A polycyclic pyridazinone derivative as an SOS1 inhibitor, its preparation method and use

Abstract

Provided are polycyclic pyridazinone derivatives as SOS1 inhibitors, their preparation methods, and uses. The polycyclic pyridazinone derivatives have a structure as shown in Formula (I) and have significant effects on inhibiting the RAS signaling pathway and treating cancers, including pancreatic cancer, colorectal cancer, lung cancer, hepatocellular carcinoma, renal cancer, gastric cancer, and bile duct cancer.

Description

Technical Field

[0001] The present invention belongs to the technical field of polycyclic pyridazinone derivatives, and particularly relates to a polycyclic pyridazinone derivative as an SOS1 inhibitor, a preparation method and use thereof. Background Art

[0002] The RAS family of proteins belongs to a class of small GTPases, encompassing three subfamilies: KRAS, NRAS, and HRAS. Mutated RAS genes are important oncogenes, found in 20-30% of human tumors, particularly pancreatic, colorectal, and lung cancers. Each RAS subtype exists in a balance between a GTP-bound active state and a GDP-bound inactive state. GTPase-activating proteins (GAPs) convert GTP to GDP, shifting RAS proteins to an inactive state. Guanine nucleotide exchange factors (GEFs) facilitate GDP release and GTP binding, shifting RAS proteins to an active state. Activation of RAS proteins promotes cell proliferation, apoptosis evasion, and metabolic reprogramming through the RAS-RAF-MEK-ERK and RAS-PI3K-PDK1-AKT signaling pathways, thereby contributing to tumor development and progression.

[0003] SOS1 (son of sevenless 1) is a key guanine nucleotide exchange factor (GEF) that binds to RAS protein, promoting the binding of RAS protein to GTP and shifting RAS protein to an activated state. Recent studies have found that SOS1 inhibitors can not only inhibit the growth of RAS mutant cells, but also produce a synergistic effect with MEK inhibitors, significantly inhibiting KRAS-driven tumors. 1-2 The development of SOS1 inhibitors has become a research hotspot, and SOS1 inhibitors of different structural types have been reported in many patents, such as WO2018172250, WO2019201848, WO2018115380, WO2019122129, WO2020173935, WO2020180768 and WO2020180770.

[0004] However, there are still uncertainties in the effectiveness, safety or selectivity of the compounds and experimental drugs disclosed in these prior arts. Therefore, it is necessary to research and develop new selective SOS1 inhibitors.

[0005] References:

[0006] 1. Hillig et al. Discovery of poetent SOS1inhibitors that block RASactivation via disruption of the RAS-SOS1interaction. PNAS. 116, 2251-2560 (2019).

[0007] 2. Hofmann et al. BI-3406, a potent and selective SOS1::KRAS interactioninhibitor, is effective in KRAS-driven cancers through combined MEKinhibition. Cancer Discov. CD-20-0142 (2020). Summary of the Invention

[0008] In order to solve the above-mentioned problems of the prior art, the object of the present invention is to provide a polycyclic pyridazinone derivative, a pharmaceutically acceptable salt thereof, a tautomer or a stereoisomer thereof, so as to screen out compounds having excellent properties in terms of efficacy, safety and selectivity as SOS1 inhibitors.

[0009] Another object of the present invention is to provide a method for preparing the derivative, its pharmaceutically acceptable salt, its tautomer or its stereoisomer.

[0010] In order to achieve the purpose of the invention, the present invention adopts the following technical solutions:

[0011] In a first aspect, the present invention provides a polycyclic pyridazinone derivative, a pharmaceutically acceptable salt thereof, a tautomer thereof or a stereoisomer thereof, wherein the structure of the polycyclic pyridazinone derivative is shown in formula (I):

[0012]

[0013] Where: R 1 is selected from hydrogen or methyl;

[0014] R 2 Selected from C1-C3 alkyl, -OR 21, halogen, 3-7 membered cycloalkyl, 5-7 membered cycloalkenyl, 6-10 membered fused cycloalkyl, 7-10 membered bridged cycloalkyl, 7-10 membered spirocycloalkyl, 4-7 membered heterocyclyl, 5-7 membered heterocyclyl, 6-10 membered fused heterocyclyl, 7-10 membered bridged heterocyclyl, 7-10 membered spiroheterocyclyl, wherein 3-7 membered cycloalkyl, 5-7 membered cycloalkenyl, 6-10 membered fused cycloalkyl, 7-10 membered bridged cycloalkyl, 7-10 membered spirocycloalkyl, 4-7 membered heterocyclyl, 5-7 membered heterocyclyl, 6-10 membered fused heterocyclyl, 7-10 membered bridged heterocyclyl, 7-10 membered spiroheterocyclyl are optionally replaced by 1-3 R 22 replaced by;

[0015] R 21 is selected from H, C1-C3 alkyl, 3-7 membered cycloalkyl, 4-7 membered heterocyclic group, wherein C1-C3 alkyl, 3-7 membered cycloalkyl, 4-7 membered heterocyclic group is optionally substituted by 1-3 R 22 replaced by;

[0016] R 22 Selected from C1-C3 alkyl, hydroxy, halogen, cyano, -NR a R b 、C1-C3 alkoxy、-C(O)R a 、-C(O)OR a 、-OC(O)R a 、-NR b C(O)R a 、-NR b C(O)OR a 、-C(O)NR a R b , phenyl, 5-6 membered heteroaryl and =O, wherein the alkyl, alkoxy, phenyl, 5-6 membered heteroaryl are optionally further substituted with 1-3 halogen, C1-C3 alkyl, hydroxy, cyano, amino and C1-C3 alkoxy;

[0017] R a and R b Independently selected from H, substituted or unsubstituted C1-C3 alkyl, substituted or unsubstituted 3-6 membered cycloalkyl or substituted or unsubstituted 4-7 membered heterocyclyl; "substituted" herein means optionally substituted with 1-3 substituents selected from C1-C3 alkyl, hydroxy, halogen, cyano, amino or alkoxy;

[0018] Q is selected from N or -CR 3 ;

[0019] R 3 Selected from H, C1-C3 alkyl, halogen, cyano or -OR 21 ;

[0020] AR is selected from 6-10 membered aryl or 5-10 membered heteroaryl, wherein the aryl or heteroaryl is optionally substituted by 1-4 R c replaced by;

[0021] R c Selected from H, halogen, C1-C4 alkyl, C1-C4 haloalkyl, hydroxy-C1-C4 alkyl, hydroxy-C1-C4 haloalkyl, 3-6 membered cycloalkyl, 4-7 membered heterocyclyl, -OR 21 、-NR a R b NR a R b -C1-C4 alkyl, NR a R b -C1-C4 haloalkyl, 6-10 membered aryl or 5-10 membered heteroaryl, wherein the 6-10 membered aryl or 5-10 membered heteroaryl is optionally substituted by 1-4 R d replaced by;

[0022] R d Selected from H, halogen, C1-C4 alkyl, C1-C4 haloalkyl, hydroxy-C1-C4 alkyl, hydroxy-C1-C4 haloalkyl, 3-6 membered cycloalkyl, 4-7 membered heterocyclyl, -OR 21 、-NR a R b NR a R b -C1-C4 alkyl, NR a R b -C1-C4 haloalkyl;

[0023] The number of heteroatoms in the heterocyclic group, heteroaryl group, heterocycloalkenyl group, fused heterocyclic group, bridged heterocyclic group and spiro heterocyclic group in the formula (I) is 1-7 and is selected from one or more of oxygen, nitrogen, sulfur and S(O)m, where m is 1 or 2.

[0024] Preferably, the structure of the polycyclic pyridazinone derivative is as shown in formula (II):

[0025]

[0026] Among them, R 1 、R 2 , Q and R c With the same defined range as above; n=1-4 (eg, n=1, n=2, n=3, n=4).

[0027] Preferably, for the compound of formula (II), the phenyl group is optionally replaced by 1-3 R c When the R c When the number of R is 2-3, the R cCan be the same or different;

[0028] and / or, when the R c When it is a C1-C4 haloalkyl group, the halogen atom is fluorine;

[0029] and / or, when the R c When it is a halogen, the halogen atom is fluorine;

[0030] and / or, when the R c -NR a R b When the R a and R b Can be the same or different.

[0031] Preferably, the structure of the polycyclic pyridazinone derivative is as shown in formula (III):

[0032]

[0033] Among them, R 1 、R 2 and R c With the same defined range as above; n=1-4 (eg, n=1, n=2, n=3, n=4).

[0034] Preferably, for the compound of formula (III), the R 2 is optionally replaced by 1-3 R 22 When the substituted 4-7 membered heterocyclic group, the R 22 When it is 2-3, R 22 Same or different;

[0035] and / or, the R 2 is optionally replaced by 1-3 R 22 When the substituted 4-7 membered heterocyclic group contains 1-2 heteroatoms;

[0036] and / or, the R 2 is optionally replaced by 1-3 R 22 When the substituted 4-7 membered heterocyclic group is a substituted 4-7 membered heterocyclic group, the heteroatom of the heterocyclic group is nitrogen and / or oxygen;

[0037] and / or, the R 2 is optionally replaced by 1-3 R 22 In the case of a substituted 4-7 membered heterocyclic group, when the heterocyclic group has two heteroatoms, the two heteroatoms are the same or different;

[0038] and / or, the R 2 is optionally replaced by 1-3 R 22 When the substituted 4-7 membered heterocyclic group, the R 22Selected from C1-C3 alkyl, -NR a R b 、-C(O)R a 、-NR b C(O)R a 、-NR b C(O)OR a 、-C(O)NR a R b And = O.

[0039] Preferably, the structure of the polycyclic pyridazinone derivative is as shown in formula (IV):

[0040]

[0041] Among them, R 1 、R 2 、R 3 and R c With the same defined range as above; n=1-4 (eg, n=1, n=2, n=3, n=4).

[0042] Preferably, for the compound of formula (IV), the R 2 is optionally replaced by 1-3 R 22 When the substituted 4-7 membered heterocyclic group, the R 22 When it is 2-3, R 22 Same or different;

[0043] and / or, the R 2 is optionally replaced by 1-3 R 22 When the substituted 4-7 membered heterocyclic group contains 1-2 heteroatoms;

[0044] and / or, the R 2 is optionally replaced by 1-3 R 22 When the substituted 4-7 membered heterocyclic group is a substituted 4-7 membered heterocyclic group, the heteroatom of the heterocyclic group is nitrogen and / or oxygen;

[0045] and / or, the R 2 is optionally replaced by 1-3 R 22 In the case of a substituted 4-7 membered heterocyclic group, when the heterocyclic group has two heteroatoms, the two heteroatoms are the same or different;

[0046] and / or, the R 2 is optionally replaced by 1-3 R 22 When the substituted 4-7 membered heterocyclic group, the R 22 Selected from C1-C3 alkyl, -NR a R b 、-C(O)R a 、-NR bC(O)R a 、-NR b C(O)OR a 、-C(O)NR a R b and = O;

[0047] and / or, the R 3 For-OR 21 When R 21 is selected from unsubstituted C1-C3 alkyl or unsubstituted 3-7 membered cycloalkyl.

[0048] Preferably, the structure of the polycyclic pyridazinone derivative is as shown in formula (V):

[0049]

[0050] Among them, R 1 、R 2 , Q and R d With the same defined range as above; n=1-4 (eg, n=1, n=2, n=3, n=4).

[0051] Preferably, the R d 1-NR a R b When the R a and R b Independently selected from H, substituted or unsubstituted C1-C3 alkyl;

[0052] and / or, the R 2 is optionally replaced by 1-3 R 22 When the substituted 4-7 membered heterocyclic group, the R 22 When it is 2-3, R 22 Same or different;

[0053] and / or, the R 2 is optionally replaced by 1-3 R 22 When the substituted 4-7 membered heterocyclic group contains 1-2 heteroatoms;

[0054] and / or, the R 2 is optionally replaced by 1-3 R 22 When the substituted 4-7 membered heterocyclic group is a substituted 4-7 membered heterocyclic group, the heteroatom of the heterocyclic group is nitrogen and / or oxygen;

[0055] and / or, the R 2 is optionally replaced by 1-3 R 22 In the case of a substituted 4-7 membered heterocyclic group, when the heterocyclic group has two heteroatoms, the two heteroatoms are the same or different;

[0056] and / or, the R2 is optionally replaced by 1-3 R 22 When the substituted 4-7 membered heterocyclic group, the R 22 Selected from C1-C3 alkyl, -NR a R b 、-C(O)R a 、-NR b C(O)R a 、-NR b C(O)OR a 、-C(O)NR a R b and = O;

[0057] and / or, the R 2 For-OR 21 When R 21 is a 4-7 membered heterocyclic group;

[0058] and / or, the R 21 When it is a 4-7 membered heterocyclic group, the 4-7 membered heterocyclic group is a 5-6 membered heterocyclic group;

[0059] and / or, the R 21 When it is a 4-7 membered heterocyclic group, the heteroatom of the heterocyclic group is nitrogen and / or oxygen;

[0060] and / or, the R 21 When it is a 4-7 membered heterocyclic group, the heterocyclic group contains 1-2 heteroatoms;

[0061] and / or, the R 21 When it is a 4- to 7-membered heterocyclic group, when the heterocyclic group has two hetero atoms, the two hetero atoms are the same or different.

[0062] Preferably, the structure of the polycyclic pyridazinone derivative is as shown in formula (VI):

[0063]

[0064] Among them, R 1 、R 2 and R d With the same defined range as above; n=1-4 (eg, n=1, n=2, n=3, n=4).

[0065] Preferably, when the R 2 For-OR 21 , the R 21 is a 4-7 membered heterocyclic group;

[0066] and / or, when the R 21 When it is a 4-7 membered heterocyclic group, the 4-7 membered heterocyclic group is a 5-7 membered heterocyclic group;

[0067] and / or, when the R 21 When it is a 4-7 membered heterocyclic group, the heteroatom of the heterocyclic group is nitrogen and / or oxygen;

[0068] and / or, when the R 21 When it is a 4-7 membered heterocyclic group, the heterocyclic group contains two heteroatoms;

[0069] and / or, when the R 21 When it is a 4-7 membered heterocyclic group, the two heteroatoms are the same or different;

[0070] and / or, the R d Selected from halogen, C1-C4 alkyl, -NR a R b 、-OR 21 NR a R b -C1-C4 alkyl;

[0071] and / or, the R d 1-NR a R b -C1-C4 alkyl, the R a and R b Independently selected from H, substituted or unsubstituted C1-C3 alkyl.

[0072] Preferably, the structure of the polycyclic pyridazinone derivative is as shown in formula (VII):

[0073]

[0074] Among them, R 1 、R 2 、R 3 and R d With the same defined range as above; n=1-4 (eg, n=1, n=2, n=3, n=4).

[0075] Preferably, when the R 2 For-OR 21 , the R 21 is a 4-7 membered heterocyclic group;

[0076] and / or, when the R 21 When it is a 4-7 membered heterocyclic group, the 4-7 membered heterocyclic group is a 5-7 membered heterocyclic group;

[0077] and / or, when the R 21 When it is a 4-7 membered heterocyclic group, the heteroatom of the heterocyclic group is nitrogen and / or oxygen;

[0078] and / or, when the R 21When it is a 4-7 membered heterocyclic group, the heterocyclic group contains two heteroatoms;

[0079] and / or, when the R 21 When it is a 4-7 membered heterocyclic group, the two heteroatoms are the same or different;

[0080] and / or, the R d Selected from halogen, C1-C4 alkyl, -NR a R b 、-OR 21 NR a R b -C1-C4 alkyl;

[0081] and / or, the R d 1 NR a R b -C1-C4 alkyl, the R a and R b Independently selected from H, substituted or unsubstituted C1-C3 alkyl.

[0082] Further preferably, the polycyclic pyridazinone derivative is selected from any one of the following structures:

[0083]

[0084]

[0085] Typical compounds of the present invention include but are not limited to the compounds in the following table:

[0086]

[0087]

[0088]

[0089]

[0090]

[0091]

[0092] In a second aspect, the present invention provides a method for preparing the polycyclic pyridazinone derivatives, pharmaceutically acceptable salts thereof, tautomers thereof or stereoisomers thereof as described in the first aspect, which is selected from one of the following two schemes:

[0093] Option 1

[0094] The method for preparing the compound of general formula (I) of the present invention or its stereoisomers, tautomers or pharmaceutically acceptable salts thereof comprises the following steps:

[0095] Preparation of key intermediate (IA):

[0096]

[0097] In the first step, an aromatic compound of formula (I-1) is subjected to metal-catalyzed cross-coupling to obtain a compound of formula (I-2);

[0098] In the second step, the compound of general formula (I-2) reacts under catalyst conditions to obtain a chiral sulfonimide compound of general formula (I-3);

[0099] In the third step, the compound of general formula (I-3) is reduced with a metal reducing agent to obtain a chiral compound of general formula (I-4);

[0100] In the fourth step, the compound of general formula (I-4) is subjected to sulfonamide cleavage under acidic conditions to obtain a chiral benzylamine compound of general formula (IA);

[0101] Among them, X is halogen, preferably bromine.

[0102] Preparation of key intermediate (IB), method 1:

[0103]

[0104] In the first step, the compound of general formula (I-5) is subjected to a diazotization reaction to obtain a compound of general formula (I-6);

[0105] In the second step, the compound of general formula (I-6) and cuprous cyanide are subjected to a substitution reaction to obtain a compound of general formula (I-7);

[0106] In the third step, the cyano group in the compound of formula (I-7) is converted into an amidine, which is then subjected to an ester exchange reaction with the ester group to obtain a compound of formula (IB);

[0107] Among them, X, X 1 is halogen, X is preferably bromine, X 1 Preferably iodine;

[0108] Preparation of key intermediate (IB), method 2:

[0109]

[0110] In the first step, the compound of general formula (I-12) is subjected to a halogenation reaction to obtain a compound of general formula (I-13);

[0111] In the second step, the compound of general formula (I-13) is subjected to an esterification reaction to obtain a compound of general formula (I-6);

[0112] In the third step, the compound of general formula (I-6) is subjected to a substitution reaction to obtain a compound of general formula (I-14);

[0113] In the fourth step, the compound of the general formula (I-14) is subjected to an oxidation reaction to obtain a compound of the general formula (I-15);

[0114] In the fifth step, the compound of the general formula (I-15) is reacted with hydroxylamine sulfonic acid to obtain a compound of the general formula (I-16);

[0115] Step 6: The compound of formula (I-16) is subjected to an Abnormal Beckmann rearrangement reaction to obtain a compound of formula (I-7);

[0116] In the seventh step, the cyano group in the compound of general formula (I-7) is converted into an amidine, which is then subjected to an ester exchange reaction with the ester group to obtain a compound of general formula (IB);

[0117] Among them, X, X 1 is halogen, X is preferably chlorine, X 1 Preferably iodine;

[0118] Preparation of general formula (I):

[0119]

[0120] In the first step, the compound of general formula (IA) and the compound of general formula (IB) are subjected to an imine addition reaction to obtain a compound of general formula (I-8);

[0121] In the second step, the compound of the general formula (I-8) and hydrazine hydrate are subjected to an imine addition reaction and then a ring expansion reaction to obtain a compound of the general formula (I-9);

[0122] In the third step, the compound of general formula (I-9) and the compound of general formula (I-10) are subjected to substitution reaction to obtain the compound of general formula (I-11) (such as R 1 =H, this step is omitted);

[0123] In the fourth step, the compound of the general formula (I-11) and the compound of the general formula (I-25) are reacted under alkaline conditions in the presence of a metal catalyst and a ligand to obtain a compound of the general formula (I);

[0124] Among them, X, X 1 is halogen, X is preferably bromine and chlorine, X 1 Preferably iodine; W is H, Q, AR, R 1 and R 2 With the same limitations as above.

[0125] Option 2

[0126] The method for preparing the compound of general formula (I) of the present invention or its stereoisomers, tautomers or pharmaceutically acceptable salts thereof comprises the following steps:

[0127]

[0128] In the first step, the compound of general formula (I-17) is subjected to a substitution reaction to obtain a compound of general formula (I-18);

[0129] In the second step, the compound of general formula (I-18) is deprotected under acidic conditions to obtain a compound of general formula (I-19);

[0130] In the third step, the compound of general formula (I-19) and the compound of general formula (I-26) are reacted under alkaline conditions to obtain the compound of general formula (I-20);

[0131] In the fourth step, the compound of the general formula (I-20) is subjected to an oxidation reaction to obtain a compound of the general formula (I-21);

[0132] In the fifth step, the compound of the general formula (I-21) is subjected to Bouveault aldehyde synthesis reaction to obtain the compound of the general formula (I-22);

[0133] Step 6: The compound of formula (I-22) and hydrazine hydrate undergo an addition cyclization reaction to obtain a compound of formula (I-23);

[0134] In the seventh step, the compound of the general formula (I-23) is subjected to a substitution reaction to obtain a compound of the general formula (I-24);

[0135] In the eighth step, the compound of the general formula (I-24) and the compound of the general formula (I-10) are subjected to a substitution reaction to obtain a compound of the general formula (I-Bb) (such as R 1 =H, this step is omitted);

[0136] In the ninth step, the compound of formula (I-Bb) and the compound of formula (IA) are reacted under alkaline conditions in the presence of a metal catalyst and a ligand to obtain a compound of formula (I);

[0137] Among them, X 2 、X 3 、X 4 is halogen, X 2 、X 3 Preferably bromine, X 4 Preferably iodine; Q is selected from N or CR 3 ; R 3 Selected from H, C1-C3 alkyl, halogen, cyano or -OR 21 ; R 1 Selected from hydrogen or methyl; AR and R 2 With the same limitations as above.

[0138] For the above preparation method, the reagent providing alkaline conditions is selected from an organic base or an inorganic base, the organic base is triethylamine, N,N-diisopropylethylamine, n-butyllithium, lithium diisopropylamide, lithium bistrimethylsilylamide, sodium tert-butoxide, sodium methoxide and potassium tert-butoxide, and the inorganic base is sodium hydride, potassium phosphate, sodium carbonate, potassium carbonate, potassium acetate, cesium carbonate, sodium hydroxide, potassium hydroxide, sodium bicarbonate and lithium hydroxide. One or more;

[0139] The reagent providing acidic conditions is one or more of hydrogen chloride, a 1,4-dioxane solution of hydrogen chloride, a methanol solution of hydrogen chloride, trifluoroacetic acid, formic acid, acetic acid, hydrochloric acid, sulfuric acid, methanesulfonic acid, nitric acid and phosphoric acid;

[0140] The metal catalyst is one or more of palladium / carbon, Raney nickel, tetrakistriphenylphosphine palladium, palladium dichloride, palladium acetate, [1,1'-bis(diphenylphosphino)ferrocene]dichloropalladium (Pd(dppf)Cl2), [1,1'-bis(diphenylphosphino)ferrocene]dichloropalladium dichloromethane complex, bistriphenylphosphine palladium dichloride (Pd(PPh3)Cl2) and tris(dibenzylideneacetone)dipalladium (Pd2(dba)3);

[0141] The ligand is one or more of 2-dicyclohexylphosphino-2,6'-dimethoxybiphenyl (SPhos), 4,5-bisdiphenylphosphino-9,9-dimethylxanthene (XantPhos), 2-dicyclohexylphosphino-2,4,6-triisopropylbiphenyl (XPhos), 2-dicyclohexylphosphino-2'-(N,N-dimethylamino)-biphenyl (DavePhos), 1,1'-bis(diphenylphosphino)ferrocene (Dppf) and 1,1'-binaphthyl-2,2'-bisdiphenylphosphine (BINAP), preferably 1,1'-binaphthyl-2,2'-bisdiphenylphosphine (BINAP);

[0142] The reducing agent is one or more of sodium borohydride, potassium borohydride, sodium cyanoborohydride, sodium triacetoxyborohydride, and lithium aluminum tetrahydride;

[0143] The oxidizing agent is one or more of potassium permanganate, manganese dioxide, potassium dichromate, sodium dichromate and potassium osmate;

[0144] The above reaction is preferably carried out in a solvent, and the solvent used is one or more of N,N-dimethylformamide, N-methylpyrrolidone, dimethyl sulfoxide, 1,4-dioxane, water, tetrahydrofuran, dichloromethane, 1,2-dichloroethane, methanol, ethanol, toluene, petroleum ether, ethyl acetate, n-hexane and acetone.

[0145] In a third aspect, the present invention provides a pharmaceutical composition comprising the polycyclic pyridazinone derivative according to the first aspect, a pharmaceutically acceptable salt thereof, a tautomer thereof or a stereoisomer thereof;

[0146] Preferably, the pharmaceutical composition further comprises a pharmaceutically acceptable carrier and / or excipient.

[0147] In a fourth aspect, the present invention provides a use of the polycyclic pyridazinone derivative according to the first aspect, a pharmaceutically acceptable salt thereof, a tautomer or a stereoisomer thereof, or the pharmaceutical composition according to the third aspect in the preparation of a drug for treating cancer or in the preparation of an SOS1 inhibitor;

[0148] Preferably, the cancer is pancreatic cancer, colorectal cancer, lung cancer, hepatocellular carcinoma, renal cancer, gastric cancer or bile duct cancer.

[0149] In a fifth aspect, the present invention provides a method for preventing and / or treating cancer, comprising administering to a human a therapeutically effective amount of the polycyclic pyridazinone derivative as described in the first aspect, a pharmaceutically acceptable salt thereof, a tautomer or a stereoisomer thereof, or the pharmaceutical composition as described in the third aspect.

[0150] Explanation of terms

[0151] Unless otherwise stated, some of the terms used in the specification and claims of the present invention are defined as follows:

[0152] "Alkyl" refers to a saturated aliphatic hydrocarbon group, comprising 1-20 carbon atoms, or 1-10 carbon atoms, or 1-6 carbon atoms, or 1-4 carbon atoms, or 1-3 carbon atoms, or 1-2 carbon atoms, a saturated straight or branched monovalent hydrocarbon group, wherein the alkyl group may be independently optionally substituted with one or more substituents described herein. Further examples of alkyl groups include, but are not limited to, methyl, ethyl, n-propyl, isopropyl, n-butyl, isobutyl, tert-butyl, sec-butyl, n-pentyl, 1,1-dimethylpropyl, 1,2-dimethylpropyl, 2,2-dimethylpropyl, 1-ethylpropyl, 2-methylbutyl, 3-methylbutyl, n-hexyl, 1-ethyl-2-methylpropyl, 1,1,2-trimethylpropyl, 1,1-dimethylbutyl, 1,2-dimethylbutyl, 2,2-dimethylbutyl, 1,3-dimethylbutyl, 2-ethylbutyl, 2-methylpentyl, 3-methylpentyl, 4-methylpentyl, 2,3-dimethylbutyl, etc. The alkyl group can be optionally substituted or unsubstituted.

[0153] "Alkenyl" refers to a linear or branched monovalent hydrocarbon group of 2 to 12 carbon atoms, or 2 to 8 carbon atoms, or 2 to 6 carbon atoms, or 2 to 4 carbon atoms, wherein at least one C—C is sp 2Double bond, wherein the alkenyl group can be independently optionally substituted with one or more substituents described in the present invention, specific examples of which include, but are not limited to, vinyl, allyl, and butylene, etc. The alkenyl group can be optionally substituted or unsubstituted.

[0154] "Cycloalkyl" refers to a saturated or partially unsaturated monocyclic or polycyclic hydrocarbon substituent, wherein the cycloalkyl ring comprises 3 to 20 carbon atoms, preferably 3 to 12 carbon atoms, and more preferably 3 to 6 carbon atoms. Non-limiting examples of monocyclic cycloalkyls include, but are not limited to, cyclopropyl, cyclobutyl, cyclopentyl, cyclopentenyl, cyclohexyl, cyclohexenyl, cyclohexadienyl, cycloheptyl, cycloheptatrienyl, cyclooctyl, and the like; polycyclic cycloalkyls include spirocyclic, fused, and bridged cycloalkyls. Cycloalkyls may be optionally substituted or unsubstituted.

[0155] "Spiroalkyl" refers to a polycyclic group with 5 to 18 members, two or more ring structures, and one carbon atom (called spiro atom) shared between the monocyclic rings, containing one or more double bonds in the ring, but no ring has a completely conjugated π electron aromatic system. Preferably, it is 6 to 14 members, more preferably 7 to 10 members. Spiroalkyl is divided into single spiro, double spiro or multiple spiroalkyl according to the number of shared spiro atoms between the rings, preferably single spiro and double spiroalkyl, preferably 4 / 5 members, 4 / 6 members, 5 / 5 members or 5 / 6 members. Non-limiting examples of "spiroalkyl" include, but are not limited to:

[0156]

[0157] "Fused cycloalkyl" refers to a 5- to 18-membered, all-carbon polycyclic group containing two or more cyclic structures sharing a pair of carbon atoms. One or more rings may contain one or more double bonds, but no ring has a completely conjugated π electron aromatic system. It is preferably 6- to 12-membered, and more preferably 7- to 10-membered. Depending on the number of constituent rings, it can be classified as a bicyclic, tricyclic, tetracyclic or polycyclic fused cycloalkyl, preferably a bicyclic or tricyclic, and more preferably a 5-membered / 5-membered or 5-membered / 6-membered bicyclic alkyl. Non-limiting examples of "fused cycloalkyl" include, but are not limited to:

[0158]

[0159] "Bridged cycloalkyl" refers to a 5- to 18-membered, all-carbon polycyclic group containing two or more cyclic structures that share two carbon atoms that are not directly connected to each other. One or more rings may contain one or more double bonds, but no ring has a completely conjugated π electron aromatic system. It is preferably 6- to 12-membered, and more preferably 7- to 10-membered. Depending on the number of constituent rings, it can be classified as a bicyclic, tricyclic, tetracyclic or polycyclic bridged cycloalkyl group, preferably a bicyclic, tricyclic or tetracyclic group, and more preferably a bicyclic or tricyclic group. Non-limiting examples of "bridged cycloalkyl" include, but are not limited to:

[0160]

[0161] The cycloalkyl ring can be fused to an aryl, heteroaryl or heterocyclyl ring, wherein the ring attached to the parent structure is a cycloalkyl, non-limiting examples of which include indanyl, tetrahydronaphthyl, benzocycloheptanyl and the like.

[0162] "Heterocyclyl," "heterocycle," or "heterocyclic" are used interchangeably herein to refer to a saturated or partially unsaturated monocyclic, bicyclic, or tricyclic non-aromatic heterocyclic group containing 3 to 12 ring atoms, wherein at least one ring atom is a heteroatom, such as an oxygen, nitrogen, or sulfur atom. Preferably, the heterocyclic group has a 5- to 7-membered monocyclic ring or a 7- to 10-membered bicyclic or tricyclic ring, which may contain 1, 2, or 3 atoms selected from nitrogen, oxygen, and / or sulfur. Examples of "heterocyclyl" include, but are not limited to, morpholinyl, oxetanyl, thiomorpholinyl, tetrahydropyranyl, 1,1-dioxo-thiomorpholinyl, piperidinyl, 2-oxo-piperidinyl, pyrrolidinyl, 2-oxo-pyrrolidinyl, piperazin-2-one, 8-oxa-3-aza-bicyclo[3.2.1]octyl, and piperazinyl. The heterocyclyl ring may be fused to an aryl, heteroaryl or cycloalkyl ring, wherein the ring attached to the parent structure is the heterocyclyl. The heterocyclyl group may be optionally substituted or unsubstituted.

[0163] "Spiro heterocyclyl" refers to a polycyclic group with 5 to 18 members, two or more ring structures, and one atom shared between the rings, containing one or more double bonds in the ring, but no ring has a completely conjugated π electron aromatic system, wherein one or more ring atoms are selected from nitrogen, oxygen, sulfur or S(O) m The heteroatom of m=1 or 2, the remaining ring atoms are carbon, and m=1 or 2. Preferably it is 6 to 14 members, more preferably 7 to 10 members. According to the number of spiro atoms shared between the rings, the spiro heterocyclic group is divided into a monospiro heterocyclic group, a bispiro heterocyclic group or a polyspiro heterocyclic group, preferably a monospiro heterocyclic group and a bispiro heterocyclic group. More preferably, it is a 4-membered / 4-membered, 4-membered / 5-membered, 4-membered / 6-membered, 5-membered / 5-membered or 5-membered / 6-membered monospiro heterocyclic group. Non-limiting examples of "spiro heterocyclic group" include, but are not limited to:

[0164]

[0165] "Fused heterocyclic group" refers to an all-carbon polycyclic group containing two or more ring structures sharing a pair of atoms, one or more rings may contain one or more double bonds, but no ring has a completely conjugated π-electron aromatic system, wherein one or more ring atoms are selected from nitrogen, oxygen, sulfur or S(O) mThe heteroatom of the ring is 1, the remaining ring atoms are carbon, and m = 1 or 2. Preferably, it is 6 to 14 members, more preferably 7 to 10 members. According to the number of constituent rings, it can be divided into a bicyclic, tricyclic, tetracyclic or polycyclic fused heterocyclic group, preferably a bicyclic or tricyclic group, more preferably a 5-membered / 5-membered or 5-membered / 6-membered bicyclic fused heterocyclic group. Non-limiting examples of "fused heterocyclic group" include but are not limited to:

[0166]

[0167] "Bridged heterocyclic group" refers to a polycyclic group of 5 to 18 members, containing two or more ring structures, sharing two atoms that are not directly connected to each other, one or more rings may contain one or more double bonds, but no ring has a completely conjugated π-electron aromatic system, wherein one or more ring atoms are selected from nitrogen, oxygen, sulfur or S(O) m The heteroatom of the cyclic ring is 1 or 2, and the remaining ring atoms are carbon atoms, and m = 1 or 2. It is preferably 6 to 14-membered, more preferably 7 to 10-membered. Depending on the number of constituent rings, it can be classified as a bicyclic, tricyclic, tetracyclic or polycyclic bridged heterocyclic group, preferably a bicyclic, tricyclic or tetracyclic group, and more preferably a bicyclic or tricyclic group. Non-limiting examples of "bridged heterocyclic groups" include, but are not limited to:

[0168]

[0169] "Aryl" refers to a carbocyclic aromatic system containing one or two rings, wherein the rings may be joined together in a fused manner. The term "aryl" includes aromatic groups such as phenyl, naphthyl, and tetrahydronaphthyl. Preferably, aryl is C6-C 10 Aryl, more preferably phenyl and naphthyl, most preferably phenyl. Aryl can be substituted or unsubstituted. The "aryl" can be fused with a heteroaryl, a heterocyclic group or a cycloalkyl group, wherein the aryl ring is connected to the parent structure. Non-limiting examples include but are not limited to:

[0170]

[0171] "Heteroaryl" refers to an aromatic 5 to 6-membered monocyclic or 9 to 10-membered bicyclic ring, which may contain 1 to 4 atoms selected from nitrogen, oxygen and / or sulfur. The embodiment of "heteroaryl" includes, but is not limited to, furyl, pyridyl, 2-oxo-1,2-dihydropyridyl, pyridazinyl, pyrimidinyl, pyrazinyl, thienyl, isoxazolyl, oxazolyl, oxadiazolyl, imidazolyl, pyrrolyl, pyrazolyl, triazolyl, tetrazolyl, thiazolyl, isothiazolyl, 1,2,3-thiadiazolyl, benzodioxolyl, benzimidazolyl, indolyl, isoindolyl, 1,3-dioxo-isoindolyl, quinolyl, indazolyl, benzisothiazolyl, benzoxazolyl and benzisoxazolyl. Heteroaryl can be optionally substituted or unsubstituted. The heteroaryl ring may be fused to an aryl, heterocyclyl or cycloalkyl ring, wherein the ring connected to the parent structure is a heteroaryl ring, non-limiting examples include but are not limited to:

[0172]

[0173] "Alkoxy" refers to a group (alkyl-O-). Alkyl is defined herein. C1-C6 alkoxy groups are preferred. Examples include, but are not limited to, methoxy, ethoxy, n-propoxy, isopropoxy, n-butoxy, isobutoxy, and tert-butoxy.

[0174] "Haloalkyl" refers to an alkyl group having one or more halogen substituents, wherein the alkyl group has the meaning as defined herein. Examples of haloalkyl groups include, but are not limited to, fluoromethyl, difluoromethyl, trifluoromethyl, perfluoroethyl, 1,1-dichloroethyl, 1,2-dichloropropyl, and the like.

[0175] "Hydroxy" refers to an -OH group.

[0176] "Halogen" refers to fluorine, chlorine, bromine and iodine, with fluorine, chlorine and bromine being preferred.

[0177] "Amino" refers to -NH2.

[0178] "Cyano" refers to -CN.

[0179] "Nitro" refers to -NO2.

[0180] "Benzyl" refers to -CH2-phenyl.

[0181] "Carboxyl" refers to -C(O)OH.

[0182] "Acetyl" refers to -C(O)CH3 or Ac.

[0183] "Carboxylate" refers to -C(O)O(alkyl) or (cycloalkyl), wherein alkyl and cycloalkyl are as defined above.

[0184] "Optional" means that the event it describes may but need not occur. For example, "AR is optionally replaced by 1 to more R c The term "substituted" implies that the AR group may be replaced by one or more R c Replaced or not by R c Replacement situation.

[0185] "Substituted" means that one or more hydrogen atoms, preferably up to 5, more preferably 1-3 hydrogen atoms, in a group are replaced independently of one another by a corresponding number of substituents. It goes without saying that the substituents are only in their possible chemical positions, and those skilled in the art can determine (by experiment or theory) possible or impossible substitutions without undue effort. For example, an amino or hydroxyl group with free hydrogen may be unstable when combined with a carbon atom with an unsaturated (e.g., olefinic) bond.

[0186] As used herein, "substituted" or "substituted", unless otherwise specified, means that a group may be substituted by one or more groups selected from the following groups: alkyl, alkenyl, alkynyl, alkoxy, alkylthio, alkylamino, halogen, mercapto, hydroxy, nitro, cyano, cycloalkyl, heterocyclyl, aryl, heteroaryl, cycloalkoxy, heterocycloalkyloxy, cycloalkylthio, heterocycloalkylthio, amino, haloalkyl, hydroxyalkyl, carboxyl, carboxylate, =O, -C(O)R b 、-OC(O)R b 、-NR b R b 、-C(O)NR b R b 、-NR b C(O)R b 、-S(O)NR b R b or -S(O)2NR b R b , where R b The definition of is as described in the general formula (I).

[0187] As used herein, the terms "subject," "individual," or "patient" are used interchangeably and refer to any animal, including mice, rats, other rodents, rabbits, dogs, cats, pigs, cattle, sheep, horses, primates, and humans. In some embodiments, the patient is a human. In some embodiments, the subject has experienced and / or displayed at least one symptom of the disease or disease to be treated and / or prevented. In some embodiments, the subject has been determined or diagnosed with a cancer having a KRAS G12 or G13 mutation (e.g., as determined by an FDA-approved regulatory agency, such as an FDA-approved assay or kit). In some embodiments, the subject has a tumor that is positive for a KRAS G12C mutation, a KRAS G12D mutation, a KRAS G12S mutation, a KRAS G12V mutation, a KRAS G12A mutation, a KRAS G13D mutation, or a KRAS G13C mutation (e.g., as determined by a regulatory agency-approved assay or kit). The research subject can be a patient with a tumor that is positive for a KRAS G12C mutation, a KRAS G12D mutation, a KRAS G12V mutation, a KRASG12S mutation, a KRAS G12A mutation, a KRAS G13D mutation, or a KRAS G13C mutation (e.g., an approved regulatory agency-such as an FDA-approved, assay, or kit). The subject can be a subject whose tumor has a KRAS G12C mutation, a KRAS G12D mutation, a KRAS G12V mutation, a KRAS G12S mutation, a KRAS G12A mutation, a KRAS G13D mutation, or a KRAS G13C mutation (e.g., the tumor is determined by an FDA-approved regulatory agency, kit, or assay). In some embodiments, the subject is suspected of having a cancer associated with the KRAS G12 or G13 gene. In some embodiments, the subject has a clinical record indicating that the subject has a tumor with a KRAS G12C mutation (and an optional clinical record indicating that the subject should be treated with any composition provided herein).

[0188] As used herein, the term "pediatric patient" refers to a patient under 16 years of age at the time of diagnosis or treatment. The term "child" can be further divided into the following subcategories: neonates (from birth to the first month of life); infants (1 month to 2 years); children (2 years to 12 years); and adolescents (12 years to 21 years (up to but not including the 22nd birthday)). Berhman RE, Kliegman R, Arvin AM, Nelson WE. Nelson Textbook of Pediatrics, 15th ed. Philadelphia: WB Saunders, Inc., 1996; Rudolph AM, et al. Rudolph's Pediatrics, 21st ed. New York: McGrow-Hill, 2002; and Avery MD, First LR. Pediatric Medicine, 2nd ed. Baltimore: Williams & Wilkins; 1994.

[0189] As used herein, an "effective amount" of a compound refers to an amount sufficient to negatively regulate or inhibit SOS 1 enzyme activity.

[0190] As used herein, a "therapeutically effective dose" of a compound is an amount sufficient to ameliorate or in some way reduce symptoms, halt or reverse disease progression, or downregulate or inhibit the activity of SOS 1. Such a dose can be used as a single dose or taken according to a regimen to be effective.

[0191] As used herein, "treat" or "treating" means ameliorating or otherwise altering in any way the symptoms or pathology of a patient's condition, disorder or disease.

[0192] As used herein, "amelioration of the symptoms of a particular disease by administration of a particular compound or pharmaceutical composition" refers to any reduction, whether permanent or temporary, long-lasting or transient, attributable to or associated with the administration of that composition.

[0193] The definitions and conventions used in this invention for stereochemistry are generally based on the following references:

[0194] SP Parker, Ed., McGraw-Hill Dictionary of Chemical Terms (1984) McGraw-Hill Book Company, New York; and Eliel, E. and Wilen, S., "Stereochemistry of Organic Compounds", John Wiley & Sons, Inc., New York, 1994. The compounds of the present invention may contain asymmetric centers or chiral centers and therefore exist as different stereoisomers. All stereoisomeric forms of the compounds of the present invention, including but not limited to diastereomers, enantiomers, atropisomers, and mixtures thereof, such as racemic mixtures, constitute part of the present invention. Diastereomers can be separated into individual diastereomers on the basis of their physical chemical differences by methods such as chromatography, crystallization, distillation, or sublimation. Enantiomers can be separated by converting a chiral isomeric mixture into a diastereomeric mixture by reacting the diastereomers with an appropriate optically active compound (e.g., a chiral auxiliary, such as a chiral alcohol or Mosher's acid chloride) and converting the individual diastereomers into the corresponding pure enantiomers. The intermediates and compounds of the present invention may also exist in different tautomeric forms, and all such forms are encompassed by the present invention. Many organic compounds exist in optically active forms, meaning they have the ability to rotate the plane of plane-polarized light. When describing optically active compounds, the prefixes D, L, R, and S are used to denote the absolute configuration of the chiral center of the molecule. The prefixes d, l, (+), and (-) are used to designate the sign of rotation of plane-polarized light in the compound, with (-) or l indicating that the compound is levorotatory and the prefix (+) or d indicating that the compound is dextrorotatory. These stereoisomers have the same order of attachment of their atoms or groups of atoms, but differ in their stereostructure. Specific stereoisomers may be enantiomers, and mixtures of isomers are often referred to as enantiomeric mixtures. A 50:50 mixture of enantiomers is called a racemic mixture or racemate, which can result in a lack of stereoselectivity or stereospecificity during chemical reactions. The terms "racemic mixture" and "racemate" refer to an equimolar mixture of two enantiomers that lacks optical activity.

[0195] "Tautomers" or "tautomeric forms" refer to structural isomers of different energies that can be interconverted via a low energy barrier. For example, proton tautomers (i.e., prototropic tautomers) include interconversions via proton migration, such as keto-enol and imine-enamine isomerizations. Valence (chemical valence) tautomers include interconversions by reorganization of bonding electrons. Unless otherwise indicated, the structural formulas described herein include all isomeric forms (e.g., enantiomers, diastereomers, and geometric isomers): for example, R and S configurations containing asymmetric centers, (Z) and (E) isomers of double bonds, and (Z) and (E) conformational isomers. Therefore, single stereochemical isomers of the compounds of the present invention or mixtures of their enantiomers, diastereomers, or geometric isomers are within the scope of the present invention.

[0196] "Pharmaceutically acceptable salts" refer to salts of the compounds of the present invention that are safe and effective for use in humans or animals. Salts of the compounds can be prepared by using a sufficient amount of a base or acid in a pure solution or a suitable inert solvent to form the corresponding addition salt. Pharmaceutically acceptable base addition salts include sodium, potassium, calcium, ammonium, organic amino or magnesium salts, and pharmaceutically acceptable acid addition salts include inorganic and organic acid salts, such as hydrochloric acid, hydrobromic acid, carbonic acid, bicarbonate, phosphoric acid, monohydrogen phosphate, dihydrogen phosphate, sulfuric acid, monohydrogen sulfate, acetic acid, maleic acid, malonic acid, succinic acid, butenedioic acid, phthalic acid, benzenesulfonic acid, p-toluenesulfonic acid, citric acid, and methanesulfonic acid (see Berge et al., "Pharmaceutical Salts," Journal of Pharmaceutical Science 66:1-19 (1977)).

[0197] The present invention provides a novel SOS1 inhibitor. Test results show that the polycyclic pyridazinone derivative exhibits excellent SOS1 inhibitory activity, as well as excellent safety and selectivity. It can be used to prepare drugs for treating cancer, especially pancreatic cancer, colorectal cancer, lung cancer, hepatocellular carcinoma, kidney cancer, gastric cancer, and bile duct cancer. BRIEF DESCRIPTION OF THE DRAWINGS

[0198] Figure 1 This is a graph showing the effects of the compounds of the present invention on the KRAS / ERK1 / 2 signal transduction pathway in K-562 cells.

[0199] Figure 2 This is a graph showing the effects of the compounds of the present invention on the KRAS / ERK1 / 2 signal transduction pathway in K-562 cells. DETAILED DESCRIPTION

[0200] The method of the present invention is described below by means of specific examples to make the technical solution of the present invention easier to understand and grasp, but the present invention is not limited thereto. 1 H NMR spectra were obtained using a Bruker instrument (400 MHz), and chemical shifts are expressed in ppm using tetramethylsilane as an internal standard (0.00 ppm). 1 H NMR notation: s = singlet, d = doublet, t = triplet, q = quartet, m = multiplet, br = broad, dd = doublet of a doublet, dt = doublet of a triplet. Coupling constants, when given, are in Hz.

[0201] Mass spectra were obtained using LC / MS instrumentation, using ESI as the ionization method.

[0202] High performance liquid chromatograph model: Agilent 1260, Thermo Fisher U3000; chromatographic column model: Waters xbrige C18 (4.6*150 mm, 3.5 μm); mobile phase: A: ACN, B: Water (0.1% H3PO4); flow rate: 1.0 mL / min; gradient: 5% A for 1 min, increase to 20% A within 4 min, increase to 80% A within 8 min, 80% A for 2 min, back to 5% A within 0.1 min; wavelength: 220 nm; column oven: 35°C.

[0203] The thin layer chromatography silica gel plate uses Yantai Huanghai HSGF254 or Qingdao GF254 silica gel plate. The specification of the silica gel plate used in thin layer chromatography (TLC) is 0.2mm-0.3mm, and the specification used for thin layer chromatography separation and purification products is 0.4mm-0.5mm.

[0204] Column chromatography generally uses Yantai Huanghai silica gel 200-300 mesh silica gel as the carrier.

[0205] In the following examples, unless otherwise indicated, all temperatures are in degrees Celsius. Unless otherwise indicated, various starting materials and reagents are commercially available or synthesized according to known methods. Commercially available raw materials and reagents are used directly without further purification. Unless otherwise indicated, commercial manufacturers include but are not limited to Sinopharm Group, J&K Technology Co., Ltd., Tokyo Pharmaceuticals (Shanghai) Co., Ltd., Shanghai Bid Pharmaceutical Technology Co., Ltd. and Shanghai Myrel Chemical Technology Co., Ltd.

[0206] CD3OD: deuterated methanol

[0207] CDCl3: deuterated chloroform

[0208] DMSO-d6: deuterated dimethyl sulfoxide

[0209] Pd2(dba)3: tris(dibenzylideneacetone)dipalladium

[0210] Pd(dppf)Cl2:[1,1'-bis(diphenylphosphino)ferrocene]palladium dichloride

[0211] XantPhos: 4,5-bis(diphenylphosphino)-9,9-dimethylxanthene

[0212] XPhos: 2-dicyclohexylphosphino-2,4,6-triisopropylbiphenyl

[0213] HATU: 2-(7-benzotriazole oxide)-N,N,N',N'-tetramethyluronium hexafluorophosphate

[0214] TLC: Thin layer chromatography

[0215] HPLC: High Performance Liquid Chromatography

[0216] purity

[0217] &:and

[0218] Hydrogen atmosphere means that the reaction bottle is connected to a hydrogen balloon with a capacity of about 1L.

[0219] Unless otherwise specified in the examples, the solution in the reaction refers to an aqueous solution.

[0220] Unless otherwise specified in the examples, the reaction temperature is room temperature, 20°C-30°C.

[0221] The reaction progress in the examples was monitored by thin layer chromatography (TLC). The developing solvent used in the reaction and the eluent system for column chromatography or thin layer chromatography used to purify the compound included: A: petroleum ether and ethyl acetate system; B: dichloromethane and methanol system; C: n-hexane:ethyl acetate; wherein the volume ratio of the solvents varies according to the polarity of the compound and can be adjusted by adding a small amount of acidic or alkaline reagents, such as acetic acid or triethylamine.

[0222] Preparation of intermediates

[0223] Intermediate 1

[0224] (R)-1-(3-nitro-5-(trifluoromethyl)phenyl)ethylamine IN-1

[0225]

[0226] Step 1-(3-nitro-5-(trifluoromethyl)phenyl)ethan-1-one IN-1b

[0227] 3-Bromo-5-nitrobenzotrifluoride IN-1a (2.0 g, 7.41 mmol), tributyl(1-ethoxyethylene)tin (3.5 g, 9.69 mmol), and bistriphenylphosphine palladium dichloride (520 mg, 0.74 mmol) were added sequentially to toluene (25 mL). Under nitrogen, the mixture was heated to 100°C overnight. TLC indicated the reaction was complete. The reaction solution was cooled to room temperature, and hydrochloric acid (15 mL, 3N) was added. The mixture was stirred for 30 minutes, filtered through a pad of Celite, and the filtrate was separated. The aqueous phase was extracted with ethyl acetate. The combined organic phases were dried over anhydrous sodium sulfate, concentrated, and the crude product was purified by silica gel column chromatography to afford the title compound IN-1b (1.25 g, 72% yield) as a yellow oil.

[0228] 1 H NMR (400MHz, CDCl3) δ8.94(s,1H),8.68(s,1H),8.53(s,1H),2.75(s,3H).

[0229] Step 2 (R,Z)-2-methyl-N-(1-(3-nitro-5-(trifluoromethyl)phenyl)ethylidene)propane-2-sulfenamide IN-1c

[0230] A mixture of compound IN-1b (1.25 g, 5.36 mmol), (R)-(+)-tert-butylsulfenamide (974 mg, 8.04 mmol), and tetraethyl titanate (10 mL, 47.70 mmol) was heated to 80°C for 3 hours. TLC indicated a small amount of starting material remaining. The reaction mixture was cooled to room temperature, poured into ice water (60 mL), and extracted with ethyl acetate. The combined organic phases were washed with saturated brine, dried over anhydrous sodium sulfate, and concentrated. The crude product was purified by silica gel column chromatography to afford the title compound IN-1c (1.01 g, 56% yield) as a yellow oil.

[0231] Step 3: (R)-2-methyl-N-((R)-1-(3-nitro-5-(trifluoromethyl)phenyl)ethyl)propane-2-sulfenamide IN-1d

[0232] Compound IN-1c (260 mg, 0.77 mmol) was dissolved in tetrahydrofuran (2.5 mL) and water (0.05 mL). The temperature was cooled to -60°C, and sodium borohydride (74 mg, 1.95 mmol) was added portionwise. After addition, stirring was continued at -60°C for 1 hour. TLC indicated the reaction was complete. The reaction solution was quenched by dropwise addition of water and extracted with ethyl acetate. The combined organic phases were washed with saturated brine, dried over anhydrous sodium sulfate, and concentrated. The crude product was purified by silica gel column chromatography to afford the title compound IN-1d (150 mg, 58% yield) as a white solid.

[0233] 1H NMR (400MHz, CDCl3) δ8.43-8.42(m,2H),7.95(s,1H),4.75-4.69(m,1H),3.55(d,J=4.4Hz,1H),1.61(d,J=6.8Hz,3H),1.25(s,9H).

[0234] Step 4 (R)-1-(3-nitro-5-(trifluoromethyl)phenyl)ethylamine IN-1

[0235] Compound IN-1d (164 mg, 0.48 mmol) was dissolved in tetrahydrofuran (3 mL) and concentrated hydrochloric acid (0.5 mL) was added dropwise. The mixture was allowed to react at room temperature for 1 hour. TLC indicated the reaction was complete. The reaction mixture was adjusted to pH 8 by adding saturated aqueous sodium carbonate solution. The mixture was extracted with ethyl acetate, and the organic phases were combined, washed with saturated brine, dried over anhydrous sodium sulfate, and concentrated to afford the title compound IN-1 (105 mg, crude) as a yellow oil, which was used directly in the next step.

[0236] LC-MS: m / z = 235.1 [M+H] +

[0237] 1 H NMR (400MHz, DMSO-d6) δ8.56(s,1H),8.32(s,1H),8.24(s,1H),4.24(q,J=6.8Hz,1H),2.22(br,2H),1.30(d,J=2.8Hz,3H).

[0238] Intermediate 2

[0239] (R)-6-Bromo-4-((1-(3-nitro-5-(trifluoromethyl)phenyl)ethyl)amino)phthalazin-1(2H)-one IN-2

[0240]

[0241] Step 1: 4-bromo-2-iodobenzoic acid methyl ester IN-2b

[0242] Methyl 2-amino-4-bromobenzoate IN-2a (2.0 g, 8.70 mmol) was dispersed in hydrochloric acid (20 mL, 120 mmol, 6 M) and cooled to approximately 0°C. A solution of sodium nitrite (360 mg, 5.22 mmol) in water (1 mL) was added dropwise. The reaction was continued for 1 hour. Potassium iodide (1.4 g, 8.43 mmol) was then added dropwise. The reaction was continued at room temperature for 2 hours. TLC indicated that the reaction was essentially complete. The reaction solution was poured into water and extracted with ethyl acetate. The organic phases were combined, washed with saturated brine, dried over anhydrous sodium sulfate, and concentrated. The crude product was purified by silica gel column chromatography to afford the title compound IN-2b (2.1 g, 70% yield) as a yellow oil.

[0243] Step 2: Methyl 4-bromo-2-cyanobenzoate IN-2c

[0244] Compound IN-2b (2.1 g, 6.16 mmol) was dissolved in N-methylpyrrolidone (10 mL). Cuprous cyanide (834 mg, 9.31 mmol) was added at room temperature and the reaction temperature was raised to 60°C overnight. TLC confirmed the complete reaction. The reaction mixture was cooled to room temperature, filtered through a pad of celite, and the filter cake was washed. The filtrate was poured into water and extracted with ethyl acetate. The combined organic phases were washed with saturated brine, dried over anhydrous sodium sulfate, and concentrated. The crude product was purified by silica gel column chromatography to afford the title compound IN-2c (1.2 g, 80% yield) as a white solid.

[0245] Step 3 5-Bromo-3-aminoisoindol-1-one IN-2d

[0246] Compound IN-2c (1.2 g, 5.00 mmol) was dissolved in methanol (20 mL). Under nitrogen, the mixture was cooled to 0°C and ammonia was introduced until saturated. The reaction mixture was then slowly returned to room temperature and allowed to react for 48 hours. TLC analysis indicated that the reaction was essentially complete. The reaction mixture was concentrated to afford the title compound IN-2d (1.05 g, crude) as a yellow solid, which was used directly in the next step.

[0247] LC-MS: m / z = 225.0 [M+H] +

[0248] 1 H NMR (400MHz, DMSO-d6) δ10.27(br,1H),9.69(br,1H),8.30(s,1H),7.88(dd,J=8.0,1.6Hz,1H),7.64(d,J=8.0Hz,1H).

[0249] Step 4 (R)-5-bromo-3-((1-(3-nitro-5-(trifluoromethyl)phenyl)ethyl)amino)-1H-isoindol-1-one IN-2e

[0250] Compound IN-2d (1.05 g, crude) and intermediate IN-1 (1.6 g, 6.83 mmol) were dissolved in isopropanol (50 mL) and heated to 90°C for 48 hours. TLC analysis indicated residual starting material. The reaction mixture was concentrated to afford the title compound IN-2e (2.3 g, crude) as a pale yellow solid, which was used directly in the next step.

[0251] LC-MS: m / z=442.0[M+H] +

[0252] Step 5 (R)-6-bromo-4-((1-(3-nitro-5-(trifluoromethyl)phenyl)ethyl)amino)phthalazin-1(2H)-one IN-2

[0253] Compound IN-2e (2.25 g, crude) was dissolved in ethanol (40 mL). Hydrazine hydrate (636 mg, 10.16 mmol, 80%) was added at room temperature and heated to 40°C for 2 hours. TLC confirmed complete reaction. The reaction mixture was cooled to room temperature, filtered, and the filtrate was concentrated. The crude product was purified by silica gel column chromatography to afford the title compound IN-2 (1.1 g, 48% yield over three steps) as a yellow solid.

[0254] LCMS: m / z = 457.0 [M+H] +

[0255] Intermediate 3

[0256] (R)-7-Chloro-1-((1-(3-nitro-5-(trifluoromethyl)phenyl)ethyl)amino)pyrido[3,4-d]pyridazin-4-(3H)-one IN-3

[0257]

[0258] Step 1 6-chloro-4-iodonicotinic acid IN-3b

[0259] 2,2,6,6-Tetramethylpiperidine (11.2 g, 79.29 mmol) was dissolved in tetrahydrofuran (100 mL) and cooled to approximately -60°C under nitrogen. n-Butyllithium (31.7 mL, 79.25 mmol, 2.5 M) was added dropwise. After addition, the temperature was slowly returned to 0°C and allowed to react for half an hour. The mixture was then cooled to approximately -60°C and 6-chloronicotinic acid IN-3a (5.0 g, 31.74 mmol) was added portionwise. Stirring was continued at -60°C for 1 hour. A solution of iodine (9.7 g, 38.22 mmol) in tetrahydrofuran (50 mL) was added dropwise. After addition, the temperature was slowly returned to room temperature and allowed to react overnight. TLC indicated that most of the starting material had reacted. The reaction mixture was quenched with 1N hydrochloric acid, filtered, and the filter cake was washed and dried to afford the title compound IN-3b (4.7 g, crude) as a brownish-yellow solid, which was used directly in the next step.

[0260] Step 2 6-chloro-4-iodonicotinic acid methyl ester IN-3c

[0261] Compound IN-3b (4.7 g, crude) was dissolved in N,N-dimethylformamide (30 mL), and potassium carbonate (6.9 g, 49.92 mmol) was added. After stirring at room temperature for 10 minutes, iodomethane (4.7 g, 33.11 mmol) was added and allowed to react overnight. TLC confirmed the complete reaction. The reaction mixture was added with water and extracted with ethyl acetate. The combined organic phases were washed with saturated brine, dried over anhydrous sodium sulfate, and concentrated. The crude product was purified by silica gel column chromatography to afford the title compound IN-3c (2.0 g, 21% yield over two steps) as a pale yellow solid.

[0262] Step 3 6-chloro-4-vinylnicotinate methyl ester IN-3d

[0263] Compound IN-3c (3.0 g, 10.08 mmol) and potassium vinyl trifluoroacetate (1.35 g, 10.08 mmol) were dissolved in 1,4-dioxane (36 mL) and water (12 mL). Triethylamine (3.06 g, 30.24 mmol) and Pd(dppf)Cl2 dichloromethane complex (200 mg, 0.24 mmol) were added at room temperature. The atmosphere was purged with nitrogen three times and heated to 80°C for 5 hours. LCMS monitored the reaction for completion. The reaction mixture was cooled to room temperature, water was added, and extraction with ethyl acetate was performed. The organic phases were combined, washed with saturated brine, dried over anhydrous sodium sulfate, and concentrated. The crude product was purified by silica gel column chromatography to afford the title compound IN-3d (1.9 g, 95% yield) as a pale yellow solid.

[0264] LC-MS: m / z = 198.1 [M+H] +

[0265] Step 4: 6-chloro-4-formylnicotinate methyl ester IN-3e

[0266] Compound IN-3d (1.8 g, 9.11 mmol) was dissolved in a mixture of ethanol (20 mL), tetrahydrofuran (10 mL), and water (10 mL). Sodium periodate (19.2 g, 89.76 mmol) was added at room temperature. A catalytic amount of potassium osmate in water (a small amount) was added dropwise with stirring. After addition, the reaction was continued at room temperature for 0.5 hours. TLC confirmed the reaction was complete. The reaction solution was added with water and extracted with ethyl acetate. The organic phases were combined, washed with saturated brine, dried over anhydrous sodium sulfate, and concentrated. The crude product was purified by column chromatography to yield the title compound IN-3e (1.3 g, 72% yield), a pale yellow oil (solidified upon standing).

[0267] Step 5 (E)-6-chloro-4-((hydroxyimino)methyl)nicotinate IN-3f

[0268] Compound IN-3e (1.0 g, 5.01 mmol) and hydroxylaminesulfonic acid (1.70 g, 15.04 mmol) were dissolved in water (15 mL) and allowed to react overnight at room temperature. TLC confirmed the reaction was complete. The reaction mixture was added with water and extracted with ethyl acetate. The combined organic phases were washed with saturated brine, dried over anhydrous sodium sulfate, and concentrated to afford the title compound IN-3f (1.05 g, crude) as an off-white solid, which was used directly in the next step.

[0269] LCMS: m / z = 215.1 [M+H] +

[0270] Step 6: 6-chloro-4-cyanonicotinoic acid methyl ester IN-3g

[0271] Compound IN-3f (1.05 g, crude) was dissolved in phosphorus oxychloride (10 mL) and heated to 100°C for 2 hours. The reaction was complete by TLC. The reaction mixture was cooled to room temperature and concentrated. The residue was poured into ice water and neutralized with aqueous sodium bicarbonate. The mixture was extracted with ethyl acetate. The combined organic phases were washed with saturated brine, dried over anhydrous sodium sulfate, and concentrated. The crude product was purified by silica gel column chromatography to afford the title compound IN-3g (800 mg, 81% yield over two steps) as a white solid.

[0272] LCMS: m / z = 197.1 [M+H] +

[0273] Step 7: 6-chloro-1-imino-1,2-dihydro-3H-pyrrolo[3,4-c]pyridin-3-one IN-3h

[0274] Compound IN-3g (800 mg, 4.07 mmol) was dissolved in methanol (60 mL), cooled to approximately 0°C, and ammonia gas was introduced until saturated. The reaction solution was slowly warmed to room temperature and allowed to react overnight. TLC confirmed complete reaction. The reaction solution was concentrated to afford the title compound IN-3h (766 mg, crude) as a pale yellow solid, which was used directly in the next step.

[0275] LCMS: m / z = 182.1 [M+H] +

[0276] Step 8 (R)-6-chloro-1-((1-(3-nitro-5-(trifluoromethyl)phenyl)ethyl)amino)-3H-pyrrolo[3,4-c]pyridin-3-one IN-3i

[0277] Compound IN-3h (766 mg, crude) was dissolved in isopropanol (30 mL). Intermediate IN-1 (988 mg, 4.22 mmol) was added at room temperature. The mixture was heated to 85°C overnight. TLC confirmed complete reaction. The reaction mixture was cooled to room temperature and concentrated to afford the title compound IN-3i (1.75 g, crude) as a yellow solid, which was used directly in the next step.

[0278] LCMS: m / z=399.1[M+H] +

[0279] Step 9 (R)-7-chloro-1-((1-(3-nitro-5-(trifluoromethyl)phenyl)ethyl)amino)pyrido[3,4-d]pyridazin-4-(3H)-one IN-3

[0280] Compound IN-3i (1.75 g, crude) was dissolved in ethanol (15 mL). Hydrazine hydrate (579 mg, 9.25 mmol, 80%) was added at room temperature and the mixture was heated to 40°C for 1 hour. TLC confirmed complete reaction. The reaction mixture was cooled to room temperature and concentrated. The crude product was purified by silica gel column chromatography to afford the title compound IN-3 (1.0 g, 59% yield over three steps) as a yellow solid.

[0281] LCMS: m / z=414.4[M+H] +

[0282] 1 H NMR(400MHz,DMSO-d6)δ11.86(s,1H),9.19(s,1H),8.54(s,1H),8.43(s,1H),8.34( s,1H),8.26(s,1H),7.41(d,J=2.8Hz,1H),5.18-5.08(m,1H),1.57(d,J=2.8Hz,3H).

[0283] Example 1

[0284] 4-(((R)-1-(3-amino-5-(trifluoromethyl)phenyl)ethyl)amino)-7-methoxy-2-methyl-6-(((S)-tetrahydrofuran-3-yl)oxy)phthalazin-1(2H)-one 1

[0285]

[0286] Step 1 5-Bromo-4-formyl-2-methoxyphenyl acetate 1b

[0287] Dissolve acetyl vanillin 1a (20.0 g, 103.0 mmol) and potassium bromide (39.96 g, 335.8 mmol) in water (100 mL), cool to 0°C, and add liquid bromine (18.27 g, 114.3 mmol) dropwise. After addition, warm to room temperature and stir for 15 hours. TLC shows a small amount of starting material remaining. Filter the reaction mixture, wash the filter cake with water, and dry it to obtain the title compound 1b (crude, aqueous product), which is used directly in the next step.

[0288] Step 2: 2-Bromo-4-hydroxy-5-methoxybenzaldehyde 1c

[0289] Compound 1b (crude) was dissolved in concentrated hydrochloric acid (400 mL, 6N) and heated to 90°C overnight. TLC indicated that the reaction was essentially complete. The reaction mixture was cooled to room temperature, filtered, and the filter cake was washed and dried to obtain the title compound 1c (16.8 g, crude), which was used directly in the next step.

[0290] Step 3 (R)-3-Hydroxytetrahydrofuran p-toluenesulfonate 1e

[0291] (R)-3-Hydroxytetrahydrofuran 1d (1.0 g, 11.35 mmol) was dissolved in dichloromethane (8 mL), cooled to 0°C, and pyridine (2.5 g, 31.61 mmol) was added. p-Toluenesulfonyl chloride (3.03 g, 15.89 mmol) was added portionwise. The mixture was allowed to warm to room temperature and react overnight. The reaction mixture was concentrated and diluted with water. The aqueous phase was adjusted to pH 1 with hydrochloric acid (3N). The mixture was extracted with ethyl acetate. The combined organic phases were washed with saturated brine, dried over anhydrous sodium sulfate, and concentrated. The crude product was purified by silica gel column chromatography to afford the title compound 1e (2.57 g, 93% yield over three steps) as a colorless oil.

[0292] 1 H NMR (400MHz, CDCl3) δ7.78 (d, J = 8.4Hz, 2H), 7.35 (d, J = 8.4Hz, 2H), 5.11-5.09 (m, 1H), 3.91-3.77 (m, 4H), 2.45 (s, 3H), 2.10-2.02 (m, 2H).

[0293] Step 4 (S)-2-bromo-5-methoxy-4-((tetrahydrofuran-3-yl)oxy)benzaldehyde 1f

[0294] Compound 1c (3.7 g, crude), compound 1e (4.3 g, 17.75 mmol), and cesium carbonate (7.8 g, 24.0 mmol) were dispersed in N,N-dimethylformamide (20 mL) and heated to 100°C overnight. LCMS confirmed the complete reaction (TLC showed a single spot for the starting material and product). The reaction mixture was cooled to room temperature, diluted with ethyl acetate, washed with saturated brine, dried over anhydrous sodium sulfate, and concentrated to afford the title compound 1f (4.7 g, crude), which was used directly in the next step.

[0295] Step 5: (S)-2-bromo-5-methoxy-4-((tetrahydrofuran-3-yl)oxy)benzoic acid 1g

[0296] Potassium permanganate (1.94 g, 12.29 mmol) and compound 1f (3.7 g, crude) were dispersed in water (120 mL), heated to 75°C, and stirred for 5 hours. The reaction was complete by TLC. The reaction mixture was cooled to room temperature, diluted with water, and filtered. The filter cake was washed with water and ethyl acetate. The filtrate was separated, the organic phase discarded, and the aqueous phase extracted once with ethyl acetate. The aqueous phases were combined and acidified with dilute hydrochloric acid (1N). A large amount of white solid precipitated. Filter, wash, and dry the filter cake to obtain 1 g (2.0 g, crude) of the title compound, which was used directly in the next step.

[0297] LC-MS: m / z=317.02[M+H] +

[0298] Step 6: (S)-2-Formyl-5-methoxy-4-((tetrahydrofuran-3-yl)oxy)benzoic acid 1h

[0299] Compound 1g (2.4 g, crude) was dissolved in tetrahydrofuran (50 mL) and cooled to -78°C under nitrogen. n-Butyllithium (6.8 mL, 17.0 mmol, 2.5 M solution in n-hexane) was added dropwise. After addition, the mixture was stirred at -78°C for half an hour. N,N-dimethylformamide (719 mg, 9.84 mmol) was added dropwise. The mixture was slowly warmed to room temperature. TLC confirmed the reaction was complete. The reaction mixture was adjusted to weak acidity with dilute hydrochloric acid (1N). Extraction was performed with ethyl acetate. The organic phases were combined, washed with saturated brine, dried over anhydrous sodium sulfate, and concentrated. The crude product was purified by silica gel column chromatography to afford the title compound 1h (1.4 g, 21% yield over three steps).

[0300] Step 7 (S)-7-methoxy-6-((tetrahydrofuran-3-yl)oxy)phthalazin-1(2H)-one 1i

[0301] Compound 1h (1.0 g, 3.76 mmol) and aqueous hydrazine hydrate solution (1.96 g, 31.36 mmol, 80%) were dissolved in isopropanol (10 mL) and stirred at 80°C for 3 hours. TLC indicated that the reaction was essentially complete. The reaction solution was cooled to room temperature, diluted with water, and concentrated to remove the isopropanol. A solid precipitated, which was filtered, washed, and dried to obtain the title compound 1i (800 mg, crude), which was used directly in the next step.

[0302] LC-MS: m / z = 263.1 [M+H] +

[0303] Step 8 (S)-4-bromo-7-methoxy-6-((tetrahydrofuran-3-yl)oxy)phthalazin-1(2H)-one 1j

[0304] Compound 1i (500 mg, crude), benzyltrimethylammonium tribromide (1.49 g, 3.82 mmol), and potassium carbonate (528 mg, 3.82 mmol) were dispersed in N,N-dimethylformamide (10 mL). The mixture was heated to 40°C and stirred for 5 hours. TLC indicated that the reaction was essentially complete. The reaction mixture was diluted with ethyl acetate, washed with saturated brine, washed with water, and concentrated. The crude product was purified by Prep-TLC to afford the title compound 1j (538 mg, 67% yield over two steps).

[0305] Step 9 (S)-4-bromo-7-methoxy-2-methyl-6-((tetrahydrofuran-3-yl)oxy)phthalazin-1(2H)-one 1k

[0306] Compound 1j (492 mg, 1.44 mmol) was dissolved in N,N-dimethylformamide (5 mL), and sodium hydride (699 mg, 1.73 mmol, 60%) was added. The mixture was stirred at room temperature for 30 minutes. Iodomethane (409 mg, 2.88 mmol) was then added and stirred at room temperature for 2 hours. TLC indicated that the reaction was essentially complete. Water was added to the reaction solution, and a white solid precipitated. The filter cake was washed and dried to obtain the title compound 1k (461 mg, crude product), which was used directly in the next step.

[0307] Step 10: 7-methoxy-2-methyl-4-(((R)-1-(3-nitro-5-(trifluoromethyl)phenyl)ethyl)amino)-6-(((S)-tetrahydrofuran-3-yl)oxy)phthalazin-1(2H)-one

[0308] Compound 1k (161 mg, crude), intermediate IN-1 (130 mg, 0.56 mmol), potassium tert-butoxide (101 mg, 0.90 mmol), BINAP (13 mg, 0.02 mmol), and Pd(dba) (21 mg, 0.02 mmol) were dispersed in 1,4-dioxane (5 mL) and stirred overnight at 100°C under nitrogen. TLC confirmed the complete reaction. The reaction mixture was cooled to room temperature, filtered through a pad of Celite, and the filter cake was washed with ethyl acetate. The filtrate was concentrated, and the crude product was purified by Prep-TLC to afford the title compound 1l (50 mg, 20% yield over two steps).

[0309] LC-MS: m / z = 509.2 [M+H] +

[0310] Step 11: 4-(((R)-1-(3-amino-5-(trifluoromethyl)phenyl)ethyl)amino)-7-methoxy-2-methyl-6-(((S)-tetrahydrofuran-3-yl)oxy)phthalazin-1(2H)-one 1

[0311] Compound 11 (52 mg, 0.10 mmol) and palladium / carbon (5 mg, 10%) were dispersed in methanol (5 mL) and reacted at room temperature under a hydrogen atmosphere for 3 hours. TLC analysis indicated that the reaction was essentially complete. The reaction mixture was filtered through celite, the filter cake was washed with methanol, and the filtrate was concentrated. The crude product was purified by Prep-TLC to afford the title compound 1 (22 mg, 46% yield).

[0312] LC-MS: m / z = 479.2 [M+H] +

[0313] 1 H NMR (400MHz, CD3OD) δ7.70(s,1H),7.59(s,1H),6.98(s,2H),6.78(s,1H),5.32-5.29(m,1H),4.99(q,J=6.8Hz,1H),4.08- 3.99(m,3H),3.98-3.89(m,4H),3.56(s,3H),2.40-2.31(m,1H),2.24-2.18(m,1H),1.58(d,J=6.8Hz,3H).(98.25% purity by HPLC)

[0314] Example 2

[0315] (R)-4-((1-(3-amino-5-(trifluoromethyl)phenyl)ethyl)amino)-2-methyl-6-(4-methylpiperazin-1-yl)phthalazin-1(2H)-one 2

[0316]

[0317] Step 1 (R)-6-bromo-2-methyl-4-((1-(3-nitro-5-(trifluoromethyl)phenyl)ethyl)amino)phthalazin-1(2H)-one 2a

[0318] Intermediate IN-2 (300 mg, 0.66 mmol) was dissolved in N,N-dimethylformamide (5 mL), potassium carbonate (182 mg, 1.32 mmol) was added, and the mixture was stirred at room temperature for 5 minutes. Methyl iodide (280 mg, 1.97 mmol) was added and allowed to react overnight at room temperature. TLC showed that the reaction was complete. The reaction solution was added with water and extracted with ethyl acetate. The organic phases were combined, washed with saturated brine, dried over anhydrous sodium sulfate, and concentrated to obtain the title compound 2a (280 mg, crude product) as a yellow solid, which was used directly in the next step.

[0319] Step 2 (R)-2-Methyl-6-(4-methylpiperazin-1-yl)-4-((1-(3-nitro-5-(trifluoromethyl)phenyl)ethyl)amino)phthalazin-1(2H)-one 2c

[0320] Compound 2a (280 mg, crude) and N-methylpiperazine 2b (298 mg, 2.97 mmol) were dissolved in 1,4-dioxane (30 mL). Cesium carbonate (384 mg, 1.18 mmol), BINAP (60 mg, 0.10 mmol), and Pd2(dba)3 (60 mg, 0.06 mmol) were added at room temperature. The atmosphere was purged with nitrogen three times and heated to 100°C overnight. TLC confirmed complete reaction. The reaction mixture was cooled to room temperature and concentrated. The crude product was purified by silica gel column chromatography to afford the title compound 2c (240 mg, 74% yield over two steps) as a brownish-yellow solid.

[0321] LCMS: m / z=491.2[M+H] +

[0322] Step 3 (R)-4-((1-(3-amino-5-(trifluoromethyl)phenyl)ethyl)amino)-2-methyl-6-(4-methylpiperazin-1-yl)phthalazin-1(2H)-one 2

[0323] Compound 2c (240 mg, 0.49 mmol) was dissolved in ethyl acetate (10 mL) and methanol (3 mL). Palladium / carbon (50 mg, 10%) was added, followed by the dropwise addition of concentrated hydrochloric acid (3 drops). The mixture was allowed to react at room temperature under a hydrogen atmosphere for 2 hours. TLC confirmed the complete reaction. The reaction solution was neutralized with methanolic ammonia solution, filtered through a pad of Celite, and the filtrate was concentrated. The crude product was purified by Prep-TLC to afford the title compound 2 (126 mg, 56% yield) as a pale yellow solid.

[0324] LCMS: m / z=461.3[M+H] +

[0325] 1 H NMR (400MHz, DMSO-d6) δ8.01(d,J=9.2Hz,1H),7.48(d,J=2.0Hz,1H),7.40(dd,J=9.2,2.0Hz,1H),6.91(d,J=7.2Hz,1H),6.85(s,2H),6 .67(s,1H),5.49(s,2H),4.96-4.89(m,1H),3.46(s,4H),3.39(s,3H),2.58(s,4H),2.31(s,3H),1.51(d,J=7.2Hz,3H).(99.72% purity by HPLC)

[0326] Example 3

[0327] (R)-4-((1-(3-amino-5-(trifluoromethyl)phenyl)ethyl)amino)-6-(4-methylpiperazin-1-yl)phthalazin-1(2H)-one 3

[0328]

[0329] Step 1 (R)-6-(4-methylpiperazin-1-yl)-4-((1-(3-nitro-5-(trifluoromethyl)phenyl)ethyl)amino)phthalazin-1(2H)-one 3a

[0330] Intermediate IN-2 (200 mg, 0.44 mmol) and N-methylpiperidine 2b (219 mg, 2.19 mmol) were dissolved in 1,4-dioxane (10 mL). Cesium carbonate (430 mg, 1.32 mmol), BINAP (40 mg, 0.06 mmol), and Pd2(dba)3 (40 mg, 0.04 mmol) were added at room temperature. The atmosphere was purged with nitrogen three times and heated to 100°C overnight. TLC confirmed complete reaction. The reaction mixture was cooled to room temperature and concentrated. The crude product was purified by silica gel column chromatography to afford the title compound 3a (48 mg, 23% yield) as a brownish-yellow solid.

[0331] Step 2 (R)-4-((1-(3-amino-5-(trifluoromethyl)phenyl)ethyl)amino)-6-(4-methylpiperazin-1-yl)phthalazin-1(2H)-one 3

[0332] Compound 3a (48 mg, 0.10 mmol) was dissolved in ethyl acetate (10 mL) and methanol (3 mL). Palladium / carbon (50 mg, 10%) was added, followed by the dropwise addition of concentrated hydrochloric acid (2 drops). The mixture was allowed to react at room temperature under a hydrogen atmosphere for 2 hours. TLC confirmed the complete reaction. The reaction mixture was neutralized with methanolic ammonia solution, filtered through a pad of Celite, and the filtrate was concentrated. The crude product was purified by Prep-TLC to afford the title compound 3 (20 mg, 44% yield) as a pale yellow solid.

[0333] LCMS: m / z=447.3[M+H] +

[0334] 1 H NMR (400MHz, DMSO-d6) δ11.11(s,1H),7.99(d,J=9.2Hz,1H),7.50(s,1H),7.40(d,J=9.2Hz,1H),6.83-6.72(m,3H),6.6 6(s,1H),5.47(s,2H),4.95-4.83(m,1H),3.47(s,4H),2.58(s,3H),2.31(s,3H),1.49(d,J=6.8Hz,3H).(97.85% purity by HPLC)

[0335] Example 4

[0336] (R)-1-((1-(3-amino-5-(trifluoromethyl)phenyl)ethyl)amino)-3-methyl-7-(4-methylpiperazin-1-yl)pyrido[3,4-d]pyridazin-4(3H)-one 4

[0337]

[0338] Step 1: (R)-7-chloro-3-methyl-1-((1-(3-nitro-5-(trifluoromethyl)phenyl)ethyl)amino)pyrido[3,4-d]pyridazin-4-(3H)-one 4a

[0339] Intermediate IN-3 (460 mg, 1.11 mmol) was dissolved in N,N-dimethylformamide (5 mL), and potassium carbonate (268 mg, 1.94 mmol) was added. After stirring at room temperature for 5 minutes, iodomethane (275 mg, 1.94 mmol) was added and the mixture was stirred at room temperature for 2 hours. TLC analysis showed that the reaction was almost complete. The reaction solution was added with water and extracted with ethyl acetate. The organic phases were combined, washed with saturated brine, dried over anhydrous sodium sulfate, and concentrated to afford the title compound 4a (430 mg, 90% yield) as a yellow solid.

[0340] LCMS: m / z = 428.2 [M+H] +

[0341] Step 2 (R)-3-Methyl-7-(4-methylpiperazin-1-yl)-1-((1-(3-nitro-5-(trifluoromethyl)phenyl)ethyl)amino)pyrido[3,4-d]pyridazin-4-(3H)-one 4b

[0342] Compound 4a (150 mg, 0.35 mmol) was dissolved in dioxane (10 mL). Cesium carbonate (352 mg, 1.08 mmol), N-methylpiperazine (180 mg, 1.80 mmol), Pd2(dba)3 (50 mg, 0.05 mmol), and BINAP (50 mg, 0.08 mmol) were added at room temperature. The atmosphere was purged with nitrogen three times and heated to 100°C for 5 hours. TLC confirmed complete reaction. The reaction mixture was cooled to room temperature and concentrated. The crude product was purified by silica gel column chromatography to afford the title compound 4b (105 mg, 61% yield) as a yellow solid.

[0343] LCMS: m / z = 492.2 [M+H] +

[0344] Step 3 (R)-1-((1-(3-amino-5-(trifluoromethyl)phenyl)ethyl)amino)-3-methyl-7-(4-methylpiperazin-1-yl)pyrido[3,4-d]pyridazin-4(3H)-one 4

[0345] Compound 4b (105 mg, 0.21 mmol) was dissolved in ethyl acetate (10 mL) and methanol (2 mL). Concentrated hydrochloric acid (3 drops) and 10% palladium / carbon (catalytic amount) were added. The mixture was allowed to react at room temperature under a hydrogen atmosphere for 2 hours. TLC confirmed the complete reaction. The reaction solution was neutralized with methanolic ammonia solution, filtered through a pad of celite, and the filter cake was washed. The filtrate was concentrated, and the crude product was purified by Prep-TLC to afford the title compound 4 (70 mg, 70% yield) as a yellow solid.

[0346] LCMS: m / z=462.3[M+H] +

[0347] 1 H NMR(400MHz,CD3OD)δ9.04(s,1H),7.29(s,1H),6.96(s,2H),6.78(s,1H),4.94(q,J=6.8Hz,1 H),3.97(s,4H),3.49(s,3H),2.97(s,4H),2.64(s,3H),1.58(d,J=6.8Hz,3H).(98.96% purity by HPLC)

[0348] Example 5

[0349] (R)-1-((1-(3-amino-5-(trifluoromethyl)phenyl)ethyl)amino)-7-(4-methylpiperazin-1-yl)pyrido[3,4-d]pyridazin-4-(3H)-one 5

[0350]

[0351] Step 1: (R)-7-(4-methylpiperazin-1-yl)-1-((1-(3-nitro-5-(trifluoromethyl)phenyl)ethyl)amino)pyrido[3,4-d]pyridazin-4(3H)-one 5a

[0352] Intermediate IN-3 (150 mg, 0.36 mmol) was dissolved in 1,4-dioxane (10 mL). Cesium carbonate (352 mg, 1.08 mmol), N-methylpiperazine 2b (180 mg, 1.80 mmol), Pd2(dba)3 (50 mg, 0.05 mmol), and BINAP (50 mg, 0.08 mmol) were added at room temperature. The atmosphere was purged with nitrogen three times and heated to 100°C overnight. TLC confirmed complete reaction. The reaction mixture was cooled to room temperature and concentrated. The crude product was purified by silica gel column chromatography to afford the title compound 5a (40 mg, crude) as a yellow solid (TLC purity, approximately 60%), which was used directly in the next step.

[0353] Step 2 (R)-1-((1-(3-amino-5-(trifluoromethyl)phenyl)ethyl)amino)-7-(4-methylpiperazin-1-yl)pyrido[3,4-d]pyridazin-4-(3H)-one 5

[0354] Compound 5a (40 mg, crude) was dissolved in ethyl acetate (5 mL) and methanol (1 mL). Concentrated hydrochloric acid (2 drops) and 10% palladium / carbon (catalytic amount) were added. The mixture was allowed to react at room temperature under a hydrogen atmosphere for 2 hours. TLC confirmed the complete reaction. The reaction solution was neutralized with ammonia and methanol, filtered through a pad of celite, and the filter cake was washed. The filtrate was concentrated, and the crude product was purified by Prep-TLC to afford the title compound 5 (12 mg, 7% yield over two steps) as a yellow solid.

[0355] LCMS: m / z=448.3[M+H] +

[0356] 1H NMR (400MHz, CD3OD) δ9.04(s,1H),7.27(s,1H),6.92(s,2H),6.77(s,1H),4.94(q,J=6. 8Hz, 1H), 3.95 (s, 4H), 2.90 (s, 4H), 2.60 (s, 3H), 1.57 (d, J = 7.2Hz, 3H). (97.91% purity by HPLC)

[0357] Example 6

[0358] (R)-4-((1-(3-amino-5-(trifluoromethyl)phenyl)ethyl)amino)-6-morpholinophthalazin-1(2H)-one 6

[0359]

[0360] Step 1 (R)-6-morpholino-4-((1-(3-nitro-5-(trifluoromethyl)phenyl)ethyl)amino)phthalazin-1(2H)-one 6b

[0361] Under nitrogen, intermediate IN-2 (200 mg, 0.44 mmol) and morpholine 6a (381 mg, 4.37 mmol) were dissolved in 1,4-dioxane (20 mL). Pd(dba) (82 mg, 0.09 mmol), BINAP (82 mg, 0.13 mmol), and cesium carbonate (428 mg, 1.31 mmol) were added at room temperature. The mixture was heated to 100°C and allowed to react overnight. TLC confirmed the reaction was complete. The reaction solution was concentrated, and the crude product was purified by silica gel column chromatography to afford the title compound 6b (55 mg, 27% yield).

[0362] LC-MS: m / z = 464.2 [M+H] +

[0363] Step 2 (R)-4-((1-(3-amino-5-(trifluoromethyl)phenyl)ethyl)amino)-6-morpholinophthalazin-1(2H)-one 6

[0364] Compound 6b (55 mg, 0.12 mmol) was dissolved in methanol (10 mL), and palladium / carbon (20 mg, 10%) was added. The mixture was reacted at room temperature under a hydrogen atmosphere for 2 hours. TLC confirmed the reaction was complete. The reaction mixture was filtered, the filtrate was concentrated, and the crude product was purified by Prep-TLC to afford the title compound 6 (16 mg, 31% yield).

[0365] LC-MS: m / z = 434.2 [M+H] +

[0366] 1H NMR (400MHz, CD3OD) δ8.12(d,J=8.8Hz,1H),7.45(s,1H),7.39(d,J=9.2Hz,1H),6.98(s,2H),6.80(s,1 H),4.98(q,J=6.8Hz,1H),3.91-3.83(m,4H),3.50-3.40(m,4H),1.57(d,J=6.8Hz,3H).(96.33% purity by HPLC)

[0367] Example 7

[0368] (R)-4-((1-(3-amino-5-(trifluoromethyl)phenyl)ethyl)amino)-6-(4-methyl-3-oxopiperazin-1-yl)phthalazin-1(2H)-one 7

[0369]

[0370] Step 1 Benzyl 3-oxopiperazine-1-carboxylate 7b

[0371] 2-Piperazinone 7a (1.0 g, 9.99 mmol) was dissolved in ethyl acetate (20 mL). Water (20 mL) and sodium carbonate (3.2 g, 30.19 mmol) were added. The mixture was stirred at room temperature for 10 minutes, cooled to 0°C, and benzyl chloroformate (2.1 g, 12.31 mmol) was slowly added dropwise. The mixture was stirred overnight at room temperature. TLC confirmed the reaction was complete. The reaction solution was extracted with ethyl acetate, and the combined organic phases were washed with saturated brine, dried over anhydrous sodium sulfate, and concentrated. The crude product was purified by silica gel column chromatography to afford the title compound 7b (2.3 g, 98% yield).

[0372] LC-MS: m / z = 235.1 [M+H] +

[0373] Step 2 Benzyl 4-methyl-3-oxopiperazine-1-carboxylate 7c

[0374] Compound 7b (2.3 g, 9.82 mmol) was dissolved in N,N-dimethylformamide (30 mL) and cooled to 0°C. Sodium hydride (780 mg, 19.50 mmol, 60%) was added portionwise. Stirring was continued for 20 minutes after addition. Methyl iodide (2.8 g, 19.73 mmol) was added, and the mixture was warmed to room temperature and stirred for 2 hours. The reaction mixture was quenched with water and extracted with ethyl acetate. The organic phases were combined, washed with saturated brine, dried over anhydrous sodium sulfate, and concentrated. The crude product was purified by silica gel column chromatography to afford the title compound 7c (2.0 g, 82% yield).

[0375] LC-MS: m / z = 249.1 [M+H] +

[0376] Step 3 1-Methylpiperazin-2-one 7d

[0377] Compound 7c (2.0 g, 8.06 mmol) was dissolved in ethyl acetate (30 mL), and palladium / carbon (1.0 g, 10%) was added. The mixture was allowed to react overnight at room temperature under a hydrogen atmosphere. TLC confirmed the reaction was complete. The reaction mixture was filtered, and the filter cake was rinsed three times with ethyl acetate. The filtrate was concentrated to afford the title compound 7d (950 mg, crude product), which was used directly in the next step.

[0378] LC-MS: m / z = 115.2 [M+H] +

[0379] Step 4 (R)-6-(4-methyl-3-oxopiperazin-1-yl)-4-((1-(3-nitro-5-(trifluoromethyl)phenyl)ethyl)amine)phthalazin-1(2H)-one 7e

[0380] Compound 7d (200 mg, 0.44 mmol) and intermediate IN-2 (75 mg, 0.66 mmol) were dissolved in toluene (10 mL). Pd(dba) (80 mg, 0.09 mmol), BINAP (82 mg, 0.13 mmol), and cesium carbonate (328 mg, 1.01 mmol) were added at room temperature. The mixture was heated to 100°C for 4 hours. The reaction solution was concentrated, and the crude product was purified by silica gel column chromatography to afford the title compound 7e (38 mg, 18% yield over two steps).

[0381] LC-MS: m / z = 491.2 [M+H] +

[0382] Step 5 (R)-4-((1-(3-amino-5-(trifluoromethyl)phenyl)ethyl)amino)-6-(4-methyl-3-oxopiperazin-1-yl)phthalazin-1(2H)-one 7

[0383] Compound 7e (38 mg, 0.08 mmol) was dissolved in ethyl acetate (15 mL), and palladium / carbon (50 mg, 10%) was added. The mixture was stirred at room temperature under a hydrogen atmosphere for 2 hours. The reaction mixture was filtered, the filtrate was concentrated, and the crude product was purified by Prep-TLC to afford the title compound 7 (18 mg, 51% yield).

[0384] LC-MS: m / z = 461.2 [M+H] +

[0385] 1H NMR (400MHz, DMSO-d6) δ11.13(s,1H),8.01(d,J=8.8Hz,1H),7.56-7.35(m,2H),6.85(d,J=7.2Hz,1H),6.79(s,2H),6.66(s,1H),5.48( s,2H),5.10-4.78(m,1H),4.15-4.06(m,2H),3.82-3.69(m,2H),3.57-3.46(m,2H),2.95(s,3H),1.50(d,J=6.8Hz,3H).(98.33% purity by HPLC)

[0386] Example 8

[0387] (R)-4-((1-(3-amino-5-(trifluoromethyl)phenyl)ethyl)amino)-6-((2-(dimethylamino)ethyl)(methyl)amino)-2-methylphthalazin-1(2H)-one 8

[0388]

[0389] Step 1 (R)-6-((2-(dimethylamino)ethyl)(methyl)amino)-2-methyl-4-((1-(3-nitro-5-(trifluoromethyl)phenyl)ethyl)amino)phthalazin-1(2H)-one 8b

[0390] Compound 2a (100 mg, 0.21 mmol) and N,N,N'-trimethylethylenediamine 8a (109 mg, 1.07 mmol) were dissolved in 1,4-dioxane (10 mL). Pd2(dba)3 (20 mg, 0.04 mmol), BINAP (40 mg, 0.06 mmol), and cesium carbonate (140 mg, 0.43 mmol) were added at room temperature. The mixture was heated to 100°C for 4 hours. The reaction mixture was concentrated, and the crude product was purified by silica gel column chromatography to afford the title compound 8b (50 mg, 48% yield).

[0391] LC-MS: m / z=493.3[M+H] +

[0392] Step 2 (R)-4-((1-(3-amino-5-(trifluoromethyl)phenyl)ethyl)amino)-6-((2-(dimethylamino)ethyl)(methyl)amino)-2-methylphthalazin-1(2H)-one 8

[0393] Compound 8b (50 mg, 0.10 mmol) was dissolved in ethyl acetate (10 mL), and palladium / carbon (30 mg, 10%) was added. The mixture was stirred at room temperature overnight under a hydrogen atmosphere. The reaction mixture was filtered, the filtrate was concentrated, and the crude product was purified by Prep-TLC to afford the title compound 8 (28 mg, 60% yield).

[0394] LC-MS: m / z = 463.3 [M+H] +

[0395] 1 H NMR(400MHz, DMSO-d6)δ7.98(d,J=8.8Hz,1H),7.27-7.14(m,2H),6.93-6.84(m,3H),6.68(s,1H),5.47(s,2H),5.01-4.86 (m,1H),3.72(t,J=6.4Hz,2H),3.39(s,3H),3.10(s,3H),2.63(s,2H),2.36(s,6H),1.52(d,J=6.8Hz,3H).(98.53% purity by HPLC)

[0396] Example 9

[0397] (R)-4-((1-(3-amino-5-(trifluoromethyl)phenyl)ethyl)amino)-6-((2-(dimethylamino)ethyl)(methyl)amino)phthalazin-1(2H)-one 9

[0398]

[0399] Step 1: 6-Bromo-4-(((R)-1-(3-nitro-5-(trifluoromethyl)phenyl)ethyl)amino)-2-(tetrahydro-2H-pyran-2-yl)phthalazin-1(2H)-one 9a

[0400] Intermediate IN-2 (500 mg, 1.09 mmol) was dissolved in tetrahydrofuran (20 mL), and 3,4-dihydro-2H-pyran (276 mg, 3.28 mmol) and pyridinium p-toluenesulfonate (55 mg, 0.22 mmol) were added. The mixture was heated at reflux for 3 hours. TLC analysis showed that most of the starting material remained. Additional 3,4-dihydro-2H-pyran (276 mg, 3.28 mmol) was added, and the mixture was heated at reflux overnight. TLC analysis showed that more than half of the starting material remained unreacted. Additional 3,4-dihydro-2H-pyran (460 mg, 5.47 mmol) was added, and the mixture was heated at reflux overnight. TLC analysis showed that the reaction was complete. The reaction mixture was cooled to room temperature, concentrated, and the crude product was purified by silica gel column chromatography to obtain the title compound 9a (620 mg, 105% yield).

[0401] LC-MS: m / z=541.0[M+H] +

[0402] Step 2: 6-((2-(Dimethylamino)ethyl)(methyl)amino)-4-(((R)-1-(3-nitro-5-(trifluoromethyl)phenyl)ethyl)amino)-2-(tetrahydro-2H-pyran-2-yl)phthalazin-1(2H)-one 9b

[0403] Compound 9a (150 mg, 0.28 mmol) and N,N,N'-trimethylethylenediamine 8a (142 mg, 1.39 mmol) were dissolved in 1,4-dioxane (10 mL). Pd2(dba)3 (51 mg, 0.06 mmol), BINAP (52 mg, 0.08 mmol), and cesium carbonate (183 mg, 0.56 mmol) were added at room temperature. The mixture was heated to 100°C for 3 hours. The reaction solution was concentrated, and the crude product was purified by silica gel column chromatography to afford the title compound 9b (90 mg, 58% yield).

[0404] LC-MS: m / z = 563.3 [M+H] +

[0405] Step 3: (R)-6-((2-(dimethylamino)ethyl)(methyl)amino)-4-((1-(3-nitro-5-(trifluoromethyl)phenyl)ethyl)amino)phthalazin-1(2H)-one 9c

[0406] Compound 9b (90 mg, 0.16 mmol) was dissolved in hydrochloric acid / methanol (5 mL, 20.0 mmol, 4 M) and stirred at room temperature for 1 hour. TLC confirmed the reaction was complete. The reaction mixture was neutralized with ammonia and methanol and concentrated. The crude product was purified by Prep-TLC to afford the title compound 9c (70 mg, 82% yield).

[0407] LC-MS: m / z = 479.2 [M+H] +

[0408] Step 4 (R)-4-((1-(3-amino-5-(trifluoromethyl)phenyl)ethyl)amino)-6-((2-(dimethylamino)ethyl)(methyl)amino)phthalazin-1(2H)-one 9

[0409] Compound 9c (70 mg, 0.15 mmol) was dissolved in ethyl acetate (5 mL) and methanol (3 mL) and reacted at room temperature under a hydrogen atmosphere for 2 hours. The reaction mixture was filtered, the filtrate was concentrated, and the crude product was purified by Prep-TLC to give the title compound 9 (18 mg, yield 28%).

[0410] LC-MS: m / z = 449.3 [M+H] +

[0411] 1 H NMR (400MHz, CD3OD) δ8.13(d,J=9.2Hz,1H),7.28-7.23(m,2H),6.95(d,J=1.4Hz,2H),6.77(s,1H),5.00(q,J=6.8Hz ,1H),3.85(t,J=7.2Hz,2H),3.20(s,3H),2.99(t,J=6.8Hz,2H),2.65(s,6H),1.58(d,J=6.8Hz,3H).(98.53% purity by HPLC)

[0412] Example 10

[0413] (R)-4-((1-(3-amino-5-(trifluoromethyl)phenyl)ethyl)amino)-6-(4-(dimethylamino)piperidin-1-yl)phthalazin-1(2H)-one 10

[0414]

[0415] Step 1: 6-(4-(dimethylamino)piperidin-1-yl)-4-(((R)-1-(3-nitro-5-(trifluoromethyl)phenyl)ethyl)amino)-2-(tetrahydro-2H-pyran-2-yl)phthalazin-1(2H)-one 10b

[0416] Compound 9a (150 mg, 0.28 mmol) and 4-dimethylaminopiperidine 10a (178 mg, 1.39 mmol) were dissolved in 1,4-dioxane (10 mL). Pd(dba) (51 mg, 0.06 mmol), BINAP (52 mg, 0.08 mmol), and cesium carbonate (183 mg, 0.56 mmol) were added at room temperature. The mixture was heated to 100°C and reacted overnight. The reaction mixture was concentrated, and the crude product was purified by Prep-TLC to afford the title compound 10b (90 mg, 55% yield).

[0417] LC-MS: m / z = 589.3 [M+H] +

[0418] Step 2: (R)-6-(4-(Dimethylamino)piperidin-1-yl)-4-((1-(3-nitro-5-(trifluoromethyl)phenyl)ethyl)amino)phthalazin-1(2H)-one 10c

[0419] Compound 10b (90 mg, 0.15 mmol) was dissolved in hydrochloric acid / methanol (5 mL, 20.0 mmol, 4 M) and stirred at room temperature for 2 hours. The reaction solution was concentrated to give the title compound 10c (70 mg, crude product), which was used directly in the next step.

[0420] LC-MS: m / z = 505.3 [M+H] +

[0421] Step 3 (R)-4-((1-(3-amino-5-(trifluoromethyl)phenyl)ethyl)amino)-6-(4-(dimethylamino)piperidin-1-yl)phthalazin-1(2H)-one 10

[0422] Compound 10c (70 mg, crude) was dissolved in ethyl acetate (5 mL) and methanol (3 mL) and reacted at room temperature under a hydrogen atmosphere for 2 hours. The reaction mixture was filtered, the filtrate was concentrated, and the crude product was purified by Prep-TLC to give the title compound 10 (36 mg, 54% yield over two steps).

[0423] LC-MS: m / z=475.3[M+H] +

[0424] 1 H NMR (400MHz, CD3OD) δ8.08(d,J=8.8Hz,1H),7.45(d,J=2.4Hz,1H),7.36(dd,J= 9.2, 2.4Hz, 1H), 6.95 (s, 2H), 6.77 (s, 1H), 4.98 (q, J = 6.8Hz, 1H), 4.23 (d, J = 13. 2Hz,2H),3.35(s,1H),2.97(t,J=12.0Hz,2H),2.87-2.75(m,1H),2.53(s,6H),2 .09(d,J=12.0Hz,2H),1.71-1.61(m,2H),1.58(d,J=6.8Hz,3H).(97.80% purity by HPLC)

[0425] Example 11

[0426] N-((R)-1-(((R)-1-(3-amino-5-(trifluoromethyl)phenyl)ethyl)amino)-3-methyl-4-oxo-3,4-dihydropyrido[3,4-d]pyridazin-7-yl)pyrrolidin-3-yl)acetamide 11

[0427]

[0428] Step 1: Tert-butyl ((R)-1-(3-methyl-1-(((R)-1-(3-nitro-5-(trifluoromethyl)phenyl)ethyl)amino)-4-oxo-3,4-dihydropyrido[3,4-d]pyridazin-7-yl)pyrrolidin-3-yl)carbamate 11b

[0429] Compound 4a (130 mg, 0.30 mmol) was dissolved in 1,4-dioxane (8 mL). (R)-3-tert-Butyloxycarbonylaminopyrrolidine 11a (169 mg, 0.91 mmol), Pd2(dba)3 (40 mg, 0.04 mmol), BINAP (40 mg, 0.06 mmol), and cesium carbonate (198 mg, 0.61 mmol) were added at room temperature. The atmosphere was purged with nitrogen three times and heated to 100°C overnight. TLC indicated complete reaction of the starting materials. The reaction mixture was cooled to room temperature, water was added, and the mixture was extracted with ethyl acetate. The organic phases were combined, washed with saturated brine, dried over anhydrous sodium sulfate, and concentrated. The crude product was purified by silica gel column chromatography to afford the title compound 11b (190 mg, crude) as a yellow solid, which was used directly in the next step.

[0430] LC-MS: m / z = 578.3 [M+H] +

[0431] Step 2: 7-((R)-3-aminopyrrolidin-1-yl)-3-methyl-1-(((R)-1-(3-nitro-5-(trifluoromethyl)phenyl)ethyl)amino)pyrido[3,4-d]pyridazin-4-(3H)-one 11c

[0432] Compound 11b (190 mg, crude) was dissolved in ethanolic hydrochloric acid (3 mL, 12 mmol, 4 M) and allowed to react at room temperature for 1 hour. TLC indicated complete reaction. Aqueous sodium bicarbonate solution was added dropwise to make the reaction alkaline. The mixture was extracted with ethyl acetate. The combined organic phases were washed with saturated brine, dried over anhydrous sodium sulfate, and concentrated. The crude product was purified by Prep-TLC to afford the title compound 11c (90 mg, 63% yield over two steps) as a yellow oil.

[0433] LC-MS: m / z = 478.2 [M+H] +

[0434] Step 3: N-((R)-1-(3-methyl-1-(((R)-1-(3-nitro-5-(trifluoromethyl)phenyl)ethyl)amino)-4-oxo-3,4-dihydropyrido[3,4-d]pyridazin-7-yl)pyrrolidin-3-yl)acetamide 11d

[0435] Compound 11c (50 mg, 0.10 mmol) was dissolved in dichloromethane (3 mL), and triethylamine (44 mg, 0.43 mmol) and acetic anhydride (23 mg, 0.22 mmol) were added. The mixture was allowed to react at room temperature for 1 hour. TLC indicated complete reaction. The reaction mixture was added with water and extracted with dichloromethane. The organic phases were combined, washed with saturated brine, dried over anhydrous sodium sulfate, and concentrated. The crude product was purified by Prep-TLC to afford the title compound 11d (57 mg, crude) as a yellow solid, which was used directly in the next step.

[0436] LC-MS: m / z = 520.2 [M+H] +

[0437] Step 4: N-((R)-1-(((R)-1-(3-amino-5-(trifluoromethyl)phenyl)ethyl)amino)-3-methyl-4-oxo-3,4-dihydropyrido[3,4-d]pyridazin-7-yl)pyrrolidin-3-yl)acetamide 11

[0438] Compound 11d (57 mg, crude) was dissolved in ethanol (10 mL) and water (3 mL). Reduced iron powder (30 mg, 0.54 mmol) and ammonium chloride (27 mg, 0.50 mmol) were added at room temperature. The mixture was heated to 90°C for 2 hours. TLC indicated complete reaction. The reaction mixture was cooled to room temperature and filtered. The filtrate was added with water and extracted with ethyl acetate. The organic phases were combined, washed with saturated brine, dried over anhydrous sodium sulfate, and concentrated. The crude product was purified by Prep-TLC to afford the title compound 11 (25 mg, 47% yield over two steps) as a pale yellow solid.

[0439] LC-MS: m / z = 490.3 [M+H] +

[0440] 1 H NMR (400MHz, CD3OD) δ8.97(s,1H),6.96(s,2H),6.88(s,1H),6.78(s,1H),4.94(q,J=6.8Hz,1H),4.55-4.49(m,1H),3.85-3.80(m,1 H),3.76-3.62(m,2H),3.53-3.43(m,4H),2.37-2.28(m,1H),2.12-2.03(m,1H),1.96(s,3H),1.58(d,J=6.8Hz,3H).(97.19% purity by HPLC)

[0441] Example 12

[0442] (R)-1-((1-(3-amino-5-(trifluoromethyl)phenyl)ethyl)amino)-7-(1-methylpiperidin-4-yl)pyrido[3,4-d]pyridazin-4(3H)-one 12

[0443]

[0444] Step 1 (R)-7-(1-methyl-1,2,3,6-tetrahydropyridin-4-yl)-1-((1-(3-nitro-5-(trifluoromethyl)phenyl)ethyl)amino)pyrido[3,4-d]pyridazin-4-(3H)-one 12b

[0445] Intermediate IN-3 (130 mg, 0.31 mmol) and 1-methyl-1,2,3,6-tetrahydropyridine-4-boronic acid pinacol ester 12a (105 mg, 0.47 mmol) were dissolved in 1,4-dioxane (4 mL) and water (1 mL). Sodium carbonate (67 mg, 0.63 mmol) and Pd(dppf)Cl2 (26 mg, 0.03 mmol) were added sequentially at room temperature. The atmosphere was purged with nitrogen several times, and the mixture was heated to 100°C and stirred for 3 hours. TLC indicated complete reaction of the starting materials. The reaction mixture was cooled to room temperature, diluted with water, and extracted with ethyl acetate. The organic phases were combined, washed with saturated brine, dried over anhydrous sodium sulfate, and concentrated. The crude product was purified by silica gel column chromatography to afford the title compound 12b (130 mg, 87% yield) as a yellow solid.

[0446] LC-MS: m / z = 475.2 [M+H] +

[0447] Step 2 (R)-1-((1-(3-amino-5-(trifluoromethyl)phenyl)ethyl)amino)-7-(1-methylpiperidin-4-yl)pyrido[3,4-d]pyridazin-4(3H)-one 12

[0448] Compound 12b (120 mg, 0.25 mmol) was dissolved in ethyl acetate (4 mL) and methanol (1 mL). Concentrated hydrochloric acid (2 drops) and palladium / carbon (catalytic amount, 10%) were added. The mixture was stirred at room temperature under a hydrogen atmosphere for 5 hours. LCMS showed that the product was primarily present. The reaction mixture was adjusted to an alkaline pH by adding methanolic ammonia solution, filtered through a pad of Celite, and the filter cake was washed with methanol. The filtrate was concentrated, and the crude product was purified by Prep-TLC to afford the title compound 12 (31 mg, 27% yield) as a white solid.

[0449] LC-MS: m / z = 447.2 [M+H] +

[0450] 1 H NMR (400MHz, DMSO-d6) δ11.69(s,1H),9.32(s,1H),8.18(s,1H),7.15(d,J=6.8Hz,1H),6.79(d,J=8.0Hz,2H),6.68(s,1H),5.50( s,2H),4.90-4.83(m,1H),3.19-3.15(m,2H),3.02-2.93(m,1H),2.47(s,3H),2.03(s,4H),1.50(d,J=6.8Hz,3H).(95.98% purity by HPLC)

[0451] Example 13

[0452] (R)-1-((1-(3-amino-5-(trifluoromethyl)phenyl)ethyl)amino)-7-(4-(dimethylamino)piperidin-1-yl)pyrido[3,4-d]pyridazin-4(3H)-one 13

[0453]

[0454] Step 1: (R)-7-(4-(dimethylamino)piperidin-1-yl)-1-((1-(3-nitro-5-(trifluoromethyl)phenyl)ethyl)amino)pyrido[3,4-d]pyridazin-4(3H)-one 13a

[0455] Intermediate IN-3 (100 mg, 0.24 mmol) was dissolved in 4-dimethylaminopiperidine 10a (310 mg, 2.42 mmol). DMSO (3 drops) was added dropwise at room temperature. The mixture was heated to 80°C and stirred for 3 hours. TLC indicated complete reaction. The reaction mixture was cooled to room temperature, diluted with water, and extracted with ethyl acetate. The combined organic phases were washed with saturated brine, dried over anhydrous sodium sulfate, and concentrated to afford the title compound 13a (140 mg, crude product), which was used directly in the next step.

[0456] LC-MS: m / z = 506.3 [M+H] +

[0457] Step 2 (R)-1-((1-(3-amino-5-(trifluoromethyl)phenyl)ethyl)amino)-7-(4-(dimethylamino)piperidin-1-yl)pyrido[3,4-d]pyridazin-4(3H)-one 13

[0458] Compound 13a (130 mg, crude) was dissolved in ethanol (4 mL) and water (2 mL). Iron powder (72 mg, 1.29 mmol) and ammonium chloride (70 mg, 1.31 mmol) were added at room temperature. The mixture was heated to 80°C and stirred for 2 hours. LCMS indicated the reaction was complete. The reaction mixture was filtered while hot, and the filter cake was washed several times with ethanol. The filtrate was concentrated, and the crude product was purified by Prep-TLC to afford the title compound 13 (7.7 mg, 6% yield over two steps) as a white solid.

[0459] LC-MS: m / z=476.3[M+H] +

[0460] 1H NMR(400MHz,DMSO-d6)δ11.18(s,1H),8.93(s,1H),7.41(s,1H),6.94(d,J =6.8Hz,1H),6.78(s,2H),6.67(s,1H),5.51(s,2H),4.90-4.83(m,1H),4. 80-4.76(m,2H),3.51-3.46(m,1H),3.04-2.98(m,2H),2.74(s,6H),2.17( d,J=11.2Hz,2H),1.69-1.61(m,2H),1.50(d,J=6.8Hz,3H).(97.49% purity by HPLC)

[0461] Example 14

[0462] (R)-1-((1-(2-fluoro-3-(trifluoromethyl)phenyl)ethyl)amino)-7-(4-methylpiperazin-1-yl)pyrido[3,4-d]pyridazin-4(3H)-one 14

[0463]

[0464] Step 1-(2-Fluoro-3-(trifluoromethyl)phenylethanol 14b

[0465] 2-Fluoro-3-trifluoromethylbenzaldehyde 14a (10.0 g, 52.05 mmol) was dissolved in tetrahydrofuran (200 mL) and cooled to 0°C. Methylmagnesium bromide (9.3 g, 78.00 mmol) was slowly added dropwise. After completion of the dropwise addition, the mixture was slowly warmed to room temperature and stirred for 1 hour. TLC indicated complete reaction. The reaction mixture was poured into saturated aqueous ammonium chloride for quenching and extracted with ethyl acetate. The organic phases were combined, washed with saturated brine, dried over anhydrous sodium sulfate, and concentrated to afford the title compound 14b (11.1 g, crude product), which was used directly in the next step.

[0466] Step 2: 2-Fluoro-3-(trifluoromethyl)acetophenone 14c

[0467] Compound 14b (11.1 g, crude) was dissolved in dichloromethane (200 mL), cooled to 0°C, and the atmosphere was replaced with nitrogen several times. Dess-Martin reagent (26.9 g, 63.42 mmol) was added, and the mixture was stirred at room temperature for 1 hour. TLC indicated complete reaction. The reaction mixture was quenched by pouring into saturated aqueous sodium carbonate solution, extracted with dichloromethane, and the combined organic phases were washed with saturated brine, dried over anhydrous sodium sulfate, and concentrated. The crude product was purified by silica gel column chromatography to afford the title compound 14c (7.3 g, 68% yield over two steps) as a yellow liquid.

[0468] 1H NMR (400MHz, CDCl3) δ8.10-8.06(m,1H),7.83-7.79(m,1H),7.35(t,J=8.0Hz,1H),2.69(d,J=5.2Hz,3H).

[0469] Step 3 (R,Z)-N-(1-(2-fluoro-3-(trifluoromethyl)phenyl)ethylidene)-2-methylpropane-2-sulfenamide 14d

[0470] Compound 14c (7.3 g, 35.41 mmol) was dissolved in tetraethyl titanate (80 mL). (R)-(+)-tert-butylsulfenamide (12.9 g, 106.4 mmol) was added at room temperature. After addition, the temperature was slowly raised to 100°C and stirred for 1 hour. TLC indicated complete reaction. The reaction mixture was poured into vigorously stirred ice water and stirred for 10 minutes. The mixture was filtered, and the filter cake was repeatedly washed with ethyl acetate. The filtrate was extracted with ethyl acetate. The organic phases were combined, washed with saturated brine, dried over anhydrous sodium sulfate, and concentrated to afford the title compound 14d (11.0 g, crude product), which was used directly in the next step.

[0471] LC-MS: m / z=310.1[M+H] +

[0472] Step 4: (R)-N-((R)-1-(2-fluoro-3-(trifluoromethyl)phenyl)ethyl)-2-methylpropane-2-sulfenamide 14e

[0473] Compound 14d (11.0 g, crude) was dissolved in tetrahydrofuran (120 mL) and water (2 mL), cooled to approximately -60°C, and sodium borohydride (4.0 g, 105.7 mmol) was then added portionwise. The reaction was continued for 2 hours after addition, and TLC indicated that the starting material was consumed. The reaction mixture was quenched by pouring into ice water and extracted with ethyl acetate. The organic phase was washed several times with water and saturated brine, dried over sodium sulfate, and concentrated to obtain the crude product. The crude product was purified by silica gel column chromatography to afford the title compound 14e (3.4 g, 28% yield over two steps) as a yellow oil.

[0474] LC-MS: m / z=312.1[M+H] +

[0475] Step 5 (R)-1-(2-fluoro-3-(trifluoromethyl)phenylethylamine 14f

[0476] Compound 14e (3.4 g, 10.92 mmol) was dissolved in tetrahydrofuran (30 mL), and concentrated hydrochloric acid (800 mg, 21.8 mmol) was added. The mixture was heated to 80°C and stirred for 1 hour. TLC indicated complete reaction. The reaction mixture was cooled to room temperature and poured into saturated aqueous sodium bicarbonate to adjust the pH to alkaline. The mixture was extracted with ethyl acetate, and the organic phase was washed with saturated brine, dried over sodium sulfate, and concentrated to afford the title compound 14f (2.0 g, 87% yield) as a yellow liquid.

[0477] LC-MS: m / z = 208.1 [M+H] +

[0478] Step 6 (R)-6-chloro-1-((1-(2-fluoro-3-(trifluoromethyl)phenyl)ethyl)amino)-3H-pyrrolo[3,4-c]pyridin-3-one 14g

[0479] Compound IN-3h (400 mg, 2.20 mmol) and compound 14f (411 mg, 1.98 mmol) were dissolved in isopropanol (15 mL) and heated to 90°C overnight. TLC indicated near-complete reaction. The reaction mixture was cooled to room temperature and concentrated to afford the title compound 14g (800 mg, crude) as a yellow solid, which was used directly in the next step.

[0480] Step 7: (R)-7-chloro-1-((1-(2-fluoro-3-(trifluoromethyl)phenyl)ethyl)amino)pyrido[3,4-d]pyridazin-4-(3H)-one for 14 hours

[0481] Compound 14g (800 mg, crude) was dissolved in ethanol (10 mL). Hydrazine hydrate (300 mg, 4.79 mmol, 80%) was added at room temperature and heated to 40°C for 1 hour. TLC indicated that the reaction was essentially complete. The reaction solution was cooled to room temperature and concentrated. The crude product was purified by silica gel column chromatography to afford the title compound 14h (340 mg, 44% yield over two steps) as a yellow solid.

[0482] Step 8 (R)-1-((1-(2-fluoro-3-(trifluoromethyl)phenyl)ethyl)amino)-7-(4-methylpiperazin-1-yl)pyrido[3,4-d]pyridazin-4(3H)-one 14

[0483] Compound 14h (80 mg, 0.21 mmol) and N-methylpiperazine (105 mg, 1.05 mmol) were dissolved in toluene (10 mL). BINAP (30 mg, 0.048 mmol), palladium acetate (30 mg, 0.13 mmol), and sodium tert-butoxide (81 mg, 0.84 mmol) were added sequentially at room temperature. The atmosphere was purged with nitrogen three times and heated to 100°C for 5 hours. TLC confirmed complete reaction. The reaction mixture was cooled to room temperature and concentrated. The crude product was purified by Prep-TLC to afford the title compound 14 (20 mg, 21% yield) as an off-white solid.

[0484] LC-MS: m / z = 451.2 [M+H] +

[0485] 1 H NMR (400MHz, DMSO-d6) δ11.18(s,1H),8.92(s,1H),7.71(t,J=7.2Hz,1H),7.63(t,J=7.2Hz,1H),7.42-7.30(m,2H),7 .06(d,J=6.8Hz,1H),5.26-5.21(m,1H),3.80(s,4H),2.60(s,4H),2.35(s,3H),1.56(d,J=6.8Hz,3H).(99.25% purity by HPLC)

[0486] Example 15

[0487] (R)-1-((1-(3-(difluoromethyl)-2-fluorophenyl)ethyl)amino)-7-(4-methylpiperazin-1-yl)pyrido[3,4-d]pyridazin-4-(3H)-one 15

[0488]

[0489] Step 1-Bromo-3-difluoromethyl-2-fluorobenzene 15b

[0490] 2-Fluoro-3-bromobenzaldehyde 15a (10.0 g, 49.26 mmol) was dissolved in dichloromethane (200 mL) and cooled to 0°C. Diethylaminosulfur trifluoride (15.9 g, 98.64 mmol) was slowly added dropwise. The mixture was slowly warmed to room temperature and stirred for 1 hour. TLC indicated complete reaction. The reaction mixture was quenched with saturated aqueous sodium bicarbonate and extracted with dichloromethane. The organic phases were combined, washed with saturated brine, dried over anhydrous sodium sulfate, and concentrated. The crude product was purified by silica gel column chromatography to afford the title compound 15b (8.1 g, 73% yield) as a yellow liquid.

[0491] 1H NMR (400MHz, CDCl3) δ7.68(t,J=7.2Hz,1H),7.55(t,J=7.2Hz,1H),7.14(t,J=8.0Hz,1H),6.89(t,J=54.8Hz,1H).

[0492] Step 2 1-(3-(Difluoromethyl)-2-fluoroacetophenone 15c

[0493] Compound 15b (8.1 g, 36.00 mmol) was dissolved in 1,4-dioxane (80 mL). Triethylamine (9.1 g, 89.93 mmol) and tributyl(1-ethoxyethylene)tin (15.6 g, 43.20 mmol) were added sequentially at room temperature. Nitrogen was bubbled through the mixture for 15 minutes. Bistriphenylphosphine palladium dichloride (250 mg, 0.36 mmol) was added, and the atmosphere was replaced with nitrogen several times. The temperature was raised to 100°C and stirred for 1 hour. TLC indicated the reaction was complete. The reaction solution was cooled to room temperature, and dilute hydrochloric acid (14.4 mL, 72.0 mmol, 5 M) was added. The mixture was stirred at room temperature for 1 hour. TLC indicated the reaction was complete. The reaction solution was diluted with water and extracted with ethyl acetate. The organic phases were combined, washed with saturated brine, dried over anhydrous sodium sulfate, and concentrated. The crude product was purified by silica gel column chromatography to obtain the title compound 15c (5.6 g, 83% yield) as a yellow liquid.

[0494] 1 H NMR (400MHz, CDCl3) δ8.00(t,J=7.2Hz,1H),7.79(t,J=6.8Hz,1H),7.34(t,J=7.6Hz,1H),6.94(t,J=54.8Hz,1H),2.67(d,J=5.2Hz,3H).

[0495] Step 3 (R,Z)-N-(1-(3-(difluoromethyl)-2-fluorophenyl)ethylidene)-2-methylpropane-2-sulfenamide 15d

[0496] Compound 15c (5.6 g, 29.76 mmol) was dissolved in tetraethyl titanate (50 mL). (R)-(+)-tert-butylsulfenamide (10.8 g, 89.11 mmol) was added at room temperature. The temperature was slowly raised to 100°C and stirred for 1 hour. TLC indicated complete reaction. The reaction mixture was cooled to room temperature and poured into vigorously stirred ice water. Stirring was continued for 10 minutes, followed by filtration. The filter cake was repeatedly washed with ethyl acetate. The filtrate was extracted with ethyl acetate. The organic phases were combined, washed with saturated brine, dried over anhydrous sodium sulfate, and concentrated to afford the title compound 15d (9.5 g, crude) as a yellow oil, which was used directly in the next step.

[0497] LC-MS: m / z = 292.1 [M+H] +

[0498] Step 4: (R)-N-((R)-1-(3-(difluoromethyl)-2-fluorophenyl)ethyl)-2-methylpropane-2-sulfenamide 15e

[0499] Compound 15d (9.5 g, crude) was dissolved in tetrahydrofuran (100 mL) and water (2 mL), cooled to approximately -60°C, and sodium borohydride (3.7 mg, 97.80 mmol) was added portionwise. After addition, the temperature was slowly raised to room temperature and the reaction was continued for 2 hours. TLC indicated that the starting material was consumed. The reaction mixture was quenched with ice water and extracted with ethyl acetate. The organic phases were combined, washed with water and saturated brine, dried over anhydrous sodium sulfate, and concentrated. The crude product was purified by silica gel column chromatography to afford the title compound 15e (2.9 g, 33% yield over two steps) as a yellow oil.

[0500] LC-MS: m / z = 294.1 [M+H] +

[0501] Step 5 (R)-1-(3-(difluoromethyl)-2-fluorophenethylamine 15f

[0502] Compound 15e (2.9 g, 9.88 mmol) was dissolved in tetrahydrofuran (50 mL). Concentrated hydrochloric acid (720 mg, 19.8 mmol) was added at room temperature. The mixture was heated to 80°C and stirred for 1 hour. TLC indicated that the starting material was consumed. The reaction solution was cooled to room temperature and poured into saturated aqueous sodium bicarbonate to adjust the pH to alkaline. The mixture was extracted with ethyl acetate. The organic phases were combined, washed with saturated brine, dried over anhydrous sodium sulfate, and concentrated to obtain the title compound 15f (2.0 g, crude) as a brown liquid, which was used directly in the next step.

[0503] LC-MS: m / z = 190.1 [M+H] +

[0504] Step 6 (R)-6-chloro-1-((1-(3-(difluoromethyl)-2-fluorophenyl)ethyl)amino)-3H-pyrrolo[3,4-c]pyridin-3-one 15g

[0505] Compound IN-3h (400 mg, 2.20 mmol) and compound 15f (375 mg, 1.98 mmol) were dissolved in isopropanol (15 mL) and heated to 90°C overnight. TLC indicated near-complete reaction. The reaction mixture was cooled to room temperature and concentrated to afford the title compound 15 g (826 mg, crude) as a yellow solid, which was used directly in the next step.

[0506] Step 7: (R)-7-chloro-1-((1-(3-(difluoromethyl)-2-fluorophenyl)ethyl)amino)pyrido[3,4-d]pyridazin-4-(3H)-one for 15h

[0507] Compound 15g (800 mg, crude) was dissolved in ethanol (10 mL), and hydrazine hydrate (300 mg, 4.79 mmol, 80%) was added. The reaction mixture was heated to 40°C for 1 hour. TLC indicated that the reaction was essentially complete. The reaction mixture was cooled to room temperature and concentrated. The crude product was purified by silica gel column chromatography to afford the title compound 15h (360 mg, 46% yield over three steps) as a yellow solid.

[0508] Step 8 (R)-1-((1-(3-(difluoromethyl)-2-fluorophenyl)ethyl)amino)-7-(4-methylpiperazin-1-yl)pyrido[3,4-d]pyridazin-4-(3H)-one 15

[0509] Compound 15h (80 mg, 0.22 mmol) and N-methylpiperazine (220 mg, 2.20 mmol) were dissolved in dimethyl sulfoxide (0.5 mL) and heated to 100°C for 2 hours. TLC confirmed complete reaction. The reaction mixture was cooled to room temperature, and water was added to precipitate a solid. The solid was filtered and the filter cake washed to afford the crude product. Purification by Prep-TLC afforded the title compound 15 (43 mg, 46% yield) as a yellow solid.

[0510] LC-MS: m / z = 433.2 [M+H] +

[0511] 1 H NMR (400MHz, DMSO-d6) δ11.16(s,1H),8.91(s,1H),7.56(t,J=7.2Hz,1H),7.48(t,J=6.8Hz,1H),7. 35(s,1H),7.27(t,J=7.6Hz,1H),7.22(t,J=54.4Hz,1H),7.00(d,J=6.8Hz,1H),5.24-5.19(m,1H), 3.77(s,4H),2.52(s,4H),2.30(s,3H),1.54(d,J=6.8Hz,3H).(97.61% purity by HPLC)

[0512] Example 16

[0513] (R)-1-((1-(5-amino-3-(difluoromethyl)-2-fluorophenyl)ethyl)amino)-7-(4-methylpiperazin-1-yl)pyrido[3,4-d]pyridazin-4-(3H)-one 16

[0514]

[0515] Step 1-(3-(difluoromethyl)-2-fluoro-5-nitrophenyl)ethan-1-one 16a

[0516] Potassium nitrate (45.6 g, 0.45 mmol) was dissolved in concentrated sulfuric acid (100 mL) and stirred at room temperature for 30 minutes. The mixture was cooled to approximately 0°C and slowly added dropwise with compound 15c (8.5 g, 45.18 mmol). The mixture was stirred at 0°C for 5 minutes. TLC indicated the reaction was complete. The reaction solution was slowly added to ice water and extracted with ethyl acetate. The organic phases were combined, washed with saturated sodium bicarbonate solution and saturated brine, and concentrated. The crude product was purified on a silica gel column to afford the title compound 16a (9.0 g, 86% yield) as a pale yellow liquid.

[0517] 1 H NMR (400MHz, CDCl3) δ8.88-8.86(m,1H),8.66-8.64(m,1H),7.12-6.85(m,1H),2.73(d,J=4.8Hz,3H).

[0518] Step 2 (R,Z)-N-(1-(3-(difluoromethyl)-2-fluoro-5-nitrophenyl)ethylidene)-2-methylpropane-2-sulfenamide 16b

[0519] Compound 16a (9.0 g, 38.60 mmol) was dispersed in tetraethyl titanate (90 mL). (R)-(+)-tert-butylsulfenamide (7.0 g, 57.76 mmol) was added at room temperature. The temperature was slowly raised to 100°C and stirred for 1 hour. TLC indicated complete reaction. The reaction mixture was cooled to room temperature and poured into vigorously stirred ice water. Stirring was continued for 10 minutes, followed by filtration. The filter cake was repeatedly washed with ethyl acetate. The filtrate was extracted with ethyl acetate. The organic phases were combined, washed with saturated brine, dried over anhydrous sodium sulfate, and concentrated to afford the title compound 16b (9.0 g, crude) as a yellow oil, which was used directly in the next step.

[0520] LC-MS: m / z=337.1[M+H] +

[0521] Step 3 (R)-N-((R)-1-(3-(difluoromethyl)-2-fluoro-5-nitrophenyl)ethyl)-2-methylpropane-2-sulfenamide 16c

[0522] Compound 16b (9.0 g, crude) was dissolved in tetrahydrofuran (150 mL) and water (2 mL), cooled to approximately -60°C, and sodium borohydride (3.1 g, 81.95 mmol) was added portionwise. After addition, the temperature was slowly raised to room temperature and the reaction was continued for 2 hours. TLC indicated that the starting material was consumed. The reaction mixture was quenched with ice water and extracted with ethyl acetate. The organic phases were combined, washed with water and saturated brine, dried over anhydrous sodium sulfate, and concentrated. The crude product was purified on a silica gel column to afford the title compound 16c (2.8 g, 21% yield over two steps) as a brown oil.

[0523] LC-MS: m / z=339.1[M+H] +

[0524] Step 4 (R)-1-(3-(difluoromethyl)-2-fluoro-5-nitrophenyl)ethan-1-amine 16d

[0525] Compound 16c (2.8 g, 8.28 mmol) was dissolved in tetrahydrofuran (30 mL), and concentrated hydrochloric acid (1.4 mL, 16.52 mmol) was added. The mixture was stirred at room temperature for 1 hour. TLC indicated that the starting material was consumed. The reaction solution was cooled to room temperature and poured into saturated aqueous sodium bicarbonate to adjust the pH to alkaline. The mixture was extracted with ethyl acetate. The organic phases were combined, washed with saturated brine, dried over anhydrous sodium sulfate, and concentrated to obtain a brown liquid, which solidified on standing to form the title compound 16d (1.8 g, 95% yield).

[0526] LC-MS: m / z = 235.1 [M+H] +

[0527] 1 H NMR (400MHz, DMSO-d6) δ8.76-8.74(m,1H),8.41-8.39(m,1H),7.52-7.25(m,1H),4.42(q,J=6.4Hz,1H),2.48-2.24(m,2H),1.35(d,J=6.8Hz,3H).

[0528] Step 5 (R)-3-(1-aminoethyl)-5-(difluoromethyl)-4-fluoroaniline 16e

[0529] Compound 16d (300 mg, 1.28 mmol) was dissolved in ethanol and water (6 mL / 2 mL). Reduced iron powder (358 mg, 6.41 mmol) and ammonium chloride (346 mg, 6.69 mmol) were added at room temperature. The mixture was heated to 90°C for 2 hours. TLC indicated the reaction was complete. The reaction solution was cooled to room temperature, filtered through a pad of celite, and the filtrate was concentrated to afford the title compound 16e (600 mg, crude product), which was used directly in the next step.

[0530] Step 6 (R)-1-((1-(5-amino-3-(difluoromethyl)-2-fluorophenyl)ethyl)amino)-6-chloro-3H-pyrrolo[3,4-c]pyridin-3-one 16f

[0531] Compound 16e (200 mg, crude) and compound IN-3h (89 mg, 0.45 mmol) were dispersed in isopropanol (5 mL) and heated to 90°C for 12 hours. TLC indicated the reaction was complete. The reaction solution was cooled to room temperature and concentrated to afford the title compound 16f (180 mg, crude) as a yellow solid, which was used directly in the next step.

[0532] LC-MS: m / z=369.1[M+H] +

[0533] Step 7 (R)-1-((1-(5-amino-3-(difluoromethyl)-2-fluorophenyl)ethyl)amino)-7-chloro-pyrido[3,4-d]pyridazin-4-(3H)-one 16g

[0534] Compound 16f (180 mg, crude) was dissolved in methanol (10 mL), and hydrazine hydrate (100 mg, 1.60 mmol, 80%) was added dropwise at room temperature. The mixture was heated to 45°C and reacted for 1.5 hours. TLC analysis showed the reaction was complete. The reaction solution was cooled to room temperature, filtered, and the filtrate was concentrated. The crude product was purified by Prep-TLC to afford the title compound 16g (44 mg, 27% yield over three steps) as a yellow solid.

[0535] LC-MS: m / z = 382.1 [MH] -

[0536] Step 8 (R)-1-((1-(5-amino-3-(difluoromethyl)-2-fluorophenyl)ethyl)amino)-7-(4-methylpiperazin-1-yl)pyrido[3,4-d]pyridazin-4-(3H)-one 16

[0537] Compound 16g (20 mg, 0.05 mmol) was dissolved in DMSO (0.5 mL). N-methylpiperazine (52 mg, 0.52 mmol) was added at room temperature and heated to 85°C for 12 hours. TLC indicated the reaction was complete. The reaction solution was cooled to room temperature, water was added, and extraction with ethyl acetate was performed. The combined organic phases were washed with saturated brine, dried over anhydrous sodium sulfate, and concentrated. The crude product was purified by Prep-TLC to afford the title compound 16 (7 mg, 30% yield) as a yellow solid.

[0538] LC-MS: m / z = 448.3 [M+H] +

[0539] 1H NMR(400MHz,CD3OD)δ9.04(s,1H),7.33(s,1H),6.95-6.82(m,2H),6.76-6.72(m,1H),5.20(q ,J=6.8Hz,1H),3.99(s,4H),3.00(s,4H),2.66(s,3H),1.58(d,J=7.2Hz,3H).(97.85% purity by HPLC)

[0540] Example 17

[0541] 1-(((R)-1-(3-amino-5-(trifluoromethyl)phenyl)ethyl)amino)-7-(1-methylpiperidin-3-yl)pyrido[3,4-d]pyridazin-4(3H)-one 17

[0542]

[0543] Step 1: (R)-5-(1-((1-(3-nitro-5-(trifluoromethyl)phenyl)ethyl)amino)-4-oxo-3,4-dihydropyrido[3,4-d]pyridazin-7-yl)-3,6-dihydropyridine-1(2H)-carboxylic acid tert-butyl ester 17b

[0544] Intermediate IN-3 (110 mg, 0.27 mmol) and 1-tert-butyloxycarbonyl-3,6-dihydro-2H-pyridine-5-boronic acid pinacol ester 17a (123 mg, 0.40 mmol) were dissolved in 1,4-dioxane (4 mL) and water (1 mL). Sodium carbonate (56 mg, 0.53 mmol) and Pd(dppf)Cl2 (22 mg, 0.023 mmol) were added sequentially at room temperature. After addition, the atmosphere was purged with nitrogen several times, the temperature was raised to 100°C, and the mixture was stirred for 3 hours. TLC showed that the starting materials had reacted completely. The reaction mixture was cooled to room temperature, diluted with water, and extracted with ethyl acetate. The organic phases were combined, washed with saturated brine, dried over anhydrous sodium sulfate, and concentrated. The crude product was purified on a silica gel column to obtain the title compound 17b (120 mg, 81% yield) as a yellow solid.

[0545] LC-MS: m / z=561.3[M+H] +

[0546] Step 2: (R)-1-((1-(3-nitro-5-(trifluoromethyl)phenyl)ethyl)amino)-7-(1,2,5,6-tetrahydropyridin-3-yl)pyrido[3,4-d]pyridazin-4(3H)-one 17c

[0547] Compound 17b (120 mg, 0.21 mmol) was dissolved in dichloromethane (2 mL), and trifluoroacetic acid (1 mL) was added. The mixture was stirred at room temperature for 1 hour. TLC indicated that the starting material was consumed. The reaction solution was concentrated, and the pH was adjusted to alkaline with saturated sodium bicarbonate aqueous solution. The mixture was extracted with ethyl acetate, and the organic phases were combined, washed with saturated brine, dried over anhydrous sodium sulfate, and concentrated to afford the title compound 17c (110 mg, crude) as a yellow oil, which was used directly in the next step.

[0548] LC-MS: m / z = 461.2 [M+H] +

[0549] Step 3 (R)-7-(1-methyl-1,2,5,6-tetrahydropyridin-3-yl)-1-((1-(3-nitro-5-(trifluoromethyl)phenyl)ethyl)amino)pyrido[3,4-d]pyridazin-4(3H)-one 17d

[0550] Compound 17c (100 mg, crude) was dissolved in acetonitrile (2 mL), and potassium carbonate (60 mg, 0.43 mmol) and methyl p-toluenesulfonate (32 mg, 0.17 mmol) were added. The mixture was stirred at room temperature overnight. The reaction mixture was diluted with water and extracted with ethyl acetate. The combined organic phases were washed with saturated brine, dried over anhydrous sodium sulfate, and concentrated. The crude product was purified on a silica gel column to afford the title compound 17d (25 mg, 25% yield over two steps) as a yellow solid.

[0551] LC-MS: m / z = 475.2 [M+H] +

[0552] Step 4: 1-(((R)-1-(3-amino-5-(trifluoromethyl)phenyl)ethyl)amino)-7-(1-methylpiperidin-3-yl)pyrido[3,4-d]pyridazin-4(3H)-one 17

[0553] Compound 17d (25 mg, 0.053 mmol) was dissolved in a mixture of ethyl acetate (1 mL) and tetrahydrofuran (1 mL). Palladium on carbon (20 mg, 10%) was added at room temperature. The mixture was heated to 50°C and stirred overnight under a hydrogen atmosphere. The reaction mixture was filtered through a pad of Celite, and the filter cake was washed several times with methanol. The filtrate was concentrated, and the crude product was purified by Prep-TLC to afford the title compound 17 (10 mg, 42% yield) as a yellow solid.

[0554] LC-MS: m / z=447.3[M+H] +

[0555] 1H NMR (400MHz, CD3OD) δ9.41(s,1H),8.09(s,1H),6.92(s,2H),6.76(s,1H),4.96(q,J=6.8Hz,1H),3.38-3.32(m,2H),3.14-3.11(m,1 H),2.82-2.70(m,1H),2.56(s,3H),2.50-2.37(m,1H),2.14-2.12(m,1H),1.95-1.78(m,3H),1.57(d,J=7.2Hz,3H).(94.55% purity by HPLC)

[0556] Example 18

[0557] 1-(((R)-1-(3-amino-5-(trifluoromethyl)phenyl)ethyl)amino)-7-(6-methyl-3,6-diazabicyclo[3.1.1]hept-3-yl)pyrido[3,4-d]pyridazin-4(3H)-one 18

[0558]

[0559] Step 1: tert-Butyl 3-(1-(((R)-1-(3-nitro-5-(trifluoromethyl)phenyl)ethyl)amino)-4-oxo-3,4-dihydropyrido[3,4-d]pyridazin-7-yl)-3,6-diazabicyclo[3.1.1]heptane-6-carboxylate 18b

[0560] Intermediate IN-3 (300 mg, 0.73 mmol) and 6-(tert-butyloxycarbonyl)-3,6-diazabicyclo[3.1.1]heptane 18a (575 mg, 2.90 mmol) were dissolved in dimethyl sulfoxide (6 mL), heated to 100°C, and stirred for 2 hours. The reaction was complete by TLC. The reaction solution was cooled to room temperature and water was added. Solid precipitated and was filtered. The filter cake was washed and dried to afford the title compound 18b (400 mg, crude) as a yellow solid, which was used directly in the next step.

[0561] LC-MS: m / z = 576.3 [M+H] +

[0562] Step 2: 7-(3,6-diazabicyclo[3.1.1]hept-3-yl)-1-(((R)-1-(3-nitro-5-(trifluoromethyl)phenyl)ethyl)amino)pyrido[3,4-d]pyridazin-4(3H)-one 18c

[0563] Compound 18b (200 mg, crude) was dissolved in methanol (2 mL), and methanolic hydrochloric acid (4 mL, 4 M) was added. The mixture was stirred at room temperature for 3 hours. TLC confirmed the reaction was complete. The reaction solution was neutralized with methanolic ammonia, filtered, and the filtrate was concentrated to obtain the title compound 18c (170 mg, crude), which was used directly in the next step.

[0564] Step 3: 7-(6-methyl-3,6-diazabicyclo[3.1.1]hept-3-yl)-1-(((R)-1-(3-nitro-5-(trifluoromethyl)phenyl)ethyl)amino)pyrido[3,4-d]pyridazin-4(3H)-one 18d

[0565] Compound 18c (170 mg, crude) and methyl p-toluenesulfonate (65 mg, 0.35 mmol) were dissolved in acetonitrile (15 mL). Potassium carbonate (72 mg, 0.52 mmol) was added and stirred at room temperature overnight. TLC indicated a large amount of residual starting material. The reaction mixture was heated to 80°C and stirred for 8 hours. TLC indicated that the reaction was essentially complete. The reaction mixture was cooled to room temperature, water was added, and extraction with ethyl acetate was performed. The organic phases were combined, washed with saturated brine, dried over anhydrous sodium sulfate, and concentrated. The crude product was purified by Prep-TLC to afford the title compound 18d (37 mg, 21% yield over three steps).

[0566] LC-MS: m / z = 490.2 [M+H] +

[0567] Step 4: 1-(((R)-1-(3-amino-5-(trifluoromethyl)phenyl)ethyl)amino)-7-(6-methyl-3,6-diazabicyclo[3.1.1]hept-3-yl)pyrido[3,4-d]pyridazin-4(3H)-one 18

[0568] Compound 18d (37 mg, 0.076 mmol) was dissolved in ethanol (10 mL) and water (2 mL). Iron powder (21 mg, 0.38 mmol) and ammonium chloride (20 mg, 0.37 mmol) were added at room temperature. The mixture was heated to 90°C and stirred for 2 hours. TLC confirmed the reaction was complete. The reaction mixture was filtered while hot, and the filtrate was concentrated. The crude product was purified by Prep-TLC to afford the title compound 18 (20 mg, 57% yield).

[0569] LC-MS: m / z = 460.2 [M+H] +

[0570] 1H NMR(400MHz, DMSO-d6+D2O)δ9.03(s,1H),7.22(s,1H),6.97-6.73(m,3H),4.88(q,J=6.8Hz,1H),4.03- 3.61(m,6H),2.57(s,3H),2.32-2.22(m,1H),1.83-1.68(m,1H),1.52(d,J=6.8Hz,3H).(98.46% purity by HPLC)

[0571] Example 19

[0572] 1-(((R)-1-(3-amino-5-(trifluoromethyl)phenyl)ethyl)amino)-7-((R)-2,4-dimethylpiperazin-1-yl)pyrido[3,4-d]pyridazin-4(3H)-one 19

[0573]

[0574] Step 1 (R)-2,4-dimethylpiperazine-1-carboxylic acid tert-butyl ester 19b

[0575] Dissolve (R)-tert-Butyl 2-methylpiperazine-1-carboxylate 19a (12.0 g, 59.92 mmol) in methanol (50 mL). Add formaldehyde (9.7 g, 119.51 mmol, 37%) and palladium / carbon (500 mg, 10%). Stir at room temperature under a hydrogen atmosphere for 2.5 hours. TLC confirms the reaction is complete. The reaction mixture is filtered through a pad of Celite, and the filtrate is concentrated to afford the title compound 19b (12.2 g, crude) as a clear oil, which is used directly in the next step.

[0576] Step 2 (R)-1,3-Dimethylpiperazine dihydrochloride 19c

[0577] Compound 19b (12.2 g, crude) was dissolved in 20 mL of 4 M methanolic hydrochloric acid and stirred at room temperature for 30 minutes. TLC confirmed the reaction was complete. The reaction solution was concentrated to afford the title compound 19c (12.0 g, crude) as a slightly yellow oil, which was used directly in the next step.

[0578] Step 3 (R)-2,4-dimethylpiperazine-1-carboxylic acid benzyl ester 19d

[0579] Compound 19c (13.2 g, crude) was dissolved in tetrahydrofuran (130 mL). Water (50 mL) and sodium carbonate (37.4 g, 352.86 mmol) were added at room temperature. The temperature was lowered to 0°C, and benzyl chloroformate (20.9 g, 122.52 mmol) was added dropwise. After the mixture was warmed to room temperature and stirred for 1 hour, TLC confirmed the complete reaction. The reaction mixture was added with water and extracted with ethyl acetate. The organic phases were combined, washed with saturated brine, dried over anhydrous sodium sulfate, and concentrated. The crude product was purified by silica gel column chromatography to afford the title compound 19d (8.9 g, 54% yield over three steps) as a yellow oil.

[0580] Step 4 (R)-1,3-dimethylpiperazine 19e

[0581] Compound 19d (8.9 g, 35.84 mmol) was dissolved in tetrahydrofuran (80 mL), and palladium / carbon (2.0 g, 10%) was added. The mixture was stirred at room temperature under a hydrogen atmosphere for 1.5 hours. The reaction was complete by TLC. The reaction mixture was filtered through a pad of Celite, and the filtrate was concentrated to afford the title compound 19e (1.8 g, crude) as a yellow oil, which was used directly in the next step.

[0582] Step 5: 7-((R)-2,4-dimethylpiperazin-1-yl)-1-(((R)-1-(3-nitro-5-(trifluoromethyl)phenyl)ethyl)amino)pyrido[3,4-d]pyridazin-4(3H)-one 19f

[0583] Compound 19e (311 mg, crude) was dissolved in dimethyl sulfoxide (1.5 mL). Intermediate IN-3 (120 mg, 0.29 mmol) was added at room temperature. The mixture was heated to 120°C and allowed to react overnight. TLC confirmed the reaction was complete. The reaction solution was cooled to room temperature and water (3 mL) was added to precipitate a solid. The solid was filtered, washed, and dried to afford the title compound 19f (45 mg, crude) as a yellow solid, which was used directly in the next step.

[0584] LC-MS: m / z = 492.3 [M+H] +

[0585] Step 6: 1-(((R)-1-(3-amino-5-(trifluoromethyl)phenyl)ethyl)amino)-7-((R)-2,4-dimethylpiperazin-1-yl)pyrido[3,4-d]pyridazin-4(3H)-one 19

[0586] Compound 19f (45 mg, crude) was dispersed in ethanol and water (3 mL / 1 mL). Reduced iron powder (25 mg, 0.45 mmol) and ammonium chloride (24 mg, 0.45 mmol) were added at room temperature. The mixture was heated to 90°C for 3 hours. TLC confirmed the reaction was complete. The reaction mixture was hot and filtered through a pad of Celite. The filter cake was washed and the filtrate was concentrated. The crude product was purified by Prep-TLC to afford the title compound 19 (28 mg, 21% yield over three steps) as a yellow solid.

[0587] LC-MS: m / z=462.3[M+H] +

[0588] 1 H NMR (400MHz, CD3OD) δ9.05(s,1H),7.24(s,1H),6.99-6.87(m,2H),6.77(s,1H),5.05-4.91(m,2H),4.56(d,J=14.4Hz,1H),3.44-3.34(m,1 H),3.30-3.25(m,1H),3.22(d,J=12.0Hz,1H),2.85-2.73(m,1H),2.64(s,4H),1.57(d,J=6.8Hz,3H),1.37(d,J=6.8Hz,3H).(96.26% purity by HPLC)

[0589] Example 20

[0590] 1-(((R)-1-(3-amino-5-(trifluoromethyl)phenyl)ethyl)amino)-7-((S)-2,4-dimethylpiperazin-1-yl)pyrido[3,4-d]pyridazin-4(3H)-one 20

[0591]

[0592] Step 1-Benzyl 4-(tert-butyl)(S)-2-methylpiperazine-1,4-dicarboxylate 20b

[0593] (S)-4-N-tert-Butyloxycarbonyl-2-methylpiperazine 20a (3.0 g, 14.98 mmol) was dissolved in ethyl acetate (30 mL) and water (30 mL). Sodium bicarbonate (4.0 g, 47.61 mmol) was added at room temperature. The mixture was cooled to approximately 0°C, and benzyl chloroformate (3.8 g, 22.28 mmol) was added dropwise. The mixture was allowed to react at 0°C for 1 hour. The reaction mixture was extracted with ethyl acetate, and the combined organic phases were washed with saturated brine, dried over anhydrous sodium sulfate, and concentrated. The crude product was purified on a silica gel column to afford the title compound 20b (5.0 g, 99.8% yield) as a yellow oil.

[0594] Step 2 (S)-2-Methylpiperazine-1-carboxylic acid benzyl ester 20c

[0595] Compound 20b (5.0 g, 14.95 mmol) was dissolved in dichloromethane (50 mL), and trifluoroacetic acid (10 mL) was added. The mixture was stirred at room temperature for 1 hour. TLC indicated the reaction was complete. The reaction solution was concentrated, and the pH was adjusted to alkaline with saturated aqueous sodium bicarbonate. The mixture was extracted with ethyl acetate. The organic phases were combined, washed with saturated brine, dried over anhydrous sodium sulfate, and concentrated to afford the title compound 20c (3.7 g, crude) as a yellow oil, which was used directly in the next step.

[0596] LC-MS: m / z = 235.2 [M+H] +

[0597] Step 3 (S)-2,4-dimethylpiperazine-1-carboxylic acid benzyl ester 20d

[0598] Compound 20c (3.7 g, crude) was dissolved in formic acid (3.1 g, 67.35 mmol). Aqueous formaldehyde (2.4 g, 29.57 mmol, 37%) was added at room temperature and stirred overnight at 80°C. TLC indicated the reaction was complete. The reaction solution was cooled to room temperature, concentrated, and the pH was adjusted to alkaline with saturated aqueous sodium bicarbonate. The mixture was extracted with ethyl acetate. The combined organic phases were washed with saturated brine, dried over anhydrous sodium sulfate, and concentrated. The crude product was purified on a silica gel column to afford the title compound 20d (3.1 g, 84% yield over two steps) as a colorless liquid.

[0599] LC-MS: m / z = 249.2 [M+H] +

[0600] Step 4 (S)-1,3-dimethylpiperazine 20e

[0601] Compound 20d (3.0 g, 12.08 mmol) was dissolved in tetrahydrofuran (30 mL), and palladium / carbon (300 mg, 10%) was added. The mixture was stirred at room temperature overnight under a hydrogen atmosphere. TLC indicated complete reaction. The reaction mixture was filtered through a pad of Celite, and the filter cake was washed with ethyl acetate. The filtrate was concentrated to afford the title compound 20e (500 mg, crude) as a yellow oil, which was used directly in the next step.

[0602] Step 5: 7-((S)-2,4-dimethylpiperazin-1-yl)-1-(((R)-1-(3-nitro-5-(trifluoromethyl)phenyl)ethyl)amino)pyrido[3,4-d]pyridazin-4(3H)-one 20f

[0603] Intermediate IN-3 (80 mg, 0.19 mmol) was dissolved in dimethyl sulfoxide (0.5 mL). Compound 20e (230 mg, crude) was added at room temperature. The mixture was heated to 120°C and stirred overnight. TLC indicated that the starting material was consumed. The reaction mixture was cooled to room temperature, diluted with water, and extracted with ethyl acetate. The combined organic phases were washed with saturated brine, dried over anhydrous sodium sulfate, and concentrated. The crude product was purified on a silica gel column to afford the title compound 20f (90 mg, 95% yield) as a brown solid.

[0604] LC-MS: m / z = 492.3 [M+1] +

[0605] Step 6: 1-(((R)-1-(3-amino-5-(trifluoromethyl)phenyl)ethyl)amino)-7-((S)-2,4-dimethylpiperazin-1-yl)pyrido[3,4-d]pyridazin-4(3H)-one 20

[0606] Compound 20f (90 mg, 0.18 mmol) was dissolved in ethanol (4 mL) and water (2 mL). Iron powder (57 mg, 1.02 mmol) and ammonium chloride (55 mg, 1.03 mmol) were added at room temperature. After addition, the mixture was heated to 80°C and stirred for 1 hour. TLC indicated complete reaction. The reaction mixture was filtered while hot, and the filter cake was washed with ethanol. The filtrate was concentrated, diluted with water, and extracted with ethyl acetate. The organic phases were combined, washed with saturated brine, dried over anhydrous sodium sulfate, and concentrated. The crude product was purified by Prep-TLC to afford the title compound 20 (22 mg, 26% yield) as a yellow solid.

[0607] LC-MS: m / z=462.3[M+H] +

[0608] 1 H NMR (400MHz, CD3OD) δ9.06(s,1H),7.20(s,1H),6.92(s,2H),6.77(s,1H),4.94(q,J=6.8Hz,2H),4.47(d,J=12.4Hz,1H) ,3.37-3.33(m,1H),3.13-3.02(m,2H),2.54-2.31(m,5H),1.56(d,J=6.8Hz,3H),1.35(d,J=6.8Hz,3H).(96.68% purity by HPLC)

[0609] Example 21

[0610] 1-(((R)-1-(3-(difluoromethyl)-2-fluorophenyl)ethyl)amino)-7-(1-methylpiperidin-3-yl)pyrido[3,4-d]pyridazin-4-(3H)-one 21

[0611]

[0612] Step 1: (R)-5-(1-((1-(3-(difluoromethyl)-2-fluorophenyl)ethyl)amino)-4-oxo-3,4-dihydropyrido[3,4-d]pyridazin-7-yl)-3,6-dihydropyridine-1(2H)-carboxylic acid tert-butyl ester 21a

[0613] Compound 15h (110 mg, 0.30 mmol) and 1-tert-butyloxycarbonyl-3,6-dihydro-2H-pyridine-5-boronic acid pinacol ester 17a (113 mg, 0.37 mmol) were dissolved in 1,4-dioxane (4 mL) and water (1 mL). Sodium carbonate (52 mg, 0.49 mmol) and Pd(dppf)Cl2 (20 mg, 0.027 mmol) were added sequentially at room temperature. The atmosphere was purged with nitrogen several times, and the mixture was heated to 100°C and stirred for 3 hours. TLC indicated complete reaction of the starting materials. The reaction mixture was cooled to room temperature, diluted with water, and extracted with ethyl acetate. The organic phases were combined, washed with saturated brine, dried over anhydrous sodium sulfate, and concentrated. The crude product was purified on a silica gel column to afford the title compound 21a (120 mg, 78% yield) as a yellow solid.

[0614] LC-MS: m / z=516.3[M+H] +

[0615] Step 2 tert-Butyl 3-(1-(((R)-1-(3-(difluoromethyl)-2-fluorophenyl)ethyl)amino)-4-oxo-3,4-dihydropyrido[3,4-d]pyridazin-7-yl)piperidine-1-carboxylate 21b

[0616] Compound 21a (120 mg, 0.23 mmol) was dissolved in ethyl acetate (2 mL) and tetrahydrofuran (2 mL). Palladium / carbon (40 mg, 10%) was added at room temperature. The mixture was heated to 50°C and stirred overnight under a hydrogen atmosphere. The reaction mixture was cooled to room temperature, filtered through a pad of Celite, and the filter cake was washed with methanol. The filtrate was concentrated to afford the title compound 21b (110 mg, crude) as a white solid, which was used directly in the next step.

[0617] LC-MS: m / z = 518.3 [M+H] +

[0618] Step 3 1-(((R)-1-(3-(difluoromethyl)-2-fluorophenyl)ethyl)amino)-7-(piperidin-3-yl)pyrido[3,4-d]pyridazin-4(3H)-one 21c

[0619] Compound 21b (50 mg, crude) was dissolved in dichloromethane (2 mL) and trifluoroacetic acid (1 mL) was added. The mixture was stirred at room temperature for 1 hour. TLC indicated the reaction was complete. The reaction solution was concentrated, and the pH was adjusted to alkaline by adding methanolic ammonia (7 M). The mixture was concentrated, and the crude product was purified by Prep-TLC to afford the title compound 21c (30 mg, 69% yield over two steps) as a white solid.

[0620] LC-MS: m / z = 418.2 [M+H] +

[0621] Step 4: 1-(((R)-1-(3-(difluoromethyl)-2-fluorophenyl)ethyl)amino)-7-(1-methylpiperidin-3-yl)pyrido[3,4-d]pyridazin-4-(3H)-one 21

[0622] Compound 21c (30 mg, 0.072 mmol) was dispersed in methanol (2 mL), and aqueous formaldehyde (58 mg, 0.71 mmol, 37%) and palladium / carbon (20 mg, 10%) were added. The mixture was stirred overnight at room temperature under a hydrogen atmosphere. The reaction mixture was filtered through a pad of Celite, and the filter cake was washed several times with methanol. The filtrate was concentrated, and the crude product was purified by Prep-TLC to afford the title compound 21 (8 mg, 26% yield) as a white solid.

[0623] LC-MS: m / z=432.3[M+H] +

[0624] 1 H NMR (400MHz, CD3OD) δ9.42(s,1H),8.13(s,1H),7.57(t,J=7.2Hz,1H),7.44(t,J= 6.8Hz,1H),7.21(t,J=7.6Hz,1H),6.98(t,J=54.8Hz,1H),5.32(q,J=7.2Hz,1H),3 .45-3.31(m,2H),3.20-3.12(m,1H),2.98-2.78(m,1H),2.62(s,3H),2.54-2.42(m ,1H),2.17-2.14(m,1H),1.96-1.84(m,3H),1.63(d,J=6.8Hz,3H).(96.73% purity by HPLC)

[0625] Example 22

[0626] 1-(((R)-1-(3-(difluoromethyl)-2-fluorophenyl)ethyl)amino)-3-methyl-7-(1-methylpiperidin-3-yl)pyrido[3,4-d]pyridazin-4-(3H)-one 22

[0627]

[0628] Step 1: tert-Butyl 3-(1-(((R)-1-(3-(difluoromethyl)-2-fluorophenyl)ethyl)amino)-3-methyl-4-oxo-3,4-dihydropyrido[3,4-d]pyridazin-7-yl)piperidine-1-carboxylate 22a

[0629] Compound 21b (80 mg, 0.15 mmol) was dissolved in N,N-dimethylformamide (1 mL). Potassium carbonate (43 mg, 0.31 mmol) and iodomethane (33 mg, 0.23 mmol) were added sequentially. The mixture was stirred at room temperature for 4 hours. TLC indicated complete reaction. The reaction mixture was diluted with water and extracted with ethyl acetate. The organic phases were combined, washed with water and saturated brine, dried over anhydrous sodium sulfate, and concentrated to afford the title compound 22a (100 mg, crude) as a white solid, which was used directly in the next step.

[0630] Step 2 1-(((R)-1-(3-(difluoromethyl)-2-fluorophenyl)ethyl)amino)-3-methyl-7-(piperidin-3-yl)pyrido[3,4-d]pyridazin-4(3H)-one 22b

[0631] Compound 22a (100 mg, crude) was dissolved in dichloromethane (2 mL) and trifluoroacetic acid (1 mL) was added. The mixture was stirred at room temperature for 1 hour. TLC indicated the reaction was complete. The reaction solution was concentrated, and the pH was adjusted to alkaline by adding methanolic ammonia (7 M). The mixture was then concentrated, and the crude product was purified by Prep-TLC to afford the title compound 22b (50 mg, 75% yield over two steps) as a white solid.

[0632] LC-MS: m / z = 432.2 [M+H] +

[0633] Step 3 1-(((R)-1-(3-(difluoromethyl)-2-fluorophenyl)ethyl)amino)-3-methyl-7-(1-methylpiperidin-3-yl)pyrido[3,4-d]pyridazin-4-(3H)-one 22

[0634] Compound 22b (50 mg, 0.12 mmol) was dispersed in methanol (2 mL), and aqueous formaldehyde (94 mg, 1.16 mmol, 37%) and palladium / carbon (30 mg, 10%) were added. The mixture was stirred overnight at room temperature under a hydrogen atmosphere. The reaction mixture was filtered through a pad of Celite, and the filter cake was washed several times with methanol. The filtrate was concentrated, and the crude product was purified by Prep-TLC to afford the title compound 22 (13 mg, 25% yield) as a pale yellow solid.

[0635] LC-MS: m / z=446.3[M+H] +

[0636] 1 H NMR (400MHz, CD3OD) δ9.40(s,1H),8.07(s,1H),7.59(t,J=7.2Hz,1H),7.45(t,J=7.2H z,1H),7.21(t,J=7.6Hz,1H),7.01(t,J=54.8Hz,1H),5.31(q,J=6.8Hz,1H),3.50(s,3 H),3.28-3.18(m,2H),3.06-2.98(m,1H),2.52-2.47(m,1H),2.43(s,3H),2.29-2.15( m,1H),2.13-2.05(m,1H),1.93-1.71(m,3H),1.64(d,J=6.8Hz,3H).(99.14%purityby HPLC)

[0637] Example 23

[0638] (R)-1-(((1-(4-(2-(((methylamino)methyl)phenyl)thiophen-2-yl)ethyl)amino)-7-(4-methylpiperazin-1-yl)pyrido[3,4-d]pyridazin-4(3H)-one 23

[0639]

[0640] Step 1 (2-((methylamino)methyl)phenyl)boronic acid 23b

[0641] o-Formylboronic acid 23a (5.0 g, 33.35 mmol) was dissolved in ethanolic methylamine (30 mL, 7 M) and palladium / carbon (200 mg, 10%) was added. The mixture was reacted at room temperature under a hydrogen atmosphere for 5 hours. TLC confirmed the complete reaction. The reaction mixture was filtered through a pad of Celite, the filter cake was washed, and the filtrate was concentrated to afford the title compound 23b (5.3 g, crude) as a pale yellow, foamy solid, which was used directly in the next step.

[0642] Step 2 (2-(((tert-Butoxycarbonyl)(methyl)amino)methyl)phenyl)boronic acid 23c

[0643] Compound 23b (5.3 g, crude) was dissolved in tetrahydrofuran (100 mL) and water (20 mL). Sodium carbonate (10.3 g, 97.18 mmol) was added at room temperature. The mixture was cooled to 0°C and di-tert-butyl dicarbonate (8.5 g, 38.95 mmol) was added dropwise. The mixture was allowed to react overnight at room temperature. TLC confirmed the reaction was complete. The reaction mixture was added with water, extracted with ethyl acetate, dried over anhydrous sodium sulfate, and concentrated. The crude product was slurried with petroleum ether and ethyl acetate, filtered, and the filter cake was washed and dried to afford the title compound 23c (7.4 g, 84% yield over two steps) as a white solid.

[0644] 1 H NMR(400MHz, DMSO-d6)δ8.14(s,2H),7.50(dd,J=7.2,0.8Hz,1H),7.38-7.30(m,1H), 7.21(t,J=7.2Hz,1H),7.08(d,J=7.6Hz,1H),4.56(s,2H),2.74(s,3H),1.40(s,9H).

[0645] Step 3 (R)-1-(4-bromothiophen-2-yl)ethan-1-amine 23e

[0646] Compound 23d (1.1 g, 3.59 mmol) was dissolved in methanolic hydrochloric acid (10 mL, 4 mol / L) and allowed to react at room temperature for 1 hour. TLC confirmed the complete reaction. The reaction solution was neutralized with saturated sodium bicarbonate aqueous solution and extracted with ethyl acetate. The organic phases were combined, washed with saturated brine, dried over anhydrous sodium sulfate, and concentrated to afford the title compound 23e (1.0 g, crude) as a yellow oil, which was used directly in the next step.

[0647] Step 4: (R)-tert-Butyl (2-(5-(1-aminoethyl)thiophen-3-yl)benzyl)(methyl)carbamate 23f

[0648] Compound 23e (1.0 g, crude) and compound 23c (1.9 g, 7.17 mmol) were dissolved in 1,4-dioxane (20 mL) and water (5 mL). Sodium carbonate (1.5 g, 14.15 mmol) and Pd(dppf)Cl2 dichloromethane complex (200 mg, 0.24 mmol) were added at room temperature. The atmosphere was purged with nitrogen three times and heated to 100°C for 4 hours. TLC confirmed complete reaction. The reaction mixture was cooled to room temperature, water was added, and extraction with ethyl acetate was performed. The organic phases were combined, washed with saturated brine, dried over anhydrous sodium sulfate, and concentrated. The crude product was purified by column chromatography to afford the title compound 23f (800 mg, 64% yield over two steps) as a red oil.

[0649] LC-MS: m / z=347.2[M+H] +

[0650] Step 5: tert-Butyl (R)-(2-(5-(1-((6-chloro-3-oxo-3H-pyrrolo[3,4-c]pyridin-1-yl)amino)ethyl)thiophen-3-yl)benzyl)(methyl)carbamate 23g

[0651] Compound 23f (800 mg, 2.31 mmol) and compound IN-3h (400 mg, 2.20 mmol) were dissolved in isopropanol (20 mL) and heated to 90°C with stirring overnight. TLC indicated the reaction was nearly complete. The reaction mixture was cooled to room temperature and concentrated to afford the title compound 23g (1.1 g, crude) as a brown, foamy solid, which was used directly in the next step.

[0652] Step 6: (R)-tert-butyl (2-(5-(1-((7-chloro-4-oxo-3,4-dihydropyrido[3,4-d]pyridazin-1-yl)amino)ethyl)thiophen-3-yl)benzyl)(methyl)carbamate 23h

[0653] Compound 23g (1.1 g, crude) was dissolved in methanol (15 mL). Hydrazine hydrate (269 mg, 4.30 mmol, 80%) was added at room temperature and heated to 40°C for 2 hours. The reaction was complete by TLC. The reaction mixture was cooled to room temperature, filtered, and the filtrate was concentrated. The crude product was purified by silica gel column chromatography to afford the title compound 23h as a yellow-brown oil (910 mg, 79% yield over two steps).

[0654] Step 7: Tert-butyl (R)-methyl (2-(5-(1-((7-(4-methylpiperazin-1-yl)-4-oxo-3,4-dihydropyrido[3,4-d]pyridazin-1-yl)amino)ethyl)thiophen-3-yl)benzyl)carbamate 23i

[0655] Compound 23h (100 mg, 0.17 mmol) and N-methylpiperazine 2b (95 mg, 0.95 mmol) were dissolved in dimethyl sulfoxide (1 mL) and heated to 80°C for 2 hours. The reaction was complete by TLC. The reaction solution was cooled to room temperature, added with water, and extracted with ethyl acetate. The organic phases were combined, washed with saturated brine, dried over anhydrous sodium sulfate, and concentrated to afford the title compound 23i (75 mg, crude) as a yellow solid, which was used directly in the next step.

[0656] Step 8 (R)-1-(((1-(4-(2-(((methylamino)methyl)phenyl)thiophen-2-yl)ethyl)amino)-7-(4-methylpiperazin-1-yl)pyrido[3,4-d]pyridazin-4(3H)-one 23

[0657] Compound 23i (75 mg, crude) was dissolved in a hydrochloric acid / 1,4-dioxane solution (3 mL, 4 M) and allowed to react at room temperature for 1 hour. TLC confirmed the reaction was complete. The reaction solution was neutralized with saturated aqueous sodium carbonate and extracted with ethyl acetate. The combined organic phases were washed with saturated brine, dried over anhydrous sodium sulfate, and concentrated. The crude product was purified by Prep-TLC to afford the title compound 23 (17 mg, 20% yield over two steps) as a yellow solid.

[0658] LC-MS: m / z = 490.3 [M+H] +

[0659] 1 H NMR (400MHz, DMSO-d6) δ11.31(s,1H),8.93(s,1H),7.50-7.46(m,1H),7.44(d,J=1.2Hz,1H),7.34-7.29(m,3H),7.23(d,J=8.0Hz,2H),7.00(d,J= 7.6Hz,1H),5.41-5.25(m,1H),3.76-3.69(m,4H),3.68(s,2H),2.47-2.4 0(m,4H),2.28(s,3H),2.24(s,3H),1.67(d,J=6.8Hz,3H).(99.72% purity by HPLC)

[0660] Example 24

[0661] 1-(((R)-1-(4-(2-((methylamino)methyl)phenyl)thiophen-2-yl)ethyl)amino)-7-(((S)-tetrahydrofuran-3-yl)oxy)pyrido[3,4-d]pyridazin-4(3H)-one 24

[0662]

[0663] First step: tert-Butyl methyl(2-(5-((R)-1-((4-oxo-7-(((S)-tetrahydrofuran-3-yl)oxy)-3,4-dihydropyrido[3,4-d]pyridazin-1-yl)amino)ethyl)thiophen-3-yl)benzyl)carbamate 24a

[0664] (S)-(+)-3-Hydroxytetrahydrofuran (1 mL) was cooled to 0°C, and sodium hydroxide (76 mg, 1.90 mmol, 60%) was added. The mixture was stirred at room temperature for 1 hour, and the reaction solution turned from an emulsion to an orange transparent solution. Compound 23h (100 mg, 0.19 mmol) was added, and the temperature was raised to 80°C and stirred overnight. The reaction solution was quenched with water and extracted with ethyl acetate. The organic phases were combined, washed with water and saturated brine, dried over anhydrous sodium sulfate, and concentrated. The crude product was purified on a silica gel column to obtain the title compound 24a (100 mg, 91% yield) as a white solid.

[0665] LC-MS: m / z = 578.3 [M+H] +

[0666] Step 2 1-(((R)-1-(4-(2-((methylamino)methyl)phenyl)thiophen-2-yl)ethyl)amino)-7-(((S)-tetrahydrofuran-3-yl)oxy)pyrido[3,4-d]pyridazin-4(3H)-one 24

[0667] Compound 24a (100 mg, 0.17 mmol) was dissolved in a hydrochloric acid / 1,4-dioxane solution (1 mL, 4 M) and stirred at room temperature for 1 hour. TLC indicated the disappearance of the starting material. The reaction mixture was adjusted to an alkaline pH by adding saturated sodium bicarbonate aqueous solution. The mixture was extracted with ethyl acetate. The combined organic phases were washed with saturated brine, dried over anhydrous sodium sulfate, and concentrated. The crude product was purified by Prep-TLC to afford the title compound 24 (7 mg, 8% yield) as a yellow solid.

[0668] LC-MS: m / z = 478.2 [M+H] +

[0669] 1 H NMR(400MHz,CD3OD)δ9.12(s,1H),7.44-7.36(m,2H),7.34-7.25(m,3H),7.14(s,1H),7.11(s,1H),5.77-5.68(m,1H),5.39 -5.29(m,1H),4.07-3.80(m,6H),2.39-2.30(m,1H),2.24(s,3H),2.20-2.14(m,1H),1.72(d,J=6.8Hz,3H).(88.28%purity by HPLC)

[0670] Example 25

[0671] 1-(((R)-1-(3-amino-5-(trifluoromethyl)phenyl)ethyl)amino)-7-(3-(dimethylamino)pyrrolidin-1-yl)pyrido[3,4-d]pyridazin-4(3H)-one 25

[0672]

[0673] Step 1: tert-Butyl 3-(dimethylamino)pyrrolidine-1-carboxylate 25b

[0674] Tert-butyl 3-aminopyrrolidine-1-carboxylate 25a (800 mg, 4.30 mmol) was dispersed in methanol (10 mL). Aqueous formaldehyde (3.5 g, 43.12 mmol, 37%) and palladium / carbon (200 mg, 10%) were added, and the mixture was stirred overnight at room temperature under a hydrogen atmosphere. The reaction mixture was filtered through a pad of celite, and the filter cake was washed several times with methanol. The filtrate was concentrated, and the crude product was purified by silica gel column chromatography to afford the title compound 25b (900 mg, 98% yield) as a yellow liquid.

[0675] LC-MS: m / z = 215.2 [M+H] +

[0676] 1 H NMR (400MHz, CDCl3) δ3.52-3.44(m,1H),3.23-3.17(m,1H),3.04-2.96(m,1H),2. 63-2.54(m,1H),2.19(s,6H),2.01-1.95(m,1H),1.74-1.55(m,2H),1.39(s,6H).

[0677] Step 2: N,N-dimethylpyrrolidin-3-amine 25c

[0678] Compound 25b (900 mg, 4.20 mmol) was dissolved in a hydrochloric acid / 1,4-dioxane solution (10 mL, 4 M) and stirred at room temperature for 1 hour. TLC indicated complete reaction. The reaction mixture was concentrated to afford the title compound 25c (700 mg, crude) as a brown liquid, which was used directly in the next step.

[0679] LC-MS: m / z = 115.2 [M+H] +

[0680] Step 3: 7-(3-(Dimethylamino)pyrrolidin-1-yl)-1-(((R)-1-(3-nitro-5-(trifluoromethyl)phenyl)ethyl)amino)pyrido[3,4-d]pyridazin-4(3H)-one 25d

[0681] Intermediate IN-3 (70 mg, 0.17 mmol) was dissolved in dimethyl sulfoxide (0.5 mL). Compound 25c (58 mg, crude) and N,N-diisopropylethylamine (87 mg, 0.68 mmol) were added at room temperature. The mixture was heated to 80°C and stirred overnight. TLC confirmed the disappearance of the starting material. The reaction mixture was added with water and extracted with ethyl acetate. The combined organic phases were washed with saturated brine, dried over anhydrous sodium sulfate, and concentrated. The crude product was purified on a silica gel column to afford the title compound 25d (50 mg, 60% yield) as a brown solid.

[0682] LC-MS: m / z = 492.2 [M+H] +

[0683] Step 4: 1-(((R)-1-(3-amino-5-(trifluoromethyl)phenyl)ethyl)amino)-7-(3-(dimethylamino)pyrrolidin-1-yl)pyrido[3,4-d]pyridazin-4(3H)-one 25

[0684] Compound 25d (50 mg, 0.10 mmol) was dissolved in ethanol (2 mL) and water (1 mL). Iron powder (28 mg, 0.50 mmol) and ammonium chloride (27 mg, 0.50 mmol) were added at room temperature. The mixture was heated to 80°C and stirred for 1 hour. TLC indicated complete reaction. The reaction mixture was filtered while hot, the filter cake was washed with ethanol, the filtrate was concentrated, diluted with water, and extracted with ethyl acetate. The organic phases were combined, washed with saturated brine, dried over anhydrous sodium sulfate, and concentrated. The crude product was purified by Prep-TLC to afford the title compound 25 (30 mg, 64% yield) as a yellow solid.

[0685] LC-MS: m / z=462.3[M+H] +

[0686] 1 H NMR (400MHz, CD3OD) δ8.98(s,1H),6.94(s,1H),6.92(s,1H),6.87(s,1H),6.77(s,1H),4.95(q,J=7.2Hz,1H),3.92-3.83(m,2H),3.57-3. 48(m,1H),3.40-3.33(m,1H),3.10-3.05(m,1H),2.44(s,6H),2.41-2.34(m,1H),2.07-1.93(m,1H),1.57(d,J=7.2Hz,3H).(99.28% purity by HPLC)

[0687] Example 26

[0688] 1-(((R)-1-(3-(difluoromethyl)-2-fluorophenyl)ethyl)amino)-7-(3-(dimethylamino)pyrrolidin-1-yl)pyrido[3,4-d]pyridazin-4(3H)-one 26

[0689]

[0690] Compound 15h (60 mg, 0.16 mmol) was dissolved in dimethyl sulfoxide (0.5 mL). Compound 25c (56 mg, 0.49 mmol) and N,N-diisopropylethylamine (84 mg, 0.65 mmol) were added at room temperature. The mixture was heated to 80°C and stirred overnight. TLC indicated the disappearance of the starting material. The reaction mixture was added with water and extracted with ethyl acetate. The combined organic phases were washed with saturated brine, dried over anhydrous sodium sulfate, and concentrated. The crude product was purified by Prep-TLC to afford the title compound 26 (25 mg, 34% yield) as a gray solid.

[0691] LC-MS: m / z = 447.2 [M+H] +

[0692] 1 H NMR(400MHz,CD3OD)δ9.00(s,1H),7.56(t,J=7.6Hz,1H),7.43(t,J=6.8Hz,1H ),7.20(t,J=8.0Hz,1H),7.11-6.84(m,2H),5.30(q,J=6.8Hz,1H),4.04-3.99 (m,1H),3.91-3.87(m,1H),3.63-3.50(m,2H),3.41-3.38(m,1H),2.62(s,6H) ,2.52-2.45(m,1H),2.17-2.07(m,1H),1.62(d,J=7.2Hz,3H).(99.44% purity by HPLC)

[0693] Example 27

[0694] 1-(((R)-1-(3-amino-5-(trifluoromethyl)phenyl)ethyl)amino)-7-(1-methylpyrrolidin-3-yl)pyrido[3,4-d]pyridazin-4(3H)-one 27

[0695]

[0696] Step 1: (R)-3-(1-((1-(3-nitro-5-(trifluoromethyl)phenyl)ethyl)amino)-4-oxo-3,4-dihydropyrido[3,4-d]pyridazin-7-yl)-2,5-dihydro-1H-pyrrole-1-carboxylic acid tert-butyl ester 27b

[0697] Intermediate IN-3 (100 mg, 0.24 mmol) was dissolved in 1,4-dioxane (5 mL) and water (1 mL). 1-tert-Butyloxycarbonyl-2,5-dihydro-1H-pyrrole-3-boronic acid pinacol ester 27a (85 mg, 0.29 mmol), sodium carbonate (76 mg, 0.72 mmol), and Pd(dppf)Cl2 dichloromethane complex (20 mg, 0.024 mmol) were added at room temperature. The atmosphere was purged with nitrogen three times and heated to 100°C for 3 hours. TLC confirmed the reaction was complete. The reaction mixture was cooled to room temperature, water was added, and extraction with ethyl acetate was performed. The organic phases were combined, washed with saturated brine, dried over anhydrous sodium sulfate, and concentrated. The crude product was purified by silica gel column chromatography to afford the title compound 27b (115 mg, 87% yield) as a yellow solid.

[0698] LC-MS: m / z=547.3[M+H] +

[0699] Step 2 tert-Butyl 3-(1-(((R)-1-(3-amino-5-(trifluoromethyl)phenyl)ethyl)amino)-4-oxo-3,4-dihydropyrido[3,4-d]pyridazin-7-yl)pyrrolidine-1-carboxylate 27c

[0700] Compound 27b (115 mg, 0.21 mmol) was dissolved in ethyl acetate (10 mL). Palladium / carbon (20 mg, 10%) was added at room temperature. The mixture was reacted at 50°C under a hydrogen atmosphere for 8 hours. LCMS confirmed the complete reaction. The reaction mixture was cooled to room temperature, filtered through a pad of Celite, and the filter cake was washed. The filtrate was concentrated to afford the title compound 27c (100 mg, crude) as a yellow solid, which was used directly in the next step.

[0701] LC-MS: m / z = 519.3 [M+H] +

[0702] Step 3 tert-Butyl 3-(1-(((R)-1-(3-acetylamino-5-(trifluoromethyl)phenyl)ethyl)amino)-4-oxo-3,4-dihydropyrido[3,4-d]pyridazin-7-yl)pyrrolidine-1-carboxylate 27d

[0703] Compound 27c (100 mg, crude) was dissolved in dichloromethane (10 mL). N,N-diisopropylethylamine (65 mg, 0.50 mmol) and acetic anhydride (38 mg, 0.37 mmol) were added at room temperature. The mixture was heated to 30°C for 4 hours. TLC indicated that the reaction was essentially complete. The reaction mixture was cooled to room temperature and extracted with dichloromethane. The organic phases were combined, washed with saturated brine, dried over anhydrous sodium sulfate, and concentrated. The crude product was purified by Prep-TLC to afford the title compound 27d (50 mg, 42% yield over two steps) as a yellow solid.

[0704] Step 4: N-(3-((1R)-1-((4-oxo-7-(pyrrolidin-3-yl)-3,4-dihydropyrido[3,4-d]pyridazin-1-yl)amino)ethyl)-5-(trifluoromethyl)phenyl)acetamide 27e

[0705] Compound 27d (50 mg, 0.096 mmol) was dissolved in dichloromethane (2 mL) and trifluoroacetic acid (1 mL) was added. The mixture was allowed to react at room temperature for 2 hours. TLC confirmed the complete reaction. The reaction solution was neutralized with saturated sodium bicarbonate aqueous solution and extracted with ethyl acetate. The organic phases were combined, washed with saturated brine, dried over anhydrous sodium sulfate, and concentrated to afford the title compound 27e (38 mg, crude) as a yellow solid, which was used directly in the next step.

[0706] Step 5: N-(3-((1R)-1-((7-(1-methylpyrrolidin-3-yl)-4-oxo-3,4-dihydropyrido[3,4-d]pyridazin-1-yl)amino)ethyl)-5-(trifluoromethyl)phenyl)acetamide 27f

[0707] Compound 27e (38 mg, crude) was dissolved in methanol (3 mL), and aqueous formaldehyde solution (3d) and palladium / carbon (15 mg, 10%) were added. The mixture was allowed to react at room temperature under a hydrogen atmosphere for 2 hours. TLC indicated that the reaction was essentially complete. The reaction mixture was filtered through a pad of celite, and the filtrate was concentrated to afford the title compound 27f (54 mg, crude), which was used directly in the next step.

[0708] Step 6 1-(((R)-1-(3-amino-5-(trifluoromethyl)phenyl)ethyl)amino)-7-(1-methylpyrrolidin-3-yl)pyrido[3,4-d]pyridazin-4(3H)-one 27

[0709] Compound 27f (54 mg, crude) was dissolved in ethanol (3 mL). Aqueous sodium hydroxide (3 mL, 12 mmol, 4 M) was added at room temperature and heated to 80°C overnight. TLC indicated near-complete reaction. The reaction mixture was cooled to room temperature, added with water, and extracted with ethyl acetate. The combined organic phases were washed with saturated brine, dried over anhydrous sodium sulfate, and concentrated. The crude product was purified by Prep-TLC to afford the title compound 27 (8 mg, 19% yield over three steps) as a pale yellow solid.

[0710] LC-MS: m / z = 433.2 [M+H] +

[0711] 1 HNMR(400MHz,CD3OD)δ9.43(s,1H),8.07(s,1H),6.93(s,2H),6.77(s,1H),4.95(q,J=6.8Hz,1H),4.05-3.75(m,1H),3.40 -3.36(m,1H),3.12-3.06(m,3H),2.63(s,3H),2.59-2.42(m,1H),2.27-2.22(m,1H),1.58(d,J=6.8Hz,3H).(97.64% purity by HPLC)

[0712] Example 28

[0713] 1-(((R)-1-(3-amino-5-(trifluoromethyl)phenyl)ethyl)amino)-3-methyl-7-(1-methylpiperidin-3-yl)pyrido[3,4-d]pyridazin-4(3H)-one 28

[0714]

[0715] Step 1: (R)-5-(3-methyl-1-((1-(3-nitro-5-(trifluoromethyl)phenyl)ethyl)amino)-4-oxo-3,4-dihydropyrido[3,4-d]pyridazin-7-yl)-3,6-dihydropyridine-1(2H)-carboxylic acid tert-butyl ester 28a

[0716] Compound 4a (200 mg, 0.47 mmol) and 1-tert-butyloxycarbonyl-3,6-dihydro-2H-pyridine-5-boronic acid pinacol ester 17a (217 mg, 0.70 mmol) were dissolved in 1,4-dioxane (15 mL) and water (3 mL). Pd(dppf)Cl2 dichloromethane complex (18 mg, 0.022 mmol) and sodium carbonate (148 mg, 1.40 mmol) were added at room temperature. The mixture was heated to 100°C and stirred for 1 hour. TLC confirmed the reaction was complete. The reaction solution was cooled to room temperature, water was added, and extraction with ethyl acetate was performed. The organic phases were combined, washed with saturated brine, dried over anhydrous sodium sulfate, and concentrated. The crude product was purified by silica gel column chromatography to afford the title compound 28a (282 mg, crude) as a yellow solid, which was used directly in the next step.

[0717] LC-MS: m / z = 575.3 [M+H] +

[0718] Step 2 tert-Butyl 3-(1-(((R)-1-(3-amino-5-(trifluoromethyl)phenyl)ethyl)amino)-3-methyl-4-oxo-3,4-dihydropyrido[3,4-d]pyridazin-7-yl)piperidine-1-carboxylate 28b

[0719] Compound 28a (282 mg, crude) was dissolved in ethyl acetate (10 mL), and palladium / carbon (100 mg, 10%) was added. The mixture was reacted at 50°C overnight under a hydrogen atmosphere. LCMS confirmed the complete reaction. The reaction mixture was cooled to room temperature, filtered through a pad of Celite, and the filter cake was washed with methanol. The filtrate was concentrated to afford the title compound 28b (290 mg, crude) as a yellow solid, which was used directly in the next step.

[0720] LC-MS: m / z=547.3[M+H] +

[0721] Step 3 tert-Butyl 3-(1-(((R)-1-(3-acetylamino-5-(trifluoromethyl)phenyl)ethyl)amino)-3-methyl-4-oxo-3,4-dihydropyrido[3,4-d]pyridazin-7-yl)piperidine-1-carboxylate 28c

[0722] Compound 28b (290 mg, crude) was dissolved in dichloromethane (3 mL). Acetic anhydride (81 mg, 0.79 mmol) and triethylamine (107 mg, 1.06 mmol) were added at room temperature. The reaction mixture was heated to 30°C for 4 hours. TLC indicated complete reaction. The reaction solution was concentrated, and Prep-TLC analysis of the crude product afforded the title compound 28c (152 mg, 55% yield over three steps) as a pale yellow solid.

[0723] LC-MS: m / z = 589.3 [M+H] +

[0724] Step 4: N-(3-((1R)-1-((3-methyl-4-oxo-7-(piperidin-3-yl)-3,4-dihydropyrido[3,4-d]pyridazin-1-yl)amino)ethyl)-5-(trifluoromethyl)phenyl)acetamide 28d

[0725] Compound 28c (152 mg, 0.26 mmol) was dissolved in dichloromethane (2 mL) and trifluoroacetic acid (2 mL) was added. The mixture was allowed to react at room temperature for 2 hours. TLC indicated complete reaction. The reaction mixture was adjusted to alkaline with saturated sodium bicarbonate aqueous solution and extracted with ethyl acetate. The organic phases were combined, washed with saturated brine, dried over anhydrous sodium sulfate, and concentrated to afford the title compound 28d (143 mg, crude) as a yellow solid, which was used directly in the next step.

[0726] Step 5: N-(3-((1R)-1-((3-methyl-7-(1-methylpiperidin-3-yl)-4-oxo-3,4-dihydropyrido[3,4-d]pyridazin-1-yl)amino)ethyl)-5-(trifluoromethyl)phenyl)acetamide 28e

[0727] Compound 28d (143 mg, crude) was dissolved in methanol (5 mL), and aqueous formaldehyde (71 mg, 0.87 mmol, 37%) and palladium / carbon (70 mg, 10%) were added. The mixture was reacted at room temperature under a hydrogen atmosphere for 2 hours. TLC indicated complete reaction. The reaction mixture was filtered through a pad of Celite, the filter cake was washed with methanol, and the filtrate was concentrated to afford the title compound 28e (190 mg, crude) as a yellow oil, which was used directly in the next step.

[0728] LC-MS: m / z=503.3[M+H] +

[0729] Step 6: 1-(((R)-1-(3-amino-5-(trifluoromethyl)phenyl)ethyl)amino)-3-methyl-7-(1-methylpiperidin-3-yl)pyrido[3,4-d]pyridazin-4(3H)-one 28

[0730] Compound 28e (190 mg, crude) was dissolved in methanol (4 mL) and aqueous sodium hydroxide solution (1.89 ml, 7.56 mmol, 4 N) was added at room temperature. The mixture was heated to 80°C and allowed to react overnight. TLC indicated complete reaction of the starting material. The reaction mixture was cooled to room temperature, added with water, and extracted with ethyl acetate. The organic phases were combined, washed with saturated brine, dried over anhydrous sodium sulfate, and concentrated. The crude product was purified by Prep-TLC to afford the title compound 28 (47 mg, 39% yield over three steps) as a white solid.

[0731] LC-MS: m / z = 461.3 [M+H] +

[0732] 1 H NMR (400MHz, CD3OD) δ9.41 (s, 1H), 8.07 (s, 1H), 6.97 (s, 2H), 6.78 (s, 1H), 4.96 (q, J = 6.8Hz, 1H), 3.56 (s, 3H), 3.38-3.26 (m, 2H), 3.15-3. 08(m,1H),2.82-2.65(m,1H),2.55(s,3H),2.50-2.36(m,1H),2.16-2.07(m,1H),1.99-1.70(m,3H),1.60(d,J=7.2Hz,3H).(99.78% purity by HPLC)

[0733] Example 29

[0734] 4-(((R)-1-(3-amino-5-(trifluoromethyl)phenyl)ethyl)amino)-6-(1-methylpiperidin-3-yl)phthalazin-1(2H)-one 29

[0735]

[0736] Step 1: (R)-5-(4-((1-(3-nitro-5-(trifluoromethyl)phenyl)ethyl)amino)-1-oxo-1,2-dihydrophthalazin-6-yl)-3,6-dihydropyridine-1(2H)-carboxylic acid tert-butyl ester 29a

[0737] Intermediate IN-2 (300 mg, 0.66 mmol) and 1-tert-butyloxycarbonyl-3,6-dihydro-2H-pyridine-5-boronic acid pinacol ester 17a (325 mg, 1.05 mmol) were dissolved in 1,4-dioxane (6 mL) and water (2 mL). Sodium carbonate (150 mg, 1.42 mmol) and Pd(dppf)Cl2 (58 mg, 0.08 mmol) were added sequentially at room temperature. The atmosphere was purged with nitrogen several times, and the mixture was heated to 100°C and stirred for 1 hour. TLC indicated complete reaction of the starting materials. The reaction mixture was cooled to room temperature, diluted with water, and extracted with ethyl acetate. The organic phases were combined, washed with saturated brine, dried over anhydrous sodium sulfate, and concentrated. The crude product was purified on a silica gel column to afford the title compound 29a (260 mg, 70% yield) as a yellow solid.

[0738] LC-MS: m / z = 560.3 [M+H] +

[0739] Step 2 tert-Butyl 3-(4-(((R)-1-(3-amino-5-(trifluoromethyl)phenyl)ethyl)amino)-1-oxo-1,2-dihydrophthalazin-6-yl)piperidine-1-carboxylate 29b

[0740] Compound 29a (250 mg, 0.45 mmol) was dissolved in ethyl acetate (8 mL), and palladium / carbon (60 mg, 10%) was added. The mixture was heated to 50°C and stirred overnight under a hydrogen atmosphere. The reaction mixture was filtered through a pad of Celite, and the filter cake was washed several times with methanol. The filtrate was concentrated, and the crude product was purified on a silica gel column to afford the title compound 29b (210 mg, 88% yield) as a white solid.

[0741] LC-MS: m / z = 532.3 [M+H] +

[0742] Step 3 tert-Butyl 3-(4-(((R)-1-(3-acetylamino-5-(trifluoromethyl)phenyl)ethyl)amino)-1-oxo-1,2-dihydrophthalazin-6-yl)piperidine-1-carboxylate 29c

[0743] Compound 29b (210 mg, 0.40 mmol) was dissolved in dichloromethane (5 mL). N,N-diisopropylethylamine (102 mg, 0.79 mmol) and acetic anhydride (61 mg, 0.60 mmol) were added sequentially at room temperature. The mixture was heated to 30°C and stirred for 5 hours. TLC indicated that the reaction was essentially complete. The reaction solution was diluted with water and extracted with dichloromethane. The organic phases were combined, washed with saturated brine, dried over anhydrous sodium sulfate, and concentrated to afford the title compound 29c (250 mg, crude) as a white solid, which was used directly in the next step.

[0744] LC-MS: m / z = 574.3 [M+H] +

[0745] Step 4: N-(3-((1R)-1-((4-oxo-7-(piperidin-3-yl)-3,4-dihydrophthalazin-1-yl)amino)ethyl)-5-(trifluoromethyl)phenyl)acetamide 29d

[0746] Compound 29c (250 mg, crude) was dissolved in dichloromethane (5 mL), and trifluoroacetic acid (2 mL) was added. The mixture was stirred at room temperature for 1 hour. TLC indicated the reaction was complete. The reaction solution was concentrated, and the pH was adjusted to a weak base with saturated sodium bicarbonate aqueous solution. The mixture was extracted with ethyl acetate, and the organic phases were combined, washed with saturated brine, dried over anhydrous sodium sulfate, and concentrated to afford the title compound 29d (200 mg, crude) as a white solid, which was used directly in the next step.

[0747] LC-MS: m / z=474.3[M+H] +

[0748] Step 5: N-(3-((1R)-1-((7-(1-methylpiperidin-3-yl)-4-oxo-3,4-dihydrophthalazin-1-yl)amino)ethyl)-5-(trifluoromethyl)phenyl)acetamide 29e

[0749] Compound 29d (200 mg, crude) was dissolved in methanol (4 mL), and aqueous formaldehyde (127 mg, 1.56 mmol, 37%) and palladium / carbon (50 mg, 10%) were added sequentially. The mixture was stirred at room temperature overnight under a hydrogen atmosphere. TLC indicated the reaction was complete. The reaction mixture was filtered through a pad of Celite, the filter cake was washed with methanol, and the filtrate was concentrated to afford the title compound 29e (200 mg, crude) as a white solid, which was used directly in the next step.

[0750] LC-MS: m / z = 488.3 [M+H] +

[0751] Step 6: 4-(((R)-1-(3-amino-5-(trifluoromethyl)phenyl)ethyl)amino)-6-(1-methylpiperidin-3-yl)phthalazin-1(2H)-one 29

[0752] Compound 29e (200 mg, crude) was dissolved in ethanol (6 mL), and aqueous sodium hydroxide (3 mL, 15 mmol, 5 M) was added at room temperature. The mixture was heated under reflux overnight. The reaction mixture was concentrated, diluted with water, and extracted with ethyl acetate. The combined organic phases were washed with saturated brine, dried over anhydrous sodium sulfate, and concentrated. The crude product was purified by Prep-TLC to afford the title compound 29 (37 mg, 21% yield over four steps) as a yellow solid.

[0753] LC-MS: m / z=446.3[M+H] +

[0754] 1 H NMR (400MHz, DMSO-d6) δ11.43(s,1H),8.21(s,1H),8.13(d,J=8.4Hz,1H),7.73( dd,J=8.4,0.8Hz,1H),6.92(d,J=6.8Hz,1H),6.80(d,J=6.0Hz,2H),6.66(s,1H) ,5.50(s,2H),4.92-4.85(m,1H),3.03-2.85(m,3H),2.25(s,3H),2.13-1.84(m, 3H),1.80-1.76(m,1H),1.71-1.58(m,2H),1.50(d,J=7.2Hz,3H).(98.90% purity by HPLC)

[0755] Example 30

[0756] 4-(((R)-1-(3-amino-5-(trifluoromethyl)phenyl)ethyl)amino)-2-methyl-6-(1-methylpiperidin-3-yl)phthalazin-1(2H)-one 30

[0757]

[0758] Step 1: (R)-5-(2-methyl-4-((1-(3-nitro-5-(trifluoromethyl)phenyl)ethyl)amino)-1-oxo-1,2-dihydrophthalazin-6-yl)-3,6-dihydropyridine-1(2H)-carboxylic acid tert-butyl ester 30a

[0759] Compound 2a (200 mg, 0.42 mmol) and 1-tert-butyloxycarbonyl-3,6-dihydro-2H-pyridine-5-boronic acid pinacol ester 17a (197 mg, 0.64 mmol) were dissolved in 1,4-dioxane (15 mL) and water (3 mL). Pd(dppf)Cl2 dichloromethane complex (18 mg, 0.022 mmol) and sodium carbonate (90 mg, 0.85 mmol) were added at room temperature. The mixture was heated to 100°C under nitrogen and stirred for 1 hour. TLC confirmed the reaction was complete. The reaction mixture was cooled to room temperature, water was added, and extraction with ethyl acetate was performed. The organic phases were combined, washed with saturated brine, dried over anhydrous sodium sulfate, and concentrated. The crude product was purified by silica gel column chromatography to afford the title compound 30a (240 mg, 99% yield).

[0760] LC-MS: m / z = 574.3 [M+H] +

[0761] Step 2 tert-Butyl 3-(4-(((R)-1-(3-amino-5-(trifluoromethyl)phenyl)ethyl)amino)-2-methyl-1-oxo-1,2-dihydrophthalazin-6-yl)piperidine-1-carboxylate 30b

[0762] Compound 30a (240 mg, 0.42 mmol) was dissolved in tetrahydrofuran (10 mL) and ethyl acetate (6 mL). Palladium / carbon (catalytic amount) was added at room temperature. The temperature was raised to 50°C under a hydrogen atmosphere and allowed to react overnight. TLC confirmed the reaction was complete. The reaction solution was cooled to room temperature, filtered through a pad of Celite, and the filtrate was concentrated to afford the title compound 30b (240 mg, crude product), which was used directly in the next step.

[0763] LC-MS: m / z=546.3[M+H] +

[0764] Step 3 tert-Butyl 3-(4-(((R)-1-(3-acetylamino-5-(trifluoromethyl)phenyl)ethyl)amino)-2-methyl-1-oxo-1,2-dihydrophthalazin-6-yl)piperidine-1-carboxylate 30c

[0765] Compound 30b (240 mg, crude) and N,N-diisopropylethylamine (108 mg, 0.84 mmol) were dissolved in 1,4-dioxane (60 mL). Acetic anhydride (64 mg, 0.63 mmol) was added and stirred at room temperature overnight. The reaction was complete by TLC. The reaction mixture was added with water and extracted with dichloromethane. The organic phases were combined, washed with saturated brine, dried over anhydrous sodium sulfate, and concentrated to afford the title compound 30c (270 mg, crude), which was used directly in the next step.

[0766] Step 4: N-(3-((1R)-1-((3-methyl-4-oxo-7-(piperidin-3-yl)-3,4-dihydrophthalazin-1-yl)amino)ethyl)-5-(trifluoromethyl)phenyl)acetamide 30d

[0767] Compound 30c (270 mg, crude) was dissolved in dichloromethane (2 mL), and trifluoroacetic acid (2 mL) was added. The mixture was stirred at room temperature for 2 hours. The reaction was complete by TLC. The reaction solution was concentrated, and the residue was neutralized with saturated aqueous sodium bicarbonate solution. The mixture was extracted with dichloromethane, and the organic phases were combined, washed with saturated brine, dried over anhydrous sodium sulfate, and concentrated to afford the title compound 30d (220 mg, crude), which was used directly in the next step.

[0768] LC-MS: m / z = 488.3 [M+H] +

[0769] Step 5: N-(3-((1R)-1-((3-methyl-7-(1-methylpiperidin-3-yl)-4-oxo-3,4-dihydrophthalazin-1-yl)amino)ethyl)-5-(trifluoromethyl)phenyl)acetamide 30e

[0770] Compound 30d (125 mg, crude) and formaldehyde (62 mg, 0.76 mmol, 37%) were dissolved in methanol (15 mL). Palladium / carbon (catalytic amount) was added and the mixture was allowed to react at room temperature under a hydrogen atmosphere for 2 hours. TLC confirmed the reaction was complete. The reaction mixture was filtered and the filtrate was concentrated to afford the title compound 30e (130 mg, crude), which was used directly in the next step.

[0771] LC-MS: m / z = 502.3 [M+H] +

[0772] Step 6: 4-(((R)-1-(3-amino-5-(trifluoromethyl)phenyl)ethyl)amino)-2-methyl-6-(1-methylpiperidin-3-yl)phthalazin-1(2H)-one 30

[0773] Compound 30e (130 mg, crude) was dissolved in ethanol (6 mL). Sodium hydroxide (103 mg, 2.58 mmol) was added at room temperature. The reaction temperature was raised to 90°C and allowed to react overnight. TLC indicated that the reaction was essentially complete. The reaction mixture was cooled to room temperature, filtered, and the filtrate was concentrated. The crude product was purified by Prep-TLC to afford the title compound 30 (40 mg, 36% yield over five steps).

[0774] LC-MS: m / z = 460.3 [M+H] +

[0775] 1 H NMR (400MHz, CD3OD) δ8.24(d,J=8.4Hz,1H),8.11(d,J=1.2Hz,1H),7.72(dd,J=1.2,8.0 Hz,1H),7.00-6.97(m,2H),6.78(s,1H),5.00(q,J=7.2Hz,1H),3.56(s,3H),3.14-3.03 (m,3H),2.44(s,3H),2.41-2.33(m,1H),2.27-2.19(m,1H),2.02-1.99(m,1H),1.95-1. 89(m,1H),1.88-1.76(m,1H),1.71-1.64(m,1H),1.60(d,J=6.8Hz,3H).(98.98% purity by HPLC)

[0776] Example 31

[0777] 1-(((R)-1-(3-amino-5-(trifluoromethyl)phenyl)ethyl)amino)-7-((R)-1-methylpiperidin-3-yl)pyrido[3,4-d]pyridazin-4(3H)-one 31-1

[0778] 1-(((R)-1-(3-amino-5-(trifluoromethyl)phenyl)ethyl)amino)-7-((S)-1-methylpiperidin-3-yl)pyrido[3,4-d]pyridazin-4(3H)-one 31-2

[0779]

[0780] Step 1: (R)-5-(1-((1-(3-amino-5-(trifluoromethyl)phenyl)ethyl)amino)-4-oxo-3,4-dihydropyrido[3,4-d]pyridazin-7-yl)-3,6-dihydropyridine-1(2H)-carboxylic acid tert-butyl ester 31a

[0781] Compound 17b (260 mg, 0.46 mmol) was dissolved in ethanol (10 mL) and water (3 mL). Reduced iron powder (129 mg, 2.31 mmol) and ammonium chloride (125 mg, 2.31 mmol) were added at room temperature. The mixture was heated to 90°C for 3 hours. TLC indicated complete reaction. The reaction mixture was hot filtered through a pad of Celite. The filtrate was added with water (10 mL) and extracted with ethyl acetate (10 mL). The combined organic phases were washed with saturated brine (10 mL), dried over anhydrous sodium sulfate, and concentrated to afford the title compound 31a (296 mg, crude) as a yellow solid, which was used directly in the next step.

[0782] Step 2: (R)-3-(1-(((R)-1-(3-amino-5-(trifluoromethyl)phenyl)ethyl)amino)-4-oxo-3,4-dihydropyrido[3,4-d]pyridazin-7-yl)piperidine-1-carboxylic acid tert-butyl ester 31b-1 & (S)-3-(1-(((R)-1-(3-amino-5-(trifluoromethyl)phenyl)ethyl)amino)-4-oxo-3,4-dihydropyrido[3,4-d]pyridazin-7-yl)piperidine-1-carboxylic acid tert-butyl ester 31b-2

[0783] Compound 31a (290 mg, crude) was dissolved in ethanol (5 mL), and palladium / carbon (50 mg, 10%) was added. The mixture was reacted at room temperature under a hydrogen atmosphere for 5 hours. TLC indicated complete reaction. The reaction mixture was filtered through a pad of Celite, and the filtrate was concentrated to afford the title compound 31b as a yellow solid (120 mg, 46% yield over two steps). Chiral resolution of compound 31b (Dacel AD-H, 30 x 250 mm, 5 μm, 20 mL / min, EtOH:Hexane = 1:9) afforded the title compound 31b-1 (peak 1, RT 15.577 min) as a pale yellow solid (40 mg, 33% yield) and the title compound 31b-2 (peak 2, RT 19.628 min) as a pale yellow solid (41 mg, 34% yield).

[0784] The configuration and properties of the compounds need further testing, and 31b-1 and 31b-2 are tentatively determined to be the above configurations.

[0785] Step 3: (R)-3-(1-(((R)-1-(3-acetylamino-5-(trifluoromethyl)phenyl)ethyl)amino)-4-oxo-3,4-dihydropyrido[3,4-d]pyridazin-7-yl)piperidine-1-carboxylic acid tert-butyl ester 31c-1 & (S)-3-(1-(((R)-1-(3-acetylamino-5-(trifluoromethyl)phenyl)ethyl)amino)-4-oxo-3,4-dihydropyrido[3,4-d]pyridazin-7-yl)piperidine-1-carboxylic acid tert-butyl ester 31c-2

[0786] Compound 31b-1 (40 mg, 0.075 mmol) was dissolved in tetrahydrofuran (6 mL). N,N-diisopropylethylamine (58 mg, 0.45 mmol) and acetic anhydride (30 mg, 0.30 mmol) were added at room temperature. The mixture was heated to 35°C for 16 hours. TLC indicated complete reaction. The reaction mixture was diluted with water (10 mL) and extracted with ethyl acetate (10 mL). The combined organic phases were washed with saturated brine (10 mL), dried over anhydrous sodium sulfate, and concentrated to afford the title compound 31c-1 (75 mg, crude) as a yellow solid, which was used directly in the next step.

[0787] Compound 31b-2 (41 mg, 0.076 mmol) was dissolved in tetrahydrofuran (6 mL). N,N-diisopropylethylamine (59 mg, 0.46 mmol) and acetic anhydride (31 mg, 0.31 mmol) were added at room temperature. The mixture was heated to 35°C for 16 hours. TLC indicated complete reaction. The reaction mixture was diluted with water (10 mL) and extracted with ethyl acetate (10 mL). The combined organic phases were washed with saturated brine (10 mL), dried over anhydrous sodium sulfate, and concentrated to afford the title compound 31c-2 (75 mg, crude) as a yellow solid, which was used directly in the next step.

[0788] LC-MS: m / z = 575.3 [M+H] +

[0789] The configuration and properties of the compounds need further testing, and 31c-1 and 31c-2 are tentatively determined to be the above configurations.

[0790] Step 4: N-(3-((R)-1-((4-oxo-7-((R)-piperidin-3-yl)-3,4-dihydropyrido[3,4-d]pyridazin-1-yl)amino)ethyl)-5-(trifluoromethyl)phenyl)acetamide 31d-1 & N-(3-((R)-1-((4-oxo-7-((S)-piperidin-3-yl)-3,4-dihydropyrido[3,4-d]pyridazin-1-yl)amino)ethyl)-5-(trifluoromethyl)phenyl)acetamide 31d-2

[0791] Compound 31c-1 (75 mg, crude) was dissolved in dichloromethane (2 mL) and trifluoroacetic acid (2 mL) was added. The mixture was allowed to react at room temperature for 2 hours. TLC indicated complete reaction. The reaction mixture was dropwise added with saturated aqueous sodium bicarbonate (15 mL) and adjusted to pH 8-9. The mixture was extracted with ethyl acetate (10 mL). The combined organic phases were washed with saturated brine (10 mL), dried over anhydrous sodium sulfate, and concentrated to afford the title compound 31d-1 (67 mg, crude) as a yellow solid, which was used directly in the next step.

[0792] Compound 31c-2 (75 mg, crude) was dissolved in dichloromethane (2 mL) and trifluoroacetic acid (2 mL) was added. The mixture was allowed to react at room temperature for 2 hours. TLC indicated complete reaction. The reaction mixture was dropwise added with saturated aqueous sodium bicarbonate (15 mL) and adjusted to pH 8-9. The mixture was extracted with ethyl acetate (10 mL). The organic phases were combined, washed with saturated brine (10 mL), dried over anhydrous sodium sulfate, and concentrated to afford the title compound 31d-2 (50 mg, crude) as a yellow solid, which was used directly in the next step.

[0793] The configuration and properties of the compounds need further testing, and 31d-1 and 31d-2 are tentatively determined to be the above configurations.

[0794] Step 5: N-(3-((R)-1-((7-((R)-1-methylpiperidin-3-yl)-4-oxo-3,4-dihydropyrido[3,4-d]pyridazin-1-yl)amino)ethyl)-5-(trifluoromethyl)phenyl)acetamide 31e-1 & N-(3-((R)-1-((7-((S)-1-methylpiperidin-3-yl)-4-oxo-3,4-dihydropyrido[3,4-d]pyridazin-1-yl)amino)ethyl)-5-(trifluoromethyl)phenyl)acetamide 31e-2

[0795] Compound 31d-1 (67 mg, crude) was dissolved in ethanol (3 mL), and aqueous formaldehyde (35 mg, 0.43 mmol, 30%) and palladium / carbon (20 mg, 10%) were added. The mixture was reacted at room temperature under a hydrogen atmosphere for 4 hours. TLC indicated complete reaction. The reaction mixture was filtered through a pad of Celite, and the filtrate was concentrated to afford the title compound 31e-1 (70 mg, crude) as a yellow solid, which was used directly in the next step.

[0796] Compound 31d-2 (50 mg, crude) was dissolved in ethanol (3 mL), and aqueous formaldehyde (35 mg, 0.43 mmol, 30%) and palladium / carbon (20 mg, 10%) were added. The mixture was reacted at room temperature under a hydrogen atmosphere for 4 hours. TLC indicated complete reaction. The reaction mixture was filtered through a pad of Celite, and the filtrate was concentrated to afford the title compound 31e-2 (60 mg, crude) as a yellow solid, which was used directly in the next step.

[0797] The configuration and properties of the compounds need further testing, and 31e-1 and 31e-2 are tentatively determined to be the above configurations.

[0798] Step 6 1-(((R)-1-(3-amino-5-(trifluoromethyl)phenyl)ethyl)amino)-7-((R)-1-methylpiperidin-3-yl)pyrido[3,4-d]pyridazin-4(3H)-one 31-1 & 1-(((R)-1-(3-amino-5-(trifluoromethyl)phenyl)ethyl)amino)-7-((S)-1-methylpiperidin-3-yl)pyrido[3,4-d]pyridazin-4(3H)-one 31-2

[0799] Compound 31e-1 (70 mg, crude) was dissolved in methanol (3 mL). Aqueous sodium hydroxide (2 mL, 8 mmol, 4 N) was added at room temperature and heated to 70°C for 16 hours. TLC indicated complete reaction. The reaction mixture was cooled to room temperature, diluted with water (10 mL), and extracted with ethyl acetate (10 mL). The combined organic phases were washed with saturated brine (10 mL), dried over anhydrous sodium sulfate, and concentrated. The crude product was purified by Prep-TLC (dichloromethane / methanolic ammonia solution = 10 / 1) to afford the title compound 31-1 (15 mg, 45% yield over four steps) as a pale yellow solid.

[0800] LC-MS: m / z = 447.2 [M+H] +

[0801] 1H NMR (400MHz, CD3OD) δ9.43(s,1H),8.13(s,1H),6.96-6.89(m,2H),6.79-6.75(m,1H),4.96(q,J=6.8H z,1H),3.49-3.39(m,2H),3.28-3.20(m,1H),3.18-2.99(m,1H),2.83-2.62(m,4H),2.24-2.12(m,1H), 2.03-1.82(m,3H),1.58(d,J=6.8Hz,3H).(96.67% purity by HPLC)

[0802] Compound 31e-2 (60 mg, crude) was dissolved in methanol (3 mL). Aqueous sodium hydroxide (2 mL, 8 mmol, 4 N) was added at room temperature and heated to 70°C for 16 hours. TLC indicated complete reaction. The reaction mixture was cooled to room temperature, diluted with water (10 mL), and extracted with ethyl acetate (10 mL). The combined organic phases were washed with saturated brine (10 mL), dried over anhydrous sodium sulfate, and concentrated. The crude product was purified by Prep-TLC (dichloromethane / methanolic ammonia solution = 10 / 1) to afford the title compound 31-2 (13 mg, 37% yield over four steps) as a pale yellow solid.

[0803] LC-MS: m / z = 447.2 [M+H] +

[0804] 1 H NMR (400MHz, CD3OD) δ9.41(s,1H),8.09(s,1H),6.98-6.88(m,2H),6.81-6.73(m,1H),4.95(q,J=6.8Hz,1H),3.30-3.20(m,2H),3.13-3.0 1(m,1H),2.75-2.58(m,1H),2.51(s,3H),2.44-2.28(m,1H),2.16-2.05(m,1H),1.97-1.71(m,3H),1.58(d,J=6.8Hz,3H).(99.69% purity by HPLC)

[0805] The configuration and properties of the compounds need further testing, and 31-1 and 31-2 are tentatively determined to be the above configurations.

[0806] IC of the compound in Test Example 1 for inhibition of K-562 cell proliferation 50 Determination

[0807] The human chronic myeloid leukemia K-562 (CCL-243) cells used in the present invention were purchased from American Type Culture Collection (ATCC) and grown in RPMI 1640 medium containing 10% fetal bovine serum (FBS) and 1% double-antibody at 37° C. and 5% CO 2 .

[0808] The inhibitory effect of the compound on the proliferation of K-562 cells cultured in vitro was determined by the following method:

[0809] 1) Cell seeding: K-562 cells in good logarithmic growth phase were seeded into a 96-well plate at 20,000 cells / well in 90 μL, and cultured at 37° C. and 5% CO 2 for 24 hours.

[0810] 2) Dosing: Serially dilute the compound to be tested in complete culture medium. Add 10 μL of the diluted compound to 90 μL of cells to achieve final concentrations of 10,000, 3,000, 1,000, 300, 100, 30, 10, 3, and 1 nM. Also include corresponding vehicle controls. Incubate in a 37°C, 5% CO2 cell culture incubator for 96 hours.

[0811] 3) Detection: Add 10 μL of 5 mg / mL MTT working solution (ABCONE, M9609) to each well. Incubate at 37°C for 4 hours. Then, add a triplex solution (10% SDS, 0.5% isopropanol, 0.1 mol / L HCl) until the cell lysate is completely dissolved. OD570 and OD690 values ​​are read using a TECAN SPARK microplate reader.

[0812] 4) Calculation: Calculate the cell growth inhibition rate using the following formula:

[0813] Inhibition rate = (control well OD570nm-OD690nm - Dosage hole OD570nm-OD690nm ) / control well OD570nm-OD690nm ×100%

[0814] Graphpad Prism 5.0 software was used to calculate the IC according to the compound concentration and the corresponding inhibition rate. 50 The test results are shown in Table 1.

[0815] Table 1 IC values ​​of the compounds of the present invention for inhibition of K-562 cell proliferation 50 (nM)

[0816] Compound number <![CDATA[IC 50 (nM)]]> Compound number <![CDATA[IC 50 (nM)]]> Compound number <![CDATA[IC 50 (nM)]]> BI3406 35.2 11 1619 22 107.7 1 735.7 12 134 23 142.8 2 822.7 13 104 24 231.9 3 111.1 14 362.3 25 125.9 4 150.8 15 279.25 26 137.6 5 44.7 16 43.8 27 91.3 6 2489 17 33.1 28 66.65 7 5917 18 60.0 29 51.0 8 1599 19 63.4 30 143.9 9 988 20 39.3 31-1 43.4 10 121.4 21 97.7 31-2 13.1

[0817] Conclusion: As shown in Table 1, the compounds of the present invention have an inhibitory effect on the proliferation of K-562 cells, and the activities of many compounds are comparable to that of BI-3402.

[0818] Effect of Test Example 2 Compounds on the Phosphorylation Level of KRAS Downstream Signaling Molecule ERK1 / 2 in K-562 Cells

[0819] The effects of the compounds of the present invention on the phosphorylation level of ERK1 / 2 in K-562 cells were detected by the following method:

[0820] 1) Cell inoculation: K-562 cells in good logarithmic growth phase were seeded with 1*10 6 Each well was inoculated into a six-well plate and cultured overnight at 37°C and 5% CO2.

[0821] 2) Drug addition: The compound to be tested was serially diluted in complete culture medium and added to the cells to achieve final concentrations of 1000, 100, 10, and 1 nM. The cells were cultured in a 37°C, 5% CO2 cell culture incubator for 24 hours.

[0822] 3) Protein Sample Preparation: Cell suspension was collected and centrifuged at 500 g for 5 minutes. The supernatant was discarded and the cells were washed three times with PBS. The cells were lysed with 100 μL of 1× SDS gel loading buffer (50 mM Tris-HCl (pH 6.8), 100 mM DTT, 2% SDS, 10% glycerol, 0.1% bromophenol blue). The cell lysate was denatured by heating at 100°C for 10 minutes.

[0823] 4) Western blot: The protein samples were subjected to SDS-PAGE electrophoresis. After electrophoresis, the proteins were transferred to a PVDF membrane using a wet transfer system. The PVDF membrane was blocked in blocking solution (5% skim milk powder diluted in TBS / T) at room temperature for 1 hour, followed by I and II antibody reactions. After washing, the membrane was stained with Immobilon Western HRP Substrate luminal reagent and photographed using a Western Blot imager (Tanon, 4600). The following is the information of the antibodies used: p-ERK1 / 2 (CST:4370); ERK1 / 2 (CST:9102); β-tubulin (CST:2146); GAPDH (CST:5174). The results of the effects of the compounds on the phosphorylation level of ERK1 / 2 in K-562 cells are shown in Figure 4. Figure 1 and Figure 2 .

[0824] Conclusion: Compounds 5, 20, 31-1, and 31-2 in the examples of the present invention have a significant inhibitory effect on the phosphorylation of ERK1 / 2 in K-562 cells; the inhibitory activity is concentration-dependent; overall, the activity is roughly equivalent to that of BI-3406.

[0825] Inhibitory activity of the compound of Test Example 3 against SOS1

[0826] Experimental steps:

[0827] 1. Compound Treatment: Prepare the compound at a 400-fold final concentration. For example, if the final concentration is 5 μM, prepare it at a 400-fold concentration, i.e., 2 mM. Use an automated microwell pipette to dilute the compound to the desired concentration points.

[0828] 2. Transfer compounds to a 384-well reaction plate: Use an ultrasonic nanoliter liquid handling system to transfer 50 nL of the diluted compounds from the Echo 384-well plate to a 384-well reaction plate. Transfer 50 nL of 100% DMSO to both the negative and positive controls.

[0829] 3. Prepare and transfer a 4x Tag1-SOS1 solution: Use the Diluent provided in the kit (KRAS-G12C / SOS1BINGDING ASSAYKIT (Cisbio, Cat. No. 63ADK000CB16PEG)) to prepare a 4x Tag1-SOS1 solution, transfer 5 ul to a 384-well reaction plate, transfer 5 ul of Diluent to the negative control wells instead of the enzyme solution, and centrifuge at 1000 rpm for 1 minute.

[0830] 4. Prepare 4x Tag2-KRAS G12C solution: Use Diluent provided in the kit to prepare 4x Tag2-KRASG12C solution, transfer 5ul to a 384-well reaction plate, and centrifuge at 1000rpm for 1 minute.

[0831] 5. Transfer 2x detection solution: Prepare 2x Anti-Tag1-Tb3+ and Anti-Tag2-XL665 solutions using the Detection Buffer provided in the kit. Transfer 10 μl to a 384-well reaction plate. Centrifuge at 1000 rpm for 1 minute and incubate at room temperature for 60 minutes.

[0832] 6. Reading: Use Envision microplate reader to read the fluorescence signal value (Ex665 / Em615).

[0833] 7. Inhibition rate calculation and IC50 fitting

[0834] The values ​​were copied from the plate reader, where the maximum value refers to the reading of the positive control and the minimum value refers to the reading of the negative control. Inhibition rate (%) = (maximum value - sample value) / (maximum value - minimum value) × 100%.

[0835] The data were imported into MS Excel and IC50 values ​​were fitted using XLFit excel add-in version 5.4.0.8;

[0836] Table 2 Inhibitory activity of compounds against SOS1

[0837] Compound number <![CDATA[IC 50 (nM)]]> Compound number <![CDATA[IC 50 (nM)]]> Compound number <![CDATA[IC 50 (nM)]]> 3 15 4 13 5 7.6 10 16 12 9.7 13 7.8 14 18 15 14 16 8.0 17 7.7 18 12 19 6.8 20 6.0 21 20 22 29 23 5.4 24 18 25 11 26 30 28 12 29 14 30 23

[0838] Test Example 4 Compound Stability Test on Mouse and Human Liver Microsomes

[0839] Experimental steps:

[0840] (1) Take out liver microsomes (20 mg protein / mL) from the -80℃ freezer, place them in a 37℃ water bath thermostat for pre-incubation for 3 minutes, and thaw them for use.

[0841] (2) Prepare the mixed solution of the incubation system (excluding β-NADPH) according to the proportions of the above “Composition of the experimental incubation system”.

[0842] (3) Prepare 100 μM working solution of the test compound and set aside.

[0843] (4) Control group (without β-NADPH): 25 μL of PB solution was added to 75 μL of the incubation system mixture described in (2), and the mixture was vortexed for 30 seconds to mix well. The total reaction volume was 100 μL, and duplicate samples were prepared. The mixture was placed in a 37°C water bath in a constant temperature oscillator for incubation, and the timing was started. The sampling time points were 0 min and 60 min.

[0844] (5) Sample group: Take 25 μL of β-NADPH solution (4 mM) and add it to 75 μL of the reaction system described in (2). Vortex for 30 seconds to mix. The total reaction volume is 100 μL. Repeat the sample. Place it in a 37℃ water bath constant temperature oscillator for incubation and start the timer. The sampling time points are 0 min, 5 min, 15 min, 30 min, and 60 min.

[0845] (6) At each time point, remove the sample tube and add 300 μL of cold terminator (including internal standard) to terminate the reaction.

[0846] (7) Vortex and centrifuge.

[0847] (8) Take 150 μL of the supernatant and add 150 μL of water, vortex mix, and inject for LC-MS / MS analysis.

[0848] Data analysis: The following first-order kinetic formula was used to calculate the half-life (t 1 / 2 ) and clearance (CL)

[0849] C t =C0*e-kt

[0850] C t =(1 / 2)*C0

[0851] t 1 / 2 =ln2 / k=0.693 / k

[0852] CL=V d *k

[0853] Vd=1 / protein content in liver microsomes

[0854] CL int(liver) =CL int(mic) × liver weight to body weight ratio × liver microsomal protein concentration per gram of liver

[0855] The parameters in the formula are shown in Table 3:

[0856] Table 3 Common parameters of liver and blood of mice, rats and humans

[0857]

[0858] The experimental results are shown in Table 4:

[0859] Table 4 Stability of compounds in liver microsomes of different species

[0860]

[0861] In the human liver microsome stability experiment, it can be seen that the human microsome stability of multiple compounds is comparable to that of BI-3406, and the mouse liver microsome stability is superior to that of BI-3406, which is more conducive to drug research.

[0862] Pharmacokinetic properties of the compound in test example 5 in mice

[0863] The compound of Example 20 (10 mg / kg) was orally administered to ICR mice (female, n=3) that had fasted overnight. Blood samples were collected before administration and at 0.25 hours, 0.5 hours, 1 hour, 2 hours, 4 hours, and 8 hours after administration, and the serum was obtained by centrifugation (4500 rpm) for 10 min at 4°C. 100 μL MeOH / ACN (1:1, v / v) was added to 10 μL serum to precipitate the mixture, which was then vortexed for 1 min and centrifuged (11000 rpm) for 5 min to obtain the supernatant. 20 μL of the supernatant was dissolved in 20 μL ACN / H2O (1:1, v / v) and analyzed by ultra-performance liquid chromatography. The results are shown in Table 5:

[0864] Table 5 Pharmacokinetic characteristics of compound 20 in mice

[0865]

[0866] As shown in Table 5, compound 20 has a good plasma exposure in mice.

[0867] The applicant states that the present invention uses the above-mentioned embodiments to illustrate the polycyclic pyridazinone derivatives of the present invention as SOS1 inhibitors, their preparation methods, and applications. However, the present invention is not limited to the above-mentioned embodiments, and it does not necessarily rely on the above-mentioned embodiments for implementation. Those skilled in the art should understand that any improvements to the present invention, equivalent substitutions for the raw materials of the present invention, addition of auxiliary ingredients, and selection of specific methods, etc., fall within the scope of protection and disclosure of the present invention.

[0868] The preferred embodiments of the present invention are described in detail above. However, the present invention is not limited to the specific details in the above embodiments. Within the technical concept of the present invention, various simple modifications can be made to the technical solution of the present invention, and these simple modifications all fall within the scope of protection of the present invention.

[0869] It should also be noted that the various specific technical features described in the above specific embodiments can be combined in any appropriate manner without contradiction. In order to avoid unnecessary repetition, the present invention will not further describe various possible combinations.

Claims

1. A polycyclic pyridazinone derivative or a pharmaceutically acceptable salt thereof, wherein the polycyclic pyridazinone derivative is selected from any one of the following structures:

2. A pharmaceutical composition, characterized in that The pharmaceutical composition comprises the polycyclic pyridazinone derivative according to claim 1, or a pharmaceutically acceptable salt thereof.

3. The composition according to claim 2, further comprising a pharmaceutically acceptable carrier and / or excipient.

4. Use of the polycyclic pyridazinone derivative according to claim 1, or a pharmaceutically acceptable salt thereof, or the pharmaceutical composition according to claim 2 in the preparation of a medicament for treating cancer or in the preparation of an SOS1 inhibitor.

5. The use according to claim 4, wherein the cancer is pancreatic cancer, colorectal cancer, lung cancer, hepatocellular carcinoma, kidney cancer, gastric cancer or bile duct cancer.

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