, isoleucine or glycine); p is 0, 1, 2, 3, or 4; q is 1, 2, 3, 4, 5, 6, 7, 8, 9, 10, 11, 12, 13, 14, 15, 16, 17, or 18; s is 0 or 1 ; and r is 1, 2, 3, 4, 5, or 6.

[0098] Preferably in formula II’, Rbis maleimido, R2is Me, and R3is Cl or F.

[0099] In another embodiment, there is provided a drug-linker compound of the formula (II”) where R1, R2, R3, X, T, t, p, Rc, q, and Rbare as defined in formula (II). Rbpreferably is maleimido, R2is Me, and R3is Cl or F.

[0100] A preferred drug -linker for compounds of this invention where X is O (z.e., hydroxy-terminated ones) is according to formula (III): where R1, R2, R3, p, Rc, q, s, r, and Rbare as defined in formula (II’). Preferably, Rbis maleimido, R2is Me, and R3is Cl or F. Also preferably, p is 4 and the tetrapeptide represented by (NH(RC)HCC(=O))4 is Gly-Gly-Phe-Gly (GGFG).

[0101] Another preferred drug -linker for compounds of this invention where X is O (z.e., hydroxy terminated ones) is according to formula (III’): where R1, R2, R3, p, Rc, q, s, r, and Rbare as defined in formula (IF). Preferably, Rbis maleimido, R2is Me, and R3is Cl or F.

[0102] A preferred drug -linker for compounds of this invention where X is NH (z.e., amine terminated ones) is according to formula (IV) : where R1, R2, R3, p, Rc, q, s, r, and Rbare as defined in formula (IF). Preferably, Rbis maleimido, R2is Me, and R3is Cl or F. o

[0103] The presence of the PEG group (- [NHCH2CH2-(OCH2CH2)q-C]s-) is optional, as illustrated by the

[0104] This disclosure also provides a drug-linker compound comprising homocamptothecin proper, BN 80927, or an analogue thereof, which drug-linker compound is of formula (Ila):

[0105] R5is H, Cl, F, (CH^H, OCCH^^H, or SCCH^H;

[0106] R6is H, Cl, F, (CH^H, OCCH^^H, or SCCH^H;

[0107] L is a linker; and

[0108] Rbis -NH2, -OH, -CO2H, -SH, maleimido, cyclooctyne, azido (-N3), hydroxylamino (-ONH2) or N-hydroxysuccinimido. Preferably, Rbis maleimido.

[0109] Examples or drug -linkers according to formula (Ila) include: where x is an integer from 2 to 10, inclusive, y (when present) is an integer from 3 to 6, inclusive, Gly is glycyl and Phe is phenylalanyl. A preferred embodiment is

[0110] In a preferred embodiment, the drug-linker compound is of formula (Ila’): wherein R4, R5, R6, T, t, p, Rc, s, q, r, and Rbare as defined hereinabove. Preferably, Rbis maleimido. In another preferred embodiment, the drug-linker compound is of formula (Ila”): wherein R4, R5, R6, T, t, p, Rc, q, and Rbare as defined hereinabove. Preferably, Rbis maleimido.

[0111] A preferred drug -linker compound is of formula (Illa): wherein R4, R5, R6, p, Rc, s, q, r, and Rbare as defined hereinabove. Preferably, p is 4 and the tetrapeptide represented by (NH(Rc)HCC(=0))4 is Gly-Gly-Phe-Gly (GGFG). Preferably, Rbis maleimido.

[0112] O

[0113] The presence of the PEG group (- [NHCH2CH2-(OCH2CH2)q-C]s-) is optional, as illustrated by the following embodiment:

[0114] In the above formula, R2is H, Cl, F, (CH2)1.3H, O(CH2)1.3H, or S(CH2)1.3H; and R3is H, Cl, F, (CFy^H, O(CH2)1.3H, or S(CH2)1.3H. Preferably, R2is Me and R3is Cl or F. Most preferably, R2is Me and R3is F.

[0115] Another preferred drug -linker compound is of formula (Illa’): wherein R4, R5, R6, p, Rc, s, q, r, and Rbare as defined hereinabove. Preferably, Rbis maleimido.

[0116] CONJUGATES

[0117] This disclosure provides a conjugate (ADC) according to formula (V): where R1, R2, R3, and X are as defined in respect of formula (I) hereinabove, L’ is a linker, Ab is an antibody, and m is 1, 2, 3, 4, 5, 6, 7 or 8. Preferably, R2is Me and R3is Cl or F. where R1, R2, R3, and X are as defined in respect of formula (I) hereinabove, T, t, p, Rc, s, q, and r are as defined in respect of formula (IF) hereinabove, Ab is an antibody, and m is 1, 2, 3, 4, 5, 6, 7, or 8. wherein R1, X, T, t, p, Rc, q, Ab, and m are as defined hereinabove.

[0118] A preferred conjugate for compounds of this invention where X is O (z.e., hydroxy-terminated ones) is according to formula (VI): where R1, R2, R3, p, Rc, q, s, and r are as defined in formula (IF), Ab is an antibody, and m is 1, 2, 3, 4, 5, 6, 7, or 8. Preferably, R2is Me, and R3is Cl or F. Also preferably, p is 4 and the tetrapeptide represented by (NH(RC)HCC(=O))4is Gly-Gly-Phe-Gly (GGFG).

[0119] O

[0120] The presence of the PEG group (- [NHCH2CH2-(OCH2CH2)q-C]s-) is optional, as illustrated by the

[0121] In the above formula, R2is H, Cl, F, (Chy^H, OCCFy^H, or S(CH2)|.3H: and R3is H, Cl, F, (CH2)I.3H, OCCFy^H, or SCCFy^H. Preferably, R2is Me and R3is Cl or F. Most preferably, R2is Me and R3is F.

[0122] Another preferred conjugate for compounds of this invention where X is O (z.e., hydroxy terminated ones) is according to formula (VF): where R1, R2, R3, p, Rc, q, s, and r are as defined in formula (IF), Ab is an antibody, and m is 1, 2, 3, 4, 5, 6, 7, or 8. Preferably, R2is Me, and R3is Cl or F.

[0123] A preferred conjugate for compounds of this invention where X is NH (z.e., amine terminated ones) is according to formula (VII):

[0124] where R1, R2, R3, p, Rc, q, s, and r are as defined in formula (IF), Ab is an antibody, and m is 1, 2, 3, 4, 5, 6, 7, or 8. Preferably, R2is Me, and R3is Cl or F (especially F).

[0125] This invention also provides an antibody-linker-drug conjugate of formula (Va):

[0126] wherein R4, R5, R6, T, t, p, Rc, s, q, r, Ab, and m are as defined hereinabove.

[0127] Another preferred conjugate is of formula (Va”): wherein R4, R5, R6, T, t, p, Rc, q, Ab, and m are as defined hereinabove. wherein R4, R5, R6, p, Rc, s, q, r, Ab and m are as defined hereinabove. O

[0128] The presence of the PEG group (- [NHCH2CH2-(OCH2CH2)q-C]s-) is optional, as illustrated by the following embodiment: wherein R4, R5, R6, p, Rc, s, q, r, Ab and m are as defined hereinabove.

[0129] PHARMACEUTICAL COMPOSITIONS AND ADMINISTRATION

[0130] In another aspect, there is provided a pharmaceutical composition comprising a compound of as disclosed herein, or of a conjugate thereof, formulated together with a pharmaceutically acceptable carrier or excipient. It may optionally contain one or more additional pharmaceutically active ingredients, such as a biologic or a small molecule drug. The pharmaceutical compositions can be administered in a combination therapy with another therapeutic agent, especially an anti -cancer agent. The pharmaceutical composition may comprise one or more excipients. Excipients that may be used include carriers, surface active agents, thickening or emulsifying agents, solid binders, dispersion or suspension aids, solubilizers, colorants, flavoring agents, coatings, disintegrating agents, lubricants, sweeteners, preservatives, isotonic agents, and combinations thereof.

[0131] Preferably, a pharmaceutical composition is suitable for intravenous, intramuscular, subcutaneous, parenteral, spinal or epidermal administration (e.g., by injection or infusion). Depending on the route of administration, the ADC may be coated in a material to protect it from the action of acids and other natural conditions that may inactivate it. The phrase "parenteral administration" means modes of administration other than enteral and topical administration, usually by injection, and includes, without limitation, intravenous, intramuscular, intraarterial, intrathecal, intracapsular, intraorbital, intracardiac, intradermal, intraperitoneal, transtracheal, subcutaneous, subcuticular, intraarticular, subcapsular, subarachnoid, intraspinal, epidural and intrastemal injection and infusion.

[0132] Pharmaceutical compositions can be in the form of sterile aqueous solutions or dispersions. They can also be formulated in a microemulsion, liposome, or other ordered structure suitable to achieve high drug concentration. The compositions can also be provided in the form of lyophilates, for reconstitution in water prior to administration.

[0133] The amount of active ingredient which can be combined with a carrier material to produce a single dosage form will vary depending upon the subject being treated and the particular mode of administration and will generally be that amount of the composition which produces a therapeutic effect. Generally, out of one hundred per cent, this amount will range from about 0.01 per cent to about ninety-nine percent of active ingredient, preferably from about 0. 1 per cent to about 70 per cent, most preferably from about 1 per cent to about 30 per cent of active ingredient in combination with a pharmaceutically acceptable carrier.

[0134] Dosage regimens are adjusted to provide a therapeutic response. For example, a single bolus may be administered, several divided doses may be administered over time, or the dose may be proportionally reduced or increased as indicated by the exigencies of the situation. It is especially advantageous to formulate parenteral compositions in dosage unit form for ease of administration and uniformity of dosage. “Dosage unit form" refers to physically discrete units suited as unitary dosages for the subjects to be treated; each unit containing a predetermined quantity of active compound calculated to produce the desired therapeutic response, in association with the required pharmaceutical carrier.

[0135] The dosage ranges from about 0.0001 to 100 mg / kg, and more usually 0.01 to 5 mg / kg, of the host body weight. For example, dosages can be 0.3 mg / kg body weight, 1 mg / kg body weight, 3 mg / kg body weight, 5 mg / kg body weight or 10 mg / kg body weight or within the range of 1-10 mg / kg, or alternatively 0. 1 to 5 mg / kg. Exemplary treatment regimens are administration once per week, once every two weeks, once every three weeks, once every four weeks, once a month, once every 3 months, or once every three to 6 months. Preferred dosage regimens include 1 mg / kg body weight or 3 mg / kg body weight via intravenous administration, using one of the following dosing schedules: (i) every four weeks for six dosages, then every three months; (ii) every three weeks; (iii) 3 mg / kg body weight once followed by 1 mg / kg body weight every three weeks.

[0136] A "therapeutically effective amount" of a compound of the invention preferably results in a decrease in severity of disease symptoms, an increase in frequency and duration of disease symptom-free periods, or a prevention of impairment or disability due to the disease affliction. For example, for the treatment of tumor-bearing subjects, a “therapeutically effective amount” preferably inhibits tumor growth by at least about 20%, more preferably by at least about 40%, even more preferably by at least about 60%, and still more preferably by at least about 80% relative to untreated subjects. A therapeutically effective amount of a therapeutic compound can decrease tumor size, or otherwise ameliorate symptoms in a subject, which is typically a human but can be another mammal. Where two or more therapeutic agents are administered in a combination treatment, “therapeutically effective amount” refers to the efficacy of the combination as a whole, and not each agent individually.

[0137] Therapeutic compositions can be administered via medical devices such as (1) needleless hypodermic injection devices; (2) micro-infusion pumps; (3) transdermal devices; (4) infusion devices; and (5) osmotic devices.

[0138] Thus, there is provided a pharmaceutical formulation comprising a conjugate according to formula (V), (V’), (V”), (VI), (VI’) or (VII) and a pharmaceutically acceptable excipient. There is also provided a pharmaceutical formulation comprising a conjugate according to formula (Va), (Va’), (Va”), (Via), or (Via’) and a pharmaceutically acceptable excipient.

[0139] INDUSTRIAL APPLICABILITY AND USES

[0140] Topoisomerase inhibitors of this invention can be used to induce cell death (e.g., cancer cell death). One embodiment of this invention is a method of killing a cell (e.g. , a cancer cell) which comprises contacting a sufficient amount of a compound disclosed herein with the cell.

[0141] Compounds of this invention can also serve as payloads in ADCs which are administered for the treatment of cancer. Thus, one embodiment of this invention provides a method of treating cancer in a patient suffering from such cancer, comprising administering to the patient a therapeutically effective amount of a conjugate disclosed herein (e.g., according to formula (V), (V’), (V”), (VI), (VI’) or (VII)). There is also provided a method of treating cancer in a patient suffering from cancer, comprising administering to the patient a therapeutically effective amount of a conjugate according to formula (Va), (Va’), (Va”), (Via), or (Via’).

[0142] As used herein, the terms “treat,” “treating” and “treatment” contemplate an action that occurs while a patient is suffering from a specified disease or disorder, which reduces the severity of the disease or disorder or retards or slows the progression of the disease or disorder. Examples of cancers that may be treated by methods of this invention include acute myeloid leukemia, adrenocortical carcinoma, Kaposi sarcoma, lymphoma, anal cancer, appendix cancer, teratoid / rhabdoid tumor, basal cell carcinoma, bile duct cancer, bladder cancer, bone cancer, brain cancer, breast cancer, bronchial tumor, carcinoid tumor, cardiac tumor, cervical cancer, chordoma, chronic lymphocytic leukemia, chronic myeloproliferative neoplasm, colon cancer, colorectal cancer, craniopharyngioma, bile duct cancer, endometrial cancer, ependymoma, esophageal cancer, esthesioneuroblastoma, Ewing sarcoma, eye cancer, fallopian tube cancer, gallbladder cancer, gastrointestinal carcinoid tumor, gastrointestinal stromal tumor, germ cell tumor, hairy cell leukemia, head and neck cancer, heart cancer, liver cancer, hypophamgeal cancer, pancreatic cancer, kidney cancer, laryngeal cancer, chronic myelogenous leukemia, lip and oral cavity cancer, lung cancer, melanoma, Merkel cell carcinoma, mesothelioma, mouth cancer, oral cancer, osteosarcoma, ovarian cancer, penile cancer, pharyngeal cancer, prostate cancer, rectal cancer, salivary gland cancer, skin cancer, small intestine cancer, soft tissue sarcoma, testicular cancer, throat cancer, thyroid cancer, urethral cancer, uterine cancer, vaginal cancer, and vulvar cancer.

[0143] The ADCs of this invention can be administered individually, in combination with another ADC, or in combination with another anticancer agent.

[0144] EXAMPLES

[0145] The practice of this invention can be further understood by reference to the following examples, which are provided by way of illustration and not of limitation.

[0146] Schemes 1, 2, and 3

[0147] Example 1-17 illustrates the synthesis of BN 80927. The synthesis is a convergent synthesis involving three schemes (Schemes 1, 2, and 3). BN 80927 serves as a reference compound and several intermediates used in its synthesis are also used to make compounds of this invention.

[0148] Scheme 1

[0149] Example 1 - Compound 2

[0150] To a stirred mixture of 2-chloropyridine-4-carboxylic acid (1) (50 g, 317.360 mmol, 1 equiv, CAS 6313- 54-8) and DMF (1.96 mL, 25.389 mmol, 0.08 equiv) in ACN (500 mL) was added SOCI2 (49.08 g, 412.568 mmol, 1.3 equiv) dropwise at room temperature under nitrogen atmosphere. The mixture was stirred at 85 °C for 1 h. The mixture was concentrated under reduced pressure. The residue was dissolved in DCM (500 mL), and cooled to 0 °C, TEA (38.54 g, 380.832 mmol, 1.2 equiv) and propan-2 -amine (20.64 g, 349.096 mmol, 1.1 equiv) were added. The mixture was stirred at 0 °C for 2 h. The mixture was poured into water (500 mL), and the phases were separated. The organic layer was washed with brine (100 mL), dried over anhydrous sodium sulfate and fdtered. The fdtrate was concentrated under reduced pressure to afford 2-chloro-N-isopropylpyridine-4-carboxamide (2) (61.2 g, 97.01% yield) as a brown solid.

[0151] MS (ESI, m / zy. 199.05 [M + H]+.JH NMR (400 MHz, CDCL3) 6 8.52 - 8.46 (m, 1H), 7.66 - 7.61 (m, 1H), 7.55 - 7.49 (m, 1H), 6.05 (s, 1H), 4.32 - 4.22 (m, 1H), 1.28 (d, J = 6.4 Hz, 6H).

[0152] Example 2 - Compound 3

[0153] To a stirred mixture of 2-chloro-A-isopropylpyridine-4-carboxamide (2) (30 g, 151.019 mmol, 1 equiv) in MeOH (168 mL) was added NaOMe (27.20 g, 151.020 mmol, 5 equiv, 30%) at room temperature under nitrogen atmosphere. The mixture was stirred at 80 °C for 1 h. The reaction was quenched by the addition of aq. NH4CI (sat., 600 mL), extracted with DCM (3 x 600 mL). The combined organic layers were washed with brine (1000 mL), dried over anhydrous Na2SC>4. After filtration, the filtrate was concentrated under reduced pressure to afford A-isopropyl-2-methoxypyridine-4-carboxamide (3) (25 g, 85.23% yield, 95.6% purity) as an off-white solid.

[0154] MS (ESI, m / z): 195.10 [M + H]+.

[0155] 'HNMR (300 MHz, CDCL3) δ 8.29 - 8.21 (m, 1H), 7.19 - 7.11 (m, 1H), 7.07 - 7.01 (m, 1H), 6.00 (s, 1H), 4.35 - 4.17 (m, 1H), 3.96 (s, 3H), 1.26 (d, J= 6.6 Hz, 6H).

[0156] Example 3 - Compound 4

[0157] To a stirred mixture of A-isopropyl-2-methoxypyridine-4-carboxamide (3) (25 g, 128.711 mmol, 1 equiv) and TMEDA (32.91 g, 283.164 mmol, 2.2 equiv) in 2-methoxy-2 -methylpropane (615 mL) was added n- BuLi (2.5 M, 154.45 mL, 386.133 mmol, 3 equiv) dropwise at -78 °C under nitrogen atmosphere. The mixture was stirred at -78 °C for 3 h and subsequently at -22 °C for 3 h. DMF (32.93 g, 450.489 mmol, 3.5 equiv) was added at -78 °C. The mixture was stirred at -78 °C for 16 h. The reaction was quenched by the addition of aq. NH4CI (sat., 300 mL) at 0 °C. The aqueous layer was extracted with DCM (3 x 600 mL). The combined organic layers were washed with brine (1000 mL), dried over anhydrous Na2SC>4. After filtration, the filtrate was concentrated under reduced pressure to afford 3 -hydroxy-2 -isopropyl-4- methoxy-3H-pyrrolo[3,4-c]pyridin-l-one (4) (48 g crude) as a brown oil.

[0158] MS (ESI, m / zy. 223.10 [M + H]+.

[0159] 'H NMR (300 MHz, CDCL3) δ 8.29 (d, J= 5.1 Hz, 1H), 7.95 (s, 1H), 7.23 (d, J= 5.1 Hz, 1H), 5.30 (s, 1H), 4.43 - 4.29 (m, 1H), 4.05 (s, 3H), 2.96 - 2.83 (m, 6H).

[0160] Example 4 - Compound 5

[0161] To a stirred mixture of 3-hydroxy-2-isopropyl-4-methoxy-3H-pyrrolo[3,4-c]pyridin-l-one (4) (48 g, 215.979 mmol, 1 equiv) in 2-propanol (720 mL) and water (240 mL) was added NaBEL (12.26 g, 323.969 mmol, 1.5 equiv) at room temperature under nitrogen atmosphere. The mixture was stirred at room temperature for 3 h. Acetone (84.16 mL, 1144.689 mmol, 5.3 equiv) was added at 0 °C. The mixture was stirred at room temperature for 0.5 h. Then the mixture was poured into 2 M HC1 (1025.90 mL, 2051.801 mmol, 9.5 equiv) at 0 °C. The mixture was stirred at room temperature for 20 min. The mixture was then heated to 50 °C overnight and subsequently cooled to 0 °C. Dipotassium hydrogen phosphate was added to adjust the pH to 3.0, and the 2-propanol was removed under reduced pressure. Water was added until the salts were dissolved, and the mixture was extracted with dichloromethane (3 x 1500 mL). The combined organic phases were dried over anhydrous Na2SC>4. After filtration, the filtrate was concentrated under reduced pressure. The residue was purified by silica gel column chromatography, eluted with PE / EA (1 / 1) to afford 4-methoxy-3H-furo[3,4-c]pyridin-l-one (5) (13 g, 36.45% yield, 96.3% purity) as an off-white solid.

[0162] MS (ESI, m / zy. 166.00 [M + H]+.

[0163] 'HNMR (400 MHz, CDCL3) δ 8.38 - 8.32 (m, 1H), 7.37 (d, J= 5.2 Hz, 1H), 5.29 (d, J= 0.8 Hz, 2H), 4.07 (s, 3H).

[0164] Example 5 - Compound 6

[0165] To a stirred mixture of triethylaluminium (2 M, 43.29 mL, 86.589 mmol, 1.1 equiv) was added ethyllithium (1 M, 86.59 mL, 86.589 mmol, 1.1 equiv) dropwise at 0 °C under nitrogen atmosphere. After 15 min at 0 °C, THF (280 mL, precooled to -40 °C) was added rapidly via a cannula. A solution of 4- methoxy-3H-furo[3,4-c]pyridin-l-one (5) (13 g, 78.717 mmol, 1 equiv) in THF (280 mL) was subsequently added rapidly at -40 °C. After 10 min at -40 °C, additional ethyllithium (86.59 mL, 86.589 mmol, 1.1 equiv) was added slowly. The mixture was allowed to warm to -15 °C within 3 h, and the reaction was afterward quenched by addition of MeOH (15.94 mL, 393.585 mmol, 5 equiv). After 30 min, the mixture was poured into aqueous potassium sodiumtartrate (sat., 4500 mL) and was extracted with dichloromethane (3 x 1500 mL), dried over anhydrous Na2SO4. After fdtration, the filtrate was concentrated under reduced pressure. The residue was purified by RP -flash with the following conditions (Column: AQ-C18Column, 330 g, 60 A, 40-60 pm; Mobile Phase A: water, Mobile Phase B: MeCN; Flow rate: 100 mL / min; 0% B to 100% B in 35 min; Detector: UV 254 / 210 nm.) to afford l-ethyl-4- methoxy-3H-furo[3,4-c]pyridin- 1 -ol (6) (7.8 g, 50.76% yield, 98.7% purity) as an off-white solid.

[0166] MS (ESI, m / zy 196.05 [M + H]+.

[0167] 'HNMR (300 MHz, CDCL3) δ 8.16 (d, J= 5.2 Hz, 1H), 6.91 (d, J= 5.2 Hz, 1H), 5.11 - 4.96 (m, 2H), 4.00 (s, 3H), 2.11 - 2.04 (m, 2H), 0.91 - 0.81 (m, 3H).

[0168] Example 6 - Compound 7

[0169] To a stirred mixture of l -cthyl-4-mcthoxy-37 / -furo|3.4-c|pyridin- l -ol (6) (5 g, 25.612 mmol, 1 equiv) and imidazole (5.23 g, 76.836 mmol, 3 equiv) in DMF (41.75 mL) was added TBSC1 (11.58 g, 76.836 mmol, 3 equiv) at 0 °C under nitrogen atmosphere. The mixture was stirred at 20 °C for 16 h. The mixture was diluted with heptane (500 mL) and washed with water (2 x 300 mL), dried over anhydrous Na2SO4. After fdtration, the fdtrate was concentrated under reduced pressure to afford l-(3-{[(tert- butyldimethylsilyl)oxy]methyl}-2-methoxypyridin-4-yl)propan-l-one (7) (7.9 g, 99.67% yield, 98.0% purity) as a colorless oil. MS (ESI, m / zy. 310.15 [M + H]+.

[0170] 'HNMR (300 MHz, CDCL3) δ 8.10 (d, J= 5.1 Hz, 1H), 6.72 (d, J= 5.1 Hz, 1H), 4.76 (s, 2H), 3.96 (s, 3H), 2.84 - 2.74 (m, 2H), 1.18 - 1.10 (m, 3H), 0.87 (s, 9H), 0.05 (s, 6H).

[0171] Example 7 - Compound 8

[0172] To a stirred mixture of LiHMDS (1 M, 9.69 mL, 9.693 mmol, 3 equiv) was added (4J?)-3 -acetyl -4-phenyl- l,3-oxazolidin-2-one (19) (1989.26 mg, 9.693 mmol, 3 equiv; see synthesis below) in THF (7.5 mL) at - 78 °C under nitrogen atmosphere. The mixture was stirred at -78 °C for 2 h. Then l-(3-{[(tert- butyldimethylsilyl)oxy]methyl}-2-methoxypyridin-4-yl)propan-l-one (7) (1 g, 3.231 mmol, 1 equiv) in THF (6.7 mL) was added at -95 °C (addition time: 60 min). The mixture was stirred at -95 °C for 0.5 h and then -78 °C for 1 h. The reaction was quenched by addition of aqueous 0.5 M HC1 (50 mL). The mixture was extracted with DCM (3 x 50 mL), and the combined extracts were dried over Na2SC>4. After filtration, the filtrate was concentrated under reduced pressure. The residue was purified by silica gel column chromatography, eluted with 25% EA in PE to afford (4J?)-3-[(3J?)-3-(3-{ [(tert- butyldimethylsilyl)oxy]methyl}-2-methoxypyridin-4-yl)-3-hydroxypentanoyl]-4-phenyl-l,3-oxazolidin-2- one (8) (800 mg, 48% yield) as a light yellow oil.

[0173] MS (ESI, m / z 515.30 [M + H]+.

[0174] 'HNMR (300 MHz, CDCL3) δ 7.95 - 7.94 (m, 1H), 7.29 - 7.27 (m, 3H), 7.05 (d, J= 7.8 Hz, 2H), 6.81 - 6.80 (m, 1H), 5.43 - 5.41 (m, 1H), 5.33 - 5.31 (m, 1H), 4.95 (s, 2H), 4.66 - 4.60 (m, 1H), 4.23 - 4.18 (m, 1H), 4.15 (s, 3H), 4.05 - 4.00 (m, 1H), 3.28 (d, J = 16.8 Hz, 1H), 1.95 - 1.84 (m, 2H), 0.88 (s, 9H), 0.79 (d, J= 7.2 Hz, 2H), 0.08 (s, 3H), 0.04 (s, 3H).

[0175] Example 8 - Compound 9

[0176] To a stirred solution of (4 / ?)-3-|(3 / ?)-3-(3-{|( / -biityldimcthylsilyl)oxy ]methyl}-2-methoxypyridin-4-yl)- 3-hydroxypentanoyl]-4-phenyl-l,3-oxazolidin-2-one (8) (1.9 g, 3.692 mmol, 1 equiv) in THF (30 mL) under argon atmosphere was added H2O2 (1.26 g, 36.920 mmol, 10 equiv) at 0 °C, followed by LiOH (0.44 g, 18.460 mmol, 5 equiv). The resulting mixture was stirred at 0 °C for 30 min. To the reaction was added a solution of aqueous NaOH (2 M, 125 mL) at 0 °C and stirred for 2 h at 25 °C. The reaction was washed with TBME (9 x 100 mL). The aqueous phase was acidified with aqueous HC1 (2 M) to adjust the pH to 3.0 and extracted with DCM / EtOH (4 : 1, 9 x lOOmL). The combined organic fractions was dried over anhydrous solium sulfated and filtered. The filtrate was concentrated under reduced pressure to afford crude (3 / ?)-3-hydroxy-3-|3-(hydroxymcthyl)-2-mcthoxypyridin-4-yl |pcntanoic acid (9) (800 mg) as a colorless viscous oil.

[0177] MS (ESI, m / z): 256.05 [M + H]+. Example 9 - Compound 10

[0178] To a stirred solution of (3 / ?)-3-hydroxy-3-|3-(hydroxymcthyl)-2-mcthoxypyridin-4-yl |pcntanoic acid (800 mg, 3.134 mmol, 1 equiv) in 1,2-dimethoxyethane (12 mL) under argon atmosphere was added hydrobromic acid (48%) (0.8 mL) at 25 °C. The resulting mixture was stirred at 25 °C for 15 min. The resulting mixture was stirred 50 °C for 18 h. The precipitate was collected by fdtration and washed with TBME (2 x 2.6 mL), then with acetone (2.6 mL), water (2 x 3.2 mL), finally again with acetone (2 x 2.6 mL) and dried in vacuo to afford (5 / ?)-5-cthyl-5-hydroxy- l / / .4 / / .8 / / -oxcpino|3.4-c|pyridinc-3.9-dione (400 mg, 57.18% yield, 99.9% purity) as an off-white solid.

[0179] MS (ESI, m / z): 224.15 [M + H]+In addition to use as an intermediate in the synthesis of Compound (22a), Compound 10 was used as a common intermediate in subsequent syntheses.

[0180] Scheme 2

[0181] Example 10 Compound 12

[0182] To a mixture of 3-chloro-p-toluidine (20 g, 141.243 mmol, 1 equiv, CAS 95-74-9) in DCE (400 mL) were added boron trichloride (1 M in DCM, 155.37 mL, 155.367 mmol, 1.1 equiv), chloroacetonitrile (12.80 g, 169.492 mmol, 1.2 equiv, CAS 107-14-2) and AICL (24.48 g, 183.616 mmol, 1.3 equiv, added portionwise) at 0 °C under argon atmosphere. The resulting mixture was warmed to 20 °C, then stirred for 2 h at 85 °C. The reaction was quenched with cold 2 N aq. HC1 (600 mL) at 0 °C. The resulting mixture was stirred for 30 min at 20 °C, then extracted with DCM (5 x 800 mL). The combined organic layers were washed with brine (sat., 1 L), dried over Na2SC>4, concentrated under reduced pressure. The residue was purified by silica gel column chromatography, eluted with 0-13% EA in PE to afford l-(2-amino-4- chloro-5-methylphenyl)-2 -chloroethanone (12) (5 g, 16.23%) as a yellow solid.

[0183] MS (ESI, m / zy. 218.00 [M + H]+.

[0184] 'H NMR (300 MHz, DMSO4) 6 7.71 (s, 1H), 7.19 (s, 2H), 6.90 (s, 1H), 5.01 (s, 2H), 2.20 (s, 3H).

[0185] Example 11 - Compound 13

[0186] To a mixture of l-(2-amino-4-chloro-5-methylphenyl)-2-chloroethanone (12) (4 g, 18.342 mmol, 1 equiv) in ACN (85 mL) were added TEA (3.71 g, 36.684 mmol, 2 equiv) and ethyl 3-chloro-3-oxopropanoate (5.52 g, 36.684 mmol, 2 equiv) dropwise at 0 °C under argon atmosphere. The resulting mixture was stirred for 3 h at 20 °C. Then sodium ethoxide (7.49 g, 22.010 mmol, 1.2 equiv, 20%) in EtOH (85 mL) was added. The resulting mixture was stirred for 16 h at 25 °C. The mixture was filtered, the solid formed was washed with EtOH (2 mL), water (2 mL), EtOH (2 x 2 mL) and Et2O (2 x 5 mL) to afford ethyl 7- chloro-4-(chloromethyl)-6-methyl-2-oxo-lH-quinoline-3-carboxylate (13) (4 g, 69.40% yield, 95% purity) as an off-white solid.

[0187] MS (ESI, m / z): 314.00 [M + H]+.

[0188] 'HNMR (300 MHz, DMSO-t / 6) 6 12.27 (s, 1H), 7.93 (s, 1H), 7.41 (s, lH), 4.87 (s, 2H), 4.38 - 4.31 (m, 2H), 2.41 (s, 3H), 1.30 (t, J= 7.2 Hz, 3H).

[0189] Example 12 Compound 14

[0190] A mixture of ethyl 7-chloro-4-(chloromethyl)-6-methyl-2-oxo-lH-quinoline-3-carboxylate (13) (4 g, 12.732 mmol, 1 equiv) in phosphoroyl trichloride (60 mL) was refluxed for 4 h under argon atmosphere. The mixture was cooled and concentrated, diisopropyl ether (60 mL) was added, the solid formed was filtered, and washed with Et2O (10 mL) and PE (10 mL), dried to afford ethyl 2,7-dichloro-4- (chloromethyl)-6-methylquinoline-3-carboxylate (14) (3.1 g, 73.20% yield, 97% purity) as a light grey solid.

[0191] MS (ESI, m / zy. 331.85 [M + H]+.

[0192] 'HNMR (300 MHz, DMSO4) 6 8.37 (d, J= 1.2 Hz, 1H), 8.18 (s, 1H), 5.15 (s, 2H), 4.53 - 4.46 (m, 2H), 2.59 (s, 3H), 1.38 (t, J= 7.2 Hz, 3H).

[0193] Example 13 Compound 15

[0194] To a mixture of ethyl 2,7-dichloro-4-(chloromethyl)-6-methylquinoline-3-carboxylate (14) (2 g, 6.013 mmol, 1 equiv) in DCM (50 mL) was added diisobutylaluminum hydride (1.0 M in DCM) (24.05 mL, 24.052 mmol, 4 equiv) at 0 °C under argon atmosphere. The resulting mixture was stirred for 3 h at 20 °C. The reaction mixture was poured into an aqueous solution of patassium tartrate (20% wt%, 120 mL). The resulting mixture was stirred for 1 h and extracted with DCM (2 x 100 mL). The combined organic layers were washed with brine (sat., 50 mL), dried over Na2SC>4, concentrated. Et2O (10 mL) was added, the solid formed was fdtered, and washed with Et2O (10 mL), dried to afford [2,7-dichloro-4-(chloromethyl)- 6-methylquinolin-3-yl]methanol (15) (1 g, 57.23% yield, 96% purity) as an off-white solid.

[0195] MS (ESI, m / zy. 289.95 [M + H]+.

[0196] 'H NMR (300 MHz, DMSO4) 6 8.30 (s, 1H), 8.09 (s, 1H), 5.52 (s, 1H), 5.37 (s, 2H), 4.84 (s, 2H), 2.57 (s, 3H).

[0197] In addition to being used as an intermediate in the synthesis of Compound (A), Compound (15) was used in the synthesis of other compounds of this invention.

[0198] Example 14 Compound 16

[0199] To a mixture of [2,7-dichloro-4-(chloromethyl)-6-methylquinolin-3-yl]methanol (15) (100 mg, 0.344 mmol, 1.00 equiv) in THF (0.5 mL) was added 4-methylpiperidine (119.46 mg, 1.204 mmol, 3.5 equiv) at 0 °C under argon atmosphere. The resulting mixture was stirred for 2 h at 20 °C. The mixture was extracted with DCM (100 mL), washed with water (30 mL), dried over Na2SC>4, concentrated under reduced pressure. The residue was triturated with Et2O (2 mL), dried to afford {2,7-dichloro-6-methyl-4- [(4-methylpiperidin-l-yl)methyl]quinolin-3-yl} methanol (16) (100 mg, 82.25% yield, 98% purity) as an off-white solid.

[0200] MS (ESI, m / zy 353.20 [M + H]+.

[0201] 'H NMR (300 MHz, DMSO4) 6 8.04 (s, 1H), 7.97 (s, 1H), 6.64 (s, 1H), 4.96 (s, 2H), 4.07 (s, 2H), 2.88 - 2.85 (m, 2H), 2.58 (s, 3H), 2.31 - 2.17 (m, 2H), 1.65 - 1.63 (m, 2H), 1.40 - 1.35 (m, 1H), 1.29 - 1.09 (m, 2H), 0.90 (d, J= 6.6 Hz, 3H).

[0202] Example 15 Compound 17

[0203] To a stirred mixture of {2,7-dichloro-6-methyl-4-[(4-methylpiperidin-l-yl)methyl]quinolin-3-yl}methanol (16) (80 mg, 0.226 mmol, 1 equiv), (57?)-5-ethyl-5-hydroxy-lH,4H,8H-oxepino[3,4-c]pyridine-3, 9-dione (10) (50.55 mg, 0.226 mmol, 1 equiv) and PPh; (65.33 mg, 0.249 mmol, 1.1 equiv) in dioxane (1.5 mL) under argon atmosphere was added DIAD (68.68 mg, 0.339 mmol, 1.5 equiv) at 25 °C. The resulting mixture was stirred at 25 °C for 18 h. The reaction was concentrated under reduced pressure. The residue was purified by prep-TLC, developed using DCM / MeOH (20 / 1) to afford (57?)-8-({2,7-dichloro-6- methyl-4- [(4-methylpiperidin- 1 -yl)methyl]quinolin-3 -yl }methyl)-5 -ethyl-5 -hydroxy- 1 H,4H-oxepino [3 ,4- c]pyridine-3, 9-dione (17) (60 mg, 47.44% yield, 93% purity) as an off-white solid.

[0204] MS ESI (Wz) = 558.20, 560.25 [M + H]+.

[0205] Example 16 BN 80927

[0206] To a stirred solution of (5 / ?)-8-( {2.7-dichloro-6-mcthyl-4-|(4-mcthylpipcridin- l -yl)mcthyl |qllinolin-3- yl [mcthyl)-5-cthyl-5-hydroxy- l / / .4 / / -oxcpino|3.4-c|pyridinc-3.9-dione (17) (60 mg, 0.107 mmol, 1 equiv) in acetonitrile (1.5 mL) under argon atmosphere were added Pd(OAc)2 (3.62 mg, 0.016 mmol, 0.15 equiv), PPhs (14.09 mg, 0.053 mmol, 0.5 equiv), KOAc (15.82 mg, 0.161 mmol, 1.5 equiv) and tetrabutylammonium bromide (38.10 mg, 0.118 mmol, 1.1 equiv) at 25 °C. The resulting mixture was stirred at 90 °C for 16 h. The precipitate was collected by filtration and washed with MeCN (2 x 2 mL), water (2 x 2 mL), then with acetone (2 mL) and Et20 (2 x 2 mL). The solid was purified by Combi-flash with the following: Column: AQ-C18Column, 12 g, 60 A, 40 - 60 pm; Mobile Phase A: water, Mobile Phase B: MeCN; Flow rate: 40 mL / min; 0% B to 100% B in 15 min; Detector: UV 254 / 210 nm; The fractions containing desired product were combined and concentrated under reduced pressure to afford (20 / ?)-6-chloro-20-cthyl-20-hydroxy-7-mcthyl- l 0-|(4-mcthylpipcridin- l -yl)mcthyl |- 17-oxa-3. 13- diazapentacyclo[11.9.0.0A{2,l l}.0A{4,9}.0A{ 15,21}]docosa-l(22),2,4,6,8,10,15(21)-heptaene-14,18- dione (BN 80927) (5.2 mg, 9.27% yield, 96.0% purity) as a light yellow solid.

[0207] MS ESI (m / z) = 522.35, 524.35 [M + H]+.

[0208] ’H-NMR: (400 MHz, DMSO-J6) δ 8.42 (s, 1H), 8.21 (s, 1H), 7.37 (s, 1H), 6.05 (s, 1H), 5.53 (d, J= 15.1 Hz, 1H), 5.39 (d, J= 15.2 Hz, 1H), 5.32 (s, 2H), 4.12 - 3.99 (m, 2H), 3.49 (d, J= 13.7 Hz, 1H), 3.04 (d, J = 13.9 Hz, 1H), 2.84 (d, J= 11.0 Hz, 2H), 2.59 (s, 3H), 2.18 - 2.05 (m, 2H), 1.88 - 1.81 (m, 1H), 1.57 (d, J = 12.4 Hz, 2H), 1.43 - 1.04 (m, 4H), 0.92 - 0.82 (m, 6H).

[0209] Scheme 3

[0210] Example 17 Compound 19

[0211] To a stirred solution of (47?)-4-phenyl-l,3-oxazolidin-2-one (18) (5 g, 30.642 mmol, 1 equiv, CAS 90319- 52-1) in THF (100 mL) was added w-butyllithium (2.5 M in n-hexane) (12.9 mL, 32.174 mmol, 1.05 equiv) at 0 °C within 10 min. The resulting mixture was stirred at 0 °C for 50 min. Then acetyl chloride (2.89 g, 36.770 mmol, 1.2 equiv) was then added within 1 min. After 3.5 h, the reaction was stopped by addition of saturated aqueous NH4CI (25 mL) and the mixture was extracted with ethyl acetate (75 mL). The organic phase was washed with aqueous NaHCOs (50 mL) and brine (20 mL), dried over anhydrous Na2SC>4. After filtration, the filtrate was concentrated under reduced pressure, the residue was purified by silica gel column chromatography, eluted with 0-37% ethyl acetate in petroleum ether to afford (4 / ?)-3 - acetyl-4-phenyl-l,3-oxazolidin-2-one (19) (3.4 g, 54% yield) as a white solid.

[0212] MS (ESI, m / zy. 206.15 [M + H]+.

[0213] ‘HNMR (300 MHz, CDCL3) δ 7.44 - 7.26 (m, 5H), 5.42 - 5.40 (m, 1H), 4.69 (t, J= 8.8 Hz, 1H), 4.29 - 4.26 (m, 1H), 2.53 (s, 3H). Scheme 4

[0214] The synthesis of Compound 22a (and its formate salt) is described in Scheme 4 and Examples 18-21.

[0215] Scheme 4

[0216] Example 18 Compound 20

[0217] To a mixture of [2,7-dichloro-4-(chloromethyl)-6-methylquinolin-3-yl]methanol (15)(200 mg, 0.688 mmol, 1 equiv) in THF (1 mL) was added 4-{[(tert-butyldimethylsilyl)oxy]methyl}piperidine (552.73 mg, 2.408 mmol, 3.5 equiv) dropwise at 0 °C under argon atmosphere. The resulting mixture was stirred at room temperature for 2 h under argon atmosphere. The resulting mixture was diluted with water (50 mL). The resulting mixture was extracted with DCM (3 x 50 mL). The combined organic layers were washed with water (60 mL), dried over anhydrous Na2SC>4. After filtration, the filtrate was concentrated under reduced pressure. The crude product was re -crystallized from Et2O (4 mL) to afford {4-[(4-{[(tert- butyldimethylsilyl)oxy]methyl}piperidin-l-yl)methyl]-2,7-dichloro-6-methylquinolin-3-yl}methanol (20) (222 mg, 66.70% yield, 88.9% purity) as off-white solid.

[0218] MS ESI (m / z) =483.25 [M + H]+.

[0219] Example 19 - Compound 21

[0220] To a stirred mixture of {4-[(4-{[(tert-butyldimethylsilyl)oxy ]methyl }piperidin-l-yl)methyl ]-2,7-dichloro- 6-methylquinolin-3-yl}methanol (20) (140 mg, 0.290 mmol, 1 equiv), (5R)-5-ethyl-5-hydroxy-lH,4H,8H- oxepino [3 ,4-c]pyridine-3, 9-dione (10) (50.55 mg, 0.226 mmol, 1 equiv), and PPh; (83.53 mg, 0.319 mmol, 1.1 equiv) in DMF (3 mL) under argon atmosphere was added DIAD (87.82 mg, 0.435 mmol, 1.5 equiv) at 25 °C. The resulting mixture was stirred at 25 °C for 1 h. The reaction was purified by Combi- flash with the following: Column: AQ-C18Column, 25 g, 60 A, 40 - 60 pm; Mobile Phase A: water, Mobile Phase B: MeCN; Flow rate: 40 mb / min; 0% B to 100% B in 15 min; Detector: UV 254 / 210 nm; The fractions containing desired product were combined and concentrated under reduced pressure to afford (57?)-8-({4-[(4-{[(tert-butyldimethylsilyl)oxy]methyl}piperidin-l-yl)methyl]-2,7-dichloro-6- methylquinolin-3 -yl }methyl)-5 -ethyl-5 -hydroxy- 177,477-oxepino [3 ,4-c]pyridine-3 ,9-dione (21 ) (80 mg, 40.12% yield, 98% purity) as a light yellow solid.

[0221] MS ESI (m / z) = 688.30, 690.30 [M + H]+.

[0222] Example 20 - Compound 22

[0223] To a stirred solution of (57?)-8-({4-[(4-{[(tert-butyldimethylsilyl)oxy]methyl}piperidin-l-yl)methyl]-2,7- dichloro-6-methylquinolin-3-yl}methyl)-5-ethyl-5-hydroxy- 177, 477-oxepino[3,4-c]pyridine-3, 9-dione (21) (80 mg, 0.116 mmol, 1 equiv) in acetonitrile (4 mb) under argon atmosphere were added PPhs (15.23 mg, 0.058 mmol, 0.5 equiv) and tetrabutylammonium bromide (41.19 mg, 0.128 mmol, 1.1 equiv) at 25 °C. The resulting mixture was stirred at 80 °C for 16 h. The reaction was purified by Combi-flash with the following: Column: AQ-C18Column, 25 g, 60 A, 40 - 60 pm; Mobile Phase A: water, Mobile Phase B: MeOH; Flow rate: 40 mb / min; 0% B to 100% B in 20 min; Detector: UV 254 / 210 nm; The fractions containing desired product were combined and concentrated under reduced pressure to afford (207?)- 10- [(4-{[(tert-butyldimethylsilyl)oxy]methyl}piperidin-l-yl)methyl]-6-chloro-20-ethyl-20-hydroxy-7- methyl-17-oxa-3,13-diazapentacyclo[11.9.0.0A{2,l l}.0A{4,9}.0A{ 15,21}]docosa-l(22),2,4,6,8,10,15(21)- heptaene-14, 18-dione (22) (25 mg, 33.00% yield, 95% purity) as a light yellow solid.

[0224] MS ESI (m / z) = 652.35, 654.40 [M + H]+.

[0225] Example 21 - Compound 22a

[0226] To a stirred solution of (207?)-10-[(4-{[(tert-butyldimethylsilyl)oxy]methyl}piperidin-l-yl)methyl]-6- chloro-20-ethyl-20-hydroxy-7-methyl-17-oxa-3,13-diazapentacyclo- [11.9.0.0A{2,l l}.0A{4,9}.0A{ 15,21}]docosa-l(22),2,4,6,8,10,15(21)-heptaene-14,18-dione (21) (40 mg, 0.061 mmol, 1 equiv) in DMSO (2 mb) under argon atmosphere was added EtsN-SHF (296.57 mg, 1.830 mmol, 30 equiv) at 25 °C. The resulting mixture was stirred at 35 °C for 3 h. The reaction was purified by Combi-flash with the following: Column: AQ-C18Column, 12 g, 60 A, 40 - 60 pm; Mobile Phase A: water, Mobile Phase B: MeCN; Flow rate: 40 mb / min; 0% B to 100% B in 20 min; Detector: UV 254 / 210 nm; The fractions containing desired product were combined and concentrated under reduced pressure to afford (20 / ?)-6-chloro-20-cthy 1 -20-hydroxy- 10- { [4-(hydroxymethyl)piperidin- 1 -yl]methyl} -7 - methyl-17-oxa-3,13-diazapentacyclo[11.9.0.0A{2,l l}.0A{4,9}.0A{ 15,21}]docosa-l(22),2,4,6,8,10,15(21)- heptaene-14, 18-dione (B) (15 mg, 45.46% yield, 91% purity) as a light yellow solid, which was purified by prep-HPEC with following conditions: Column: Xselect CSH Prep Column, 19 x 250 mm, 5 pm; Mobile Phase A: Water (0.1% FA), Mobile Phase B: MeCN; Flow rate: 25 mb / min; Gradient: 13% B to 43% B in 10 min, Detector: 254 / 220 nm; RT1: 9.15 min. The product-containing fractions were combined, ro / o-cvaporatcd in vacuo to afford (207?)-6-chloro-20-ethyl-20-hydroxy-10-{[4- (hydroxymethyl)piperidin- 1 -yl]methyl } -7 -methyl- 17 -oxa-3 , 13-diazapentacyclo- [11.9.0.0A{2,l l}.0A{4,9}.0A{ 15,21}]docosa-l(22),2,4,6,8,10,15(21)-heptaene-14,18-dione formate (22a) (7.1 mg, 99.6% purity) as an off-white solid.

[0227] MS ESI (m / z) = 538.25, 539.75 [M + H]+.

[0228] ’H-NMR: (400 MHz, DMSO-J6) δ 8.41 (s, 1H), 8.38 (s, 1H), 8.20 (s, 1H), 7.37 (s, 1H), 6.04 (s, 1H), 5.53 (d, J= 15.2 Hz, 1H), 5.39 (d, J= 15.1 Hz, 1H), 5.31 (s, 2H), 4.42 (s, 1H), 4.12 - 4.00 (m, 2H), 3.53 - 3.45 (m, 1H), 3.24 (d, J= 6.2 Hz, 2H), 3.05 (d, J= 13.9 Hz, 1H), 2.87 (d, J= 10.7 Hz, 2H), 2.59 (s, 3H), 2.12 (t, J= 11.2 Hz, 2H), 1.85 (q, J= 7.4 Hz, 2H), 1.63 (d, J= 11.8 Hz, 2H), 1.45 - 1.32 (m, 2H), 1.19 - 1.04 (m, 2H), 0.86 (t, J= 7.4 Hz, 3H).

[0229] Scheme 5

[0230] The synthesis of Compound (25a) is described in Scheme 5 and Examples 22-25.

[0231] Scheme 5

[0232] Example 22 - Compound 23

[0233] To a mixture of [2,7-dichloro-4-(chloromethyl)-6-methylquinolin-3-yl]methanol (15) (200 mg, 0.688 mmol, 1 equiv) in THF (1 mL) was added 4-[(tert-butyldimethylsilyl)oxy]piperidine (518.94 mg, 2.408 mmol, 3.5 equiv) dropwise at 0 °C under argon atmosphere. The resulting mixture was stirred at room temperature for 2 h under argon atmosphere. The resulting mixture was diluted with water (50 mL). The resulting mixture was extracted with DCM (3 x 50 mL). The combined organic layers were washed with water (60 mL), dried over anhydrous Na2SC>4. After filtration, the filtrate was concentrated under reduced pressure. The crude product was re-crystallized from Et2O (4 mL) to afford [4-({4-[(tert- butyldimethylsilyl)oxy]piperidin-l-yl}methyl)-2,7-dichloro-6-methylquinolin-3-yl]methanol (23) (250 mg, 77.36% yield, 98.0% purity) as off-white solid.

[0234] 'HNMR (400 MHz, DMSO-J6) δ 8.38 (s, 1H), 8.00 (s, 1H), 5.69 (s, 1H), 4.81 (d, J= 5.1 Hz, 2H), 4.07 (s, 2H), 3.69 (s, 1H), 2.65 (s, 2H), 2.52 (s, 3H), 2.31 (s, 2H), 1.64 (d, J = HJEz, 2H), 1.31 (d, J= 11.2 Hz, 2H), 0.83 (s, 9H), 0.00 (s, 6H).

[0235] Example 23 - Compound 24

[0236] To a stirred mixture of [4-({4-[(tert-butyldimethylsilyl)oxy]piperidin-l-yl}methyl)-2,7-dichloro-6- methylquinolin-3-yl]methanol (23) (160 mg, 0.341 mmol, 1 equiv), (57?)-5-ethyl-5-hydroxy-lH,4H,8H- oxepino[3,4-c]pyridine-3, 9-dione (10) (76.07 mg, 0.341 mmol, 1 equiv) and PPhs (98.32 mg, 0.375 mmol, 1.1 equiv) in dioxane (4 mL) under argon atmosphere was added DIAD (103.36 mg, 0.512 mmol, 1.5 equiv) at 25 °C. The resulting mixture was stirred at 25 °C for 18 h. The reaction was concentrated under reduced pressure. The residue was purified by silica gel column chromatography, eluted with ethyl acetate / dichloromethane (1 ~ 80%) to afford (57?)-8-{[4-({4-[(tert-butyldimethylsilyl)oxy]piperidin-l- yl }methyl)-2,7-dichloro-6-methylquinolin-3-yl]methyl } -5 -ethyl-5 -hydroxy- 1 H,4H-oxepino [3,4- c]pyridine-3, 9-dione (24) (140 mg, 60.89% yield, 100% purity) as an off-white solid.

[0237] MS ESI (m / z) = 674.30, 676.30 [M + H]+.

[0238] Example 24 - Compound 25

[0239] To a stirred solution of (57?)-8-{[4-({4-[(tert-butyldimethylsilyl)oxy]piperidin-l-yl}methyl)-2,7-dichloro- 6-methylquinolin-3-yl]methyl } -5 -ethyl-5 -hydroxy- 1 H,4H-oxepino [3 ,4-c]pyridine-3 ,9-dione (24) (140 mg, 0.207 mmol, 1 equiv) in acetonitrile (3 mL) under argon atmosphere were added Pd(OAc)2 (6.99 mg, 0.031 mmol, 0.15 equiv), PPhs (27.21 mg, 0.103 mmol, 0.5 equiv), KOAc (30.54 mg, 0.310 mmol, 1.5 equiv) and tetrabutylammonium bromide (73.58 mg, 0.228 mmol, 1.1 equiv) at 25 °C. The resulting mixture was stirred at 80 °C for 16 h. The precipitate was collected by filtration and washed with MeCN (2 x 2 mL), water (2 x 2 mL), then with acetone (2 mL) and Et2O (2 x 2 mL). The solid was dried in vacuo to afford (207?)-10-({4-[(tert-butyldimethylsilyl)oxy ]piperidin-l-yl }methyl)-6-chloro-20-ethyl-20- hydroxy-7-methyl-17-oxa-3,13-diazapentacyclo[11.9.0.0A{2,l l}.0A{4,9}.0A{15,21}]docosa- l(22),2,4,6,8,10,15(21)-heptaene-14,18-dione (25) (60 mg, 43.95% yield, 97% purity) as a light brown solid.

[0240] MS ESI (m / z) = 638.40, 640.30 [M + H]+.

[0241] Example 25 - Compound 25a

[0242] To a stirred solution of (207?)-10-({4-[(tert-butyldimethylsilyl)oxy]piperidin-l-yl}methyl)-6-chloro-20- ethyl-20-hydroxy-7-methyl-17-oxa-3,13-diazapentacyclo[11.9.0.0A{2,l l}.0A{4,9}.0A{15,21}]docosa- l(22),2,4,6,8,10,15(21)-heptaene-14,18-dione (24) (60 mg, 0.094 mmol, 1 equiv) in THE (1.5 mL) and DMSO (1.5 mL) under argon atmosphere was added EtsN-3HF (303.09 mg, 1.880 mmol, 20 equiv) at 25 °C. The resulting mixture was stirred at 25 °C for 40 h. The reaction was purified by Combi-flash with the following: Column: AQ-C18Column, 12 g, 60 A, 40 - 60 pm; Mobile Phase A: water, Mobile Phase B: MeCN; Flow rate: 40 mb / min; 0% B to 100% B in 15 min; Detector: UV 254 / 210 nm; The fractions containing desired product were combined and concentrated under reduced pressure to afford (207?)-6- chloro-20-ethyl-20-hydroxy- 10- [(4-hydroxypiperidin- 1 -yl)methyl] -7-methyl- 17-oxa-3 , 13 - diazapentacyclo [11.9.O.OA{2,1 l}.0A{4,9}.0A{ 15,21}]docosa-l(22), 2, 4, 6, 8,10, 15(21)-heptaene-14, 18- dione (25a) (25.5 mg, 51.77% yield, 98.4% purity) as a light yellow solid.

[0243] MS ESI = 524.25, 526.25 [M + H]+.

[0244] ’H-NMR: (400 MHz, DMSO-J6) δ 8.43 (s, 1H), 8.21 (s, 1H), 7.37 (s, 1H), 6.05 (s, 1H), 5.53 (d, J= 15.1 Hz, 1H), 5.40 (d, J= 15.1 Hz, 1H), 5.32 (s, 2H), 4.59 (d, J= 4.2 Hz, 1H), 4.12 - 4.00 (m, 2H), 3.49 (d, J= 13.8 Hz, 2H), 3.04 (d, J= 14.0 Hz, 1H), 2.81- 2.72 (m, 2H), 2.59 (s, 3H), 2.28 - 2.18 (m, 2H), 1.88 - 1.80 (m, 2H), 1.75 - 1.67 (m, 2H), 1.44 - 1.35 (m, 2H), 0.86 (t, J= 7.4 Hz, 3H).

[0245] Scheme 6

[0246] The synthesis of Compound 28a is described in Scheme 6 and Examples 26-29.

[0247] Scheme 6

[0248] Example 26 Compound 26

[0249] To a mixture of [2,7-dichloro-4-(chloromethyl)-6-methylquinolin-3-yl]methanol (15) (200 mg, 0.688 mmol, 1 equiv) in THF (1 mL) was added tert-butyl A-(pipcridin-4-yl)carbamatc (482.49 mg, 2.408 mmol, 3.5 equiv) dropwise at 0 °C under argon atmosphere. The resulting mixture was stirred at room temperature for 2 h. The reaction mixture was quenched with water (60 mL) and extracted with DCM (3 x 60 mL). The combined organic layers were dried over Na2SC>4, concentrated under reduced pressure. The residue was triturated with Et2O (5 mL) to afford tert-butyl N-( 1 - { [2,7-dichloro-3-(hydroxymethyl)-6- mcthylquinolin-4-yl |mcthyl [pipcridin-4-yl)carbamatc (26) (280 mg, 89.53% yield, 97.8% purity) as an off-white solid.

[0250] MS (ESI, m / z): 454.00 [M + H]+.

[0251] 'HNMR (400 MHz, DMSO-J6) δ 8.39 (s, 1H), 8.02 (s, 1H), 6.78 (d, J= 8.0 Hz, 1H), 5.69 (s, 1H), 4.83 (s, 2H), 4.09 (s, 2H), 2.76 - 2.74 (m, 2H), 2.55 (s, 3H), 2.49 - 2.35 (m, 2H), 2.30 - 2.20 (m, 1H), 1.79 - 1.58 (m, 4H), 1.36 (s, 9H).

[0252] Example 27 - Compound 27

[0253] To a mixture of (5 / ?)-5-cthyl-5-hydroxy-IH.4H.8H-oxcpino|3.4-c|pyridinc-3.9-dionc (50 mg, 0.224 mmol, 1 equiv), tert-butyl A-(l-{[2,7-dichloro-3-(hydroxymethyl)-6-methylquinolin-4- yl]methyl}piperidin-4-yl)carbamate (26) (101.78 mg, 0.224 mmol, 1 equiv) and PPhs (64.62 mg, 0.246 mmol, 1.1 equiv) in 1,4-dioxane (3 mb) was added DIAD (67.94 mg, 0.336 mmol, 1.5 equiv) at 10 °C under argon atmosphere. The resulting mixture was stirred for 15 h at 20 °C. The mixture was concentrated under reduced pressure. The residue was purified by Prep-TLC, eluted with MeOH / DCM (1 / 15) to afford tert-butyl A'-{ l-|(2.7-dichloro-3-{|(5 / ?)-5-cthyl-5-hydroxy-3.9-dioxo-IH.4H-oxcpino|3.4- c]pyridin-8-yl]methyl}-6-methylquinolin-4-yl)methyl]piperidin-4-yl}carbamate (27) (60 mg, 40.61% yield, 91% purity) as a light grey solid.

[0254] MS (ESI, m / z): 659.15 [M + H]+.

[0255] Example 28 - Compound 28

[0256] A mixture of tert-butyl N-{ I -|(2.7-dichloro-3-{ |(5 / ?)-5-cthyl-5-hydroxy-3.9-dioxo- IH.4H-oxcpino|3.4- c]pyridin-8-yl]methyl}-6-methylquinolin-4-yl)methyl]piperidin-4-yl}carbamate (27) (55 mg, 0.083 mmol, 1 equiv), Pd(OAc)2 (1.87 mg, 0.008 mmol, 0.1 equiv), PPhs (10.94 mg, 0.042 mmol, 0.5 equiv), tetrabutylammonium bromide (29.57 mg, 0.091 mmol, 1.1 equiv) and KOAc (12.28 mg, 0.124 mmol, 1.5 equiv) in acetonitrile (1.5 mb) was stirred at 80 °C for 16 h under argon atmosphere. The resulting mixture was concentrated. The residue was purified by Prep-TLC, eluted with MeOH / DCM (1 / 10) to afford tert-butyl A'-( l-{|(20 / ?)-6-chloro-20-cthyl-20-hydroxy-7-mcthyl- 14. 18-dioxo- 17-oxa-3. 13- diazapentacyclo[11.9.0.0A{2,l l}.0A{4,9}.0A{15,21}]docosa-l(22),2,4,6,8,10,15(21)-heptaen-10- yl]methyl}piperidin-4-yl)carbamate (28) (40 mg, 76.98% yield, 100% purity) as a light yellow solid.

[0257] MS (ESI, m / z): 623.20 [M + H]+.

[0258] Example 29 - Compound 28a

[0259] To a mixture of tert-butyl A-(l-{[(207?)-6-chloro-20-ethyl-20-hydroxy-7-methyl-14,18-dioxo-17-oxa- 3,13-diazapentacyclo[11.9.0.0A{2,l l}.0A{4,9}.0A{15,21}]docosa-l(22),2,4,6,8,10,15(21)-heptaen-10- yl]methyl}piperidin-4-yl)carbamate (28) (40 mg, 0.064 mmol, 1 equiv) in DCM (2 mb) was added trifluoroacetic acid (0.7 mb) at 0 °C under argon atmosphere. The resulting mixture was stirred for 30 min at 20 °C. The mixture was concentrated. The residue was purified by RP-flash, eluted with 0-100% MeOH in 5 M aq. NH4HCO3 to afford (20 / ?)- 10-|(4-aminopipcridin- 1 -yl)mcthyl |-6-chloro-20-cthyl-20- hydroxy-7-methyl-17-oxa-3,13-diazapentacyclo[11.9.0.0A{2,l l}.0A{4,9}.0A{ 15,21}]docosa- l(22),2,4,6,8,10,15(21)-heptaene-14,18-dione (28a) (24.0 mg, 71.49% yield, 96.3% purity) as a light yellow solid.

[0260] MS (ESI, m / z): 523.50 [M + H]+.

[0261] 'HNMR (400 MHz, DMSO-rA) 6 8.41 (s, 1H), 8.20 (s, 1H), 7.37 (s, 1H), 6.06 (s, 1H), 5.53 (d, J = 14.8 Hz, 1H), 5.40 (d, J = 15.2 Hz, 1H), 5.31 (s, 2H), 4.10 - 4.00 (m, 2H), 3.53 - 3.46 (m, 2H), 3.05 (d, J= 13.6 Hz, 1H), 2.82 - 2.80 (m, 2H), 2.58 (s, 3H), 2.25 - 2.11 (m, 2H), 1.92 - 1.79 (m, 2H), 1.68 - 1.66 (m, 2H), 1.37 - 1.18 (m, 4H), 0.90 - 0.82 (m, 3H).

[0262] Scheme 7

[0263] The synthesis of Compound 34 (and its formate salt) is described in Scheme 7 and Examples 30-34.

[0264] Scheme 7

[0265] Example 30 - Compound 30

[0266] To a mixture of 3-(aminomethyl)cyclobutan-l-ol hydrochloride (300 mg, 2.180 mmol, 1 equiv, CAS 1427386-91-1) and imidazole (445.25 mg, 6.540 mmol, 3 equiv) in DCM (3 mL) was added tert- butyl(chloro)dimethylsilane (394.30 mg, 2.616 mmol, 1.2 equiv) at 0 °C under argon atmosphere. The resulting mixture was stirred at room temperature for 3 h. The reaction was quenched with aq. NaHCOs (sat., 60 mL), extracted with DCM (3 x 60 mL). The combined organic layers were washed with brine (10 mL), dried over anhydrous Na2SC>4. After filtration, the filtrate was concentrated under reduced pressure to afford crude l-{3-[(tert-butyldimethylsilyl)oxy]cyclobutyl}methanamine (30) (390 mg, 83.05% yield, 85.56% purity) as a yellow oil.

[0267] MS (ESI, m / zy. 216.10 [M + H]+.

[0268] Example 31 Compound 31

[0269] To a mixture of [2,7-dichloro-4-(chloromethyl)-6-methylquinolin-3-yl]methanol (15) (140 mg, 0.482 mmol, 1.00 equiv) in THF (0.7 mb) was added l-{3-[(tert-butyldimethylsilyl)oxy]- cyclobutyl}methanamine (15) (363.26 mg, 1.687 mmol, 3.5 equiv) dropwise at 0 °C under argon atmosphere. The resulting mixture was stirred at room temperature for 2 h. The reaction was quenched with water (40 mL), extracted with DCM (3 x 40 mb). The combined organic layers were washed with brine (40 mL), dried over anhydrous Na2SO4. After filtration, the filtrate was concentrated under reduced pressure. The residue was purified by silica gel column chromatography, eluted with PE / EA (3 / 1) to afford (4-{[({3-[(tert-butyldimethylsilyl)oxy]cyclobutyl}methyl)amino]methyl}-2,7-dichloro-6- methylquinolin-3-yl)methanol (31) (180 mg, 79.57% yield, 98.4% purity) as an off-white solid.

[0270] MS (ESI, m / z): 469.10 [M + H]+.

[0271] ‘HNMR (300 MHz, DMSO4) 6 8.31 (s, 1H), 8.02 (s, 1H), 4.83 (s, 2H), 4.23 (s, 2H), 4.16 - 4.07 (m, 1H), 2.70 (d, J= 6.9 Hz, 2H), 2.56 (s, 3H), 2.39 - 2.24 (m, 2H), 1.95 - 1.78 (m, 1H), 1.58 - 1.45 (m, 2H), 0.86 (s, 9H), 0.02 (s, 6H).

[0272] Example 32 - Compound 32

[0273] To a stirred mixture of (4-{[({3-[(tert-butyldimethylsilyl)oxy]cyclobutyl}methyl)amino]methyl}-2,7- dichloro-6-methylquinolin-3-yl)methanol (31) (180 mg, 0.383 mmol, 1 equiv), (5 / ?)-5-cthyl-5-hydroxy- lH,4H,8H-oxepino[3,4-c]pyridine-3, 9-dione (10) (85.58 mg, 0.383 mmol, 1.00 equiv) and PPhs (110.61 mg, 0.421 mmol, 1.1 equiv) in 1,4-dioxane (7 mL) was added DIAD (116.28 mg, 0.575 mmol, 1.5 equiv) dropwise at 0 °C under argon atmosphere. The resulting mixture was stirred at room temperature for 16 h. The resulting mixture was concentrated under reduced pressure. The residue was purified by Prep-TLC, eluted with DCM / MeOH (15 / 1) to afford (57?)-8-[(4-{[({3-[(tert- butyldimethylsilyl)oxy] cyclobutyl }methyl)amino]methyl } -2, 7-dichloro-6-methylquinolin-3 -yl)methyl] -5 - ethyl-5-hydroxy-lH,4H-oxepino[3,4-c]pyridine-3, 9-dione (32) (110 mg, 42.52% yield, 99.9% purity) as an off-white solid.

[0274] MS (ESI, m / zy 674.25 [M + H]+.

[0275] Example 33 Compound 33

[0276] A mixture of (57?)-8-[(4-{[({3-[(tert-butyldimethylsilyl)oxy]cyclobutyl}methyl)amino]methyl}-2,7- dichloro-6-methylquinolin-3-yl)methyl]-5-ethyl-5-hydroxy-lH,4H-oxepino[3,4-c]pyridine-3, 9-dione (32) (55 mg, 0.082 mmol, 1 equiv), PPhs (10.69 mg, 0.041 mmol, 0.5 equiv), Pd(OAc)2 (1.87 mg, 0.008 mmol, 0.1 equiv), KOAc (12.00 mg, 0.123 mmol, 1.5 equiv) and tetrabutylammonium bromide (28.91 mg, 0.090 mmol, 1.1 equiv) in MeCN (1.33 mL) was stirred at 80 °C for 8 h under argon atmosphere. The resulting mixture was concentrated under reduced pressure. The residue was purified by Prep-TLC, eluted with DCM / MeOH (14 / 1) to afford (207?)-10-{ [({3-[(tert- butyldimethylsilyl)oxy] cyclobutyl }methyl)amino]methyl } -6-chloro-20-ethyl-20-hydroxy-7 -methyl- 17- oxa-3,13-diazapentacyclo[11.9.0.0A{2,l l}.0A{4,9}.0A{15,21}]docosa-l(22),2,4,6,8,10,15(21)-heptaene- 14,18-dione (33) (27 mg, 51.90% yield, 73% purity) as a light yellow solid.

[0277] MS (ESI, m / zy. 638.35 [M + H]+.

[0278] Example 34 Compound 34

[0279] To a mixture of (207?)-10-{[({3-[(tert-butyldimethylsilyl)oxy]cyclobutyl}methyl)amino]methyl}-6- chloro-20-ethyl-20-hydroxy-7-methyl-17-oxa-3,13-diazapentacyclo- [11.9.0.0A{2,l l}.0A{4,9}.0A{15,21}]docosa-l(22),2,4,6,8,10,15(21)-heptaene-14,18-dione (33) (27 mg, 0.042 mmol, 1 equiv) in DMSO (0.25 mL) was added EtsN.3HF (0.25 mL) at 20 °C. The resulting mixture was stirred for 30 min at 65 °C. The mixture was cooled and purified by RP -flash, eluted with 0- 40% ACN in aq. 0.1% FA to afford (20 / ?)-6-chloro-20-cthyl-20-hydroxy-l0-( {|(3- hydroxycyclobutyl)methyl] amino }methyl)-7 -methyl- 17 -oxa-3 , 13 -diazapentacyclo- [11.9.0.0A{2,l l}.0A{4,9}.0A{15,21}]docosa-l(22),2,4,6,8,10,15(21)-heptaene-14,18-dione formic acid (34) (8.0 mg, 33.18% yield, 98.4% purity) as a light yellow solid.

[0280] MS (ESI, m / zy 524.50 [M + H]+.

[0281] 'H NMR (300 MHz, DMSO4) 8 8.39 - 8.37 (m, 2H), 8.19 (s, 1H), 7.36 (s, 1H), 6.05 (s, 1H), 5.57 - 5.35 (m, 4H), 4.27 (s, 2H), 3.95 - 3.86 (m, 1H), 3.50 (s, 1H), 3.06 - 3.05 (m, 1H), 2.63 - 2.61 (m, 2H), 2.58 (s, 3H), 2.29 - 2.22 (m, 2H), 1.90 - 1.79 (m, 3H), 1.50 - 1.40 (m, 2H), 1.23 (s, 1H), 0.89 - 0.83 (m, 3H).

[0282] Example 35 - Potency Data

[0283] The following protocol was used for evaluating the anti-cancer cell properties of compounds of this disclosure.

[0284] All tested compounds were 3-fold serial diluted from 1 mM for 11 doses in DMSO. For the tested compounds and reference with 0.1% DMSO, the final testing concentrations were: 1000, 333.3, 111.1, 37.04, 12.35, 4.115, 1.372, 0.457, 0.152, 0.051, 0.017 nM.

[0285] Aliquots of 200 nl serial dilution were added into 96-well cell plate by Echo. A 96-well culture plate was seeded with the test cancer cells. Then 200 pl of cell suspension at 7,500 cells / well were dispensed into the 96-well microplate.

[0286] The cells were incubated for 96 h in a 37 °C water incubator with 95% air and 5% CO2. For detection, 100 pL medium was removed and add 100 pL reagent (Celltiter Gio assay kit) per well and shake plates (avoiding light) for 3 min on a plate shaker. The plates were incubated (avoiding light) at room temperature for 30 min. The luminescence was read by Envision.

[0287] Assay robustness check was performed with VC and PC data: LUM: Chemiluminescence signal for each well.

[0288] HC: Average chemiluminescence signal of high control (Cells with medium+DMSO).

[0289] LC: Average chemiluminescence signal of low control(Blank-Only medium).

[0290] GraphPad Prism 8 software was used to graph the results for each compound:

[0291] Y= (HC-A) / (HC-LC)* 100% where Y = % inhibition

[0292] X = dilution ratio

[0293] A = test wells raw data

[0294] Selected compounds were tested for efficacy in inhibiting the growth of various cancer cell lines, alongside comparative compounds, using the above protocol. Results are provided in Table I. Exatecan and MMAE are well-known anticancer agents whose structures are provided below. BN 80927 has been discussed supra.

[0295] Exatecan MMAE

[0296]

[0297] Scheme 8

[0298] The synthesis of compound 39 is described in Scheme 8 and Examples 36-39.

[0299] Scheme 8

[0300] Example 36 Compound 36

[0301] To a solution of [2-bromo-4-(bromomethyl)-7-fluoro-6-methylquinolin-3-yl]methanol (35, CAS 305452- 39-1, CN 117164601) (1 g, 2.755 mmol, 1 equiv) in THF (10 mb) was added tert-butyl A-(piperidin-4- yl)carbamate (1.10 g, 5.510 mmol, 2.0 equiv) at room temperature under Ar. The resulting mixture was stirred at room temperature for 1 h. The mixture was concentrated under reduced pressure and the residue was purified by silica gel column chromatography, eluted with ethyl acetate in petroleum ether (0 - 50%) to afford tert-butyl N-( 1 - { [2-bromo-7 -fluoro-3 -(hydroxymethyl)-6-methylquinolin-4-yl]methyl }piperidin- 4-yl)carbamate (36) (1.24 g, 93.32% yield, 95% purity) as a white solid.

[0302] MS: m / z = 482.00, 484.00 [M + H]+.

[0303] ’H NMR (300 MHz, CDCh), 57.93 (d, J= 7.9 Hz, 1H), 7.65 (d, J= 10.2 Hz, 1H), 5.00 (s, 2H), 4.14 (s, 2H), 3.69 - 3.41 (m, 1H), 3.00 - 2.76 (m, 2H), 2.50 (s, 3H), 2.46 - 2.31 (m, 2H), 2.02 - 1.87 (m, 2H), 1.47 - 1.38 (m, 11H).19F NMR (282 MHz, CDCh) 8 -111.13.

[0304] Example 37 - Compound 37

[0305] To a solution of PPhs (87.00 mg, 0.331 mmol, 1.6 equiv) in THF (1 mb) was added DIAD (62.88 mg, 0.310 mmol, 1.5 equiv) at 0 °C, and the mixture was stirred for 20 min at this temperature. Then a solution of (5R)-5-ethyl-5-hydroxy-lH,4H,8H-oxepino[3,4-c]pyridine-3, 9-dione (10) (46.28 mg, 0.207 mmol, 1 equiv) in THF (0.5 mb) and a solution of tert-butyl A'-( I -{ |2-bromo-7-fluoro-3-(hydroxymcthyl)- 6-methylquinolin-4-yl]methyl}piperidin-4-yl)carbamate (36) (100 mg, 0.207 mmol, 1 equiv) in THF (0.5 mb) were added to the above reaction mixture at 0 °C under Ar. The resulting mixture was stirred at room temperature for 1 h. The resulting mixture was concentrated under reduced pressure. The residue was purified by silica gel column chromatography, eluted with THF in petroleum ether (0 - 50%) to afford tert-butyl N-{ l-[(2-bromo-3-{[(5R)-5-ethyl-5-hydroxy-3,9-dioxo-lH,4H-oxepino[3,4-c]pyridin-8- yl]methyl}-7-fluoro-6-methylquinolin-4-yl)methyl]piperidin-4-yl}carbamate (37) (103.3 mg, 72.47% yield, 95% purity) as an off-white solid.

[0306] MS: m / z = 687.40, 689.40 [M + H]+.

[0307] ’H NMR (300 MHz, CDCh) 8 8.09 (d, J= 8.0 Hz, 1H), 7.72 - 7.61 (m, 2H), 7.58 - 7.42 (m, 1H), 7.02 - 6.91 (m, 1H), 6.33 (d, J = 7.5 Hz, 1H), 4.60 - 4.51 (m, 1H), 3.92 (s, 2H), 3.80 - 3.69 (m, 2H), 3.44 - 3.35 (m, 1H), 3.33 - 3.20 (m, 1H), 3.17 - 3.03 (m, 2H), 2.75 - 2.64 (m, 1H), 2.60 - 2.45 (m, 5H), 2.31 - 2.20 (m, 1H), 2.18 - 2.02 (m, 2H), 1.93 - 1.80 (m, 4H), 1.41 (s, 9H), 0.93 (t, J= 7.4 Hz, 3H).19F NMR (282 MHz, CDCh) 8 -109.52.

[0308] Example 38 - Compound 38

[0309] To a solution of tert-butyl JV-{l-[(2-bromo-3-{[(5R)-5-ethyl-5-hydroxy-3,9-dioxo-lH,4H-oxepino[3,4- c]pyridin-8-yl]methyl } -7 -fluoro-6-methylquinolin-4-yl)methyl]piperidin-4-yl } carbamate (37) ( 157 mg, 0.228 mmol, 1 equiv), KOAc (33.61 mg, 0.342 mmol, 1.5 equiv) and PPhs (35.93 mg, 0.137 mmol, 0.6 equiv) in MeCN (8 mb) was added Pd(OAc)2 (10.25 mg, 0.046 mmol, 0.2 equiv) at room temperature under Ar. The resulting mixture was stirred at 80 °C for 16 h. The resulting mixture was concentrated under reduced pressure. The residue was purified by silica gel column chromatography, eluted with THF in DCM (0 - 30%) to afford tert-butyl JV-(l-{[(20R)-20-ethyl-6-fluoro-20-hydroxy-7-methyl-14,18-dioxo- 17-oxa-3,13-diazapentacyclo-[11.9.0.0A{2,l l}.0A{4,9}.0A{15,21}]docosa-l(22),2,4,6,8,10,15(21)- heptaen-10-yl]methyl}piperidin-4-yl)carbamate (38) (120 mg, 86.63% yield, 95% purity) as a yellow solid. MS: m / z = 607.15 [M + H]+.

[0310] 'HNMR flOO MHz, CDC13) δ 7.86 (d, J= 8.0 Hz, 1H), 7.17 (s, 1H), 6.98 (s, 1H), 6.79 (d, J= 10.5 Hz, 1H), 5.74 - 5.55 (m, 2H), 5.40 (d, J= 15.7 Hz, 1H), 5.12 - 4.90 (m, 2H), 4.66 (d, J= 8.3 Hz, 1H), 4.15 (d, J= 14.7 Hz, 1H), 3.92 (d, J= 14.8 Hz, 1H), 3.66 (s, 1H), 3.50 - 3.28 (m, 3H), 3.01 (d, J= 10.9 Hz, 1H), 2.59 - 2.48 (m, 1H), 2.45 - 2.35 (m, 1H), 2.09 (d, J= 11.6 Hz, 2H), 2.07 - 1.92 (m, 2H), 1.91 - 1.78 (m, 1H), 1.74 - 1.54 (m, 2H), 1.43 (s, 9H), 1.03 (t, J= 7.4 Hz, 3H).19F NMR (376 MHz, CDCh) 5 -111.98.

[0311] Example 39 - Compound 39

[0312] To a solution of tert-butyl JV-(l-{[(207?)-20-ethyl-6-fluoro-20-hydroxy-7-methyl-14,18-dioxo-17-oxa- 3,13-diazapentacyclo[11.9.0.0A{2,l l}.0A{4,9}.0A{ 15,21}]docosa-l(22),2,4,6,8,10,15(21)-heptaen-10- yl]methyl}piperidin-4-yl)carbamate (38) (100 mg, 0.165 mmol, 1 equiv) in THF (1 mL) was added HC1 (4 M in 1,4-dioxane, 1 mL) at room temperature under Ar. The resulting mixture was stirred at 25 °C for 1 h. The resulting mixture was concentrated under reduced pressure. The residue was purified reversed- phase flash chromatography with the following conditions: column, C18silica gel; mobile phase, MeCN in water (0.05% HC1), 0% to 100% gradient in 35 min; detector, UV 254 nm & 210 nm to afford (20R)-10- [(4-aminopiperidin- 1 -yl)methyl] -20-ethyl-6-fluoro-20-hydroxy-7 -methyl- 17 -oxa-3 ,13- diazapentacyclo[11.9.0.0A{2,l l}.0A{4,9}.0A{ 15,21}]docosa-l(22),2,4,6,8,10,15(21)-heptaene-14,18- dione (39), as its hydrochloride (5.8 mg, 6.48% yield, 99.5% purity) as a yellow solid.

[0313] MS: m / z = 507.10 [M + H]+.

[0314] ’H NMR (300 MHz, DMSO-tL) 5 11.30 - 10.80 (m, 1H), 8.76 - 8.57 (m, 1H), 8.54 - 8.24 (m, 2H), 7.93 (d, J= 10.6 Hz, 1H), 7.41 (s, 1H), 5.72 - 5.59 (m, 1H), 5.59 - 5.37 (m, 2H), 5.07 - 4.78 (m, 2H), 3.49 (d, J= 13.9 Hz, 9H), 3.08 (d, J= 13.8 Hz, 1H), 2.57 (s, 3H), 2.18 - 2.00 (m, 3H), 1.86 (q, J= 7.3 Hz, 2H), 0.87 (t, J= 7.3 Hz, 3H).19F NMR (282 MHz, DMSO-tL) 5 -112.44.

[0315] Scheme 9

[0316] The synthesis of compound 40 is described in Scheme 9 and Example 40.

[0317] Scheme 9 Example 40 - Compound 40

[0318] To a solution of glycolic acid (2.80 mg, 0.037 mmol, 1.0 equiv) and HATU (21.01 mg, 0.055 mmol, 1.5 equiv) in A'.A'-dimcthylacctamidc (0.2 mL) were added (20 / ?)- l 0-|(4-aminopipcridin- l -yl)mcthyl |-20- ethyl-6-fluoro-20-hydroxy-7 -methyl- 17 -oxa-3 , 13 -diazapentacyclo- [11.9.0.0A{2,l l}.0A{4,9}.0A{ 15,21}]docosa-l(22),2,4,6,8,10,15(21)-heptaene-14,18-dione (37, as its hydrochloride) (20 mg, 0.037 mmol, 1 equiv) and A'.A'-diisopropylcthylaminc (9.52 mg, 0.074 mmol, 2.0 equiv) at room temperature under Ar. The resulting mixture was stirred at 25 °C for 1 h. The mixture was purified by reversed-phase flash chromatography with the following conditions: column, C1840 g silica gel; mobile phase, MeCN in water (0.05% HC1), 0% to 100% gradient in 35 min; detector, UV 254 nm & 210 nm to afford A'-( l -{ |(20 / ?)-20-cthyl-6-fluoro-20-hydroxy-7-mcthyl- l4. 18-dioxo- l 7-oxa-3. 13- diazapentacyclo[11.9.0.0A{2,l l}.0A{4,9}.0A{ 15,21}]docosa-l(22),2,4,6,8,10,15(21)-heptaen-10- yl]methyl}piperidin-4-yl)-2-hydroxyacetamide (40, as its hydrochloride) (5.9 mg, 26.65% yield, 98.3% purity) as an orange solid.

[0319] MS: m / z = 565.10 [M + H]+.

[0320] 1H NMR (300 MHz, DMSO-J6) δ 8.57 - 8.48 (m, 1H), 7.93 (d, J= 10.5 Hz, 1H), 7.41 (s, 1H), 5.63 - 5.32 (m, 4H), 4.94 (s, 2H), 4.01 - 3.76 (m, 5H), 3.49 (d, J = 15.1 Hz, 5H), 3.12 - 3.00 (m, 1H), 2.56 (s, 3H), 2.10 - 1.76 (m, 6H), 0.85 (t, J= 7.4 Hz, 3H).19F NMR (282 MHz, DMSO-J6) δ -112.08.

[0321] Scheme 10

[0322] The synthesis of Compound 45 is described in Scheme 10 and Examples 41-45.

[0323] Scheme 10 Example 41 - Compound 41

[0324] To a vial charged with [2-bromo-4-(bromomethyl)-7-fluoro-6-methylquinolin-3-yl]methanol (35) (1 g, 2.755 mmol, 1 equiv) was added NtL (7 M in MeOH, 100 mL) and the mixture was stirred at 40 °C for 30 min under nitrogen atmosphere. The mixture was concentrated under reduced pressure. The residue was purified by silica gel column chromatography using 10% - 40% gradient of ethyl acetate in petroleum ether as eluent to afford [4-(aminomethyl)-2-bromo-7-fluoro-6-methylquinolin-3-yl]methanol (41) (650 mg, 78.88% yield, 95% purity) as a light yellow solid.

[0325] MS: m / z = 299.00, 300.95 [M + H]+.

[0326] 'HNMR (300 MHz, DMSO-J6) δ 8.31 (d, J= 8.1 Hz, 1H), 7.77 (d, J= 10.6 Hz, 1H), 7.56 - 6.84 (m, 2H), 4.96 - 4.85 (m, 2H), 4.61 (s, 2H), 3.45 - 3.34 (m, 1H), 2.49 (s, 3H).

[0327] Example 42 - Compound 42

[0328] To a solution of [4-(aminomethyl)-2-bromo-7-fluoro-6-methylquinolin-3-yl]methanol (41) (650 mg, 2.173 mmol, 1 equiv) in THF (4 mL) was added BOC2O (1896.91 mg, 8.692 mmol, 4 equiv) at 25°C under nitrogen atmosphere followed by the addition of A'.A'-Diisopropylcthylaminc (842.51 mg, 6.519 mmol, 3 equiv) dropwise at 25°C. The mixture was stirred for 2 h. The mixture was concentrated in vacuo. The residue was purified by silica gel column chromatography, eluted with 0 - 50% of ethyl acetate in petroleum ether as eluent to afford tert-butyl A'-{|2-bromo-7-fluoro-3-(hydroxymcthyl)-6-mcthylqiiinolin-

[0329] 4-yl]methyl}carbamate (42) (600 mg, 69.16% yield, 95% purity) as a light yellow solid.

[0330] MS: m / z = 399.10, 401.10 [M + H]+.

[0331] ’HNMR (300 MHz, DMSO-J6) δ 8.21 (d, J= 8.3 Hz, 1H), 7.70 (d, J= 10.6 Hz, 1H), 7.47 (s, 1H), 5.26 (t, J= 5.4 Hz, 1H), 4.90 (d, J= 5.4 Hz, 2H), 4.73 (d, J= 5.8 Hz, 2H), 2.43 (d, J= 1.8 Hz, 3H), 1.37 (s, 9H).

[0332] Example 43 - Compound 43

[0333] To a solution of tert-butyl A'-{|2-bromo-7-fluoro-3-(hydroxymcthyl)-6-mcthylqiiinolin-4- yl]methyl}carbamate (42) (550 mg, 1.378 mmol, 1 equiv), (5R)-5-ethyl-5-hydroxy-lH,4H,8H- oxepino[3,4-c]pyridine-3, 9-dione (10) (307.51 mg, 1.378 mmol, 1 equiv) and PPhs (541.98 mg, 2.067 mmol, 1.5 equiv) in DMF (10 mL) was added DIAD (249.26 mg, 1.929mmol, 1.4 equiv) at 0 °C under a nitrogen atmosphere. After stirring for 1 h at 25 °C, the reaction mixture was quenched by water (100 mL) and extracted with ethyl acetate (100 mL x 3). then the combined organic layers were washed with brine (50 mL x 3). The organic layer was dried over anhydrous sodium sulfate and filtered. The filtrate was concentrated in vacuo. The residue was purified by combi flash with the following conditions: SiO?

[0334] Column, 330 g ethyl acetate in petroleum ether 0 - 80% to afford tert-butyl A'-|(2-bromo-3-{|(5 / ?)-5-cthyl-

[0335] 5-hydroxy-3,9-dioxo-lH,4H-oxepino[3,4-c]pyridin-8-yl ]methyl}-7-fluoro-6-methylquinolin-4- yl)methyl] carbamate (43) (600 mg, 72.06% yield, 95% purity) as a white solid.

[0336] MS: m / z = 604.35, 606.35 [M + H]+. ’H NMR (400 MHz, DMSO-tL) 5 8.28 (d, J= 8.3 Hz, 1H), 8.21 (d, J= 5.5 Hz, 1H), 7.76 (d, J= 10.4 Hz, 1H), 7.64 (t, J= 5.6 Hz, 1H), 7.24 (d, J= 5.5 Hz, 1H), 5.81 (s, 1H), 5.76 (d, J= 4.8 Hz, 2H), 5.42 - 5.22 (m, 2H), 4.79 (d, J= 5.7 Hz, 2H), 3.37 - 3.33 (m, 1H), 3.07 (d, J= 13.8 Hz, 1H), 2.46 (s, 3H), 1.81 - 1.64 (m, 2H), 1.32 (d, J= 6.3 Hz, 9H), 0.80 (t, J= 7.4 Hz, 3H).19F NMR (376 MHz, DMSO-tL) 5 -111.32.

[0337] Example 44 - Compound 44

[0338] To a solution of tert-butyl A-|(2-bromo-3-{ |(5 / ?)-5-cthyl-5-hydroxy-3.9-dioxo- IH.4H-oxcpino|3.4- c]pyridin-8-yl]methyl}-7-fluoro-6-methylquinolin-4-yl)methyl]carbamate (43) (380 mg, 0.629 mmol, 1 equiv), KOAc (92.54 mg, 0.944 mmol, 1.5 equiv) and PPhs (98.88 mg, 0.377 mmol, 0.6 equiv) in acetonitrile (12 mb) was added Pd(OAc)2 (28.23 mg, 0.126 mmol, 0.2 equiv) under a nitrogen atmosphere. After stirring for 2 h at 80 °C, the resulting mixture was concentrated under reduced pressure. The residue was purified by silica gel column chromatography using 0% - 10% gradient of MeOH in DCM to afford tert-butyl A'-{ |(20 / ?)-20-cthyl-6-fluoro-20-hydroxy-7-mcthyl- 14. 18-dioxo- 17-oxa-3. 13- diazapentacyclo[11.9.0.0A{2,l l}.0A{4,9}.0A{ 15,21}]docosa-l(22),2,4,6,8,10,15(21)-heptaen-10- yl]methyl}carbamate (44) (220 mg, 66.84% yield, 95% purity) as a yellow solid.

[0339] MS: m / z = 524.40 [M + H]+.

[0340] Example 45 - Compound 45

[0341] To a solution of tert-butyl A-{ |(20 / ?)-20-cthyl-6-fluoro-20-hydroxy-7-mcthyl- 14. 18-dioxo- 17-oxa-3. 13- diazapentacyclo[11.9.0.0A{2,l l}.0A{4,9}.0A{ 15,21}]docosa-l(22),2,4,6,8,10,15(21)-heptaen-10- yl]methyl}carbamate (44) (20 mg, 0.038 mmol, 1 equiv) in tetrahydrofuran (0.2 mL) was added HC1 (4 M in dioxane, 1 mL) at 25 °C and the mixture was stirred for 1 hour under nitrogen atmosphere. The mixture was concentrated under reduced pressure. The residue was purified by reversed-phase flash chromatography with the following conditions: column, C18silica gel; 40 g mobile phase, MeCN in water (0.05% HC1), 2% to 100% gradient in 30 min; detector, UV 254 nm & 210 nm to afford (20R)-10- (aminomethyl)-20-ethyl-6-fluoro-20-hydroxy-7-methyl-17-oxa-3,13- diazapentacyclo[11.9.0.0A{2,l l}.0A{4,9}.0A{ 15,21}]docosa-l(22),2,4,6,8,10,15(21)-heptaene-14,18- dione (45, as its hydrochloride) (5.3 mg, 30.17% yield, 98.6% purity) as a yellow solid.

[0342] MS: m / z = 424.20 [M + H]+.

[0343] ’H NMR (400 MHz, DMSO-tL) 5 8.69 (d, J= 6.4 Hz, 3H), 8.44 (d, J= 8.0 Hz, 1H), 7.94 (d, J= 10.7 Hz, 1H), 7.40 (s, 1H), 6.21 - 5.90 (m, 1H), 5.60 - 5.55 (m, 2H), 5.52 (s, 1H), 5.41 (d, J= 15.1 Hz, 1H), 4.68 (d, J= 5.9 Hz, 2H), 3.48 (d, J= 13.8 Hz, 1H), 3.07 (d, J= 13.8 Hz, 1H), 2.55 (s, 3H), 1.86 (q, J= 7.4 Hz, 2H), 0.87 (t, J = 7.4 Hz, 3H).19F NMR (376 MHz, DMSO-tL) 5 -112.21.

[0344] Scheme 11

[0345] The synthesis of Compounds 46 and 47 is described in Scheme 11 and Examples 46-47. Scheme 11

[0346] To a solution of 3 -hydroxy cyclobutane- 1 -carboxylic acid (7.95 mg, 0.068 mmol, 1 equiv) in N.N- dimethylacetamide (0.3 mL) was added HATU (24.80 mg, 0.065mmol, 1.0 equiv) and the mixture was stirred for 5 min at room temperature. Then (20 / ?)-l0-(aminomcthyl)-20-cthyl-6-fluoro-20-hydroxy-7- methyl-17-oxa-3,13-diazapentacyclo[11.9.0.0A{2,l l}.0A{4,9}.0A{15,21}]docosa-l(22),2,4,6,8,10,15(21)- heptaene-14, 18-dione hydrochloride (45) (20 mg, 0.043 mmol, 1 equiv) and A'.A'-Diisopropylcthylaminc (17.70 mg, 0.136mmol, 2.0 equiv) were added, and the mixture was stirred at 25 °C for 2 h under nitrogen atmosphere. The mixture was purified by reversed-phase flash chromatography with the following conditions: column, C1840 g silica gel; Mobile phase, MeCN in Water (0.05% HC1), 2% to 100% gradient in 30 min; detector, UV 254 nm & 210 nm to afford ( / ?)-A'-((5-cthyl-9-fluoro-5-hydroxy-IO-mcthyl-3. l5- dioxo-4,5, 13, 15-tetrahydro-lH,3H-oxepino[3',4':6,7]indolizino[ l.2- / ?|quinolin-l2-yl)mcthyl)-3- hydroxycyclobutane-1 -carboxamide (46) (5.4 mg, 22.25% yield, 99.6% purity) as a light yellow solid.

[0347] MS: m / z = 522.20 [M + H]+.

[0348] ’HNMR (300 MHz, DMSO-t / e) δ 8.56 (t, J= 5.6 Hz, 1H), 8.31 (d, J= 8.2 Hz, 1H), 7.85 (dd, J= 10.7, 2.7 Hz, 1H), 7.37 (s, 1H), 5.53 (d, J= 15.2 Hz, 1H), 5.46 - 5.31 (m, 3H), 4.83 (d, J= 5.7 Hz, 2H), 4.01 - 3.85 (m, 1H), 3.47 (d, J= 13.8 Hz, 1H), 3.14 - 2.96 (m, 1H),2.5O (s, 3H). 2.46 - 2.34 (m, 1H), 2.32 - 2.21 (m, 2H), 2.03 - 1.90 (m, 2H), 1.90 - 1.78 (m, 2H), 0.86 (t, J= 7.4 Hz, 3H),19F NMR (282 MHz, DMSO - d6) 3 -112.91.

[0349] Example 47 - Compound 47

[0350] To a solution of glycolic acid (5.39 mg, 0.071 mmol, 1.0 equiv) and HATU (26.94 mg, 0.071 mmol, 1.0 equiv) in A'.A'-dimcthylacctamidc (0.3 mL) was added (20 / ?)-l0-(aminomcthyl)-20-cthyl-6-fluoro-20- hydroxy-7-methyl-17-oxa-3,13-diazapentacyclo[11.9.0.0A{2,l l}.0A{4,9}.0A{ 15,21}]-docosa- l(22),2,4,6,8,10,15(21)-heptaene-14,18-dione hydrochloride (45) (30 mg, 0.065 mmol, 1 equiv) and N.N- diisopropylethylamine (18.31 mg, 0.142 mmol, 2.0 equiv) at room temperature under Ar. The resulting mixture was stirred at 25 °C for 1 h. The reaction mixture was purified by reversed-phase flash chromatography with the following conditions: column, 40 g C18silica gel; mobile phase, MeCN in water (0.05% HC1), 0% to 100% gradient in 35 min; detector, UV 254 nm & 210 nm to afford (R)-A-((5 -ethyl - 9-fluoro-5 -hydroxy- 10-methyl-3 , 15 -dioxo-4,5 , 13,15 -tetrahydro- 1 H,3H-oxepino [3 ',4' : 6,7]indolizino [1,2- b]quinolin-12-yl)methyl)-2 -hydroxyacetamide (47) (11.7 mg, 34.63% yield, 96.0% purity) as a white solid.

[0351] MS: m!z = 482.10 [M + H]+.1

[0352] H NMR (300 MHz, DMSO-d6) δ 8.74 (t, J= 6.1 Hz, 1H), 8.45 (d, J= 8.3 Hz, 1H), 7.85 (d, J= 10.8 Hz, 1H), 7.37 (s, 1H), 6.04 (s, 1H), 5.63 - 5.37 (m, 5H), 4.83 (d, J= 6.0 Hz, 2H), 3.84 (d, J= 5.7 Hz, 2H), 3.48 (d, J= 13.8 Hz, 1H), 3.06 (d, J= 13.8 Hz, 1H), 2.50 (s, 3H), 1.85 (d, J= 7.5 Hz, 2H), 0.86 (t, J= 7.4 Hz, 3H).19F NMR (282 MHz, DMSO-d6) δ -112.99.

[0353] Scheme 12

[0354] The synthesis of Compound 51 is described in Scheme 12 and Examples 48-50.

[0355] Scheme 12

[0356] To a stirred solution of 2- [(tert-butyldimethylsilyl)oxy] ethanol (48) (656.12 mg, 3.721 mmol, 1.5 equiv) in DCM (4 mb) was added DIEA (641.23 mg, 4.962 mmol, 2 equiv) and 4-nitrophenyl carbonochloridate (500 mg, 2.481 mmol, 1 equiv) in portions at 25 °C under Argon atmosphere. The reaction was monitored by LCMS. The resulting mixture was concentrated under reduced pressure. The residue was purified by silica gel column chromatography, eluted with petroleum ether : DCM (1 : 1) to afford 2-[(tert- butyldimethylsilyl)oxy] ethyl 4-nitrophenyl carbonate (49) (500 mg, 59.03% yield, 95% purity) as a colorless oil.

[0357] MS: m / z = 342.35 [M + H]+.

[0358] 'HNMR (400 MHz, CDC13) δ 8.34 - 8.25 (m, 2H), 7.43 - 7.33 (m, 2H), 4.40 - 4.34 (m, 2H), 3.94 - 3.89 (m, 2H), 0.91 (s, 9H), 0.10 (s, 6H).

[0359] Example 49 - Compound 50

[0360] A mixture of (20 / ?)-l0-(aminomcthyl)-20-cthyl-6-fluoro-20-hydroxy-7-mcthyl-l 7-oxa-3. l3- diazapentacyclo[11.9.0.0A{2,l l}.0A{4,9}.0A{15,21}]docosa-l(22),2,4,6,8,10,15(21)-heptaene-14,18- dione hydrochloride (45) (100 mg, 0.217 mmol, 1 equiv) , 2-|( / c / 7-biityldimcthylsilyl)oxy |cthyl 4- nitrophenyl carbonate (49) (74.24 mg, 0.217 mmol, 1.00 equiv) and DIEA (56.21 mg, 0.434 mmol, 2 equiv) in A.A-dimcthylacctamidc (2.5 mb, 5.378 mmol) was stirred at 25 °C for 2 h under Argon atmosphere. The mixture was purified by reverse phase flash with the following conditions: MeCN : H2O (0.05% FA), 5% - 100% to afford 2-((tert-butyldimethylsilyl)oxy)ethyl ( / ?)-((5-cthyl-9-fluoro-5-hydroxy- 10-methyl-3, 15-dioxo-4,5, 13, 15 -tetrahydro- lH,3H-oxepino[3',4':6,7]indolizino[ 1.2- / ? |qninolin- 12- yl)methyl) carbamate (50) (100 mg, 73.49% yield, 87.06% purity) as a white solid.

[0361] MS: m / z = 626.25 [M + H]+.

[0362] Example 50 - Compound 51

[0363] A mixture of 2-[(tert-butyldimethylsilyl)oxy]ethyl A-{|(20R)-20-cthyl-6-fluoro-20-hydroxy-7-mcthyl- 14,18-dioxo-17-oxa-3,13-diazapentacyclo[11.9.0.0A{2,l l}.0A{4,9}.0A{15,21}]docosa- l(22),2,4,6,8,10,15(21)-heptaen-10-yl]methyl}carbamate (50) (75 mg, 0.120 mmol, 1 equiv) in triethylamine trihydrofluoride (0.75 mb) and DMSO (0.75 mb) was stirred at 65 °C for 1 h. The mixture was purified by reverse phase flash with the following conditions MeCN in water (0.05% HC1), 5% - 100% to afford 2-hydroxyethyl A'-{|(20 / ?)-20-cthyl-6-fluoro-20-hydroxy-7-mcthyl-l4. l 8-dioxo-l 7-oxa- 3,13-diazapentacyclo[11.9.0.0A{2,l l}.0A{4,9}.0A{15,21}]docosa-l(22),2,4,6,8,10,15(21)-heptaen-10- yl]methyl}carbamate (51) (20.7 mg, 33.36% yield, 97.8% purity) as a light yellow solid.

[0364] MS: m / z = 512.15 [M + H]+.

[0365] ’HNMR (400 MHz, DMSO-J6) δ 8.33 (d, J= 8.1 Hz, 1H), 8.14 (t, J= 6.0 Hz, 1H), 7.91 - 7.82 (m, 1H), 7.37 (s, 1H), 6.04 (s, 1H), 5.53 (d, J= 15.2 Hz, 1H), 5.47 - 5.32 (m, 3H), 4.85 - 4.65 (m, 2H), 4.03 - 3.87 (m, 2H), 3.55 - 3.50 (m, 2H), 3.48 (d, J= 13.8 Hz, 2H), 3.05 (d, J= 13.6 Hz, 1H), 2.51 (s, 3H), 1.85 (q, J = 7.4 Hz, 2H), 0.86 (t, J= 7.4 Hz, 3H).19F NMR (376 MHz, DMSO-J6) δ -112.89.

[0366] Scheme 13

[0367] The synthesis of Compound 54 is described in Scheme 13 and Examples 51-53. Scheme 13

[0368] To a stirred mixture of 2-bromo-4-(bromomethyl)-7-fluoro-6-methylquinolin-3-yl]methanol (35) (300 mg, 0.826 mmol, 1 equiv) in THF (3 mb) under argon atmosphere was added 4-methylpiperidine (245.88 mg, 2.478 mmol, 3 equiv) at 0 °C. The resulting mixture was stirred at 0 °C for 1 h. The mixture was purified by silica gel column chromatography, eluted with ethyl acetate in petroleum ether (0 - 30%) to afford {2- bromo-7-fhioro-6-methyl-4-[(4-methylpiperidin-l-yl)methyl]quinolin-3-yl}methanol (52) (256.5 mg, 81.4% yield, 98.4% purity) as a white solid.

[0369] MS: m / z = 381.25, 383.25 [M + H]+.

[0370] 1HNMR (400 MHz, CDC13) δ 7.94 (dd, J= 7.9, 1.2 Hz, 1H), 7.62 (d, J= 10.2 Hz, 1H), 6.53 (s, 1H), 4.96 (s, 2H), 4.06 (s, 2H), 2.90 - 2.79 (m, 2H), 2.49 (s, 3H), 2.24 - 2.17 (m, 2H), 1.68 - 1.57 (m, 2H), 1.51 - 1.35 (m, 1H), 1.16 - 1.2 (m, 2H), 0.89 (d, J= 6.5 Hz, 3H).19F NMR (376 MHz, CDC13) 8 -111.55.

[0371] Example 52 - Compound 53

[0372] To a solution of PPhs (210.50 mg, 0.802 mmol, 1.5 equiv) in THF (2.0 mb) was added DIAD (151.46 mg, 0.749 mmol, 1.4 equiv) at 0 °C and the mixture was stirred for 30 min under nitrogen atmosphere followed by the addition of (5R)-5-ethyl-5-hydroxy-lH,4H,8H-oxepino[3,4-c]pyridine-3, 9-dione (10) (119.43 mg, 0.535 mmol, 1 equiv) and {2-bromo-7-fluoro-6-methyl-4-[(4-methylpiperidin-l- yl)methyl]quinolin-3-yl}methanol (52) (204 mg, 0.535 mmol, 1 equiv) in portions at 25 °C. The resulting mixture was stirred at 25°C for 2 h. The mixture was purified by silica gel column chromatography, eluted with ethyl acetate in petroleum ether (0 - 70%) to afford (5R)-8-({2-bromo-7-fluoro-6-methyl-4-[(4- methylpiperidin- 1 -yl)methyl] quinolin-3 -yl }methyl)-5 -ethyl-5 -hydroxy- 1 H,4H-oxepino [3 ,4-c]pyridine- 3, 9-dione (53) (150 mg, 47.80% yield, 95% purity) as a white solid. MS: m / z = 586.20, 588.20. [M + H]+.

[0373] 'HNMR (400 MHz, CDC13) δ 8.14 (d, J= 8.0 Hz, 1H), 7.64 (d, J= 10.1 Hz, 1H), 6.95 (d, J= 7.5 Hz, 1H), 6.32 (d, J= 7.5 Hz, 1H), 5.68 (d, J= 15.3 Hz, 1H), 5.48 (d, J= 2.0 Hz, 2H), 5.26 (d, J= 15.3 Hz, 1H), 3.90 (q, J= 13.4 Hz, 2H), 3.42 (d, J= 13.5 Hz, 1H), 3.04 (d, J= 13.6 Hz, 1H), 2.73 - 2.63 (m, 2H), 2.56 (s, 1H), 2.50 (s, 3H), 2.16 - 2.05 (m, 2H), 1.89 - 1.82 (m, 2H), 1.54 - 1.43 (m, 2H), 0.92 (d, J= 7.4 Hz, 4H), 0.83 (d, J= 6.5 Hz, 3H).19F NMR (376 MHz, CDC13) δ -109.85.

[0374] Example 53 Compound 54

[0375] To a solution of (5 / ?)-8-( {2-bromo-7-fluoro-6-mcthyl-4-|(4-mcthylpipcridin- 1 -yl (methyl |quinolin-3- yl}methyl)-5-ethyl-5-hydroxy-lH,4H-oxepino[3,4-c]pyridine-3, 9-dione (53) (150 mg, 0.256 mmol, 1 equiv), PPhs (33.54 mg, 0.128 mmol, 0.50 equiv), KOAc (37.65 mg, 0.384 mmol, 1.5 equiv) in MeCN (7.5 mL) was added Pd(OAc)2 (5.74 mg, 0.026 mmol, 0.1 equiv) at room temperature under Argon. The resulting mixture was stirred at 80 °C for 16 h. The resulting mixture was concentrated under reduced pressure. The residue was purified by silica gel column chromatography, eluted with 0 - 100% (DCM : MeOH = 20 : 1) in DCM to afford the product. The product was further purified by reverse phase flash with the following conditions: 0 - 100% MeCN in water (0.05% HC1) to afford (20 / ?)-20-cthyl-6-fluoro- 20-hydroxy-7 -methyl- 10-[(4-methylpiperidin- 1 -yl)methyl] - 17-oxa-3 , 13 -diazapentacyclo [11.9.0.0A{2,l l}.0A{4,9}.0A{ 15,21}]docosa-l(22),2,4,6,8,10,15(21)-heptaene-14,18-dione (54, as its hydrochloride )(34.5 mg, 24.89% yield, 99.4% purity) as a yellow solid.

[0376] MS: m / z = 506.10 [M + H]+.

[0377] ’H NMR (400 MHz, DMSO-J6) δ 10.27 (s, 1H), 8.60 (d, J= 8.0 Hz, 1H), 7.94 (d, J= 10.6 Hz, 1H), 7.41 (s, 1H), 5.65 - 5.47 (m, 3H), 5.41 (d, J= 15.1 Hz, 1H), 4.99 - 4.83 (m, 2H), 3.57 - 3.40 (m, 3H), 3.31 (q, J= 10.5 Hz, 2H), 3.07 (d, J= 13.8 Hz, 1H), 2.56 (s, 3H), 1.85 (q, J= 7.4 Hz, 2H), 1.78 - 1.67 (m, 2H), 1.66 - 1.47 (m, 3H), 0.94 - 0.80 (m, 6H).19F NMR (376 MHz, DMSO-J6) δ -112.43.

[0378] Scheme 14

[0379] The synthesis of compound (58) is described in Scheme 14 and Examples 54-57.

[0380] Scheme 14

[0381] Example 44 - Compound 55

[0382] To a solution of [2-bromo-4-(bromomethyl)-7-fluoro-6-methylquinolin-3-yl]methanol (35) (500 mg, 1.377 mmol, 1 equiv) in THF (5 mb) was added 4-{[(tert-butyldimethylsilyl)oxy]methyl}piperidine (1011.23 mg, 4.406 mmol, 3.2 equiv) at room temperature under Ar. The resulting mixture was stirred at room temperature for 1 h. The residue was purified by silica gel column chromatography using 10% - 50% gradient of ethyl acetate in petroleum ether to afford {2-bromo-4-[(4-{[(tert- butyldimethylsilyl)oxy]methyl }piperidin- 1 -yl)methyl] -7 -fluoro-6-methylquinolin-3 -y I [ methanol (55) (607.4 mg, 86.21% yield, 95% purity) as a white solid.

[0383] MS: m / z = 511.15, 512.75 [M + H]+.

[0384] 1HNMR (400 MHz, CDC13) δ 7.94 (d, J= 7.8 Hz, 1H), 7.63 (d, J= 10.3 Hz, 1H), 6.50 (s, 1H), 4.98 (s, 2H), 4.09 (s, 2H), 3.52 - 3.28 (m, 2H), 3.00 - 2.84 (m, 2H), 2.49 (s, 3H), 2.23 (t, J= 11.4 Hz, 2H), 1.86 - 1.66 (m, 2H), 1.64 - 1.48 (m, 1H), 1.28 - 1.06 (m, 2H), 0.85 (s, 9H), -0.00 (s, 6H).19F NMR (376 MHz, CDC13) δ -111.48.

[0385] Example 55 - Compound 56

[0386] To a solution of PPhs (205.10 mg, 0.782 mmol, 1.6 equiv) in THF (2 mb) was added DIAD (148.23 mg, 0.734 mmol, 1.5 equiv) at 0 °C under Ar. The mixture was stirred for 20 min. Then a solution of (5R)-5- ethyl-5-hydroxy-lH,4H,8H-oxepino[3,4-c]pyridine-3, 9-dione (10) (109.09 mg, 0.489 mmol, 1.0 equiv) in THF (0.5 mb) and a solution of {2-bromo-4-[(4-{[(tert-butyldimethylsilyl)-oxy]methyl }piperidin-l- yl)methyl]-7-fhroro-6-methylquinolin-3-yl}methanol (55) (250 mg, 0.489 mmol, 1 equiv) in THF (0.5 mb) were added to the above reaction mixture at 0 °C under Ar. The resulting mixture was stirred at room temperature for 1 h. The resulting mixture was concentrated under reduced pressure. The residue was purified by silica gel column chromatography using 10% - 50% gradient of ethyl acetate in petroleum ether to afford (5R)-8-({2-bromo-4-[(4-{[(tert-butyldimethylsilyl)oxy]methyl}piperidin-l-yl)methyl]-7- fhioro-6-methylquinolin-3-yl}methyl)-5-ethyl-5-hydroxy-lH,4H-oxepino[3,4-c]pyridine-3, 9-dione (56) (178 mg, 50.82% yield, 95% purity) as an off-white solid.

[0387] MS: m / z = 716.40, 718.40 [M + H]+.

[0388] 1HNMR (300 MHz, CDC13) δ 8.15 (d, J= 8.0 Hz, 1H), 7.63 (d, J= 10.1 Hz, 1H), 6.96 (d, J= 7.5 Hz, 1H), 6.31 (d, J= 7.5 Hz, 1H), 5.66 (d, J= 15.3 Hz, 1H), 5.50 (s, 2H), 5.26 (d, J= 15.3 Hz, 1H), 4.00 - 3.82 (m, 2H), 3.44 - 3.29 (m, 3H), 3.03 (d, J= 13.6 Hz, 1H), 2.79 - 2.66 (m, 2H), 2.53 (s, 1H), 2.50 (s, 3H), 2.18 - 2.07 (m, 2H), 1.84 (q, J= 7.5 Hz, 2H), 1.66 - 1.60 (m, 1H), 1.51 - 1.36 (m, 1H), 0.91 (t, J = 7.6 Hz, 5H), 0.85 (s, 9H), 0.00 (s, 6H).19F NMR (282 MHz, CDC13) δ -109.85.

[0389] Example 56 Compound 57

[0390] To a solution of (5R)-8-({2-bromo-4-[(4-{[(tert-butyldimethylsilyl)oxy ]methyl }piperidin-l-yl)methyl ]-7- fhioro-6-methylquinolin-3-yl}methyl)-5-ethyl-5-hydroxy-lH,4H-oxepino[3,4-c]pyridine-3, 9-dione (56) (108 mg, 0.151 mmol, 1 equiv), KO Ac (22.18 mg, 0.226 mmol, 1.5 equiv) and PPhs (19.76 mg, 0.075 mmol, 0.5 equiv) in MeCN (6 mL) was added Pd(OAc)2 (3.38 mg, 0.015 mmol, 0.1 equiv) at room temperature under Ar. The resulting mixture was stirred at room temperature for 1 h. The resulting mixture was concentrated under reduced pressure. The residue was purified by silica gel column chromatography, eluted with 0 - 30% THF in DCM to afford (20R)-10-[(4-{[(tert- butyldimethylsilyl)oxy]methyl}piperidin-l-yl)methyl]-20-ethyl-6-fluoro-20-hydroxy-7-methyl-17-oxa- 3,13-diazapentacyclo[11.9.0.0A{2,l l}.0A{4,9}.0A{15,21}]docosa-l(22),2,4,6,8,10,15(21)-heptaene- 14, 18-dione (57) (98 mg, 91% yield, 89% purity) as a light yellow solid.

[0391] MS: m / z = 636.25 [M + H]+.

[0392] 'HNMR (300 MHz, DMSO-J6) δ 8.35 (d, J= 8.5 Hz, 1H), 7.81 (d, J= 10.8 Hz, 1H), 7.35 (s, 1H), 6.01 (s, 1H), 5.52 (d, J= 15.1 Hz, 1H), 5.38 (d, J= 15.1 Hz, 1H), 5.28 (s, 2H), 4.03 (s, 2H), 3.51 - 3.37 (m, 3H), 3.04 (d, J= 13.9 Hz, 1H), 2.87 (d, J= 10.8 Hz, 2H), 2.49 (s, 3H). 2.10 (d, J= 11.3 Hz, 2H), 1.84 (d, J= 7.6 Hz, 2H), 1.69 - 1.57 (m, 2H), 1.50 - 1.38 (m, 1H), 1.20 - 1.09 (m, 2H), 0.84 (d, J= 3.3 Hz, 12H), 0.00 (s, 6H),19F NMR (282 MHz, DMSO-tL) 5 -113.08.

[0393] Example 57 - Compound 58

[0394] To a solution of (20R)-10-[(4-{[(tert-butyldimethylsilyl)oxy]methyl}piperidin-l-yl)methyl]-20-ethyl-6- fluoro-20-hydroxy-7-methyl-17-oxa-3,13-diazapentacyclo-[11.9.0.0A{2,l l}.0A{4,9}.0A{15,21}]docosa- l(22),2,4,6,8,10,15(21)-heptaene-14,18-dione (57) (93 mg, 0.146 mmol, 1 equiv) in THF (0.5 mL) was added HC1 (4 M in 1,4-dioxane, 1 mL) at room temperature under Ar. The resulting mixture was stirred at 25 °C for 1 h. The resulting mixture was concentrated under reduced pressure. The residue was purified by reversed-phase flash chromatography with the following conditions: column, C18silica gel; mobile phase, MeCN in Water (0.05% HC1), 0% to 100% gradient in 35 min; detector, UV 254 nm & 210 nm to afford (207?)-20-ethyl-6-fluoro-20-hydroxy- 10- { [4-(hydroxymethyl)piperidin- 1 -yl]methyl } -7 -methyl- 17- oxa-3,13-diazapentacyclo[11.9.0.0A{2,l l}.0A{4,9}.0A{15,21}]docosa-l(22),2,4,6,8,10,15(21)-heptaene- 14,18-dione hydrochloride (58) (48.1 mg, 58.93% yield, 98.9% purity) as an orange solid.

[0395] MS: m / z = 522.10 [M + H]+.

[0396] 'HNMR (300 MHz, DMSO-J6) δ 10.08 (s, 1H), 8.58 (d, J= 7.9 Hz, 1H), 7.95 (d, J= 10.6 Hz, 1H), 7.41 (s, 1H), 5.55 (d, J= 18.8 Hz, 3H), 5.41 (d, J= 15.1 Hz, 1H), 5.00 - 4.88 (m, 4H), 3.49 (d, J= 13.6 Hz, 3H), 3.39 - 3.20 (m, 4H), 3.07 (d, J= 13.7 Hz, 1H), 2.57 (s, 3H), 1.91 - 1.67 (m, 4H), 1.68 - 1.49 (m, 3H), 0.86 (t, J= 7.2 Hz, 3H).19F NMR (282 MHz, DMSO-J6) δ -112.36.

[0397] Scheme 15

[0398] The synthesis of Compound 62 is described in Scheme 15 and Examples 58-61.

[0399] Scheme 15

[0400] Example 58 - Compound 59

[0401] To a solution of [2-bromo-4-(bromomethyl)-7-fluoro-6-methylquinolin-3-yl]methanol (35) (440 mg, 1.212 mmol, 1 equiv) in THF (3.6 mb) was added l-{3-[(tert-butyldimethylsilyl)oxy]cyclobutyl}- methanamine (913.81 mg, 4.242 mmol, 3.5 equiv) at room temperature under Ar. The resulting mixture was stirred at room temperature for 1 h. The residue was purified by silica gel column chromatography using 10% - 40% gradient of ethyl acetate in petroleum ether as eluent to afford (2-bromo-4-{[({3-[(tert- butyldimethylsilyl)oxy] cyclobutyl }methyl)amino]methyl } -7 -fluoro-6-methylquinolin-3 -yl)methanol (59) (200 mg, 33.17% yield, 95% purity) as a light yellow oil.

[0402] MS: m / z = 497.25, 499.30 [M + H]+.

[0403] ’H NMR (300 MHz, DMSO-J6) δ 8.25 (d, J= 8.3 Hz, 1H), 7.66 (d, J= 10.6 Hz, 1H), 4.82 (s, 2H), 4.25 - 4.17 (m, 2H), 4.16 - 3.99 (m, 1H), 2.76 - 2.62 (m, 2H), 2.45 (s, 3H), 2.37 - 2.17 (m, 2H), 2.08 - 1.97 (m, 1H), 1.96 - 1.78 (m, 1H), 1.58 - 1.43 (m, 2H), 1.29 - 1.13 (m, 1H), 0.84 (s, 9H), 0.00 (s, 6H).19F NMR (282 MHz, DMSO- d6) 5 -112.49

[0404] Example 59 - Compound 60

[0405] To a solution of (2-bromo-4-{[({3-[(tert-butyldimethylsilyl)oxy ]cyclobutyl }methyl)amino]methyl}-7- fluoro-6-methylquinolin-3-yl)methanol (59) (240 mg, 0.482 mmol, 1 equiv), (5 / ?)-5-cthyl-5-hydroxy- 177, 477, 877-oxepino[3,4-c]pyridine-3, 9-dione (10) (107.68 mg, 0.482 mmol, 1 equiv) and PPhs (202.44 mg, 0.771 mmol, 1.6 equiv) in DMF (5 mL) was added DIAD (146.32 mg, 0.723 mmol, 1.5 equiv) at 0 °C under Ar. The resulting mixture was heated to room temperature for 1 h. The reaction mixture quenched with water (50 mL) and extracted with ethyl acetate (50 mL x 3). The combined organic layers were dried over sodium sulfate and fdtered. The fdtrate was concentrated under reduced pressure. The residue was purified by silica gel column chromatography, eluted with 0 - 50% THF in petroleum ether to afford (57?)- 8-[(2-bromo-4-{[({3-[( / ert-butyldimethylsilyl)oxy]cyclobutyl}methyl)amino]methyl}-7-fluoro-6- methylquinolin-3-yl)methyl] -5 -ethyl-5 -hydroxy- 177, 477-oxepino[3,4-c]pyridine-3, 9-dione (60) (100 mg, 29.50% yield, 95% purity) as an off-white solid.

[0406] MS: m / z = 702.20, 704.20 [M + H]+.

[0407] ’H NMR (300 MHz, CDC13) δ 8.30 - 7.98 (m, 1H), 7.95 - 7.72 (m, 1H), 7.62 - 7.49 (m, 1H), 7.36 - 7.10 (m, 1H), 6.65 - 6.30 (m, 1H), 5.80 - 5.56 (m, 1H), 5.58 - 5.29 (m, 1H), 5.27 - 5.07 (m, 1H), 5.05 - 4.86 (m, 2H), 4.59 - 4.20 (m, 3H), 4.20 - 3.96 (m, 1H), 3.46 - 3.30 (m, 1H), 3.16 (s, 2H), 3.09 - 2.93 (m, 1H), 2.91 - 2.67 (m, 1H), 2.54 - 2.42 (m, 2H), 2.42 - 2.16 (m, 1H), 2.04 - 1.76 (m, 3H), 1.40 - 1.10 (m, 14H), 0.99 - 0.80 (m, 3H), 0.07 - -0.07 (m, 3H).19F NMR (282 MHz, CDC13) δ -109.27.

[0408] Example 60 - Compound 61

[0409] To a solution of (57?)-8-[(2-bromo-4-{[({3-[( / ert-butyldimethylsilyl)oxy]cyclobutyl}methyl)amino]- methyl}-7-fluoro-6-methylquinolin-3-yl)methyl]-5-ethyl-5-hydroxy-177,477-oxepino[3,4-c]pyridine-3,9- dione (60) (130 mg, 0.185 mmol, 1 equiv), PPhs (24.10 mg, 0.092 mmol, 0.5 equiv), KOAc (27.23 mg, 0.277 mmol, 1.5 equiv) in MeCN (6 mL) was added Pd(OAc)2 (4.15 mg, 0.018 mmol, 0.1 equiv) under nitrogen atmosphere. The mixture was stirred at 80 °C for 1 h. The reaction mixture concentrated under reduced pressure. The residue was purified by preparative TLC (DCM : MeOH = 10 : 1) to afford (20 / ?)- 10-{[({3-[( / ert-butyldimethylsilyl)oxy]cyclobutyl}methyl)amino]-methyl}-20-ethyl-6-fluoro-20-hydroxy- 7 -methyl- 17-oxa-3, 13-diazapentacyclo-[l 1.9.0.0A{2, 11} ,0A{4,9} ,0A{ 15,21 }]docosa- l(22),2,4,6,8,10,15(21)-heptaene-14,18-dione (61) (30 mg, 26% yield, 95% purity) as an off-white solid. MS: m!z = 622.50 [M + H]+.

[0410] Example 61 - Compound 62

[0411] To a solution of (207?)-10-{[({3-[(tert-butyldimethylsilyl)oxy]cyclobutyl}methyl)amino]methyl}-20- ethyl-6-fluoro-20-hydroxy-7-methyl-17-oxa-3,13-diazapentacyclo- [11.9.0.0A{2,l l}.0A{4,9}.0A{ 15,21}]docosa-l(22),2,4,6,8,10,15(21)-heptaene-14,18-dione (61) (30 mg, 0.048 mmol, 1 equiv) in DMSO (0.3 mb) was added triethylamine trihydrofluoride (0.3 mb) at 25 °C under nitrogen atmosphere. The resulting mixture was heated to 65 °C for 30 min. The mixture was purified by reversed-phase flash chromatography with the following conditions: column, C18silica gel; mobile phase, MeCN in water (0.05% HC1), 2% to 100% gradient in 30 min; detector, UV 254 nm & 210 nm to afford (207?)-20-ethyl-6-fluoro-20-hydroxy-10-({[(3-hydroxycyclobutyl)methyl]amino}methyl)-7- methyl- 17 -oxa-3 , 13 -diazapentacyclo- [ 11.9.0.0A{ 2, 11 } .0A{ 4, 9 } .0A{ 15 ,21 } ] docosa- l(22),2,4,6,8,10,15(21)-heptaene-14,18-dione (62, as its hydrochloride) (8.4 mg, 32.00% yield, 99.0% purity) as an orange solid.

[0412] MS: m / z = 508.25 [M + H]+.

[0413] 'HNMR (300 MHz, DMSO- d6) δ 9.43 (s, 2H), 8.45 (d, J= 8.0 Hz, 1H), 7.95 (d, J= 10.6 Hz, 1H), 7.40 (s, 1H), 5.76 - 5.30 (m, 4H), 4.85 - 4.54 (m, 3H), 4.07 - 3.91 (m, 3H), 3.48 (d, J= 13.8 Hz, 1H), 3.33 - 3.18 (m, 2H), 3.14 - 3.00 (m, 1H), 2.46 - 2.31 (m, 4H), 2.26 - 2.10 (m, 1H), 1.86 (d, J= 7.8 Hz, 2H), 1.71 - 1.57 (m, 2H), 0.87 (t, J= 7.3 Hz, 3H).19F NMR (282 MHz, DMSO-d6) δ -112.26.

[0414] Scheme 16

[0415] The synthesis of Compound 66 is described in Scheme 16 and Examples 62-65.

[0416] Scheme 16

[0417] Example 62 - Compound 63

[0418] To a solution of [2-bromo-4-(bromomethyl)-7-fluoro-6-methylquinolin-3-yl]methanol (35) (500 mg, 1.377 mmol, 1 equiv) in THF (5 mb) was added 4-[(tert-butyldimethylsilyl)oxy]piperidine (890.07 mg, 4.131 mmol, 3 equiv) at room temperature under Ar. The resulting mixture was stirred at room temperature for 1 h. The mixture was purified by silica gel column chromatography using 0% - 50% gradient of ethyl acetate in petroleum ether as eluent to afford [2-bromo-4-({4-[(tert- butyldimethylsilyl)oxy]piperidin-l-yl}methyl)-7-fluoro-6-methylquinolin-3-yl]methanol (63) (588.4 mg, 85.87% yield, 95% purity) as a white solid.

[0419] MS: m / z = 497.30, 499.30 [M + H]+.

[0420] Example 63 - Compound 64

[0421] To a solution of PPhs (476.58 mg, 1.818 mmol, 1.6 equiv) in THF (3 mb) was added DIAD (344.45 mg, 1.704 mmol, 1.5 equiv) at 0 °C under Ar. Then a solution of (57?)-5-ethyl-5-hydroxy-lH,4H,8H- oxepino[3,4-c]pyridine-3, 9-dione (10) (253.50 mg, 1.136 mmol, 1 equiv) in THF (2 mb) and a solution of [2-bromo-4-({4-[(tert-butyldimethylsilyl)oxy]piperidin-l-yl}methyl)-7-fluoro-6-methylquinolin-3- yl]methanol (63) (565 mg, 1.136 mmol, 1 equiv) in THF (3 mb) were added to above reaction mixture at 0 °C under Ar. The resulting mixture was stirred at room temperature for 1 h. The resulting mixture was concentrated under reduced pressure. The residue was purified by silica gel column chromatography using 10% - 50% gradient of ethyl acetate in petroleum ether as eluent to afford (57?)-8-{[2-bromo-4-({4-[(tert- butyldimethylsilyl)oxy]piperidin- 1 -yl }methyl)-7 -fluoro-6-methylquinolin-3 -yl]methyl } -5 -ethyl-5 - hydroxy- lH,4H-oxepino[3,4-c]pyridine-3, 9-dione (64) (577 mg, 72.30% yield, 95% purity) as a white solid.

[0422] MS: m / z = 702.40, 704.40 [M + H]+.

[0423] 'HNMR (300 MHz, CDC13) δ 8.07 (d, J= 7.9 Hz, 1H), 7.66 (d, J= 10.1 Hz, 1H), 6.90 (d, J= 7.4 Hz, 1H), 6.34 (d, J= 7.5 Hz, 1H), 5.65 - 5.35 (m, 3H), 3.98 - 3.88 (m, 2H), 3.63 - 3.49 (m, 1H), 3.30 (d, J = 13.4 Hz, 1H), 3.11 (d, J= 13.4 Hz, 1H), 3.02 - 2.92 (m, 1H), 2.76 - 2.64 (m, 1H), 2.49 (s, 3H), 2.45 - 2.34 (m, 1H), 2.27 - 2.13 (m, 1H), 2.10 - 1.98 (m, 1H), 1.95 - 1.77 (m, 2H), 1.55 - 1.46 (m, 1H), 1.22 - 1.05 (m, 2H), 1.00 - 0.92 (m, 4H), 0.86 (s, 9H), 0.02 (s, 6H).19F NMR (282 MHz, CDC13) 5 -109.63.

[0424] Example 64 - Compound 65

[0425] To a solution of (5R)-8-{[2-bromo-4-({4-[(tert-butyldimethylsilyl)oxy]piperidin-l-yl}methyl)-7-fluoro-6- methylquinolin-3 -yl]methyl } -5 -ethyl-5 -hydroxy- 1 H,4H-oxepino [3 ,4-c]pyridine-3 ,9-dione (64)(485 mg, 0.690 mmol, 1 equiv) in MeCN (50 mb) were added Pd(OAc)2 (30.99 mg, 0.138 mmol, 0.2 equiv), KO Ac (101.60 mg, 1.035 mmol, 1.5 equiv) and PPhs (108.61 mg, 0.414 mmol, 0.6 equiv) at room temperature under Ar. The resulting mixture was stirred at 80 °C for 6 h. The resulting mixture was concentrated under reduced pressure. The residue was purified by silica gel column chromatography, eluted with 0 - 20% THF in DCM to afford (20R)-10-({4-[(tert-butyldimethylsilyl)oxy]piperidin-l-yl}methyl)-20-ethyl- 6-fluoro-20-hydroxy-7-methyl-17-oxa-3,13-diazapentacyclo[11.9.0.0A{2,l l}.0A{4,9}.0A{15,21}]docosa- l(22),2,4,6,8,10,15(21)-heptaene-14,18-dione (65) (213.5 mg, 49.75% yield, 95% purity) as a light yellow solid.

[0426] MS: m / z = 622.15 [M + H]+.

[0427] ’HNMR (300 MHz, DMSO-J6) δ 8.41 - 8.32 (m, 1H), 7.82 (d, J= 10.8 Hz, 1H), 7.36 (s, 1H), 6.02 (s, 1H), 5.58 - 5.35 (m, 2H), 5.31 (s, 2H), 4.07 (s, 2H), 3.85 - 3.69 (m, 1H), 3.48 (d, J= 13.7 Hz, 1H), 3.06 (d, J= 13.9 Hz, 1H), 2.79 - 2.67 (m, 2H), 2.49 (s, 3H), 2.40 - 2.26 (m, 2H), 1.91 - 1.80 (m, 2H), 1.78 - 1.65 (m, 2H), 1.48 (s, 1H), 0.90 - 0.83 (m, 12H), 0.06 (s, 6H).19F NMR (282 MHz, DMSO-J6) 5 -113.06.

[0428] Example 65 - Compound 66

[0429] To a vial charged with (20R)-10-({4-[(tert-butyldimethylsilyl)oxy]piperidin-l-yl}methyl)-20-ethyl-6- fluoro-20-hydroxy-7-methyl-17-oxa-3,13-diazapentacyclo-[11.9.0.0A{2,l l}.0A{4,9}.0A{15,21}]docosa- l(22),2,4,6,8,10,15(21)-heptaene-14,18-dione (65) (101 mg, 0.162 mmol, 1 equiv) was added HC1 (4 M in 1,4-dioxane, 1.5 mb) at room temperature under Ar. The resulting mixture was stirred at 25 °C for 1 h. The resulting mixture was concentrated under reduced pressure. The residue was purified by reversed- phase flash chromatography with the following conditions: column, C18silica gel; mobile phase, MeCN in Water (0.05% HC1), 0% to 100% gradient in 35 min; detector, UV 254 nm & 210 nm to afford (20R)-20- ethyl-6-fluoro-20-hydroxy- 10- [(4-hydroxypiperidin- 1 -yl)methyl] -7-methyl- 17 -oxa-3 , 13 -diazapentacyclo- [11.9.0.0A{2,l l}.0A{4,9}.0A{15,21}]docosa-l(22),2,4,6,8,10,15(21)-heptaene-14,18-dione (66, as its hydrochloride) (42.6 mg, 48.21% yield, 99.1% purity) as an orange solid. MS: m / z = 508.10 [M + H]+.

[0430] ’H NMR (300 MHz, DMSO-J6) δ 10.53 - 10.27 (m, 1H), 8.61 (t, J= 7.9 Hz, 1H), 7.94 (d, J= 10.6 Hz, 1H), 7.41 (s, 1H), 5.71 - 5.47 (m, 3H), 5.46 - 5.37 (m, 1H), 5.09 - 4.96 (m, 2H), 4.96 - 4.83 (m, 2H), 3.93 (s, 1H), 3.64 - 3.43 (m, 3H), 3.40 - 3.25 (m, 2H), 3.14 - 3.00 (m, 1H), 2.57 (s, 3H), 2.09 (t, J= 13.1 Hz, 1H), 1.98 - 1.63 (m, 5H), 0.86 (t, J= 7.3 Hz, 3H).19F NMR (282 MHz, DMSO-J6) δ -112.45.

[0431] Example 66 - Additional Potency Data

[0432] Following the protocol of Example 35, biological activity data was obtained for the following compounds.

[0433] Example 67 PAMPA Permeability Data The bystander effect of exatecan-based ADCs refers to the ability of the released payload to diffuse into neighboring tumor cells that may not express the target antigen and kill them. This effect enhances the overall antitumor efficacy of the ADC, especially in tumors with heterogeneous antigen expression. This way, tumor cells that express other antigens or are low expressors of the target antigen can also be killed.

[0434] The key characteristics of an exatecan-based ADC having efficacious bystander effect are: 1 . Membrane permeable payload, meaning that once the payload is released inside the targeted tumor cell (after internalization and cleavage of the linker), it can diffuse out of the targeted cell and affect nearby cells. Exatecan and its analogues are cell-permeable molecules.

[0435] 2. Potent topoisomerase I inhibition. Exatecan interferes with DNA replication and transcription, leading to double strand breaks and consequent cell death - even in neighboring cells that didn’t bind the ADC directly (because they did not express the antigen of the antibody on the ADC).

[0436] 3. Cleavable linker, such as a peptide linker. Such linkers are designed to be cleaved by lysosomal enzymes or tumor-specific proteases, releasing the free payload into the cytoplasm and then to the tumor microenvironment, where it can contact neighboring cells.

[0437] The parallel artificial membrane permeability assay (PAMPA) is a measure of cell permeability of a compound. PAMPA values greater or equal to 1 are considered good, while PAMPA values between 1 and zero are considered moderate.

[0438] PAMPA values were measured as follows:

[0439] A stock solution of compound to serve as positive controls (testosterone, methotrexate, BN 80927, and exatecan) and test compounds was prepared in DMSO at a concentration of 10 mM. The solution was further diluted with PBS (pH 7.4), to afford a final concentration of 10 pM. These stock solutions (after dilution) were tested for PAMPA values as follows: a) A 1.8 % solution (w / v) of lecithin in dodecane was prepared and sonicated to ensure complete dissolution. b) A 5 pL aliquot of the lecitihin / dodecane mixture was carefully pipetted into each acceptor plate well (top compartment), avoiding pipette tip contact with the membrane. c) Immediately after the application to the artificial membrane (within 10 minutes) 300 pL of PBS (pH 7.4) solution was added to each well of the acceptor plate. Then 300 pL of drug (or control) containing solution was added to each well of the donor plate (bottom compartment) in triplicate. d) Slowly and carefully the acceptor plate was placed onto the donor plate, making sure that the underside of the membrane is in contact with the drug (or control) containing solutions in all wells. e) Replace the plate lid and incubate at 25 °C for 16 hr. f) After incubation, 50 pL aliquots from each well of the acceptor and donor plate are transferred into a 96-well plate. 200 pL of methanol containing 100 nM alprazolam, 200 nM labetalol, 200 nM caffeine, and 200 nM diclofenac was added to each well. g) The plate lid was covered. Vortex at 750 rpm for 100 seconds. Samples were centrifuged at 3,220 g for 20 minutes. The compound concentrations were determine by LC-MS / MS.

[0440] The effective permeability (Pe), in units of centimeter / second, can be calculated using the following equation: where

[0441] C = (VDx VA) / [(VD + VA) x t x A] where

[0442] VD = volume of donor compartment (0.30 mL)

[0443] VA = volume of acceptor compartment (0.30 mL)

[0444] A = fdter area (0.24 cm2for multi-screen permeability filter plate) t = incubation time

[0445] The PAMPA permeability data for compounds of this invention are provided in Table III:

[0446]

[0447] * After 16 hr incubation, the signal responses of these test compounds on the apical side were below the limit of detection. So 50 was used as the peak area value of the analyte in the calculation.

[0448] Compounds of this invention preferably have a PAMPA permeability value of between 3.0 and 0. Example 68 - Human Liver Microsome Data

[0449] In the event that there is premature cleavage of the ADC, leading to release of the payload away from the tumor microenvironment, it is desirable that it be cleared away quickly, to avoid systemic toxicity. An assay using human liver microsomes is used to measure the clearance rate of compounds, with a lower ti / 2 being indicative of faster clearance. The human microsome assay was performed as follows: 1. A master solution was prepared as follows:

[0450] Reagent Stock concentration Volume Final Concentration

[0451] Phosphate buffer 114.94 mM 69.6 pL 111.11 mM

[0452] Ultra-pure water 0.4 pL

[0453] Human liver microsomes 20 mg / mL 2 pL 0.556 mg / mL

[0454] 2. A stock solution of 10 mM NADPH was prepared in ultra-pure water.

[0455] 3. 80 nL aliquots of test compounds (or control, verapamil or exatecan) dissolved in DMSO were added to 72 pL of master solution.

[0456] 4. Reaction was started by the addition of 8 pL of the 10 mM NADPH solution. The final concentration of the NADPH was 1 mM. The negative control samples were prepared by replacing the NADPH solution with 8 pL of ultra-pure water. (The negative control was used to exclude the otherwise misleading factor that resulted from the instability of the test compound itself.) Samples with the NADPH were prepared in duplicate, while single samples of the negative control were prepared. The final concentration of the test compounds (or control) was 1 pM and the DMSO concentration in the incubation system was 0.1%. The mixture was pre-warmed at 37 °C for 10 minutes.

[0457] 5. Four 10 pL aliquots (40 pL in total) were taken from the reaction solution into one well of a new plate containing 120 pL of cold acetonitrile with internal standards (IS, 200 nM labetalol, 100 nM tolbutamide, and 100 nM ketoprofen). The same operation was done after 15, 30, 45, and 60 min incubation at 37 °C. Samples were centrifuged at 3,220 g for 45 min. 40 pL aliquots of the supernatant were mixed with 40 pL of ultra-pure water and then used for LC-MS / MS analysis.

[0458] Using Microsoft Excel, peak areas were determined from the extracted ion chromatograms. The slope value, k, was determined by linear regression of the natural logarithm of the remaining percentage of parent drug versus incubation time curve. The in vitro half-life (in vitro ti / 2) was determined from the slope value: in vitro ti / 2 = - (0.693 / k)

[0459] Conversion of the in vitro ti / 2 (min) into the in vitro intrinsic clearance (in vitro CLmt, in pL / min / mg protein) calculated from the following equation (mean of duplicate determinations):

[0460] 0.683 volume of incubation (pt) amount of proteins (mg)

[0461] The results are presented in Table IV :

[0462] Preferably, compounds of this invention have a ti / 2 of less than or equal to 100 min, more preferably less than or equal to 30 min. Alternatively, compounds of this invention have an in vitro CLmt less than or equal to 50 pL / min / mg protein, more preferably less than or equal to 20 pL / min / mg protein. The foregoing detailed description of the invention includes passages that are chiefly or exclusively concerned with particular parts or aspects of the invention. It is to be understood that this is for clarity and convenience, that a particular feature may be relevant in more than just the passage in which it is disclosed, and that the disclosure herein includes all the appropriate combinations of information found in the different passages. Similarly, although the various figures and descriptions herein relate to specific embodiments of the invention, it is to be understood that where a specific feature is disclosed in the context of a particular figure or embodiment, such feature can also be used, to the extent appropriate, in the context of another figure or embodiment, in combination with another feature, or in the invention in general. Further, while the present invention has been particularly described in terms of certain preferred embodiments, the invention is not limited to such preferred embodiments. Rather, the scope of the invention is defined by the appended claims.

Claims

CLAIMSWhat is claimed is:

1. A compound according to formula (I)or a pharmaceutically acceptable salt thereof, whereinR2is H, Cl, F, (CH^H, ©(CH^^H, or SCCH^H;R3is H, Cl, F, (CH^H, ©(CH^^H, or SCCH^H; or R2and R3in combination formeach X is independently O or NH; andY is CH orN; where the open valence of R1that is bonded to -XH is denoted by an * and the other end of R1that is bonded to the aromatic core of the compound of formula (I) is denoted by a wiggly line ( ).

2. The compound or pharmaceutically acceptable salt of claim 1, having a structure according to formula (F)R2is H, Cl, F, (CH2)1-3H, O(CH2)!.3H, or S(CH2)!.3H;R3is H, Cl, F, (CH^H, O(CH2)!.3H, or SCCH^H; or R2and R3in combination formeach X is independently O or NH;Y is CH orN; where the open valence of R1that is bonded to -NH, is denoted by an * and the other end of R1that is bonded to the aromatic core of the compound of formula (F) is denoted by a wiggly line ().The compound or pharmaceutically acceptable salt of claim 1, having a structure according to formula (I”)R2is H, Cl, F, (CH^H, ©(CH^^H, or SCCH^^H;R3is H, Cl, F, (CH^H, ©(CH^^H, or SCCH^^H; or R2and R3in combination formeach X is independently O or NH;Y is CH orN; where the open valence of R1that is bonded to -NH2 is denoted by an * and the other end of R1that is bonded to the aromatic core of the compound of formula (F) is denoted by a wiggly line ( ); with the provision thatis excluded.

4. The compound or pharmaceutically acceptable salt of any of the previous claims with the provisos that: when R1is absent, R2is methyl and R3is fluoro, X is not O or NH; and when X is O, R2is methyl and R3is fluoro, R1is not5. The compound or pharmaceutically acceptable salt of any of the previous claims with the proviso that the compound is not6. The compound or pharmaceutically acceptable salt of any of the previous claims, wherein R2is Me and R3is Cl or F.

7. The compound of claim 1, which is selected from the group consisting of:and pharmaceutically acceptable salts thereof.. The compound of claim 7, which is selected from the group consisting ofand pharmaceutically acceptable salts thereof.

9. The compound of claim 1, which is selected from the group consisting ofand pharmaceutically acceptable salts thereof.

10. A compound of the formula:or a pharmaceutically acceptable salt thereof.

11. The compound of any of claims 1-9, which compound has a parallel artificial membrane permeability assay (PAMPA) permeability value of less than or equal to 3.0 (e.g., less than 2.0, 1.0, 0.5, 0.4, 0.3, 0.2, or 0.1).

12. The compound of any of claims 1-9 or 11, which compound has a human liver microsome clearance ti / 2 of less than or equal to 100 minutes (e.g., less than 75, 50, or 30 minutes).

13. The compound of any of claims 1-9, 11 or 12, which compound has an in vitro CLmt less than or equal to 50 pL / min / mg protein (e.g., less than or equal to 30, 20, or 10 minutes).

14. A drug -linker compound having a structure according to formula (II):whereinR2is H, Cl, F, (CH^H, ©(CH^^H, or SCCH^H;R3is H, Cl, F, (CH^H, ©(CH^^H, or SCCH^H; or R2and R3in combination formeach X is independently O or NH;Y is CH orN; where the open valence of R1that is bonded to XLRbis denoted by an * and the other end of R1that is bonded to the aromatic core of the compound of formula (I) is denoted by a wiggly line ( •~v'' );L is a linker group; andRbis -NH2, -OH, -CO2H, -SH, maleimido, cyclooctyne, azido (-N3), hydroxylamino (-ONH2) orN -hydroxy succinimido .

15. The drug-linker compound according to claim 14, which is of formula (II’):whereinT is a self-immolating group; t is 0, 1, or 2;Rcis the side chain of an amino acid selected from the group consisting of alanine, arginine, asparagine, aspartic acid, citrulline, cysteine, glutamic acid, glutamine, glycine, histidine, isoleucine, leucine, lysine, methionine, norleucine, norvaline, ornithine, phenyalanine, proline, serine, threonine, tryptophan, tyrosine, and valine; p is 0, 1, 2, 3, or 4; q is 1, 2, 3, 4, 5, 6, 7, 8, 9, 10, 11, 12, 13, 14, 15, 16, 17, or 18; s is 0 or 1 ; and r is 1, 2, 3, 4, 5, or 6.

16. The drug-linker compound according to claim 14, which is of formula (III):

18. The drug-linker compound according to claim 14, which is of formula (IV):

19. The drug-linker compound according to claim 14, which is of formula (II”):whereinT is a self-immolating group; t is 0, 1, or 2;Rcis the side chain of an amino acid selected from the group consisting of alanine, arginine, asparagine, aspartic acid, citrulline, cysteine, glutamic acid, glutamine, glycine, histidine, isoleucine, leucine, lysine, methionine, norleucine, norvaline, ornithine, phenyalanine, proline, serine, threonine, tryptophan, tyrosine, and valine; p is 0, 1, 2, 3, or 4; and q is 1, 2, 3, 4, 5, 6, 7, 8, 9, 10, 11, 12, 13, 14, 15, 16, 17, or 18.

20. An antibody-drug conjugate having a structure according to formula (V):whereinR2is H, Cl, F, (CH^H, ©(CH^^H, SCCH^^H;R3is H, Cl, F, (CH^H, ©(CH^^H, SCCH^^H; or R2and R3in combination formeach X is independently O or NH;Y is CH orN; where the open valence of R1that is bonded to XL’ Ab is denoted by an * and the other end of R1that is bonded to the aromatic core of the compound of formula (I) is denoted by a wiggly line ( );L’ is a linker;Ab is an antibody; and m is 1, 2, 3, 4, 5, 6, 7 or 8.whereinT is a self-immolating group; t is 0, 1, or 2;Rcis the side chain of an amino acid selected from the group consisting of alanine, arginine, asparagine, aspartic acid, citrulline, cysteine, glutamic acid, glutamine, glycine, histidine, isoleucine, leucine, lysine, methionine, norleucine, norvaline, ornithine, phenyalanine, proline, serine, threonine, tryptophan, tyrosine, and valine; p is 0, 1, 2, 3, or 4; q is 1, 2, 3, 4, 5, 6, 7, 8, 9, 10, 11, 12, 13, 14, 15, 16, 17, or 18; s is 0 or 1; and r is 1, 2, 3, 4, 5, or 6.

23. The antibody-drug conjugate according to claim 20, which is of formula (VI’):whereinT is a self-immolating group; t is 0, 1, or 2;Rcis the side chain of an amino acid selected from the group consisting of alanine, arginine, asparagine, aspartic acid, citrulline, cysteine, glutamic acid, glutamine, glycine, histidine, isoleucine, leucine, lysine, methionine, norleucine, norvaline, ornithine, phenyalanine, proline, serine, threonine, tryptophan, tyrosine, and valine; p is 0, 1, 2, 3, or 4; and q is 1, 2, 3, 4, 5, 6, 7, 8, 9, 10, 11, 12, 13, 14, 15, 16, 17, or 18.

25. The antibody-drug conjugate according to claim 20, which is of formula (VII):

26. A drug -linker compound of formula (Ila):R5is H, Cl, F, (CH^H, O(CH2)1.3H, or S(CH2)1.3H;R6is H, Cl, F, (CH^H, OWA or SCCH^H;Rbis -NH2, -OH, -CO2H, -SH, maleimido, cyclooctyne, azido (-N3), hydroxylamino (-ONH2) or N-hydroxysuccinimido; andL is a linker.

27. An antibody-drug conjugate of formula (Va):R5is H, Cl, F, (CH^H, OCCH^^H, or SCCH^H;R6is H, Cl, F, (CH^H, OCCH^^H, or SCCH^H;L’ is a linker,Ab is an antibody and m is 1, 2, 3, 4, 5, 6, 7, or 8.

28. The antibody-drug conjugate of any of claims 20-25 or 27, wherein the antibody Ab recognizes an antigen selected from the group consisting of GPC3, PTK7, DLL3, Claudin 6, Claudin 1, Claudin 18.2, ROR1, ROR2, AXL, PSMA, B7H3 (CD276), B7H4, B7H7, LIV-1, MUC 1, CSPG4, 5T4, Trop2, Nectin4, TF, HER3, CEACAM5, Cadherin 17, Cadherin 6, STn, ITGB6 (integrin beta 6), SEZ6, EGFR, mesothelin, CD70, CD74, CDCP1, FRalpha, cMet, GUCY2C, EFNA4, HER2, EphA5, Napi2B, 5T4, uPARAP, CD30, CD33, CD79b, CD22, CD19, CD46, CD56, BCMA, and GPRC5D.

28. A method of treating cancer, comprising administering to a patient in need thereof a therapeutically effective amount of an antibody-drug conjugate of any of claims 20-25 or 27.

30. A pharmaceutical formulation comprising an antibody-drug conjugate of any of claims 20-25 or27 and a pharmaceutically acceptable excipien

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