Compositions and methods for activating pyruvate kinase

Abstract

Provided herein are compositions and methods for activating pyruvate kinase (e.g., in a subject). In particular, provided herein are compositions and methods for treating a disease or disorder (e.g., an ocular disease, a hematological condition, or a cancer) using a pyruvate kinase activator.

Description

Technical Field

[0001] This article provides compositions and methods for activating pyruvate kinase (e.g., in a subject). In particular, this article provides compositions and methods for treating diseases or conditions (e.g., eye diseases, blood disorders, or cancer) using pyruvate kinase activators. Background Technology

[0002] Photoreceptor death is the ultimate cause of vision loss in many retinal diseases, including retinal detachment, retinal dystrophy, and age-related macular degeneration (AMD). AMD affects 17 million people in the United States, with a potential annual market size of $40 billion (Wong, W. Let et al. Lancet Glob. Health 2, e106-116 (2014)). Retinal dystrophy (including retinitis pigmentosa (RP)) affects 100,000 people in the United States, with a potential annual market size of $480 million. Retinal detachment affects 750,000 people globally, with a market size of $150 million (Haimann, MH, et al. Arch. Ophthalmol. Chic. Ill 1960 100, 289–292 (1982)). The patient experience of poor visual function and blindness leads to lifelong vision services, loss of productivity for patients and caregivers, and a decline in quality of life. In the United States, the annual supplemental medical costs due to vision impairment are valued at $5.5 billion, and the value of the loss of quality of life is valued at $10.5 billion annually (Frick, KD, et al. Arch. Ophthalmol. Chic. Ill 1960 125, 544–550 (2007)). No successful treatment options exist to prevent photoreceptor death in retinal diseases.

[0003] There is an urgent unmet need for neuroprotective mechanisms to improve photoreceptor survival and related disorders. Summary of the Invention

[0004] This article provides compositions and methods for activating pyruvate kinase (e.g., in a subject). In particular, this article provides compositions and methods for treating diseases or conditions (e.g., eye diseases, blood disorders, or cancer) using pyruvate kinase activators.

[0005] Metabolic reprogramming of photoreceptors is a treatment solution for vision loss associated with age-related macular degeneration, retinal dystrophy, retinal degeneration, diabetic retinopathy, and retinal detachment. N. et al. Cell 161, 817–832 (2015); Zhang, L. et al. J. Clin. Invest. 126, 4659–4673 (2016)). PKM2 (a key regulator of aerobic glycolysis and energy metabolism in photoreceptors) is reprogrammed to enhance metabolism and energy production through activation by small molecule activators (including ML-265) by promoting catabolism in cells (Anastasiou, D. et al. Nat. Chem. Biol. 8, 839–847 (2012; Wubben et al. Sci Rep. 2017 and Wubben et al. Sci Rep. 2020)). Limiting PKM2 expression and aerobic glycolysis of intraocular photoreceptors reduces any potential off-target effects after ocular delivery, increases treatment specificity, and expands the therapeutic window (Rajala, RVS, et al. Sci. Rep. 6, 37727 (2016); Lindsay, KJ et al. Proc. Natl. Acad. Sci. USA 111, 15579–15584 (2014)).

[0006] Therefore, in some embodiments, this document provides pyruvate kinase activators for, for example, treating diseases or conditions (e.g., eye disorders). For example, in some embodiments, a composition is provided comprising: compounds selected from the following

[0007]

[0008] X1, X2, X3, and X4 are independently selected from CH, CO, N, NH, S, or O;

[0009] Z represents a vacancy, bond, optional substitution of a C1-6 alkyl group, -O-, -S-, -CH2-, -CHR5-, -CR5R6-, or -(CH2). n -、-(CHR5) n -、-(CR5R6) n-, -S(=O)CH2, -S(=O)2CH2-, -NR5-, -NR5C(=O)-, -C(O)NR5-, -C(=O)-, -OC(=O)-, -C(=O)O -, -NR5C(=O)O-, -OC(=O)NR5-, -NR5C(=O)NR5-, -OC(R5)2-, -C(R5)2O-, -NR4C(R5)2-, C(R6 )2NR5-、-S(=O)-、S(=O)2-、-S(=O)2O-、-OS(=O)2-、-S(=O)2NR5-、-NR5S(=O)2-、-S(=O)NR5-、-NR5S(=O-、-OS(=O)NR5-、-NR5S(=O)O- or –S(=O)(=NR5)-, where the connection point with R1 or R2 is located on the left and n=1-6;

[0010] R5 and R6 are each independently hydrogen, halogen, -CN, OR7, NR7R8, -N(R7)C(=O)R8, -C(=O)N(R7), -C(=O)R7, -C(=O)OR7, -SR7, -S(=O)R7, -S(=O)2R7 or any optimally substituted –C1-C6 alkyl;

[0011] R7 and R8 are each independently hydrogen, any optimally substituted –C1-C6 alkyl; or alternatively, R7 and R8 together are either an optionally substituted C1-C6 monocyclic cycloalkyl ring or an optionally substituted monocyclic heterocycle.

[0012] Y represents an empty space, -CH2-, -CHR9-, or -CR9R. 10 -、-(CH2) n -、-(CHR9) n -、-(CR9R 10 ) n -, -C(=O)-, -S(=O)-, -S(=O)2-, where n=1-6;

[0013] R9 and R 10 Each is independently hydrogen, halogen, -CN, or any optimally substituted –C1-C6 alkyl; or alternatively R9 and R 10 They can be used together as optional substituted C1-C6 monocyclic cycloalkyl rings or optional substituted monocyclic heterocycles;

[0014] R1 is -H, -F, -Cl, -Br, -NO2, -CN, -NH2, -NHR 11 -NR 11 R 12 -OH, -OR 11Optionally substituted C1-C6 alkyl, Optionally substituted monocyclic cycloalkyl, Optionally substituted bicyclic cycloalkyl, Optionally substituted tricyclic cycloalkyl, Optionally substituted monocyclic heterocycle, Optionally substituted bicyclic heterocycle or Optionally substituted tricyclic heterocycle;

[0015] R2 is -H, -F, -Cl, -Br, -NO2, -CN, -NO2, -NH2, -NHR 11 -NR 11 R 12 -OH, -OR 11 R2 is -H, -F, -Cl, -Br, -NO2, -CN, -NO2, -NH2, -NHR 11 -NR 11 R 12 -OH, -OR 11 Optionally substituted C1-C6 alkyl, Optionally substituted monocyclic cycloalkyl, Optionally substituted bicyclic cycloalkyl, Optionally substituted tricyclic cycloalkyl, Optionally substituted monocyclic heterocycle, Optionally substituted bicyclic heterocycle, Optionally substituted tricyclic heterocycle;

[0016] R3 is -H, -F, -Cl, -Br, -NO2, -CN, -NO2, -NH2, -NHR 11 -NR 11 R 12 -OH, -OR 11 Optionally substituted C1-C6 alkyl, Optionally substituted monocyclic cycloalkyl, Optionally substituted bicyclic cycloalkyl, Optionally substituted tricyclic cycloalkyl, Optionally substituted monocyclic heterocycle, Optionally substituted bicyclic heterocycle, Optionally substituted tricyclic heterocycle;

[0017] R 11 and R 12 Each of them independently is an optional substituted C1-C6 alkyl, an optional substituted monocyclic cycloalkyl, an optional substituted bicyclic cycloalkyl, an optional substituted tricyclic cycloalkyl, an optional substituted monocyclic heterocycle, an optional substituted bicyclic heterocycle, or an optional substituted tricyclic heterocycle;

[0018] R4 represents hydrogen, -CH3, and -CHR. 13 -CR 13 R 14 -S(=O)R 13 -S(=O)2R 13 Optionally substituted alkyl groups, preferably substituted haloalkyl groups, preferably substituted alkenyl groups, preferably substituted alkynyl groups, preferably substituted cycloalkyl groups, preferably substituted heterocyclic groups, preferably substituted aryl groups, -C(=O)R 15 Or nitrogen protecting group; wherein;

[0019] R 13 and R14 Each of these can be independently a optionally substituted C1-C6 alkyl, an optionally substituted monocyclic cycloalkyl, an optionally substituted bicyclic cycloalkyl, an optionally substituted tricyclic cycloalkyl, an optionally substituted monocyclic heterocycle, an optionally substituted bicyclic heterocycle, or an optionally substituted tricyclic heterocycle.

[0020] R 15 It is hydrogen, -CH3, optionally substituted alkyl, preferably substituted haloalkyl, preferably substituted alkenyl, preferably substituted alkynyl, preferably substituted cycloalkyl, preferably substituted heterocyclic, preferably substituted aryl, or a pharmaceutically acceptable salt thereof.

[0021] In some implementations, R2-Z- is H and R1-Z- is selected from;

[0022] Z represents an empty space, a bond, or an optional substitution C. 1-6 Alkyl, -O-, -S-, -CH2-, -CHR5-, -CR5R6-, -(CH2) n -、-(CHR5) n -、-(CR5R6) n -、-S(=O)CH2、-NR5-、-NR5C(=O)-、-C(O)NR5-、-C(=O)-、-S(=O)-、where the connection point with R1 is located on the left and n=1-6;

[0023] R5 and R6 are each independently hydrogen, -CN, OR7, NR7R8, or any optimally substituted –C1-C6 alkyl; or alternatively, R7 and R8 may together be an optionally substituted C1-C6 monocyclic cycloalkyl ring or an optionally substituted monocyclic heterocycle; wherein;

[0024] R7 and R8 are each independently hydrogen, any optimally substituted –C1-C6 alkyl; or alternatively, R7 and R8 may be together as an optionally substituted C1-C6 monocyclic cycloalkyl ring or an optionally substituted monocyclic heterocycle.

[0025] Y represents an empty space, -CH2-, -CHR9-, or -CR9R. 10 -、-(CH2) n -、-(CHR9) n -、-(CR9R 10 ) n -, where n = 1-3;

[0026] R9 and R 10 Each is independently hydrogen or any optimally substituted –C1-C6 alkyl; or alternatively R9 and R 10 They can be used together as optional substituted C1-C6 monocyclic cycloalkyl rings or optional substituted monocyclic heterocycles;

[0027] R1 is -H, -F, -Cl, -Br, -NO2, -CN, -NO2, -NH2, -NHR 11 -NR 11 R 12 -OH, -OR 11 Optionally substituted C1-C6 alkyl, Optionally substituted monocyclic cycloalkyl, Optionally substituted bicyclic cycloalkyl, Optionally substituted tricyclic cycloalkyl, Optionally substituted monocyclic heterocycle, Optionally substituted bicyclic heterocycle or Optionally substituted tricyclic heterocycle;

[0028] R2 is -H, -F, -Cl, -Br, -NO2, -CN, -NO2, -NH2, -NHR 11 -NR 11 R 12 -OH, -OR 11 Optionally substituted C1-C6 alkyl, Optionally substituted monocyclic cycloalkyl, Optionally substituted bicyclic cycloalkyl, Optionally substituted tricyclic cycloalkyl, Optionally substituted monocyclic heterocycle, Optionally substituted bicyclic heterocycle or Optionally substituted tricyclic heterocycle;

[0029] R3 is -H, -F, -Cl, -Br, -NO2, -CN, -NO2, -NH2, -NHR 11 -NR 11 R 12 -OH, -OR 11 Optionally substituted C1-C6 alkyl, Optionally substituted monocyclic cycloalkyl, Optionally substituted bicyclic cycloalkyl, Optionally substituted tricyclic cycloalkyl, Optionally substituted monocyclic heterocycle, Optionally substituted bicyclic heterocycle, Optionally substituted tricyclic heterocycle;

[0030] R 11 and R 12 Each of them independently is an optional substituted C1-C6 alkyl, an optional substituted monocyclic cycloalkyl, an optional substituted bicyclic cycloalkyl, an optional substituted tricyclic cycloalkyl, an optional substituted monocyclic heterocycle, an optional substituted bicyclic heterocycle, or an optional substituted tricyclic heterocycle;

[0031] R4 is –CH3.

[0032] In some exemplary embodiments, Z-R1, Z-R2, Z-R3, and Y-R3 are selected individually or in combination, for example, Where R 16 and R 17 It can be located at the ortho, meta, or para position of the aryl ring and is selected from H, -OCH3, C1-C4 alkyl, -NH2, -halogen, -CN, -OH, -S(=O)Me, -S(=O)2Me, -CH2OMe, -CH2NR 18 R 19 , where R18 and R 19 Selected from –H, C1-C4 alkyl, optimally substituted aryl or heterocyclic groups, or those forming carbocyclic or heterocyclic rings together.

[0033] In some embodiments, the compound is a compound of formula I, wherein Z-R2 is H, Z-R1 is not 8-OCH3, and R4 is not... And Y-R3 is not In some embodiments, the compound is a compound of formula I, wherein Z-R2 is H, Z-R1 is not 8-Cl, and R4 is not CH3 or And Y-R3 is not CH3 or In some embodiments, the compound is a compound of formula I and R2 is not. Or CH3, and Y-R3 is not In some embodiments, the compound is a compound of formula I and Z-R1 is not. X1 is not CO, R2 is not CH3, and Y-R3 is not In some embodiments, the compound is not Where R1 and R2 are independently CH3 or In some embodiments, the compound is selected from the compounds described in Table 2. In some embodiments, the compound is:

[0034] Wherein R1 is a substituted monocyclic or bicyclic carbocyclic or heterocyclic moiety and R2 is a substituted monocyclic or bicyclic heterocyclic moiety.

[0035] In some exemplary embodiments, the compound is

[0036]

[0037]

[0038]

[0039]

[0040]

[0041]

[0042] In some embodiments, the compound is

[0043]

[0044] in

[0045] X is an empty space or a selection from -H, -CH2-, -CHR3-, -CR3R4-, or -(CH2). n -、-(CHR3) n -、-(CR3R4) n -, where n–1–6;

[0046] R1 is selected from, for example, -H, -CN, -NO2, -NH2, -NHR3, NR3R4, -OH, OR3, -SOR3, -SO2R3, optionally substituted C1-C6 alkyl, optionally substituted monocyclic cycloalkyl, optionally substituted bicyclic cycloalkyl, optionally substituted tricyclic cycloalkyl, optionally substituted monocyclic heterocycle, optionally substituted bicyclic heterocycle or optionally substituted tricyclic heterocycle;

[0047] R2 is selected from, for example, -H, -CN, -NO2, -NH2, -NHR3, NR3R4, -OH, OR3, -SOR3, -SO2R3, optionally substituted C1-C6 alkyl, optionally substituted monocyclic cycloalkyl, optionally substituted bicyclic cycloalkyl, optionally substituted monocyclic heterocycle, optionally substituted bicyclic heterocycle or optionally substituted tricyclic heterocycle.

[0048] R3 and R4 are each independently selected from the group consisting of, for example, optionally substituted C1-C6 alkyl, optionally substituted monocyclic cycloalkyl, optionally substituted bicyclic cycloalkyl, optionally substituted tricyclic cycloalkyl, optionally substituted monocyclic heterocycle, optionally substituted bicyclic heterocycle or optionally substituted tricyclic heterocycle; and.

[0049] In some implementations, R1 is selected from, for example,

[0050] And R2 is selected from, for example,

[0051]

[0052] In another embodiment, the compound is

[0053]

[0054] R1 is selected from, for example, -H, -F, -Cl, -Br, -CN, -NO2, -NH2, -NHR3, -NR3R4, -OH, -OR3, optionally substituted C1-C6 alkyl, optionally substituted monocyclic cycloalkyl, optionally substituted bicyclic cycloalkyl, optionally substituted tricyclic cycloalkyl, optionally substituted monocyclic heterocycle, optionally substituted bicyclic heterocycle or optionally substituted tricyclic heterocycle;

[0055] R2 is selected from, for example, -H, -CH3, -(CH2). n -R5-、-(CHR3) n -R5-、-(CR3R4) n -R5-, optionally substituted C1-C6 alkyl, optionally substituted monocyclic cycloalkyl, optionally substituted bicyclic cycloalkyl, optionally substituted monocyclic heterocycle, optionally substituted bicyclic heterocycle or optionally substituted tricyclic heterocycle;

[0056] R3 and R4 are each independently selected from, for example, optionally substituted C1-C6 alkyl groups, optionally substituted monocyclic cycloalkyl groups, optionally substituted bicyclic cycloalkyl groups, optionally substituted tricyclic cycloalkyl groups, optionally substituted monocyclic heterocycles, optionally substituted bicyclic heterocycles, or optionally substituted tricyclic heterocycles; and

[0057] R5 is -H, -F, -Cl, -Br, -NO2, -CN, -NO2, -NH2, -NHR3, -NR3R4, -OH, -OR3, optionally substituted C1-C6 alkyl, optionally substituted monocyclic cycloalkyl, optionally substituted bicyclic cycloalkyl, optionally substituted tricyclic cycloalkyl, optionally substituted monocyclic heterocycle, optionally substituted bicyclic heterocycle, optionally substituted tricyclic heterocycle.

[0058] In some embodiments, R1 is selected from, for example, OCH3, OH, NH2, NCH3, N(CH3)2,

[0059]

[0060] And R2 is selected from, for example,

[0061]

[0062] In some embodiments, one or more hydrogen atoms in any of the above compounds are replaced with deuterium.

[0063] In some embodiments, the composition is a pharmaceutical composition. In some embodiments, the composition is formulated for injection, oral delivery, or as eye drops. In some embodiments, the composition includes a pharmaceutically acceptable carrier. In some embodiments, the composition is a pyruvate kinase (e.g., PKM1 or PKM2) activator.

[0064] In another embodiment, this document provides a method for activating pyruvate kinase in a subject (e.g., the subject's eye), the method comprising administering a compound described herein to the subject (e.g., the subject's eye), wherein the administration activates pyruvate kinase. In some embodiments, activation treats or alleviates symptoms of a disease or condition (e.g., an eye disorder, cancer, or a blood disorder) in the subject. Exemplary eye disorders include, but are not limited to, vision loss, retinal dystrophy, macular degeneration, retinal degeneration, diabetic retinopathy, retinal detachment, or proliferative vitreoretinopathy. Exemplary blood disorders include, but are not limited to, anemia, hemolytic anemia, sickle cell disease, thalassemia, hereditary spherocytosis, hereditary elliptic polycythemia, abetalipoproteinemia, or Bassen-Kornzweig syndrome.

[0065] Another implementation provides a method for treating a disease or condition, the method comprising: administering the pyruvate kinase activator described herein to a subject in need, wherein the administration treats or alleviates the symptoms of the subject's disease or condition.

[0066] Another embodiment provides a method for treating an eye disease, the method comprising:

[0067] The compound described herein is administered to the eye of a subject in need, wherein the administration treats or alleviates symptoms of an eye condition in the subject. In some embodiments, the administration prevents or reduces photoreceptor cell death in the subject's eye. In some embodiments, the administration prevents or reduces proliferative vitreoretinopathy in the subject's eye. In some embodiments, the activator is formulated for injection (e.g., intravitreal injection), oral delivery, or as eye drops.

[0068] Another embodiment provides a method for treating an eye disease, the method comprising:

[0069] Administering the drug to the eyes of subjects who require it, selected from The compound in which R 1 It is hydrogen, optionally substituted alkyl, optionally substituted haloalkyl, optionally substituted alkenyl, optionally substituted ynyl, optionally substituted cycloalkyl, optionally substituted heterocyclic, optionally substituted aryl, -OR o1 -C(=O)R c1 Or nitrogen protecting group; wherein: R 01 It is a hydrogen, optionally substituted alkyl, or oxygen protecting group; R c1 It is an substituted alkyl group or -N(R) cn )2, where R cn Each instance is independently hydrogen, -C 1-6Alkyl or nitrogen protecting groups;

[0070] R 2 Q and R are each independently a optionally substituted 5- or 6-membered monocyclic heteroaryl group; a and R b Each of these can be independently hydrogen, halogen, -CN, -NO2, -N3, optionally substituted alkyl, or -OR. o3 -N(R n1 )2、-C(=O)N(R n1 )2 or -C(=O)R c2 ; or alternatively R a and R b They can form optionally substituted cycloalkyl or optionally substituted heterocyclic groups together with the carbon atoms to which they are attached; wherein: R n1 Each instance is independently hydrogen, optionally substituted -C1-C6 alkyl, or nitrogen-protecting group; R o3 It is hydrogen, optionally substituted -C1-C6 alkyl or oxygen protecting group; and R c2 It is an optionally substituted -C1-C6 alkyl group; and R j and R k Each is independently hydrogen, halogen, -CN, -OR o7 -N(R) n5 )2、-N(R n5 )C(=O)R c5 -C(=O)N(R) n5 )2、-C(=O)R c5 -C(=0)0R o7 -SR js -S(=0)2R js -S(=O)R js Or optionally substituted -C1-C6 alkyl; or alternatively R j and R k They can form C=O, optionally substituted C1-C6 monocyclic cycloalkyl rings, or optionally substituted C3-C6 monocyclic heterocyclic rings together with the carbon atoms they are attached to; wherein: R n5 Each instance is independently hydrogen, optionally substituted -C1-C6 alkyl, -OR o8 Or nitrogen-protecting group, wherein R o8 It is hydrogen, optionally substituted -C1-C6 alkyl or oxygen protecting group; R o7 Each instance is independently hydrogen, optionally substituted -C1-C6 alkyl, or oxygen protecting group; R c5 Each instance is independently an optionally substituted -C1-C6 alkyl group; and R js Each instance is independently an optionally substituted -C1-C6 alkyl group, optionally substituted C 6-12aryl, optionally substituted heteroaryl or thioprotective groups, or including the formula Compounds wherein Q is hydrogen, an optionally substituted alkyl group, an optionally substituted alkenyl group, an optionally substituted alkynyl group, an optionally substituted cycloalkyl group, an optionally substituted heterocyclic group, an optionally substituted aryl group, or an optionally substituted heteroaryl group; R 1 It is hydrogen, optionally substituted alkyl, optionally substituted haloalkyl, optionally substituted alkenyl, optionally substituted ynyl, optionally substituted cycloalkyl, optionally substituted heterocyclic, optionally substituted aryl, -OR 01 -C(=0)R cl Or nitrogen-protected group; L 1 It is a bond, an optional substituted alkylene group -0-, -S-, -S-CH2--S(=0)CH2--S(=0)2CH2--NR 3 -、-NR 3 C(=0)-、-C(=0)NR 3 -, —C(=0)-, -OC(=0)-, -C(=0)0--NR 3 C(=0)0-、-OC(=0)NR 3 -、-NR 3 C(=0)NR 3 -、-OC(R)2-、-C(R)20-、-NR 3 C(R 4 )2-、-C(R 4 )2NR 3 -, -S(=0)2--S(=0)--S(=0)20--OS(=0)2--S(=0)0--OS(=0)-,-S(=0)2NR 3 -、-NR 3 S(=0)2--S(=0)NR 3 -、-NR 3 S(=0)-、-NR 3 S(=0)20-、-OS(=0)2NR 3 -、-NR 3 S(=0)0-、-OS(=0)NR 3 -or-S(=0)(=NR) 3 )-, where R 2 The connection point is located on the left-hand side; L 2 It is a bond, an optionally substituted alkylene group, -C(=0)-, -S(=0)2- or -S(=0)-, wherein the connection point with Q is on the right-hand side; R 2It is hydrogen, halogen, optionally substituted alkyl, optionally substituted alkoxy, optionally substituted alkenyl, optionally substituted alkynyl, optionally substituted cycloalkyl, optionally substituted heterocyclic, optionally substituted aryl or optionally substituted heteroaryl, or nitrogen-protecting group (when L 1 -NR 3 -、-NR 3 C(=0)-、-NR 3 C(=0)0--NR 3 C(R)2--NR 3 S(=0)2-、-NR 3 S(=0)-、-NR 3 C(=0)NR 3 -、-NR 3 S(=0)20-or-NR 3 When S(=0)0-), oxygen protecting group (when L 1 It is -0-, -OC(=0)-, -OC(=0)NR 3 -、-OC(R 4 )2--OS(=0)2-、-OS(=0)2NR 3 -、-OS(=0)NR 3 -or-OS(=0)-) or sulfur protecting group (when L 1 (is -S-time); R 3 Each instance is independently hydrogen, -OR o2 Optionally substituted alkyl, optionally substituted alkenyl, optionally substituted alkynyl, optionally substituted cycloalkyl, optionally substituted heterocyclic, optionally substituted aryl, optionally substituted heteroaryl or nitrogen-protecting group; R o1 and R o2 Each instance is independently a hydrogen, optionally substituted alkyl, or oxygen protecting group; R cl Each instance is independently an optionally substituted alkyl group or -N(R) cn )2, where R cn Each instance is independently a hydrogen, -Ci-6 alkyl, or nitrogen-protecting group; and R 4 Each instance is independently hydrogen, optionally substituted alkyl, optionally substituted alkenyl, optionally substituted alkynyl, optionally substituted cycloalkyl, optionally substituted heterocyclic, optionally substituted aryl, or optionally substituted heteroaryl; provided that when Li and L2 are optionally substituted methylene, Q and R 2 Not all are optionally substituted 5- or 6-membered monocyclic heteroaryl groups. This drug is used to treat or alleviate symptoms of ocular diseases in the subjects.

[0071] This document describes alternative implementation methods.

[0072] definition

[0073] To facilitate understanding of this disclosure, several terms and phrases are defined below:

[0074] As used herein, the term "aliphatic" refers to groups including, but not limited to, alkyl, alkenyl, alkynyl, and alicyclic groups.

[0075] The term "halogen" or "halogen" refers to any group consisting of fluorine, chlorine, bromine, or iodine.

[0076] As used herein, the term "alkyl" refers to an unsaturated carbon chain substituent group. Typically, alkyl groups have the general formula C1. n H 2n+1 Exemplary alkyl groups include, but are not limited to, methyl (CH3), ethyl (C2H5), propyl (C3H7), butyl (C4H9), and pentyl (C5H5). 11 )wait.

[0077] The term "alkenyl" refers to a monovalent straight-chain or branched hydrocarbon chain containing 2-12 carbon atoms and having one or more double bonds. Examples of alkenyl groups include, but are not limited to, allyl, propenyl, 2-butenyl, 3-hexenyl, and 3-octenyl groups. Optionally, one of the carbon atoms in the double bond may be the linking point of the alkenyl substituent. In some respects, the term "alkenyl" refers to a monovalent straight-chain or branched hydrocarbon chain containing 2-6 carbon atoms and having one or more double bonds. In other respects, the term "alkenyl" refers to a monovalent straight-chain or branched hydrocarbon chain containing 2-4 carbon atoms and having one or more double bonds.

[0078] The term "alkynyl" refers to a monovalent straight-chain or branched hydrocarbon chain containing 2-12 carbon atoms and characterized by having one or more triple bonds. Examples of alkynyl groups include, but are not limited to, ethynyl, propynyl, and 3-hexynyl. Optionally, one of the carbon atoms in the triple bond may be the linking point of the alkynyl substituent.

[0079] As used herein, the term "aryl" refers to a single aromatic ring (such as a benzene ring), or two or more aromatic rings (e.g., biphenyl, naphthalene, anthracene), or an aromatic ring and one or more non-aromatic rings. The aryl group may optionally be substituted with a lower aliphatic group (e.g., alkyl, alkenyl, alkynyl, or alicyclic). Furthermore, the aliphatic and aryl groups may be further substituted with one or more functional groups (including, but not limited to, chemical moieties comprising N, S, O, -NH2, -NHCOCH3, -OH, lower alkoxy (C1-C4), and halogens (-F, -Cl, -Br, or -I)).

[0080] As used herein, the term "substituted aliphatic" refers to an alkane, alkene, alkyne, or alicyclic moiety in which at least one aliphatic hydrogen atom has been replaced, for example, by a halogen, amino, hydroxyl, nitro, thio, ketone, aldehyde, ester, amide, lower aliphatic, substituted lower aliphatic, or cyclic (aryl, substituted aryl, cycloaliphatic, or substituted cycloaliphatic, etc.). Examples of such moiety include, but are not limited to, 1-chloroethyl.

[0081] As used herein, the term "substituted aryl" refers to an aromatic ring or fused aromatic ring system consisting of at least one aromatic ring, wherein at least one hydrogen atom on the ring carbon has been replaced, for example, by a halogen, amino, hydroxyl, nitro, thio, ketone, aldehyde, ester, amide, lower aliphatic, substituted lower aliphatic, or ring (aryl, substituted aryl, cycloaliphatic, or substituted cycloaliphatic). Examples of such a system include, but are not limited to, hydroxyphenyl.

[0082] As used herein, the term "cycloaliphatic" refers to an aliphatic structure that comprises a fused ring system. Examples of such structures include, but are not limited to, naphthalenes.

[0083] As used herein, the term "substituted cycloaliphatic" refers to a cycloaliphatic structure in which at least one aliphatic hydrogen atom has been replaced by a halogen, nitro, thio, amino, hydroxyl, ketone, aldehyde, ester, amide, lower aliphatic, substituted lower aliphatic, or ring (aryl, substituted aryl, cycloaliphatic, or substituted cycloaliphatic). Examples of such structures include, but are not limited to, 1-chlorodecane, bicycloheptane, octane, and nonane (e.g., norbornyl).

[0084] As used herein, the term "heterocycle" means, for example, an aromatic or non-aromatic ring containing one or more heteroatoms. Heteratoms may be the same as or different from each other. Examples of heteroatoms include, but are not limited to, nitrogen, oxygen, and sulfur. Aromatic and non-aromatic heterocycles are well known in the art. Some non-limiting examples of aromatic heterocycles include pyridine, pyrimidine, indole, purine, quinoline, and isoquinoline. Non-limiting examples of non-aromatic heterocyclic compounds include piperidine, piperazine, morpholine, pyrrolidine, and pyrazolidine. Examples of oxygen-containing heterocycles include, but are not limited to, furan, ethylene oxide, 2H-pyran, 4H-pyran, 2H-chromogenene, and benzofuran. Examples of sulfur-containing heterocycles include, but are not limited to, thiophene, benzothiophene, and p-thiazine. Examples of nitrogen-containing rings include, but are not limited to, pyrrole, pyrrolidine, pyrazole, pyrazolidine, imidazole, imidazoline, imidazoline, pyridine, piperidine, pyrazine, piperazine, pyrimidine, indole, purine, benzimidazole, quinoline, isoquinoline, triazole, and triazine. Examples of heterocycles containing two different heteroatoms include, but are not limited to, phenothiazine, morpholine, p-thiazine, oxazine, oxazole, thiazine, and thiazole. The heterocycle may optionally be further substituted with one or more groups selected from aliphatic, nitro, acetyl (i.e., -C(=O)-CH3), or aryl groups.

[0085] As used herein, the term "substituted heterocycle" refers to a heterocyclic structure in which at least one ring carbon atom is replaced by oxygen, nitrogen, or sulfur, and at least one aliphatic hydrogen atom is replaced by a halogen, hydroxyl, thio, nitro, amino, ketone, aldehyde, ester, amide, lower aliphatic, substituted lower aliphatic, or ring (aryl, substituted aryl, cycloaliphatic, or substituted cycloaliphatic). Examples of such heterocycles include, but are not limited to, 2-chloropyranyl.

[0086] As used herein, the term "connector" refers to an organic or inorganic molecule that links multiple functional units of a molecule. In some embodiments, a connector is a single part or chain that contains up to and includes eight consecutive atoms connecting two different structural parts, wherein such atoms are, for example, carbon, nitrogen, oxygen, or sulfur.

[0087] As used herein, the term "lower alkyl-substituted amino" refers to any alkyl unit comprising up to and including eight carbon atoms, wherein one of the aliphatic hydrogen atoms is replaced by an amino group. Examples of such units include, but are not limited to, ethylamino.

[0088] As used herein, the term “derivative” of a compound refers to a chemically modified compound in which the chemical modification occurs on the functional groups or main chain of the compound.

[0089] As used herein, the term "subject" refers to an organism to be treated by the methods of this disclosure. Such an organism preferably includes, but is not limited to, mammals (e.g., rodents, apes, equines, bovines, suidae, canines, felines, etc.), and most preferably includes humans. In the context of this disclosure, the term "subject" generally refers to an individual who will receive or has received treatment for a condition characterized by an eye disease (e.g., administration of a compound of this disclosure and optionally one or more other agents).

[0090] As used in this article, the term “diagnosed” refers to a disease identified by its signs and symptoms (e.g., resistance to conventional treatments) or by genetic analysis, pathological analysis, histological analysis, etc.

[0091] As used herein, the term "effective amount" refers to an amount of compound (e.g., compounds of this disclosure) sufficient to achieve a beneficial or desired result. Effective amounts may be administered in one or more doses, applications, or dosages, and are not limited to a particular formulation or route of administration.

[0092] As used herein, the term "co-administration" refers to administering at least two agents (e.g., compounds of this disclosure) or therapies to a subject. In some embodiments, co-administration of two or more agents / therapies is simultaneous. In some embodiments, the first agent / therapy is administered before the second agent / therapy. Those skilled in the art will understand that the formulations and / or routes of administration of the various agents / therapies used can vary. Those skilled in the art can readily determine the appropriate dosage for co-administration. In some embodiments, when agents / therapies are co-administered, each agent / therapy is administered at a lower dose than would be appropriate for its individual administration. Therefore, co-administration is particularly desirable in embodiments where co-administration of agents / therapies reduces the necessary dose of one or more known potentially harmful (e.g., toxic) agents.

[0093] As used herein, the term "toxic" refers to any adverse or harmful effect on cells or tissues compared to the same cells or tissues before administration of the toxic substance.

[0094] As used herein, the term "pharmaceutical composition" refers to a combination of an active agent and a carrier (inert or active) that makes the composition particularly suitable for diagnostic or therapeutic use in vivo or in vitro.

[0095] As used herein, the term “pharmaceutically acceptable carrier” means any standard pharmaceutical carrier, such as phosphate-buffered saline solutions, water, emulsions (e.g., oil / water or water / oil emulsions), and various types of wetting agents. Compositions may also include stabilizers and preservatives. Examples include carriers, stabilizers, and adjuvants. (See, for example, Martin, Remington's Pharmaceutical Sciences, 15th Ed., Mack Publ. Co., Easton, PA

[1975] ).

[0096] As used herein, the term "pharmaceutically acceptable salt" means any pharmaceutically acceptable salt (e.g., acid or base) of the compounds of this disclosure that, when administered to a subject, provides the compounds of this disclosure or their active metabolites or residues. As those skilled in the art will recognize, "salts" of the compounds of this disclosure can be derived from inorganic or organic acids and bases. Examples of acids include, but are not limited to, hydrochloric acid, hydrobromic acid, sulfuric acid, nitric acid, perchloric acid, fumaric acid, maleic acid, phosphoric acid, glycolic acid, lactic acid, salicylic acid, succinic acid, toluene-p-sulfonic acid, tartaric acid, acetic acid, citric acid, methanesulfonic acid, ethanesulfonic acid, formic acid, benzoic acid, malonic acid, naphthalene-2-sulfonic acid, benzenesulfonic acid, etc. In obtaining the compounds of this disclosure and their pharmaceutically acceptable acid addition salts, other acids, such as oxalic acid, although not pharmaceutically acceptable on their own, may be used as intermediates in the preparation of the salts.

[0097] Examples of bases include, but are not limited to, alkali metal (e.g., sodium) hydroxides, alkaline earth metal (e.g., magnesium) hydroxides, ammonia, and those of the formula NW4. + Compounds in which W is C 1-4 Alkyl groups, etc.

[0098] Examples of salts include, but are not limited to: acetates, adipates, alginates, aspartates, benzoates, benzenesulfonates, hydrogen sulfates, butates, citrates, camphorates, camphorsulfonates, cyclopentanepropionates, digluconates, dodecyl sulfates, ethanesulfonates, fumarates, flucoheptanoates, glycerophosphates, hemisulfates, heptanates, hexanoates, hydrochlorides, hydrobromide, hydroiodates, 2-hydroxyethanesulfonates, lactates, maleates, methanesulfonates, 2-naphthalenesulfonates, nicotinates, oxalates, palmates, pectates, persulfates, phenylpropionates, picrates, neopentanoates, propionates, succinates, tartrates, thiocyanates, toluenesulfonates, undecanoates, etc. Other examples of salts include those with suitable cations such as Na+. + NH4 + and NW4 + (where W is C) 1-4 The anions of the compounds disclosed herein are composed of alkyl groups and other compounds.

[0099] For therapeutic use, salts of the compounds disclosed herein are considered pharmaceutically acceptable. However, it has also been found that salts of non-pharmaceutically acceptable acids and bases can be used, for example, in the preparation or purification of pharmaceutically acceptable compounds.

[0100] As used herein, the term "sample" is used in its broadest sense. In a sense, this means including specimens or cultures obtained from any source, as well as biological and environmental samples. Biological samples can be obtained from animals (including humans) and include fluids, solids, tissues, and gases. Biological samples include blood products such as plasma, serum, etc. Environmental samples include environmental materials such as surface substances, soil, water, and industrial samples. However, such examples should not be construed as limiting the types of samples applicable to this disclosure.

[0101] As used herein, the term "purified" or "purified" means the removal of unwanted components from a sample. As used herein, the term "substantially purified" means that at least 60%, preferably 75%, and most preferably 90% or more of the molecules that are free from other components with which they normally associate.

[0102] The term "test compound" refers to any chemical entity, pharmaceutical agent, etc., that can be used to treat or prevent a disease, ailment, disorder, or morbidity affecting bodily function, or otherwise alter the physiological or cellular state of a sample (e.g., pyruvate kinase levels). Test compounds include both known and potential therapeutic compounds. A test compound can be determined to be therapeutic by using the screening methods of this disclosure. "Known therapeutic compound" refers to a therapeutic compound that has been shown (e.g., through animal studies or prior experience with human administration) to be effective in such treatment or prevention. Detailed Implementation

[0103] This article provides compositions and methods for activating pyruvate kinase (e.g., in a subject). In particular, this article provides compositions and methods for treating diseases or conditions (e.g., eye diseases, blood disorders, or cancer) using pyruvate kinase activators.

[0104] Metabolic reprogramming of photoreceptors is a therapeutic solution for vision loss associated with AMD, retinal dystrophy, and retinal detachment. -Ali, N. et al. Cell 161, 817–832 (2015); Zhang, L. et al. J. Clin. Invest. 126, 4659–4673 (2016); Include Wubben et al. Sci Rep. 2017; Wubben et al. Sci Rep. 2020). PKM2 (a key regulator of aerobic glycolysis and energy metabolism in photoreceptors) is activated by small molecule activators (e.g., those described herein) to reprogram metabolism and enhance energy production by favoring catabolistic activity in cells. Limiting PKM2 expression and aerobic glycolysis in intraocular photoreceptors reduces any potential off-target effects after ocular delivery, increases therapeutic specificity, and expands the therapeutic window.

[0105] Similar to cells with high metabolic demands (including tumor cells), photoreceptors maintain PKM2 expression (Rajala et al., ibid.; Lindsay et al., ibid.; Ng, SK et al. Clin. Experiment. Ophthalmol. 43, 367–376 (2015)). This contrasts sharply with other terminally differentiated neurons, which express only the constitutively active PKM1 isoform (Jurica, MS et al. Struct. Lond. Engl. 1993 6, 195–210 (1998)). Unlike PKM1, PKM2 activity is tightly regulated in the cell. As a tetramer, PKM2 exhibits high catalytic activity and is involved in ATP synthesis and catabolism. The non-tetrameric form exhibits low catalytic activity and is involved in the anabolism of metabolic intermediates and shuttle pathways to biosynthesis (Gui, DY, et al. Sci. Signal. 6, pe7 (2013); Wong, N., et al. Cancer Lett. 356, 184–191 (2015); Yang, W. & Lu, ZJ Cell Sci. 128, 1655–1660 (2015)). A mouse model of selective deletion of the PKM2 isoform in photoreceptors demonstrated a net increase in compensatory PKM1 isoform expression and overall PKM activity in the retina (see, for example, WO 2019 / 079541; incorporated herein by reference in its entirety). This mouse model showed reduced retinal cell death under acute photoreceptor degeneration induced by retinal detachment. The model also showed reduced phosphorylation of PKM2, which promoted tetramerization and increased enzyme activity in the rodent retina during photoreceptor stress. Therefore, the metabolic reprogramming observed in the retinas of photoreceptor-specific PKM2 knockout mice mimics PKM2 activation following nutrient deprivation, circumventing acute apoptotic stress by replacing constitutively active PKM1 (Wubben et al. Sci Rep. 2017). Furthermore, it has been demonstrated that small-molecule activation of PKM2 with ML-265 also circumvents photoreceptor cell apoptosis in an external retinal stress model without any long-term toxic effects on the retina (Wubben et al. Sci Rep. 2020).

[0106] Therefore, this article provides compositions and methods for activating pyruvate kinases (e.g., PKM1 or PKM2) to reprogram photoreceptor metabolism and block apoptosis, thereby preventing vision loss in many retinal diseases and treating other diseases or conditions (e.g., blood disorders or cancer).

[0107] I. Activator

[0108] This article provides pyruvate kinase (e.g., PKM1 or PKM2) activators. Exemplary, non-limiting examples are provided below.

[0109] In some embodiments, the compound is selected from...

[0110]

[0111] X1, X2, X3, and X4 are independently selected from CH, CO, N, NH, S, or O;

[0112] Z represents a vacancy, bond, optional substitution of a C1-6 alkyl group, -O-, -S-, -CH2-, -CHR5-, -CR5R6-, or -(CH2). n -、-(CHR5) n -、-(CR5R6) n -, -S(=O)CH2, -S(=O)2CH2-, -NR5-, -NR5C(=O)-, -C(O)NR5-, -C(=O)-, -OC(=O)-, -C(=O)O -, -NR5C(=O)O-, -OC(=O)NR5-, -NR5C(=O)NR5-, -OC(R5)2-, -C(R5)2O-, -NR4C(R5)2-, C(R6 )2NR5-、-S(=O)-、S(=O)2-、-S(=O)2O-、-OS(=O)2-、-S(=O)2NR5-、-NR5S(=O)2-、-S(=O)NR5-、-NR5S(=O-、-OS(=O)NR5-、-NR5S(=O)O- or –S(=O)(=NR5)-, where the connection point with R1 or R2 is located on the left and n=1-6;

[0113] R5 and R6 are each independently hydrogen, halogen, -CN, OR7, NR7R8, -N(R7)C(=O)R8, -C(=O)N(R7), -C(=O)R7, -C(=O)OR7, -SR7, -S(=O)R7, -S(=O)2R7 or any optimally substituted –C1-C6 alkyl;

[0114] R7 and R8 are each independently hydrogen, any optimally substituted –C1-C6 alkyl; or alternatively, R7 and R8 together are either an optionally substituted C1-C6 monocyclic cycloalkyl ring or an optionally substituted monocyclic heterocycle.

[0115] Y represents an empty space, -CH2-, -CHR9-, or -CR9R. 10 -、-(CH2) n -、-(CHR9) n -、-(CR9R 10 ) n-, -C(=O)-, -S(=O)-, -S(=O)2-, where n=1-6;

[0116] R9 and R 10 Each is independently hydrogen, halogen, -CN, or any optimally substituted –C1-C6 alkyl; or alternatively R9 and R 10 They can be used together as optional substituted C1-C6 monocyclic cycloalkyl rings or optional substituted monocyclic heterocycles;

[0117] R1 is -H, -F, -Cl, -Br, -NO2, -CN, -NH2, -NHR 11 -NR 11 R 12 -OH, -OR 11 -CH2-, -CHR9-, -CR9R 10 -(CH2) n -、-(CHR9) n -、-(CR9R 10 ) n - Optionally substituted C1-C6 alkyl, Optionally substituted monocyclic cycloalkyl, Optionally substituted bicyclic cycloalkyl, Optionally substituted tricyclic cycloalkyl, Optionally substituted monocyclic heterocycle, Optionally substituted bicyclic heterocycle or Optionally substituted tricyclic heterocycle;

[0118] R2 is -H, -F, -Cl, -Br, -NO2, -CN, -NO2, -NH2, -NHR 11 -NR 11 R 12 -OH, -OR 11 Optionally substituted C1-C6 alkyl, Optionally substituted monocyclic cycloalkyl, Optionally substituted bicyclic cycloalkyl, Optionally substituted tricyclic cycloalkyl, Optionally substituted monocyclic heterocycle, Optionally substituted bicyclic heterocycle, Optionally substituted tricyclic heterocycle;

[0119] R3 is -H, -F, -Cl, -Br, -NO2, -CN, -NO2, -NH2, -NHR 11 -NR 11 R 12 -OH, -OR 11 Optionally substituted C1-C6 alkyl, Optionally substituted monocyclic cycloalkyl, Optionally substituted bicyclic cycloalkyl, Optionally substituted tricyclic cycloalkyl, Optionally substituted monocyclic heterocycle, Optionally substituted bicyclic heterocycle, Optionally substituted tricyclic heterocycle;

[0120] R 11 and R 12Each of them independently is an optional substituted C1-C6 alkyl, an optional substituted monocyclic cycloalkyl, an optional substituted bicyclic cycloalkyl, an optional substituted tricyclic cycloalkyl, an optional substituted monocyclic heterocycle, an optional substituted bicyclic heterocycle, or an optional substituted tricyclic heterocycle;

[0121] R4 represents hydrogen, -CH3, and -CHR. 13 -CR 13 R 14 -S(=O)R 13 -S(=O)2R 13 Optionally substituted alkyl groups, preferably substituted haloalkyl groups, preferably substituted alkenyl groups, preferably substituted alkynyl groups, preferably substituted cycloalkyl groups, preferably substituted heterocyclic groups, preferably substituted aryl groups, -C(=O)R 15 Or nitrogen protecting group; wherein;

[0122] R 13 and R 14 Each of these can be independently a optionally substituted C1-C6 alkyl, an optionally substituted monocyclic cycloalkyl, an optionally substituted bicyclic cycloalkyl, an optionally substituted tricyclic cycloalkyl, an optionally substituted monocyclic heterocycle, an optionally substituted bicyclic heterocycle, or an optionally substituted tricyclic heterocycle.

[0123] R 15 It is hydrogen, -CH3, optionally substituted alkyl, preferably substituted haloalkyl, preferably substituted alkenyl, preferably substituted alkynyl, preferably substituted cycloalkyl, preferably substituted heterocyclic, preferably substituted aryl, or a pharmaceutically acceptable salt thereof.

[0124] In some implementations, R2-Z- is H and R1-Z- is selected from;

[0125] Z represents an empty space, a bond, or an optional substitution C. 1-6 Alkyl, -O-, -S-, -CH2-, -CHR5-, -CR5R6-, -(CH2) n -、-(CHR5) n -、-(CR5R6) n -、-S(=O)CH2、-NR5-、-NR5C(=O)-、-C(O)NR5-、-C(=O)-、-S(=O)-、where the connection point with R1 is located on the left and n=1-6;

[0126] R5 and R6 are each independently hydrogen, -CN, OR7, NR7R8, or any optimally substituted –C1-C6 alkyl; or alternatively, R7 and R8 may together be an optionally substituted C1-C6 monocyclic cycloalkyl ring or an optionally substituted monocyclic heterocycle; wherein;

[0127] R7 and R8 are each independently hydrogen, any optimally substituted –C1-C6 alkyl; or alternatively, R7 and R8 may be together as an optionally substituted C1-C6 monocyclic cycloalkyl ring or an optionally substituted monocyclic heterocycle.

[0128] Y represents an empty space, -CH2-, -CHR9-, or -CR9R. 10 -、-(CH2) n -、-(CHR9) n -、-(CR9R 10 ) n -, where n = 1-3;

[0129] R9 and R 10 Each is independently hydrogen or any optimally substituted –C1-C6 alkyl; or alternatively R9 and R 10 They can be used together as optional substituted C1-C6 monocyclic cycloalkyl rings or optional substituted monocyclic heterocycles;

[0130] R1 is -H, -F, -Cl, -Br, -NO2, -CN, -NO2, -NH2, -NHR 11 -NR 11 R 12 -OH, -OR 11 -CH2-, -CHR9-, -CR9R 10 -、-(CH2) n -、-(CHR9) n -、-(CR9R 10 ) n - Optionally substituted C1-C6 alkyl, Optionally substituted monocyclic cycloalkyl, Optionally substituted bicyclic cycloalkyl, Optionally substituted tricyclic cycloalkyl, Optionally substituted monocyclic heterocycle, Optionally substituted bicyclic heterocycle or Optionally substituted tricyclic heterocycle;

[0131] R2 is -H, -F, -Cl, -Br, -NO2, -CN, -NO2, -NH2, -NHR 11 -NR 11 R 12 -OH, -OR 11 -CH2-, -CHR9-, -CR9R 10 -、-(CH2) n -、-(CHR9) n -、-(CR9R 10 ) n - Optionally substituted C1-C6 alkyl, Optionally substituted monocyclic cycloalkyl, Optionally substituted bicyclic cycloalkyl, Optionally substituted tricyclic cycloalkyl, Optionally substituted monocyclic heterocycle, Optionally substituted bicyclic heterocycle or Optionally substituted tricyclic heterocycle;

[0132] R3 is -H, -F, -Cl, -Br, -NO2, -CN, -NO2, -NH2, -NHR 11 -NR 11 R 12 -OH, -OR 11 -CH2-, -CHR9-, -CR9R 10 -、-(CH2) n -、-(CHR9) n -、-(CR9R 10 ) n - Optionally substituted C1-C6 alkyl, Optionally substituted monocyclic cycloalkyl, Optionally substituted bicyclic cycloalkyl, Optionally substituted tricyclic cycloalkyl, Optionally substituted monocyclic heterocycle, Optionally substituted bicyclic heterocycle, Optionally substituted tricyclic heterocycle;

[0133] R 11 and R 12 Each of them independently is an optional substituted C1-C6 alkyl, an optional substituted monocyclic cycloalkyl, an optional substituted bicyclic cycloalkyl, an optional substituted tricyclic cycloalkyl, an optional substituted monocyclic heterocycle, an optional substituted bicyclic heterocycle, or an optional substituted tricyclic heterocycle;

[0134] R4 is –CH3.

[0135] In some exemplary embodiments, Z-R1, Z-R2, Z-R3, and Y-R3 are selected individually or in combination, for example, Where R 16 and R 17 It can be located at the ortho, meta, or para position of the aryl ring and is selected from H, -OCH3, C1-C4 alkyl, -NH2, -halogen, -CN, -OH, -S(=O)Me, -S(=O)2Me, -CH2OMe, -CH2NR 18 R 19 , where R 18 and R 19 Selected from -H, C1-C4 alkyl, optimally substituted aryl or heterocyclic, or together forming a carbocyclic or heterocyclic ring;

[0136] In some embodiments, the compound is a compound of formula I, wherein Z-R2 is H, Z-R1 is not 8-OCH3, and R4 is not... And Y-R3 is not In some embodiments, the compound is a compound of formula I, wherein Z-R2 is H, Z-R1 is not 8-Cl, and R4 is not CH3 or And Y-R3 is not CH3 or In some embodiments, the compound is a compound of formula I and R2 is not. Or CH3, and Y-R3 is not In some embodiments, the compound is a compound of formula I and Z-R1 is not. X1 is not CO, R2 is not CH3, and Y-R3 is not In some embodiments, the compound is not Where R1 and R2 are independently CH3 or In some embodiments, the compound is selected from the compounds described in Table 2. In some embodiments, the compound is:

[0137]

[0138] Wherein R1 is a substituted monocyclic or bicyclic carbocyclic or heterocyclic moiety and R2 is a substituted monocyclic or bicyclic heterocyclic moiety.

[0139] In some exemplary embodiments, the compound is

[0140]

[0141]

[0142]

[0143]

[0144]

[0145]

[0146]

[0147] In some embodiments, the compound is

[0148]

[0149] in

[0150] X is an empty space or a selection from -H, -CH2-, -CHR3-, -CR3R4-, or -(CH2). n -、-(CHR3) n -、-(CR3R4) n -, where n–1–6;

[0151] R1 is selected from, for example, -H, -CN, -NO2, -NH2, -NHR3, NR3R4, -OH, OR3, -SOR3, -SO2R3, optionally substituted C1-C6 alkyl, optionally substituted monocyclic cycloalkyl, optionally substituted bicyclic cycloalkyl, optionally substituted tricyclic cycloalkyl, optionally substituted monocyclic heterocycle, optionally substituted bicyclic heterocycle or optionally substituted tricyclic heterocycle;

[0152] R2 is selected from, for example, -H, -CN, -NO2, -NH2, -NHR3, NR3R4, -OH, OR3, -SOR3, -SO2R3, optionally substituted C1-C6 alkyl, optionally substituted monocyclic cycloalkyl, optionally substituted bicyclic cycloalkyl, optionally substituted monocyclic heterocycle, optionally substituted bicyclic heterocycle or optionally substituted tricyclic heterocycle.

[0153] R3 and R4 are each independently selected from the group consisting of, for example, optionally substituted C1-C6 alkyl, optionally substituted monocyclic cycloalkyl, optionally substituted bicyclic cycloalkyl, optionally substituted tricyclic cycloalkyl, optionally substituted monocyclic heterocycle, optionally substituted bicyclic heterocycle, or optionally substituted tricyclic heterocycle; and

[0154] In some implementations, R1 is selected from, for example, And R2 is selected from, for example,

[0155] In another embodiment, the compound is

[0156]

[0157] R1 is selected from, for example, -H, -F, -Cl, -Br, -CN, -NO2, -NH2, -NHR3, -NR3R4, -OH, -OR3, optionally substituted C1-C6 alkyl, optionally substituted monocyclic cycloalkyl, optionally substituted bicyclic cycloalkyl, optionally substituted tricyclic cycloalkyl, optionally substituted monocyclic heterocycle, optionally substituted bicyclic heterocycle or optionally substituted tricyclic heterocycle;

[0158] R2 is selected from, for example, -H, -CH3, -(CH2). n -R5, -(CHR3) n -R5、-(CR3R4) n -R5, optionally substituted C1-C6 alkyl, optionally substituted monocyclic cycloalkyl, optionally substituted bicyclic cycloalkyl, optionally substituted monocyclic heterocycle, optionally substituted bicyclic heterocycle or optionally substituted tricyclic heterocycle;

[0159] R3 and R4 are each independently selected from, for example, optionally substituted C1-C6 alkyl groups, optionally substituted monocyclic cycloalkyl groups, optionally substituted bicyclic cycloalkyl groups, optionally substituted tricyclic cycloalkyl groups, optionally substituted monocyclic heterocycles, optionally substituted bicyclic heterocycles, or optionally substituted tricyclic heterocycles; and

[0160] R5 is -H, -F, -Cl, -Br, -NO2, -CN, -NO2, -NH2, -NHR3, -NR3R4, -OH, -OR3, optionally substituted C1-C6 alkyl, optionally substituted monocyclic cycloalkyl, optionally substituted bicyclic cycloalkyl, optionally substituted tricyclic cycloalkyl, optionally substituted monocyclic heterocycle, optionally substituted bicyclic heterocycle, optionally substituted tricyclic heterocycle.

[0161] In some embodiments, R1 is selected from, for example, OCH3, OH, NH2, NCH3, N(CH3)2,

[0162]

[0163] And R2 is selected from, for example,

[0164]

[0165] In some embodiments, one or more hydrogen atoms or any of the aforementioned compounds are replaced with deuterium (D) (see, for example, Cargnin et al., Future Medicinal Chemistry; (2019) 11(16), 2039–2042)). Since D has twice the mass of H, the CD bond is more resistant to oxidative processes, such as those catalyzed by CYP450 or other metabolically involved enzymes, while generally retaining very similar steric properties. Therefore, isosteric substitutions of HD corresponding to the oxidizable soft spot generally preserve pharmacodynamics while improving drug pharmacokinetics, affecting half-life and / or area under the curve, and ultimately influencing dosage and / or dosing regimens.

[0166] The following Example 1 describes the synthesis and activity of the compound.

[0167] II. Treatment methods

[0168] In one embodiment, a method for treating diseases, symptoms, or ailments as described herein is provided.

[0169] (For example, treatment), the method includes administering the compound, the pharmaceutically significant aspect of the compound.

[0170] An acceptable salt or a pharmaceutical composition comprising the compounds described herein.

[0171] The compounds and compositions described herein may be administered, for example, in vitro or ex vivo to cultured cells, or, for example, in vivo to a subject, to treat and / or diagnose a variety of conditions, including those described below.

[0172] In some embodiments, methods and uses of the aforementioned pyruvate kinase (e.g., PKM2) activator in the treatment of ocular diseases are provided. In some embodiments, the ocular disease is, for example, retinal dystrophy, vision loss, macular degeneration, retinal detachment, or proliferative vitreoretinopathy. In some embodiments, the administration prevents or reduces photoreceptor cell death in the subject's eye.

[0173] In some embodiments, this disclosure provides compositions, kits, systems, and / or methods for preventing, inhibiting, blocking, and / or reducing (e.g., in human subjects with this need) photoreceptor cell death. In some embodiments, the activator inhibits photoreceptor cell apoptosis. In some embodiments, photoreceptor death and / or apoptosis are caused by retinal detachment, age-related macular degeneration, trauma, inflammation, uveitis, diabetes, hereditary retinal degeneration, and / or diseases affecting photoreceptor cells. In some embodiments, photoreceptor death and / or apoptosis are caused by retinal detachment. In some embodiments, retinal detachment is caused by one or more underlying diseases, disorders, or conditions (e.g., age-related macular degeneration, trauma, inflammation, uveitis, diabetes, hereditary retinal degeneration, etc.). In some embodiments, this disclosure has been found to enhance photoreceptor viability and / or inhibit photoreceptor death in a variety of conditions and / or diseases, including but not limited to macular degeneration (e.g., dry, wet, non-exudative, or exudative / neovascular), hereditary retinal degeneration (e.g., retinitis pigmentosa, Stargardt's disease, Usher syndrome, etc.), inflammatory eye diseases (e.g., uveitis), ocular infections (e.g., bacterial, fungal, viral), autoimmune retinitis (e.g., caused by infection), trauma, diabetic retinopathy, choroidal neovascularization, retinal ischemia, retinal vascular occlusive diseases (e.g., retinal vein branch occlusion, central retinal vein occlusion, retinal artery branch occlusion, central retinal artery occlusion, etc.), pathological myopia, angiostripes, macular edema (e.g., of any etiology), and / or central serous chorioretinopathy.

[0174] In some embodiments, this disclosure provides a method for treating a patient suffering from such retinal detachment and / or retinal disease and requiring treatment. In some embodiments, a pharmaceutical composition comprising at least one PKM2 activator described herein is delivered to such a patient in an amount and location sufficient to treat the disease or ailment. In some embodiments, the activator (or a pharmaceutical composition comprising such an activator) may be delivered systemically or locally to the patient, and a person of ordinary skill in treating such a patient is capable of determining the most appropriate route of delivery, timing, and therapeutic dose. It should be understood that the application of the method of treating the patient is most preferably intended to substantially alleviate or even eliminate such symptoms; however, as with many medical treatments, the application of the method is considered successful if, during, after, or otherwise as a result, the patient's symptoms of the disease or ailment subside to a definite degree.

[0175] In some embodiments, the compounds described herein are used to treat blood disorders (e.g., those described herein). In one embodiment of this disclosure, a method is provided for increasing the lifespan of red blood cells (RBCs) when needed, the method comprising contacting blood with an effective amount of the compounds described herein or pharmaceutically acceptable salts thereof; or contacting a pharmaceutically acceptable composition comprising the compounds described herein or pharmaceutically acceptable salts thereof and a pharmaceutically acceptable carrier.

[0176] In another embodiment, the compound or pharmaceutical composition is added directly to whole blood or concentrated red blood cells (e.g., in vitro). In yet another embodiment, the compound or pharmaceutical composition is administered to a subject who requires it.

[0177] In one embodiment of this disclosure, a method is provided for adjusting the level of 2,3-diphosphoglycerate in the blood when needed, by contacting the blood with an effective amount of the compound described herein or a pharmaceutically acceptable salt thereof; or a pharmaceutically acceptable composition comprising the compound described herein or a pharmaceutically acceptable salt thereof and a pharmaceutically acceptable carrier.

[0178] In one embodiment of this disclosure, a method for treating sickle cell disease is provided, the method comprising administering to a subject in need an effective amount of the compound described herein or a pharmaceutically acceptable salt thereof; or a pharmaceutically acceptable composition comprising the compound described herein or a pharmaceutically acceptable salt thereof and a pharmaceutically acceptable carrier.

[0179] As used herein, sickle cell disease (SCD), hemoglobin SS disease, and sickle cell anemia are used interchangeably. Sickle cell disease (SCD) describes a group of inherited red blood cell disorders. In some embodiments, a subject with SCD has an abnormal hemoglobin in their red blood cells, referred to as hemoglobin S or sickle hemoglobin. In some embodiments, a subject with SCD has at least one abnormal gene that causes the body to produce hemoglobin S. In some embodiments, a subject with SCD has two hemoglobin S genes, namely hemoglobin SS.

[0180] In one embodiment of this disclosure, a method for treating a subject with pyruvate kinase deficiency (PKD) is provided, the method comprising administering to the subject an effective amount of the compound described herein or a pharmaceutically acceptable salt thereof; or a pharmaceutically acceptable composition comprising the compound described herein or a pharmaceutically acceptable salt thereof and a pharmaceutically acceptable carrier.

[0181] In one embodiment of this disclosure, a method of treating anemia in a subject is provided, the method comprising administering to the subject an effective amount of the compound described herein or a pharmaceutically acceptable salt thereof; or a pharmaceutically acceptable composition comprising the compound described herein or a pharmaceutically acceptable salt thereof and a pharmaceutically acceptable carrier. In some embodiments, the anemia is erythropoietic anemia, such as type I, II, III, or IV congenital erythropoietic anemia. In some embodiments, the anemia is hemolytic anemia. In some embodiments, the hemolytic anemia is a congenital and / or hereditary form of hemolytic anemia, such as PKD, sickle cell disease, thalassemia (e.g., α or β), hereditary spherocytosis, hereditary elliptic polycythemia, paroxysmal nocturnal hemoglobinuria, or aβ-lipoproteinemia (Bassen-Kornzweig syndrome). In some embodiments, the hemolytic anemia is acquired hemolytic anemia, such as autoimmune hemolytic anemia or drug-induced hemolytic anemia. In some implementations, hemolytic anemia is anemia that is part of a multisystem disease, such as congenital erythropoietic purpura, Fanconi anemia, and Diamond-Blackfan anemia.

[0182] As used herein, the term “anemia” refers to a deficiency of red blood cells (RBCs) and / or hemoglobin. As used herein, anemia includes all types of clinical anemia, such as (but not limited to): microcytic anemia, iron deficiency anemia, hemoglobinopathies, heme synthesis defects, globin synthesis defects, sideroblast defects, normocytic anemia, anemia of chronic disease, aplastic anemia, hemolytic anemia, macrocytic anemia, megaloblastic 10 anemia, pernicious anemia, dimorphic anemia, anemia of preterm infants, Fanconi anemia, hereditary spherocytosis, sickle cell disease, warm-type autoimmune hemolytic anemia, cold agglutinin hemolytic anemia, osteosclerosis, thalassemia, and myelodysplastic syndromes.

[0183] In some embodiments, this document provides a method for increasing the amount of hemoglobin in a subject by administering an effective amount of a compound described herein or a pharmaceutically acceptable salt thereof; or a pharmaceutically acceptable composition comprising a compound described herein or a pharmaceutically acceptable salt thereof and a pharmaceutically acceptable carrier. In some embodiments, the provided method increases the hemoglobin concentration in a subject.

[0184] In one embodiment of this disclosure, a method for treating hemolytic anemia is provided, the method comprising administering to a subject an effective amount of the compound described herein or a pharmaceutically acceptable salt thereof; or a pharmaceutically acceptable composition comprising the compound described herein or a pharmaceutically acceptable salt thereof and a pharmaceutically acceptable carrier. In other embodiments, the hemolytic anemia is hereditary and / or congenital hemolytic anemia, acquired hemolytic anemia, or anemia as part of a multisystem disease. In some embodiments, the hemolytic anemia is congenital. In some embodiments, the hemolytic anemia is hereditary (e.g., nonspherocytic hemolytic anemia or hereditary spherocytosis).

[0185] In one embodiment of this disclosure, a method is provided for treating thalassemia; hereditary spherocytosis; hereditary ellipocytosis; abetalipoproteinemia or Basen-Kornzweig syndrome; paroxysmal nocturnal hemoglobinuria; acquired hemolytic anemia (e.g., congenital anemia (e.g., enzyme disease)); sickle cell disease; or anemia of chronic disease, the method comprising administering to a subject an effective amount of the compound described herein or a pharmaceutically acceptable salt thereof; or a pharmaceutically acceptable composition comprising the compound described herein or a pharmaceutically acceptable salt thereof and a pharmaceutically acceptable carrier. In one embodiment, acquired hemolytic anemia includes congenital anemia. In some embodiments, the method provided is for treating thalassemia. In some embodiments, the thalassemia is β-thalassemia.

[0186] In some embodiments, the compounds described herein are used to treat proliferative disorders (e.g., those described herein). In some embodiments, a method of treating a proliferative disorder is provided, the method comprising administering to a subject an effective amount of a compound described herein or a pharmaceutically acceptable salt thereof; or a pharmaceutically acceptable composition comprising a compound described herein or a pharmaceutically acceptable salt thereof and a pharmaceutically acceptable carrier. As used herein, “proliferative disorder” refers to a disease resulting from abnormal growth or expansion due to cell proliferation (Walker, Cambridge Dictionary of Biology; Cambridge University Press: Cambridge, UK, 1990). Proliferative disorders may be associated with: 1) pathological proliferation of normal quiescent cells; 2) pathological displacement of cells from their normal location (e.g., metastasis of neoplastic cells); 3) pathological expression of proteolytic enzymes (such as matrix metalloproteinases (e.g., collagenase, gelatinase, and elastase)); or 4) pathological angiogenesis, such as proliferative retinopathy and tumor metastasis. Exemplary proliferative disorders include cancer (i.e., “malignant growths”), benign growths, angiogenesis, inflammatory diseases, and autoimmune diseases. In some embodiments, the proliferative disease is cancer. In some embodiments, the proliferative disease is an autoimmune disease. In some embodiments, the proliferative disease is proliferative vitreoretinopathy.

[0187] The terms “vesicle” and “tumor” are used interchangeably herein and refer to an abnormal mass of tissue that grows beyond and is not coordinated with the growth of normal tissue. A vegetation or tumor may be “benign” or “malignant” depending on the following characteristics: degree of cell differentiation (including morphology and function), growth rate, local invasion, and metastasis. “Benign vegetations” are typically well-differentiated, grow more slowly than malignant vegetations, and remain confined to the primary site. Furthermore, benign vegetations do not have the ability to infiltrate, invade, or metastasize to distant sites. Exemplary benign vegetations include, but are not limited to, lipomas, chondromas, adenomas, acanthomas, senile hemangiomas, seborrheic keratosis, lentigines, and sebaceous hyperplasia. In some cases, certain “benign” tumors may later develop into malignant vegetations, possibly due to additional genetic changes in a subset of the tumor’s vesicular cells, and these tumors are referred to as “premalignant vegetations.” An exemplary premalignant vegetation is a teratoma.

[0188] Conversely, "malignant growths" are typically poorly differentiated (anaplastic) and characterized by rapid growth accompanied by progressive infiltration, invasion, and destruction of surrounding tissues. Furthermore, malignant growths often have the ability to metastasize to distant sites. The terms "metastasis," "metastatic," or "metastasize" refer to the spread or migration of cancer cells from a primary or original tumor to another organ or tissue, and are usually identified by the presence of a "secondary tumor" or "secondary cell mass" of the tissue type in which the primary or original tumor is located, rather than those in the organ or tissue where the secondary (metastatic) tumor is located. For example, prostate cancer that has metastasized to bone is called metastatic prostate cancer and includes cancerous prostate cancer cells growing in bone tissue.

[0189] The term "cancer" refers to a class of diseases characterized by the development of abnormal cells that proliferate uncontrollably and have the ability to infiltrate and destroy normal body tissues. See, for example, Stedman's Medical Dictionary, 25th ed.; Hensyl ed.; Williams & Wilkins: Philadelphia, 1990. Exemplary cancers include solid tumors, soft tissue tumors, and their metastases. The disclosed methods can also be used to treat non-solid cancers. Exemplary solid tumors include malignant tumors of various organ systems (e.g., sarcomas, adenocarcinomas, and carcinomas), such as those of the lungs, breasts, lymph nodes, gastrointestinal tract (e.g., colon), and genitourinary tracts (e.g., kidney, urothelial, or testicular tumors), pharynx, prostate, and ovaries. Exemplary adenocarcinomas include colorectal cancer, renal cell carcinoma, liver cancer, non-small cell lung cancer, and small bowel cancer. Other exemplary cancers include: acute lymphoblastic leukemia (ALL), adults; acute lymphoblastic leukemia (ALL), children; acute myeloid leukemia (AML), adults; adrenocortical carcinoma; adrenocortical carcinoma, children; AIDS-related lymphoma; AIDS-related malignancies; anal cancer; astrocytoma, cerebellum in children; astrocytoma, brain in children; bile duct cancer, extrahepatic; bladder cancer; bladder cancer, children; bone cancer, osteosarcoma / malignant fibrous histiocytoma; brainstem glioma, children; brain tumors, adults; brain tumors, brainstem glioma, children; brain tumors, cerebellar astrocytoma, children; brain tumors, astrocytoma / malignant glioma, children; brain tumors, ependymoma, children; brain tumors, medulloblastoma, children; brain tumors, supratentorial primitive neuroectodermal tumors. Tumors, Children; Brain Tumors, Visual Pathway and Hypothalamic Gliomas, Children; Brain Tumors, Children (Other); Breast Cancer; Breast Cancer and Pregnancy; Breast Cancer, Children; Breast Cancer, Men; Bronchial Adenoma / Carcinoid, Children; Carcinoid Tumor, Children; Carcinoid Tumor, Gastrointestinal Tract; Cancer, Adrenal Cortex; Cancer, Pancreatic Islet Cells; Carcinoma of Unknown Primacy; Central Nervous System Lymphoma, Primary; Cerebellar Astrocytoma, Children; Brain Astrocytoma / Malignant Gliomas, Children; Cervical Cancer; Childhood Cancer; Chronic Lymphocytic Leukemia; Chronic Myeloid Leukemia; Chronic Myeloproliferative Disorder; Tenosynovial Clear Cell Sarcoma; Colon Cancer; Colorectal Cancer, Children; Cutaneous T-cell Lymphoma; Endometrial Cancer; Ependymoma, Children; Epithelial Cancer, Ovarian; Esophageal Cancer; Esophageal Cancer, Children; Ewing's Tumor Family (of Tumors); Extracranial germ cell tumors, children; Extragonadal germ cell tumors; Extrahepatic bile duct carcinoma; Ocular cancer, intraocular melanoma; Ocular cancer, retinoblastoma; Gallbladder cancer; Gastric (stomach) cancer; Gastric (stomach) cancer, children; Gastrointestinal carcinoid tumors; Germ cell tumors, extracranial, children;Germ cell tumors, extragonadal; germ cell tumors, ovarian; gestational trophoblastic tumors; gliomas, pediatric brainstem; gliomas, pediatric visual pathway and hypothalamus; hairy cell leukemia; head and neck cancer; hepatocellular carcinoma, adult (primary); hepatocellular carcinoma, pediatric (primary); Hodgkin's lymphoma, adult; Hodgkin's lymphoma, pediatric; Hodgkin's lymphoma during pregnancy; hypopharyngeal cancer; hypothalamic and visual pathway gliomas, pediatric; intraocular melanoma; islet cell carcinoma (endocrine pancreas); Kaposi's sarcoma; renal cell carcinoma; laryngeal cancer; laryngeal cancer, pediatric; leukemia, acute lymphoblastic leukemia, adult; leukemia, acute lymphoblastic leukemia, pediatric; leukemia, acute myeloid leukemia, adult; leukemia... Blood disorders, acute myeloid, children; Leukemia, chronic lymphocytic; Leukemia, chronic myeloid; Leukemia, hairy cell; Lip and oral cancer; Liver cancer, adults (primary); Liver cancer, children (primary); Lung cancer, non-small cell; Lung cancer, small cell; Lymphocytic leukemia, acute in adults; Lymphocytic leukemia, acute in children; Lymphocytic leukemia, chronic; Lymphoma, AIDS-related; Lymphoma, central nervous system (primary); Lymphoma, cutaneous T-cell; Lymphoma, Hodgkin's, adults; Lymphoma, Hodgkin's, children; Lymphoma, Hodgkin's, during pregnancy; Lymphoma, non-Hodgkin's, adults; Lymphoma, non-Hodgkin's, children; Lymphoma, non-Hodgkin's, during pregnancy; Lymphoma, primary central nervous system; Macroglobulin Leukemia, Waldenstrom's; Male breast cancer; Malignant mesothelioma, adult; Malignant mesothelioma, child; Malignant thymoma; Medulloblastoma, child; Melanoma; Melanoma, intraocular; Merkel cell carcinoma; Mesothelioma, malignant; Metastatic squamous neck carcinoma with occult primary; Multiple endocrine tumor syndrome, child; Multiple myeloma / plasmacytoma; Mycosis fungoides; Myelodysplastic syndrome; Myeloid leukemia, chronic; Myeloid leukemia, acute in child; Multiple myeloma; Myeloproliferative disorder, chronic; Nasal and paranasal sinus carcinoma; Nasopharyngeal carcinoma; Nasopharyngeal carcinoma, child; Neuroblastoma; Non-Hodgkin lymphoma, adult; Non-Hodgkin lymphoma, child; Non-Hodgkin lymphoma during pregnancy Hodgkin's lymphoma; non-small cell lung cancer; oral cancer, children; oral and lip cancer; oropharyngeal cancer; osteosarcoma / malignant fibrous histiocytoma of bone; ovarian cancer, children; ovarian epithelial cancer; ovarian germ cell tumors; low-potential ovarian tumors; pancreatic cancer; pancreatic cancer, children; pancreatic cancer, islet cells; paranasal sinus and nasal cavity cancer; parathyroid cancer; penile cancer; pheochromocytoma; pineal and supratentorial primitive neuroectodermal tumors, children; pituitary adenoma; plasmacytoma / multiple myeloma; pleural pulmonary blastoma; pregnancy and breast cancer; pregnancy and Hodgkin's lymphoma; pregnancy and non-Hodgkin's lymphoma; primary central nervous system lymphoma; primary liver cancer, adults; primary liver cancer, children; prostate cancer; rectal cancer; renal cell (kidney) carcinoma; renal cell carcinoma, children;Renal pelvis and ureter, transitional cell carcinoma; retinoblastoma; rhabdomyosarcoma, pediatric; salivary gland carcinoma; salivary gland carcinoma, pediatric; sarcoma, Ewing tumor family; Kaposi's sarcoma; sarcoma (osteosarcoma) / malignant fibrous histiocytoma of bone; sarcoma, rhabdomyosarcoma, pediatric; sarcoma, soft tissue, adult; sarcoma, soft tissue, pediatric; Sezary syndrome; skin cancer; skin cancer, pediatric; skin cancer (melanoma); skin cancer, Merkel cell; small cell lung cancer; small bowel cancer; soft tissue sarcoma, adult; soft tissue sarcoma, pediatric; squamous neck carcinoma with occult lesions Primary and metastatic cancers; gastric (stomach) cancer; gastric (stomach) cancer, children; supratentorial primitive neuroectodermal tumor, children; T-cell lymphoma, skin; testicular cancer; thymoma, children; thymoma, malignant; thyroid cancer; thyroid cancer, children; transitional cell carcinoma of the renal pelvis and ureter; trophoblastic tumor, pregnancy; cancer of unknown primary site, children; rare childhood cancers; transitional cell carcinoma of the ureter and renal pelvis; urethral cancer; uterine sarcoma; vaginal cancer; visual pathway and hypothalamic glioma, children; vulvar cancer; Waldenström macroglobulinemia; and Wilms' tumor. Metastasis of the above cancers can also be treated or prevented according to the methods described herein.

[0190] Compositions can be formulated for topical (e.g., ocular; intraocular space; etc.), parenteral, oral, or topical administration. For example, parenteral formulations may include immediate-release or sustained-release liquid formulations, dry powders, emulsions, suspensions, or any other standard formulations or components thereof. Oral formulations of pharmaceutical compositions may be, for example, liquid solutions, such as an effective amount of the composition dissolved in a diluent (e.g., water, saline, fruit juice, etc.), a suspension in a suitable liquid, or a suitable emulsion. Oral formulations may also be delivered in tablet form and may include excipients, colorants, diluents, buffers, wetting agents, preservatives, flavoring agents, and pharmacologically compatible excipients. Topical formulations may include compounds to enhance the absorption or penetration of the active ingredient through the skin or tissue or other affected areas, such as dimethyl sulfoxide and related analogues. Pharmaceutical compositions may also be delivered topically using transdermal devices (such as patches or pumps, which may include compositions in suitable solvent systems with adhesive systems, such as acrylic emulsions and polyester patches). Compositions may be delivered via eye drops or other topical ocular delivery methods. The composition can be delivered intraocularly at any location in the eye, including, for example, the vitreous cavity, anterior chamber, etc. The composition can be delivered intravitreally, as is commonly done with intravitreal injections of Lucentis (ranibizumab), Avastin (bevacizumab), triamcinolone acetonide, antibiotics, etc. The composition can be delivered periocularly (e.g., to tissues surrounding the eyeball but within the bony orbit). The composition can be delivered via an intraocular implant (e.g., a ganciclovir implant, a fluocinolone implant, etc.). In intraocular implant delivery, a device containing the composition of this disclosure is surgically implanted (e.g., intravitreal), and the drug is released into the eye (e.g., at a predetermined rate). The composition can be administered using encapsulated cell technology (e.g., via Neurotech), in which genetically modified cells are engineered to produce and secrete the composition of this disclosure. The composition can be delivered via transscleral drug delivery using a device that is sutured or placed next to the eyeball, which slowly elutes the drug, allowing the composition to diffuse into the eye.

[0191] In some implementations, the PKM2 activator is co-administered with another treatment for retinal detachment or macular degeneration (e.g., laser or other surgical procedures, ranibizumab, vitamins or nutritional supplements), blood disorders, or cancer (e.g., chemotherapy or radiation therapy).

[0192] In some embodiments, this disclosure provides for the co-administration of two or more of the anti-apoptotic and / or photoreceptor protective compositions described herein. In some embodiments, this disclosure provides for the co-administration of one or more of the anti-apoptotic and / or photoreceptor protective compositions described herein with one or more additional pharmaceutical compositions for treating the conditions described herein (e.g., retinal detachment, blood disorders, or cancer).

[0193] Pharmaceutical compounds can be administered in the form of compositions, formulated according to good pharmaceutical practice with pharmaceutically acceptable carriers and optional excipients, adjuvants, etc. Pharmaceutical compositions can be in solid, semi-solid, or liquid dosage forms: such as powders, solutions, elixirs, syrups, suspensions, creams, drops, pastes, and sprays. As those skilled in the art will recognize, the form of the composition is determined depending on the chosen route of administration (e.g., eye drops, injection, etc.). Generally, unit dosage forms of the compound or agent are preferred to achieve easy and accurate administration of the active pharmaceutical compound. Typically, therapeutically effective pharmaceutical compounds are present in such dosage forms at concentration levels ranging from about 0.5% to about 99% by weight of the total composition: for example, in an amount sufficient to provide the desired unit dose. In some embodiments, the pharmaceutical composition can be administered in single or multiple doses. The specific route of administration and dosage regimen will be determined by a technician based on the condition of the individual to be treated and the individual's response to treatment. In some embodiments, the pharmaceutical composition of a unit dosage form for administration to a subject comprises a pharmaceutical compound and one or more non-toxic, pharmaceutically acceptable carriers, adjuvants, or loads. As mentioned above, the amount of active ingredient that can be combined with such materials to produce a single dosage form will vary depending on various factors. A variety of materials are available as carriers, adjuvants, and loads in the compositions of this disclosure, such as those available in the pharmaceutical industry. Injectable formulations, such as oily solutions, suspensions, or emulsions, can be formulated as known in the art, using suitable dispersants or wetting agents and suspending agents as needed. Sterile injectable formulations can be prepared using non-toxic, parenteral-acceptable diluents or solvents, such as sterile, pyrogen-free water or 1,3-butanediol. Other acceptable loads and solvents that can be used include 5% dextran injection, Ringer's injection, and isotonic sodium chloride injection (as described in USP / NF). Furthermore, sterile, non-volatile oils can generally be used as solvents or suspension media. For this purpose, any mild, non-volatile oil can be used, including synthetic monoglycerides, diglycerides, or triglycerides. Fatty acids (such as oleic acid) can also be used to prepare injectable compositions.

[0194] In some embodiments, the compositions of this disclosure (e.g., small molecule PKM2 activators) are delivered optically, for example, using the techniques described herein and / or other techniques (e.g., injection, topical administration, etc.) (see, for example, Janoria et al. Expert Opinion on Drug Delivery. July 2007, Vol.4, No.4, Pages 371-388; Ghate & Edelhauser. Expert Opin Drug Deliv. 2006 March; 3(2):275-87; Bourges et al. Adv Drug Deliv Rev. 2006 November 15; 58(11):1182-202. Epub 2006 Sep.22; Gomes DosSantos et al. Curr Pharm Biotechnol. 2005 February; 6(1):7-15; all of which are incorporated herein by reference).

[0195] In some embodiments, the composition (e.g., a small molecule PKM2 activator) is provided as part of a kit. In some embodiments, the kit of this disclosure comprises one or more compositions and / or pharmaceutical compositions. In some embodiments, the kit comprises a composition configured for co-administration with one or more other compositions (e.g., pharmaceutical compositions). In some embodiments, one or more compositions are co-administered with one or more other agents to effectively protect photoreceptors and / or inhibit apoptosis or treat blood disorders or cancer.

[0196] experiment

[0197] The following examples are provided to demonstrate and further illustrate certain preferred embodiments and aspects of this disclosure, and should not be construed as limiting the scope of this disclosure.

[0198] Example 1

[0199] Exemplary compounds are described below with reference to the illustrative synthetic schemes for their general preparation and subsequent specific examples. Those skilled in the art will recognize that, in order to obtain the various compounds described herein, starting materials can be suitably selected such that the final desired substituent will be carried out via the reaction scheme with or without appropriate protection to produce the desired product. Alternatively, it may be necessary or desirable to substitute a suitable group for the final desired substituent, which can be carried by the reaction scheme and, where appropriate, replaced by the desired substituent. The reaction can be carried out between the melting point and the reflux temperature of the solvent, and preferably between 0°C and the reflux temperature of the solvent. The reaction can be heated using conventional heating or microwave heating. The reaction can also be carried out in a sealed pressure vessel at a temperature above the normal reflux temperature of the solvent.

[0200] Option 1

[0201]

[0202] Compound II (Scheme 1) was obtained from a commercially available source or generated from commercially available compound I by reaction with ethyl azide under standard Hemetsberger conditions. The conversion of compound II to compound III was carried out using standard conditions with the Vilsmeier-Haack reagent.

[0203] Option 2

[0204]

[0205] In cases where compound II is incompatible with the Wilsmeier-Hacker reaction conditions, compound III is generated via the synthetic sequence shown in Scheme 2. Compound II is treated with N-bromosuccinimide, N-chlorosuccinimide, or N-iodosuccinimide to generate compound IV, where X is Cl, Br, or I. Compound IV is converted to compound V by treatment with trimethylvinyltin under standard Stille cross-coupling. The conversion of compound V to compound III is achieved by treatment with ozone gas to promote olefin cleavage followed by treatment with triphenylphosphine to provide the desired aldehyde. Alternatively, compound V is converted to compound III via olefin dihydroxylation and metal-mediated cleavage. Suitable reagents compatible with the substrate are selected to complete the desired dihydroxylation and cleavage events.

[0206] Option 3

[0207]

[0208] If compound I, in which the desired "Z" moiety is not commercially available, the desired compound can be generated from the desired precursor compound VI via SnAR or a suitable metal-mediated cross-coupling reaction, where X is a functional group or a protected functional group, which will allow for the desired reactivity and functional group incorporation. In some cases, the protecting group is removed prior to the reaction.

[0209] Option 4

[0210]

[0211] If compound II, in which the desired "Z" moiety is not commercially available, the desired compound can be generated from the desired precursor compound VI via SnAR or a suitable metal-mediated cross-coupling reaction, where X is a functional group or a protected functional group, which will allow for the desired reactivity and functional group incorporation. In some cases, the protecting group is removed prior to the reaction.

[0212] Option 5

[0213]

[0214] Compound III is converted to compound VIII by treatment with methyl iodine and potassium carbonate in dimethylformamide. Other bases (such as cesium carbonate, sodium hydride, etc.) provide the same result. Treatment of compound VIII with hydrazine in ethoxyethanol at elevated temperatures leads to the formation of compound IX. Compound X is generated by treating compound IX with potassium tert-butoxide or an equivalent base and the desired alkylating agent. The alkylating agent may contain the desired functionality masked by the protecting group to promote the desired reactivity.

[0215] Chemical:

[0216] In obtaining the compounds described in the following examples and the corresponding analytical data, unless otherwise stated, the following experimental and analytical protocols were followed.

[0217] Unless otherwise specified, the reaction mixture is magnetically stirred at room temperature (rt) under a nitrogen atmosphere. Solutions that are “dried” are typically dried with a drying agent such as Na₂SO₄ or MgSO₄. Mixtures, solutions, and extracts that are “concentrated” are typically concentrated under reduced pressure on a rotary evaporator.

[0218] Normal-phase rapid column chromatography (FCC) is performed on silica gel (SiO2) using a pre-packed cartridge, with elution using a specified solvent.

[0219] At 400 MHz on a Bruker 400Ascend™ spectrometer 1 H frequency and 100MHz 13NMR spectra were obtained at C600 Hz. Chemical shifts (δ) were reported as parts per million (ppm) relative to an internal standard. The final products were purified on a preparative HPLC column (Waters 2545, quaternary gradient module) with a SunFire Prep C18 OBD 5 μm 50 × 100 mm reversed-phase column. The mobile phase consisted of gradient solvent A (H2O with 0.1% TFA) and solvent B (MeCN with 0.1% TFA) at a flow rate of 60 mL / min, with an increase in solvent B of 1% / min. All final compounds were determined to be ≥95% pure, as determined by Waters ACQUITY UPLC using a reversed-phase column (SunFire, C18-5 μm, 4.6 × 150 mm) and gradient solvents A (H2O with 0.1% TFA) and B (CH3CN with 0.1% TFA). ESI mass spectrometry analysis was performed on a Thermo-Scientific LCQ Fleet mass spectrometer.

[0220] Generate chemical names using ChemDraw Ultra 6.0.2 (CambridgeSoft Corp., Cambridge, MA) or ACD / Name Version 9 (Advanced 5 Chemistry Development, Toronto, Ontario, Canada).

[0221] Intermediate A: Methyl 3-formyl-6-methyl-1H-indole-2-carboxylate.

[0222]

[0223] Phosphorus oxychloride (0.30 mL, 3.18 mmol) was added dropwise to DMF (6 mL) at 0 °C. The resulting reaction mixture was warmed to room temperature and stirred for 10 minutes. The reaction mixture was cooled to 0 °C, and methyl 6-methyl-1h-indole-2-carboxylate (503 mg, 2.66 mmol) was added dropwise to DMF (3 mL). The reaction mixture was heated to 80 °C and stirred for 1.5 hours, cooled to room temperature, and poured into 50 mL of ice water. The resulting precipitate was collected by vacuum filtration to provide the desired product (478 mg, 83%). 1 H NMR (400MHz, DMSO-d6) δ12.76(s,1H),10.59(s,1H),8.12(d,J=8.3Hz,1H),7.37–7.34(m,1H),7.15(dd,J=8.3,1.4Hz,1H),3.99(s,3H),2.44(s,3H).

[0224] Intermediate B: Methyl 3-formyl-1,6-dimethyl-1H-indole-2-carboxylate.

[0225]

[0226] To a solution of methyl 3-formyl-6-methyl-1H-indole-2-carboxylate (478 mg, 2.20 mmol) in DMF (4 mL), K₂CO₃ (911 mg, 6.60 mmol) and methyl iodoformate (0.15 mL, 2.42 mmol) were added. The reaction mixture was stirred overnight at room temperature and then poured into water (40 mL). The resulting precipitate was collected by vacuum filtration, providing the desired product (451 mg, 87%). 1 H NMR (400MHz, DMSO-d6) δ10.41(s,1H),8.18(d,J=8.2Hz,1H),7.58–7.52(m,1H),7.26–7.17(m,1H),4.02(s,3H),4.00(s,3H),2.49(s,3H).

[0227] Intermediate C: 5,7-dimethyl-3,5-dihydro-4H-pyridazin[4,5-b]indol-4-one.

[0228]

[0229] Hydrazine (7.81 mmol) was added to a solution of methyl 3-formyl-1,6-dimethyl-1H-indole-2-carboxylate (451 mg, 1.95 mmol) in ethoxyethanol (5 mL), and the reaction mixture was heated to 135 °C overnight. The reaction mixture was cooled to room temperature and water was added. The reaction mixture was extracted with EtOAc (3 × 10 mL). The organic compounds were combined, dried, concentrated, and purified by rapid column chromatography (0–20% MeOH in DCM) to provide the desired product (239 mg, 58%). 1 H NMR (400MHz, DMSO-d6) δ12.75(s,1H),8.71(s,1H),8.07(d,J=8.1Hz,1H),7.58–7.52(m,1H),7.23(dd,J=8.1,1.4Hz,1H),4.25(s,3H),2.54(s,3H).

[0230] Example 1: 3-Benzyl-5,7-dimethyl-3,5-dihydro-4H-pyridazin[4,5-b]indol-4-one.

[0231]

[0232] Benzyl bromide (51 mg, 0.30 mmol) was added to a solution of 5,7-dimethyl-3,5-dihydro-4H-pyridazino[4,5-b]indol-4-one (53 mg, 0.25 mmol) and KOtBu (35 mg, 0.29 mmol) in DMF (5 mL) at 0 °C. The reaction mixture was warmed to room temperature and stirred overnight. Water was added and the reaction mixture was extracted with DCM (3 × 10 mL). The organic compounds were combined, dried, concentrated, and purified by rapid column chromatography (0-100% EtOAc in hexane) to provide the desired product (45 mg, 61%). 1 H NMR (400MHz, DMSO-d6) δ8.78 (s, 1H), 8.08 (d, J = 8.0Hz, 1H), 7.57 (s, 1H), 7. 36–7.31(m,4H),7.31–7.22(m,2H),5.41(s,2H),4.26(s,3H),2.54(s,3H).

[0233] Example 2: 3-(4-methoxybenzyl)-5,7-dimethyl-3,5-dihydro-4H-pyridazin[4,5-b]indol-4-one.

[0234]

[0235] Example 2 was prepared in a manner similar to that of Example 1. 1 H NMR(400MHz,DMSO-d6)δ8.76(s,1H),8.07(d,J=8.2Hz,1H),7.56(s,1H),7.35–7.28(m,2H),7.2 4(dd,J=8.2,1.4Hz,1H),6.93–6.86(m,2H),5.33(s,2H),4.26(s,3H),3.72(s,3H),2.54(s,3H).

[0236] Intermediate D: tert-butyl (3-((5,7-dimethyl-4-oxoyne-4,5-dihydro-3H-pyridazino[4,5-b]indol-3-yl)methyl)phenyl)carbamate).

[0237]

[0238] Intermediate D was prepared in a manner similar to that of Example 1. 1H NMR(400MHz, DMSO-d6)δ9.31(s,1H),8.79(s,1H),8.09(d,J=8.2Hz,1H),7.57(s,1H),7.44–7.38(m,1H),7.39–7.31( m,1H),7.26–7.22(m,1H),7.22–7.16(m,1H),6.95–6.89(m,1H),5.34(s,2H),4.26(s,3H),2.54(s,3H),1.44(s,9H).

[0239] Example 3: 3-(3-aminobenzyl)-5,7-dimethyl-3,5-dihydro-4H-pyridazin[4,5-b]indol-4-one.

[0240]

[0241] To a solution of tert-butyl (3-((5,7-dimethyl-4-oxoylide-4,5-dihydro-3H-pyridazino[4,5-b]indol-3-yl)methyl)phenyl)carbamate (53 mg, 0.13 mmol) in DCM (5 mL), HCl (1 mL, 4 M HCl in dioxane) was added and the reaction mixture was stirred at room temperature for 1 hour. The reaction mixture was diluted with DCM (10 mL), washed with saturated sodium bicarbonate aqueous solution (2 × 5 mL), dried, concentrated, and purified by hplc to provide the desired product (24 mg, 60%).

[0242] Intermediate E: Methyl 3-formyl-6-(methylthio)-1H-indole-2-carboxylic acid.

[0243]

[0244] Intermediate E is generated in a similar manner to intermediate A. 1 H NMR(400MHz,DMSO-d6)δ12.79(s,1H),10.58(s,1H),8.15(d,J=8.5Hz,1H),7 .36(d,J=1.6Hz,1H),7.23(dd,J=8.6,1.7Hz,1H),3.99(s,3H),2.54(s,3H).

[0245] Intermediate F: Methyl 3-formyl-1-methyl-6-(methylthio)-1H-indole-2-carboxylic acid ester.

[0246]

[0247] Intermediate F is generated in a similar manner to intermediate B. 1H NMR (400MHz, DMSO-d6) δ10.41(s,1H),8.19(dd,J=8.5,0.6Hz,1H),7.57(d,J=1 .5Hz,1H),7.27(dd,J=8.5,1.6Hz,1H),4.05(s,3H),4.00(s,3H),2.60(s,3H).

[0248] Intermediate G: 5-methyl-7-(meththio)-3,5-dihydro-4H-pyridazin[4,5-b]indol-4-one.

[0249]

[0250] Intermediate G was prepared in a manner similar to that used for intermediate C. 1 H NMR(400MHz,DMSO-d6)δ12.90–12.71(m,1H),8.72(s,1H),8.11(dd,J=8.4,0.6Hz,1H ),7.57(dd,J=1.7,0.7Hz,1H),7.29(dd,J=8.4,1.6Hz,1H),4.27(s,3H),2.63(s,3H).

[0251] Example 4: 3-Benzyl-5-methyl-7-(methylthio)-3,5-dihydro-4H-pyridazin[4,5-b]indol-4-one.

[0252]

[0253] Example 4 was prepared in a manner similar to that of Example 1. 1 H NMR(400MHz,DMSO-d6)δ8.79(s,1H),8.11(dd,J=8.4,0.6Hz,1H),7.57(dd,J =1.6,0.6Hz,1H),7.36–7.25(m,6H),5.41(s,2H),4.28(s,3H),2.63(s,3H).

[0254] Example 5: 3-Benzyl-5-methyl-7-(methylsulfinyl)-3,5-dihydro-4H-pyridazino[4,5-b]indol-4-one.

[0255]

[0256] Add 3-chloroperoxybenzoic acid (13.9 mg, 0.081 mmol) to a solution of 3-benzyl-5-methyl-7-(methylthio)-3,5-dihydro-4H-pyridazino[4,5-b]indol-4-one (27.0 mg, 0.081 mmol) in dichloromethane (10 mL). Stir the reaction mixture overnight at room temperature. Dilute the reaction mixture with DCM (20 mL), extract with saturated sodium bicarbonate aqueous solution (3 × 10 mL), dry, concentrate, and purify by rapid column chromatography (0-100% EtOAc in hexane) to provide the desired product (10 mg, 53%). 1 H NMR (400MHz, DMSO-d6) δ8.89(s,1H),8.41(dd,J=8.4,0.7Hz,1H),8.11(dd,J=1.4,0.7Hz,1H),7.67( dd,J=8.3,1.4Hz,1H),7.37–7.31(m,4H),7.31–7.25(m,1H),5.43(s,2H),4.35(s,3H),2.85(s,3H).

[0257] Intermediate H: (3-((5-methyl-7-(methylthio)-4-oxonyl-4,5-dihydro-3H-pyridazino[4,5-b]indol-3-yl)methyl)phenyl)tert-butyl carbamate.

[0258]

[0259] Intermediate H was prepared from 5-methyl-7-(methylthio)-3,5-dihydro-4H-pyridazino[4,5-b]indol-4-one in a manner similar to that in Example 1. 1 H NMR(400MHz,DMSO-d6)δ9.31(s,1H),8.79(s,1H),8.15–8.10(m,1H),7.59–7.54(m,1H),7.44–7.39(m,1H),7.33–7.26 (m,1H),7.22–7.16(m,1H),7.17–7.07(m,1H),6.94–6.89(m,1H),5.34(s,2H),4.27(s,3H),2.63(s,3H),1.45(s,9H).

[0260] Example 6: 3-Benzyl-5-methyl-7-(methylthio)-3,5-dihydro-4H-pyridazin[4,5-b]indol-4-one.

[0261]

[0262] Example 6 was prepared from tert-butyl (3-((5-methyl-7-(methylthio)-4-oxonyl-4,5-dihydro-3H-pyridazino[4,5-b]indol-3-yl)methyl)phenyl)carbamate in a manner similar to that of Example 3. 1 HNMR (400MHz, DMSO-d6) δ8.77(s,1H),8.12(dd,J=8.4,0.6Hz,1H),7.57(d,J=1.6Hz,1H),7.30(dd,J=8. 4,1.6Hz,1H),7.03–6.90(m,1H),6.54–6.39(m,3H),5.24(s,2H),5.05(s,2H),4.28(s,3H),2.63(s,3H).

[0263] Intermediate I: (3-((5-methyl-7-(methylsulfinyl)-4-oxonyl-4,5-dihydro-3H-pyridazino[4,5-b]indol-3-yl)methyl)phenyl)tert-butyl carbamate.

[0264]

[0265] Intermediate I was generated from tert-butyl (3-((5-methyl-7-(methylthio)-4-oxonyl-4,5-dihydro-3H-pyridazino[4,5-b]indol-3-yl)methyl)phenyl)carbamate in a manner similar to that in Example 5. 1 HNMR(400MHz,DMSO-d6)δ9.31(s,1H),8.90(s,1H),8.45–8.39(m,1H),8.14–8.08(m,1H),7.71–7.64(m,1H),7.44–7.3 9(m,1H),7.39–7.33(m,1H),7.23–7.17(m,1H),6.96–6.89(m,1H),5.36(s,2H),4.34(s,3H),2.86(s,3H),1.45(s,9H).

[0266] Example 7: 3-(3-aminobenzyl)-5-methyl-7-(methylsulfinyl)-3,5-dihydro-4H-pyridazino[4,5-b]indol-4-one.

[0267]

[0268] Example 7 was produced from (3-((5-methyl-7-(methylsulfinyl)-4-oxonyl-4,5-dihydro-3H-pyridazino[4,5-b]indol-3-yl)methyl)phenyl)carbamate tert-butyl ester in a manner similar to that of Example 3. 1H NMR (400MHz, DMSO-d6) δ8.91(s,1H),8.42(d,J=8.3Hz,1H),8.12(d,J=1.3Hz,1H),7.73–7.64(m,1H ),7.28–7.20(m,1H),7.03–6.94(m,1H),6.93–6.83(m,2H),5.38(s,2H),4.35(s,3H),2.85(s,3H).

[0269] Intermediate J: Methyl 3-formyl-6-methoxy-1H-indole-2-carboxylate.

[0270]

[0271] Intermediate J was generated from commercially available methyl 6-methoxy-1h-indole-2-carboxylate in a manner similar to that of intermediate A. 1 H NMR (400MHz, DMSO-d6) δ12.67(s,1H),10.58(s,1H),8.13–8.08(m,1H),7.00–6.93(m,2H),3.98(s,3H),3.82(s,3H).

[0272] Intermediate K: Methyl 3-formyl-6-methoxy-1-methyl-1H-indole-2-carboxylate.

[0273]

[0274] Intermediate K is generated from methyl 3-formyl-6-methoxy-1H-indole-2-carboxylate in a manner similar to that of intermediate B. 1 H NMR(400MHz,DMSO-d6)δ10.42(s,1H),8.16(d,J=8.8Hz,1H),7.27–7.19(m,1H) ,7.00(dd,J=8.9,2.3Hz,1H),4.03(d,J=1.4Hz,3H),3.99(s,3H),3.88(s,3H).

[0275] Intermediate L: 7-methoxy-5-methyl-3,5-dihydro-4H-pyridazin[4,5-b]indol-4-one.

[0276]

[0277] Intermediate L was generated from methyl 3-formyl-6-methoxy-1-methyl-1H-indole-2-carboxylate in a similar manner to intermediate C. ¹H NMR (400 MHz, DMSO-d6) δ 12.69 (s, 1H), 8.67 (s, 1H), 8.06 (d, J = 8.8 Hz, 1H), 7.23 (d, J = 2.2 Hz, 1H), 7.01 (dd, J = 8.7, 2.2 Hz, 1H), 4.24 (s, 3H), 3.91 (s, 3H).

[0278] Intermediate M: tert-butyl (3-((7-methoxy-5-methyl-4-oxo-4,5-dihydro-3H-pyridazino[4,5-b]indol-3-yl)methyl)phenyl)carbamate).

[0279]

[0280] Intermediate M was generated from 7-methoxy-5-methyl-3,5-dihydro-4H-pyridazino[4,5-b]indole-4-one in a manner similar to that in Example 1. 1 H NMR (400MHz, DMSO-d6) δ9.29(s,1H),8.75(s,1H),8.08(d,J=8.8Hz,1H),7.44–7.31(m,2H),7.25(d,J=2.2Hz,1H),7. 22–7.15(m,1H),7.02(dd,J=8.8,2.2Hz,1H),6.96–6.85(m,1H),5.34(s,2H),4.26(s,3H),3.92(s,3H),1.44(s,9H).

[0281] Example 8: 3-(3-aminobenzyl)-7-methoxy-5-methyl-3,5-dihydro-4H-pyridazin[4,5-b]indol-4-one.

[0282]

[0283] Example 8 was produced from (3-((7-methoxy-5-methyl-4-oxo-4,5-dihydro-3H-pyridazino[4,5-b]indol-3-yl)methyl)phenyl)carbamate tert-butyl ester in a manner similar to that of Example 3. 1HNMR (400MHz, DMSO-d6) δ8.73(s,1H),8.08(d,J=8.8Hz,1H),7.24(d,J=2.2Hz,1H),7.02(dd,J=8.8,2 .2Hz,1H),6.99–6.89(m,1H),6.51–6.41(m,3H),5.24(s,2H),5.08(s,2H),4.26(s,3H),3.92(s,3H).

[0284] Example 9: 7-methoxy-5-methyl-3-((4-oxoylide-4H-pyrido[1,2-a]pyrimidin-2-yl)methyl)-3,5-dihydro-4H-pyridazino[4,5-b]indol-4-one.

[0285]

[0286] Example 9 was generated from 7-methoxy-5-methyl-3,5-dihydro-4H-pyridazino[4,5-b]indol-4-one in a manner similar to that of Example 3. 1 H NMR (400MHz, DMSO-d6) δ8.96–8.92(m,1H),8.82(s,1H),8.11(d,J=8.8Hz,1H),8.02–7.94(m,1H),7.70–7.64(m,1H) ,7.40–7.33(m,1H),7.29–7.25(m,1H),7.08–7.02(m,1H),5.94(s,1H),5.44–5.35(m,2H),4.26(s,3H),3.93(s,3H).

[0287] Example 10: 3-(imidazo[1,2-a]pyrimidin-2-ylmethyl)-7-methoxy-5-methyl-3,5-dihydro-4H-pyridazin[4,5-b]indol-4-one.

[0288]

[0289] Example 10 was generated from 7-methoxy-5-methyl-3,5-dihydro-4H-pyridazino[4,5-b]indole-4-one in a manner similar to that of Example 3. 1H NMR (400MHz, DMSO-d6) δ8.87(dd,J=6.8,2.0Hz,1H),8.77(s,1H),8.49(dd,J=4.1,2.0Hz,1H),8.10(d,J=8.8Hz,1 H),7.69(d,J=0.8Hz,1H),7.26(d,J=2.2Hz,1H),7.07–6.99(m,2H),5.60–5.50(m,2H),4.27(s,3H),3.92(s,3H).

[0290] Example 11: 3-(imidazo[1,2-a]pyridin-2-ylmethyl)-7-methoxy-5-methyl-3,5-dihydro-4H-pyridazin[4,5-b]indol-4-one.

[0291]

[0292] Example 11 was generated from 7-methoxy-5-methyl-3,5-dihydro-4H-pyridazino[4,5-b]indole-4-one in a manner similar to that of Example 3. 1 H NMR (400MHz, DMSO-d6) δ8.74(s,1H),8.49–8.43(m,1H),8.08(dd,J=8.8,0.5Hz,1H),7.73(d,J=0.8Hz,1H),7.51–7.44(m,1H),7.25(d, J=2.1Hz,1H),7.20(ddd,J=9.1,6.7,1.3Hz,1H),7.02(dd,J=8.7,2.2Hz,1H),6.90–6.81(m,1H),5.50(s,2H),4.27(s,3H),3.92(s,3H).

[0293] Example 12: 7-Methoxy-5-methyl-3-(quinolin-6-ylmethyl)-3,5-dihydro-4H-pyridazin[4,5-b]indol-4-one.

[0294]

[0295] Example 12 was generated from 7-methoxy-5-methyl-3,5-dihydro-4H-pyridazino[4,5-b]indole-4-one in a manner similar to that of Example 3. 1H NMR (400MHz, DMSO-d6) δ8.87(dd,J=4.2,1.7Hz,1H),8.80(s,1H),8.37–8.32(m,1H),8.09(d,J=8.8Hz,1H),8.00(d,J=8.7Hz,1H),7.89–7.83(m,1H) ,7.76(dd,J=8.7,2.0Hz,1H),7.51(dd,J=8.3,4.2Hz,1H),7.25(d,J=2.2H z, 1H), 7.03 (dd, J = 8.7, 2.2Hz, 1H), 5.61 (s, 2H), 4.27 (s, 3H), 3.92 (s, 3H).

[0296] Example 13: 7-Methoxy-5-methyl-3-((5-methyl-1,3,4-oxadiazol-2-yl)methyl)-3,5-dihydro-4H-pyridazin[4,5-b]indol-4-one.

[0297]

[0298] Example 13 was generated from 7-methoxy-5-methyl-3,5-dihydro-4H-pyridazino[4,5-b]indole-4-one in a manner similar to that of Example 3. 1 H NMR(400MHz,DMSO-d6)δ8.78(s,1H),8.13–8.04(m,1H),7.26(d,J=2.1Hz,1H),7 .04(dd,J=8.8,2.2Hz,1H),5.62(s,2H),4.24(s,3H),3.92(s,3H),2.47(s,3H).

[0299] Example 14: 7-Methoxy-5-methyl-3-(pyrimidin-2-ylmethyl)-3,5-dihydro-4H-pyridazin[4,5-b]indol-4-one.

[0300]

[0301] Example 14 was generated from 7-methoxy-5-methyl-3,5-dihydro-4H-pyridazino[4,5-b]indole-4-one in a manner similar to that of Example 3. 1 H NMR (400MHz, DMSO-d6) δ8.80–8.71(m,3H),8.10(d,J=8.8Hz,1H),7.41(t,J=4.9Hz,1H),7 .26(d,J=2.2Hz,1H),7.04(dd,J=8.8,2.2Hz,1H),5.60(s,2H),4.24(s,3H),3.92(s,3H).

[0302] Example 15: 3-(3-aminobenzyl)-7-hydroxy-5-methyl-3,5-dihydro-4H-pyridazin[4,5-b]indol-4-one.

[0303]

[0304] At 0 °C, 1 M tribromoborane in 1.00 mL of DCM was added to a solution of tert-butyl (3-((7-methoxy-5-methyl-4-oxoylide-4,5-dihydro-3H-pyridazino[4,5-b]indol-3-yl)methyl)phenyl)carbamate (76.4 mg, 0.176 mmol) in 10 mL of DCM, and the reaction mixture was warmed to room temperature for 3 hours. A saturated aqueous solution of sodium bicarbonate (10 mL) was added, and the resulting reaction mixture was extracted with DCM. The organic layers were combined, dried, concentrated, and purified by rapid column chromatography (0-20% MeOH in DCM) to provide the desired product (32 mg, 57%). 1 H NMR (400MHz, DMSO-d6) δ9.95 (s, 1H), 8.68 (s, 1H), 7.98 (d, J = 8.6Hz, 1H), 7. 00–6.85(m,3H),6.50–6.38(m,2H),5.23(s,2H),5.02(s,2H),4.18(s,3H).

[0305] Intermediate N: ethyl 2-methyl-6H-thieno[2,3-b]pyrrole-5-carboxylate.

[0306]

[0307] Ethyl 2-azidoethyl (1.33 g, 10.3 mmol) was added to a solution of 5-methylthiophene-3-carboxaldehyde (1.00 g, 7.93 mmol) in ethanol (50 mL) at 0 °C, followed by dropwise addition of sodium ethoxide (10.3 mmol) in ethanol. The reaction mixture was warmed to room temperature and stirred overnight. A saturated aqueous solution of ammonium chloride (50 mL) was added, and the resulting reaction mixture was extracted with DCM (3 × 20 mL). The organic matter was dried, concentrated, dissolved in toluene, and refluxed for 1.5 h. The reaction mixture was concentrated and purified by rapid column chromatography (0-30% EtOAc in hexane) to provide the desired product. 1 H NMR(400MHz,DMSO-d6)δ12.10(s,1H),6.86(d,J=2.0Hz,1H),6.71(dt,J=1.6,1 .2Hz, 1H), 4.26 (q, J = 7.1Hz, 2H), 2.45 (d, J = 1.3Hz, 3H), 1.30 (t, J = 7.1Hz, 3H).

[0308] Intermediate O: ethyl 4-formyl-2-methyl-6H-thieno[2,3-b]pyrrole-5-carboxylate.

[0309]

[0310] Intermediate O was generated from ethyl 2-methyl-6H-thiopheno[2,3-b]pyrrole-5-carboxylate in a manner similar to that of intermediate A. 1 H NMR (400MHz, DMSO-d6) δ12.97(s,1H),10.42(s,1H),7.11–6.97(m,1H),4.39(q,J=7.1Hz,2H),3.30(s,3H),1.37(t,J=7.1Hz,3H).

[0311] Intermediate P: ethyl 4-formyl-2,6-dimethyl-6H-thieno[2,3-b]pyrrole-5-carboxylate.

[0312]

[0313] Intermediate P was generated from ethyl 4-formyl-2-methyl-6H-thieno[2,3-b]pyrrole-5-carboxylate in a manner similar to that of intermediate B. 1 H NMR (400MHz, DMSO-d6) δ10.35(s,1H),7.11–7.07(m,1H),4.40(q,J=7.1Hz,2H),3.99(s,3H),2.56–2.53(m,3H),1.37(t,J=7.1Hz,3H).

[0314] Intermediate Q: 2,8-dimethyl-6,8-dihydro-7H-thieno[3',2':4,5]pyrrolo[2,3-d]pyridazin-7-one.

[0315]

[0316] Intermediate Q was generated from ethyl 4-formyl-2,6-dimethyl-6H-thieno[2,3-b]pyrrole-5-carboxylate in a manner similar to that of intermediate C. 1 H NMR (400MHz, DMSO-d6) δ12.52(s,1H),8.40(s,1H),7.15(s,1H),4.18(s,3H),2.57(s,3H).

[0317] Example 16: 6-(2-fluorobenzyl)-2,8-dimethyl-6,8-dihydro-7H-thieno[3',2':4,5]pyrrolo[2,3-d]pyridazin-7-one.

[0318]

[0319] Example 16 was produced from 2,8-dimethyl-6,8-dihydro-7H-thieno[3',2':4,5]pyrrolo[2,3-d]pyridazine-7-one in a manner similar to that of Example 1. 1 H NMR (400MHz, DMSO-d6) δ8.49(s,1H),7.38–7.30(m,1H),7.26–7.11(m,4H),5.42(s,2H),4.20(s,3H),2.58(s,3H).

[0320] Intermediate R: ethyl 3-formyl-1H-pyrrolo[2,3-b]pyridine-2-carboxylate.

[0321]

[0322] Hexamethylenetetramine (812 mg, 5.78 mmol) was added to a solution of 1H-pyrrolo[2,3-B]pyridine-2-carboxylate (1.00 g, 5.25 mmol) in acetic acid (3 mL) and water (6 mL), and the resulting reaction mixture was heated to 120 °C for 9 hours. The reaction mixture was cooled to room temperature and its volume was halved. The reaction mixture was cooled to 0 °C and the resulting precipitate was collected by filtration to provide the desired product (52.2 mg, 5%). 1 H NMR (400MHz, DMSO-d6) δ13.44(s,1H),10.59(s,1H),8.59(dd,J=8.0,1.7Hz,1H),8.55(dd,J =4.6, 1.7Hz, 1H), 7.38 (dd, J = 8.0, 4.6Hz, 1H), 4.47 (q, J = 7.1Hz, 2H), 1.41 (t, J = 7.1Hz, 3H).

[0323] Intermediate S: ethyl 3-formyl-1-methyl-1H-pyrrolo[2,3-b]pyridine-2-carboxylate.

[0324]

[0325] Intermediate S was generated from ethyl 3-formyl-1H-pyrrolo[2,3-b]pyridine-2-carboxylate in a manner similar to that of intermediate B. 1H NMR (400MHz, chloroform-d) δ10.64(s,1H),8.78(dd,J=8.0,1.7Hz,1H),8.58(dd,J=4.6,1.7Hz,1H ), 7.33 (dd, J = 8.0, 4.6Hz, 1H), 4.57 (q, J = 7.2Hz, 2H), 4.22 (s, 4H), 1.51 (t, J = 7.1Hz, 4H).

[0326] Intermediate T: 9-methyl-7,9-dihydro-8H-pyrido[3',2':4,5]pyrrolo[2,3-d]pyridazin-8-one.

[0327]

[0328] Intermediate T is generated from ethyl 3-formyl-1-methyl-1H-pyrrolo[2,3-b]pyridine-2-carboxylate in a manner similar to that of intermediate C.

[0329] Example 17: 7-(2-fluorobenzyl)-9-methyl-7,9-dihydro-8H-pyrido[3',2':4,5]pyrrolo[2,3-d]pyridazin-8-one.

[0330]

[0331] Example 17 was generated from 9-methyl-7,9-dihydro-8H-pyrido[3',2':4,5]pyrrolo[2,3-d]pyridazin-8-one in a manner similar to that of Example 1. 1 H NMR (400MHz, DMSO-d6) δ8.86(d,J=1.6Hz,1H),8.73–8.64(m,2H),7.55–7.45(m,1H),7.40–7.24(m,5H),5.44(s,2H),4.29(d,J=1.5Hz,3H).

[0332] Intermediate U: Ethyl 6-methyl-1H-pyrrolo[2,3-b]pyridine-2-carboxylate

[0333]

[0334] Intermediate U is generated from commercially available 3-pyridinecarboxaldehyde in a manner similar to that of intermediate N. 1 ¹H NMR (400MHz, methanol-d⁴) δ 8.01 (d, J = 8.1Hz, 1H), 7.16 (s, 1H), 7.09 (d, J = 8.2Hz, 1H), 4.42 (q, J = 7.1Hz, 2H), 2.63 (s, 3H), 1.43 (t, J = 7.1Hz, 3H).

[0335] Intermediate V: ethyl 3-bromo-6-methyl-1H-pyrrolo[2,3-b]pyridine-2-carboxylate.

[0336]

[0337] N-bromosuccinimide (94 mg, 525 mmol) was added to a solution of ethyl 6-methyl-1H-pyrrolo[2,3-b]pyridine-2-carboxylate (102 mg, 500 mmol) in DCM (10 mL). The reaction mixture was stirred overnight at room temperature. A saturated aqueous solution of sodium bicarbonate was added and the resulting reaction mixture was extracted with DCM (3 × 10 mL). The organic layers were combined, dried, concentrated, and purified by rapid column chromatography (0-100% EtOAc in hexane) to provide the desired product (96 mg, 68%). 1 HNMR (400MHz, DMSO-d6) δ12.67(s,1H),7.88(dd,J=8.2,0.8Hz,1H),7.17(d,J=8.2Hz,1H),4.37(q,J=7.1Hz,2H),2.60(s,3H),1.37(t,J=7.1Hz,3H).

[0338] Intermediate W: ethyl 3-bromo-1,6-dimethyl-1H-pyrrolo[2,3-b]pyridine-2-carboxylate.

[0339]

[0340] Intermediate W was generated from ethyl 3-bromo-6-methyl-1H-pyrrolo[2,3-b]pyridine-2-carboxylate in a manner similar to that of intermediate B. 1 ¹H NMR (400MHz, methanol-d⁴) δ 7.91 (d, J = 8.2 Hz, 1H), 7.18 (d, J = 8.1 Hz, 1H), 4.47 (q, J = 7.1 Hz, 2H), 4.10 (s, 3H), 2.68 (s, 3H), 1.47 (t, J = 7.1 Hz, 3H).

[0341] Intermediate X: ethyl 1,6-dimethyl-3-vinyl-1H-pyrrolo[2,3-b]pyridine-2-carboxylate.

[0342]

[0343] To a solution of ethyl 3-bromo-1,6-dimethyl-1H-pyrrolo[2,3-b]pyridine-2-carboxylate (71 mg, 0.24 mmol) in DMF, tributyl(vinyl)stanane (83 mg, 0.26 mmol) and tetrakis(triphenylphosphine)palladium(O) (27 mg, 23 mmol) were added, and nitrogen was bubbled through the reaction mixture for 15 minutes. The reaction mixture was heated to 100 °C overnight. The reaction mixture was cooled to room temperature and water was added. The resulting reaction mixture was extracted with EtOAc (3 × 15 mL), and the organic layers were combined, dried, concentrated, and purified by FCC (0-100% EtOAc in hexane) to provide the desired product (57 mg, 99%). 1 H NMR (400MHz, methanol-d4) δ8.27(d,J=8.3Hz,1H),7.43(dd,J=18.0,11.5Hz,1H),7.13(d,J=8.2Hz,1H),5.83(dd, J=18.0,1.5Hz,1H),5.43(dd,J=11.5,1.5Hz,1H),4.46(q,J=7.1Hz,2H),4.07(s,3H),1.46(t,J=7.1Hz,3H).

[0344] Intermediate Y: 2,9-dimethyl-7,9-dihydro-8H-pyrido[3',2':4,5]pyrrolo[2,3-d]pyridazin-8-one.

[0345]

[0346] Ozone was gently bubbled through a solution of ethyl 1,6-dimethyl-3-vinyl-1H-pyrrolo[2,3-b]pyridine-2-carboxylate (71 mg, 0.29 mmol) in DCM (10 mL) cooled to -78 °C. When the starting material was indicated to have been consumed by TLC, triphenylphosphine (303 mg, 1.15 mmol) was added, and the reaction mixture was warmed to room temperature and stirred for 4 hours. The reaction mixture was concentrated, dissolved in ethoxyethanol (5 mL), and hydrazine (116 mg, 1.15 mmol) was added. The reaction mixture was heated to 135 °C and stirred overnight. The reaction mixture was cooled to room temperature, water was added, and the resulting reaction mixture was extracted with EtOAc (3 × 10 mL). The organic layers were combined, dried, concentrated, and purified by rapid column chromatography (0-100% EtOAc in hexane) to provide the desired product (45 mg, 73%). 1H NMR (400MHz, DMSO-d6) δ12.87(s,1H),8.73(s,1H),8.52(d,J=8.1Hz,1H),7.34(d,J=8.1Hz,1H),4.23(s,3H),2.68(s,3H).

[0347] Example 18: 7-(3-fluorobenzyl)-2,9-dimethyl-7,9-dihydro-8H-pyrido[3',2':4,5]pyrrolo[2,3-d]pyridazin-8-one.

[0348]

[0349] Example 18 was produced from 2,9-dimethyl-7,9-dihydro-8H-pyrido[3',2':4,5]pyrrolo[2,3-d]pyridazin-8-one in a manner similar to that of Example 1. 1 H NMR (400MHz, DMSO-d6) δ8.81 (s, 1H), 8.53 (d, J = 8.1Hz, 1H), 7.44–7.31 (m, 2H), 7.2 8–7.19(m,2H),7.15(td,J=7.4,1.2Hz,1H),5.48(s,2H),4.24(s,3H),2.69(s,3H).

[0350] Intermediate Z: Methyl 5-fluoro-3-formyl-6-methoxy-1H-indole-2-carboxylate

[0351]

[0352] Intermediate Z was generated from commercially available starting materials in a similar manner to intermediate A. ¹H NMR (400 MHz, DMSO-d6) δ 12.85 (s, ¹H), 10.55 (s, ¹H), 7.89 (d, J = 11.6 Hz, ¹H), 7.12 (d, J = 7.4 Hz, ¹H), 3.98 (s, ³H), 3.91 (s, ³H).

[0353] Intermediate AA: Methyl 5-fluoro-3-formyl-6-methoxy-1-methyl-1H-indole-2-carboxylate

[0354]

[0355] Intermediate AA was generated in a similar manner to intermediate B. ¹H NMR (400 MHz, DMSO-d6) δ 10.38 (s, ¹H), 7.93 (d, J = 11.5 Hz, ¹H), 7.45 (d, J = 7.3 Hz, ¹H), 4.05 (s, 3H), 3.98 (s, 3H), 3.96 (s, 3H).

[0356] Intermediate AB: 8-fluoro-7-methoxy-5-methyl-3,5-dihydro-4H-pyridazin[4,5-b]indol-4-one

[0357]

[0358] Intermediate AB was generated in a similar manner to intermediate C. ¹H NMR (400 MHz, DMSO-d6) δ 12.72 (s, ¹H), 8.65 (s, ¹H), 8.06 (d, J = 11.3 Hz, ¹H), 7.46 (d, J = 7.2 Hz, ¹H), 4.26 (s, 3H), 4.00 (s, 3H).

[0359] Example 19: (3-((8-fluoro-7-methoxy-5-methyl-4-oxoylide-4,5-dihydro-3H-pyridazin[4,5-b]indol-3-yl)methyl)phenyl)tert-butyl carbamate

[0360]

[0361] Example 19 is generated in a manner similar to that of intermediate D.

[0362] Example 20: 3-(3-aminobenzyl)-8-fluoro-7-methoxy-5-methyl-3,5-dihydro-4H-pyridazin[4,5-b]indol-4-one

[0363]

[0364] Example 20 was generated in a manner similar to that of Example 3. ¹H NMR (400MHz, DMSO-d6) δ 8.71 (s, ¹H), 8.08 (d, J = 11.3Hz, ¹H), 7.48 (d, J = 7.2Hz, ¹H), 6.94 (t, J = 7.9Hz, ¹H), 6.50–6.40 (m, ³H), 5.23 (s, ²H), 5.04 (s, ²H), 4.28 (s, ³H), 4.01 (s, ³H).

[0365] Example 21: 3-(3-aminobenzyl)-8-fluoro-7-hydroxy-5-methyl-3,5-dihydro-4H-pyridazin[4,5-b]indol-4-one

[0366]

[0367] Example 21 was generated in a manner similar to that of Example 15. ¹H NMR (400 MHz, methanol-d⁴) δ 8.54 (s, ¹H), 7.68 (d, J = 10.8 Hz, ¹H), 7.10–7.02 (m, ¹H), 6.95 (d, J = 7.4 Hz, ¹H), 6.69 (dt, J = 8.7, 1.6 Hz, 2H), 6.63 (ddd, J = 7.9, 2.2, 1.1 Hz, 1H), 5.51 (s, 3H), 5.40–5.33 (m, 2H), 4.20 (s, 2H), 3.37 (s, ¹H).

[0368] Intermediate AC: Ethyl 3-formyl-7-methoxy-1H-indole-2-carboxylate

[0369]

[0370] Intermediate AC was generated in a similar manner to intermediate A. ¹H NMR (400 MHz, DMSO-d⁶) δ 12.92 (s, ¹H), 10.59 (s, ¹H), 7.85–7.74 (m, ¹H), 7.23 (t, J = 8.0 Hz, ¹H), 6.95 (dd, J = 7.8, 0.8 Hz, ¹H), 4.43 (q, J = 7.1 Hz, 2H), 3.96 (s, 3H), 1.40 (t, J = 7.1 Hz, 3H).

[0371] Intermediate AD: Ethyl 3-formyl-7-methoxy-1-methyl-1H-indole-2-carboxylate

[0372]

[0373] Intermediate AD was generated in a similar manner to intermediate B. ¹H NMR (400 MHz, DMSO-d⁶) δ 10.36 (s, ¹H), 7.89 (dd, J = 8.1, 0.9 Hz, ¹H), 7.25 (t, J = 8.0 Hz, ¹H), 6.98 (dd, J = 7.9, 0.9 Hz, ¹H), 4.47 (q, J = 7.1 Hz, 2H), 4.27 (s, 3H), 3.95 (s, 3H), 1.42–1.36 (m, 3H).

[0374] Intermediate AE: 6-methoxy-5-methyl-3,5-dihydro-4H-pyridazin[4,5-b]indol-4-one

[0375]

[0376] Intermediate AE was generated in a similar manner to intermediate C. ¹H NMR (400 MHz, DMSO-d6) δ 12.75 (s, ¹H), 8.71 (s, ¹H), 7.74 (dd, J = 8.0, 0.9 Hz, ¹H), 7.27 (t, J = 7.9 Hz, ¹H), 7.09 (dd, J = 7.9, 0.9 Hz, ¹H), 4.53 (s, ³H), 3.97 (s, ³H).

[0377] Example 22: tert-butyl (3-((6-methoxy-5-methyl-4-oxoylide-4,5-dihydro-3H-pyridazin[4,5-b]indol-3-yl)methyl)phenyl)carbamate

[0378]

[0379] Example 22 was generated in a manner similar to that of intermediate D. ¹H NMR (400MHz, DMSO-d6) δ 9.31 (s, 1H), 8.79 (s, 1H), 7.76 (dd, J = 8.1, 0.9Hz, 1H), 7.39 (s, 1H), 7.36 (d, J = 8.3Hz, 1H), 7.28 (t, J = 7.9Hz, 1H), 7.19 (t, J = 7.8Hz, 1H), 7.12–7.08 (m, 1H), 6.91 (d, J = 7.7Hz, 1H), 5.33 (s, 2H), 4.54 (s, 3H), 3.97 (s, 3H), 1.44 (s, 9H).

[0380] Example 23: 3-(3-aminobenzyl)-6-methoxy-5-methyl-3,5-dihydro-4H-pyridazin[4,5-b]indol-4-one

[0381]

[0382] Example 23 was generated in a manner similar to that of Example 3. ¹H NMR (400MHz, DMSO-d6) δ 8.77 (s, ¹H), 7.75 (dd, J = 8.1, 0.9Hz, ¹H), 7.28 (t, J = 7.9Hz, ¹H), 7.09 (dd, J = 7.9, 0.9Hz, ¹H), 6.94 (t, J = 7.9Hz, ¹H), 6.50–6.39 (m, ³H), 5.24 (s, 2H), 5.05 (s, 2H), 4.55 (s, 3H), 3.97 (s, 3H).

[0383] Intermediate AF: Methyl 6-bromo-2-methyl-4H-pyrrolo[3,2-d]thiazole-5-carboxylate

[0384]

[0385] Intermediate AF was generated in a manner similar to that of intermediate V. ¹H NMR (400 MHz, DMSO-d6) δ 12.67 (s, ¹H), 3.84 (s, ³H), 2.72 (s, ³H).

[0386] Intermediate AG: Methyl 6-bromo-2,4-dimethyl-4H-pyrrolo[3,2-d]thiazole-5-carboxylate

[0387]

[0388] Intermediate AG was generated in a similar manner to intermediate W. ¹H NMR (400 MHz, DMSO-d6) δ 3.98 (s, 3H), 3.84 (s, 3H), 2.74 (s, 3H).

[0389] Intermediate AH: Methyl 2,4-dimethyl-6-vinyl-4H-pyrrolo[3,2-d]thiazole-5-carboxylate

[0390]

[0391] Intermediate AH was generated in a manner similar to that of intermediate X. ¹H NMR (400 MHz, DMSO-d6) δ 7.30 (dd, J = 17.6, 11.2 Hz, ¹H), 6.43 (dd, J = 17.6, 2.4 Hz, ¹H), 5.43 (dd, J = 11.1, 2.4 Hz, ¹H), 3.94 (s, 3H), 3.85 (s, 3H), 2.76 (s, 3H).

[0392] Intermediate AI: 2,4-dimethyl-4,6-dihydro-5H-thiazo[4',5':4,5]pyrrolo[2,3-d]pyridazin-5-one

[0393]

[0394] Intermediate AI was generated in a similar manner to intermediate C. ¹H NMR (400MHz, DMSO-d6) δ 12.65 (s, ¹H), 8.45 (s, ¹H), 4.26 (s, ³H), 2.82 (s, ³H).

[0395] Example 24: (3-((2,4-dimethyl-5-oxoyne-4,5-dihydro-6H-thiazo[4',5':4,5]pyrrolo[2,3-d]pyridazin-6-yl)methyl)phenyl)tert-butyl carbamate

[0396]

[0397] Example 24 is generated in a manner similar to that of intermediate D.

[0398] Example 25: 6-(3-aminobenzyl)-2,4-dimethyl-4,6-dihydro-5H-thiazo[4',5':4,5]pyrrolo[2,3-d]pyridazin-5-one

[0399]

[0400] Example 25 was generated in a manner similar to that of Example 3. ¹H NMR (400MHz, DMSO-d6) δ 8.51 (s, ¹H), 6.94 (t, J = 7.6Hz, ¹H), 6.51–6.39 (m, ³H), 5.20 (s, ²H), 5.05 (s, ²H), 4.28 (s, ³H), 2.82 (s, ³H).

[0401] Intermediate AJ: Ethyl 3-formyl-5-methoxy-1H-indole-2-carboxylate

[0402]

[0403] Intermediate AJ was generated in a similar manner to intermediate A. ¹H NMR (400MHz, DMSO-d6) δ 12.77 (s, ¹H), 10.60 (s, ¹H), 7.70 (d, J = 2.5Hz, ¹H), 7.48 (dd, J = 9.0, 0.5Hz, ¹H), 7.05 (dd, J = 9.0, 2.5Hz, ¹H), 4.45 (q, J = 7.1Hz, 2H), 3.81 (s, 3H), 1.40 (t, J = 7.1Hz, 3H).

[0404] Intermediate AK: Ethyl 3-formyl-5-methoxy-1-methyl-1H-indole-2-carboxylate

[0405]

[0406] Intermediate AH was generated in a similar manner to intermediate B. ¹H NMR (400 MHz, DMSO-d⁶) δ 10.44 (s, ¹H), 7.77 (d, J = 2.5 Hz, ¹H), 7.67 (d, J = 9.2 Hz, ¹H), 7.11 (dd, J = 9.1, 2.6 Hz, ¹H), 4.46 (q, J = 7.1 Hz, 2H), 4.04 (s, 3H), 3.83 (s, 3H), 1.40 (t, J = 7.1 Hz, 3H).

[0407] Intermediate AL: 8-methoxy-5-methyl-3,5-dihydro-4H-pyridazin[4,5-b]indol-4-one

[0408]

[0409] Intermediate AL was generated in a similar manner to intermediate C. ¹H NMR (400 MHz, DMSO-d⁶) δ 12.71 (s, ¹H), 8.73 (s, ¹H), 7.75 (d, J = 2.4 Hz, ¹H), 7.71–7.62 (m, ¹H), 7.23 (dd, J = 9.0, 2.5 Hz, ¹H), 4.25 (s, ³H), 3.87 (s, ³H).

[0410] Example 26: (3-((8-methoxy-5-methyl-4-oxoylide-4,5-dihydro-3H-pyridazin[4,5-b]indol-3-yl)methyl)phenyl)tert-butyl carbamate

[0411]

[0412] Example 26 was generated in a manner similar to that of intermediate D. ¹H NMR (400MHz, DMSO-d6) δ 9.31 (s, ¹H), 8.80 (s, ¹H), 7.77 (d, J = 2.5Hz, ¹H), 7.69 (dd, J = 9.2, 0.5Hz, ¹H), 7.39 (s, ¹H), 7.35 (d, J = 8.3Hz, ¹H), 7.24 (dd, J = 9.1, 2.5Hz, ¹H), 7.19 (t, J = 7.8Hz, ¹H), 6.91 (dt, J = 7.7, 1.2Hz, ¹H), 5.33 (s, 2H), 4.26 (s, 3H), 3.87 (s, 3H), 1.44 (s, 9H).

[0413] Example 27: 3-(3-aminobenzyl)-8-methoxy-5-methyl-3,5-dihydro-4H-pyridazin[4,5-b]indol-4-one

[0414]

[0415] Example 27 was generated in a manner similar to that of Example 3.

[0416] Example 28: 7-Methoxy-5-methyl-3-((6-methylpyridin-2-yl)methyl)-3,5-dihydro-4H-pyridazin[4,5-b]indol-4-one

[0417]

[0418] Example 28 was generated in a manner similar to that of Example 15. ¹H NMR (400MHz, DMSO-d6) δ 8.77 (s, ¹H), 8.09 (d, J = 8.8 Hz, ¹H), 7.60 (t, J = 7.7 Hz, ¹H), 7.26 (d, J = 2.2 Hz, ¹H), 7.14 (d, J = 7.6 Hz, ¹H), 7.04 (dd, J = 8.8, 2.2 Hz, ¹H), 6.82 (d, J = 7.7 Hz, ¹H), 5.45 (s, 2H), 4.25 (s, 3H), 3.92 (s, 3H), 2.45 (s, 3H).

[0419] Example 29: 7-hydroxy-5-methyl-3-((6-methylpyridin-2-yl)methyl)-3,5-dihydro-4H-pyridazin[4,5-b]indol-4-one

[0420]

[0421] Example 29 was generated in a manner similar to that of Example 15. ¹H NMR (400MHz, DMSO-d6) δ 9.97(s, ¹H), 8.72(s, ¹H), 8.00(d, J = 8.6Hz, ¹H), 7.59(t, J = 7.7Hz, ¹H), 7.13(d, J = 7.7Hz, ¹H), 6.97(d, J = 2.0Hz, ¹H), 6.92(dd, J = 8.6, 2.1Hz, ¹H), 6.80(d, J = 7.8Hz, ¹H), 5.44(s, 2H), 4.17(s, 3H), 2.45(s, 3H).

[0422] Intermediate AM: 7-methoxy-5-methyl-3-((1-((2-(trimethylsilyl)ethoxy)methyl)-1H-indazol-4-yl)methyl)-3,5-dihydro-4H-pyridazin[4,5-b]indol-4-one

[0423]

[0424] Intermediate AM was generated in a similar manner to intermediate D. ¹H NMR (400MHz, DMSO-d⁶) δ 8.77 (s, ¹H), 8.25 (d, J = 1.0 Hz, ¹H), 8.07 (d, J = 8.7 Hz, ¹H), 7.66 (d, J = 8.5 Hz, ¹H), 7.38 (dd, J = 8.5, 7.0 Hz, ¹H), 7.24 (d, J = 2.0 Hz, ¹H), 7.07–6.99 (m, 2H), 5.77–5.70 (m, 4H), 4.26 (s, 3H), 3.91 (s, 3H), 3.56–3.47 (m, 2H), 0.85–0.72 (m, 2H), -0.07–-0.15 (m, 9H).

[0425] Example 30: 3-((1H-indazol-4-yl)methyl)-7-methoxy-5-methyl-3,5-dihydro-4H-pyridazino[4,5-b]indol-4-one

[0426]

[0427] TFA (1 mL) was added to a solution of 7-methoxy-5-methyl-3-((1-((2(trimethylsilyl)ethoxy)methyl)-1H-indazol-4-yl)methyl)-3,5-dihydro-4H-pyridazino[4,5-b]indol-4-one (38.2 mg, 78.0 μmol) in DCM (5 mL), and the reaction mixture was stirred at room temperature for 4 hours. The reaction mixture was diluted with DCM (10 mL), washed with saturated sodium bicarbonate aqueous solution (2 × 5 mL), dried, concentrated, and purified by rapid column chromatography (0-100% EtOAc in hexane).

[0428] Intermediate AN: 5-methyl-7-(meththio)-3-((1-((2-(trimethylsilyl)ethoxy)methyl)-1H-indazol-4-yl)methyl)-3,5-dihydro-4H-pyridazin[4,5-b]indol-4-one

[0429]

[0430] Intermediate AN is generated in a manner similar to that of Example 1. 1H NMR (400MHz, DMSO-d6) δ8.81(s,1H),8.26(d,J=0.9Hz,1H),8.11(d,J=8.5Hz,1 H),7.66(d,J=8.4Hz,1H),7.58(d,J=1.6Hz,1H),7.38(dd,J=8.4,7.1Hz,1H),7. 30(dd,J=8.4,1.6Hz,1H),7.06(d,J=7.1Hz,1H),5.74(d,J=8.2Hz,2H),4.28(s, 3H), 3.56–3.46 (m, 3H), 2.63 (s, 2H), 0.78 (dd, J = 8.6, 7.6Hz, 2H), -0.12 (s, 9H).

[0431] Example 31: 3-((1H-indazol-4-yl)methyl)-5-methyl-7-(methylthio)-3,5-dihydro-4H-pyridazin[4,5-b]indol-4-one

[0432]

[0433] Example 31 was generated in a manner similar to that of Example AN.

[0434] Intermediate AO: 5-methyl-7-(methylsulfinyl)-3-((1-((2-(trimethylsilyl)ethoxy)methyl)-1H-indazol-4-yl)methyl)-3,5-dihydro-4H-pyridazin[4,5-b]indol-4-one

[0435]

[0436] The intermediate AO was generated in a manner similar to that of Example 1.

[0437] Example 32: 3-((1H-indazol-4-yl)methyl)-5-methyl-7-(methylsulfinyl)-3,5-dihydro-4H-pyridazin[4,5-b]indol-4-one

[0438]

[0439] Example 32 was generated in a manner similar to that of Example 31.

[0440] Intermediate AP: Methyl 6-chloro-1-methyl-1H-pyrrolo[2,3-b]pyridine-2-carboxylate

[0441]

[0442] Intermediate AP was generated from commercially available materials in a manner similar to that of intermediate B. ¹H NMR (400 MHz, DMSO-d6) δ 8.22 (d, J = 8.3 Hz, ¹H), 7.33 (s, ¹H), 7.29 (d, J = 8.3 Hz, ¹H), 4.02 (s, ³H), 3.89 (s, ³H).

[0443] Intermediate AQ: methyl 6-(2-fluorobenzyl)-1-methyl-1H-pyrrolo[2,3-b]pyridine-2-carboxylate.

[0444]

[0445] Tetra(triphenylphosphine)palladium(O) (9.602 mg, 8.309 μmol) was added to a solution of methyl 6-chloro-1-methyl-1H-pyrrolo[2,3-b]pyridine-2-carboxylate (105.9 mg, 471.4 μmol) in a sealed tube under a nitrogen atmosphere, followed by the addition of zinc(II) chloride (2-fluorobenzyl)chloride (111.5 mg, 530.9 μmol) in THF. The reaction mixture was heated to 100 °C for 2 hours. The reaction mixture was cooled and water (30 mL) was added. The reaction mixture was extracted with EtOAc (3 × 30 mL). The organic compounds were combined, dried, concentrated, and purified by rapid column chromatography (0-50% EtOAc in hexane) to give the desired product (42.1 mg, 30.6%). 1H NMR(400MHz, DMSO-d6)δ8.06(d,J=8.1Hz,1H),7.37(td,J=7.7,1.9Hz,1H),7.32–7.26(m,1H), 7.25(s,1H),7.22–7.13(m,2H),7.07(d,J=8.1Hz,1H),4.25(s,2H),4.02(s,3H),3.88(s,3H).

[0446] Intermediate AR: Methyl 3-bromo-6-(2-fluorobenzyl)-1-methyl-1H-pyrrolo[2,3-b]pyridine-2-carboxylate

[0447]

[0448] Intermediate AR was generated in a manner similar to that of intermediate V. ¹H NMR (400 MHz, DMSO-d⁶) δ 7.96 (d, J = 8.2 Hz, ¹H), 7.37 (td, J = 7.6, 1.7 Hz, ¹H), 7.33–7.27 (m, ¹H), 7.22–7.13 (m, ³H), 4.30–4.27 (m, ²H), 4.00 (s, ³H), 3.93 (s, ³H).

[0449] Intermediate AS: Methyl 6-(2-fluorobenzyl)-1-methyl-3-vinyl-1H-pyrrolo[2,3-b]pyridine-2-carboxylate

[0450]

[0451] The intermediate AS is generated in a similar manner to the intermediate AR.

[0452] Intermediate AT: Methyl 6-(2-fluorobenzyl)-3-formyl-1-methyl-1H-pyrrolo[2,3-b]pyridine-2-carboxylate

[0453]

[0454] Intermediate AT is generated in a similar manner to intermediate Y.

[0455] Intermediate AU: 2-(2-fluorobenzyl)-9-methyl-7,9-dihydro-8H-pyrido[3',2':4,5]pyrrolo[2,3-d]pyridazin-8-one

[0456]

[0457] The intermediate AU is generated in a similar manner to intermediate C.

[0458] Example 33: 2-(2-fluorobenzyl)-9-methyl-7-((6-methylpyridin-2-yl)methyl)-7,9-dihydro-8H-pyrido[3',2':4,5]pyrrolo[2,3-d]pyridazin-8-one

[0459]

[0460] Example 33 was generated in a manner similar to that of Example 1.

[0461] Intermediate AV: 2-(2-fluorobenzyl)-9-methyl-7-((1-((2-(trimethylsilyl)ethoxy)methyl)-1H-indazol-4-yl)methyl)-7,9-dihydro-8H-pyrido[3',2':4,5]pyrrolo[2,3-d]pyridazin-8-one

[0462]

[0463] The intermediate AV is generated in a manner similar to that of Example 1.

[0464] Example 34: 7-((1H-indazol-4-yl)methyl)-2-(2-fluorobenzyl)-9-methyl-7,9-dihydro-8H-pyrido[3',2':4,5]pyrrolo[2,3-d]pyridazin-8-one

[0465]

[0466] Example 34 was generated in a manner similar to that of Example 31.

[0467] Intermediate AW: methyl 6-(2-fluorophenoxy)-1-methyl-1H-pyrrolo[2,3-b]pyridine-2-carboxylate.

[0468]

[0469] Under a nitrogen atmosphere, methyl 6-chloro-1-methyl-1H-pyrrolo[2,3-b]pyridine-2-carboxylate (105.9 mg, 471.4 μmol) in toluene was added to a sealed tube containing 1-fluoro-2-hydroxybenzene (68.70 mg, 54.70 μmol), tris(dibenzylacetone)dipalladium (34.54 mg, 37.71 μmol), 2-(dicyclohexylphosphino)-2',4',6'-tris(isopropyl)biphenyl (35.96 mg, 75.43 μmol), and potassium carbonate (143.3 mg, 1.037 mmol). The reaction mixture was heated to 110 °C and stirred overnight. The reaction mixture was cooled and water (30 mL) was added. The reaction mixture was extracted with ethyl EtOAc (3 × 30 mL). The organic compounds were combined, dried, concentrated, and purified by rapid column chromatography (0-100% EtOAc in hexane) to give the desired product (95.2 mg, 67.2%). ¹H NMR (400 MHz, DMSO-d6) δ 8.21 (d, J = 8.5 Hz, 1H), 7.47–7.38 (m, 2H), 7.37–7.29 (m, 2H), 7.28 (d, J = 5.0 Hz, 1H), 6.96 (d, J = 8.5 Hz, 1H), 3.85 (s, 3H), 3.75 (s, 3H).

[0470] Intermediate AX: Methyl 3-bromo-6-(2-fluorophenoxy)-1-methyl-1H-pyrrolo[2,3-b]pyridine-2-carboxylate

[0471]

[0472] Intermediate AX was generated in a similar manner to intermediate V. ¹H NMR (400 MHz, DMSO-d⁶) δ 8.10 (d, J = 8.6 Hz, ¹H), 7.46–7.40 (m, 2H), 7.39–7.27 (m, 2H), 7.06 (d, J = 8.6 Hz, 1H), 3.91 (s, 3H), 3.74 (s, 3H).

[0473] Intermediate AY: Methyl 6-(2-fluorophenoxy)-1-methyl-3-vinyl-1H-pyrrolo[2,3-b]pyridine-2-carboxylate

[0474]

[0475] Intermediate AY was generated in a similar manner to intermediate X. ¹H NMR (400MHz, DMSO-d6) δ 8.50 (d, J = 8.7Hz, ¹H), 7.46–7.42 (m, ¹H), 7.42–7.38 (m, ¹H), 7.38–7.27 (m, ³H), 6.98 (d, J = 8.6Hz, ¹H), 5.91 (dd, J = 18.1, 1.4Hz, ¹H), 5.46 (dd, J = 11.5, 1.3Hz, ¹H), 3.89 (s, ³H), 3.71 (s, ³H).

[0476] Intermediate AZ: 2-(2-fluorophenoxy)-9-methyl-7,9-dihydro-8H-pyrido[3',2':4,5]pyrrolo[2,3-d]pyridazin-8-one

[0477]

[0478] Intermediate AZ is generated in a similar manner to intermediate C.

[0479] Example 35: 2-(2-fluorophenoxy)-9-methyl-7-((6-methylpyridin-2-yl)methyl)-7,9-dihydro-8H-pyrido[3',2':4,5]pyrrolo[2,3-d]pyridazin-8-one

[0480]

[0481] Example 35 was generated in a manner similar to that of Example 1.

[0482] Intermediate BA: 2-(2-fluorophenoxy)-9-methyl-7-((1-((2-(trimethylsilyl)ethoxy)methyl)-1H-indazol-4-yl)methyl)-7,9-dihydro-8H-pyrido[3',2':4,5]pyrrolo[2,3-d]pyridazin-8-one

[0483]

[0484] The intermediate BA is generated in a manner similar to that of Example 1.

[0485] Example 36: 2-(2-fluorophenoxy)-9-methyl-7-((1-((2-(trimethylsilyl)ethoxy)methyl)-1H-indazol-4-yl)methyl)-7,9-dihydro-8H-pyrido[3',2':4,5]pyrrolo[2,3-d]pyridazin-8-one

[0486]

[0487] Example 36 is generated in a similar manner to intermediate 31. NMR is required. Planned Rxn.

[0488] Example 37: 7-Methoxy-5-methyl-3-(3-nitrobenzyl)-3,5-dihydro-4H-pyridazin[4,5-b]indol-4-one

[0489]

[0490] Example 37 was generated in a manner similar to that of Example 1. ¹H NMR (400MHz, DMSO-d6) δ 8.80 (s, ¹H), 8.20–8.14 (m, 2H), 8.09 (d, J = 8.8 Hz, 1H), 7.78 (dt, J = 7.6, 1.4 Hz, 1H), 7.65 (t, J = 7.9 Hz, 1H), 7.25 (d, J = 2.2 Hz, 1H), 7.03 (dd, J = 8.8, 2.2 Hz, 1H), 5.55 (s, 2H), 4.26 (s, 3H), 3.92 (s, 3H).

[0491] Example 38: 7-Hydroxy-5-methyl-3-(3-nitrobenzyl)-3,5-dihydro-4H-pyridazin[4,5-b]indol-4-one

[0492]

[0493] Example 38 was generated in a manner similar to that of Example 15. ¹H NMR (400MHz, DMSO-d6) δ 10.00 (s, ¹H), 8.75 (s, ¹H), 8.21–8.13 (m, 2H), 7.99 (d, J = 8.6Hz, ¹H), 7.77 (dt, J = 7.7, 1.3Hz, 1H), 7.65 (td, J = 7.7, 0.8Hz, 1H), 6.96 (d, J = 1.9Hz, 1H), 6.91 (dd, J = 8.6, 2.1Hz, 1H), 5.54 (s, 2H), 4.17 (s, 3H).

[0494] Example 39: 5-Methyl-3-(3-nitrobenzyl)-7-((6-nitropyridin-2-yl)oxy)-3,5-dihydro-4H-pyridazin[4,5-b]indol-4-one

[0495]

[0496] Example 39 is generated in a manner similar to intermediate XXX2.

[0497] Example 40: 5-Methyl-3-(3-nitrobenzyl)-7-((6-nitropyridin-2-yl)oxy)-3,5-dihydro-4H-pyridazin[4,5-b]indol-4-one

[0498]

[0499] Example 40 is generated in a manner similar to that of Example XXX3.

[0500] Example 41: 5-Methyl-3-((6-methylpyridin-2-yl)methyl)-7-(3-nitrophenoxy)-3,5-dihydro-4H-pyridazin[4,5-b]indol-4-one

[0501]

[0502] Example 42: 7-(3-aminophenoxy)-5-methyl-3-((6-methylpyridin-2-yl)methyl)-3,5-dihydro-4H-pyridazin[4,5-b]indol-4-one

[0503]

[0504] Example 43: 5-Methyl-3-((6-methylpyridin-2-yl)methyl)-7-((6-nitropyridin-2-yl)oxy)-3,5-dihydro-4H-pyridazin[4,5-b]indol-4-one

[0505]

[0506] Example 44: 7-((6-aminopyridin-2-yl)oxy)-5-methyl-3-((6-methylpyridin-2-yl)methyl)-3,5-dihydro-4H-pyridazin[4,5-b]indol-4-one

[0507]

[0508] Example 45: 3-(2-fluorobenzyl)-7-methoxy-5-methyl-3,5-dihydro-4H-pyridazin[4,5-b]indol-4-one

[0509]

[0510] Example 46: 3-(2-fluorobenzyl)-7-hydroxy-5-methyl-3,5-dihydro-4H-pyridazin[4,5-b]indol-4-one

[0511]

[0512] Example 47: 3-(2-fluorobenzyl)-5-methyl-7-((6-nitropyridin-2-yl)oxy)-3,5-dihydro-4H-pyridazin[4,5-b]indol-4-one

[0513]

[0514] Example 48: 7-((6-aminopyridin-2-yl)oxy)-3-(2-fluorobenzyl)-5-methyl-3,5-dihydro-4H-pyridazin[4,5-b]indol-4-one

[0515]

[0516] Intermediate AC: 7-hydroxy-5-methyl-3-((1-((2-(trimethylsilyl)ethoxy)methyl)-1H-indazol-4-yl)methyl)-3,5-dihydro-4H-pyridazin[4,5-b]indol-4-one

[0517]

[0518] Intermediate AD: 5-methyl-7-(3-nitrophenoxy)-3-((1-((2-(trimethylsilyl)ethoxy)methyl)-1H-indazol-4-yl)methyl)-3,5-dihydro-4H-pyridazin[4,5-b]indol-4-one

[0519]

[0520] Intermediate AE: 7-(3-aminophenoxy)-5-methyl-3-((1-((2-(trimethylsilyl)ethoxy)methyl)-1H-indazol-4-yl)methyl)-3,5-dihydro-4H-pyridazin[4,5-b]indol-4-one

[0521]

[0522] Example 49: 3-((1H-indazol-4-yl)methyl)-7-(3-aminophenoxy)-5-methyl-3,5-dihydro-4H-pyridazin[4,5-b]indol-4-one

[0523]

[0524] Intermediate AF: 5-methyl-7-((6-nitropyridin-2-yl)oxy)-3-((1-((2-(trimethylsilyl)ethoxy)methyl)-1H-indazol-4-yl)methyl)-3,5-dihydro-4H-pyridazin[4,5-b]indol-4-one

[0525]

[0526] Intermediate AG: 7-((6-aminopyridin-2-yl)oxy)-5-methyl-3-((1-((2-(trimethylsilyl)ethoxy)methyl)-1H-indazol-4-yl)methyl)-3,5-dihydro-4H-pyridazin[4,5-b]indol-4-one

[0527]

[0528] Example 50: 3-((1H-indazol-4-yl)methyl)-7-((6-aminopyridin-2-yl)oxy)-5-methyl-3,5-dihydro-4H-pyridazin[4,5-b]indol-4-one

[0529]

[0530] Example 51: 3-((1H-pyrazol-3-yl)methyl)-8-fluoro-7-methoxy-5-methyl-3,5-dihydro-4H-pyridazin[4,5-b]indol-4-one

[0531]

[0532] Example 51 was prepared by treating intermediate AB (30 mg, 0.121 mmol) with CsCO3 (119 mg, 364 mmol) in DMF (3 mL) and methanesulfonic acid (1-(tetrahydro-2H-pyran-2-yl)-1H-pyrazol-3-yl)methyl ester (31.6 mg, 0.121 mmol). The reaction mixture was stirred for 48 hours and water was added. The reaction mixture was extracted with EtOAc (5 mL × 3) and the organic matter was dried, concentrated and purified by FCC (0-100% EtOAc in hexane) to provide 8-fluoro-7-methoxy-5-methyl-3-((1-(tetrahydro-2H-pyran-2-yl)-1H-pyrazol-3-yl)methyl)-3,5-dihydro-4H-pyridazino[4,5-b]indol-4-one. The 8-fluoro-7-methoxy-5-methyl-3-((1-(tetrahydro-2H-pyran-2-yl)-1H-pyrazol-3-yl)methyl)-3,5-dihydro-4H-pyridazino[4,5-b]indol-4-one was then dissolved in methanol (3 mL) and treated with 0.4 mL of 4M HCl in dioxane. The resulting reaction mixture was diluted with EtOAc and washed with a saturated aqueous sodium bicarbonate solution. The organic matter was dried, concentrated, and purified by FCC (0-100% EtOAc in hexane) to provide the desired product. 1H NMR (400MHz, DMSO-d6) δ12.63(s,1H),8.68(s,1H),8.07(d,J=11.2Hz,1H),7.62(s,1H) ,7.47(d,J=7.3Hz,1H),6.13(s,1H),5.39(d,J=19.9Hz,2H),4.28(s,3H),4.00(s,3H).

[0533] Example 52: 8-Fluoro-7-methoxy-5-methyl-3-((6-methylpyridin-2-yl)methyl)-3,5-dihydro-4H-pyridazin[4,5-b]indol-4-one

[0534]

[0535] Example 52 was prepared from intermediate AB and commercially available 2-(chloromethyl)-6-methylpyridine in a manner similar to that of Example 1. ¹H NMR (400 MHz, DMSO-d6) δ 12.63 (s, 1H), 8.68 (s, 1H), 8.07 (d, J = 11.2 Hz, 1H), 7.62 (s, 1H), 7.47 (d, J = 7.3 Hz, 1H), 6.13 (s, 1H), 5.39 (d, J = 19.9 Hz, 2H), 4.28 (s, 3H), 4.00 (s, 3H).

[0536] Example 53: 3-(3-aminobenzyl)-8-fluoro-7-methoxy-5-methyl-3,5-dihydro-4H-pyridazin[4,5-b]indol-4-one

[0537]

[0538] Example 53 was generated from intermediate AB using similar reagents and procedures as outlined in Example 3. ¹H NMR (400 MHz, DMSO-d6) δ 8.71 (s, ¹H), 8.08 (d, J = 11.3 Hz, ¹H), 7.48 (d, J = 7.2 Hz, ¹H), 6.94 (dd, J = 7.9 Hz, ¹H), 6.54–6.30 (m, 4H), 5.23 (s, 2H), 5.04 (s, 2H), 4.28 (s, 3H), 4.01 (s, 3H).

[0539] Example 54: 3-(3-aminobenzyl)-8-fluoro-7-hydroxy-5-methyl-3,5-dihydro-4H-pyridazin[4,5-b]indol-4-one

[0540]

[0541] Example 54 was generated from Example 53 using a similar procedure to that used to generate Example 15. ¹H NMR (400 MHz, methanol-d⁴) δ 8.54 (s, ¹H), 7.68 (d, J = 10.8 Hz, ¹H), 7.10–7.01 (m, ¹H), 6.95 (d, J = 7.4 Hz, ¹H), 6.75–6.58 (m, 3H), 5.74–5.70 (m, ¹H), 5.51 (s, 3H), 5.44–5.32 (m, 2H), 4.20 (s, 3H).

[0542] Example 55: 3-(2-fluorobenzyl)-8-methoxy-5-methyl-3,5-dihydro-4H-pyridazin[4,5-b]indol-4-one

[0543]

[0544] Example 55 was prepared by treating intermediate AL (203 mg, 0.89 mmol) with KOtBu (199 mg, 1.77 mmol) in DMF and 1-(chloromethyl)-2-fluorobenzene (141 mg, 0.98 mmol). The reaction mixture was stirred overnight at room temperature and water was added. The resulting precipitate was filtered to provide the desired product (205 mg). ¹H NMR (400 MHz, DMSO-d6) δ 8.81 (s, 1H), 7.76 (d, J = 2.5 Hz, 1H), 7.69 (dd, J = 9.1, 0.6 Hz, 1H), 7.34 (dddd, J = 8.9, 7.3, 5.4, 2.1 Hz, 1H), 7.30–7.09 (m, 4H), 5.46 (s, 2H), 4.26 (s, 3H), 3.87 (s, 3H).

[0545] Example 56: 3-(2-fluorobenzyl)-8-hydroxy-5-methyl-3,5-dihydro-4H-pyridazin[4,5-b]indol-4-one

[0546]

[0547] Example 56 was generated from Example 53 using a similar procedure to that used to generate Example 15. ¹H NMR (400MHz, DMSO-d6) δ 9.46 (s, ¹H), 8.73 (s, ¹H), 7.59 (d, J = 8.9 Hz, ¹H), 7.46 (dd, J = 2.4, 0.6 Hz, ¹H), 7.39–7.27 (m, ¹H), 7.26–7.06 (m, 4H), 5.45 (s, 2H), 4.23 (s, 3H).

[0548] Example 57: 3-(2-fluorobenzyl)-7-((6-fluoropyridin-2-yl)oxy)-5-methyl-3,5-dihydro-4H-pyridazin[4,5-b]indol-4-one

[0549]

[0550] Example 57 was generated by treating Example 56 with 2,6-difluoropyridine (11.7 mg, 0.102 mmol) and KOtBu (32 mg, 0.232 mmol) in DMF. The reaction mixture was stirred overnight at 100 °C, cooled to room temperature, and water was added to the reaction mixture. The reaction mixture was diluted with EtOAc, washed with a saturated aqueous sodium bicarbonate solution, dried, concentrated, and purified by DCC (0-100% EtOAc in hexane) to provide the desired product (36 mg). 1H NMR (400MHz, DMSO-d6) δ9.46 (s, 1H), 8.73 (s, 1H), 7.59 (d, J = 8.9Hz, 1H), 7.46 (dd, J=2.4,0.6Hz,1H),7.39–7.27(m,1H),7.26–7.06(m,4H),5.45(s,2H),4.23(s,3H).

[0551] Example 58: 8-Methoxy-5-methyl-3-((6-methylpyridin-2-yl)methyl)-3,5-dihydro-4H-pyridazin[4,5-b]indol-4-one

[0552]

[0553] Example 58 was generated using a similar procedure to that used to generate Example 55, wherein commercially available 2-(chloromethyl)-6-methylpyridine was used to provide the desired product. ¹H NMR (400 MHz, DMSO-d6) δ 9.44 (s, ¹H), 8.74 (s, ¹H), 7.80–6.66 (m, 6H), 5.44 (s, 2H), 4.23 (s, 3H), 2.45 (s, 3H).

[0554] Example 59: 8-Hydroxy-5-methyl-3-((6-methylpyridin-2-yl)methyl)-3,5-dihydro-4H-pyridazin[4,5-b]indol-4-one

[0555]

[0556] Example 59 was generated from Example 53 using a similar procedure to that used to generate Example 15. ¹H NMR (400MHz, DMSO-d6) δ 9.44 (s, ¹H), 8.74 (s, ¹H), 7.80–6.66 (m, 6H), 5.44 (s, 2H), 4.23 (s, 3H), 2.45 (s, 3H).

[0557] Intermediate AH: 5-methyl-3-((6-methylpyridin-2-yl)methyl)-8-((1-(tetrahydro-2H-pyran-2-yl)-1H-pyrazolo[4,3-b]pyridin-5-yl)oxy)-3,5-dihydro-4H-pyridazino[4,5-b]indol-4-one

[0558]

[0559] Example AH was generated by treating Example 59 (15 mg, 0.47 mmol) with 5-bromo-1-(tetrahydro-2H-pyran-2-yl)-1H-pyrazolo[4,3-b]pyridine (16 mg, 0.56 mmol) and CsCO3 (31 mg, 94 mmol) in DMF (3 mL). The reaction mixture was heated to 120 °C overnight. The reaction mixture was cooled to room temperature, diluted with EtOAc, washed with saturated sodium bicarbonate aqueous solution, dried, concentrated, and purified by DCC (0-100% EtOAc in hexane) to provide the desired product (5 mg). 1HNMR (400MHz, chloroform-d) δ8.52 (s, 1H), 8.14–8.06 (m, 2H), 7.88 (dd, J = 2.3, 0.6Hz ,1H),7.59(dd,J=9.0,0.6Hz,1H),7.55–7.44(m,2H),7.12–7.02(m,2H),6.94– 6.87(m,1H),5.66(s,2H),5.63(dd,J=8.5,3.7Hz,1H),4.41(s,3H),4.18–4.06 (m,1H),3.82–3.71(m,1H),2.58(s,3H),2.29–2.14(m,2H),1.87–1.56(m,4H).

[0560] Example 60: 8-((1H-pyrazolo[4,3-b]pyridin-5-yl)oxy)-5-methyl-3-((6-methylpyridin-2-yl)methyl)-3,5-dihydro-4H-pyridazin[4,5-b]indol-4-one

[0561]

[0562] Example 60 was generated by treating intermediate AH (5 mg, 0.009 mmol) in methanol (1 mL) with 4 M HCl in dioxane (0.2 mL). The reaction mixture was stirred overnight, diluted with EtOAc, washed with saturated sodium bicarbonate aqueous solution, dried, concentrated, and purified by FCC (0-100% EtOAc in hexane). 1H NMR (400MHz, chloroform-d) δ8.52(s,1H),8.05(s,1H),7.92(dd,J=8.9,1.0Hz,1H),7.87(dd,J=2.2,0.6Hz,1H),7.60(dd,J=9.0,0.6Hz,1 H),7.57–7.43(m,2H),7.14(d,J=9.0Hz,1H),7.05(d,J=7.6Hz,1H),6.91(d,J=7.7Hz,1H),5.67(s,2H),4.41(s,3H),2.58(s,3H).

[0563] Example 61: 7-((6-fluoropyridin-2-yl)oxy)-5-methyl-3-((6-methylpyridin-2-yl)methyl)-3,5-dihydro-4H-pyridazin[4,5-b]indol-4-one

[0564]

[0565] Example 61 was generated from Example 59 in a similar manner to Example 57. ¹H NMR (400MHz, DMSO-d6) δ 8.83 (s, ¹H), 8.10 (dd, J = 2.3, 0.6Hz, ¹H), 8.04 (dt, J = 8.6, 7.9Hz, ¹H), 7.86 (dd, J = 9.0, 0.6Hz, ¹H), 7.60 (t, J = 7.7Hz, ¹H), 7.47 (dd, J = 9.0, 2.4Hz, ¹H), 7.14 (d, J = 7.7Hz, ¹H), 7.01–6.94 (m, ¹H), 6.92–6.87 (m, ¹H), 6.84 (d, J = 7.8Hz, ¹H), 5.47 (s, 2H), 4.32 (s, 3H), 2.45 (s, 3H).

[0566] Intermediate AI: 5-methyl-3-((6-methylpyridin-2-yl)methyl)-7-((1-(tetrahydro-2H-pyran-2-yl)-1H-pyrazolo[4,3-b]pyridin-5-yl)oxy)-3,5-dihydro-4H-pyridazin[4,5-b]indol-4-one

[0567]

[0568] Intermediate AI was generated from Example 29 using a procedure similar to that used to generate intermediate AH. ¹H NMR (400MHz, DMSO-d6) δ 8.86 (s, ¹H), 8.44 (d, J = 0.9Hz, ¹H), 8.30–8.21 (m, 2H), 7.67 (d, J = 2.0Hz, 1H), 7.61 (t, J = 7.7Hz, 1H), 7.26 (dd, J = 8.6, 2.0Hz, 1H), 7.16 (t, J = 8.7Hz, 2H), 6.85 (d, J = 7.8Hz, 1H), 5.69 (d d,J=9.6,2.5Hz,1H),5.48(s,2H),4.25(s,3H),3.96(d,J=12.0Hz,1H),3.70(dt,J=11.6,6.7Hz,1H),2. 46(s,3H),2.26–2.11(m,1H),1.99(d,J=6.1Hz,2H),1.81–1.65(m,1H),1.58(dq,J=9.8,5.7,4.8Hz,2H).

[0569] Example 62: 7-((1H-pyrazolo[4,3-b]pyridin-5-yl)oxy)-5-methyl-3-((6-methylpyridin-2-yl)methyl)-3,5-dihydro-4H-pyridazin[4,5-b]indol-4-one

[0570]

[0571] Example 62 was generated from intermediate AI using a procedure similar to that used to generate Example 60. ¹H NMR (400MHz, DMSO-d6) δ 13.32 (s, ¹H), 8.85 (s, ¹H), 8.26 (dd, J = 8.7, 0.6Hz, ¹H), 8.14 (dd, J = 8.9, 1.0Hz, ¹H), 8.03–7.99 (m, ¹H), 7.66–7.55 (m, 2H), 7.24 (dd, J = 8.6, 2.0Hz, 1H), 7.19 (d, J = 8.9Hz, 1H), 7.15 (d, J = 7.6Hz, 1H), 6.85 (d, J = 7.7Hz, 1H), 5.48 (s, 2H), 4.24 (s, 3H), 2.46 (s, 3H).

[0572] Intermediate AJ: 5-methyl-3-((6-methylpyridin-2-yl)methyl)-7-((6-((2-(trimethylsilyl)ethoxy)methoxy)pyridin-2-yl)oxy)-3,5-dihydro-4H-pyridazin[4,5-b]indol-4-one

[0573]

[0574] Intermediate AJ was generated from Example 29 by treatment with 2-chloro-6-((2-(trimethylsilyl)ethoxy)methoxy)pyridine and under similar reaction conditions as used in Example 57.

[0575] Example 63: 7-((6-hydroxypyridin-2-yl)oxy)-5-methyl-3-((6-methylpyridin-2-yl)methyl)-3,5-dihydro-4H-pyridazin[4,5-b]indol-4-one

[0576]

[0577] Example 63 was generated by treating intermediate AJ (28.9 mg, 0.53 mmol) in DCM (3 mL) with 4 M HCl in dioxane (1.0 mL). The reaction mixture was stirred at room temperature for 1 hour. Water and K2CO3 (50 mg) were added and the reaction mixture was stirred for 1 hour. The reaction mixture was diluted with DCM, washed with water, dried, concentrated, and purified by FCC (0-100% EtOAc in hexane) to provide the desired product (7.3 mg). 1H NMR (400MHz, chloroform-d) δ8.46(s,1H),7.97(dd,J=8.6,0.6Hz,1H),7.63–7.54(m,1H),7.49(t,J=7.7Hz,1H),7.24(d,J=2.0Hz,1H),7.17(dd,J=8.7,2. 1Hz,1H),7.05(d,J=7.7Hz,1H),6.84(d,J=7.8Hz,1H),6.48(dd,J=8.3,0 .7Hz, 1H), 6.13 (d, J = 7.7Hz, 1H), 5.59 (s, 2H), 4.28 (s, 3H), 2.58 (s, 3H).

[0578] Intermediate AK: 5-methyl-3-((6-methylpyridin-2-yl)methyl)-7-((1-(tetrahydro-2H-pyran-2-yl)-1H-pyrazolo[4,3-c]pyridin-4-yl)oxy)-3,5-dihydro-4H-pyridazin[4,5-b]indol-4-one

[0579]

[0580] Intermediate AK was generated from Example 29 using a procedure similar to that used to generate intermediate AI. ¹H NMR (400MHz, DMSO-d6) δ 8.88 (s, ¹H), 8.35 (d, J = 0.8Hz, ¹H), 8.29 (dd, J = 8.7, 0.6Hz, ¹H), 7.88 (d, J = 6.0Hz, ¹H), 7.78–7.73 (m, ¹H), 7.61 (t, J = 7.7Hz, ¹H), 7.51 (dd, J = 6.1, 1.0Hz, ¹H), 7.31 (dd, J = 8.6, 2.0Hz, ¹H), 7.15 (d ,J=7.6Hz,1H),6.86(d,J=7.8Hz,1H),5.95–5.89(m,1H),5.48(s,2H),3.92(d,J=11.2Hz,1H),3.86–3.71(m, 1H), 2.55(d,J=5.3Hz,1H),2.45(s,3H),2.43–2.30(m,1H),2.01(d,J=14.8Hz,2H),1.77(s,1H),1.61(s,0H).

[0581] Example 64: 7-((1H-pyrazolo[4,3-c]pyridin-4-yl)oxy)-5-methyl-3-((6-methylpyridin-2-yl)methyl)-3,5-dihydro-4H-pyridazin[4,5-b]indol-4-one

[0582]

[0583] Example 64 was generated from intermediate AK in a manner similar to that used in Example 60. ¹H NMR (400MHz, chloroform-d) δ 10.55 (s, ¹H), 8.58 (s, ¹H), 8.29 (s, ¹H), 8.09 (dd, J = 8.6, 0.6Hz, ¹H), 7.95 (d, J = 6.0Hz, ¹H), 7.52 (t, J = 7.7Hz, ¹H), 7.48 (dd, J = 2.0, 0.6Hz, ¹H), 7.32 (dd, J = 8.6, 2.0Hz, ¹H), 7.15 (d, J = 6.1Hz, ¹H), 7.06 (d, J = 7.7Hz, ¹H), 6.92 (d, J = 7.7Hz, ¹H), 5.68 (s, 2H), 4.35 (s, 3H), 2.58 (s, 3H).

[0584] Intermediate AL: 5-methyl-3-((6-methylpyridin-2-yl)methyl)-7-((1-(tetrahydro-2H-pyran-2-yl)-1H-pyrazolo[3,4-d]pyrimidin-4-yl)oxy)-3,5-dihydro-4H-pyridazin[4,5-b]indol-4-one

[0585]

[0586] Intermediate AL was generated from Example 29 by treatment with 4-chloro-1-(tetrahydro-2H-pyran-2-yl)-1H-pyrazolo[3,4-d]pyrimidine using a procedure similar to that used to generate intermediate AI. ¹H NMR (400 MHz, DMSO-d6) δ 8.90 (s, 1H), 8.60 (s, 1H), 8.38–8.30 (m, 1H), 8.26 (d, J = 0.5 Hz, 1H), 7.92–7.85 (m, 1H), 7.61 (t, J = 7.7 Hz, 1H), 7.40 (dd, J = 8.6, 2.0 Hz, 1H), 7.15 (d, J = 7.6 Hz, 1H), 6.87 (d, J = 7.85 Hz, 1H), 6.87 (d, J = 7.85 Hz, 1H). .8Hz,1H),6.01(d,J=10.3Hz,1H),5.48(s,2H),4.26(s,3H),3.98(d,J=11.8Hz,1H),3.74(d,J=12. 8Hz,1H),2.71–2.65(m,1H),2.58–2.54(m,1H),2.45(s,3H),2.35–2.31(m,1H),2.13–1.41(m,2H).

[0587] Example 65: 7-((1H-pyrazolo[3,4-d]pyrimidin-4-yl)oxy)-5-methyl-3-((6-methylpyridin-2-yl)methyl)-3,5-dihydro-4H-pyridazin[4,5-b]indol-4-one

[0588]

[0589] Example 65 was generated from intermediate AL in a manner similar to that used in Example 60. ¹H NMR (400MHz, DMSO-d⁶) δ 14.20 (s, ¹H), 8.89 (s, ¹H), 8.53 (s, ¹H), 8.34 (d, J = 8.6Hz, ¹H), 8.17 (d, J = 1.3Hz, ¹H), 7.87 (d, J = 2.0Hz, ¹H), 7.61 (t, J = 7.7Hz, ¹H), 7.39 (dd, J = 8.6, 2.0Hz, 1H), 7.15 (d, J = 7.6Hz, 1H), 6.86 (d, J = 7.8Hz, 1H), 5.48 (s, 2H), 4.26 (s, 3H), 2.45 (s, 3H).

[0590] Example 66: 7-((6-fluoropyridin-2-yl)oxy)-5-methyl-3-((6-methylpyridin-2-yl)methyl)-3,5-dihydro-4H-pyridazin[4,5-b]indol-4-one

[0591]

[0592] Example 66 was generated from Example 29 using a procedure similar to that used in Example 57. ¹H NMR (400MHz, DMSO-d⁶) δ 8.87 (s, ¹H), 8.29 (d, J = 8.6Hz, ¹H), 8.06 (q, J = 8.2Hz, ¹H), 7.68 (d, J = 2.0Hz, ¹H), 7.61 (t, J = 7.7Hz, ¹H), 7.24 (dd, J = 8.6, 2.1Hz, ¹H), 7.15 (d, J = 7.7Hz, ¹H), 7.01 (dd, J = 7.9, 1.6Hz, ¹H), 6.92 (dd, J = 7.9, 2.4Hz, ¹H), 6.84 (d, J = 7.8Hz, ¹H), 5.47 (s, 2H), 4.25 (s, 3H), 2.45 (s, 3H).

[0593] Example 67: 7-((6-chloropyridin-2-yl)oxy)-5-methyl-3-((6-methylpyridin-2-yl)methyl)-3,5-dihydro-4H-pyridazin[4,5-b]indol-4-one

[0594]

[0595] Example 67 was generated by treating Example 29 with 2-chloro-6-nitropyridine using a procedure similar to that used in Example 57. ¹H NMR (400 MHz, DMSO-d6) δ 8.88 (s, 1H), 8.29 (d, J = 8.7 Hz, 1H), 8.00–7.92 (m, 1H), 7.73 (s, 1H), 7.68 (d, J = 2.0 Hz, 1H), 7.35–7.20 (m, 3H), 7.10 (d, J = 8.2 Hz, 1H), 6.95 (s, 1H), 5.52 (s, 2H), 4.25 (s, 3H), 2.45 (s, 3H).

[0596] Example 68: 3-(2-fluorobenzyl)-7-methoxy-5-methyl-3,5-dihydro-4H-pyridazin[4,5-b]indol-4-one

[0597]

[0598] Example 68 was generated by treating intermediate L with 1-(chloromethyl)-2-fluorobenzene used in DMF and KOtBu. The reaction mixture was stirred overnight at room temperature, water was added, and the resulting precipitate was collected by filtration to provide the desired product. ¹H NMR (400 MHz, DMSO-d6) δ 8.76 (s, 1H), 8.08 (dd, J = 8.8, 0.5 Hz, 1H), 7.37–7.30 (m, 1H), 7.27–7.10 (m, 5H), 7.03 (dd, J = 8.8, 2.2 Hz, 1H), 5.46 (s, 2H), 4.25 (s, 3H), 3.92 (s, 3H).

[0599] Example 69: 3-(2-fluorobenzyl)-7-hydroxy-5-methyl-3,5-dihydro-4H-pyridazin[4,5-b]indol-4-one

[0600]

[0601] Example 69 was generated from Example 68 using a similar procedure to that used to generate Example 15. ¹H NMR (400MHz, DMSO-d6) δ 9.99 (s, ¹H), 8.71 (s, ¹H), 8.02–7.95 (m, ¹H), 7.39–7.26 (m, ¹H), 7.27–7.10 (m, ³H), 6.96 (dd, J = 2.1, 0.6Hz, ¹H), 6.91 (dd, J = 8.6, 2.1Hz, ¹H), 5.45 (s, ²H), 4.17 (s, ³H).

[0602] Intermediate AM: 3-(2-fluorobenzyl)-5-methyl-7-((1-(tetrahydro-2H-pyran-2-yl)-1H-pyrazolo[4,3-b]pyridin-5-yl)oxy)-3,5-dihydro-4H-pyridazino[4,5-b]indol-4-one

[0603]

[0604] Intermediate AM is generated from Example 69 using a procedure similar to that used to generate intermediate AI.

[0605] Example 70: 7-((1H-pyrazolo[4,3-b]pyridin-5-yl)oxy)-3-(2-fluorobenzyl)-5-methyl-3,5-dihydro-4H-pyridazin[4,5-b]indol-4-one

[0606]

[0607] Example 70 was generated from intermediate AM in a manner similar to that used in Example 60. ¹H NMR (400MHz, DMSO-d6) δ 13.32 (s, ¹H), 8.84 (s, ¹H), 8.24 (dd, J = 8.6, 0.6Hz, ¹H), 8.14 (dd, J = 8.9, 1.0Hz, ¹H), 8.01 (t, J = 1.2Hz, ¹H), 7.64–7.58 (m, ¹H), 7.40–7.31 (m, ¹H), 7.28–7.09 (m, 5H), 5.49 (s, 2H), 4.24 (s, 3H).

[0608] Intermediate AN: 3-(2-fluorobenzyl)-5-methyl-7-((1-(tetrahydro-2H-pyran-2-yl)-1H-pyrazolo[4,3-c]pyridin-4-yl)oxy)-3,5-dihydro-4H-pyridazin[4,5-b]indol-4-one

[0609]

[0610] Intermediate AN was generated in Example 69 by treating it with 5-bromo-1-(tetrahydro-2H-pyran-2-yl)-1H-pyrazolo[4,3-b]pyridine using a similar procedure to that used in the generation of intermediate AI. ¹H NMR (400 MHz, DMSO-d6) δ 8.86 (s, 1H), 8.34 (d, J = 0.7 Hz, 1H), 8.27 (d, J = 8.6 Hz, 1H), 7.88 (d, J = 6.1 Hz, 1H), 7.75 (d, J = 2.0 Hz, 1H), 7.50 (dd, J = 6.1, 0.9 Hz, 1H), 7.41–7.11 (m, 5H), 5.96–5. 88(m,1H),5.49(s,2H),4.25(s,3H),3.92(d,J=11.6Hz,1H),3.82(d,J=28.1Hz,1H),2.70–2 .65(m,1H),2.55(d,J=5.3Hz,2H),2.46(s,0H),2.37–2.28(m,1H),2.03(s,2H),1.61(s,1H).

[0611] Example 71: 7-((1H-pyrazolo[4,3-c]pyridin-4-yl)oxy)-3-(2-fluorobenzyl)-5-methyl-3,5-dihydro-4H-pyridazin[4,5-b]indol-4-one

[0612]

[0613] Example 71 was generated from intermediate AN in a manner similar to that of Example 60. ¹H NMR (400 MHz, chloroform-d) δ 10.72 (s, ¹H), 8.54 (s, ¹H), 8.26 (d, J = 1.0 Hz, ¹H), 8.06 (dd, J = 8.6, 0.6 Hz, ¹H), 7.94 (d, J = 6.0 Hz, ¹H), 7.47 (dd, J = 2.1, 0.5 Hz, ¹H), 7.36–7.23 (m, ³H), 7.17–7.04 (m, ³H), 5.62 (s, ²H), 4.34 (s, ³H).

[0614] Intermediate AO: 3-(2-fluorobenzyl)-5-methyl-7-((6-((2-(trimethylsilyl)ethoxy)methoxy)pyridin-2-yl)oxy)-3,5-dihydro-4H-pyridazin[4,5-b]indol-4-one

[0615]

[0616] Intermediate AO was generated from Example 69 using the procedure employed to generate intermediate AJ. ¹H NMR (400MHz, DMSO-d6) δ 8.87 (s, ¹H), 8.34 (d, J = 8.7 Hz, ¹H), 7.73 (d, J = 2.1 Hz, ¹H), 7.40–7.11 (m, 6H), 6.16 (dd, J = 9.2, 1.1 Hz, 1H), 5.58 (s, 2H), 5.48 (s, 2H), 5.33 (dd, J = 7.6, 1.1 Hz, 1H), 4.26 (s, 3H), 3.72 (dd, J = 8.5, 7.5 Hz, 2H), 0.91 (dd, J = 8.4, 7.6 Hz, 2H), -0.02 (s, 9H).

[0617] Example 72: 3-(2-fluorobenzyl)-7-((6-hydroxypyridin-2-yl)oxy)-5-methyl-3,5-dihydro-4H-pyridazin[4,5-b]indol-4-one

[0618]

[0619] Example 72 was generated from intermediate AO using a similar procedure to that used in Example 63. ¹H NMR (400MHz, DMSO-d⁶) δ 10.79 (s, ¹H), 8.83 (s, ¹H), 8.23 ​​(d, J = 8.7Hz, ¹H), 7.69 (s, ¹H), 7.58 (s, ¹H), 7.43–7.29 (m, ¹H), 7.28–7.10 (m, 4H), 6.55–6.31 (m, 2H), 5.48 (s, 2H), 4.24 (s, 3H).

[0620] Intermediate AP: 3-(2-fluorobenzyl)-5-methyl-7-((1-(tetrahydro-2H-pyran-2-yl)-1H-pyrazolo[3,4-d]pyrimidin-4-yl)oxy)-3,5-dihydro-4H-pyridazin[4,5-b]indol-4-one

[0621]

[0622] Intermediate AP was generated from Example 69 using a similar procedure to that used to generate intermediate AN. ¹H NMR (400MHz, DMSO-d6) δ 8.89 (s, ¹H), 8.60 (s, ¹H), 8.33 (d, J = 8.7Hz, ¹H), 8.25 (s, ¹H), 7.87 (d, J = 2.1Hz, ¹H), 7.43–7.32 (m, 2H), 7.29–7.11 (m, 3H), 6.01 (dd, J = 10.2, 2.5Hz, ¹H), 5.49 (s ,2H),4.26(s,3H),3.98(d,J=12.0Hz,1H),3.72(t,J=12.6Hz,1H),2.70–2.65(m,1H),2.58– 2.54(m,1H),2.36–2.30(m,1H),2.05(d,J=13.0Hz,1H),1.94(d,J=17.0Hz,1H),1.60(s,1H).

[0623] Example 73: 7-((1H-pyrazolo[3,4-d]pyrimidin-4-yl)oxy)-3-(2-fluorobenzyl)-5-methyl-3,5-dihydro-4H-pyridazin[4,5-b]indol-4-one

[0624]

[0625] Example 73 was generated from intermediate AP using a similar procedure to that used in Example 71. ¹H NMR (400 MHz, chloroform-d) δ 11.38 (s, ¹H), 8.63 (s, ¹H), 8.57 (s, ¹H), 8.13 (t, J = 4.3 Hz, 2H), 7.48 (d, J = 2.0 Hz, 1H), 7.36–7.24 (m, 3H), 7.17–7.06 (m, 2H), 5.63 (s, 2H), 4.38 (s, 3H).

[0626] Example 74: 3-(2-fluorobenzyl)-7-((6-fluoropyridin-2-yl)oxy)-5-methyl-3,5-dihydro-4H-pyridazin[4,5-b]indol-4-one

[0627]

[0628] Example 74 was generated from Example 69 using a procedure similar to that used in Example 57. ¹H NMR (400MHz, DMSO-d6) δ 8.86 (s, ¹H), 8.28 (d, J = 8.6Hz, ¹H), 8.06 (q, J = 8.1Hz, ¹H), 7.67 (d, J = 1.9Hz, ¹H), 7.41–7.29 (m, ¹H), 7.29–7.10 (m, 4H), 7.00 (dd, J = 8.0, 1.6Hz, ¹H), 6.92 (dd, J = 7.8, 2.5Hz, ¹H), 5.48 (s, 2H), 4.25 (s, 3H).

[0629] Example 75: 7-Methoxy-5-methyl-3-(3-nitrobenzyl)-3,5-dihydro-4H-pyridazin[4,5-b]indol-4-one

[0630]

[0631] Example 75 was generated by treating intermediate L with 1-(chloromethyl)-3-nitrobenzene using a similar scheme to that used in Example 69. ¹H NMR (400 MHz, DMSO-d6) δ 8.80 (s, 1H), 8.21–8.13 (m, 2H), 8.09 (d, J = 8.8 Hz, 1H), 7.82–7.75 (m, 1H), 7.69–7.59 (m, 1H), 7.25 (d, J = 2.2 Hz, 1H), 7.03 (dd, J = 8.8, 2.2 Hz, 1H), 5.55 (s, 2H), 4.26 (s, 3H), 3.92 (s, 3H).

[0632] Example 76: 7-Hydroxy-5-methyl-3-(3-nitrobenzyl)-3,5-dihydro-4H-pyridazin[4,5-b]indol-4-one

[0633]

[0634] Example 76 was generated from Example 75 using a similar procedure to that used to generate Example 15. ¹H NMR (400MHz, DMSO-d6) δ 10.00 (s, 1H), 8.75 (s, 1H), 8.20–8.12 (m, 2H), 7.99 (d, J = 8.6Hz, 1H), 7.82–7.71 (m, 1H), 7.71–7.59 (m, 1H), 6.96 (d, J = 2.0Hz, 1H), 6.94–6.87 (m, 1H), 5.54 (s, 2H), 4.17 (s, 3H).

[0635] Example 77: 7-((6-chloropyridin-2-yl)oxy)-5-methyl-3-(3-nitrobenzyl)-3,5-dihydro-4H-pyridazin[4,5-b]indol-4-one

[0636]

[0637] Example 77 was generated by treating Example 76 with 2-chloro-6-nitropyridine using a procedure similar to that used in Example 57. ¹H NMR (400 MHz, DMSO-d6) δ 8.89 (s, 1H), 8.28 (dd, J = 8.7, 0.6 Hz, 1H), 8.22–8.12 (m, 2H), 7.94 (dd, J = 8.1, 7.7 Hz, 1H), 7.79 (d, J = 7.9 Hz, 1H), 7.70–7.61 (m, 2H), 7.29 (dd, J = 7.6, 0.6 Hz, 1H), 7.23 (dd, J = 8.6, 2.1 Hz, 1H), 7.09 (dd, J = 8.2, 0.6 Hz, 1H), 5.57 (s, 2H), 4.25 (s, 3H).

[0638] Example 78: 3-(3-aminobenzyl)-7-((6-chloropyridin-2-yl)oxy)-5-methyl-3,5-dihydro-4H-pyridazin[4,5-b]indol-4-one

[0639]

[0640] Example 77 (23.5 mg, 0.051 mmol) was prepared by treating Example 77 (23.5 mg, 0.051 mmol) with stannous chloride dihydrate (57.4 mg, 0.254 mmol), 1,4-dioxane (3 mL), ethanol (3 mL), and water (0.1 mL). The reaction mixture was heated to 100 °C for 5 hours and then cooled to room temperature. The reaction mixture was filtered, diluted with EtOAc, washed with a saturated aqueous bicarbonate solution, dried, concentrated, and purified by FCC (0-10% MeOH in DCM) to provide the desired product (11 mg). 1H NMR(400MHz, DMSO-d6)δ8.83(s,1H),8.27(dd,J=8.7,0.6Hz,1H),7.99–7.90(m,1H),7.66(d,J=2.0Hz,1H),7.28(dd,J=7.6,0.6Hz,1H) ,7.22(dd,J=8.6,2.1Hz,1H),7.09(dd,J=8.2,0.6Hz,1H),7.00–6.89(m,1H),6.52–6.37(m,3H),5.27(s,2H),5.03(s,2H),4.26(s,3H).

[0641] Example 79: 5-Methyl-3-(3-nitrobenzyl)-7-(3-nitrophenoxy)-3,5-dihydro-4H-pyridazin[4,5-b]indol-4-one

[0642]

[0643] Example 79 was generated by treating Example 76 with m-nitrobenzofluoride under similar conditions to those for Example 57. 1H NMR (400MHz, chloroform-d) δ8.55(s,1H),8.32(t,J=2.0Hz,1H),8.17(ddd,J=8.2,2.3,1 .1Hz,1H),8.06(dd,J=8.7,0.6Hz,1H),8.02(ddd,J=8.2,2.2,1.0Hz,1H),7.85(t ,J=2.3Hz,1H),7.82(ddd,J=7.7,1.8,1.1Hz,1H),7.55(td,J=8.1,7.3Hz,2H),7 .41(ddd,J=8.3,2.5,1.0Hz,1H),7.22–7.18(m,1H),7.15(ddd,J=8.6,2.1Hz,1H).

[0644] Example 80: 3-(3-aminobenzyl)-7-(3-aminophenoxy)-5-methyl-3,5-dihydro-4H-pyridazin[4,5-b]indol-4-one

[0645]

[0646] Example 80 was generated from Example 79 using a similar procedure to that used to generate Example 78. ¹H NMR (400 MHz, chloroform-d) δ 8.48 (s, ¹H), 7.97–7.87 (m, ¹H), 7.23–7.03 (m, 4H), 6.96–6.74 (m, 2H), 6.67–6.56 (m, ¹H), 6.55–6.30 (m, 3H), 5.43 (s, 2H), 4.27 (s, 3H).

[0647] Intermediate AO: 7-methoxy-3,5-dimethyl-3,5-dihydro-4H-pyridazin[4,5-b]indol-4-one

[0648]

[0649] Intermediate I (110 mg, 0.48 mmol) was treated with methyl iodide (75.6 mg, 0.53 mmol) and KOtBu (109 mg, 0.97 mmol) in DMF to generate intermediate AO. The reaction mixture was heated to 60 °C for 7 hours, cooled to room temperature, and water was added. The resulting reaction mixture was extracted with DCM, and the organic matter was combined, dried, concentrated, and purified by FCC (0-5% MeOH in DCM) to provide the desired product (64 mg). ¹H NMR (400 MHz, DMSO-d6) δ 8.68 (d, J = 0.6 Hz, 1H), 8.06 (d, J = 8.7 Hz, 1H), 7.23 (d, J = 2.2 Hz, 1H), 7.01 (dd, J = 8.7, 2.2 Hz, 1H), 4.25 (s, 3H), 3.91 (s, 3H), 3.78 (s, 3H).

[0650] Intermediate AQ: 7-hydroxy-3,5-dimethyl-3,5-dihydro-4H-pyridazin[4,5-b]indol-4-one

[0651]

[0652] Intermediate AQ was generated from intermediate AO using a similar procedure to that used to generate Example 15. ¹H NMR (400MHz, DMSO-d6) δ 9.94 (s, ¹H), 8.63 (s, ¹H), 8.02–7.91 (m, ¹H), 6.97–6.92 (m, ¹H), 6.92–6.85 (m, ¹H), 4.17 (s, ³H), 3.77 (s, ³H).

[0653] Example 81: 7-((6-chloropyridin-2-yl)oxy)-3,5-dimethyl-3,5-dihydro-4H-pyridazin[4,5-b]indol-4-one

[0654]

[0655] Example 81 was generated by treating intermediate AQ with 2-chloro-6-nitropyridine using a procedure similar to that used in Example 57. ¹H NMR (400 MHz, DMSO-d6) δ 8.78 (s, ¹H), 8.27 (d, J = 8.6 Hz, ¹H), 7.98–7.90 (m, ¹H), 7.65 (d, J = 2.0 Hz, ¹H), 7.28 (d, J = 7.7 Hz, ¹H), 7.24–7.19 (m, ¹H), 7.11–7.05 (m, ¹H), 4.25 (s, ³H), 3.81 (s, ³H).

[0656] Intermediate AR: Methyl 6-hydroxy-1-methyl-1H-indole-2-carboxylate

[0657]

[0658] Intermediate AQ was generated from commercially available methyl 6-methoxy-1-methyl-1H-indole-2-carboxylate using a similar procedure to that used to generate Example 15. ¹H NMR (400 MHz, DMSO-d6) δ 9.56 (s, 1H), 7.47 (d, J = 8.5 Hz, 1H), 7.16 (d, J = 0.9 Hz, 1H), 6.79 (dt, J = 2.1, 0.7 Hz, 1H), 6.69 (dd, J = 8.6, 2.1 Hz, 1H), 3.90 (s, 3H), 3.82 (s, 3H).

[0659] Intermediate AS: Methyl 6-(benzyloxy)-1-methyl-1H-indole-2-carboxylate

[0660]

[0661] Intermediate AS was generated by treating intermediate AR (1.12 g, 4.56 mmol) with sodium hydride (327 mg, 8.18 mmol) in DMF, followed by treatment with benzyl bromide (1.12 g, 6.55 mmol). The reaction mixture was stirred overnight at room temperature and water was added. The resulting reaction mixture was extracted with EtOAc, and the organics were combined, dried, and purified by FCC (0-50% EtOAc in hexane) to provide the desired product (1.37 g). 1H NMR(400MHz,DMSO-d6)δ7.57(d,J=8.7Hz,1H),7.54–7.46(m,2H),7.46–7.28(m,3H),7 .25–7.17(m,2H),6.86(dd,J=8.7,2.2Hz,1H),5.20(s,2H),3.99(s,3H),3.83(s,3H).

[0662] Intermediate AT: methyl 6-(benzyloxy)-3-formyl-1-methyl-1H-indole-2-carboxylate

[0663]

[0664] Intermediate AT was generated from intermediate AS using a similar scheme to that used in the generation of intermediate B. ¹H NMR (400 MHz, DMSO-d⁶) δ 10.42 (s, ¹H), 8.17 (d, J = 8.8 Hz, ¹H), 7.68–7.31 (m, 6H), 7.08 (dd, J = 8.9, 2.2 Hz, ¹H), 5.22 (s, 2H), 4.03 (s, 3H), 3.99 (s, 3H).

[0665] Intermediate AU: 7-(benzyloxy)-5-methyl-3,5-dihydro-4H-pyridazin[4,5-b]indol-4-one

[0666]

[0667] Intermediate AU was generated from intermediate AT using a similar scheme to that used in the generation of intermediate C. ¹H NMR (400 MHz, DMSO-d⁶) δ 12.72 (s, ¹H), 8.69 (s, ¹H), 8.09 (d, J = 8.8 Hz, ¹H), 7.68–7.28 (m, 6H), 7.09 (dd, J = 8.8, 2.2 Hz, ¹H), 5.26 (s, 2H), 4.24 (s, 3H).

[0668] Intermediate AV: (6-((7-(benzyloxy)-5-methyl-4-oxoylide-4,5-dihydro-3H-pyridazino[4,5-b]indol-3-yl)methyl)pyridin-2-yl)di-tert-butyl carbamate

[0669]

[0670] Intermediate AV was generated by treating intermediate AU with 2-[bis(tert-butoxycarbonyl)amino]-6-(bromomethyl)pyridine under reaction conditions similar to those used in Example 1. ¹H NMR (400 MHz, DMSO-d6) δ 8.75 (s, 1H), 8.11 (dd, J = 8.7, 2.2 Hz, 1H), 7.82 (t, J = 7.8 Hz, 1H), 7.56–7.50 (m, 2H), 7.48–7.33 (m, 4H), 7.25 (dd, J = 7.9, 0.8 Hz, 1H), 7.15–7.08 (m, 2H), 5.46 (s, 2H), 5.27 (s, 2H), 4.24 (s, 3H), 1.31 (s, 18H).

[0671] Intermediate AW: (6-((7-hydroxy-5-methyl-4-oxoylide-4,5-dihydro-3H-pyridazin[4,5-b]indol-3-yl)methyl)pyridin-2-yl)di-tert-butyl carbamate

[0672]

[0673] Intermediate AV (408 mg, 0.67 mmol) was treated with 10% Pd / C (41 mg) in ethanol for 12 hours under a hydrogen atmosphere to generate intermediate AW. The reaction mixture was filtered through diatomaceous earth and concentrated to provide the desired product that could be used without purification. 1H NMR(400MHz, DMSO-d6)δ8.85(s,1H),8.33–8.24(m,2H),7.91–7.78(m,2H),7.77–7.72(m,1H),7.51(dd,J=6.1,0.9Hz,1H),7.34–7.24(m,2 H),7.17–7.12(m,1H),5.97–5.89(m,1H),5.49(s,2H),4.24(s,3H),3 .98–3.86(m,1H),3.84–3.70(m,1H),2.14–1.57(m,6H),1.32(s,18H).

[0674] Intermediate AX: (6-((5-methyl-4-oxoylide-7-((1-(tetrahydro-2H-pyran-2-yl)-1H-pyrazolo[4,3-c]pyridin-4-yl)oxy)-4,5-dihydro-3H-pyridazino[4,5-b]indol-3-yl)methyl)pyridin-2-yl)di-tert-butyl carbamate

[0675]

[0676] Intermediate AX was generated by treating intermediate AW with 5-bromo-1-(tetrahydro-2H-pyran-2-yl)-1H-pyrazolo[4,3-b]pyridine using a similar procedure to that used in the generation of intermediate AI. 1H NMR(400MHz, DMSO-d6)δ8.85(s,1H),8.33–8.24(m,2H),7.91–7.78(m,2H),7.77–7.72(m,1H),7.51(dd,J=6.1,0.9Hz,1H),7.34–7.24(m,2 H),7.17–7.12(m,1H),5.97–5.89(m,1H),5.49(s,2H),4.24(s,3H),3 .98–3.86(m,1H),3.84–3.70(m,1H),2.14–1.57(m,6H),1.32(s,18H).

[0677] Example 82: 7-((1H-pyrazolo[4,3-c]pyridin-4-yl)oxy)-3-((6-aminopyridin-2-yl)methyl)-5-methyl-3,5-dihydro-4H-pyridazin[4,5-b]indol-4-one

[0678]

[0679] Example 82 was generated by treating intermediate AX (16 mg, 0.22 mmol) in methanol (2 mL) with 4 M HCl in dioxane (0.3 mL) and stirring the reaction mixture at room temperature for 8 hours. The reaction mixture was purified by hplc to provide the desired product (3.2 mg). ¹H NMR (400 MHz, methanol-d⁴) δ 8.76 (s, 1H), 8.25 (d, J = 8.6 Hz, 1H), 8.13 (s, 1H), 7.84 (d, J = 6.1 Hz, 1H), 7.66 (d, J = 2.0 Hz, 1H), 7.39 (dd, J = 8.3, 7.3 Hz, 1H), 7.34–7.27 (m, 2H), 6.48 (d, J = 8.1 Hz, 1H), 6.34 (d, J = 7.4 Hz, 1H), 5.42 (s, 2H), 4.33 (s, 3H).

[0680] Intermediate AY: (6-((7-(((6-fluoropyridin-2-yl)oxy)-5-methyl-4-oxylidene-4,5-dihydro-3H-pyridazino[4,5-b]indol-3-yl)methyl)pyridin-2-yl)di-tert-butyl carbamate

[0681]

[0682] Intermediate AY was generated from intermediate AW using a procedure similar to that used in Example 57. ¹H NMR (400 MHz, chloroform-d) δ 8.55 (s, ¹H), 8.04 (dd, J = 8.6, 0.6 Hz, ¹H), 7.83 (q, J = 8.0 Hz, ¹H), 7.38 (dd, J = 8.2, 7.4 Hz, ¹H), 7.34 (dd, J = 2.1, 0.6 Hz, ¹H), 7.19 (dd, J = 8.6, 2.0 Hz, 1H). H),6.85(ddd,J=7.9,1.5,0.6Hz,1H),6.68(ddd,J=7.8,2.7,0.6Hz,1H),6.52(dq,J= 7.4, 0.6Hz, 1H), 6.40 (dd, J = 8.2, 0.8Hz, 1H), 5.50 (s, 2H), 4.46 (s, 2H), 4.34 (s, 3H).

[0683] Example 83: 3-((6-aminopyridin-2-yl)methyl)-7-((6-fluoropyridin-2-yl)oxy)-5-methyl-3,5-dihydro-4H-pyridazin[4,5-b]indol-4-one

[0684]

[0685] Example 83 was generated from intermediate AY using a procedure similar to that used in Example 82. ¹H NMR (400 MHz, chloroform-d) δ 8.55 (s, ¹H), 8.04 (dd, J = 8.6, 0.6 Hz, ¹H), 7.83 (q, J = 8.0 Hz, ¹H), 7.38 (dd, J = 8.2, 7.4 Hz, ¹H), 7.34 (dd, J = 2.1, 0.6 Hz, ¹H), 7.1 9(dd,J=8.6,2.0Hz,1H),6.85(ddd,J=7.9,1.5,0.6Hz,1H),6.68(ddd,J=7.8,2.7,0.6Hz,1H),6 .52(dq,J=7.4,0.6Hz,1H),6.40(dd,J=8.2,0.8Hz,1H),5.50(s,2H),4.46(s,2H),4.34(s,3H).

[0686] Intermediate AZ: 7-(benzyloxy)-5-methyl-3-((1-((2-(trimethylsilyl)ethoxy)methyl)-1H-indazol-4-yl)methyl)-3,5-dihydro-4H-pyridazin[4,5-b]indol-4-one

[0687]

[0688] Intermediate AZ was generated by treating intermediate AU (215 mg, 0.70 mmol) with 4-(bromomethyl)-1-((2-(trimethylsilyl)ethoxy)methyl)-1H-indazole (289 mg, 0.85 mmol) and K₂CO₃ (243 mg, 1.79 mmol) in DMF. The reaction mixture was stirred overnight at room temperature and water was added. The reaction mixture was diluted with EtOAc, washed with a saturated aqueous bicarbonate solution, dried, concentrated, and purified by FCC (0-100% EtOAc in hexane) to provide the desired product (220 mg). 1H NMR (400MHz, DMSO-d6) δ8.78(s,1H),8.26(d,J=0.9Hz,1H),8.09(d,J=8.8Hz,1H),7.66(d,J=8.5Hz,1H),7.56–7.33(m,7H),7.14 –7.00(m,2H),5.75(d,J=2.0Hz,2H),5.72(s,2H),5.26(s,2H),4.25(s,3H),3.56–3.46(m,2H),0.85–0.66(m,2H),-0.12(s,9H).

[0689] Intermediate BA: 7-hydroxy-5-methyl-3-((1-((2-(trimethylsilyl)ethoxy)methyl)-1H-indazol-4-yl)methyl)-3,5-dihydro-4H-pyridazin[4,5-b]indol-4-one

[0690]

[0691] The intermediate BA is generated using a procedure similar to that used to generate the intermediate AW. 1HNMR (400MHz, DMSO-d6) δ9.98 (s, 1H), 8.72 (s, 1H), 8.24 (d, J = 0.9Hz, 1H), 7. 98(d,J=8.6Hz,1H),7.66(d,J=8.4Hz,1H),7.38(dd,J=8.5,7.1Hz,1H),7.07– 7.00(m,1H),6.95(d,J=2.0Hz,1H),6.90(dd,J=8.6,2.1Hz,1H),5.74(s,2H), 5.71(s,2H),4.18(s,3H),3.57–3.46(m,2H),0.84–0.72(m,2H),-0.12(s,9H).

[0692] Intermediate BB: 5-methyl-7-((6-((2-(trimethylsilyl)ethoxy)methoxy)pyridin-2-yl)oxy)-3-((1-((2-(trimethylsilyl)ethoxy)methyl)-1H-indazol-4-yl)methyl)-3,5-dihydro-4H-pyridazin[4,5-b]indol-4-one

[0693]

[0694] Intermediate BB was generated from intermediate BA using a similar procedure to that used to generate intermediate AJ. ¹H NMR (400MHz, DMSO-d6) δ 8.85 (s, ¹H), 8.53 (d, J = 1.1Hz, ¹H), 8.32 (dd, J = 8.7, 0.5Hz, ¹H), 7.72 (d, J = 2.1Hz, ¹H), 7.58 (d, J = 8.7Hz, ¹H), 7.35 (dd, J = 9.1, 7.6Hz, ¹H), 7.29–7.20 (m, 2H), 6.95 (dd, J = 6.8, 0.9Hz, ¹H), 6.15 (d d,J=9.1,1.1Hz,1H),5.74(d,J=12.0Hz,2H),5.65(s,2H),5.57(s,2H),5.30(dd,J=7.6,1.1Hz,1H),4.27(s, 3H),3.76–3.68(m,2H),3.59–3.50(m,2H),0.97–0.87(m,2H),0.84–0.74(m,2H),-0.02(s,9H),-0.17(s,9H).

[0695] Example 84: 3-((1H-indazol-4-yl)methyl)-7-((6-hydroxypyridin-2-yl)oxy)-5-methyl-3,5-dihydro-4H-pyridazin[4,5-b]indol-4-one

[0696]

[0697] Example 84 was generated by treating intermediate BB (21.2 mg, 0.030 mmol) in methanol (2 mL) with 4 M HCl in dioxane (0.5 mL). The reaction mixture was stirred at room temperature for 3 hours. The reaction mixture was diluted with EtOAc, washed with saturated sodium bicarbonate aqueous solution, dried, concentrated, and purified by FCC (0-10% MeOH in DCM) to provide the desired product (4.2 mg). 1H NMR (400MHz, DMSO-d6) δ13.12(s,1H),10.77(s,1H),8.84(s,1H),8.22(d,J=8.7Hz,1H),8.19–8.13(m,1H),7.68(t,J=7.8Hz,1H),7.57(d,J=2.0Hz, 1H),7.47(d,J=8.4Hz,1H),7.30(dd,J=8.4,7.0Hz,1H),7.15(dd,J=8.7,2 .1Hz,1H),7.06–6.93(m,1H),6.57–6.29(m,2H),5.72(s,2H),4.25(s,3H).

[0698] Intermediate BC: 7-((6-fluoropyridin-2-yl)oxy)-5-methyl-3-((1-((2-(trimethylsilyl)ethoxy)methyl)-1H-indazol-4-yl)methyl)-3,5-dihydro-4H-pyridazin[4,5-b]indol-4-one

[0699]

[0700] Intermediate BC is generated from intermediate BA using a procedure similar to that used in generating Example 82. 1HNMR (400MHz, chloroform-d) δ8.52 (s, 1H), 8.44 (d, J = 1.0Hz, 1H), 8.00 (dd, J = 8.6, 0.6H z,1H),7.87–7.78(m,1H),7.74–7.66(m,1H),7.35–7.30(m,2H),7.25–7.20(m,1 H),7.18(dd,J=8.6,2.0Hz,1H),6.88–6.82(m,1H),6.69–6.65(m,1H),5.74(s,2 H),5.73(s,2H),4.34(s,3H),3.68–3.59(m,2H),0.99–0.90(m,2H),0.03(s,9H).

[0701] Example 85: 3-((1H-indazol-4-yl)methyl)-7-((6-fluoropyridin-2-yl)oxy)-5-methyl-3,5-dihydro-4H-pyridazin[4,5-b]indol-4-one

[0702]

[0703] Example 85 was generated using a procedure similar to that used to generate Example 84. ¹H NMR (400MHz, DMSO-d6) δ 13.12 (s, ¹H), 8.86 (s, ¹H), 8.26 (dd, J = 8.6, 0.5Hz, ¹H), 8.19–8.14 (m, ¹H), 8.11–8.01 (m, ¹H), 7.66 (dd, J = 2.1, 0.6Hz, ¹H), 7.47 (d, J = 8.4Hz, ¹H), 7.30 (dd, J = 8.4, 7.0Hz, ¹H), 7.22 (dd, J = 8.6, 2.1Hz, ¹H), 7.06–6.96 (m, 2H), 6.92 (dd, J = 7.8, 2.4Hz, 1H), 5.73 (s, 2H), 4.26 (s, 3H).

[0704] Intermediate BD: 7-(benzyloxy)-5-methyl-3-((1-(tetrahydro-2H-pyran-2-yl)-1H-pyrazol-3-yl)methyl)-3,5-dihydro-4H-pyridazin[4,5-b]indol-4-one

[0705]

[0706] Intermediate BD was generated by treating intermediate AU (637 mg, 2.09 mmol) with CsCO3 (1.70 g, 5.22 mmol) in DMF (8 mL) and methyl methanesulfonic acid (1-(tetrahydro-2H-pyran-2-yl)-1H-pyrazol-3-yl) ester (652 mg, 2.52 mmol). The reaction mixture was stirred for 48 hours and water was added. The reaction mixture was extracted with EtOAc (5 mL × 3), and the organic matter was dried, concentrated, and purified by FCC (0-100% EtOAc in hexane) to provide the desired product. 1H NMR (400MHz, DMSO-d6) δ8.72 (s, 1H), 8.08 (d, J = 8.8Hz, 1H), 7.77 (dd, J = 3.3, 2. 4Hz,1H),7.57–7.51(m,2H),7.47–7.31(m,3H),7.15–7.07(m,1H),6.23(d,J=2 .4Hz,1H),6.16(d,J=2.4Hz,1H),5.34–5.29(m,1H),5.26(s,2H),4.40(d,J=5. 9Hz,2H),4.25(s,3H),3.98–3.81(m,1H),3.68–3.53(m,1H),2.16–1.40(m,6H).

[0707] Intermediate BE: 7-hydroxy-5-methyl-3-((1-(tetrahydro-2H-pyran-2-yl)-1H-pyrazol-3-yl)methyl)-3,5-dihydro-4H-pyridazin[4,5-b]indol-4-one

[0708]

[0709] The intermediate BE was generated using a procedure similar to that used to generate the intermediate AW. ¹H NMR (400MHz, DMSO-d⁶) δ 9.95 (s, ¹H), 8.66 (s, ¹H), 8.03–7.94 (m, ¹H), 6.95 (dd, J = 2.1, 0.6 Hz, ¹H), 6.90 (dd, J = 8.6, 2.1 Hz, ¹H), 6.23 (d, J = 2.4 Hz, ¹H), 6.15 (d, J = 2.4 Hz, ¹H), 4.40 (d, J = 5.8 Hz, 2H), 4.18 (s, 3H), 3.98–3.85 (m, ¹H), 3.67–3.49 (m, ¹H), 2.16–1.39 (m, 6H).

[0710] Intermediate BF: 7-((6-fluoropyridin-2-yl)oxy)-5-methyl-3-((1-(tetrahydro-2H-pyran-2-yl)-1H-pyrazol-3-yl)methyl)-3,5-dihydro-4H-pyridazin[4,5-b]indol-4-one

[0711]

[0712] Intermediate BF was generated from intermediate BE using a procedure similar to that used in Example 82. ¹H NMR (400MHz, DMSO-d6) δ 8.81 (s, ¹H), 8.27 (dd, J = 8.7, 0.6Hz, ¹H), 8.06 (dt, J = 8.6, 7.9Hz, ¹H), 7.79 (d, J = 2.4Hz, ¹H), 7.66 (dd, J = 2.1, 0.5Hz, ¹H), 7.22 (dd, J = 8.6, 2.1Hz, ¹H). 7.03–6.96(m,1H),6.92(dd,J=7.9,2.4Hz,1H),6.17(d,J=2.4Hz,1H),5.76(s,2H),5.39 –5.27(m,1H),4.26(s,3H),3.91(d,J=11.7Hz,1H),3.64–3.53(m,1H),2.19–1.45(m,6H).

[0713] Example 86: 3-((1H-pyrazol-3-yl)methyl)-7-((6-fluoropyridin-2-yl)oxy)-5-methyl-3,5-dihydro-4H-pyridazin[4,5-b]indol-4-one

[0714]

[0715] Example 86 was generated from intermediate BF in a manner similar to that used in Example 60. ¹H NMR (400MHz, DMSO-d6) δ 12.65 (s, ¹H), 8.80 (s, ¹H), 8.27 (d, J = 8.6 Hz, ¹H), 8.14–7.99 (m, ¹H), 7.70–7.56 (m, 2H), 7.22 (dd, J = 8.6, 2.1 Hz, 1H), 7.00 (dd, J = 7.9, 1.6 Hz, 1H), 6.92 (dd, J = 7.9, 2.4 Hz, 1H), 6.14 (s, ¹H), 5.40 (s, 2H), 4.26 (s, 3H).

[0716] Intermediate BG: 5-methyl-3-((1-(tetrahydro-2H-pyran-2-yl)-1H-pyrazol-3-yl)methyl)-7-((1-(tetrahydro-2H-pyran-2-yl)-1H-pyrazolo[4,3-c]pyridin-4-yl)oxy)-3,5-dihydro-4H-pyridazino[4,5-b]indol-4-one

[0717]

[0718] The intermediate BG is generated from the intermediate BE in a manner similar to that used in generating the intermediate AX.

[0719] Example 87: 3-((1H-pyrazol-3-yl)methyl)-7-((1H-pyrazolo[4,3-c]pyridin-4-yl)oxy)-5-methyl-3,5-dihydro-4H-pyridazin[4,5-b]indol-4-one

[0720]

[0721] Example 87 was generated from intermediate BG in a manner similar to that used in Example 60. ¹H NMR (400MHz, DMSO-d6) δ 13.63 (s, ¹H), 12.65 (s, ¹H), 8.80 (s, ¹H), 8.33–8.23 (m, 2H), 7.84–7.78 (m, ¹H), 7.76–7.70 (m, ¹H), 7.64 (s, ¹H), 7.33–7.24 (m, 3H), 5.42 (d, J = 21.1Hz, 2H), 4.26 (s, 3H).

[0722] Intermediate BH: 7-(benzyloxy)-3-(4-methoxybenzyl)-5-methyl-3,5-dihydro-4H-pyridazin[4,5-b]indol-4-one

[0723]

[0724] Intermediate BH was generated by treating intermediate AU (260 mg, 0.85 mmol) in DMF (6 mL) with 1-(chloromethyl)-4-methoxybenzene (134 mg, 0.85 mmol) and K₂CO₃ (294 mg, 2.13 mmol). The reaction mixture was stirred overnight at room temperature, diluted with EtOAc, washed with water, dried, concentrated, and purified by FCC (0-100% EtOAc in hexane) to provide the desired product (270 mg). 1H NMR(400MHz, DMSO-d6)δ8.73(s,1H),8.08(d,J=8.8Hz,1H),7.56–7.49(m,2H),7.47–7.40(m,1H),7.40–7.33(m,1H),7.33–7.25 (m,2H),7.27–7.20(m,2H),7.09(dd,J=8.8,2.2Hz,1H),6.92–6.84(m,2H),5.32(s,2H),5.26(s,2H),4.25(s,3H),3.72(s,3H).

[0725] Intermediate BI: 7-hydroxy-3-(4-methoxybenzyl)-5-methyl-3,5-dihydro-4H-pyridazin[4,5-b]indol-4-one

[0726]

[0727] Intermediate BI was generated using a procedure similar to that used to generate intermediate AW. ¹H NMR (400MHz, DMSO-d⁶) δ 9.97 (s, ¹H), 8.68 (s, ¹H), 7.97 (d, J = 8.6 Hz, ¹H), 7.32–7.23 (m, 2H), 6.93–6.82 (m, 4H), 5.31 (s, 2H), 4.17 (s, 3H), 3.72 (s, 3H).

[0728] Example 88: 7-((6-fluoropyridin-2-yl)oxy)-3-(4-methoxybenzyl)-5-methyl-3,5-dihydro-4H-pyridazin[4,5-b]indol-4-one

[0729]

[0730] Example 88 was generated from intermediate BI using a procedure similar to that used in Example 82. ¹H NMR (400MHz, DMSO-d⁶) δ 8.83 (d, J = 2.9Hz, 1H), 8.26 (dd, J = 8.5, 2.7Hz, 1H), 8.06 (q, J = 8.5, 7.9Hz, 1H), 7.66 (s, 1H), 7.31 (d, J = 8.1Hz, 2H), 7.22 (d, J = 9.0Hz, 1H), 7.00 (d, J = 8.2Hz, 1H), 6.96–6.86 (m, 3H), 5.35 (d, J = 3.0Hz, 2H), 4.25 (d, J = 2.9Hz, 3H), 3.73 (d, J = 2.8Hz, 3H).

[0731] Intermediate BJ: 7-((6-fluoropyridin-2-yl)oxy)-5-methyl-3,5-dihydro-4H-pyridazin[4,5-b]indol-4-one

[0732]

[0733] Intermediate BJ was prepared by dissolving Example 88 (75.2 mg, 0.175 mmol) in trifluoroacetic acid (3 mL) and heating the resulting reaction mixture to 60 °C for 5 hours. The reaction mixture was concentrated, dissolved in DCM, washed with saturated sodium bicarbonate aqueous solution, dried, concentrated, and purified by FCC (0-100% EtOAc in hexane) to provide the desired product (43 mg). 1H NMR (400MHz, DMSO-d6) δ12.82(s,1H),8.77(s,1H),8.26(dd,J=8.6,0.6Hz,1H),8.06(dt,J=8.6,7.9Hz,1H),7.65(dd,J=2.1 ,0.6Hz,1H),7.21(dd,J=8.6,2.1Hz,1H),6.99(ddd,J=8.1,1.8,0.6Hz,1H),6.92(ddd,J=8.0,2.5,0.5Hz,1H),4.24(s,3H).

[0734] Intermediate BK: (3-((7-((6-fluoropyridin-2-yl)oxy)-5-methyl-4-oxoylide-4,5-dihydro-3H-pyridazino[4,5-b]indol-3-yl)methyl)phenyl)carbamate bis-tert-butyl

[0735]

[0736] The intermediate BK is generated using a procedure similar to that used to generate the intermediate AV.

[0737] Example 89: 3-(3-aminobenzyl)-7-((6-fluoropyridin-2-yl)oxy)-5-methyl-3,5-dihydro-4H-pyridazin[4,5-b]indol-4-one

[0738]

[0739] Example 89 was generated from intermediate BK using a procedure similar to that used in Example 82. ¹H NMR (400 MHz, chloroform-d) δ 8.50 (s, ¹H), 8.04–7.99 (m, ¹H), 7.86–7.78 (m, ¹H), 7.34–7.30 (m, ¹H), 7.21–7.16 (m, ¹H), 7.16–7.09 (m, ¹H), 6.89–6.81 (m, 2H), 6.81–6.77 (m, ¹H), 6.71–6.65 (m, ¹H), 6.64–6.59 (m, ¹H), 5.43 (s, 2H), 4.33 (s, 3H).

[0740] Example 90: 7-((6-fluoropyridin-2-yl)oxy)-3-(3-methoxybenzyl)-5-methyl-3,5-dihydro-4H-pyridazin[4,5-b]indol-4-one

[0741]

[0742] Example 90 was generated from intermediate BJ using a procedure similar to that used to generate intermediate BH. ¹H NMR (400MHz, DMSO-d6) δ 8.84 (s, ¹H), 8.30–8.24 (m, ¹H), 8.06 (dt, J = 8.6, 7.9 Hz, ¹H), 7.66 (dd, J = 2.1, 0.5 Hz, ¹H), 7.28–7.20 (m, 2H), 7.00 (dd, J = 8.0, 1.6 Hz, 1H), 6.94–6.76 (m, 4H), 5.39 (s, 2H), 4.25 (s, 3H), 3.73 (d, J = 1.2 Hz, 3H).

[0743] Intermediate BL: 4-Fluoro-2-iodo-5-(methoxymethyl)aniline

[0744]

[0745] Intermediate BL was generated by treating 4-fluoro-3-(methoxymethyl)aniline (1.84 g, 11.89 mmol) in acetic acid with 1-iodopyrrolidine-2,5-dione (3.08 g, 13.78 mmol), and the reaction mixture was stirred at room temperature for 3 h. The reaction mixture was diluted with EtOAc, washed with saturated sodium thiosulfate aqueous solution, and purified by FCC (0–50% EtOAc in hexane) to provide the desired product. ¹H NMR (400 MHz, DMSO-d6) δ 7.39 (d, J = 9.3 Hz, 1H), 6.83–6.75 (m, 1H), 5.08 (s, 2H), 4.36–4.26 (m, 2H), 3.29 (s, 3H).

[0746] Intermediate BM: N-(4-fluoro-2-iodo-5-(methoxymethyl)phenyl)-4-methylbenzenesulfonamide

[0747]

[0748] Intermediate BM was generated by treating intermediate BL (2.30 g, 8.18 mmol) in DCM (50 mL) with pyridine (1.00 mL, 12.27 mmol) and 4-methylbenzenesulfonyl chloride (1.56 g, 8.18 mmol), and the reaction mixture was stirred overnight at room temperature. The reaction mixture was washed with saturated sodium bicarbonate aqueous solution, dried, concentrated, and purified by FCC (0–100% EtOAc in hexane) to provide the desired product. ¹H NMR (400 MHz, DMSO-d6) δ 9.69 (s, 1H), 7.73–7.66 (m, 1H), 7.62–7.53 (m, 2H), 7.43–7.34 (m, 2H), 7.02–6.96 (m, 1H), 4.36–4.27 (m, 2H), 3.19 (s, 3H), 2.38 (s, 3H).

[0749] Intermediate BN: Ethyl 5-fluoro-6-(methoxymethyl)-1-toluenesulfonyl-1H-indole-2-carboxylate

[0750]

[0751] Intermediate BN was generated by treating intermediate BM (3.11 g, 7.15 mmol) in degassed THF (50 mL) with Pd(P(Ph3)4) (413 mg, 0.36 mmol), ethyl propynate (2.10 g, 21.44 mol), DIPEA (4.62 g, 35.73 mmol), and ZnBr2 (4.83 g, 21.44 mmol). The reaction mixture was heated to 80 °C overnight, cooled to room temperature, and filtered through diatomaceous earth. The filtrate was washed with saturated sodium bicarbonate, dried, concentrated, and purified by FCC (0-100% EtOAc in hexane) to provide the desired product (1.76 g). 1H NMR (400MHz, DMSO-d6) δ8.07–8.02(m,1H),7.85–7.81(m,2H),7.51(d,J=9.8Hz,1H),7.46–7.41(m,2H),7.35(d ,J=0.8Hz,1H),4.60(d,J=1.2Hz,2H),4.36(q,J=7.1Hz,2H),3.35(s,3H),2.36(s,3H),1.32(t,J=7.1Hz,3H).zz

[0752] Intermediate BO: Ethyl 5-fluoro-6-(methoxymethyl)-1H-indole-2-carboxylate

[0753]

[0754] Intermediate BO was generated by treating intermediate BN (2.30 g, 5.68 mmol) in ethanol (25 mL) and THF (25 mL) with potassium hydroxide (796 mg, 14.19 mmol). The reaction mixture was stirred at room temperature for 2 hours. The reaction mixture was diluted with EtOAc, washed with water, dried, concentrated, and purified by FCC (0-10% MeOH in DCM) to provide the desired product (4.62 g). 1H NMR (400MHz, DMSO-d6) δ12.00(s,1H),7.47(dd,J=6.2,0.9Hz,1H),7.43(d,J=10.8Hz,1H),7.12(d d,J=2.2,0.9Hz,1H),4.58–4.51(m,2H),4.35(q,J=7.1Hz,2H),3.34(s,3H),1.35(t,J=7.1Hz,3H).

[0755] Intermediate BP: Ethyl 5-fluoro-6-(methoxymethyl)-1-methyl-1H-indole-2-carboxylate

[0756]

[0757] Intermediate BP was generated by treating intermediate BO (50.4 mg, 0.20 mmol) in DMF (3 mL) with K₂CO₃ (69.3 mg, 0.50 mmol) and MeI (31.3 mg, 0.22 mmol). The resulting reaction mixture was stirred overnight at room temperature. The reaction mixture was diluted with EtOAc, washed with water, dried, concentrated, and purified by FCC (0-50% EtOAc in hexane) to provide the desired product. 1HNMR (400MHz, DMSO-d6) δ7.64(d,J=6.0Hz,1H),7.47(d,J=10.5Hz,1H),7.24(d,J=0.8Hz ,1H),4.57(s,2H),4.33(q,J=7.1Hz,2H),4.04(s,3H),3.36(s,3H),1.34(t,J=7.1Hz,3H).

[0758] Intermediate BQ: Ethyl 5-fluoro-3-formyl-6-(methoxymethyl)-1-methyl-1H-indole-2-carboxylate

[0759]

[0760] Intermediate BQ was generated from intermediate BP using a similar procedure to that used to generate intermediate A. z1H NMR (400MHz, DMSO-d6) δ 10.42 (s, 1H), 7.96 (d, J = 10.5Hz, 1H), 7.82 (d, J = 6.0Hz, 1H), 4.60 (t, J = 0.9Hz, 2H), 4.48 (qd, J = 7.1, 1.7Hz, 2H), 4.07 (s, 3H), 3.38 (s, 3H), 1.41 (td, J = 7.1, 1.0Hz, 3H).

[0761] Intermediate BR: 8-fluoro-7-(methoxymethyl)-5-methyl-3,5-dihydro-4H-pyridazin[4,5-b]indol-4-one

[0762]

[0763] Intermediate BR was generated from intermediate BQ using a similar procedure to that used to generate intermediate C. ¹H NMR (400MHz, DMSO-d⁶) δ 12.80 (s, ¹H), 8.72 (s, ¹H), 8.07 (d, J = 10.2 Hz, ¹H), 7.80 (d, J = 5.9 Hz, ¹H), 4.65 (d, J = 1.2 Hz, 2H), 4.29 (s, 3H), 3.40 (s, 3H).

[0764] Example 91: 8-Fluoro-7-(methoxymethyl)-5-methyl-3-((6-methylpyridin-2-yl)methyl)-3,5-dihydro-4H-pyridazin[4,5-b]indol-4-one

[0765]

[0766] Example 91 was prepared by treating intermediate BR (25.2 mg, 0.097 mmol) in DMF (3 mL) with 2-(bromomethyl)-6-methylpyridine (19.7 mg, 0.106 mmol) and Cs₂CO₃ (78.6 mg, 0.241 mmol). The reaction mixture was stirred overnight at room temperature, diluted with EtOAc, washed with water, dried, concentrated, and purified by FCC (0-100% EtOAc in hexane) to provide the desired product (23 mg). 1H NMR (400MHz, DMSO-d6) δ8.82(s,1H),8.10(d,J=10.2Hz,1H),7.83(d,J=5.8Hz,1H),7.60(t,J=7.7Hz,1H),7.14(d,J=7 .6Hz, 1H), 6.85 (d, J = 7.7Hz, 1H), 5.46 (s, 2H), 4.66 (d, J = 1.0Hz, 2H), 4.30 (s, 3H), 3.39 (d, J = 13.3Hz, 3H), 2.45 (s, 3H).

[0767] Intermediate BS: 7-(bromomethyl)-8-fluoro-5-methyl-3-((6-methylpyridin-2-yl)methyl)-3,5-dihydro-4H-pyridazin[4,5-b]indol-4-one

[0768]

[0769] Intermediate BS was generated by treating DCM (5 mL) in Example 91 (150 mg, 0.41 mmol) with BBr3 (461 mg, 1.843 mmol) and stirring the reaction mixture at room temperature for 2 hours. The reaction mixture was diluted with DCM, washed with saturated sodium bicarbonate aqueous solution, dried, concentrated, and purified by FCC (0-100% EtOAc in hexane) to provide the desired product (120 mg). 1H NMR (400MHz, DMSO-d6) δ8.82(s,1H),8.15(d,J=10.2Hz,1H),8.04(d,J=6.1Hz,1H),7.61(t,J=7.7Hz,1H), 7.15(d,J=7.7Hz,1H),6.86(d,J=7.8Hz,1H),5.46(s,2H),4.92(d,J=1.2Hz,2H),4.28(s,3H),2.44(s,3H).

[0770] Example 92: 8-Fluoro-5-methyl-3-((6-methylpyridin-2-yl)methyl)-7-(morpholinomethyl)-3,5-dihydro-4H-pyridazin[4,5-b]indol-4-one

[0771]

[0772] Example 92 was prepared by treating intermediate BS (38.1 mg, 0.068 mmol) in DCM (3 mL) with triethylamine (27.4 mg, 0.271 mmol) and morpholine (11.8 mg, 0.135 mmol), and stirring the reaction mixture overnight at room temperature. The reaction mixture was diluted with DCM, washed with saturated sodium bicarbonate aqueous solution, dried, concentrated, and purified by FCC (0-20% MeOH in DCM) to provide the desired product (23 mg). 1H NMR (400MHz, DMSO-d6) δ8.81(s,1H),8.08(d,J=10.1Hz,1H),7.78(d,J=5.9Hz,1H),7.60(t,J=7.7Hz,1H),7.14(d,J=7. 6Hz, 1H), 6.84 (d, J = 7.7Hz, 1H), 4.29 (s, 3H), 3.75–3.69 (m, 2H), 3.65–3.57 (m, 6H), 2.47 (t, J = 4.6Hz, 4H), 2.44 (s, 3H).

[0773] Example 93: 8-Fluoro-7-(((2-hydroxyethyl)amino)methyl)-5-methyl-3-((6-methylpyridin-2-yl)methyl)-3,5-dihydro-4H-pyridazin[4,5-b]indol-4-one

[0774]

[0775] Example 93 was generated from intermediate BS using a procedure similar to that used to generate Example 92. ¹H NMR (400MHz, DMSO-d⁶) δ 8.80 (s, ¹H), 8.05 (d, J = 10.3 Hz, ¹H), 7.85 (d, J = 6.0 Hz, ¹H), 7.60 (t, J = 7.7 Hz, ¹H), 7.14 (d, J = 7.6 Hz, ¹H), 6.84 (d, J = 7.7 Hz, ¹H), 5.46 (s, 2H), 4.52 (t, J = 5.4 Hz, 1H), 4.29 (s, 3H), 3.95 (s, 2H), 3.52 (q, J = 5.6 Hz, 2H), 2.67 (t, J = 5.7 Hz, 2H), 2.53–2.51 (m, ¹H), 2.45 (s, 3H).

[0776] Example 94: 8-Fluoro-5-methyl-3-((6-methylpyridin-2-yl)methyl)-7-(((tetrahydro-2H-pyran-4-yl)amino)methyl)-3,5-dihydro-4H-pyridazin[4,5-b]indol-4-one

[0777]

[0778] Example 94 was generated from intermediate BS using a procedure similar to that used to generate Example 92. ¹H NMR (400 MHz, methanol-d⁴) δ 7.87 (s, ¹H), 7.09 (d, J = 10.0 Hz, ¹H), 7.00 (d, J = 5.9 Hz, ¹H), 6.81 (t, J = 7.8 Hz, ¹H), 6.36 (d, J = 7.6 Hz, ¹H), 6.11 (d, J = 7.8 Hz, ¹H), 4.75 (s, 2H), 3 .52(s,3H),3.36–3.28(m,2H),3.17(dd,J=11.4,4.5Hz,2H),2.67–2.57(m,2H),2 .51(s,1H),2.16–2.04(m,1H),1.72(s,3H),1.23–1.13(m,2H),0.84–0.67(m,2H).

[0779] Example 95: 8-Fluoro-5-methyl-3-((6-methylpyridin-2-yl)methyl)-7-(pyrrolidone-1-ylmethyl)-3,5-dihydro-4H-pyridazin[4,5-b]indol-4-one

[0780]

[0781] Example 95 was generated from intermediate BS using a procedure similar to that used to generate Example 92. ¹H NMR (400MHz, DMSO-d⁶) δ 8.80 (s, ¹H), 8.06 (d, J = 10.2 Hz, ¹H), 7.77 (d, J = 5.9 Hz, ¹H), 7.60 (t, J = 7.7 Hz, ¹H), 7.14 (d, J = 7.6 Hz, ¹H), 6.84 (d, J = 7.7 Hz, ¹H), 5.46 (s, 2H), 4.29 (s, 3H), 3.82 (s, 2H), 2.54 (s, 4H), 2.44 (s, 3H), 1.73 (s, 4H).

[0782] Example 96: 7-((dimethylamino)methyl)-8-fluoro-5-methyl-3-((6-methylpyridin-2-yl)methyl)-3,5-dihydro-4H-pyridazin[4,5-b]indol-4-one

[0783]

[0784] Example 96 was generated from intermediate BS using a procedure similar to that used to generate Example 92. ¹H NMR (400MHz, DMSO-d⁶) δ 8.81 (s, ¹H), 8.07 (d, J = 10.2 Hz, ¹H), 7.77 (d, J = 5.9 Hz, ¹H), 7.60 (t, J = 7.7 Hz, ¹H), 7.14 (d, J = 7.7 Hz, ¹H), 6.84 (d, J = 7.8 Hz, ¹H), 5.46 (s, 2H), 4.29 (s, 3H), 3.68–3.60 (m, 2H), 2.44 (s, 3H), 2.23 (d, J = 1.5 Hz, 6H).

[0785] Example 97: 7-((6-fluoropyridin-2-yl)oxy)-5-methyl-3-(2-morpholinoethyl)-3,5-dihydro-4H-pyridazin[4,5-b]indol-4-one

[0786]

[0787] Example 97 was prepared by treating intermediate BJ (10.1 mg, 0.033 mmol) in DMF (3 mL) with potassium carbonate (11.2 mg, 0.081 mmol) and 4-(2-chloroethyl)morpholine (4.9 mg, 0.033 mmol), and stirring the reaction mixture overnight at room temperature. The reaction mixture was diluted with EtOAc, washed with water, dried, concentrated, and purified by FCC (0-30% MeOH, DCM) to provide the desired product (9.3 mg). 1H NMR (400MHz, DMSO-d6) δ8.81(s,1H),8.32–8.24(m,1H),8.12–8.02(m,1H),7.65(d,J=2.0Hz,1H),7.22(dd,J=8.7,2.1Hz,1H),7.00(dd,J=7 .9,1.6Hz,1H),6.92(dd,J=7.8,2.4Hz,1H),4.36(t,J=6.9Hz,2H),4.25(s,3H),3.58–3.51(m,4H),2.79–2.67(m,2H),2.47(t,J=4.5Hz,4H).

[0788] Example 98: 3-(2-(dimethylamino)ethyl)-7-((6-fluoropyridin-2-yl)oxy)-5-methyl-3,5-dihydro-4H-pyridazin[4,5-b]indol-4-one

[0789]

[0790] Example 98 was generated from intermediate BJ using a similar procedure to that used to generate Example 97. ¹H NMR (400MHz, DMSO-d6) δ 8.80 (s, ¹H), 8.26 (dd, J = 8.6, 0.5Hz, ¹H), 8.12–7.99 (m, ¹H), 7.67–7.61 (m, ¹H), 7.21 (dd, J = 8.6, 2.1Hz, ¹H), 6.99 (dd, J = 7.9, 1.6Hz, ¹H), 6.92 (dd, J = 7.8, 2.4Hz, ¹H), 4.38–4.29 (m, 2H), 4.25 (s, 3H), 2.74–2.64 (m, 2H), 2.22 (s, 6H).

[0791] Intermediate BT: 3-(2-(benzyloxy)ethyl)-7-((6-fluoropyridin-2-yl)oxy)-5-methyl-3,5-dihydro-4H-pyridazin[4,5-b]indol-4-one

[0792]

[0793] Intermediate BT is generated from intermediate BJ using a similar procedure to that used to generate Example 97.

[0794] Example 99: 7-((6-fluoropyridin-2-yl)oxy)-3-(2-hydroxyethyl)-5-methyl-3,5-dihydro-4H-pyridazin[4,5-b]indol-4-one

[0795]

[0796] Example 99 was prepared by treating intermediate BS (20.2 mg, 0.045 mmol) in EtOH (5 mL) with 10% Pd / C (5.0 mg), and the reaction mixture was placed under a hydrogen atmosphere and stirred overnight. The reaction mixture was filtered through diatomaceous earth and concentrated to provide the desired product (12 mg). 1H NMR(400MHz,)δ8.80(s,1H),8.26(dd,J=8.7,0.6Hz,1H),8.10–8.02(m,1H),7.68–7.63(m,1H),7.21(dd,J=8.6,2.1Hz,1H),7 .00(dd,J=7.8,1.6Hz,1H),6.92(dd,J=7.9,2.4Hz,1H),4.86–4.78(m,1H),4.33–4.27(m,2H),4.26(s,3H),3.83–3.73(m,2H).

[0797] Intermediate BU: 4-(2-(7-((6-fluoropyridin-2-yl)oxy)-5-methyl-4-oxoylide-4,5-dihydro-3H-pyridazino[4,5-b]indol-3-yl)ethyl)piperazine-1-carboxylic acid tert-butyl ester

[0798]

[0799] Intermediate BU is generated from intermediate BJ using a similar procedure to that used to generate Example 97. 1HNMR(400MHz,DMSO-d6)δ8.80(s,1H),8.26(dd,J=8.6,0.5Hz,1H),8.11–7 .98(m,1H),7.64(dd,J=2.1,0.5Hz,1H),7.21(dd,J=8.6,2.1Hz,1H),6.99(d d,J=8.0,1.6Hz,1H),6.91(dd,J=7.9,2.5Hz,1H),4.42–4.30(m,2H),4.25(s ,3H),3.32–3.23(m,4H),2.82–2.72(m,2H),2.48–2.39(m,5H),1.39(s,9H).

[0800] Example 100: 7-((6-fluoropyridin-2-yl)oxy)-5-methyl-3-(2-(piperazin-1-yl)ethyl)-3,5-dihydro-4H-pyridazin[4,5-b]indol-4-one

[0801]

[0802] Example 100 was prepared by treating intermediate BU (23.1 mg, 0.044 mmol) in DCM (5 mL) with 4 M HCl in dioxane (138 mg, 1.11 mol) and stirring the reaction mixture at room temperature for 5 hours. The reaction mixture was concentrated, dissolved in methanol (5 mL), and passed through an alkaline column to provide the desired compound (15.1 mg). 1H NMR(400MHz,DMSO-d6)δ8.80(s,1H),8.26(d,J=8.6Hz,1H),8.11–8.01(m,1 H),7.65(d,J=2.1Hz,1H),7.22(dd,J=8.7,2.1Hz,1H),7.00(dd,J=7.9,1.7H z,1H),6.92(dd,J=7.9,2.4Hz,1H),4.40–4.29(m,2H),4.25(s,3H),2.76–2 .63(m,4H),2.56(p,J=1.9Hz,2H),2.47–2.39(m,4H),2.33(p,J=1.9Hz,1H).

[0803] Example 101: 8-Fluoro-3-(2-Fluorobenzyl)-7-(Methoxymethyl)-5-methyl-3,5-dihydro-4H-pyridazin[4,5-b]indol-4-one

[0804]

[0805] Example 101 was generated from intermediate BR using a similar procedure to that used in Example 91. ¹H NMR (400MHz, DMSO-d6) δ 8.80 (s, ¹H), 8.08 (d, J = 10.2 Hz, ¹H), 7.82 (d, J = 5.9 Hz, ¹H), 7.34 (tdd, J = 7.4, 5.4, 2.0 Hz, ¹H), 7.29–7.09 (m, 3H), 5.46 (s, 2H), 4.68–4.60 (m, 2H), 4.29 (s, 3H), 3.40 (s, 3H).

[0806] Intermediate BU: 7-(bromomethyl)-8-fluoro-3-(2-fluorobenzyl)-5-methyl-3,5-dihydro-4H-pyridazin[4,5-b]indol-4-one

[0807]

[0808] Intermediate BU was generated from Example 101 using a similar procedure to that used to generate Example BS. ¹H NMR (400MHz, DMSO-d6) δ 8.80 (s, ¹H), 8.13 (d, J = 10.2 Hz, ¹H), 8.03 (d, J = 6.2 Hz, ¹H), 7.41–7.30 (m, ¹H), 7.30–7.07 (m, 3H), 5.46 (s, 2H), 4.91 (d, J = 1.1 Hz, 2H), 4.28 (s, 3H).

[0809] Example 102: 8-Fluoro-3-(2-Fluorobenzyl)-5-methyl-7-(morpholinomethyl)-3,5-dihydro-4H-pyridazin[4,5-b]indol-4-one

[0810]

[0811] Example 102 was generated from intermediate BU using a procedure similar to that used to generate Example 92. ¹H NMR (400MHz, DMSO-d⁶) δ 8.79 (s, ¹H), 8.06 (d, J = 10.1 Hz, ¹H), 7.78 (d, J = 5.9 Hz, ¹H), 7.41–7.31 (m, ¹H), 7.30–7.10 (m, 3H), 5.47 (s, 2H), 4.29 (s, 3H), 3.71 (s, 2H), 3.64–3.55 (m, 4H), 2.49–2.42 (m, 4H).

[0812] Example 103: 8-Fluoro-3-(2-Fluorobenzyl)-7-(((2-hydroxyethyl)amino)methyl)-5-methyl-3,5-dihydro-4H-pyridazin[4,5-b]indol-4-one

[0813]

[0814] Example 103 was generated from intermediate BU using a procedure similar to that used to generate Example 92. ¹H NMR (400MHz, DMSO-d⁶) δ 8.79 (s, ¹H), 8.05 (d, J = 10.2 Hz, ¹H), 7.88 (d, J = 6.0 Hz, ¹H), 7.42–7.32 (m, ¹H), 7.28–7.17 (m, 2H), 7.14 (td, J = 7.4, 1.2 Hz, ¹H), 5.47 (s, 2H), 4.63 (s, ¹H), 4.48–2.04 (bs, ¹H), 4.29 (s, 3H), 4.01 (s, 2H), 3.54 (q, J = 5.5 Hz, 2H), 2.77–2.70 (m, 2H).

[0815] Example 104: 8-Fluoro-3-(2-Fluorobenzyl)-5-methyl-7-(((tetrahydro-2H-pyran-4-yl)amino)methyl)-3,5-dihydro-4H-pyridazin[4,5-b]indol-4-one

[0816]

[0817] Example 104 was generated from intermediate BU using a procedure similar to that used to generate Example 92. ¹H NMR (400MHz, DMSO-d⁶) δ 8.78 (s, ¹H), 8.02 (d, J = 10.2 Hz, ¹H), 7.86 (d, J = 6.0 Hz, ¹H), 7.40–7.29 (m, ¹H), 7.27–7.10 (m, 3H), 5.47 (s, 2H), 4.28 (s, 3H), 3.96 (s, 2H), 3.91–3.79 (m, 2H), 3.30–3.23 (m, 2H), 2.74–2.63 (m, ¹H), 2.18 (s, ¹H), 1.83 (d, J = 13.0 Hz, 2H), 1.32 (tt, J = 20.4, 10.3 Hz, 2H).

[0818] Example 105: 8-Fluoro-3-(2-fluorobenzyl)-5-methyl-7-(pyrrolidone-1-ylmethyl)-3,5-dihydro-4H-pyridazin[4,5-b]indol-4-one

[0819]

[0820] Example 105 was generated from intermediate BU using a procedure similar to that used to generate Example 92. ¹H NMR (400MHz, DMSO-d⁶) δ 8.79 (s, ¹H), 8.04 (d, J = 10.1 Hz, ¹H), 7.77 (d, J = 5.9 Hz, ¹H), 7.40–7.29 (m, ¹H), 7.28–7.08 (m, 3H), 5.47 (s, 2H), 4.29 (s, 3H), 3.87–3.77 (m, 2H), 2.59–2.52 (m, 4H), 1.80–1.66 (m, 4H).

[0821] Example 106: 7-((dimethylamino)methyl)-8-fluoro-3-(2-fluorobenzyl)-5-methyl-3,5-dihydro-4H-pyridazin[4,5-b]indol-4-one

[0822]

[0823] Example 106 was generated from intermediate BU using a procedure similar to that used to generate Example 92. ¹H NMR (400MHz, DMSO-d⁶) δ 8.79 (s, ¹H), 8.05 (d, J = 10.2 Hz, ¹H), 7.76 (d, J = 5.9 Hz, ¹H), 7.38–7.30 (m, ¹H), 7.26–7.09 (m, 3H), 5.47 (s, 2H), 4.29 (s, 3H), 3.63 (d, J = 1.2 Hz, 2H), 2.23 (s, 6H).

[0824] Intermediate BV: 4-((8-fluoro-3-(2-fluorobenzyl)-5-methyl-4-oxoylide-4,5-dihydro-3H-pyridazino[4,5-b]indol-7-yl)methyl)-1,4-diazacycloheptane-1-carboxylic acid tert-butyl ester

[0825]

[0826] Intermediate BV was generated from intermediate BU using a procedure similar to that used to generate Example 92. ¹H NMR (400MHz, DMSO-d6) δ 8.79 (s, ¹H), 8.05 (d, J = 10.2 Hz, ¹H), 7.79 (d, J = 5.9 Hz, ¹H), 7.40–7.26 (m, ¹H), 7.26–7.06 (m, 3H), 5.47 (s, 2H), 4.29 (s, 3H), 3.85 (s, 2H), 3.46–3.35 (m, 4H), 2.76–2.59 (m, 4H), 1.76 (s, 2H), 1.40 (d, J = 7.0 Hz, 9H).

[0827] Example 107: 7-((1,4-diazacycloheptane-1-yl)methyl)-8-fluoro-3-(2-fluorobenzyl)-5-methyl-3,5-dihydro-4H-pyridazin[4,5-b]indol-4-one

[0828]

[0829] Example 107 was prepared by dissolving intermediate BV (43.8 mg, 0.082 mmol) in trifluoroacetic acid (3 mL). The reaction mixture was stirred at room temperature for 30 minutes and concentrated to provide the desired product (35 mg). ¹H NMR (400 MHz, DMSO-d6) δ 8.97 (s, 2H), 8.85 (s, 1H), 8.31–8.20 (m, 1H), 8.07–8.01 (m, 1H), 7.40–7.31 (m, 1H), 7.29–7.18 (m, 2H), 7.18–7.10 (m, 1H), 5.48 (s, 2H), 4.55 (s, 2H), 4.31 (s, 3H), 3.72–3.07 (m, 9H), 2.24–2.04 (m, 2H).

[0830] Intermediate BW: 4-((8-fluoro-3-(2-fluorobenzyl)-5-methyl-4-oxoylide-4,5-dihydro-3H-pyridazino[4,5-b]indol-7-yl)methyl)piperazine-1-carboxylic acid tert-butyl ester

[0831]

[0832] Intermediate BW was generated from intermediate BU using a procedure similar to that used to generate Example 92. ¹H NMR (400MHz, DMSO-d6) δ 8.79 (s, ¹H), 8.06 (d, J = 10.1 Hz, ¹H), 7.77 (d, J = 5.8 Hz, ¹H), 7.40–7.29 (m, ¹H), 7.28–7.10 (m, 3H), 5.47 (s, 2H), 4.29 (s, 3H), 3.73 (s, 2H), 3.41–3.32 (m, 4H), 2.46–2.39 (m, 4H), 1.39 (s, 9H).

[0833] Example 108: 8-Fluoro-3-(2-Fluorobenzyl)-5-methyl-7-(piperazin-1-ylmethyl)-3,5-dihydro-4H-pyridazin[4,5-b]indol-4-one

[0834]

[0835] Example 108 was generated from intermediate BW using a procedure similar to that used to generate Example 107. ¹H NMR (400MHz, DMSO-d6) δ 8.81 (s, ¹H), 8.66 (s, 2H), 8.13 (d, J = 10.1 Hz, ¹H), 7.87 (d, J = 5.9 Hz, 1H), 7.41–7.30 (m, ¹H), 7.26–7.08 (m, 3H), 5.47 (s, 2H), 4.30 (s, 3H), 4.00 (s, 2H), 3.24–3.15 (m, 4H), 2.95–2.79 (m, 4H).

[0836] Example 109: 5-Methyl-3-((6-methylpyridin-2-yl)methyl)-7-(2-morpholinoethoxy)-3,4a,5,9b-tetrahydro-4H-pyridazin[4,5-b]indol-4-one

[0837]

[0838] Example 109 was generated by treating Example 29 (30.3 mg, 0.095 mmol) in DMF (3 mL) with Cs₂CO₃ (92.5 mg, 0.284 mmol) and 4-(2-chloroethyl)-morpholine hydrochloride (17.6 mg, 0.095 mmol), and the resulting reaction mixture was allowed to be stirred overnight at room temperature. The reaction mixture was diluted with EtOAc, washed with water, dried, concentrated, and purified by FCC (0-100% EtOAc in hexane) to provide the desired product (32 mg). 1H NMR (400MHz, DMSO-d6) δ8.76(s,1H),8.08(d,J=8.7Hz,1H),7.64–7.53(m,1H),7.29(d,J=2.2Hz,1H),7.14(d,J=7.6Hz,1H),7.04(dd,J=8.8, 2.2Hz,1H),6.82(d,J=7.8Hz,1H),5.45(s,2H),4.29–4.22(m,5H),3.66–3.56(m,4H),2.81–2.73(m,2H),2.51(q,J=1.9Hz,4H),2.45(s,3H).

[0839] Example 110: 3-(2-fluorobenzyl)-5-methyl-7-(2-morpholinoethoxy)-3,4a,5,9b-tetrahydro-4H-pyridazin[4,5-b]indol-4-one

[0840]

[0841] Example 110 was generated from Example 46 using a procedure similar to that used to generate Example 109. ¹H NMR (400MHz, DMSO-d6) δ 8.75 (s, ¹H), 8.07 (d, J = 8.8 Hz, ¹H), 7.38–7.31 (m, ¹H), 7.29 (d, J = 2.2 Hz, ¹H), 7.27–7.09 (m, 3H), 7.03 (dd, J = 8.8, 2.2 Hz, 1H), 5.46 (s, 2H), 4.29–4.19 (m, 5H), 3.65–3.57 (m, 4H), 2.81–2.74 (m, 2H), 2.56–2.50 (m, 4H).

[0842] Example 2

[0843] Pharmacological Examples

[0844] Pyruvate kinase activity assay. Recombinant enzyme. A sequential enzyme-coupled assay using lactate dehydrogenase (LDH) and measuring NADH depletion by absorbance at 340 nm was used to determine pyruvate kinase activity. For the AC50 assay using ML-265 (the activator concentration required to achieve half-maximal activation), assays were performed in 96 wells using a 200 μL / well assay volume, with final concentrations of 20 nM human recombinant PKM2 (Sigma, SAE0021), different concentrations of ML-265, 0.5 mM PEP, 1 mM ADP, 0.2 mM NADH, and 8 U of lactate dehydrogenase (LDH) in assay buffers of 50 mM Tris-HCl (pH 7.4), 100 mM KCl, and 5 mM MgCl2. Decrease in absorbance at 340 nm was monitored using a SPECTROstar Omega microplate reader (BMG LABTECH Inc., Cary, NC, USA). Initial velocity was calculated using MARS software. The data were normalized to the PKM2 activity after DMSO (dimethyl sulfoxide) treatment.

[0845] Cell Culture. For 661W cell line experiments, change the medium before starting treatment with DMSO or ML-265. Incubate cells with DMSO or different concentrations of ML-265 for 2 hours. Lyse and homogenize cells in RIPA lysis and extraction buffer (catalog number: 89900, Life Technologies Corporation, Grand Island, NY) with protease inhibitor (Complete-Mini, Roche Diagnostics, Indianapolis, IN) and remove cell debris by centrifugation at 10,000 rpm for 10 minutes. Ten μL of supernatant was used to assess pyruvate kinase activity, and the activity was normalized to total protein content as previously described.

[0846] The results are shown in Table 1 below.

[0847] Table 1

[0848] Example EC50(nM) Maximum activation (%) Cellular EC50 (nM) Maximum activation (%) 1 117 212 15 431 2 106 90 3 428 193 28 310 4 527 267 22 309 5 192 226 13 295 6 7 63 222 18 355 8 117 271 25 324 9 10 11 110 170 12 131 123 13 14 NA NA 15 40 254 124 296 16 40 231 58 279 17 74 157 17 281 18 41 264 23 292 20 15 199 67 229 21 25 299 75 273 23 22 196 33 288 27 26 307 52 97 225 66 305 53 15 199 37 229 54 53 299 75 273 57 62 234 81 370 60 67 243 54 334 61 76 293 120 223 62 82 221 134 376 63 31 219 78 302 64 69 165 73 272 65 43 191 115 262 66 36 154 121 323 67 49 277 147 290 70 82 191 162 210 71 28 165 69 278 72 55 206 72 222 73 22 232 104 189 74 62 234 81 370 78 35 195 47 247 80 45 286 235 325 81 49 336 259 259 85 63 330 76 315 86 63 195 172 352 87 50 270 95 317 90 72 218 60 202

[0849] Example 3

[0850] Table 2 shows other compounds. Compounds can be synthesized using the methods described herein or other suitable synthetic protocols.

[0851] Table 2

[0852]

[0853]

[0854]

[0855]

[0856]

[0857]

[0858]

[0859]

[0860]

[0861]

[0862]

[0863]

[0864]

[0865]

[0866] All publications and patents mentioned in the foregoing specification are incorporated herein by reference. Various modifications and variations to the methods and systems described in this disclosure will be apparent to those skilled in the art without departing from the scope and spirit of this disclosure. Although this disclosure has been described in conjunction with specific preferred embodiments, it should be understood that the claimed disclosure should not be unduly limited to such specific embodiments. In fact, various modifications to the described modes used in carrying out this disclosure, which will be apparent to those skilled in the art, are intended to be within the scope of the following claims.

Claims

1. A composition comprising: or , Among them, R1 is selected from OCH3, OH, N(CH3)2, , and The group is formed, and R2 is chosen freely. , and A group that is formed.

2. The composition according to claim 1, comprising: ， Among them, R1 is selected from OCH3, OH, N(CH3)2, , and The group is formed, and R2 is chosen freely. , and A group that is formed.

3. The composition according to claim 1, wherein, The compound is selected from the group consisting of: , , , , , , , , , , , and .

4. The composition according to claim 3, wherein, The compound is selected from the group consisting of: , , , , , , , and .

5. The composition according to claim 3, wherein, The compound is selected from the group consisting of: , , and .

6. The composition according to claim 3, wherein, The compound is .

7. The composition according to claim 3, wherein, The compound is .

8. The composition according to claim 3, wherein, The compound is .

9. The composition according to claim 3, wherein, The compound is .

10. A composition comprising compounds selected from the group consisting of: , , , and .

11. The composition according to any one of claims 1-10, wherein, At least one hydrogen atom in the compound is replaced by deuterium.

12. The composition according to any one of claims 1-11, wherein, The composition is a pharmaceutical composition.

13. The composition according to any one of claims 1-12, wherein, The composition is formulated for injection, oral delivery, or as eye drops.

14. The composition according to any one of claims 1-12, wherein, The composition includes a pharmaceutically acceptable carrier.

15. The composition according to any one of claims 1-12, wherein, The composition is a pyruvate kinase activator.

16. The composition according to claim 15, wherein, The pyruvate kinase is selected from the group consisting of PKM1 and PKM2.

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