A method for C-C bond coupling in a DNA-encoded compound library

Abstract

This invention relates to a method for constructing a DNA-encoded compound library. 3 -sp 3 The C-C bond coupling method involves the formation of On-DNA sp from primary amine compounds, aromatic aldehyde compounds, and On-DNA alkenyl compounds via the Mannich reaction and photo-redox reaction. 3 -sp 3 C-C bond coupling yields On-DNA sp. 3 -sp 3 C-C bond coupling compounds. This method features simple post-reaction processing, is environmentally friendly, and enables large-scale activation of amines as synthetic modules, making it suitable for the synthesis of DNA-encoded compound libraries using multi-well plates.

Description

Technical Field

[0001] This invention belongs to the field of organic synthesis technology, specifically relating to a method for constructing a DNA-encoded compound library. 3 -sp 3 The C / C bond coupling method involves the formation of On-DNA sp from primary amine compounds, aromatic aldehyde compounds, and On-DNA alkenyl compounds via the Mannich reaction and photo-redox reaction. 3 -sp 3 The method of C / C bond coupling yields On-DNA sp 3 -sp 3 CC-linked coupling compounds. Background Technology

[0002] In drug development, especially in the development of new drugs, high-throughput screening targeting biological targets is one of the main methods for rapidly obtaining lead compounds. However, traditional high-throughput screening based on single molecules is time-consuming, requires huge equipment investment, and has a limited number of compounds (millions). Furthermore, the construction of compound libraries requires decades of accumulation, which limits the efficiency and possibility of lead compound discovery. In recent years, DNA-encoded compound library technology (WO2005058479, WO2018166532, CN103882532) has emerged. This technology combines combinatorial chemistry and molecular biology techniques, adding a DNA tag to each compound at the molecular level. It can synthesize compound libraries of up to hundreds of millions in a very short time, becoming the trend of next-generation compound library screening technology. It has begun to be widely used in the pharmaceutical industry, producing many positive effects (Accounts of Chemical Research, 2014, 47, 1247-1255).

[0003] DNA-encoded compound libraries enable the rapid generation of giant compound libraries through combinatorial chemistry, and allow for high-throughput screening of lead compounds, making lead compound screening faster and more efficient than ever before. One of the challenges in constructing DNA-encoded compound libraries is the need for high-yield synthesis of chemically diverse small molecules on DNA. Since DNA requires specific conditions (solvent, pH, temperature, ion concentration) to maintain stability, and the On-DNA reactions used in the construction of DNA-encoded compound libraries also need to achieve high yields, the types of reagents, reaction types, and reaction conditions of chemical reactions performed on DNA (referred to as On-DNA reactions) directly affect the richness and selectivity of DNA-encoded compound libraries. Therefore, developing DNA-compatible chemical reactions has become a long-term exploration and research direction for DNA-encoded compound library technology, directly impacting the application and commercial value of DNA-encoded compound libraries.

[0004] Constructing sp in a DNA-encoded compound library 3 -sp 3 The coupling of C and C bonds is of great significance. Therefore, it is desirable to develop a new sp... suitable for high-volume operation of multi-well plates. 3 -sp 3 The C / C bond coupling method can increase the diversity of DNA-encoded compound libraries and further enhance the application value of DNA-encoded compound library technology. Summary of the Invention

[0005] The method of this invention can achieve On-DNA sp 3 -sp 3 The coupling of C and C bonds is simple to handle after the reaction, environmentally friendly, and can activate amines on a large scale as synthetic modules, making it suitable for the synthesis of DNA-encoded compound libraries using multi-well plates.

[0006] This invention provides a method for constructing a DNA-encoded compound library. 3 -sp 3 The method of C / C bond coupling is characterized by using primary amine compounds, aromatic aldehyde compounds, and On-DNA alkenyl compounds via Mannich reaction and photo-redox reaction to achieve sp... 3 -sp 3 CC-linked coupling yields On-DNA sp 3 -sp 3 CC-coupled products.

[0007] The primary amine compound has the structural formula NH2-R. 1 The aromatic aldehyde compound has the structural formula CHO-Ar 1 The structural formula of the On-DNA alkenyl compound is as follows:

[0008] Ar 1 Selected from groups with a molecular weight of less than 1000 that are directly bonded to the carbon atom of the aldehyde group;

[0009] R 1 Selected from groups with a molecular weight of less than 1000 that are directly bonded to an amino nitrogen atom;

[0010] R 2 Selected from groups with a molecular weight of less than 1000 that are directly linked to DNA;

[0011] R 3 R 4 Each group is independently selected from groups with a molecular weight of less than 1000 that are directly bonded to a carbon atom in a double bond;

[0012] The oligonucleotides in the DNA are 10–200 bp in length.

[0013] The DNA in the structural formula comprises single-stranded or double-stranded nucleotide chains obtained by polymerizing artificially modified and / or unmodified nucleotide monomers, which are bonded to R by one or more chemical bonds or groups. 2 Connected. When there is one chemical bond, it refers to the connection between DNA and R in the structural formula. 2 Directly connected; when there are multiple chemical bonds, it refers to the DNA and R in the structural formula. 2 They are linked by multiple chemical bonds, for example, DNA and R... 2 The amino groups of DNA are linked by a methylene group (-CH2-), i.e., by two chemical bonds; or DNA and R... 2 The amino group of DNA is linked by a carbonyl group (-CO-), which is also a connection between two chemical bonds; or DNA and R... 2 The amino group of DNA is linked by a methylene carbonyl group (-CH2CO-), which is also linked by three consecutive chemical bonds.

[0014] Preferably, DNA and R 2 The amino groups of DNA are linked by a carbonyl group (-CO-).

[0015] Preferred, Ar 1 The aromatic rings are 6-10 nucleotides or 5-10 nucleotides, and the aromatic rings and heteroaromatic rings can be independently converted by one or more R... 5 replace.

[0016] R 1 Selected from -C1~C 10 Alkyl, -C 0～4 Alkylene (3- to 10-membered carbon ring), -C 0～4 Alkylene-(3- to 10-membered heterocycles), -C 0～4 alkylene-(6-12 membered aromatic rings), -C 0～4 alkylene-(5-12 membered heteroaryl rings), -C 0～4 Alkylene-(5-12 membered spirocyclic), -C 0～4 Alkylene-(5-12 membered spiroheterocycle), -C 0～4 Alkylene-(6- to 10-membered bridged ring), -C 0～4 Alkylene (6- to 10-membered bridged heterocycle), wherein the alkyl, alkylene, carbocyclic, heterocyclic, aromatic, heteroaromatic, spirocyclic, spiroheterocyclic, bridged ring, or bridged heterocycle may be further optionally bounded by one, two, three, or four independent Rs. b replace.

[0017] R 2 Selected from aryl, 6-10 aryl, and 5-10 heteroaryl groups, wherein the aryl and heteroaryl groups may be further optionally coupled with one, two, three, or four independent R groups. 6replace.

[0018] R 3 R 4 Each group is independently selected from hydrogen, halogen, cyano, nitro, alkenyl, alkynyl, -C1 to C1. 10 Alkyl or halogen-substituted C1-C 10 Alkyl, 3-10 membered cycloalkyl, 3-10 membered heterocycloalkyl, 6-10 membered aryl, 5-10 membered heteroaryl, -OR c -NR d R e -OC(O)R a -C(O)OR a -NR a C(O)R b -NR a S(O)2R b -S(O)2R c -S(O)2NR d R e -C(O)R c -C(O)NR d R e The alkyl, alkenyl, alkynyl, cycloalkyl, heterocycloalkyl, aryl, and heteroaryl groups may be further optionally surrounded by one, two, three, or four independent R groups. 6 replace.

[0019] R 5 Selected from hydrogen, halogen, cyano, nitro, alkenyl, alkynyl, aldehyde, borate ester, -C1~C 10 Alkyl or halogen-substituted C1-C 10 Alkyl, 3-10 membered cycloalkyl, 3-10 membered heterocycloalkyl, 6-10 membered aryl, 5-12 membered heteroaryl, -OR c -NR d R e -OC(O)R a -C(O)OR a -NR a C(O)R b -NR a S(O)2R b -S(O)2R c -S(O)2NR d R e -C(O)R c -C(O)NR d R e The alkyl, alkenyl, alkynyl, cycloalkyl, heterocycloalkyl, aryl, and heteroaryl groups may be further optionally surrounded by one, two, three, or four independent R groups. 6 replace.

[0020] R b Selected from halogen, cyano, nitro, oxo, -C2~C 10 alkenyl, -C2~C 10 Alkyne group, aldehyde group, -C1~C 10 Alkyl or halogen-substituted C1-C 10 Alkyl or amino substituted C1-C 10 Alkyl, 3-10 membered cycloalkyl, 3-10 membered heterocycloalkyl, 6-12 membered aryl, 5-12 membered heteroaryl, -OR c -NR d R e -OC(O)R a -C(O)OR a -NR a C(O)R c -NR a C(O)OR c -S(O)2R c -S(O)2NR d R e -C(O)R c -C(O)NR d R e The alkyl, cycloalkyl, heterocycloalkyl, aryl, and heteroaryl groups may be optionally further distinguished by one, two, three, or four independent R groups. 6 replace.

[0021] R 6 Selected from hydrogen, halogen, cyano, nitro, oxo, amino, -NR a R c Aldehyde group, halogen-substituted aryl group, -C 1～6 Alkyl, -C 2～6 alkenyl, -C 2～6 Alkyne- and halogen-substituted C 1～6 Alkyl, halogen-substituted C 2～6 Alkenyl, halogen-substituted C 2～6 Alkyne group, -OR c -C(O)OR a .

[0022] R a R c R d R e Selected from hydrogen, -C1 to C 10 Alkyl or halogen-substituted C1-C 10 Alkyl, 3-10 membered cycloalkyl, 3-10 membered heterocycloalkyl, 6-12 membered aryl, 5-12 membered heteroaryl.

[0023] Furthermore, Ar 1Selected from: X is selected from NH, O, S; Y, X1, Y1, Y2 are selected from CH, N; the Ar 1 It can be further used by one or more R 5 replace.

[0024] More specifically, Ar 1 Selected from:

[0025] More specifically, R 1 Selected from:

[0026] More specifically, R 2 Selected from key,

[0027] More specifically, R 3 Selected from H and methyl.

[0028] More specifically, R 4 Selected from H,

[0029] More specifically, the structural formula of the On-DNA alkenyl compound is specifically selected from:

[0030] The method provided by this invention includes the following steps:

[0031]

[0032] Step 1: Add NH2-R 1 and CHO-Ar 1 It dissolves in a solvent and, under heating conditions, directly forms the compound shown in Formula I;

[0033] Step 2: Combine the compound shown in Formula I with... Dissolved in a solvent, protected by N2, and directly yielded by the compound shown in Formula II under the action of a metallic iridium catalyst, phosphate, and light.

[0034] Furthermore, the solvent is selected from one or a mixture of several of the following: acetonitrile, methanol, ethanol, DMF, DMA, DMSO, THF, water, inorganic salt buffer, organic acid buffer, and organic base buffer.

[0035] Furthermore, the reaction temperature in step 1 is 60℃~140℃, preferably 60℃, 80℃, or 100℃.

[0036] Furthermore, the reaction time of step 1 is 1 to 24 hours, preferably 2 hours, 4 hours, 8 hours, 12 hours, or 16 hours.

[0037] Furthermore, the NH2-R 1 The solution concentration is 0.1–0.8 M, and the CHO-Ar 1 The solution concentration is 0.1–0.8 M, preferably, the NH2-R 1 The solution concentration is 0.4M, and the CHO-Ar 1 The solution concentration is 0.4M.

[0038] Further, the volume ratio of the reaction reagents in step 1 is amine (NH2-R) 1 ) / aldehyde (CHO-Ar 1 The volume ratio of the reaction reagents in step 1 is 1 / 1 to 3 / 1, preferably, the volume ratio of the reagents is amine (NH2-R) to amine (NH2-R). 1 ) / aldehyde (CHO-Ar 1 = 1 / 1.

[0039] Further, the reaction temperature of step 2 is 20℃~100℃, preferably 25℃, 35℃, 45℃, 55℃, 65℃, 75℃, 85℃, or 95℃.

[0040] Furthermore, the reaction time of step 2 is 1 to 24 hours, preferably 3 hours, 6 hours, 9 hours, 12 hours, 15 hours, or 18 hours.

[0041] Further, the light intensity in step 2 is 5 to 13.6V, preferably 13.6V.

[0042] Further, the illumination wavelength in step 2 is 400-500nm, preferably 420nm or 450nm.

[0043] Furthermore, the aforementioned The molar equivalent of the compound represented by Formula I is 1, the molar equivalent of the compound represented by Formula I is 10 to 500, the molar equivalent of the iridium catalyst is 0.1 to 2, and the molar equivalent of the phosphate is 0 to 500; preferably, the molar equivalent of the compound represented by Formula I is 120 or 240; preferably, the molar equivalent of the iridium catalyst is 0.5, 1, 1.5, or 2; preferably, the molar equivalent of the phosphate is 0, 60, 120, 240, or 480.

[0044] Further, the iridium metal catalyst is specifically selected from bis[2-(2,4-difluorophenyl)-5-trifluoromethylpyridine][2-2'-bipyridine]iridium di(hexafluorophosphate), bis[2-(2,4-difluorophenyl)-5-trifluoromethylpyridine][2-2'-bi(4-tert-butylpyridine)]iridium di(hexafluorophosphate), bis[2-(2,4-difluorophenyl)-5-methylpyridine][2,2'-bi(tetra-tert-butylpyridine)]iridium di(hexafluorophosphate), 4,4'-bis(1,1-dimethylethyl)-2,2'-bipyridyl-KN,KN]bis[5-fluoro-2-(5-methyl-2-pyridyl-KN)phenylKC]iridium hexafluorophosphate, One or more of (2,2'-bipyridine)bis(2-phenylpyridine)iridium(III) hexafluorophosphate and (4,4'-di-tert-butyl-2,2'-bipyridine)bis[(2-pyridyl)phenyl]iridium(III) hexafluorophosphate.

[0045] Furthermore, the phosphate is selected from one or more of potassium phosphate, potassium dihydrogen phosphate, and dipotassium hydrogen phosphate.

[0046] Furthermore, the method is used for batch operations of perforated plates.

[0047] Furthermore, the method is used for the synthesis of DNA-encoded compound libraries for multi-well plates.

[0048] This invention provides a method for constructing a DNA-encoded compound library. 3 -sp 3 The C-C bond coupling method utilizes primary amine compounds, aromatic aldehyde compounds, and On-DNA alkenyl compounds to achieve sp... 3 -sp 3 CC-linked coupling. This method is of great significance for the synthesis of DNA-encoded compound libraries. It is simple to process after the reaction, environmentally friendly, and can activate amines on a large scale as synthetic modules. It is suitable for the synthesis of DNA-encoded compound libraries using multi-well plates.

[0049] Regarding the definition of terms used in this invention: Unless otherwise stated, the initial definitions provided for groups or terms herein apply to the groups or terms used throughout this specification; for terms not specifically defined herein, the meanings that a person skilled in the art would give them should be given based on the disclosure and context.

[0050] The term "substitution" refers to the replacement of one or more hydrogen atoms in a molecule by other different atoms or molecules.

[0051] The minimum and maximum carbon atom content in hydrocarbon groups are indicated by a prefix, for example, the prefix (C a～ Cb Alkyl groups indicate any alkyl group containing one to two carbon atoms ("a" to "b"). Therefore, for example, C 1～ C 12 Alkyl groups are straight-chain or branched alkyl groups containing 1 to 12 carbon atoms.

[0052] The term "alkyl" refers to a saturated straight-chain or branched hydrocarbon group consisting of a carbon atom, such as methyl-CH3, ethyl-CH2CH3, methylene-CH2-, or isopropyl; the alkyl group can also be part of other groups, such as C 1～6 Alkoxy, C 1～6 Alkylamino.

[0053] The term "alkylene" refers to a saturated, straight-chain or branched, non-bridged divalent alkyl group containing 1 to 20 carbon atoms.

[0054] The term "halogen" refers to fluorine, chlorine, bromine, or iodine.

[0055] The term "carbocyclic" or "cycloalkyl" refers to a saturated or partially saturated cyclic group having a number of carbon atoms and no heterocyclic atoms, and having a single ring or multiple rings (including fused, bridged, and spirocyclic systems).

[0056] The term "heterocyclic" or "heterocyclic alkyl" refers to a non-aromatic cyclic group consisting of a carbon atom and a heteroatom selected from nitrogen, oxygen, sulfur, boron, silicon, etc., that is saturated or partially unsaturated (containing one or two double bonds). This cyclic group can be monocyclic or polycyclic.

[0057] The terms "aromatic ring" and "aryl" refer to aromatic single or multiple cyclic groups composed of carbon atoms that do not contain heteroatoms.

[0058] The term "heteroaromatic ring" or "heteroaryl" refers to a single or multiple cyclic aromatic ring groups formed by replacing a carbon atom on at least one ring with a heteroatom selected from nitrogen, oxygen, or sulfur, including monocyclic heterocyclic alkyl rings fused to a phenyl or heteroaryl group.

[0059] The term "spirocycle" refers to a spool of multiple carbon atoms that are either saturated or partially unsaturated (containing one or two double bonds) and do not contain cyclic heteroatoms. Examples of spirocycles include... Etc. A "spiroheterocycle" is a spool of carbon atoms and multiple rings (containing one or two double bonds) selected from heteroatoms such as nitrogen, oxygen, sulfur, boron, and silicon, either saturated or partially unsaturated. Examples include... wait.

[0060] The term "bridging ring" refers to multiple rings (containing one or two double bonds) that are saturated or partially unsaturated and composed of multiple carbon atoms without cyclic heteroatoms, and examples include... Adamantyl groups, etc., "bridged heterocycles" refer to multiple saturated or partially unsaturated rings (containing one or two double bonds) connected by carbon atoms and heteroatoms selected from nitrogen, oxygen, sulfur, boron, silicon, etc. Examples include wait.

[0061] The term "primary amine compounds" refers to NH3 in which one hydrogen atom is replaced by another non-hydrogen group.

[0062] The term "aromatic aldehydes" refers to compounds in which the aldehyde group is directly attached to an aryl or heteroaryl group.

[0063] Obviously, based on the above description of the present invention, and according to common technical knowledge and conventional methods in the field, various other modifications, substitutions or alterations can be made without departing from the basic technical concept of the present invention.

[0064] The following detailed embodiments further illustrate the above-described content of the present invention. However, this should not be construed as limiting the scope of the present invention to the following examples. All technologies implemented based on the above-described content of the present invention fall within the scope of the present invention. Attached image description:

[0065] Figure 1 The 27 On-DNA sps prepared in Example 2 3 -sp 3 Conversion rate distribution of CC-coupled compounds. Detailed Implementation

[0066] The technical solution of the present invention will now be clearly and completely described with reference to the accompanying drawings. Obviously, the described embodiments are only some, not all, of the embodiments of the present invention. All other embodiments obtained by those skilled in the art based on the embodiments of the present invention without creative effort are within the scope of protection of the present invention.

[0067] The raw materials and equipment used in this invention are all known products, obtained by purchasing commercially available products.

[0068] HATU: (2-(7-benzotriazole oxide)-N,N,N',N'-tetramethylurea hexafluorophosphate). DMSO: dimethyl sulfoxide. DMA: N,N-dimethylacetamide. DIPAE: N,N-diisopropylethylamine.

[0069] In this invention, DNA-NH2 refers to a DNA structure with a -NH2 linker formed by single-stranded or double-stranded DNA and a linker group, such as the DNA-NH2 structure of "compound1" in WO2005058479. Other examples include the following DNA structures:

[0070]

[0071] Where A is adenine, T is thymine, C is cytosine, and G is guanine.

[0072] Example 1. On-DNA sp 3 -sp 3 CC key coupling

[0073] (1) Synthesis of On-DNA alkenyl compounds

[0074] DNA-NH2 was dissolved in 250 mM borate buffer (pH 9.4) to prepare a 1 mM solution, which was then aliquoted into EP tubes and reacted with acrylic acid using HATU as a condensation activator to obtain the corresponding On-DNA alkenyl compound (Reference: ACS. Combinatorial. Science., 2016, 18, 8, 438-443). After the reaction was completed, only ethanol precipitation was performed, and the solution was concentrated, vacuumed, and directly used in the next step of the reaction.

[0075]

[0076] (2) Synthesis of Schiff bases

[0077] Primary amines react with aromatic aldehydes under heating conditions to form Schiff bases:

[0078]

[0079] 1-Adamantane and 2,4,6-trimethylbenzaldehyde were separately dissolved in DMSO to prepare 0.4 M solutions. The 1-adamantane and 2,4,6-trimethylbenzaldehyde solutions were mixed thoroughly at a volume ratio of 1:1 and heated in a metal bath at 80°C for 2 hours. The reaction was monitored by LCMS. After the reaction was completed, no post-treatment was performed, and the solution was immediately used for the next reaction.

[0080] (3) On-DNA sp 3 -sp 3 CC key coupling

[0081] On-DNA alkenyl compounds and Schiff bases are oxidized and reduced to C-C bonded compounds under the action of iridium catalysts, phosphates, and light irradiation.

[0082]

[0083] The On-DNA alkenyl compound obtained in (1) was dissolved in ultrapure water to prepare a 1 mM solution. Then, 4 μL of potassium phosphate solution (480 equivalents, 1.2 M ultrapure water), 1 μL of bis[2-(2,4-difluorophenyl)-5-trifluoromethylpyridine][2-2'-bipyridine]iridium di(hexafluorophosphate) salt solution (1 equivalent, 10 mM DMSO), and 12 μL of adamantane Schiff base compound solution obtained in (2) (240 equivalents, 0.2 M DMSO) were added to the solution (10 μL, 10 nmol) in sequence. Finally, 8 μL of DMSO was added to make the volume ratio of DMSO to water in the reaction system 3:2. After mixing evenly, the reaction was carried out under N2 protection and irradiated with an LED blue lamp (450 nm, 13.6 V) for 3 hours.

[0084] After the reaction was complete, ethanol precipitation was performed: 10% of the total volume of 5M sodium chloride solution was added to the reaction system, followed by 3 times the total volume of anhydrous ethanol. After shaking and mixing, the reaction was placed in dry ice and frozen for 2 hours. Then, it was centrifuged at 12,000 rpm for half an hour, the supernatant was discarded, and the precipitate was dissolved in deionized water to obtain a solution of the On-DNA compound. The solution was sent to LCMS to confirm that the conversion rate of the photo-redox reaction was 76%.

[0085] Example 2. On-DNA sp 3 -sp 3 Application of CC-key coupling in library construction

[0086] (1) Synthesis of On-DNA alkenyl compounds

[0087] DNA-NH2 was dissolved in 250mM borate buffer (pH 9.4) to prepare a 1mM solution, which was then aliquoted into EP tubes. The solution was then reacted with alkenyl compounds of different structures using HATU as a condensation activator to obtain the corresponding On-DNA alkenyl compounds (see Example 1). After the reaction was completed, only ethanol precipitation was performed, and the solution was concentrated and vacuumed before being used directly in the next reaction.

[0088]

[0089] (2) Synthesis of Schiff bases

[0090] Primary amines with different structures react with aromatic aldehydes under heating conditions to form Schiff bases:

[0091]

[0092] Primary amine compounds and aromatic aldehydes were separately dissolved in DMSO to prepare 0.4 M solutions. The primary amine and aromatic aldehyde solutions were mixed thoroughly at a volume ratio of 1:1 and heated in a metal bath at 80°C for 2 hours. The reaction was monitored by LCMS. After the reaction was completed, no post-processing was performed, and the solution was immediately used for the next reaction.

[0093] (3) On-DNA sp 3 -sp 3 CC key coupling

[0094] The different types of On-DNA alkenyl compounds obtained in reaction (1) were mixed evenly together, dissolved in ultrapure water, and prepared into a 1 mM concentration solution. The mixture was then evenly distributed into 96-well plates, and the different Schiff base compounds obtained in reaction (2) were oxidized and reduced to C-C bond-coupled compounds under the action of iridium catalyst, phosphate, and light irradiation.

[0095]

[0096] 4 μL of potassium phosphate solution (480 equivalents, 1.2 M ultrapure water), 1 μL of bis[2-(2,4-difluorophenyl)-5-trifluoromethylpyridine][2-2'-bipyridine]iridium di(hexafluorophosphate) salt solution (1 equivalent, 10 mM DMSO), and 12 μL of different Schiff base compounds obtained in (2) (240 equivalents, 0.2 M DMSO) were added sequentially to a mixture of On-DNA alkenyl groups (10 μL, 10 nmol) to make the volume ratio of DMSO to water in the reaction system 3:2. After mixing thoroughly, the mixture was protected with N2 and irradiated with a blue LED lamp (450 nm, 13.6 V) for 3 hours.

[0097] After the reaction was complete, ethanol precipitation was performed: 10% (5M) sodium chloride solution was added to the reaction mixture, followed by three times the total volume of anhydrous ethanol. The mixture was shaken thoroughly and then frozen on dry ice for 2 hours. Afterward, it was centrifuged at 4000 rpm for half an hour, and the supernatant was discarded to obtain On-DNA sp. 3 -sp 3 The CC-linked compound precipitated, and the precipitate was dissolved in deionized water to obtain a solution of the On-DNA compound. The conversion rate of the photo-redox reaction was confirmed by LCMS. The specific conversion rates of each compound are shown in [reference needed]. Figure 1 .

[0098] In summary, the above embodiments and accompanying drawings are merely preferred embodiments of the present invention and are not intended to limit the scope of protection of the present invention. Any modifications, equivalent substitutions, improvements, etc., made within the spirit and principles of the present invention should be within the scope of protection of the present invention.

Claims

1. A method for constructing a DNA-encoded compound library sp 3 -sp 3 The method of C / C bond coupling is characterized by... This method utilizes primary amine compounds, aromatic aldehyde compounds, and on-DNA alkenyl compounds via the Mannich reaction and photo-redox reaction to achieve sp. 3 -sp 3 CC-linked coupling yields On-DNA sp 3 -sp 3 CC-linked coupling compounds; the primary amine compounds have the structural formula NH2-R. 1 The aromatic aldehyde compound has the structural formula CHO-Ar 1 The structural formula of the On-DNA alkenyl compound is specifically selected from: , , , , , , , , , , , , , , , In the structural formula, the DNA comprises single-stranded or double-stranded nucleotide chains obtained by polymerizing artificially modified and / or unmodified nucleotide monomers; the Ar 1 The aromatic rings are 6-10 nucleotides or 5-10 nucleotides, and the aromatic rings and heterocyclic rings are independently bounded by one or more R... 5 Replaced or not replaced; R 1 Selected from -C1~C 10 Alkyl, -C 0~4 Alkylene rings (3- to 10-membered carbon rings), -C 0~4 Alkylene (3- to 10-membered heterocycles), -C 0~4 Alkylene-(6~12-membered aromatic ring), -C 0~4 Alkylene-(5~12-membered heteroaryl ring), -C 0~4 Alkylene-(5~12-membered spirocyclic), -C 0~4 Alkylene-(5-12 membered spiroheterocycle), -C 0~4 Alkylene-(6~10-membered bridged ring), -C 0~4 Alkylene (6-10-membered bridged heterocycle), wherein the alkyl, alkylene, carbocyclic, heterocyclic, aromatic, heteroaromatic, spirocyclic, spiroheterocyclic, bridged ring, or bridged heterocycle is further optionally bounded by one, two, three, or four independent Rs. b Replaced or not replaced; R 5 Selected from hydrogen, halogen, cyano, nitro, alkenyl, alkynyl, aldehyde, borate ester, -C1~C 10 Alkyl, 3-10 membered cycloalkyl, 3-10 membered heterocycloalkyl, 6-10 membered aryl, 5-12 membered heteroaryl, -OR c -NR d R e -OC(O)R a -C(O)OR a -NR a C(O)R b -NR a S(O)2R b -S(O)2R c -S(O)2NR d R e -C(O)R c -C(O)NR d R e The alkyl, alkenyl, alkynyl, cycloalkyl, heterocycloalkyl, aryl, and heteroaryl groups are further optionally separated by one, two, three, or four independent R groups. 6 Replaced or not replaced; R b Selected from halogen, cyano, nitro, oxo, -C2~C 10 alkenyl, -C2~C 10 Alkyne group, aldehyde group, -C1~C 10 Alkyl, 3-10 membered cycloalkyl, 3-10 membered heterocycloalkyl, 6-12 membered aryl, 5-12 membered heteroaryl, -OR c -NR d R e -OC(O)R a -C(O)OR a -NR a C(O)R c -NR a C(O)OR c -S(O)2R c -S(O)2NR d R e -C(O)R c -C(O)NR d R e The alkyl, cycloalkyl, heterocycloalkyl, aryl, and heteroaryl groups are further optionally separated by one, two, three, or four independent R groups. 6 Replaced or not replaced; R 6 Selected from hydrogen, halogen, cyano, oxo, nitro, -NR a R c Aldehyde group, -C 1~6 Alkyl, -C 2~6 alkenyl, -C 2~6 alkynyl group, -OR c -C(O)OR a , or R 6 Aryl groups substituted with halogens; R a R c R d R e Selected from hydrogen, -C1~C 10 Alkyl, 3-10 membered cycloalkyl, 3-10 membered heterocycloalkyl, 6-12 membered aryl, 5-12 membered heteroaryl; The method includes the following steps: Step 1: Add NH2-R 1 and CHO-Ar 1 Dissolving in DMSO and reacting at 80–100°C for 2–24 hours directly yields the compound shown in Formula I, wherein the volume ratio of the reactants is amine (NH2-R). 1 ) / aldehyde (CHO-Ar 1 ) = 1 / 1 ~ 3 / 1; Step 2: Combine the compound shown in Formula I with... Dissolving in an aqueous solvent, under N2 protection, and under the irradiation of a metallic iridium catalyst with a molar equivalent of 1-2, a phosphate with a molar equivalent of 480, a light intensity of 5-13.6 V, and a light wavelength of 450 nm, and at a reaction temperature of 20℃-100℃ and a reaction time of 3-24 hours, the compound shown in Formula II is directly generated. The molar equivalent is 1, and the molar equivalent of the compound shown in Formula I is 240 to 500.

2. The method according to claim 1, characterized in that: The Ar 1 Selected from ; The R 1 Selected from , , , , , , , , , , , , , , , , , , , , , , , , , , , , , , , , , , , , , , , , , , , , , , , , , , , , , , , , , , , .

3. The method according to claim 1, characterized in that: The iridium metal catalyst is selected from bis[2-(2,4-difluorophenyl)-5-trifluoromethylpyridine][2-2'-bipyridine]iridium di(hexafluorophosphate), bis[2-(2,4-difluorophenyl)-5-trifluoromethylpyridine][2-2'-bi(4-tert-butylpyridine)]iridium di(hexafluorophosphate), and bis[2-(2,4-difluorophenyl)-5-methylpyridine][2,2'-bi(tetra-tert-butylpyridine)]iridium di(hexafluorophosphate). Salt, one or more of 4,4'-bis(1,1-dimethylethyl)-2,2'-bipyridyl-KN,KN]bis[5-fluoro-2-(5-methyl-2-pyridyl-KN)phenylKC]iridium hexafluorophosphate, (2,2'-bipyridine)bis(2-phenylpyridine)iridium(III) hexafluorophosphate, and (4,4'-di-tert-butyl-2,2'-bipyridine)bis[(2-pyridyl)phenyl]iridium(III) hexafluorophosphate.

4. The method according to claim 1, characterized in that: The molar equivalents of the iridium metal catalyst are 1, 1.5, and 2.

5. The method according to claim 1, characterized in that: The phosphate is selected from one or more of potassium phosphate, potassium dihydrogen phosphate, and dipotassium hydrogen phosphate.

6. The method according to any one of claims 1 to 5, characterized in that: The method is used for batch operations of multi-hole plates.

7. The method according to any one of claims 1 to 5, characterized in that: The method is used for the synthesis of DNA-encoded compound libraries for multi-well plates.

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