C-N coupling method using [Cu2 (mu-salbene) 2] complex

Abstract

The present invention relates to a method for coupling aryl halides with N-nucleophilic reagent compounds, characterized in that the coupling is carried out in the presence of a binuclear copper (II) complex with a substituted salbed-type ligand of formula (I). Also described are novel binuclear copper (II) complexes with a substituted salbed-type ligand within the scope of Formula (I).

Description

[0001] This invention relates to a novel method for coupling aryl halides with N-nucleophilic compounds in the presence of a dinuclear copper(II) complex of formula I with a substituted salben-type ligand.

[0002]

[0003] Where R 1 and R 2 For the same or different, and for C that is optionally substituted 1-12 -alkyl, aryl or heteroaryl, or hydrogen, and

[0004] G 1 G 2 G 3 and G 4 For the same or different, and representing hydrogen or selected from C 1-12 -alkyl, C 1-12 -alkoxy group, halogen C 1-12 -alkyl, halogen, mono- or di-C 1-12 -alkylamino, carboxyl, C 1-12 One or more substituents of alkoxycarbonyl or nitro; or G 1 G 2 G 3 and G 4 Each ring independently forms a fused aryl ring with the phenyl ring to which it is attached, having two or three rings, wherein the two or three rings are optionally selected from C 1-6 -alkyl, C 1-6 -alkoxy, halogen, mono- or di-C 1-6 -One or more substituents of alkylamino or nitro groups.

[0005] The present invention also relates to novel binuclear copper(II) complexes of formula I with substituted salben-type ligands, wherein

[0006] R 1 and R 2 For the same, and for substituents of formula IIa, IIb or IIc

[0007]

[0008] in

[0009] R 3 R 4 R 5 R 6 and R 7 For the same or different, and for hydrogen, C1-12 -alkyl, C 1-12 -alkoxy groups, halogens, halogen-C 1-12 -alkyl, mono- or di-C 1-12 -alkylamino, nitro, or R 3 and R 4 、or R 4 and R 5 、or R 5 and R 6 、or R 6 and R 7 Together with the phenyl rings to which they are attached, they form a fused aryl ring having two or three rings, which optionally have been selected from C 1-6 -alkyl, C 1-6 -alkoxy, halogen, mono- or di-C 1-6 -One or more substituents of alkylamino or nitro groups.

[0010] X 1 X 2 X 3 X 4 and X 5 They may be the same or different, and represent at least one heteroatom selected from nitrogen, oxygen or sulfur;

[0011] n is 0 or 1;

[0012] R 8 R 9 and R 10 For the same or different, and for hydrogen, C 1-12 -alkyl, halogen C 1-12 -alkyl, hydroxyl, hydroxy-C 1-12 -alkyl, halogen, or R 8 and R 9 Together they form saturated carbon rings or heterocycles, or R 8 R 9 and R 10 Together they form unsaturated carbon rings.

[0013] The condition is to exclude R. 1 and R 2 It is hydrogen, or R 1 and R 2 Having equation IIa and where R 3 R 4 R 6 and R 7 For hydrogen and R at the same time 5 Compounds that are hydrogen, chlorine, methyl, or nitro groups.

[0014] The object of this invention is to provide an alternative method for CN-coupling, namely, to couple aryl halides to N-nucleophiles using a binuclear copper(II) complex of Formula I with a substituted salben-type ligand, and to provide a novel binuclear copper(II) complex of Formula I with a substituted salben-type ligand capable of catalyzing such coupling.

[0015] This objective can be achieved using the method outlined below, which involves coupling an aryl halide with an N-nucleophile compound in the presence of a binuclear copper(II) complex of formula I with a substituted salben-type ligand.

[0016] The following definitions are set forth to illustrate and define the meaning and scope of various terms used to describe the present invention.

[0017] Term "C" 1-12 "-alkyl" refers to a branched or straight-chain monovalent saturated aliphatic hydrocarbon group with one to twelve carbon atoms, preferably one to eight, and more preferably one to four carbon atoms. The term is further illustrated by examples of groups such as methyl, ethyl, n-propyl, isopropyl, n-butyl, isobutyl, sec-butyl, or tert-butyl, pentyl, or hexyl, heptyl, octyl, nonyl, decyl, undecyl, and dodecyl and their isomers.

[0018] Term "C" 1-12 "-alkoxy" refers to a C atom bonded to an oxygen atom as defined above. 1-12 -alkyl. Typical examples are methoxy or ethoxy.

[0019] The term "single- or bi-C" 1-12 "-alkylamino" refers to C as defined above. 1-12 - An amino group that is mono- or di-substituted with an alkyl group. Typical examples are N-methylamino, N-ethylamino, N,N-dimethylamino, or N,N-diethylamino.

[0020] The term "halogen" refers to fluorine, chlorine, bromine, or iodine, preferably fluorine and chlorine, but specifically refers to bromine and iodine in relation to CN coupling.

[0021] The term "halogen-C" 1-12 "-alkyl" refers to a C14 alkyl group substituted with one or more halogens as defined above. 1-12 -alkyl groups, such as trichloromethyl, 1,1-dichloroethyl or 1,1-dichlorooctyl.

[0022] The term "aryl" refers to an aromatic carbide ring, such as a phenyl, naphthyl, anthracene, or phenanthrene ring, preferably a phenyl ring.

[0023] The term "heteroaryl" refers to an aromatic 5- to 6-membered monocyclic or 9- to 10-membered bicyclic ring that may contain 1, 2, or 3 heteroatoms selected from nitrogen, oxygen, and / or sulfur, such as pyridyl, pyridinyl, pyrazolyl, pyrimidinyl, benzimidazolyl, quinolinyl and isoquinolinyl, thiophene, furanyl, pyrroleyl, isoxazolyl, oxazolyl, thiazolyl, or imidazolyl.

[0024] The term "optionally replaced" refers to a selection from C 1-12 -alkyl, C 1-12 -alkoxy groups, halogens, halogen C 1-12 -alkyl, mono- or di-C 1-12 -One or more substituents of alkylamino or nitro groups.

[0025] Spiral key

[0026] “ "

[0027] represent" "or" ", and thus indicates the chirality of the molecule, but also represents a mixture of enantiomers.

[0028] When a chiral carbon is present in a chemical structure, it means that all stereoisomers associated with that chiral carbon are included in the structure as pure stereoisomers and mixtures thereof.

[0029] method:

[0030] As summarized above, the present invention relates to a novel method for coupling aryl halides with N-nucleophilic compounds in the presence of a binuclear copper(II) complex of formula I with a substituted salben-type ligand.

[0031] Aryl halides:

[0032] Suitable aryl halides can be defined using Formula III.

[0033]

[0034] in

[0035] X 6 X 7 X 8 X 9 and X 10 They may be the same or different, and represent carbon or at least one heteroatom selected from nitrogen, oxygen or sulfur;

[0036] R 19 R 20 R21 R 22 and R 23 For the same or different, and for hydrogen, C 1-12 -alkyl, C 2-12 -Alkenyl, C 2-12 -Alynyl group, C 1-12 -alkoxy groups, halogens, halogen-C 1-12 -alkyl, mono- or di-C 1-12 -alkylamino or cyano; or R 19 and R 20 ,or R 20 and R 21 、or R 21 and R 22 、or R 22 and R 23 Together with the rings they are attached to, they form a fused aryl ring having two or three rings, which optionally are selected from C 1-12 -alkyl, C 1-12 -alkoxy, halogen, mono- or di-C 1-6 -One or more substituents of alkylamino or nitro groups are used;

[0037] n is 0 or 1, and

[0038] Z represents a halogen atom.

[0039] In preferred aryl halides

[0040] X 6 X 7 X 8 X 9 and X 10 They can be the same or different, and represent either carbon or a nitrogen atom;

[0041] R 19 R 20 R 21 R 22 and R 23 They can be the same or different, and can be hydrogen, methyl, methoxy or cyano;

[0042] n is 1, and

[0043] Z is chlorine, bromine, or iodine, preferably bromine or iodine.

[0044] The aryl halides of Formula III are generally commercially available or can be synthesized according to procedures known to those skilled in the art.

[0045] N-nucleophile:

[0046] Suitable N-nucleophiles can be selected from compounds of the following formula.

[0047]

[0048] in

[0049] R 11 R 12 R 13 R 14 R 15 R 16 R 17 and R 18 For the same or different, and for hydrogen, C 1-12 -alkyl, C 1-12 -alkoxy, aryl, halogen, halogen-C 1-12 -alkyl, nitro, hydroxyl, mono- or di-C 1-12 -alkylamino; or R 15 and R 16 Together with the nitrogen atoms they are attached to, they form saturated 5- or 6-membered heterocycles;

[0050] X 11 X 12 X 13 and X 14 For the same or different, and indicating optional C 1-12 -alkyl, aryl-C 1-12 -alkyl, C 1-12 -alkoxy groups, halogens, halogen-C 1-12 - An alkyl-substituted carbon atom and at least one other heteroatom selected from nitrogen, oxygen or sulfur.

[0051] Preferred N-nucleophiles have the formulas IVc, IVd, and IVe, wherein

[0052] R 15 R 16 R 17 and R 18 For the same or different, and for hydrogen, C 1-6 -alkyl, aryl; or R 15 and R 16 Together with the nitrogen atoms they are attached to, they form saturated heterocyclic 5- or 6-membered heterocycles;

[0053] X 11 X 12 X 13 and X 14 For the same or different, and indicating optional C 1-12 -alkyl, aryl-C1-12 - An alkyl-substituted carbon atom, and at least one other heteroatom selected from nitrogen, oxygen or sulfur.

[0054] catalyst:

[0055] The method of the present invention is carried out in the presence of a binuclear copper(II) complex having a substituted salben-type ligand of Formula I.

[0056]

[0057] Where R 1 and R 2 For the same or different, and for C that is optionally substituted 1-12 -alkyl, aryl or heteroaryl, or hydrogen, and

[0058] G 1 G 2 G 3 and G 4 For the same or different, and representing hydrogen or selected from C 1-12 -alkyl, C 1-12 -alkoxy groups, halogens, halogen C 1-12 -alkyl, mono- or di-C 1-12 -alkylamino, carboxyl, C 1-12 -One or more substituents of alkoxycarbonyl or nitro; or G 1 G 2 G 3 and G 4 Each ring independently forms a fused aryl ring with the phenyl ring to which it is attached, having two or three rings, wherein the two or three rings are optionally selected from C 1-6 -alkyl, C 1-6 -alkoxy, halogen, mono- or di-C 1-6 -One or more substituents of alkylamino or nitro groups.

[0059] In a preferred embodiment, G 1 G 2 G 3 and G 4 For the same or different, and representing hydrogen or one or more substituents selected from methyl, ethyl, tert-butyl, methoxy, ethoxy, fluorine, chlorine, bromine, trifluoromethyl, N / N-dimethylamino, N / N-diethylamino, carboxyl, methoxycarbonyl, ethoxycarbonyl, isopropoxycarbonyl, n-butoxycarbonyl, tert-butoxycarbonyl, nitro; or G 1 G 2 G 3 and G 4They independently form fused aryl rings together with the phenyl rings to which they are attached.

[0060] In a preferred embodiment, R 1 and R 2 For the same, and for substituents of formula IIa, IIb or IIc

[0061]

[0062] in

[0063] R 3 R 4 R 5 R 6 and R 7 For the same or different, and for hydrogen, C 1-12 -alkyl, C 1-12 -alkoxy groups, halogens, halogen-C 1-12 -alkyl, mono- or di-C 1-12 -alkylamino, nitro, or R 3 and R 4 ,or R 4 and R 5 、or R 5 and R 6 、or R 6 and R 7 Together with the phenyl rings to which they are attached, they form a fused aryl ring having two or three rings, which optionally have been selected from C 1-6 -alkyl, C 1-6 -alkoxy, halogen, mono- or di-C 1-6 -One or more substituents of alkylamino or nitro groups are used for substitution.

[0064] X 1 X 2 X 3 X 4 and X 5 They may be the same or different, and represent at least one heteroatom selected from nitrogen, oxygen or sulfur;

[0065] n is 0 or 1;

[0066] R 8 R 9 and R 10 For the same or different, and for hydrogen, C 1-12 -alkyl, halogen C 1-12 -alkyl, hydroxyl, hydroxy-C 1-12 -alkyl, halogen; or R 8and R 9 Together they form saturated carbon rings or heterocycles; or R 8 R 9 and R 10 Together they form an unsaturated carbon ring.

[0067] The preferred substituents of formula IIa are those substituents, wherein R 3 R 4 R 5 R 6 and R 7 For the same or different, and for hydrogen, methyl, ethyl, tert-butyl, methoxy, ethoxy, chlorine, bromine, trifluoromethyl, N / A-dimethylamino, N / A-diethylamino, nitro; or R 3 and R 4 ,or R 4 and R 5 、or R 5 and R 6 、or R 6 and R 7 Together with the phenyl rings they are attached to, they form fused aryl rings selected from 2-naphthalene, anthracene, or phenanthrene.

[0068] Specifically, instances of formula IIa

[0069] ●R 3 To R 7 It is hydrogen, or

[0070] ●R 3 It is methyl, methoxy, chloro, or nitro, and R 4 To R 7 It is hydrogen, or

[0071] ●R 5 It is methyl, tert-butyl, methoxy, chloro, nitro, or N,N-dimethylamino, and R 3 and R 4 and R 6 and R 7 It is hydrogen, or

[0072] ●R 3 and R 5 It is methoxy or chlorine, and R 4 R 6 and R 7 It is hydrogen, or

[0073] ●R 3 and R 7It is methoxy or chlorine, and R 4 To R 6 It is hydrogen, or

[0074] ●R 3 R 5 and R 7 It is methoxy, and R 4 To R 6 It is hydrogen.

[0075] The preferred substituents of formula IIb can be selected from...

[0076]

[0077] Particularly preferred substituents have the formulas IIb1, IIb2, IIb14, and IIb17.

[0078] The preferred substituents of formula IIc are those substituents, wherein R 8 R 9 and R 10 Whether they are the same or different, and whether they are hydrogen, hydroxyl, or hydroxyl-C 1-4 -alkyl; or R 8 and R 9 Together they form a heterocyclic ring; or R 8 R 9 and R 10 Together they form an unsaturated carbon ring.

[0079] Specific examples of substituents in formula IIc are:

[0080]

[0081] Preferably, they are selected from formulas IIc1 and IIc2.

[0082] Substituents IIa and IIc are the most preferred.

[0083] The catalysts of particularly preferred formula IIa are those catalysts, wherein,

[0084] R 3 R 4 R 5 R 6 and R 7 It is hydrogen; or R 3 It is chlorinated or methoxy and R 4 R 5 R 6 and R 7 It is hydrogen; or R 5It is chlorinated or methoxy and R 3 R 4 R 6 and R 7 It is hydrogen; or R 3 and R 7 It is methoxy and R 4 R 5 and R 6 It is hydrogen.

[0085] Particularly preferred catalysts of formula IIc are those having formula IIc1.

[0086] The preferred catalysts are listed below.

[0087]

[0088] A binuclear copper(II) complex of Formula I with a substituted salben-type ligand

[0089]

[0090] in

[0091] G 1 G 2 G 3 and G 4 For the same or different, and representing hydrogen or selected from C 1-12 -alkyl, C 1126 -alkoxy groups, halogens, halogen C 1-12 -alkyl, mono- or di-C 1-12 -alkylamino, carboxyl, C 1-12 -One or more substituents of alkoxycarbonyl or nitro; or G 1 G 2 G 3 and G 4 Each ring independently forms a fused aryl ring with the phenyl ring to which it is attached, having two or three rings, wherein the two or three rings are optionally selected from C 1-6 -alkyl, C 1-6 -alkoxy, halogen, mono- or di-C 1-6 -One or more substituents of alkylamino or nitro groups are used;

[0092] R 1 and R 2 For the same, and for substituents of formula IIa, IIb or IIc

[0093]

[0094] R 3 R 4 R 5 R 6 and R 7 For the same or different, and for hydrogen, C 1-12 -alkyl, C 1-12 -alkoxy groups, halogens, halogen-C 1-12 -alkyl, mono- or di-C 1-12 -alkylamino, nitro, or R 3 and R 4 、or R 4 and R 5 、or R 5 and R 6 、or R 6 and R 7 Together with the phenyl rings to which they are attached, they form a fused aryl ring having two or three rings, which optionally have been selected from C 1-6 -alkyl, C 1-6 -alkoxy, halogen, mono- or di-C 1-6 -One or more substituents of alkylamino or nitro groups.

[0095] X 1 X 2 X 3 X 4 and X 5 They may be the same or different, and represent at least one heteroatom selected from nitrogen, oxygen or sulfur;

[0096] n is 0 or 1;

[0097] R 8 R 9 and R 10 For the same or different, and for hydrogen, C 1-12 -alkyl, halogen C 1-12 -alkyl, hydroxyl, hydroxy-C 1-12 -alkyl, halogen, or R 8 and R 9 Together they form saturated carbon rings or heterocycles, or R 8 R 9 and R 10 Together they form unsaturated carbon rings.

[0098] The condition is that R is not included. 1 and R 2 It is hydrogen, or R 1 and R 2Having equation IIa and where R 3 R 4 R 6 and R 7 For hydrogen and R at the same time 5 Compounds that are hydrogen, chlorine, methyl, or nitro groups.

[0099] The excluded compounds have been disclosed as follows:

[0100] a) Alessandro Pasini, Francesco Demartin, Olivo Piovesana, BrunettoChiari, Antonio Cinti and Ornella Crispu, J. Chem. Soc., Dalton Trans. 2000,3467–3472, DOI: 10.1039 / b003825n

[0101] The novel binuclear copper(II) complex of Formula I with substituted salben-type ligands is novel and thus constitutes a particular embodiment of the present invention.

[0102] In addition to known compounds, the preferred and specific examples outlined above are equally applicable to novel binuclear copper(II) complexes with substituted salben-type ligands.

[0103] Catalyst preparation:

[0104] The catalyst can be prepared according to the following scheme:

[0105]

[0106] H2salben-type derivatives of formula XII can be prepared by reacting an N,N'-bis-(salicylene)-2-hydroxyphenylmethane diamine derivative of formula X with a suitable aldehyde of formula XI in the presence of ammonium acetate or ammonia and a polar protic solvent such as methanol or ethanol at a temperature of -10°C to 40°C for a period of 3 h to 120 h under stirring. The resulting ligands can be separated by filtration, washing with a polar protic solvent, and drying under vacuum.

[0107] The formation of a binuclear copper(II) complex of Formula I with a substituted salben-type ligand can then be carried out by adding a copper(II) salt CuX2 (preferably acetate monohydrate) to the H2 salben-type ligand of Formula XII suspended in a polar protic solvent (preferably methanol or ethanol) under stirring at a temperature of -10°C to 40°C in the presence of a tertiary amine (preferably triethylamine) for a period of ½ h to 24 h. The resulting complex can be separated by filtration, washing with a polar protic solvent, and drying under vacuum.

[0108] CN coupling method:

[0109] The CN coupling method, namely the coupling of aryl halides with N-nucleophilic compounds in the presence of a binuclear copper(II) complex of formula I with a substituted salben-type ligand, is advantageously carried out in the presence of a suitable base.

[0110] The base may be selected from alkali metal salts of inorganic or organic acids, such as cesium carbonate, potassium phosphate, potassium carbonate, or sodium acetate, or from alkali metal alkoxides, such as sodium tert-butoxide or potassium tert-butoxide.

[0111] The preferred bases are cesium carbonate and potassium phosphate.

[0112] Typically, the alkali is applied in the following equivalents: 1 to 5, preferably 1.5 to 4.

[0113] Furthermore, the reaction is carried out in the presence of an organic solvent, which may be selected from aliphatic alcohols such as methanol, ethanol, tert-amyl alcohol, N,N-dimethylformamide, tetrahydrofuran, acetonitrile, toluene, or dimethyl sulfoxide.

[0114] Preferred organic solvents are acetonitrile, tetrahydrofuran, and dimethyl sulfoxide.

[0115] The equivalent of the aryl halide relative to 1 equivalent of the N-nucleophile can vary depending on the halide, but typically ranges from 0.2 to 3.0:1.0, preferably from 0.3 to 2.5:1.0.

[0116] The reaction temperature is typically maintained between 20°C and 200°C, preferably between 50°C and 100°C, and more preferably between 70°C and 90°C.

[0117] The catalyst loading (referring to the limiting reagent, which may be an N-nucleophile or an aryl halide) in mol% can vary between 0.1 mol% and 5.0 mol%, preferably between 0.25 mol% and 3.0 mol%.

[0118] After a reaction time of 4 to 24 hours, preferably 8 to 18 hours, the resulting arylamine can be separated by filtering out the catalyst and evaporating the organic phase. The resulting crude arylamine can be further purified by chromatography.

[0119] Example:

[0120] abbreviation:

[0121] RT = room temperature

[0122] MeOH = Methanol

[0123] Et2O = Diethyl ether

[0124] iPr2O = diisopropyl ether

[0125] Et3N = Triethylamine

[0126] EtOH = ethanol

[0127] MeCN = Acetonitrile

[0128] DMSO = dimethyl sulfoxide

[0129] THF = Tetrahydrofuran

[0130] DCM = dichloromethane

[0131] 1. Activity of copper(II) complexes as catalysts for CN coupling reactions

[0132] 1.1 Optimization Conditions

[0133] The coupling of aryl iodine with pyrazole can be achieved by reacting with 0.5 mol% [Cu2(μ-salben)2] catalyst in acetonitrile (MeCN) at 82 °C in the presence of 2 equivalents of Cs2CO3. Complete conversion was observed after 16 h, and the product could be isolated in 83% yield.

[0134]

[0135] 1.2 Optimization Research

[0136] Table 1. Optimization of temperature and catalyst loading.

[0137]

[0138] General conditions: Iodobenzene (215 µL, 1.93 mmol, 1.5 equiv), pyrrole (90 µL, 1.29 mmol, 1 equiv), Cs2CO3 (839.2 mg, 2.58 mmol, 2 equiv), [Cu2(μ-salben)2], in 1 mL MeCN. [a] Catalyst loading refers to the amount of pyrrole. [b] 1 H NMR yield, with the fraction in parentheses indicating the separation yield.

[0139] Table 2. Screening of bases in MeCN.

[0140]

[0141] General conditions: Iodobenzene (715 µL, 6.42 mmol, 1.5 equiv), pyrrole (300 µL, 4.29 mmol, 1 equiv), base (2 equiv), [Cu2(μ-salben)2], in 3.5 mL MeCN. [a] Catalyst loading refers to the amount of pyrrole. [b] 1 H NMR yield, with the fraction in parentheses indicating the separation yield. [c] Tetrahydrofuran (THF) was used as a solvent. [d] K3PO4 (3.64g, 17.18 mmol, 4 equiv).

[0142] Table 3. Screening of solvents and bases.

[0143]

[0144] General conditions: Iodobenzene (215 µL, 1.93 mmol, 1.5 equiv), pyrrole (90 µL, 1.29 mmol, 1 equiv), base (2.58 mmol, 2 equiv), [Cu2(μ-salben)2] (2.52 mg, 0.003 mmol, 0.0025 equiv), in 1 mL solvent; T = 70℃–82℃, t = 16 h. [a] DMF = dimethylformamide, DMSO = dimethyl sulfoxide, EtOH = ethanol.

[0145] Table 4. Activity of different copper(II) derivatives as catalysts.

[0146]

[0147] General conditions: Iodobenzene (215 µL, 1.93 mmol, 1.5 equiv), pyrrole (90 µL, 1.29 mmol, 1 equiv), base (2.58 mmol, 2 equiv), 1 mL MeCN; T = 82℃, t = 16 h. [a] LC / MS yield (%).

[0148] Table 5. Effects of various equivalents of alkali and iodobenzene.

[0149]

[0150] General conditions: Iodobenzene, pyrrole (90 µL, 1.29 mmol, 1 equiv), K3PO4, [Cu2(μ-salben)2] (0.006 mmol, 0.005 equiv), in 1 mL MeCN; T = 82℃, t = 16 h.

[0151] 1.3. Substrate range

[0152] Table 6. Substrate range of aryl iodides

[0153]

[0154] General conditions: aryl iodine (1.5 equiv), N-nucleophile (1 eq), Cs₂CO₃ (2 equiv), [Cu₂(μ-salben)₂] (0.5 mol %), in MeCN, c = 1 mol L –1 T = 82℃, t = 16 h.

[0155] Table 7. Substrate range of aryl bromides

[0156]

[0157] General conditions: aryl bromide (1 equiv), N-nucleophile (n equiv, as reported in the table), base, [Cu2(μ-salben)2] (2.5 mol%), base (n equiv, as reported in the table), solvent, c = 0.5 mol L –1 T = 85℃, t = 16 h.

[0158] 2. Synthesis of novel salben-type ligands

[0159] 2.1 Synthesis of H2sal(2,6-diOMe)ben

[0160]

[0161] H3salmp (0.499 g, 1.44 mmol) and 2,6-dimethoxybenzaldehyde (0.478 g, 2.87 mmol) were added to a solution of ammonium acetate (0.113 g, 1.46 mmol) in 5 mL of methanol (MeOH) to obtain a yellow suspension, which was placed at room temperature (RT) for 24 h with magnetic stirring. The yellow product was then recovered by filtration, washed with MeOH (2 × 2 mL) and diethyl ether (Et2O) (2 × 2 mL), and dried under vacuum for several hours to give 0.645 g of bright yellow powdery solid (76% yield). For C 23 H 22 Elemental analysis of N₂O₄ (MW = 390.44 g / mol): Calculated values: C 70.75%; H 5.68%; N 7.17%. Measured values: C 70.54%; H 5.79%; N 7.24%. IR spectrum, cm⁻¹ -1 (ATR): 3050-2830, 1628, 1585, 1493-1474, 1397, 1276, 1250, 1110-1067, 758. 1 H NMR, 298 K, CDCl3, 400 MHz: δ (ppm) 13.62 (2H, s), 8.55 (2H, d), 7.32-7.28(4H, m), 7.26 (1H, t), 6.95 (2H, dd), 6.87 (2H, td), 6.77 (1H,s), 6.58 (2H,d), 3.85 (3H,s). 13 C NMR, 298 K, CDCl3, 101 MHz: δ (ppm) 163.4, 161.5, 158.5, 132.6, 132.0, 130.3, 119.1, 118.6, 117.3, 115.6, 104.7, 79.6, 56.0.

[0162] 2.2 Synthesis of H2sal(2)thn

[0163]

[0164] H3salmp (3.012 g, 8.695 mmol) was added to a solution of ammonium acetate (2.024 g, 26.26 mmol) in EtOH (50 mL). Thiophene-2-carboxaldehyde (1.70 mL, 18.5 mmol) was then added to the suspension, and the reaction mixture was stirred at RT for 4 days. The resulting pale yellow solid was filtered, washed with EtOH and diisopropyl ether (iPr2O), and dried under vacuum for several hours to give 2.946 g of the title compound (67%). (For C...) 19 H 16 Elemental analysis of N₂O₂S (MW = 336.41): Calculated values: C 67.84%; H 4.79%; N 8.33%; S 9.53%. Measured values: C 67.77%; H 4.84%; N 8.15%; S 10.05%. mp: 109℃-112℃. IR spectrum, cm⁻¹ –1 (ATR): 3109, 2853, 1618, 1575, 1497, 1456, 1422, 1372, 1357, 1272, 1040, 1025, 975,897, 753, 712, 598. 1 H NMR, 298 K, acetone-d6, 400 MHz: δ (ppm) 6.44 (s, 1H), 6.94(dd, 2H), 7.08 (dd, 2H), 7.24 (dt, 2H), 7.40 (ddd, 2H), 7.51 (dd, 2H), 7.52(dd, 2H), 8.85 (s, 2H), 12.82 (s, 2H). 13 C10 NMR, 298 K, acetone-d6, 101 MHz: δ (ppm) 86.2, 116.8, 118.7, 119.0, 125.2, 126.8, 127.2, 132.8, 133.4, 144.4, 161.1, 166.5. Mass spectrometry (ESI) + In MeOH, m / z (intensity %): relative to [M + H] + Calculated value: 337.1011; Measured value: 337.1000 (100) The method described is used to prepare trispecific antibodies.

[0165] 2.3 Synthesis of H2sal(3)thn

[0166]

[0167] H3salmp (3.011 g, 8.694 mmol) was added to a solution of ammonium acetate (2.017 g, 26.17 mmol) in EtOH (30 mL). Thiophene-3-carboxaldehyde (2.012 g, 17.94 mmol) was added to the suspension, and the reaction mixture was stirred at RT for 4 days. The resulting pale yellow solid was filtered, washed with EtOH and iPr2O, and dried under vacuum to give 3.268 g of the title compound (75%). (For C...) 19 H 16 Elemental analysis of N₂O₂S (MW = 336.41): Calculated values: C 67.84%, H 4.79%, N 8.33%, S 9.53%. Measured values: C 67.18%, H 4.76%, N 8.55%, S 10.50%. mp: 94℃-96℃. IR spectrum, cm⁻¹ –1 (ATR): 3107, 3054, 3000, 2858, 1618,1576, 1500, 1456, 1424, 1368, 1273, 1043, 976, 779, 752, 733, 635. 1 H NMR, 298K, acetone-d6, 400 MHz: δ (ppm) 6.26 (s, 1H), 6.94 (d, 2H), 6.96 (dd, 2H), 7.29(dd, 2H), 7.39 (dd, 2H), 7.50 (dd, 2H), 7.54 (dd, 1H), 7.56 (m, 1H), 8.83 (s, 2H), 13.01 (s, 2H). 13 C10 NMR, 298 K, acetone-d6, 101 MHz: δ (ppm) 87.5, 117.6, 119.6, 119.7, 123.4, 127.0, 127.8, 133.5, 133.9, 143.0, 162.0, 167.0. Mass spectrometry (ESI) + In MeOH, m / z (intensity %): relative to [M + H] + Calculated value: 337.1011; Measured value: 337.1001(100) The method described is used to prepare trispecific antibodies.

[0168] 2.4.1 Synthesis of (R)-H2salmyr

[0169]

[0170] H3salmp (0.30135 g, 0.870 mmol) was added to a solution of NH4OAc (0.20471 g, 2.66 mmol) in MeOH (10 mL). (R)-Myrtol (0.25308 g, 1.68 mmol) was added to the yellow suspension and stirred at RT for 1 day, during which time H3salmp completely dissolved within 6 h, and the desired product precipitated after 12 h. The resulting pale yellow solid was filtered, washed with MeOH (3 × 3 mL) and iPr2O (3 × 3 mL), and dried under vacuum for 1 h. Yield: 0.29610 g (61%). (For C) 24 H 26 Elemental analysis of N2O2S (374.48): Calculated values ​​(%): C 76.98, H 7.00, N 7.48. Measured values: C 76.78, H 7.04, N 7.51. Melting point: 184℃-187℃. 1 ¹H NMR (400 MHz, 298 K, acetone-d6), in ppm: δ 0.86 (3H, s), 1.43 (3H, s), 1.85 (1H, d, 10 Hz), 2.00 (1H, dd, 2 – 4 Hz), 2.14 – 2.19 (1H, m), 2.57 – 2.66 (2H, m), 3.45 (1H, t, 5 Hz), 4.40 (1H, d, 8 Hz), 6.83 (1H, s), 6.86 – 6.94 (4H, m, 1 – 7 Hz), 7.30 – 7.34 (2H, m, 2 Hz), 7.37 (1H, dd, 2 – 8 Hz), 7.43 (1H, dd, 2 – 8 Hz), 8.40 (1H, s), 8.61 (1, s), 13.15 (1H,s), 13.20 (1H, s). 13C NMR (101 MHz, 298 K acetone-d6), in ppm: δ 22.3, 26.4, 27.5, 34.9, 41.5, 42.0, 45.9, 64.2, 117.4, 117.4, 119.5, 119.8, 120.1, 120.4, 132.6, 132.7, 133.0, 133.3, 138.5, 144.8, 161.6, 161.9, 163.3, 164.2. IR-ATR (cm −1 (): 2993, 2969, 2949, 2865, 1645, 1620, 1596, 1563, 1495, 1456, 1413, 1300, 1278, 1244, 1148, 1079, 971, 844, 753. Mass spectrometry (ESI) + In MeCN, m / z (intensity %) is defined for [M + H]. + Calculated value: 375.2073; Measured value: 375.2060 (100) The method described above is used to prepare trispecific antibodies.

[0171] 2.4.2 Synthesis of (R)-H2salC1myr

[0172]

[0173] (R)-Myrtol (0.1352 g, 0.900 mmol) was added to NH3 (750 μL of aqueous solution, 25%). Salicylic acid (salH) (0.2001 g, 1.64 mmol) was added to a colorless solution in MeOH (2.5 mL). The mixture was stirred at RT for 3 h to form the desired product as a precipitate. The resulting pale yellow solid was filtered, washed with MeOH (3 × 3 mL) and iPr₂O (3 × 3 mL), and dried under vacuum overnight. Yield: 0.2112 g (69%). (For C) 24 H 26 Elemental analysis of N₂O₂·0.2 H₂O (382.08): Calculated values ​​(%): C 76.24, H 7.04, N 7.41. Measured values: C 76.04, H 6.68, N 7.38. Melting point: 135℃-139℃. 1¹H NMR (400 MHz, 298 K, acetone-d6), in ppm: δ 0.79 (3H, s), 1.28 (3H, s), 1.85 (1H, d, 10 Hz), 2.09–2.14 (1H, m), 2.27–2.42 (2H, m, 6 Hz), 2.45–2.48 (2H, m), 5.49 (1H, s), 5.76 (1H, s), 6.91–6.95 (4H, m), 7.35–7.40 (2H, m), 7.48 (2H, dd, 1–8 Hz), 8.71 (1H, s), 8.73 (1H, s), 13.04 (1H, s), 13.06 (1H, s). 13 C NMR (101 MHz, 298 K acetone-d6), in ppm: δ 21.6, 26.4, 32.0, 32.3, 38.6, 41.6, 43.1, 91.7, 117.6, 119.6, 121.4, 121.4, 133.3, 133.7, 148.2, 162.0, 166.6, 166.8. IR-ATR (cm −1 ): 2989, 2969, 2927, 2877, 1620, 1574, 1494, 1457, 1413, 1365, 1274, 1212, 1150, 1039, 976, 897, 836, 751. Mass spectrometry (ESI+), in MeCN, m / z (intensity %): For [M+H]+, calculated value: 375.2073; measured value: 335.2067 (100).

[0174] 2.6 Synthesis of H2(4-OMe)salben

[0175]

[0176] Benzaldehyde (0.1631 mg, 1.537 mmol) was added to a supersaturated solution of ammonium acetate (0.1631 g, 1.5368 mmol) in MeOH (2.5 mL), and stirred at RT for 5 min under an inert atmosphere. Then, 4-OMe-salH (0.4175 g, 2.734 mmol) was added to a colorless solution, and the reaction mixture was stirred at RT for 4 h. The resulting pale gray solid was filtered, washed with MeOH (3 × 3 mL), and dried under vacuum for 1 h. Yield: 0.2615 g (49%). (For C...) 23 H 22 Elemental analysis of N₂O₄ (390.44): Calculated values ​​(%): C 70.76, H 5.68, N 7.16. Measured values: C 70.70, H 5.80, N 6.85. Melting point: 111℃-112℃. 1 ¹H NMR (400 MHz, 298 K, acetone-d6), in ppm: δ 13.44 (2H, s), 8.74 (2H, s), 7.56 (2H, d, 8 Hz), 7.47–7.43 (2H, m), 7.40 (2H, d, 8 Hz), 7.39–7.34 (1H, m), 6.51 (2H, dd, 8–2Hz), 6.46 (2H, d, 2 Hz), 6.03 (1H, s), 3.84 (6H, s). 13 C10 NMR (151 MHz, 298 K, acetone-d6) δ 164.9, 163.9, 163.5, 141.8, 133.8, 128.8, 128.2, 127.0, 112.6, 106.4, 100.8, 90.4, 54.9. IR-ATR (cm −1 (): 1610, 1570, 1452, 1316, 1288, 1240, 1168, 764. Mass spectrometry (ESI) + In MeCN, m / z (intensity %) is defined for [M + H]. + Calculated value: 391.1658; Measured value: 391.1639 (100) The method described is used to prepare trispecific antibodies.

[0177] 2.7 Synthesis of H2(5-OMe)salben

[0178]

[0179] Benzaldehyde (84.2 mg, 0.794 mmol) was added to a supersaturated solution of ammonium acetate (0.9672 g, 15.81 mmol) in MeOH (2.5 mL), and stirred at RT for 5 min under an inert atmosphere. 5-OMe-salH (201.3 mg, 1.332 mmol) was added to a colorless solution, and the reaction mixture was stirred at RT for 3 h. The resulting pale yellow solid was filtered, washed with MeOH (3 × 3 mL) and iPr₂O (3 × 3 mL), and dried under vacuum for 1 h. Yield: 0.1754 g (68%). (For C) 23 H 22 Elemental analysis of N₂O₄ (390.44): Calculated values ​​(%): C 70.76, H 5.68, N 7.16. Measured values: C 71.04, H 5.71, N 7.26. Melting point: 137℃-141℃. 1 ¹H NMR (600 MHz, 298 K, acetone-d6), in ppm: δ 12.49 (2H, s), 8.82 (2H, s), 7.59 – 7.54 (2H, m), 7.47 – 7.42 (2H, m), 7.39 – 7.33 (1H, m), 7.08 (2H, d, 3 Hz), 7.00 (2H, dd, 9– 3 Hz), 6.86 (2H, d, 9 Hz), 6.14 (1H, s), 3.76 (6H, s). 13 C10 NMR (151 MHz, 298 K, acetone-d6) δ 166.7, 156.1, 153.3, 129.8, 129.3, 127.8, 121.2, 119.3, 118.4, 116.3, 91.5, 56.1. Mass spectrometry (ESI) + In MeCN, m / z (intensity %) is defined for [M + H]. + Calculated value: 391.1658; Measured value: 391.1655 (100) The method described is used to prepare trispecific antibodies.

[0180] 2.8 Synthesis of H2(5-F)salben

[0181]

[0182] Benzaldehyde (83.2 mg, 0.784 mmol) was added to a supersaturated solution of ammonium acetate (0.8828 g, 11.442 mmol) in MeOH (2.5 mL) and stirred at RT for 5 min. 5-F-salH (206.1 mg, 1.4708 mmol) was added to a colorless solution, and the reaction mixture was stirred at RT for 3 h. The resulting yellow solid was filtered, washed with MeOH (3 × 3 mL) and iPr₂O (3 × 3 mL), and dried under vacuum for 1 h. Yield: 0.2295 g (85%). (For C...) 21 H 16 Elemental analysis of F2N2O2 (366.12): Calculated values ​​(%): C 68.45, H 4.40, N 7.65. Measured values: C 68.19, H 4.32, N 7.54. Melting point: 144℃-147℃. 1 H NMR (600 MHz, 298 K, acetone-d6)δ 12.78 (2H, d, 4 Hz), 8.86 (2H, d, 3 Hz), 7.58 (2H, d, 8 Hz), 7.46 (2H, dd,8 Hz), 7.43 – 7.35 (1H, m), 7.32 (2H, ddd, 9 – 3 – 2 Hz), 7.19 (2H, ddd, 9 – 3 Hz), 6.94 (2H, ddd, 9 – 4 – 2 Hz), 6.20 (1H, d, 2 Hz). 13 C NMR (151 MHz, 298 K, acetone-d6) δ 166.1, 158.3, 156.3 (d, J) C-F =233 Hz), 141.7, 129.8, 129.4, 127.9, 120.8 (d, 23 Hz), 119.6 (d, 8 Hz), 118.9 (d, 8 Hz), 118.3 (d, 23 Hz), 91.4. Mass spectrometry (ESI) + In MeCN, m / z (intensity %) is defined for [M + H]. + Calculated value: 367.1258; Measured value: 367.1254 (100) The method described is used to prepare trispecific antibodies.

[0183] 2.9 Synthesis of H2(5-Cl)salben

[0184]

[0185] Benzaldehyde (81.1 mg, 0.764 mmol) was added to a supersaturated solution of ammonium acetate (0.9869 g, 12.80 mmol) in MeOH (3 mL) and stirred at RT for 5 min. 5-Cl-salH (200.6 mg, 1.281 mmol) was added to a colorless solution, and the reaction mixture was stirred at RT for 3 h. The resulting yellow solid was filtered, washed with MeOH (3 × 3 mL) and iPr₂O (3 × 3 mL), and dried under vacuum for 1 h. Yield: 0.1567 g (61%). (For C) 21 H 16 Elemental analysis of Cl2N2O2·0.3 H2O (405.28): Calculated values ​​(%): C 62.33, H 4.13, N 6.92. Measured values: C 62.33, H 3.96, N 6.82. Melting point: 128℃-131℃. 1 ¹H NMR (600 MHz, 298 K acetone-d6) δ 13.05 (2H, s), 8.86 (2H, s), 7.60 – 7.54 (4H, m, 2 – 7 Hz), 7.49 – 7.40 (2H, m(dd), 7 Hz), 7.40 – 7.34 (3H, m, 2 – 9 Hz), 6.95 (2H, d, 9 Hz), 6.21 (1H, s). 13 C10 NMR (151 MHz, 298 K, acetone-d6) δ 166.1, 160.7, 141.6, 133.6, 132.5, 129.9, 129.5, 127.9, 123.8, 120.8, 119.5, 91.2. Mass spectrometry (ESI) + In MeCN, m / z (intensity %) is defined for [M + Na]. + Calculated value: 399.0667; Measured value: 399.0661 (100) The method described above is used to prepare trispecific antibodies.

[0186] 2.10 Synthesis of H2(5-Me)salben

[0187]

[0188] Benzaldehyde (84.2 mg, 0.8015 mmol) was added to a supersaturated solution of ammonium acetate (0.8636 g, 11.20 mmol) in MeOH (2.5 mL) and stirred at RT for 5 min. 5-Me-salH (214.5 mg, 1.5755 mmol) was added to a colorless solution, and the reaction mixture was stirred at RT for 3 h. The resulting yellow solid was filtered, washed with MeOH (3 × 3 mL), and dried under vacuum for 1 h. Yield: 0.2041 g (72%). (For C...) 23 H 22 Elemental analysis of N₂O₂·0.05NH₄OAc (362.29): Calculated values ​​(%): C 76.58, H 6.22, N 7.93. Measured values: C 76.38, H 5.98, N 7.80. Melting point: 125℃-130℃. 1 H NMR (400 MHz, 298 K, CDCl3) δ12.72 (2H, s), 8.53 (2H, s), 7.49 – 7.29 (5H, m), 7.16 (2H, dd, 8 – 2 Hz), 7.11 (2H, d, 2 Hz), 6.89 (2H, d, 8 Hz), 5.99 (1H, s), 2.29 (6H, s). Mass spectrometry (ESI + In MeCN, m / z (intensity %) is defined for [M + H]. + Calculated value: 359.1758; Measured value: 359.1747 (100) The method described above is used to prepare trispecific antibodies.

[0189] 2.11 Synthesis of H2(3-OMe)sal(2-Cl)ben

[0190]

[0191] 2-Cl-benzaldehyde (92.2 mg, 0.657 mmol) was added to a solution of ammonium acetate (0.8808 g, 11.40 mmol) in MeOH (10 mL) and stirred at RT for 5 min. 3-OMe-salH (0.2085 g, 1.370 mmol) was added to a colorless solution, and the reaction mixture was stirred at RT for 3 h. The resulting yellow solid was filtered, washed with MeOH (3 × 3 mL), and dried under vacuum for 1 h. Yield: 0.1431 g (51%). (For C...) 23H 21 Elemental analysis of ClN2O4 (424.12): Calculated values ​​(%): C 65.02, H 4.98, N 6.59. Measured values: C 64.56, H 4.91, N 6.54. 1 H NMR (400 MHz, 298 K, acetone-d6) δ 13.00 (2H, s), 8.91(2H, s), 7.72 (1H, dd, 1 - 8 Hz), 7.53 (1H, dd, 1 - 8 Hz), 7.49 (1H, dd, 1 -7 Hz), 7.43 (1H, dd, 2 - 7 H) 7.15 (2H, d, 8Hz), 7.11 (2H, d, 8 Hz), 6.89 (2H, dd, 8 Hz), 6.55 (1H, s), 3.85 (6H, s). 13 C NMR (101 MHz, 298 K, acetone-d6) δ 166.7, 151.5, 148.5, 138.1, 132.6, 130.0, 129.2, 127.2, 124.1, 118.8, 118.6, 118.5, 115.8, 86.3, 55.6. IR-ATR (cm −1 (): 1613, 1600, 1459, 1317, 1252, 1171, 764, 728. Mass spectrometry (ESI) + In MeCN, m / z (intensity %) is defined for [M + H]. + Calculated value: 425.1268; Measured value: 425.1248 (60) The method described above is used to prepare trispecific antibodies.

[0192] 2.12 Synthesis of H2(5-OMe)sal(2-Cl)ben

[0193]

[0194] 2-Cl-benzaldehyde (0.1322 g, 0.8689 mmol) was added to a solution of ammonium acetate (0.21665 g, 2.038 mmol) in MeOH (10 mL) and stirred at RT for 5 min. 5-OMe-salH (0.2864 g, 2.038 mmol) was added to a colorless solution, and the reaction mixture was stirred at RT for 3 h. The resulting yellow solid was filtered, washed with MeOH (3 × 3 mL), and dried under vacuum for 1 h. Yield: 0.1278 g (32%). (For C...) 23 H 21 Elemental analysis of ClN2O4 (424.12): Calculated values ​​(%): C 65.02, H 4.98, N 6.59. Measured values: C 64.78, H 4.95, N 6.51. 1 H NMR (400 MHz, 298 K, acetone-d6) δ 12.34 (2H, s), 8.85(2H, s), 7.71 (1H, dd, 8 – 2 Hz), 7.51 (1H, dd, 8 – 2 Hz), 7.47 (1H, dd, 8 –2 Hz), 7.41 (1H, dd, 8 – 2 Hz), 7.12 (2H, d, 3 Hz), 7.02 (2H, dd, 9 – 3 Hz), 6.86 (2H, d, 9 Hz), 6.54 (1H, s), 3.76 (6H, s). 13 C NMR (101 MHz, 298 K, acetone-d6) δ 166.7, 155.2, 152.5, 138.1, 132.6, 130.0, 129.2, 127.9, 120.6, 118.4, 117.6, 115.5, 115.4, 86.5, 55.2. IR-ATR (cm −1 (): 1634, 1584, 1487, 1329, 1266, 1225, 1165, 770. Mass spectrometry (ESI) + In MeCN, m / z (intensity %) is defined for [M + H]. + Calculated value: 425.1268; Measured value: 425.1248 (60) The method described above is used to prepare trispecific antibodies.

[0195] 2.13 Synthesis of H2(5-OMe)sal(2-OMe)ben

[0196]

[0197] 2-OMe-benzaldehyde (0.1349 g, 0.9584 mmol) was added to a solution of ammonium acetate (0.2294 g, 2.977 mmol) in MeOH (10 mL) and stirred at RT for 5 min. 5-OMe-salH (0.2883 g, 1.895 mmol) was added to a colorless solution, and the reaction mixture was stirred at RT for 3 h. The resulting yellow solid was filtered, washed with MeOH (3 × 3 mL), and dried under vacuum for 1 h. Yield: 0.3028 g (76%). (For C...) 24 H 24 Elemental analysis of N2O5 (420.47): Calculated values ​​(%): C 68.75, H 5.75, N 6.62. Measured values: C 69.07, H 5.83, N 6.68. 1 H NMR (400 MHz, 298 K, acetone-d6) δ 12.60 (2H, s), 8.78 (2H, s), 7.49 (1H, dd, 8 – 2 Hz), 7.35 (1H, dd, 9 Hz), 7.10 – 7.07 (3H, m), 7.03 (1H,dd, 6 Hz), 7.00 (2H, dd, 9 – 3 Hz), 6.85 (2H, d, 9 Hz), 6.50 (1H, s), 3.96 (3H, s) 3.76 (6H, s). 13 C NMR (101 MHz, 298 K, acetone-d6) δ 165.7, 156.8, 154.9, 153.2, 152.8, 129.6, 128.6, 127.5, 120.8, 120.0, 118.6, 117.5, 115.4, 111.4, 84.2, 55.2. IR-ATR (cm −1 (): 1635, 1585, 1489, 1328, 1266, 1240, 1167, 760. Mass spectrometry (ESI) + In MeCN, m / z (intensity %) is defined for [M + H]. + Calculated value: 421.1764; Measured value: 421.2400 (40) The method described above is used to prepare trispecific antibodies.

[0198] 2.14 Synthesis of H2(5-Cl)sal(2-OMe)ben

[0199]

[0200] 2-OMe-benzaldehyde (0.1287 g, 0.9449 mmol) was added to a solution of ammonium acetate (0.2173 g, 2.819 mmol) in MeOH (10 mL) and stirred at RT for 5 min. 5-Cl-salH (0.2918 g, 1.864 mmol) was added to a colorless solution, and the reaction mixture was stirred at RT for 3 h. The resulting yellow solid was filtered, washed with MeOH (3 × 3 mL), and dried under vacuum for 1 h. Yield: 0.2996 g (76%). (For C...) 22 H 18 Elemental analysis of Cl2N2O3 (429.30): Calculated values ​​(%): C 61.35, H 4.23, N 6.53. Measured values: C 61.05, H 4.26, N 6.55. Melting point (°C): 152 – 154. 1 ¹H NMR (400 MHz, 298 K, acetone-d6) δ 13.23 (2H, s), 8.90 (2H, s), 7.63 (2H, d, 3 Hz), 7.55 (1H, dd, 8 – 2 Hz), 7.36–7.30 (3H, m), 7.09 (1H, d, 9 Hz), 7.05 (1H, ddd, 8 – 1 Hz), 6.90 (2H, d, 9 Hz), 6.50 (1H, s), 3.90 (3H, s). 13 C NMR (101 MHz, 298 K, acetone-d6) δ 166.9, 162.2, 157.9, 133.8, 133.4, 130.7, 129.6, 128.5, 121.8, 119.9, 119.8, 117.7, 112.3, 85.2, 56.1. IR-ATR (cm −1 (): 1606, 1521, 1453, 1384, 1297, 1242, 1165, 748. Mass spectrometry (ESI) + In MeCN, m / z (intensity %) is defined for [M + H]. + Calculated value: 429.0773; Measured value: 429.0660 (40) The method described above is used to prepare trispecific antibodies.

[0201] 2.15 Synthesis of H2(5-OMe)sal(2-NO2)ben

[0202]

[0203] 2-NO2-benzaldehyde (0.1390 ​​g, 0.9199 mmol) was added to a solution of ammonium acetate (0.2212 g, 2.869 mmol) in MeOH (10 mL) and stirred at RT for 5 min. 5-OMe-salH (0.2796 g, 1.837 mmol) was added to a colorless solution, and the reaction mixture was stirred at RT for 3 h. The resulting yellow solid was filtered, washed with MeOH (3 × 3 mL), and dried under vacuum for 1 h. Yield: 0.1957 g (49%). (For C...) 23 H 21 Elemental analysis of N3O6 (435.42): Calculated values ​​(%): C 63.24, H 4.86, N 9.65. Measured values: C 62.98, H 4.90, N 9.25. Melting point (°C): 159 – 160. 1 H NMR (400 MHz, 298 K, acetone-d6) δ 12.20 (2H,s), 8.82 (2H, s), 8.01 (1H, d, 8 Hz), 7.97 (1H, dd, 8 – 1 Hz), 7.86 (1H, dd,7 – 1 Hz), 7.66 (1H, dd, 8 - 1 Hz), 7.11 (2H, d, 3 Hz), 7.01 (2H, dd, 9 - 3Hz), 6.85 (2H, dd, 9 Hz), 6.66 (1H, s), 3.77 (6H, s). 13 C NMR (101 MHz, 298 K, acetone-d6) δ 166.9, 156.7, 154.8, 153.4, 134.7, 133.7, 129.7, 129.6, 124.5, 120.8, 118.3, 117.6, 115.6, 84.7, 55.9. IR-ATR (cm −1 (): 1626, 1586, 1525, 1489, 1370, 1338, 1158, 767. Mass spectrometry (ESI) + In MeCN, m / z (intensity %) is defined for [M + H]. +Calculated value: 436.1483; Measured value: 436.1493 (60) The method described above is used to prepare trispecific antibodies.

[0204] 2.16 Synthesis of H2(5-Cl)sal(2-NO2)ben

[0205]

[0206] 2-NO2-benzaldehyde (0.0976 g, 0.646 mmol) was added to a solution of ammonium acetate (0.1677 g, 2.175 mmol) in MeOH (10 mL) and stirred at RT for 5 min. 5-Cl-salH (0.2096 g, 1.339 mmol) was added to a colorless solution, and the reaction mixture was stirred at RT for 3 h. The resulting yellow solid was filtered, washed with MeOH (3 × 3 mL), and dried under vacuum for 1 h. Yield: 0.477 g (50%). (For C...) 22 H 15 Elemental analysis of Cl2N3O4 (443.27): Calculated values ​​(%): C 56.77, H 3.40, N 9.46. Measured values: C 57.26, H 3.42, N 9.61. Melting point (°C): 148 - 151. 1 H NMR (400 MHz, 298 K, acetone-d6) δ12.73 (2H, s), 8.80 (2H, s), 8.04 (1H, dd, 8 – 2 Hz), 7.99 (1H, dd, 7.8 – 1.3Hz), 7.87 (1H, dd, 7 Hz), 7.69 (1H, dd, 8 Hz), 7.60 (2H, d, 3 Hz), 7.40 (2H,dd, 9 – 3 Hz), 6.95 (2H, d, 9 Hz), 6.73 (1H, s). 13 C10 NMR (101 MHz, 298 K, acetone-d6) δ 167.7, 160.4, 150.3, 135.0, 134.9, 133.3, 132.5, 130.4, 129.6, 124.9, 124.0, 120.06, 119.6, 84.5. IR-ATR (cm −1 (): 1616, 1559, 1520, 1473, 1342, 1199, 792, 751. Mass spectrometry (ESI) +In MeCN, m / z (intensity %) is defined for [M + H]. + Calculated value: 444.0417; Measured value: 444.0505 (40) The method described above is used to prepare trispecific antibodies.

[0207] 3. Synthesis of new copper(II) binuclear complexes

[0208] 3.1 Synthesis of [Cu2(μ-sal(4-tBu)ben)2]

[0209]

[0210] Et3N (0.109 g, 1.08 mmol) was added to a yellow suspension of H2sal(4-tBu)ben (0.2021 g, 0.5229 mmol) in 8 mL of EtOH. Then, solid Cu(OAc)2·H2O (0.1017 g, 0.5094 mmol) was added, and the color turned brown within minutes. The reaction mixture was placed at RT with magnetic stirring for 3 h, and the precipitate was then separated by filtration, washed with EtOH (2 × 2 mL), and dried under vacuum for several hours. Yield: 0.1882 g of brown solid (83%). (For C) 50 H 48 Elemental analysis of Cu₂N₄O₄·0.8EtOH (932.91): Calculated values ​​(%): C 66.43, H 5.70, N 6.01. Measured values: C 66.51, H 5.70, N 6.01. IR-ATR (cm⁻¹) −1 (): 3047, 2964, 1606, 1533, 1466, 1387, 1317, 1192, 1147, 1107, 981, 928, 827. Mass spectrometry (ESI) + In MeCN, m / z (intensity %) is defined for [M + H]. + Calculated value: 895.2346; Measured value: 895.2327 (100) The method described above is used to prepare trispecific antibodies.

[0211] The ligand structure has been disclosed in Rigamonti, L et al., Int. J. Mol. Sci. 2020, 21, 7882, DOI: :10.3390 / ijms21217882.

[0212] 3.2 Synthesis of [Cu2(μ-sal(2-OMe)ben)2]

[0213]

[0214] Et3N (0.185 g, 1.83 mmol) was added to a yellow suspension of H2sal(2-OMe)ben (0.329 g, 0.922 mmol) in 14 mL of EtOH. Copper acetate monohydrate (0.184 g, 0.912 mmol) was then added, and the color changed to brown / green within minutes. The reaction mixture was allowed to stand at RT with magnetic stirring for 3 h, and the precipitate was then separated by filtration, washed with EtOH (2 × 2 mL) and Et2O (2 × 2 mL), and dried under vacuum for several hours to give 0.354 g of brown solid (92% yield). For C 44 H 36 Elemental analysis of Cu₂N₄O₆ (MW = 843.9 g / mol): Calculated values: C 62.62%; H 4.30%; N 6.64%. Measured values: C 61.85%; H 4.34%; N 6.74%. IR spectrum, cm⁻¹ -1 (ATR): 3044-3010, 1601, 1586, 1531-1490, 1387, 1318, 1242, 1147-1026, 751. Mass spectrometry (ESI) + ), m / z (strength %): 843 [Cu2(sal(2-OMe)ben)2 + 1] + (100).

[0215] 3.3 Synthesis of [Cu2(μ-sal(2,6-diOMe)ben)2]

[0216]

[0217] Et3N (0.150 g, 1.48 mmol) was added to a yellow suspension of H2sal(2,6-diOMe)ben (0.290 g, 0.743 mmol) in 7 mL MeOH. Copper acetate monohydrate (0.148 g, 0.742 mmol) was then added, and the color turned brown within minutes. The reaction mixture was incubated at 0 °C with magnetic stirring for 30 minutes, and the precipitate was then separated by filtration, washed with cold MeOH (2 × 2 mL) and Et2O (2 × 2 mL), and dried under vacuum for several hours to give 0.285 g of brown solid (85% yield). (For C...) 46 H 40Elemental analysis of Cu₂N₄O₈ (MW = 903.94 g / mol): Calculated values: C 61.12%; H 4.46%; N 6.20%. Measured values: C 60.82%; H 4.57%; N 6.19%. IR spectrum, cm⁻¹ -1 (ATR): 3050-2840, 1604, 1528, 1470-1426, 1383, 1250, 1195, 1034-980, 758. Mass spectrometry (ESI) + ), m / z (strength %): 903 [Cu2(sal(2,6-diOMe)ben)2 + 1] + (100).

[0218] 3.4 Synthesis of [Cu2(μ-sal(4-OMe)ben)2]

[0219]

[0220] Et3N (0.138 g, 1.36 mmol) was added to a yellow suspension of H2sal(4-OMe)ben (0.245 g, 0.680 mmol) in 10 mL of EtOH. Copper acetate monohydrate (0.138 g, 0.690 mmol) was then added, and the color changed to green / brown within minutes. The reaction mixture was placed at RT with magnetic stirring for 3 h, and the precipitate was then separated by filtration, washed with EtOH (3 mL), iPr2O (3 mL), and dried under vacuum for several hours to give 0.198 g of brown solid (69% yield). For C 44 H 36 Elemental analysis of Cu₂N₄O₆ (MW = 843.89 g / mol): Calculated values: C 62.62%; H 4.30%; N 6.64%. Measured values: C 63.02%; H 4.34%; N 6.83%. IR spectrum, cm⁻¹ -1 (ATR): 3050-2835, 1607, 1534, 1467-1440, 1395, 1243, 1196, 1176, 1148, 1051-979, 750, 736.

[0221] 3.5 Synthesis of [Cu2(μ-sal(2-Cl)ben)2]

[0222]

[0223] Et3N (0.156 g, 1.54 mmol) was added to a yellow suspension of H2sal(2-Cl)ben (0.255 g, 0.699 mmol) in 10 mL of EtOH. Then, copper acetate monohydrate (0.154 g, 0.774 mmol) was added, and the color changed to green-brown within minutes. After stirring for 3 h, another 5 mL of EtOH was added, and the reaction mixture was then placed at RT with magnetic stirring overnight. The precipitate was then separated by filtration, washed with EtOH (2 × 2 mL) and Et2O (2 × 2 mL), and dried under vacuum for several hours to give 0.285 g of brown solid (96% yield). (For C...) 42 H 30 Elemental analysis of Cl₂Cu₂N₄O₄ (MW = 852.72 g / mol): Calculated values: C 59.16%; H 3.55%; N 6.57%. Measured values: C 59.23%; H 3.46%; N 6.80%. IR spectrum, cm⁻¹ -1 (ATR): 3058-2903, 1605, 1530, 1464-1430, 1319, 1270, 1194, 1149, 1035-978, 744.

[0224] 3.6 Synthesis of [Cu2(μ-sal(2)thn)2]

[0225]

[0226] Et3N (2.11 g, 20.8 mmol) was added to a yellow suspension of H2sal(2)thn (1.699 g, 5.051 mmol) in 75 mL of EtOH. Copper acetate monohydrate (1.012 g, 5.070 mmol) was then added, and the color turned brown within minutes. After stirring at RT for 3 h, the resulting dark brown solid was filtered, washed with EtOH (2 × 10 mL) and iPr2O (2 × 10 mL), and dried under vacuum for several hours to give 1.398 g of the title compound (70%). (For C...) 38 H 28 Elemental analysis of Cu₂N₄O₄S₂ (MW = 795.88): Calculated values: C 57.35%, H 3.55%, N 7.04%, S 8.06%. Measured values: C 57.25%, H 3.54%, N 7.03%, S 8.19%. IR spectrum, cm⁻¹ –1(ATR): 3084, 3016, 2894, 1602, 1531, 1465, 1437, 1393, 1344, 1320, 1241, 1190, 1145, 1123, 1033, 973, 750, 735, 707. Mass spectrometry (ESI) + In MeCN, m / z (intensity %) is defined for [M + H]. + Calculated value: 795.0223; Measured value: 795.0217 (100) The method described is used to prepare trispecific antibodies.

[0227] 3.7 Synthesis of [Cu2(μ-sal(3)thn)2]

[0228]

[0229] Et3N (1.81 g, 17.9 mmol) was added to a yellow suspension of H2sal(2)thn (1.706 g, 5.073 mmol) in 75 mL of EtOH. Copper acetate monohydrate (1.012 g, 5.072 mmol) was then added, and the color turned brown within minutes. After stirring at RT for 3 h, the resulting dark brown solid was filtered, washed with EtOH (2 × 10 mL) and iPr2O (2 × 10 mL), and dried under vacuum for several hours to give 1.695 g of the title compound (84%). (For C...) 38 H 28 Elemental analysis of Cu₂N₄O₄S₂ (MW = 795.88): Calculated values: C 57.35%, H 3.55%, N 7.04%, S 8.06%. Measured values: C 57.11%, H 3.52%, N 7.10%, S 8.06%. IR spectrum, cm⁻¹ –1 (ATR): 3088, 3017, 2898, 1604, 1532, 1465, 1439, 1393, 1343, 1320, 1192, 1145, 1123, 1033, 974, 845, 751, 732. Mass spectrometry (ESI) + In MeCN, m / z (intensity %) is defined for [M + H]. + Calculated value: 795.0223; Measured value: 795.0200 (100) The method described above is used to prepare trispecific antibodies.

[0230] 3.8.1 Synthesis of [Cu2(μ-(R)-salC1myr)2]

[0231]

[0232] Solid Cu(OAc)₂·H₂O (0.09931 g, 0.497 mmol) was added to a yellow suspension of (R)-H₂salC₁myr (0.20174 g, 0.539 mmol) in EtOH (8 mL) and NEt₃ (0.11112 g, 1.10 mmol), and the resulting dark green / brown mixture was stirred at RT for 3 h. The resulting dark brown solid was filtered, washed with EtOH (3 × 3 mL) and iPr₂O (3 × 3 mL), and dried under vacuum for 2 h. Yield: 0.10927 g (50%). The reaction mixture was stored in a refrigerator for 4 days, after which the title compound (0.06016 g, 28%) was further isolated. (For C...) 48 H 48 Elemental analysis of Cu₂N₄O₄ (872.03): Calculated values ​​(%): C 66.11, H 5.55, N 6.43. Measured values: C 66.22, H 5.90, N 6.30. IR-ATR (cm⁻¹) −1 (ESI mass spectrometry): 2970, 2918, 2864, 1600, 1532, 1464, 1445, 1325, 1194, 1145, 1051, 979, 917, 849, 752. + In MeCN, m / z (intensity %) is defined for [M + H]. + Calculated value: 871.2346; Measured value: 871.2319 (100) The method described is used to prepare trispecific antibodies.

[0233] 3.8.2 Synthesis of [Cu2(μ-(R)-salC1myr)2]

[0234] Solid Cu(OAc)₂·H₂O (0.0544 g, 0.273 mmol) was added to a yellow suspension of (R)-H₂salC₁myr (0.1023 g, 0.273 mmol) in EtOH (4 mL) and NEt₃ (0.0566 g, 0.559 mmol), and the resulting dark green / brown mixture was stirred at RT for 3 h. The resulting dark brown solid was filtered, washed with EtOH (3 × 3 mL) and iPr₂O (3 × 3 mL), and dried under vacuum for 2 h. Yield: 0.09696 g (82%). (For C) 48 H 48 Elemental analysis of Cu₂N₄O₄·0.55H₂O (881.94): Calculated values ​​(%): C 65.37, H 5.61, N 6.35. Measured values: C 65.32, H 5.56, N 6.35. IR-ATR (cm⁻¹) −1 ): 3015, 2974, 2926, 2827, 1608, 1533, 1464, 1440, 1348, 1318, 1246, 1194, 1147, 971, 925, 750.

[0235] 3.9 Synthesis of [Cu2(μ-(4-OMe)salben)2]

[0236]

[0237] Et3N (0.109 g, 1.08 mmol) was added to a yellow suspension of H2(4-OMe)salben (0.2333 g, 0.5974 mmol) in 8 mL of EtOH. Then, solid Cu(OAc)2·H2O (0.1216 g, 0.6090 mmol) was added, and the color turned brown within minutes. The reaction mixture was placed at RT with magnetic stirring for 3 h, and the precipitate was then separated by filtration, washed with EtOH (2 × 2 mL), and dried under vacuum for several hours. Yield: 0.1198 g light brown solid (44%). (For C) 46 H 40 Elemental analysis of Cu₂N₄O₈ (903.94): Calculated values ​​(%): C 61.12, H 4.46, N 6.20. Measured values: C 61.15, H 4.57, N 5.86. IR-ATR (cm⁻¹) −1): 3002, 2936, 2839, 1592, 1520, 1485, 1439, 1314, 1217, 1117, 1025, 975, 832.

[0238] 3.10 Synthesis of [Cu2(μ-(5-OMe)salben)2]

[0239]

[0240] Et3N (54.5 mg, 0.538 mmol) was added to a yellow suspension of H2(5-OMe)salben (0.1005 g, 0.2575 mmol) in 4 mL of EtOH. Then, solid Cu(OAc)2·H2O (0.05171 g, 0.2590 mmol) was added, and the color turned brown within minutes. The reaction mixture was placed at RT with magnetic stirring for 3 h, and the precipitate was then separated by filtration, washed with EtOH (2 × 2 mL), and dried under vacuum for several hours. Yield: 0.08951 g of brown solid (77%). (For C) 46 H 40 Elemental analysis of Cu₂N₄O₈ (903.94): Calculated values ​​(%): C 61.12, H 4.46, N 6.20. Measured values: C 60.97, H 4.43, N 6.13. IR-ATR (cm⁻¹) −1 (): 3062, 2928, 2827, 1623, 1592, 1532, 1456, 1307, 1215, 1157, 1026, 975, 813, 700. Mass spectrometry (ESI) + In MeCN, m / z (intensity %) is defined for [M + H]. + Calculated value: 903.1517; Measured value: 903.1331 (100) The method described above is used to prepare trispecific antibodies.

[0241] 3.11 Synthesis of [Cu2(μ-(5-F)salben)2]

[0242]

[0243] Et3N (57.4 mg, 0.57 mmol) was added to a yellow suspension of H2(5-F)salben (0.1004 g, 0.2742 mmol) in 4 mL of EtOH. Then, solid Cu(OAc)2·H2O (0.0564 g, 0.282 mmol) was added, and the color turned brown within minutes. The reaction mixture was placed at RT with magnetic stirring for 3 h, and the precipitate was then separated by filtration, washed with EtOH (2 × 2 mL), and dried under vacuum for several hours. Yield: 0.0945 g of brown solid (81%). (For C) 42 H 28 Elemental analysis of Cu₂F₄N₄O₄ (855.80): Calculated values ​​(%): C 58.95, H 3.30, N 6.53. Measured values: C 58.91, H 3.17, N 6.53. IR-ATR (cm⁻¹) −1 ): 3053, 3004, 2897, 1607, 1538, 1457, 1393, 1310, 1241, 1209, 1141, 1002, 925, 865, 812.

[0244] 3.12 Synthesis of [Cu2(μ-(5-Cl)salben)2]

[0245]

[0246] Et3N (105.3 mg, 1.04 mmol) was added to a yellow suspension of H2(5-Cl)salben (0.2015 g, 0.5048 mmol) in 8 mL of EtOH. Then, solid Cu(OAc)2·H2O (0.10236 g, 0.5127 mmol) was added, and the color turned brown within minutes. The reaction mixture was placed at RT with magnetic stirring for 3 h, and the precipitate was then separated by filtration, washed with EtOH (2 × 2 mL), and dried under vacuum for several hours. Yield: 0.2074 g of brown solid (89%). (For C) 42 H 28 Elemental analysis of Cu₂Cl₄N₄O₄ (921.60): Calculated values ​​(%): C 54.34, H 3.06, N 6.08. Measured values: C 54.37, H 3.03, N 5.99. IR-ATR (cm⁻¹) −1(): 3061, 2899, 1609, 1521, 1449, 1309, 1172, 1099, 976, 941, 876, 772. Mass spectrometry (ESI) + In MeCN, m / z (intensity %) is defined for [M + H]. + Calculated value: 942.9325; Measured value: 942.9382 (100) The method described above is used to prepare trispecific antibodies.

[0247] 3.13 Synthesis of [Cu2(μ-(5-Me)salben)2]

[0248]

[0249] Et3N (61.7 mg, 0.609 mmol) was added to a yellow suspension of H2(5-Me)salben (0.1019 g, 0.2843 mmol) in 4 mL of EtOH. Then, solid Cu(OAc)2·H2O (0.0568 g, 0.284 mmol) was added, and the color turned brown within minutes. The reaction mixture was placed at RT with magnetic stirring for 3 h, and the precipitate was then separated by filtration, washed with EtOH (2 × 2 mL), and dried under vacuum for several hours. Yield: 0.1001 g of brown solid (84%). (For C) 46 H 40 Elemental analysis of Cu₂N₄O₄ (839.94): Calculated values ​​(%): C 65.78, H 4.80, N 6.67. Measured values: C 65.78, H 4.75, N 6.64. IR-ATR (cm⁻¹) −1 (): 3059, 3004, 2911, 2853, 1619, 1529, 1468, 1382, 1313, 1251, 1137, 1045, 974, 824, 759. Mass spectrometry (ESI) + In MeCN, m / z (intensity %) is defined for [M + H]. + Calculated value: 839.1720; Measured value: 839.1714 (100) The method described is used to prepare trispecific antibodies.

[0250] 3.14 Synthesis of [Cu2(μ-(4-OMe)sal(2-Cl)ben)2]

[0251]

[0252] Et3N (54.5 mg, 0.538 mmol) was added to a yellow suspension of H2(4-OMe)sal(2-Cl)ben (0.1099 g, 0.2587 mmol) in 4 mL of EtOH. Then, solid Cu(OAc)2·H2O (0.05256 g, 0.2633 mmol) was added, and the color turned brown within minutes. The reaction mixture was placed at RT with magnetic stirring for 3 h, and the precipitate was then separated by filtration, washed with EtOH (2 × 2 mL), and dried under vacuum for several hours. Yield: 0.0672 g of brown solid (52%). (For C) 46 H 38 Elemental analysis of Cl₂Cu₂N₄O₈·H₂O (990.84): Calculated values ​​(%): C 55.76, H 4.07, N 5.65. Measured values: C 55.85, H 3.85, N 5.45. IR-ATR (cm⁻¹) −1 ): 3059, 3004, 2911, 2853, 1619, 1595, 1529, 1468, 1382, 1313, 1251, 1215, 1163, 1137, 1045, 974, 824, 759.

[0253] 3.15 Synthesis of [Cu2(μ-(5-OMe)sal(2-Cl)ben)2]

[0254]

[0255] Et3N (36.4 mg, 0.359 mmol) was added to a yellow suspension of H2(5-OMe)sal(2-Cl)ben (0.7146 g, 0.1685 mmol) in 3 mL of EtOH. Then, solid Cu(OAc)2·H2O (0.0330 g, 0.165 mmol) was added, and the color turned brown within minutes. The reaction mixture was placed at RT with magnetic stirring for 3 h, and the precipitate was then separated by filtration, washed with EtOH (2 × 2 mL), and dried under vacuum for several hours. Yield: 0.07628 g of brown solid (95%). (For C) 46 H 38 Elemental analysis of Cl₂Cu₂N₄O₈ (972.82): Calculated values ​​(%): C 56.79, H 3.94, N 5.76. Measured values: C 56.85, H 4.00, N 5.73. IR-ATR (cm⁻¹) −1): 2831, 1624, 1594, 1532, 1470, 1307, 1218, 1159, 1031, 973, 952, 824, 755.

[0256] 3.16 Synthesis of [Cu2(μ-(5-OMe)sal(2-OMe)ben)2]

[0257]

[0258] Et3N (98.0 mg, 0.969 mmol) was added to a yellow suspension of H2(5-OMe)sal(2-OMe)ben (0.2004 g, 0.4763 mmol) in 8 mL of EtOH. Then, solid Cu(OAc)2·H2O (0.0951 g, 0.0968 mmol) was added, and the color turned brown within minutes. The reaction mixture was placed at RT with magnetic stirring for 3 h, and the precipitate was then separated by filtration, washed with EtOH (2 × 2 mL), and dried under vacuum for several hours. Yield: 0.2091 g of brown solid (91%). (For C) 48 H 44 Cu2N4O 10 Elemental analysis of (963.99): Calculated values ​​(%): C 59.81, H 4.60, N 5.81. Measured values: C 60.28, H 4.51, N 5.82. IR-ATR (cm −1 (): 3053, 2832, 1622, 1597, 1533, 1459, 1392, 1306, 1246, 1219, 1158, 1207, 953, 824, 754. Mass spectrometry (ESI) + In MeCN, m / z (intensity %) is defined for [M + H]. + Calculated value: 963.1728; Measured value: 963.1677 (100) The method described is used to prepare trispecific antibodies.

[0259] 3.17 Synthesis of [Cu2(μ-(5-Cl)sal(2-OMe)ben)2]

[0260]

[0261] Et3N (98.0 mg, 0.969 mmol) was added to a yellow suspension of H2(5-Cl)sal(2-OMe)ben (0.1929 g, 0.1685 mmol) in 8 mL of EtOH. Then, solid Cu(OAc)2·H2O (0.0940 g, 0.471 mmol) was added, and the color turned brown within minutes. The reaction mixture was placed at RT with magnetic stirring for 3 h, and the precipitate was then separated by filtration, washed with EtOH (2 × 2 mL), and dried under vacuum for several hours. Yield: 0.2088 g of brown solid (94%). (For C) 44 H 32 Elemental analysis of Cl4Cu2N4O6 (981.65): Calculated values ​​(%): C 53.84, H 3.29, N 5.71. Measured values: C 53.74, H 3.26, N 5.60. IR-ATR (cm⁻¹) −1 ): 2960, 2839,1611, 1520, 1483, 1457, 1311, 1241, 1172, 1043, 982, 823, 740.

[0262] 3.18 Synthesis of [Cu2(μ-(5-Cl)sal(2-NO2)ben)2]

[0263]

[0264] Et3N (94.4 mg, 0.933 mmol) was added to a yellow suspension of H2(5-Cl)sal(2-NO2)ben (0.2046 g, 0.460 mmol) in 8 mL of EtOH. Then, solid Cu(OAc)2·H2O (0.0898 g, 0.450 mmol) was added, and the color turned brown within minutes. The reaction mixture was placed at RT with magnetic stirring for 3 h, and the precipitate was then separated by filtration, washed with EtOH (2 × 2 mL), and dried under vacuum for several hours. Yield: 0.2123 g of brown solid (93%). (For C) 42 H 26 Elemental analysis of Cl4Cu2N6O8 (1011.60): Calculated values ​​(%): C 49.87, H 2.59, N 8.31. Measured values: C 49.63, H 2.55, N 8.00. IR-ATR (cm⁻¹) −1): 1610, 1520, 1384, 1311, 1175, 1036, 978, 829, 765.

[0265] 3.19 Synthesis of [Cu2(μ-(5-OMe)sal(2-NO2)ben)2]

[0266]

[0267] Et3N (69.0 mg, 0.682 mmol) was added to a yellow suspension of H2(5-OMe)sal(2-NO2)ben (0.1435 g, 0.330 mmol) in 5.5 mL of EtOH. Then, solid Cu(OAc)2·H2O (0.0665 g, 0.333 mmol) was added, and the color turned brown within minutes. The reaction mixture was placed at RT with magnetic stirring for 3 h, and the precipitate was then separated by filtration, washed with EtOH (2 × 2 mL), and dried under vacuum for several hours. Yield: 0.141 g of brown solid (82%). (For C) 46 H 38 Cu2N6O 12 Elemental analysis of EtOH (1040.00): Calculated values ​​(%): C 55.44, H 4.29, N 8.08. Measured values: C 55.06, H 3.94, N 7.81. IR-ATR (cm⁻¹) −1 ): 2853,1625, 1596, 1525, 1467, 1339, 1302, 1271, 1218, 1160, 1032, 954, 811, 766.

[0268] 3.20 Synthesis of [Cu2(μ-(sal(2-NO2)ben)2]

[0269]

[0270] Et3N (58.1 mg, 0.574 mmol) was added to a yellow suspension of H2sal(2-NO2)ben (0.100 g, 0.266 mmol) in 4 mL of EtOH. Then, solid Cu(OAc)2·H2O (0.050 g, 0.275 mmol) was added, and the color turned brown within minutes. The reaction mixture was placed at RT with magnetic stirring for 3 h, and the precipitate was then separated by filtration, washed with EtOH (2 × 2 mL), and dried under vacuum for 1 h. Yield: 0.1022 g of brown solid (43.9%). (For C) 42 H 30 Elemental analysis of Cu2N6O8 (873.82): Calculated values ​​(%): C 57.73, H 3.46, N 9.62, O 14.65. Measured values: C 57.5, H 3.6, N 9.6, O 14.7. IR-ATR (cm⁻¹) −1 (): 3039, 3021, 2940, 2904, 1605, 1522, 1464, 1443, 1339, 1319, 1194, 1147, 1126, 1027, 979, 851, 751, 741, 713. Mass spectrometry (ESI) + In MeCN, m / z (intensity %): for [M + CH3CN + NH4] + ] + Calculated value: 931.1327; Measured value: 931.1322 (100) The method described is used to prepare trispecific antibodies.

[0271] 4. CN Coupling Reaction

[0272] 4.1 Coupling of iodobenzene and pyrazole using Cs2CO3 as a base

[0273] Under an inert atmosphere, iodobenzene (215 µL, 1.93 mmol, 1.5 equiv) and pyrrole (90 µL, 1.29 mmol, 1 equiv) were added to Cs₂CO₃ (839.2 mg, 2.58 mmol, 2 equiv) and [Cu₂(μ-salben)₂] (5 mg, 6.45 × 10⁻⁶) in a 10 mL tube with a threaded cap, along with pyrrole (90 µL, 1.29 mmol, 1 equiv). -3mmol (0.005 equiv) in a suspension in 1 mL MeCN. The tube was sealed and heated in an oil bath at 82 °C while stirring the reaction mixture for 16 h. The suspension was cooled to RT and then the solvent was removed under reduced pressure. The resulting residue was treated with 10 mL dichloromethane (DCM), the resulting suspension was filtered through filter paper, and the remaining solids were washed with DCM (2 × 5 mL). The organic phase was concentrated under reduced pressure to obtain a crude product as a brown liquid, which was purified by rapid column chromatography on silica gel to give 153 mg of the desired product (83% yield). 1 The H NMR spectrum was compared with commercially available samples to confirm the identity of the product. 1 H NMR, 298 K, CDCl3, 400 MHz: δ (ppm) 7.45-7.38 (4H, m), 7.27-7.23 (1H,m), 7.09 (2H, t), 6.35 (2H, t).

[0274] 4.2 Coupling of iodobenzene and pyrazole using K3PO4 as a base

[0275] Under an inert atmosphere, iodobenzene (720 µL, 6.47 mmol, 1.5 equiv) and pyrrole (300 µL, 4.29 mmol.1 equiv) were added to a suspension of K3PO4 (17.2 mmol, 3.65 g, 4 equiv) and [Cu2(μ-salben)2] (0.0429 mmol, 33.6 mg, 0.010 equiv) in 3 mL MeCN in a 50 mL tube with a threaded cap. The tube was sealed and heated in an oil bath at 82 °C, and the reaction mixture was stirred for 16 h. The suspension was cooled to RT and then the solvent was removed under reduced pressure. The resulting residue was treated with 10 mL DCM, the resulting suspension was filtered through filter paper, and the remaining solids were washed with DCM (2 × 5 mL). The organic phase was concentrated under reduced pressure to obtain a crude product as a brown liquid. The residual oil was purified on silica gel by rapid column chromatography to obtain 521 mg of the desired product (85% yield). By... 1 The H NMR spectrum was compared with commercially available samples to confirm the identity of the product. 1HNMR, 298 K, CDCl3, 400 MHz: δ (ppm) 7.45-7.38 (4H, m), 7.27-7.23 (1H, m), 7.09(2H, t), 6.35 (2H, t).

[0276] 4.3 General Procedures for Screening Solvents and Bases

[0277] Under an inert atmosphere, the base and catalyst were loaded into a 10 mL tube. Solvent (1 mL) was added, along with iodobenzene and pyrrole, to the resulting suspension. The tube was sealed and heated at the indicated temperature. After the indicated time, the tube was cooled to RT. The suspension was filtered through a 0.45 µm PTFE filter using a syringe. The solids on the filter were washed with the reaction solvent (3 × 3 mL). The filtrate was concentrated under reduced pressure to obtain the crude product. An external standard solution (0.429 mol / L dimethyl sulfone in 1 mL) was added. −1 (CDCl3 solution), 150 µL of the solution thus obtained was diluted to 700 µL with CDCl3 in an NMR tube and recorded. 1 H NMR spectra were used to calculate the yield.

[0278] 4.4 General Procedure for Coupling Aryl Halides with N-Nucleophiles

[0279] Under an inert atmosphere, the base, catalyst, and solvent are placed into a threaded-cap glass tube equipped with a stir bar. An N-nucleophile and an aryl halide are added to the suspension. The tube is sealed and heated at the indicated temperature for the indicated time. The reaction mixture is filtered through a 0.45 µm PTFE syringe filter and the filter is washed with MeCN (1–2 mL). LC-MS samples are prepared from the resulting solution. Optionally, the solvent is evaporated, and the crude product is purified by chromatography for further characterization.

[0280] 4.5 Characterization data of the product

[0281] .

Claims

1. A method for coupling an aryl halide with an N-nucleophile compound in the presence of a dinuclear copper(II) complex of formula I with a substituted salben-type ligand. Where R 1 and R 2 For the same or different, and for C that is optionally substituted 1-12 -alkyl, aryl or heteroaryl, or hydrogen, and G 1 G 2 G 3 and G 4 Whether the same or different, and whether it is hydrogen or selected from C 1-12 -alkyl, C 1-12 -alkoxy group, halogen C 1-12 -alkyl, halogen, mono- or di-C 1-12 -alkylamino, carboxyl, C 1-12 One or more substituents of alkoxycarbonyl or nitro, or G 1 G 2 G 3 and G 4 Each ring independently forms a fused aryl ring with two or three rings together with the phenyl ring to which it is attached, wherein the two or three rings are optionally selected from C 1-6 -alkyl, C 1-6 -alkoxy, halogen, mono- or di-C 1-6 -One or more substituents of alkylamino or nitro groups.

2. The method according to claim 1, wherein R 1 and R 2 Substituents of the same type and of formula IIa, IIb or IIc in R 3 R 4 R 5 R 6 and R 7 For the same or different, and for hydrogen, C 1-12 -alkyl, C 1-12 -alkoxy groups, halogens, halogen-C 1-12 -alkyl, mono- or di-C 1-12 -alkylamino, nitro, or R 3 and R 4 Or R 4 and R 5 Or R 5 and R 6 Or R 6 and R 7 Together with the phenyl rings they are attached to, they form a fused aryl ring having two or three rings, said two or three rings optionally selected from C 1-6 -alkyl, C 1-6 -alkoxy, halogen, mono- or di-C 1-6 -One or more substituents of alkylamino or nitro groups are used for substitution. X 1 X 2 X 3 X 4 and X 5 They may be the same or different, and represent at least one heteroatom selected from nitrogen, oxygen or sulfur; n is 0 or 1; R 8 R 9 and R 10 For the same or different, and for hydrogen, C 1-12 -alkyl, halogen C 1-12 -alkyl, hydroxyl, hydroxy-C 1-12 -alkyl, halogen, or R 8 and R 9 Together they form saturated carbon rings or heterocycles, or R 8 R 9 and R 10 Together they form an unsaturated carbon ring.

3. The method according to claim 1 or 2, wherein the aryl halide has formula III in X 6 X 7 X 8 X 9 and X 10 They may be the same or different, and represent carbon or at least one heteroatom selected from nitrogen, oxygen or sulfur; R 19 R 20 R 21 R 22 and R 23 For the same or different, and for hydrogen, C 1-12 -alkyl, C 2-12 -Alkenyl, C 2-12 -Alynyl group, C 1-12 -alkoxy groups, halogens, halogen-C 1-12 -alkyl, mono- or di-C 1-12 -alkylamino or cyano, or R 19 and R 20 Or R 20 and R 21 Or R 21 and R 22 Or R 22 and R 23 Together with the rings they are connected to, they form a fused aryl ring having two or three rings, said two or three rings optionally selected from C 1-6 -alkyl, C 1-6 -alkoxy, halogen, mono- or di-C 1-6 -One or more substituents of alkylamino or nitro groups are used; n is 0 or 1, and Z represents a halogen atom.

4. The method according to any one of claims 1 to 3, wherein the N-nucleophile may be selected from compounds of the following formula: in R 11 R 12 R 13 R 14 R 15 R 16 R 17 and R 18 For the same or different, and for hydrogen, C 1-12 -alkyl, C 1-12 -alkoxy, aryl, halogen, halogen-C 1-12 -alkyl, nitro, hydroxyl, mono- or di-C 1-12 -alkylamino, or R 15 and R 16 Together with the nitrogen atoms they are attached to, they form saturated heterocyclic 5- or 6-membered heterocycles; X 11 X 12 X 13 and X 14 For the same or different, and indicating optional C 1-12 -alkyl, aryl-C 1-12 -alkyl, C 1-6 -alkoxy groups, halogens, halogen-C 1-12 - An alkyl-substituted carbon atom and at least one other heteroatom selected from nitrogen, oxygen or sulfur.

5. The method according to any one of claims 1 to 4, wherein a base is present, said base being selected from alkali metal salts of inorganic or organic acids or from alkali metal alkoxides.

6. The method according to any one of claims 1 to 5, wherein the reaction is carried out in the presence of an organic solvent at a reaction temperature between 20°C and 200°C.

7. A binuclear copper(II) complex having a substituted salben-type ligand of Formula I in R 1 and R 2 For the same or different, and for C that is optionally substituted 1-12 -alkyl, aryl or heteroaryl, or hydrogen, and G 1 G 2 G 3 and G 4 For the same or different, and representing hydrogen or selected from C 1-12 -alkyl, C 1-12 -alkoxy groups, halogens, halogen C 1-12 -alkyl, mono- or di-C 1-12 -alkylamino, carboxyl, C 1-12 One or more substituents of -alkoxycarbonyl or nitro, or G 1 G 2 G 3 and G 4 Each ring independently forms a fused aryl ring with two or three rings together with the phenyl ring to which it is attached, wherein the two or three rings are optionally selected from C 1-6 -alkyl, C 1-6 -alkoxy, halogen, mono- or di-C 1-6 -One or more substituents of alkylamino or nitro groups are used; R 1 and R 2 For the same, and for substituents of formula IIa, IIb or IIc R 3 R 4 R 5 R 6 and R 7 For the same or different, and for hydrogen, C 1-12 -alkyl, C 1-12 -alkoxy groups, halogens, halogen-C 1-12 -alkyl, mono- or di-C 1-12 -alkylamino, nitro, or R 3 and R 4 Or R 4 and R 5 Or R 5 and R 6 Or R 6 and R 7 Together with the phenyl rings they are attached to, they form a fused aryl ring having two or three rings, said two or three rings optionally selected from C 1-6 -alkyl, C 1-6 -alkoxy, halogen, mono- or di-C 1-6 -One or more substituents of alkylamino or nitro groups are used; X 1 X 2 X 3 X 4 and X 5 They may be the same or different, and represent at least one heteroatom selected from nitrogen, oxygen or sulfur; n is 0 or 1; R 8 R 9 and R 10 For the same or different, and for hydrogen, C 1-12 -alkyl, halogen C 1-12 -alkyl, hydroxyl, hydroxy-C 1-12 -alkyl, halogen, or R 8 and R 9 Together they form saturated carbon rings or heterocycles, or R 8 R 9 and R 10 Together they form unsaturated carbon rings. The condition is to exclude R. 1 and R 2 It is hydrogen, or R 1 and R 2 Having the formula IIa and wherein R 3 R 4 R 6 and R 7 It is hydrogen, and R 5 Compounds that are hydrogen, chlorine, methyl, or nitro groups.

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