CN113264983A - Synthetic method of linker LND1088 for antibody-conjugated drug - Google Patents
Synthetic method of linker LND1088 for antibody-conjugated drug Download PDFInfo
- Publication number
- CN113264983A CN113264983A CN202110548700.4A CN202110548700A CN113264983A CN 113264983 A CN113264983 A CN 113264983A CN 202110548700 A CN202110548700 A CN 202110548700A CN 113264983 A CN113264983 A CN 113264983A
- Authority
- CN
- China
- Prior art keywords
- lnd1088
- lnd1067
- lnd
- stirring
- solvent
- Prior art date
- Legal status (The legal status is an assumption and is not a legal conclusion. Google has not performed a legal analysis and makes no representation as to the accuracy of the status listed.)
- Pending
Links
- 238000010189 synthetic method Methods 0.000 title claims abstract description 18
- 229940079593 drug Drugs 0.000 title claims abstract description 11
- 239000003814 drug Substances 0.000 title claims abstract description 11
- 238000006243 chemical reaction Methods 0.000 claims abstract description 72
- JGFZNNIVVJXRND-UHFFFAOYSA-N N,N-Diisopropylethylamine (DIPEA) Chemical compound CCN(C(C)C)C(C)C JGFZNNIVVJXRND-UHFFFAOYSA-N 0.000 claims abstract description 42
- ZMXDDKWLCZADIW-UHFFFAOYSA-N N,N-Dimethylformamide Chemical compound CN(C)C=O ZMXDDKWLCZADIW-UHFFFAOYSA-N 0.000 claims abstract description 36
- 230000015572 biosynthetic process Effects 0.000 claims abstract description 19
- 238000002156 mixing Methods 0.000 claims abstract description 19
- 238000003786 synthesis reaction Methods 0.000 claims abstract description 19
- HPNMFZURTQLUMO-UHFFFAOYSA-N diethylamine Chemical compound CCNCC HPNMFZURTQLUMO-UHFFFAOYSA-N 0.000 claims abstract description 14
- NQTADLQHYWFPDB-UHFFFAOYSA-N N-Hydroxysuccinimide Chemical compound ON1C(=O)CCC1=O NQTADLQHYWFPDB-UHFFFAOYSA-N 0.000 claims abstract description 9
- 238000001308 synthesis method Methods 0.000 claims abstract description 5
- FPQQSJJWHUJYPU-UHFFFAOYSA-N 3-(dimethylamino)propyliminomethylidene-ethylazanium;chloride Chemical compound Cl.CCN=C=NCCCN(C)C FPQQSJJWHUJYPU-UHFFFAOYSA-N 0.000 claims abstract description 3
- 238000003756 stirring Methods 0.000 claims description 37
- 239000002904 solvent Substances 0.000 claims description 21
- QTBSBXVTEAMEQO-UHFFFAOYSA-N Acetic acid Chemical compound CC(O)=O QTBSBXVTEAMEQO-UHFFFAOYSA-N 0.000 claims description 16
- 238000001816 cooling Methods 0.000 claims description 14
- 238000000034 method Methods 0.000 claims description 11
- 239000012295 chemical reaction liquid Substances 0.000 claims description 7
- 239000007821 HATU Substances 0.000 claims description 4
- 238000001035 drying Methods 0.000 claims description 4
- 238000010438 heat treatment Methods 0.000 claims description 4
- 230000008569 process Effects 0.000 claims description 4
- 238000005406 washing Methods 0.000 claims description 4
- -1 1-ethyl- (3-dimethylaminopropyl) Chemical group 0.000 claims description 3
- VEXZGXHMUGYJMC-UHFFFAOYSA-N Hydrochloric acid Chemical compound Cl VEXZGXHMUGYJMC-UHFFFAOYSA-N 0.000 claims description 3
- 239000007788 liquid Substances 0.000 claims description 2
- 238000004321 preservation Methods 0.000 claims description 2
- XLYOFNOQVPJJNP-UHFFFAOYSA-N water Substances O XLYOFNOQVPJJNP-UHFFFAOYSA-N 0.000 claims description 2
- LMDZBCPBFSXMTL-UHFFFAOYSA-N 1-Ethyl-3-(3-dimethylaminopropyl)carbodiimide Substances CCN=C=NCCCN(C)C LMDZBCPBFSXMTL-UHFFFAOYSA-N 0.000 claims 1
- WEVYAHXRMPXWCK-UHFFFAOYSA-N Acetonitrile Chemical compound CC#N WEVYAHXRMPXWCK-UHFFFAOYSA-N 0.000 description 36
- 238000004128 high performance liquid chromatography Methods 0.000 description 25
- 239000012071 phase Substances 0.000 description 14
- 238000001228 spectrum Methods 0.000 description 13
- 239000000047 product Substances 0.000 description 12
- 239000007787 solid Substances 0.000 description 11
- 238000000746 purification Methods 0.000 description 10
- 229940049595 antibody-drug conjugate Drugs 0.000 description 8
- 238000002514 liquid chromatography mass spectrum Methods 0.000 description 8
- XXMFJKNOJSDQBM-UHFFFAOYSA-N 2,2,2-trifluoroacetic acid;hydrate Chemical compound [OH3+].[O-]C(=O)C(F)(F)F XXMFJKNOJSDQBM-UHFFFAOYSA-N 0.000 description 6
- 239000012074 organic phase Substances 0.000 description 6
- 239000000611 antibody drug conjugate Substances 0.000 description 5
- 201000011510 cancer Diseases 0.000 description 4
- 230000002194 synthesizing effect Effects 0.000 description 4
- 239000003053 toxin Substances 0.000 description 4
- 231100000765 toxin Toxicity 0.000 description 4
- 108700012359 toxins Proteins 0.000 description 4
- IAZDPXIOMUYVGZ-UHFFFAOYSA-N Dimethylsulphoxide Chemical compound CS(C)=O IAZDPXIOMUYVGZ-UHFFFAOYSA-N 0.000 description 3
- 206010028980 Neoplasm Diseases 0.000 description 3
- 210000004027 cell Anatomy 0.000 description 3
- 231100000599 cytotoxic agent Toxicity 0.000 description 3
- 239000002619 cytotoxin Substances 0.000 description 3
- 239000003112 inhibitor Substances 0.000 description 3
- 231100000782 microtubule inhibitor Toxicity 0.000 description 3
- 101710112752 Cytotoxin Proteins 0.000 description 2
- 102000003915 DNA Topoisomerases Human genes 0.000 description 2
- 108090000323 DNA Topoisomerases Proteins 0.000 description 2
- 239000012623 DNA damaging agent Substances 0.000 description 2
- 102000004163 DNA-directed RNA polymerases Human genes 0.000 description 2
- 108090000626 DNA-directed RNA polymerases Proteins 0.000 description 2
- 229930126263 Maytansine Natural products 0.000 description 2
- PMZURENOXWZQFD-UHFFFAOYSA-L Sodium Sulfate Chemical compound [Na+].[Na+].[O-]S([O-])(=O)=O PMZURENOXWZQFD-UHFFFAOYSA-L 0.000 description 2
- FAPWRFPIFSIZLT-UHFFFAOYSA-M Sodium chloride Chemical class [Na+].[Cl-] FAPWRFPIFSIZLT-UHFFFAOYSA-M 0.000 description 2
- PQLVXDKIJBQVDF-UHFFFAOYSA-N acetic acid;hydrate Chemical compound O.CC(O)=O PQLVXDKIJBQVDF-UHFFFAOYSA-N 0.000 description 2
- 239000002253 acid Substances 0.000 description 2
- 239000002246 antineoplastic agent Substances 0.000 description 2
- 229940041181 antineoplastic drug Drugs 0.000 description 2
- 230000000052 comparative effect Effects 0.000 description 2
- 150000001875 compounds Chemical class 0.000 description 2
- 239000012043 crude product Substances 0.000 description 2
- 238000004108 freeze drying Methods 0.000 description 2
- 238000004895 liquid chromatography mass spectrometry Methods 0.000 description 2
- WKPWGQKGSOKKOO-RSFHAFMBSA-N maytansine Chemical compound CO[C@@H]([C@@]1(O)C[C@](OC(=O)N1)([C@H]([C@@H]1O[C@@]1(C)[C@@H](OC(=O)[C@H](C)N(C)C(C)=O)CC(=O)N1C)C)[H])\C=C\C=C(C)\CC2=CC(OC)=C(Cl)C1=C2 WKPWGQKGSOKKOO-RSFHAFMBSA-N 0.000 description 2
- 238000007670 refining Methods 0.000 description 2
- 238000000967 suction filtration Methods 0.000 description 2
- 229910021642 ultra pure water Inorganic materials 0.000 description 2
- 239000012498 ultrapure water Substances 0.000 description 2
- 238000005160 1H NMR spectroscopy Methods 0.000 description 1
- OHLUUHNLEMFGTQ-UHFFFAOYSA-N N-methylacetamide Chemical compound CNC(C)=O OHLUUHNLEMFGTQ-UHFFFAOYSA-N 0.000 description 1
- 230000009471 action Effects 0.000 description 1
- 125000000539 amino acid group Chemical group 0.000 description 1
- 239000000427 antigen Substances 0.000 description 1
- 108091007433 antigens Proteins 0.000 description 1
- 102000036639 antigens Human genes 0.000 description 1
- 239000008280 blood Substances 0.000 description 1
- 210000004369 blood Anatomy 0.000 description 1
- 239000003795 chemical substances by application Substances 0.000 description 1
- 229960002173 citrulline Drugs 0.000 description 1
- 230000000694 effects Effects 0.000 description 1
- 230000012202 endocytosis Effects 0.000 description 1
- 230000007062 hydrolysis Effects 0.000 description 1
- 238000006460 hydrolysis reaction Methods 0.000 description 1
- 230000002132 lysosomal effect Effects 0.000 description 1
- 238000004519 manufacturing process Methods 0.000 description 1
- 230000010534 mechanism of action Effects 0.000 description 1
- 230000004048 modification Effects 0.000 description 1
- 238000012986 modification Methods 0.000 description 1
- 238000002360 preparation method Methods 0.000 description 1
- 238000002626 targeted therapy Methods 0.000 description 1
- 230000008685 targeting Effects 0.000 description 1
Images
Classifications
-
- C—CHEMISTRY; METALLURGY
- C07—ORGANIC CHEMISTRY
- C07K—PEPTIDES
- C07K5/00—Peptides containing up to four amino acids in a fully defined sequence; Derivatives thereof
- C07K5/04—Peptides containing up to four amino acids in a fully defined sequence; Derivatives thereof containing only normal peptide links
- C07K5/10—Tetrapeptides
- C07K5/1002—Tetrapeptides with the first amino acid being neutral
- C07K5/1005—Tetrapeptides with the first amino acid being neutral and aliphatic
- C07K5/1008—Tetrapeptides with the first amino acid being neutral and aliphatic the side chain containing 0 or 1 carbon atoms, i.e. Gly, Ala
Landscapes
- Chemical & Material Sciences (AREA)
- Organic Chemistry (AREA)
- Health & Medical Sciences (AREA)
- Life Sciences & Earth Sciences (AREA)
- Biochemistry (AREA)
- Biophysics (AREA)
- General Health & Medical Sciences (AREA)
- Genetics & Genomics (AREA)
- Medicinal Chemistry (AREA)
- Molecular Biology (AREA)
- Proteomics, Peptides & Aminoacids (AREA)
- Peptides Or Proteins (AREA)
Abstract
The invention provides a synthetic method of a linker LND1088 for antibody-conjugated drugs. The synthesis method comprises the following steps: reacting LND1067-L1 with DX8951 to obtain LND 1067-1; mixing LND1067-1, diethylamine and N, N-dimethylformamide for reaction to obtain LND 1067-2; reacting LND1088-L2-1, LND1088-L2-2 and diisopropylethylamine to obtain LND 1088-L2-3; LND1088-L2-3, N-hydroxysuccinimide and EDCl are reacted to obtain LND 1088-L2; LND1067-2, LND1088-L2, and diisopropylethylamine were reacted to give LND 1088. The synthetic method avoids the synthesis of the intermediate LND1067-2-1, realizes the high-efficiency synthesis of LND1088 and is easy to amplify.
Description
Technical Field
The invention relates to a synthetic method of a linker, in particular to a synthetic method of a linker LND1088 for antibody coupled drugs, and belongs to the technical field of organic drug synthesis.
Background
Antibody Drug Conjugates (ADC) are a novel anti-tumor drug, and the principle is that cytotoxin is connected to an Antibody, and the cytotoxin is transported to a target point through recognition of a specific antigen on the surface of a cancer cell by the Antibody and entering the cancer cell through endocytosis, so that the aim of targeted therapy of malignant tumor is achieved. Compared with the traditional micromolecule antitumor drugs, the ADC has higher specificity and effectiveness due to the fact that the target recognition of the antibody and the high activity of the toxin can be used.
ADCs comprise three distinct components, namely antibodies, linkers and cytotoxins. The antibody realizes targeting, the linker ensures the stability of the ADC in the blood transportation process, and after the ADC reaches an action target, the toxin plays a role in killing cancer cells. Depending on the mechanism of action, the toxins suitable for ADCs are classified into Microtubule inhibitors (Microtubule inhibitors), DNA damaging agents (DNA damaging agents), RNA polymerase inhibitors (RNA polymerase inhibitors), and the like.
Currently, the toxins used in ADCs marketed and used in clinical trials are mainly microtubule inhibitors, mainly including compounds designed as DNA topoisomerase i (topoisomerase i) inhibitors, such as Dxd, and compounds designed based on Maytansine (Maytansine-based), such as DM1 and DM 4. In the linker context, the main applications are non-cleavable types, such as glycine-phenylalanine-glycine-citrulline (GGFG-Acid) and dibenzocyclooctynoic Acid (DBCO-Acid), which, after lysosomal hydrolysis, remain active and bind to an amino Acid residue via a linker region.
LND1088 as a linker for antibody-conjugated drugs, the structural formula is as follows:
the existing synthetic method needs to pass through the synthetic step of an intermediate LND1067-2, and the main synthetic method of LND1067-2 is as follows:
in the existing synthesis method, the synthesis yield of the intermediate LND1067-2 is low.
Disclosure of Invention
In order to solve the above-mentioned problems, an object of the present invention is to provide a method for synthesizing LND1088 with a high synthesis yield.
In order to achieve the above technical objects, the present invention first provides a method for synthesizing a linker LND1088 for an antibody-coupled drug, the method comprising:
LND1067-L1 and DX8951 react in a first solvent, and the reaction liquid passes through a reverse phase column and is purified to obtain LND 1067-1;
mixing LND1067-1, Diethylamine (DEA) and N, N-Dimethylformamide (DMF) for reaction, removing diethylamine from reaction solution under reduced pressure, adding acetic acid, passing through a reverse phase column, and purifying to obtain LND 1067-2;
LND1088-L2-1, LND1088-L2-2 and Diisopropylethylamine (DIPEA) react in a second solvent, and the reaction liquid passes through a reverse phase column and is purified to obtain LND 1088-L2-3;
reacting LND1088-L2-3, N-hydroxysuccinimide (NHS) and 1-ethyl- (3-dimethylaminopropyl) carbonyldiimine hydrochloride (EDCl) in a third solvent, washing with water, separating liquid, extracting, drying and concentrating to obtain LND 1088-L2;
LND1067-2, LND1088-L2 and diisopropylethylamine are reacted in a first solvent, and the reaction liquid is subjected to reverse phase column and purified to obtain LND 1088.
The synthesis method specifically comprises the following steps:
the synthetic method of the invention includes the step of preparing LND 1067-1.
In one embodiment of the present invention, in the preparation of LND1067-1,
and (3) cooling LND1067-L1 to-5 ℃, stirring for 3min-5min, adding HATU, and reacting for 1h-3h at 0-5 ℃ to obtain LND 1067-1.
Wherein the mixing molar equivalent ratio of LND1067-L1 to DX8951 is 1.4eq-2.0 eq: 1 eq.
The synthetic method of the invention includes the step of preparing LND 1067-2.
In one embodiment of the present invention, in preparing LND1067-2,
stirring LND1067-1 and N, N-dimethylformamide (solvent) at room temperature for 3min-5min, cooling to 0-5 ℃, stirring for 3min-5min, adding diethylamine, and reacting for 0.5h-1h under heat preservation to obtain LND 1067-2.
Wherein the mixing molar equivalent ratio of the diethylamine to the LND1067-1 is 10eq-20 eq: the mixing molar equivalent ratio of 1, N-dimethylformamide to LND1067-1 is 8eq-12 eq: 1 (i.e. 8-12 ml/g). Specifically, the mixing ratio of LND1067-1, N-dimethylformamide and diethylamine is 3.5g-4.5 g: 30mL-35 mL: 2.5g-3 g.
The synthetic method of the invention comprises the step of preparing LND 1088-L2-3.
In one embodiment of the present invention, in preparing LND1088-L2-3,
and (2) dissolving LND1088-L2-1 in a second solvent, stirring at room temperature, adding LND1088-L2-2, cooling to 0-5 ℃, stirring for 3-5min, adding diisopropylethylamine, reacting for 0.5-1h at a constant temperature, heating to room temperature, and stirring for 1-3h to obtain LND 1088-L2-3.
Wherein the mixing molar equivalent of LND1088-L2-1, LND1088-L2-2 and diisopropylethylamine is 1.0eq-1.5 eq: 1.0eq to 1.2 eq: 1.8eq-2.0 eq.
The synthetic process of the present invention includes the step of preparing LND 1088-L2.
In one embodiment of the present invention, in making LND1088-L2,
mixing LND1088-L2-3, the third solvent and N-hydroxysuccinimide, stirring at room temperature, adding EDCL, and stirring at room temperature for 3h-6h to obtain LND 1088-L2.
Wherein the mixing molar equivalent ratio of LND1088-L2-3, N-hydroxysuccinimide and EDCl is 1.0eq-1.2 eq: 1.3eq to 1.5 eq: 1.3eq to 1.5 eq.
The synthetic process of the invention includes the step of preparing LND 1088.
In one embodiment of the present invention, in preparing LND1088,
mixing LND1067-2, a first solvent and LND1088-L2, cooling to-5 ℃ to 0 ℃, stirring for 3min-5min, adding diisopropylethylamine, and reacting at-5 ℃ to 0 ℃ for 0.5h-1h to obtain LND 1088.
Wherein the mixing molar equivalent ratio of LND1067-2, LND1088-L2 and diisopropylethylamine is 1.0eq-1.2 eq: 1.0eq to 1.2 eq: 1.8eq-2.0 eq.
In one embodiment of the invention, the first solvent is DCM, THF or DMF;
the second solvent is DMF, DMA or DCM;
the third solvent is EA or DCM.
The synthetic method of the linker LND1088 for the antibody-conjugated drug avoids the linker LND1067-2 which is low in yield and difficult to purify in the prior art, realizes novel and efficient synthesis of the LND1088, is easy to amplify and is convenient for commercial production.
Drawings
FIG. 1 is an HPLC spectrum of LND1067-1 in an example of the invention.
FIG. 2 is a LCMS spectrum of LND1067-1 in an example of the invention.
FIG. 3 is a HNMR spectrum of LND1067-1 in an embodiment of the invention.
FIG. 4 is an HPLC spectrum of LND1067-2 in an example of the invention.
FIG. 5 is a LCMS spectrum of LND1067-2 in an example of the invention.
FIG. 6 is a HNMR spectrum of LND1067-2 in an embodiment of the present invention.
FIG. 7 is an HPLC spectrum of LND1088-L2-3 in an example of the invention.
FIG. 8 is a LCMS spectrum of LND1088-L2-3 in an embodiment of the invention.
FIG. 9 is an HPLC spectrum of LND1088-L2 in an example of the invention.
FIG. 10 is a LCMS spectrum of LND1088-L2 in an embodiment of the invention.
FIG. 11 is an HPLC spectrum of LND1088 in an example of the invention.
FIG. 12 is a LCMS spectrum of LND1088 in an embodiment of the invention.
FIG. 13 is a HNMR spectrum of LND1088 in an embodiment of the present invention.
Detailed Description
Example 1
The embodiment provides a method for synthesizing a linker LND1088 for antibody-conjugated drugs, which specifically comprises the following steps.
Synthesis of LND1067-1
DX8951(LND1029, 1.0eq/2.0g) is taken and put into a reaction bottle, anhydrous DMF (10V/20mL) and DIPEA (3.0eq/1.64g) are stirred for 5-10min at room temperature; LND1067-L1(3.83g/1.4eq) is added into a reaction bottle, then the reaction system is transferred into a low-temperature reactor, and the temperature is reduced to-5 ℃ and the stirring is carried out for 3-5 min; adding HATU (as condensing agent, 1.6eq/2.58 g) into the reaction system, and reacting for 1-3h at 0-5 ℃; detecting a reaction system by HPLC, and after the reaction is completed, carrying out post-treatment: the reaction system is directly purified in a medium-pressure reverse phase way (the product is obtained by purifying a 0.1 percent TFA water/acetonitrile system, and the acetonitrile ratio is 70-80 percent); the product was collected and lyophilized to give 4.05g of a yellow solid with an HPLC purity of 93.3% and a yield of 90%.
HPLC of LND1067-1 is shown in FIG. 1, LCMS (FIG. 2) and HNMR spectra (FIG. 3).
Synthesis of LND1067-2
Putting LND1067-1(1.0eq/4.0g) anhydrous DMF (8V/32mL) in a reaction bottle, and stirring at room temperature for 3-5 min; cooling the reaction system to 0-5 deg.C, and stirring for 3-5 min; adding DEA (2.75g, 3.9mL) into the reaction system, and reacting for 0.5-1h at the temperature of 0-5 ℃; detecting a reaction system by HPLC, and after the reaction is completed, carrying out post-treatment: concentrating the reaction system to remove DEA; dissolving 0.45g of acetic acid in a small amount of DMF, adding into the reaction system, and shaking up; direct medium-pressure reverse-phase purification of the reaction system (0.1% TFA water/acetonitrile system purification); the product was collected and lyophilized to give 2.87g of a yellow solid with an HPLC purity of 93.2% and a yield of 89%.
HPLC of LND1067-2 is shown in FIG. 4, LCMS (FIG. 5) and HNMR spectra (FIG. 6).
Synthesis of LND1088-L2-3
Taking LND1088-L2-1(1.0eq/1.33g) anhydrous DMF (10V/13mL), stirring at room temperature, and putting LND1088-L2-2(1.05eq/1.04g) in a reaction bottle; cooling the reaction system to 0-5 ℃, stirring for 3-5min, adding DIPEA (0.85g/2.0eq) into the reaction system, keeping the temperature at 0-5 ℃, reacting for 0.5-1h, then heating to room temperature, and stirring for 1-3 h; detecting a reaction system by HPLC, and after the reaction is completed, carrying out post-treatment: direct medium-pressure reverse-phase purification of the reaction system (0.1% TFA water/acetonitrile system purification); the product was collected and lyophilized to give 1.43g of a pale yellow viscous solid with an HPLC purity of 93.3% and a yield of 90%.
The HPLC spectrum of LND1088-L2-3 is shown in FIG. 7, and the LCMS spectrum is shown in FIG. 8.
Synthesis of LND1088-L2
LND1088-L2-3(1.0eq/1.43g), DCM (20V/28mL), NHS (1.5eq/0.45g) were added into the reaction system; stirring at room temperature; adding EDCl (1.5eq/0.75g) into the reaction system, and stirring at room temperature for 3-6 h; detecting a reaction system by HPLC, and after the reaction is completed, carrying out post-treatment: adding 10V/14mL of ultrapure water into the reaction system, stirring, separating, back-extracting the organic phase with 20V DCM, combining the organic phases, washing with 10V/14mL of saturated saline solution, and drying with anhydrous sodium sulfate for 1-2 h; suction filtration and vacuum concentration of the organic phase gave 1.68g of a pale yellow viscous solid with an HPLC purity of 96.5% and a yield of 100%.
The HPLC of LND1088-L2 is shown in FIG. 9, and the LCMS spectrum is shown in FIG. 10.
Synthesis of LND1088 DBCO-PEG4-GGFG-Dxd
LND1067-2(1.0eq/500mg) anhydrous DMF 15V/7.5mL) and LND1088-L2(1.0eq/351mg) are added into a reaction bottle; transferring the reaction system into a low-temperature reactor, cooling to-5-0 ℃, stirring for 3-5min, adding DIPEA (136mg/2.0eq) into the reaction system, and reacting for about 0.5-1h at the temperature of-5-0 ℃; detecting a reaction system by HPLC, and after the reaction is completed, carrying out post-treatment: adding 0.1mL of AcOH into the reaction system at the temperature of between 5 and 0 ℃; directly performing medium-pressure reverse-phase purification on a reaction system (purifying a 0.1% AcOH water/acetonitrile system, and obtaining a product in an acetonitrile ratio of 80-90%), collecting a product, and freeze-drying to obtain a yellow-like solid crude product LND 1088-C; medium pressure reverse phase refining to obtain light yellow solid LND1088, 470mg, HPLC purity 98.6%, yield 62.5%.
LND1088 has 1H NMR (400MHz, DMSO) δ 8.63(t, J ═ 6.6Hz, 1H), 8.50(d, J ═ 8.8Hz, 1H), 8.29(t, J ═ 5.8Hz, 1H), 8.15(t, J ═ 5.6Hz, 1H), 8.11(d, J ═ 7.9Hz, 1H), 7.99(t, J ═ 5.6Hz, 1H), 7.75(t, J ═ 9.5, 2H), 7.69 to 7.58(m, 2H), 7.48(dt, J ═ 15.4, 5.3Hz, 3H), 7.39 to 7.13(m, 10 ddh), 5.59 (td, J ═ 13.8, 6.0, 1H, 5.42(s, 2.3 Hz, 3H), 7.39 to 7.13(m, 10 ddh), 5.59 (td, J ═ 13.8, 6.0, 1H), 3H, 3, 3.29(t, J ═ 5.9Hz, 2H), 3.17(d, J ═ 18.4Hz, 2H), 3.05(ddd, J ═ 18.3, 11.7, 4.9Hz, 3H), 2.77(dd, J ═ 13.7, 9.6Hz, 1H), 2.62-2.52(m, 1H), 2.40-2.35(m, 5H), 2.20(dd, J ═ 13.7, 6.2Hz, 3H), 2.05-1.95(m, 1H), 1.91-1.81(m, 2H), 1.80-1.71(m, 1H), 0.87(t, J ═ 7.3Hz, 3H).
The HPLC spectrum of LND1088 is shown in FIG. 11, the LCMS spectrum is shown in FIG. 12, and the HNMR spectrum is shown in FIG. 13.
Comparative example 1
The comparative example provides a method for synthesizing a linker LND1088 for antibody-conjugated drugs, comprising the following steps.
Synthesis of LND1067-1
DX8951(1.0eq/2.0g) was placed in a reaction flask, and anhydrous DMF (10V/20mL) and DIPEA (3.0eq/1.64g) were stirred at room temperature for 15 min; LND1067-L1(3.83g/1.4eq) is added into a reaction bottle, then the reaction system is transferred into a low-temperature reactor, and the temperature is reduced to-5 ℃ and the stirring is carried out for 10 min; HATU (2.58g/1.6eq) is added into a reaction system and is kept at the temperature of 0-5 ℃ for reaction for 5 hours; detecting a reaction system by HPLC, and after the reaction is completed, carrying out post-treatment: the reaction system is directly purified in a medium-pressure reverse phase way (the product is obtained by purifying a 0.1 percent TFA water/acetonitrile system, and the acetonitrile ratio is 70-80 percent); the product was collected and lyophilized to give a yellow solid in 60% yield.
Synthesis of LND1067-2
Taking LND1067-1(8.0g) anhydrous DMF (8V/32mL) in a reaction bottle, and stirring at room temperature for 3-5 min; cooling the reaction system to 0-5 deg.C, and stirring for 3-5 min; DEA (5g/10.0eq) is added into a reaction system, and the temperature is kept at 0-5 ℃ for reaction for 0.5-1 h; detecting a reaction system by HPLC, and after the reaction is completed, carrying out post-treatment: concentrating the reaction system to remove DEA; dissolving 0.45g of acetic acid in a small amount of DMF, adding into the reaction system, and shaking up; direct medium-pressure reverse-phase purification of the reaction system (0.1% TFA water/acetonitrile system purification); the product was collected and lyophilized to give a yellow solid in 67% yield.
Synthesis of LND1088-L2-3
Taking LND1088-L2-1(1.0eq/1.33g) anhydrous DMF (10V/13mL), stirring at room temperature, and putting LND1088-L2-2(3g) in a reaction bottle; cooling the reaction system to 0-5 ℃, stirring for 3-5min, adding DIPEA (2g) into the reaction system, keeping the temperature at 0-5 ℃, reacting for 0.5-1h, then heating to room temperature, and stirring for 1-3 h; detecting a reaction system by HPLC, and after the reaction is completed, carrying out post-treatment: direct medium-pressure reverse-phase purification of the reaction system (0.1% TFA water/acetonitrile system purification); the product was collected and lyophilized to give a pale yellow viscous solid in 45% yield.
Synthesis of LND1088-L2
LND1088-L2-3(3g), DCM (20V/28mL), NHS (1.5eq/0.45g) were added into the reaction system; stirring at room temperature; EDCl (1.5eq/0.75g) was added to the reaction system and stirred at room temperature for 1 h; detecting a reaction system by HPLC, and after the reaction is completed, carrying out post-treatment: adding 10V/14mL of ultrapure water into the reaction system, stirring, separating, back-extracting the organic phase with 20V DCM, combining the organic phases, washing with 10V/14mL of saturated saline solution, and drying with anhydrous sodium sulfate for 1-2 h; suction filtration and organic phase vacuum concentration gave a pale yellow viscous solid with a yield of 60%.
Synthesis of LND1088 DBCO-PEG4-GGFG-Dxd
Taking LND1067-2(300mg), anhydrous DMF 15V/7.5mL) and LND1088-L2(501mg) to be added into a reaction bottle; transferring the reaction system into a low-temperature reactor, cooling to 5 ℃, stirring for 10min, adding DIPEA (136mg/2.0eq) into the reaction system, and reacting for about 0.5-1h at the temperature of-5 to 0 ℃; detecting a reaction system by HPLC, and after the reaction is completed, carrying out post-treatment: adding 0.1mL of AcOH into the reaction system at the temperature of between 5 and 0 ℃; directly performing medium-pressure reverse-phase purification on a reaction system (purifying by a 0.1% AcOH water/acetonitrile system, and obtaining a product according to the proportion of 80-90% acetonitrile), collecting a product, and freeze-drying to obtain a yellow solid-like crude product LND 1088-C; medium pressure reverse phase refining gave light yellow solid LND1088 in 35.7% yield.
The above embodiments are merely illustrative of the technical ideas and features of the present invention, and the purpose thereof is to enable those skilled in the art to understand the contents of the present invention and implement the present invention, and not to limit the protection scope of the present invention. All equivalent changes and modifications made according to the spirit of the present invention should be covered within the protection scope of the present invention.
Claims (10)
1. A method of synthesis of linker LND1088 for antibody-coupled drugs, the method of synthesis comprising:
LND1067-L1 and DX8951 react in a first solvent, and the reaction liquid passes through a reverse phase column and is purified to obtain LND 1067-1;
mixing the LND1067-1, diethylamine and N, N-dimethylformamide for reaction, decompressing reaction liquid to remove diethylamine, adding acetic acid, passing through a reverse phase column, and purifying to obtain LND 1067-2;
LND1088-L2-1, LND1088-L2-2 and diisopropylethylamine are reacted in a second solvent, and the reaction liquid passes through a reverse phase column and is purified to obtain LND 1088-L2-3;
reacting the LND1088-L2-3, N-hydroxysuccinimide and 1-ethyl- (3-dimethylaminopropyl) carbonyldiimine hydrochloride in a third solvent, washing with water, separating liquid, extracting, drying and concentrating to obtain LND 1088-L2;
and (3) reacting the LND1067-2, the LND1088-L2 and diisopropylethylamine in a first solvent, and purifying the reaction liquid through a reverse phase column to obtain LND 1088.
2. The synthetic method according to claim 1, wherein in preparing LND1067-1,
and (3) cooling LND1067-L1 to-5 ℃, stirring for 3min-5min, adding HATU, and reacting for 1h-3h at 0-5 ℃ to obtain LND 1067-1.
3. The synthesis process of claim 1 or 2, wherein the molar equivalent ratio of LND1067-L1 to DX8951 is 1.4eq-2.0 eq: 1 eq.
4. The synthetic method according to claim 1, wherein in preparing LND1067-2,
stirring LND1067-1 and N, N-dimethylformamide at room temperature for 3min-5min, cooling to 0-5 ℃, stirring for 3min-5min, adding diethylamine, and reacting for 0.5h-1h under heat preservation to obtain LND 1067-2.
5. The synthesis method according to claim 1 or 4, wherein the mixing molar equivalent ratio of diethylamine to LND1067-1 is 10eq-20 eq: 1, and the mixing molar equivalent ratio of N, N-dimethylformamide to LND1067-1 is 8eq-12 eq: 1.
6. The synthetic method according to claim 1, wherein when LND1088-L2-3 is prepared,
dissolving LND1088-L2-1 in a second solvent, stirring at room temperature, adding LND1088-L2-2, cooling to 0-5 ℃, stirring for 3-5min, adding diisopropylethylamine, reacting for 0.5-1h at constant temperature, heating to room temperature, and stirring for 1-3h to obtain LND 1088-L2-3;
preferably, the mixing equivalent weight of the LND1088-L2-1, the LND1088-L2-2 and the diisopropylethylamine is 1.0eq-1.5 eq: 1.0eq-1.2 eq: 1.8eq-2.0 eq.
7. The synthetic method according to claim 1, wherein in preparing LND1088-L2,
mixing the LND1088-L2-3, a third solvent and N-hydroxysuccinimide, stirring at room temperature, adding 1-ethyl- (3-dimethylaminopropyl) carbodiimide hydrochloride, and stirring at room temperature for 3h-6h to obtain the LND 1088-L2;
preferably, the mixing molar equivalent ratio of the LND1088-L2-3, the N-hydroxysuccinimide, and the 1-ethyl- (3-dimethylaminopropyl) carbonyldiimine hydrochloride is 1.0eq-1.2 eq: 1.3eq-1.5 eq.
8. The synthetic method according to claim 1, wherein, in preparing LND1088,
and (2) mixing the LND1067-2, the first solvent and the LND1088-L2, cooling to-5 ℃ to 0 ℃, stirring for 3min-5min, adding diisopropylethylamine, and reacting for 0.5h-1h at the temperature of-5 ℃ to 0 ℃ to obtain the LND 1088.
9. The synthesis method according to claim 1 or 8, wherein the mixing molar equivalent ratio of LND1067-2, LND1088-L2 and diisopropylethylamine is 1.0eq-1.2 eq: 1.8eq-2.0 eq.
10. The synthetic method of claim 1, wherein the first solvent is DCM, THF or DMF;
the second solvent is DMF, DMA or DCM;
the third solvent is EA or DCM.
Priority Applications (1)
| Application Number | Priority Date | Filing Date | Title |
|---|---|---|---|
| CN202110548700.4A CN113264983A (en) | 2021-05-20 | 2021-05-20 | Synthetic method of linker LND1088 for antibody-conjugated drug |
Applications Claiming Priority (1)
| Application Number | Priority Date | Filing Date | Title |
|---|---|---|---|
| CN202110548700.4A CN113264983A (en) | 2021-05-20 | 2021-05-20 | Synthetic method of linker LND1088 for antibody-conjugated drug |
Publications (1)
| Publication Number | Publication Date |
|---|---|
| CN113264983A true CN113264983A (en) | 2021-08-17 |
Family
ID=77231853
Family Applications (1)
| Application Number | Title | Priority Date | Filing Date |
|---|---|---|---|
| CN202110548700.4A Pending CN113264983A (en) | 2021-05-20 | 2021-05-20 | Synthetic method of linker LND1088 for antibody-conjugated drug |
Country Status (1)
| Country | Link |
|---|---|
| CN (1) | CN113264983A (en) |
Cited By (2)
| Publication number | Priority date | Publication date | Assignee | Title |
|---|---|---|---|---|
| CN114394960A (en) * | 2021-12-10 | 2022-04-26 | 宜昌博仁凯润药业有限公司 | Dibenzocyclooctyne-tetraethylene glycol-active ester compound, preparation method and application |
| CN114773588A (en) * | 2022-05-05 | 2022-07-22 | 武汉翱飞科技有限公司 | DBCO modified polyethylene glycol linking agent and synthesis method thereof |
Citations (2)
| Publication number | Priority date | Publication date | Assignee | Title |
|---|---|---|---|---|
| CN110291097A (en) * | 2016-11-08 | 2019-09-27 | 里珍纳龙药品有限公司 | Steroidal compounds and its protein-conjugate |
| CN111051330A (en) * | 2017-08-31 | 2020-04-21 | 第一三共株式会社 | Improved preparation method of antibody-drug conjugate |
-
2021
- 2021-05-20 CN CN202110548700.4A patent/CN113264983A/en active Pending
Patent Citations (2)
| Publication number | Priority date | Publication date | Assignee | Title |
|---|---|---|---|---|
| CN110291097A (en) * | 2016-11-08 | 2019-09-27 | 里珍纳龙药品有限公司 | Steroidal compounds and its protein-conjugate |
| CN111051330A (en) * | 2017-08-31 | 2020-04-21 | 第一三共株式会社 | Improved preparation method of antibody-drug conjugate |
Cited By (2)
| Publication number | Priority date | Publication date | Assignee | Title |
|---|---|---|---|---|
| CN114394960A (en) * | 2021-12-10 | 2022-04-26 | 宜昌博仁凯润药业有限公司 | Dibenzocyclooctyne-tetraethylene glycol-active ester compound, preparation method and application |
| CN114773588A (en) * | 2022-05-05 | 2022-07-22 | 武汉翱飞科技有限公司 | DBCO modified polyethylene glycol linking agent and synthesis method thereof |
Similar Documents
| Publication | Publication Date | Title |
|---|---|---|
| CN113264983A (en) | Synthetic method of linker LND1088 for antibody-conjugated drug | |
| US10221155B2 (en) | Method for preparing Alectinib | |
| JP2006518704A (en) | Tube lysin, method for producing the same, and tube lysin preparation | |
| CN105732622A (en) | Preparation method of apixaban | |
| CN102367291A (en) | Mono-functional group-containing multilevel branched polyethylene glycol and its synthesis method | |
| CN109400528B (en) | Synthesis method of kebomei | |
| CN113402584A (en) | Synthetic method of intermediate LND1067-L1 of antibody-coupled drug linker | |
| CN114685485B (en) | Synthesis method of pyridoimidazole thiopropionic acid anti-gout compound | |
| CN113336823A (en) | Synthetic method for antibody-conjugated drug linker LND1067 | |
| CA2368815A1 (en) | Novel synthesis and crystallization of piperazine ring-containing compounds | |
| CN101402573B (en) | Bihydroxyl-2-acyl benzene acetic acid ester, producing method and uses thereof | |
| CN111362926A (en) | Synthetic method of intermediate CLA-SN38 for antibody coupled drug and intermediate thereof | |
| CN111454230A (en) | Synthesis method of key intermediate Tuv of natural anticancer drug Tubulysins | |
| CN109232559A (en) | A kind of synthetic method of [60] fullerene dihydro carboline analog derivative | |
| CN106674330B (en) | Preparation method of 34-Dimethyl apratoxin A/E | |
| CN114874287B (en) | Synthesis method of antibody coupling drug-linker LND1042 | |
| CN114057767A (en) | Preparation method of temsirolimus | |
| CN111574463B (en) | Rivastigmine intermediate compound IV | |
| CN111574520B (en) | Riagliptin intermediate compound V | |
| CN103012328B (en) | Method for preparing second-generation taxol anticancer drug Cabazitaxel | |
| US20030088094A1 (en) | Novel synthesis and crystallization of piperazine ring-containing compounds | |
| CN111138435A (en) | Modified methotrexate and preparation method and application thereof | |
| CN110563721A (en) | Preparation method of azasetron hydrochloride | |
| CN112125889A (en) | Preparation method of 7-bromo-2- (1-methyl-1H-pyrazol-4-yl) quinoxaline | |
| CN111363005B (en) | Synthetic method for antibody-coupled drug intermediate CLB-SN38 |
Legal Events
| Date | Code | Title | Description |
|---|---|---|---|
| PB01 | Publication | ||
| PB01 | Publication | ||
| SE01 | Entry into force of request for substantive examination | ||
| SE01 | Entry into force of request for substantive examination | ||
| RJ01 | Rejection of invention patent application after publication |
Application publication date: 20210817 |
|
| RJ01 | Rejection of invention patent application after publication |