CN113004361B - Method for synthesizing On-DNA pyrazole compound - Google Patents
Method for synthesizing On-DNA pyrazole compound Download PDFInfo
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- -1 pyrazole compound Chemical class 0.000 title claims abstract description 44
- 238000000034 method Methods 0.000 title claims abstract description 29
- 230000002194 synthesizing effect Effects 0.000 title claims abstract description 12
- 238000006243 chemical reaction Methods 0.000 claims abstract description 27
- 150000001875 compounds Chemical class 0.000 claims abstract description 25
- OAKJQQAXSVQMHS-UHFFFAOYSA-N hydrazine Substances NN OAKJQQAXSVQMHS-UHFFFAOYSA-N 0.000 claims abstract description 14
- 125000000217 alkyl group Chemical group 0.000 claims description 23
- 239000000243 solution Substances 0.000 claims description 19
- 125000001424 substituent group Chemical group 0.000 claims description 15
- 125000001997 phenyl group Chemical group [H]C1=C([H])C([H])=C(*)C([H])=C1[H] 0.000 claims description 14
- 239000000126 substance Substances 0.000 claims description 11
- 229910052736 halogen Inorganic materials 0.000 claims description 10
- 150000002367 halogens Chemical class 0.000 claims description 9
- 230000015572 biosynthetic process Effects 0.000 claims description 7
- 229910052739 hydrogen Inorganic materials 0.000 claims description 7
- 238000006467 substitution reaction Methods 0.000 claims description 7
- 238000003786 synthesis reaction Methods 0.000 claims description 7
- 239000001257 hydrogen Substances 0.000 claims description 6
- 125000000449 nitro group Chemical group [O-][N+](*)=O 0.000 claims description 5
- 125000003729 nucleotide group Chemical group 0.000 claims description 5
- 150000003217 pyrazoles Chemical class 0.000 claims description 5
- UFHFLCQGNIYNRP-UHFFFAOYSA-N Hydrogen Chemical compound [H][H] UFHFLCQGNIYNRP-UHFFFAOYSA-N 0.000 claims description 4
- 229910052799 carbon Inorganic materials 0.000 claims description 4
- BNIILDVGGAEEIG-UHFFFAOYSA-L disodium hydrogen phosphate Chemical compound [Na+].[Na+].OP([O-])([O-])=O BNIILDVGGAEEIG-UHFFFAOYSA-L 0.000 claims description 4
- 125000002887 hydroxy group Chemical group [H]O* 0.000 claims description 4
- 229910000403 monosodium phosphate Inorganic materials 0.000 claims description 4
- 235000019799 monosodium phosphate Nutrition 0.000 claims description 4
- AJPJDKMHJJGVTQ-UHFFFAOYSA-M sodium dihydrogen phosphate Chemical compound [Na+].OP(O)([O-])=O AJPJDKMHJJGVTQ-UHFFFAOYSA-M 0.000 claims description 4
- 125000004306 triazinyl group Chemical group 0.000 claims description 4
- 125000004989 dicarbonyl group Chemical group 0.000 claims description 3
- 150000002429 hydrazines Chemical class 0.000 claims description 3
- 239000002773 nucleotide Substances 0.000 claims description 3
- 239000002994 raw material Substances 0.000 claims description 3
- 125000004453 alkoxycarbonyl group Chemical group 0.000 claims description 2
- 239000000178 monomer Substances 0.000 claims description 2
- 230000035484 reaction time Effects 0.000 claims description 2
- 125000005843 halogen group Chemical group 0.000 claims 1
- 239000008055 phosphate buffer solution Substances 0.000 claims 1
- 230000000379 polymerizing effect Effects 0.000 claims 1
- XLYOFNOQVPJJNP-UHFFFAOYSA-N water Substances O XLYOFNOQVPJJNP-UHFFFAOYSA-N 0.000 abstract description 8
- 239000003153 chemical reaction reagent Substances 0.000 abstract description 3
- 229910052751 metal Inorganic materials 0.000 abstract description 2
- 239000002184 metal Substances 0.000 abstract description 2
- 239000003960 organic solvent Substances 0.000 abstract description 2
- 238000007363 ring formation reaction Methods 0.000 abstract 1
- 108020004414 DNA Proteins 0.000 description 32
- IAZDPXIOMUYVGZ-UHFFFAOYSA-N Dimethylsulphoxide Chemical compound CS(C)=O IAZDPXIOMUYVGZ-UHFFFAOYSA-N 0.000 description 13
- 239000007853 buffer solution Substances 0.000 description 7
- 239000000047 product Substances 0.000 description 7
- LFQSCWFLJHTTHZ-UHFFFAOYSA-N Ethanol Chemical compound CCO LFQSCWFLJHTTHZ-UHFFFAOYSA-N 0.000 description 6
- FAPWRFPIFSIZLT-UHFFFAOYSA-M Sodium chloride Chemical compound [Na+].[Cl-] FAPWRFPIFSIZLT-UHFFFAOYSA-M 0.000 description 6
- 125000003118 aryl group Chemical group 0.000 description 6
- 125000004432 carbon atom Chemical group C* 0.000 description 6
- 125000003545 alkoxy group Chemical group 0.000 description 5
- 125000004122 cyclic group Chemical group 0.000 description 5
- 125000006618 5- to 10-membered aromatic heterocyclic group Chemical group 0.000 description 4
- 150000002611 lead compounds Chemical class 0.000 description 4
- WEVYAHXRMPXWCK-UHFFFAOYSA-N Acetonitrile Chemical compound CC#N WEVYAHXRMPXWCK-UHFFFAOYSA-N 0.000 description 3
- CURLTUGMZLYLDI-UHFFFAOYSA-N Carbon dioxide Chemical compound O=C=O CURLTUGMZLYLDI-UHFFFAOYSA-N 0.000 description 3
- 102000004190 Enzymes Human genes 0.000 description 3
- 108090000790 Enzymes Proteins 0.000 description 3
- OKKJLVBELUTLKV-UHFFFAOYSA-N Methanol Chemical compound OC OKKJLVBELUTLKV-UHFFFAOYSA-N 0.000 description 3
- 125000005037 alkyl phenyl group Chemical group 0.000 description 3
- 235000011089 carbon dioxide Nutrition 0.000 description 3
- 239000008367 deionised water Substances 0.000 description 3
- 229910021641 deionized water Inorganic materials 0.000 description 3
- 238000012869 ethanol precipitation Methods 0.000 description 3
- 230000008014 freezing Effects 0.000 description 3
- 238000007710 freezing Methods 0.000 description 3
- 125000004435 hydrogen atom Chemical group [H]* 0.000 description 3
- 238000012917 library technology Methods 0.000 description 3
- 238000004895 liquid chromatography mass spectrometry Methods 0.000 description 3
- 239000002244 precipitate Substances 0.000 description 3
- 239000011780 sodium chloride Substances 0.000 description 3
- 125000000547 substituted alkyl group Chemical group 0.000 description 3
- 239000006228 supernatant Substances 0.000 description 3
- SXGZJKUKBWWHRA-UHFFFAOYSA-N 2-(N-morpholiniumyl)ethanesulfonate Chemical compound [O-]S(=O)(=O)CC[NH+]1CCOCC1 SXGZJKUKBWWHRA-UHFFFAOYSA-N 0.000 description 2
- 102000053602 DNA Human genes 0.000 description 2
- 125000003277 amino group Chemical group 0.000 description 2
- 125000004429 atom Chemical group 0.000 description 2
- 239000000872 buffer Substances 0.000 description 2
- 125000002915 carbonyl group Chemical group [*:2]C([*:1])=O 0.000 description 2
- 125000003178 carboxy group Chemical group [H]OC(*)=O 0.000 description 2
- 239000007810 chemical reaction solvent Substances 0.000 description 2
- 208000012839 conversion disease Diseases 0.000 description 2
- 125000000753 cycloalkyl group Chemical group 0.000 description 2
- OPTASPLRGRRNAP-UHFFFAOYSA-N cytosine Chemical compound NC=1C=CNC(=O)N=1 OPTASPLRGRRNAP-UHFFFAOYSA-N 0.000 description 2
- VILAVOFMIJHSJA-UHFFFAOYSA-N dicarbon monoxide Chemical group [C]=C=O VILAVOFMIJHSJA-UHFFFAOYSA-N 0.000 description 2
- 238000009509 drug development Methods 0.000 description 2
- UYTPUPDQBNUYGX-UHFFFAOYSA-N guanine Chemical compound O=C1NC(N)=NC2=C1N=CN2 UYTPUPDQBNUYGX-UHFFFAOYSA-N 0.000 description 2
- 125000005059 halophenyl group Chemical group 0.000 description 2
- 125000005842 heteroatom Chemical group 0.000 description 2
- 125000000623 heterocyclic group Chemical group 0.000 description 2
- 238000013537 high throughput screening Methods 0.000 description 2
- 125000001183 hydrocarbyl group Chemical group 0.000 description 2
- 125000005647 linker group Chemical group 0.000 description 2
- 239000000203 mixture Substances 0.000 description 2
- 238000011160 research Methods 0.000 description 2
- 238000012216 screening Methods 0.000 description 2
- 239000002904 solvent Substances 0.000 description 2
- RWQNBRDOKXIBIV-UHFFFAOYSA-N thymine Chemical group CC1=CNC(=O)NC1=O RWQNBRDOKXIBIV-UHFFFAOYSA-N 0.000 description 2
- 125000002023 trifluoromethyl group Chemical group FC(F)(F)* 0.000 description 2
- NWZSZGALRFJKBT-KNIFDHDWSA-N (2s)-2,6-diaminohexanoic acid;(2s)-2-hydroxybutanedioic acid Chemical compound OC(=O)[C@@H](O)CC(O)=O.NCCCC[C@H](N)C(O)=O NWZSZGALRFJKBT-KNIFDHDWSA-N 0.000 description 1
- WQDQLOXQQUFECI-UHFFFAOYSA-N 2-nitropropanedial Chemical compound [O-][N+](=O)C(C=O)C=O WQDQLOXQQUFECI-UHFFFAOYSA-N 0.000 description 1
- ZCYVEMRRCGMTRW-UHFFFAOYSA-N 7553-56-2 Chemical compound [I] ZCYVEMRRCGMTRW-UHFFFAOYSA-N 0.000 description 1
- 229930024421 Adenine Natural products 0.000 description 1
- GFFGJBXGBJISGV-UHFFFAOYSA-N Adenine Chemical compound NC1=NC=NC2=C1N=CN2 GFFGJBXGBJISGV-UHFFFAOYSA-N 0.000 description 1
- WKBOTKDWSSQWDR-UHFFFAOYSA-N Bromine atom Chemical compound [Br] WKBOTKDWSSQWDR-UHFFFAOYSA-N 0.000 description 1
- ZAMOUSCENKQFHK-UHFFFAOYSA-N Chlorine atom Chemical compound [Cl] ZAMOUSCENKQFHK-UHFFFAOYSA-N 0.000 description 1
- PXGOKWXKJXAPGV-UHFFFAOYSA-N Fluorine Chemical compound FF PXGOKWXKJXAPGV-UHFFFAOYSA-N 0.000 description 1
- 108020004682 Single-Stranded DNA Proteins 0.000 description 1
- QBJRFHRDZABKOX-UHFFFAOYSA-N [N]NN Chemical group [N]NN QBJRFHRDZABKOX-UHFFFAOYSA-N 0.000 description 1
- 238000009825 accumulation Methods 0.000 description 1
- 229960000643 adenine Drugs 0.000 description 1
- 150000001335 aliphatic alkanes Chemical class 0.000 description 1
- 239000003513 alkali Substances 0.000 description 1
- 125000003282 alkyl amino group Chemical group 0.000 description 1
- 230000004075 alteration Effects 0.000 description 1
- 125000006615 aromatic heterocyclic group Chemical group 0.000 description 1
- KGBXLFKZBHKPEV-UHFFFAOYSA-N boric acid Chemical compound OB(O)O KGBXLFKZBHKPEV-UHFFFAOYSA-N 0.000 description 1
- 239000004327 boric acid Substances 0.000 description 1
- GDTBXPJZTBHREO-UHFFFAOYSA-N bromine Substances BrBr GDTBXPJZTBHREO-UHFFFAOYSA-N 0.000 description 1
- 229910052794 bromium Inorganic materials 0.000 description 1
- 238000001311 chemical methods and process Methods 0.000 description 1
- 229910052801 chlorine Inorganic materials 0.000 description 1
- 239000000460 chlorine Substances 0.000 description 1
- 238000010276 construction Methods 0.000 description 1
- 125000004093 cyano group Chemical group *C#N 0.000 description 1
- 229940104302 cytosine Drugs 0.000 description 1
- 238000011161 development Methods 0.000 description 1
- 239000012154 double-distilled water Substances 0.000 description 1
- 238000005516 engineering process Methods 0.000 description 1
- 229910052731 fluorine Inorganic materials 0.000 description 1
- 239000011737 fluorine Substances 0.000 description 1
- 230000004907 flux Effects 0.000 description 1
- IKDUDTNKRLTJSI-UHFFFAOYSA-N hydrazine monohydrate Substances O.NN IKDUDTNKRLTJSI-UHFFFAOYSA-N 0.000 description 1
- 229910017053 inorganic salt Inorganic materials 0.000 description 1
- 239000011630 iodine Substances 0.000 description 1
- 229910052740 iodine Inorganic materials 0.000 description 1
- 150000002500 ions Chemical class 0.000 description 1
- 230000007774 longterm Effects 0.000 description 1
- 125000006357 methylene carbonyl group Chemical group [H]C([H])([*:1])C([*:2])=O 0.000 description 1
- 125000001570 methylene group Chemical group [H]C([H])([*:1])[*:2] 0.000 description 1
- GRVDJDISBSALJP-UHFFFAOYSA-N methyloxidanyl Chemical class [O]C GRVDJDISBSALJP-UHFFFAOYSA-N 0.000 description 1
- 239000012046 mixed solvent Substances 0.000 description 1
- 238000012986 modification Methods 0.000 description 1
- 230000004048 modification Effects 0.000 description 1
- 238000001823 molecular biology technique Methods 0.000 description 1
- 239000002547 new drug Substances 0.000 description 1
- 229910052757 nitrogen Inorganic materials 0.000 description 1
- 150000007524 organic acids Chemical class 0.000 description 1
- 229910052760 oxygen Inorganic materials 0.000 description 1
- 125000004430 oxygen atom Chemical group O* 0.000 description 1
- 238000006116 polymerization reaction Methods 0.000 description 1
- 230000008092 positive effect Effects 0.000 description 1
- 229920006395 saturated elastomer Polymers 0.000 description 1
- 150000003384 small molecules Chemical class 0.000 description 1
- 239000000758 substrate Substances 0.000 description 1
- 229910052717 sulfur Inorganic materials 0.000 description 1
- 238000001308 synthesis method Methods 0.000 description 1
- 229940113082 thymine Drugs 0.000 description 1
Classifications
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- C—CHEMISTRY; METALLURGY
- C07—ORGANIC CHEMISTRY
- C07H—SUGARS; DERIVATIVES THEREOF; NUCLEOSIDES; NUCLEOTIDES; NUCLEIC ACIDS
- C07H21/00—Compounds containing two or more mononucleotide units having separate phosphate or polyphosphate groups linked by saccharide radicals of nucleoside groups, e.g. nucleic acids
- C07H21/04—Compounds containing two or more mononucleotide units having separate phosphate or polyphosphate groups linked by saccharide radicals of nucleoside groups, e.g. nucleic acids with deoxyribosyl as saccharide radical
-
- C—CHEMISTRY; METALLURGY
- C07—ORGANIC CHEMISTRY
- C07H—SUGARS; DERIVATIVES THEREOF; NUCLEOSIDES; NUCLEOTIDES; NUCLEIC ACIDS
- C07H1/00—Processes for the preparation of sugar derivatives
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- Chemical & Material Sciences (AREA)
- Organic Chemistry (AREA)
- Health & Medical Sciences (AREA)
- Life Sciences & Earth Sciences (AREA)
- Biochemistry (AREA)
- Molecular Biology (AREA)
- Engineering & Computer Science (AREA)
- Biotechnology (AREA)
- General Health & Medical Sciences (AREA)
- Genetics & Genomics (AREA)
- Plural Heterocyclic Compounds (AREA)
Abstract
The invention relates to a method for synthesizing an On-DNA pyrazole compound, which is characterized in that the On-DNA pyrazole compound is obtained by ring closure reaction of an On-DNA hydrazine compound and a 1,3 dicarbonyl compound, the reaction method can be carried out in a mixed water phase of an organic solvent/water phase, the operation is simple, no metal reagent is introduced, the environment is friendly, and the method is suitable for synthesizing a DNA coding compound library by using a porous plate.
Description
Technical Field
The invention belongs to the technical field of coding compound libraries, and particularly relates to a method for synthesizing an On-DNA pyrazole compound in construction of a DNA coding compound library.
Background
In drug development, especially new drug development, high throughput screening against biological targets is one of the main means to rapidly obtain lead compounds. However, conventional high throughput screening based on single molecules requires long time, huge equipment investment, limited numbers of library compounds (millions), and the build-up of compound libraries requires decades of accumulation, limiting the efficiency and possibilities of discovery of lead compounds. The recent advent of DNA-encoded compound library technology (WO 2005058479, WO2018166532, CN 103882532), combining combinatorial chemistry and molecular biology techniques, tagged each compound with a DNA tag at the molecular level, and capable of synthesizing up to hundred million classes of compound libraries in extremely short time, has become a trend for the next generation of compound library screening technology, and began to be widely used in the pharmaceutical industry, producing a number of positive effects (Accounts of Chemical Research,2014,47,1247-1255).
The DNA encoding compound library rapidly generates a huge compound library by combinatorial chemistry, and can screen the lead compound with high flux, so that the screening of the lead compound becomes unprecedented rapid and efficient. One of the challenges in constructing libraries of DNA-encoding compounds is the need to synthesize small molecules with chemical diversity on DNA in high yields. Since DNA needs to be stable under certain conditions (solvent, pH, temperature, ion concentration), higher yields are also required for the On-DNA reaction constructed from DNA encoding compound libraries. Therefore, the kind of the reagent, the kind of the reaction and the reaction condition of the chemical reaction (called On-DNA reaction for short) performed On the DNA directly influence the richness and the selectivity of the DNA coding compound library. Thus, the development of chemical reactions compatible with DNA is also a long-term research and study direction of the current DNA coding compound library technology, and directly influences the application and commercial value of the DNA coding compound library.
Pyrazole compounds are an important pharmaceutical compound framework structure, however, no method for synthesizing On-DNA pyrazole compounds by On-DNA hydrazine compounds is reported at present. Therefore, a new synthesis method of On-DNA pyrazole compounds suitable for large-scale porous plate operation needs to be developed to increase the diversity of DNA coding compound libraries and further improve the application value of the DNA coding compound library technology.
Disclosure of Invention
The invention provides a method for synthesizing an On-DNA pyrazole compound, which has mild reaction conditions and simple post-treatment, is suitable for producing a DNA coding compound library, and can obviously improve the diversity of molecules of the compound library.
The invention provides a method for synthesizing an On-DNA pyrazole compound, which is characterized in that an On-DNA hydrazine compound and a 1,3 dicarbonyl compound are adoptedAs raw materials, the On-DNA product is obtained by reaction; wherein the structural formula of the On-DNA hydrazine compound isThe structural formula of the 1,3 dicarbonyl compound is +.>The structural formula of the On-DNA product is +.>
Wherein the DNA of the formula comprises a single-or double-stranded nucleotide chain obtained by polymerization of artificially modified and/or unmodified nucleotide monomers, which is linked to R by one or more chemical bonds or groups 1 Are connected; the length of the DNA is 10-200 bp.
Wherein, the DNA in the structural formula and R 1 Connected by a chemical bond or bonds. In the case of one chemical bond, it means DNA and R in the structural formula 1 Directly connected; in the case of multiple chemical bonds, the terms DNA and R in the structural formula 1 With multiple chemical bonds spaced apart, e.g. DNA and R 1 Through a methylene group (-CH) 2 (-) are connected, namely through two chemical bonds; or DNA and R 1 The amino group of DNA is connected with the carbonyl group (-CO-) through two chemical bonds; or DNA and R 1 Through a methylene carbonyl (-CH) 2 CO-) is linked to the amino group of the DNA, also via three consecutive chemical bonds.
R 1 A group having a molecular weight of 1000 or less and directly linked to a hydrazino nitrogen atom and DNA;
r' is selected from hydrogen or a group with the molecular weight below 1000 which is directly connected with carbon atom at alpha position of carbonyl;
R 2 selected from hydrogen or a group having a molecular weight of 1000 or less directly attached to a carbonyl carbon atom;
R 3 selected from hydrogen or a group having a molecular weight of 1000 or less and directly bonded to a carbonyl carbon atom.
As preferable: said R is 1 、R 2 、R 3 Respectively selected from alkyl, substituted alkyl, 5-10 membered aryl, substituted 5-10 membered aryl, 5-10 membered aromatic heterocyclic group and substituted 5-10 membered aromatic heterocyclic group; wherein the alkyl group is C 1 ～C 20 Alkyl or C 3 ～C 8 Cycloalkyl; the number of substituents for the substituted alkyl group is one or more; the substituent of the substituted alkyl is one or more of halogen, carboxyl, hydroxyl, nitro, alkoxy, halogenated phenyl, alkylphenyl and heterocyclic groups which are independent of each other; the number of the substituent groups for substituting the 5-10 membered aryl is one or more, and the substituent groups for substituting the 5-10 membered aryl are mutually independent halogen, cyano, hydroxyl, nitro, carboxyl, alkoxy and C 1 ～C 20 One or more of alkyl and trifluoromethyl; the number of the substituent groups for substituting the 5-10 membered aromatic heterocyclic group is one or more, and the substituent groups for substituting the 5-10 membered aromatic heterocyclic group are independently selected from halogen, cyano, nitro, carboxyl, alkoxy and C 1 ～C 20 One or more of alkyl and trifluoromethyl.
Said R' is selected from hydrogen, C 1 ～C 20 Alkoxycarbonyl, halogen, carboxyl, hydroxyl, nitro, C 1 ～C 20 One or more of alkoxy, halophenyl, phenyl, alkylphenyl, heterocyclyl.
Preferably, said R 1 Selected from phenyl, substituted phenyl, triazinyl, halogen substituted triazinyl, C 1 ～C 6 Alkyl, substituted C 1 ～C 6 An alkyl group; the number of substituents of the substituted phenyl is one or more, and the substituents of the substituted phenyl are independently selected from hydroxy, nitro, halogen and C 1 ～C 6 An alkyl group; substitution C 1 ～C 6 The number of substituents of the alkyl group is one or more; substitution C 1 ～C 6 The substituents of the alkyl groups are independently selected from phenyl groups.
Said R is 2 、R 3 Respectively selected from hydrogen or C 1 ～C 6 An alkyl group.
The On-DNA hydrazine compound is specifically selected from
The 1,3 dicarbonyl compound is specifically selected from
A method for synthesizing an On-DNA pyrazole compound, the reaction steps of the reaction being: adding 10-1000 times of molar equivalent of 1,3 dicarbonyl compound into On-DNA hydrazine compound solution with molar equivalent of 1 and molar concentration of 0.5-5mM, and reacting for 0.5-24 hours at 0-100 ℃.
The reaction is carried out in a solvent, and the reaction solvent is any one or a plurality of aqueous mixed solvents of water, methanol, ethanol, acetonitrile, dimethyl sulfoxide, inorganic salt buffer solution, organic acid buffer solution and organic alkali buffer solution.
Preferably, the reaction solvent contains 2- (N-morpholino) ethanesulfonic acid (MES) buffer or disodium hydrogen phosphate/sodium dihydrogen Phosphate (PBS) buffer; the pH of the buffer solution is 5.5-9.4; preferably, the pH is 5.5.
Further, the reaction temperature of the reaction is 20 ℃,30 ℃, 40 ℃,50 ℃,60 ℃, 70 ℃ or 80 ℃.
Further, the reaction time of the reaction is 1 hour, 4 hours, 8 hours, 10 hours, 16 hours, 18 hours or 20 hours.
Further, in the method, the molar equivalent of the On-DNA hydrazine compound is 10-1000, and the molar equivalent of the 1,3 dicarbonyl compound is 1; preferably, the molar equivalent of the 1,3 dicarbonyl compound is 50 equivalents, 100 equivalents, 200 equivalents, 300 equivalents, 400 equivalents, 500 equivalents, 600 equivalents, 800 equivalents, 1000 equivalents, most preferably, the molar equivalent of the 1,3 dicarbonyl compound is 100.
Further, the above method is used for batch multi-well plate operations.
Further, the above method is used for the synthesis of DNA encoding compound libraries in multiwell plates.
The method can obtain the On-DNA pyrazole compound from the DNA coding compound library through the On-DNA hydrazine compound, can be widely applied to various On-DNA hydrazine compound substrates, and can introduce various substituted 1,3 dicarbonyl compounds in a large scale as a synthesis module. The method has high yield and single product, can be carried out in a mixed water phase of an organic solvent/water phase, is simple to operate, does not introduce metal reagents, is environment-friendly, and is suitable for synthesizing the DNA coding compound library by using a porous plate.
Definition of terms used in connection with the present invention: unless otherwise indicated, the initial definitions provided for groups or terms herein apply to the groups or terms throughout the specification; for terms not specifically defined herein, the meanings that one skilled in the art can impart based on the disclosure and the context.
"substituted" means that a hydrogen atom in a molecule is replaced by a different atom or molecule.
The minimum and maximum values of the carbon atom content in the hydrocarbon group are represented by prefixes, for example, prefixes (Ca to C b ) Alkyl indicates any alkyl group containing from "a" to "b" carbon atoms. Thus, for example, C 1 ～C 12 Alkyl refers to straight or branched chain alkyl groups containing 1 to 12 carbon atoms.
Alkyl refers to straight or branched hydrocarbon groups in the alkane molecule, e.g. methyl-CH 3 ethyl-CH 2 CH 3 methylene-CH 2 -; the alkyl group may also be part of another group, such as C 1 ～C 6 Alkoxy, C 1 ～C 6 An alkylamino group.
Cycloalkyl refers to a saturated or partially saturated cyclic group having multiple carbon atoms and no ring heteroatoms, and having a single ring or multiple rings (including fused, bridged and spiro ring systems).
The halogen is fluorine, chlorine, bromine or iodine.
Alkoxy means that the alkyl group is attached to an oxygen atom to form a substituent, e.g. methylOxy is-OCH 3 。
Halo phenyl refers to a group formed by substitution of H on phenyl with halogen.
Alkylphenyl refers to a group formed by substitution of H on phenyl with alkyl.
Aryl refers to an aromatic single cyclic or multiple cyclic group consisting of C atoms without heteroatoms.
The aromatic heterocyclic group means a single cyclic group or a plurality of cyclic groups having aromaticity and composed of 5 to 10 atoms such as C, O, S, N.
It should be apparent that, in light of the foregoing, various modifications, substitutions and alterations can be made herein without departing from the spirit and scope of the invention as defined by the appended claims.
The above-described aspects of the present invention will be described in further detail below with reference to specific embodiments in the form of examples. It should not be understood that the scope of the above subject matter of the present invention is limited to the following examples only. All techniques implemented based on the above description of the invention are within the scope of the invention.
Drawings
Fig. 1: corresponding conversion profiles of 21 On-DNA pyrazole compounds prepared in example 2.
Detailed Description
The raw materials and equipment used in the invention are all known products and are obtained by purchasing commercial products.
DMSO: dimethyl sulfoxide;
DNA-NH in the present invention 2 Is formed by single-stranded or double-stranded DNA and a linker group and carries-NH 2 DNA structure of linker, e.g. DNA-NH of "component 1" in WO2005058479 2 Structure is as follows. Also for example the following DNA structure:
wherein A is adenine, T is thymine, C is cytosine, and G is guanine.
EXAMPLE 1 Synthesis of On-DNA pyrazole Compounds
Step 1, synthesis of On-DNA hydrazine compounds
On-DNA bromoacetic acid compound (1) was dissolved in 250mM boric acid buffer solution (pH=9.4) to prepare a 1mM concentration solution (20. Mu.L, 20 nmol), and hydrazine hydrate (10000 nmol,500 equivalent of 1M double distilled water) was added to the solution, and the mixture was uniformly mixed and reacted at 30℃for 2 hours.
Ethanol precipitation is carried out after the reaction is finished: adding 5M sodium chloride solution with the total volume of 10% into the reacted solution, continuously adding absolute ethanol with the total volume of 3 times, shaking uniformly, placing the reaction in dry ice, freezing for 0.5 hour, centrifuging for half an hour at the speed of 12000rpm, pouring out supernatant, dissolving the rest precipitate with deionized water to obtain the solution of the On-DNA hydrazino compound (2), quantifying by an enzyme-labeled instrument OD, and sending LCMS to confirm that the reaction conversion rate is 95%.
Step 2, synthesis of On-DNA pyrazole Compound
The On-DNA hydrazine compound (2) was redissolved in 500mM disodium hydrogen phosphate/sodium dihydrogen Phosphate (PBS) buffer solution at pH=5.5 to prepare a 1mM concentration solution (20. Mu.L, 20 nmol), and nitromalonaldehyde (2000 nmol,100 equivalents, 0.2M DMSO) was added to the solution, and the mixture was uniformly mixed and reacted at 60℃for 16 hours.
Ethanol precipitation is carried out after the reaction is finished: adding 5M sodium chloride solution with the total volume of 10% into the reacted solution, continuously adding absolute ethanol with the total volume of 3 times, shaking uniformly, placing the reaction in dry ice, freezing for 0.5 hour, centrifuging for half an hour at the speed of 12000rpm, pouring out supernatant, dissolving the rest precipitate with deionized water to obtain the solution of the On-DNA product, quantifying by an enzyme-labeled instrument OD, and sending LCMS to confirm that the reaction conversion rate is 85%.
EXAMPLE 2 Synthesis of On-DNA pyrazole Compounds
On-DNA hydrazines were dissolved in 250mM disodium hydrogen phosphate/sodium dihydrogen Phosphate (PBS) buffer solution at ph=5.5 to prepare a 2mM concentration solution (10 μl,20 nmol), and the corresponding 1,3 dicarbonyl compounds (2000 nmol,100 equivalents, 0.2M DMSO) were added to the solution, mixed well, and reacted at 80 ℃ for 16 hours.
Ethanol precipitation is carried out after the reaction is finished: adding 5M sodium chloride solution with the total volume of 10% into the reacted solution, continuously adding absolute ethanol with the total volume of 3 times, shaking uniformly, placing the reaction in dry ice, freezing for 0.5 hour, centrifuging for half an hour at the speed of 12000rpm, pouring out supernatant, dissolving the rest precipitate with deionized water to obtain 21 solutions of On-DNA products, quantifying by an enzyme-labeled instrument OD, and sending LCMS to confirm the conversion rate of the reaction.
Claims (6)
1. A method for synthesizing On-DNA pyrazole compounds, characterized in that: the method takes On-DNA hydrazine compounds and 1,3 dicarbonyl compounds as raw materials to react to obtain On-DNA products; wherein the structural formula of the On-DNA hydrazine compound isThe structural formula of the 1,3 dicarbonyl compound is +.>The structural formula of the On-DNA product is as follows:
wherein the DNA in the structural formula comprises a single-stranded or double-stranded nucleotide chain obtained by polymerizing an artificially modified and/or unmodified nucleotide monomer,the nucleotide chain is bonded with R through one or more chemical bonds or groups 1 Are connected;
said R' is selected from H, C 1 ～C 20 Alkoxycarbonyl and nitro;
said R is 1 Selected from phenyl, substituted phenyl, triazinyl, halogen substituted triazinyl, C 1 ～C 6 Alkyl, substituted C 1 ～C 6 An alkyl group; the number of substituents of the substituted phenyl is one or more, and the substituents of the substituted phenyl are independently selected from hydroxy, nitro, halogen and C 1 ～C 6 An alkyl group; substitution C 1 ～C 6 The number of substituents of the alkyl group is one or more; substitution C 1 ～C 6 The substituents of the alkyl groups are independently selected from phenyl groups;
said R is 2 、R 3 Respectively selected from hydrogen or C 1 ～C 6 An alkyl group;
the reaction steps of the method are as follows: dissolving On-DNA hydrazine compound into disodium hydrogen phosphate/sodium dihydrogen phosphate buffer solution with pH=5.5 to prepare solution, adding 10-1000 times of 1,3 dicarbonyl compound into On-DNA hydrazine compound solution with molar equivalent of 1 and molar concentration of 0.5-5mM, and reacting at 0-100 deg.c for 0.5-24 hr until the reaction is completed.
2. The method according to claim 1, characterized in that: the reaction temperature of the method is 20 ℃,30 ℃, 40 ℃,50 ℃,60 ℃, 70 ℃ or 80 ℃.
3. The method according to claim 1, characterized in that: the reaction time of the process is 1 hour, 4 hours, 8 hours, 10 hours, 16 hours, 18 hours or 20 hours.
4. The method according to claim 1, characterized in that: in the method, the molar equivalent of the On-DNA hydrazine compound is 50 equivalents, 100 equivalents, 200 equivalents, 300 equivalents, 400 equivalents, 500 equivalents, 600 equivalents, 800 equivalents and 1000 equivalents of the 1,3 dicarbonyl compound.
5. The method according to any one of claims 1-4, wherein the method is used for batch multi-well plate operations.
6. The method according to any one of claims 1 to 4, wherein the method is used for the synthesis of a library of DNA encoding compounds of a multiwell plate.
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| WO2017108741A1 (en) * | 2015-12-23 | 2017-06-29 | Technische Universität Dortmund | Dna-encoded chemical library, use thereof and method to synthesize the library |
| CN110041390A (en) * | 2019-04-26 | 2019-07-23 | 上海药明康德新药开发有限公司 | The synthetic method of the 1,2,3- 3-triazole compounds of On-DNA in DNA encoding compound library |
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| WO2017108741A1 (en) * | 2015-12-23 | 2017-06-29 | Technische Universität Dortmund | Dna-encoded chemical library, use thereof and method to synthesize the library |
| CN110041390A (en) * | 2019-04-26 | 2019-07-23 | 上海药明康德新药开发有限公司 | The synthetic method of the 1,2,3- 3-triazole compounds of On-DNA in DNA encoding compound library |
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