Pharmaceutical synthesis process of Tovorafenib

Abstract

The invention relates to a pharmaceutical synthesis process of Tovorafenib (Tovorafenib). According to the synthesis process and the related preparation method of the chiral alpha-miscellaneous amine, prochiral ketone is directly reduced and aminated into optical pure chiral alpha-miscellaneous amine by a one-step method under a mild condition by utilizing a biocatalyst amine dehydrogenase, the defects of the existing catalysis technology can be overcome, the process route is simple, the reaction condition is mild, the yield is high, and the method is suitable for industrial production. The method has green synthesis advantages of step economy, atom economy and the like, and has the advantages of good stereoselectivity (ee value is as high as 99%) and the like. In the synthesis route, a method of condensing amino and carboxyl in an HATU and alkaline environment is mostly adopted, the operation is simple, the reaction yield is high, and the method is suitable for batch production.

Description

Technical Field

[0001] The present invention relates to the field of biomedicine technology, and in particular to a drug synthesis process of Tovorafenib. Background Art

[0002] Tovorafenib is an oral, brain-penetrating pan-RAF kinase inhibitor developed by Day One Biopharmaceuticals that can inhibit wild-type and certain mutant forms of BRAF, CRAF, and ARAF protein kinases. On October 30, 2023, Day One announced that the FDA accepted the new drug application for Tovorafenib for the treatment of recurrent or progressive pediatric low-grade glioma (pLGG) and obtained priority review. On April 23, 2024, Day One Biopharmaceuticals' Tovorafenib tablets (specification 100MG) and Tovorafenib oral suspension (specification 25MG / ML) were approved for marketing by the US FDA.

[0003] Tovorafenib's NDA is based on the results of an open-label pivotal Phase 2 trial evaluating Tovorafenib as a once-weekly monotherapy for patients with recurrent or progressive pLGG aged 6 months to 25 years. In November 2023, the results of this Phase 2 (FIREFLY-1) trial of Tovorafenib were published in Nature Medicine. The results showed that Tovorafenib may be an effective treatment for recurrent / refractory pediatric low-grade gliomas carrying BRAF mutations, bringing new hope for the treatment of pediatric low-grade gliomas. The results showed that according to independent review assessment, in patients with high-grade gliomas in neuro-oncology (RANO-HGG), the overall response rate (ORR) was 67%, reaching the pre-set primary endpoint of Group 1, with a median duration of response (DOR) of 16.6 months, and a median time to response (TTR) of 3.0 months (secondary endpoint). In patients with pediatric neuro-oncology low-grade glioma (RAPNO), the ORR was 51%, the median DOR was 13.8 months, and the median TTR was 5.3 months.

[0004] The chemical structure of Tovorafenib is shown in Formula I:

[0005]

[0006] However, the current drug synthesis process for Tovorafenib still needs to be improved. Summary of the invention

[0007] The present invention aims to solve one of the technical problems in the related art at least to a certain extent. To this end, one object of the present invention is to propose a pharmaceutical synthesis process of a compound Tovorafenib shown in Formula I. Relative to the prior art, the method described in the present invention uses a potential chiral ketone compound (compound shown in Formula 1) as a raw material, isopropylamine as an amine donor, water as a solvent, DMSO as a cosolvent, transaminase (ATASK-000250) as a biocatalyst, and PLA (pyridoxal phosphate) as a coenzyme regeneration system, and the ketone is reduced and aminated in one step to form an optically pure compound shown in Formula 2 (a chiral α-heteroamine compound), and then the compound shown in Formula 2 is subjected to a condensation reaction with the compound shown in Formula 3 to obtain a compound shown in Formula 4, and the compound shown in Formula 4 is then hydrolyzed under an alkaline environment to obtain a corresponding carboxylic acid compound (compound shown in Formula 5), ​​and then the compound shown in Formula 5 is subjected to a condensation reaction with the compound shown in Formula 6 to obtain the product compound Tovorafenib shown in Formula I.

[0008] In one aspect of the present invention, the present invention provides a pharmaceutical synthesis process of Tovorafenib, a compound represented by Formula I. According to an embodiment of the present invention, the synthesis process comprises:

[0009] (1) contacting the compound represented by Formula 1 with a transaminase to obtain the compound represented by Formula 2;

[0010] (2) contacting the compound represented by Formula 2 with the compound represented by Formula 3, HATU, and DIPEA to obtain the compound represented by Formula 4;

[0011] (3) contacting the compound represented by Formula 4 with LiOH to obtain the compound represented by Formula 5;

[0012] (4) contacting the compound represented by Formula 5 with the compound represented by Formula 6, HATU and DIPEA to obtain the compound represented by Formula I, Tovorafenib,

[0013]

[0014] The inventors have found that, by utilizing the drug synthesis process of the present invention, using the compound shown in Formula 1 as the starting material, a total of four steps of reaction can be performed to successfully synthesize and prepare the target product Tovorafenib.

[0015] The term "contact" as used herein should be understood in a broad sense, and can be any manner that allows at least two reactants to undergo a chemical reaction, such as mixing the two reactants under appropriate conditions. If necessary, the reactants to be contacted can be mixed under stirring, and thus, the type of stirring is not particularly limited, and can be mechanical stirring, i.e. stirring under the action of mechanical force.

[0016] In this document, the terms "first" and "second" are used for descriptive purposes only and should not be understood as indicating or implying relative importance or implicitly indicating the number of the indicated technical features. Thus, a feature defined as "first" or "second" may explicitly or implicitly include one or more of the features. In the description of the present invention, the meaning of "plurality" is two or more, unless otherwise clearly and specifically defined.

[0017] According to an embodiment of the present invention, the method for preparing the compound represented by Formula 2, the compound represented by Formula 4, the compound represented by Formula 5, and the compound represented by Formula I may also have at least one of the following additional technical features:

[0018] According to an embodiment of the present invention, the chemical reaction described in the present invention can be carried out according to any method known in the art. The source of the raw materials of the compound shown in Formula 2, the compound shown in Formula 4, the compound shown in Formula 5, and the compound shown in Formula 1 is not particularly limited, and it can be prepared by any known method or commercially available. For example, the cas of the compound shown in Formula 1 is: 1261080-59-4.

[0019] According to an embodiment of the present invention, in step (1), the contacting method of the compound of Formula 1 and the transaminase is not particularly limited. Thus, the efficiency of the contact reaction between the compound of Formula 1 and the transaminase can be improved, the reaction speed can be accelerated, and the efficiency of preparing the compound of Formula 2 by using this method can be further improved.

[0020] According to an embodiment of the present invention, in step (1), the following steps are included: dissolving the compound shown in Formula 1 in DMSO, placing it in a reaction bottle, adding a phosphate buffer with a pH of 7.5 and a concentration of 0.1M and a transaminase, shaking the reaction at 30°C and 250rpm for 48 hours, adding a 10M NaOH solution to the reaction solution in an ice-water bath to stop the reaction, extracting the reaction solution with ethyl acetate, filtering the mixed solution and separating the organic layer, adding saturated brine for washing, drying with anhydrous sodium sulfate, concentrating the filtrate under reduced pressure to evaporate the solvent, and purifying the concentrate with a petroleum ether / ethyl acetate mixed solvent with a volume ratio of 10:1 by silica gel column chromatography to obtain the compound shown in Formula 2. Thus, the efficiency of the contact reaction between the compound shown in Formula 1 and the transaminase can be improved, the reaction speed can be accelerated, and the efficiency of preparing the compound shown in Formula 2 by using this method can be further improved.

[0021] According to an embodiment of the present invention, in step (1), the transaminase is purchased from Codex Amine Transaminase Screening Kit ATASK-000250.

[0022] According to an embodiment of the present invention, in step (1), the phosphate buffer consists of 0.25 mM pyridoxal phosphate and 1 M isopropylamine.

[0023] According to an embodiment of the present invention, in step (1), the weight ratio (g / g) of the compound represented by Formula 1 to the transaminase is 1:0.15. Thus, the efficiency of preparing the compound represented by Formula 2 by this method can be further improved.

[0024] According to a specific embodiment of the present invention, in step (1), the following steps are included: dissolving the compound represented by formula 1 (10.0 g, 54.0 mmol) in DMSO (100 mL), placing the mixture in a 5 L reaction bottle, adding phosphate buffer (2106 mL, 0.25 mM pyridoxal phosphate, 1 M isopropylamine) with a pH of 7.5 and a concentration of 0.1 M and transaminase (1.5 g, purchased from Codex Amine Transaminase Screening Kit ATASK-000250), and shaking the mixture at 30° C. and 250 rpm for 48 hours. , 10M NaOH solution (1736mL) was added to the reaction solution under ice-water bath and stirred to stop the reaction. The reaction solution was extracted with ethyl acetate (2×2L), the mixed solution was filtered and the organic layer was separated, washed with saturated brine and dried over anhydrous sodium sulfate, the filtrate was concentrated under reduced pressure to evaporate the solvent, and the concentrate was purified by silica gel column chromatography using a mixed solvent of petroleum ether / ethyl acetate in a volume ratio of 10:1 to obtain the compound of formula 2, with an amount of 7.4g, a yield of 73.6%, a purity of 99.5% (HPLC), and an optical purity of 99.3% (chiral HPLC).

[0025] According to an embodiment of the present invention, in step (2), the contacting method of the compound represented by Formula 2 with the compound represented by Formula 3, HATU, and DIPEA is not particularly limited. Thus, the efficiency of the contact reaction between the compound represented by Formula 2 and the compound represented by Formula 3, HATU, and DIPEA can be improved, the reaction speed can be accelerated, and the efficiency of preparing the compound represented by Formula 4 using this method can be further improved.

[0026] According to an embodiment of the present invention, in step (2), the following steps are included: adding the compound of formula 2 and the compound of formula 3 to DMF at 0°C, stirring, then adding 1-[bis(dimethylamino)methylene]-1H-1,2,3-triazole [4,5-b] pyridine-3-oxide hexafluorophosphate (HATU) and N,N-diisopropylethylamine (DIPEA), the reaction solution is naturally heated to room temperature and stirred for reaction for 6 to 8 hours, after the reaction is completed, water is added to the reaction solution for washing, and then ethyl acetate is added for extraction, the organic phases are combined and then washed with saturated brine, dried over anhydrous sodium sulfate, the filtrate is concentrated under reduced pressure to evaporate the solvent, and the concentrate is purified by silica gel column chromatography using a petroleum ether / ethyl acetate mixed solvent to obtain the compound of formula 4. Thus, the efficiency of the contact reaction between the compound of formula 2 and the compound of formula 3, HATU, and DIPEA can be improved, the reaction speed can be accelerated, and the efficiency of preparing the compound of formula 4 using this method can be further improved.

[0027] According to an embodiment of the present invention, in step (2), the molar ratio of the compound represented by Formula 2 to the compound represented by Formula 3, HATU, and DIPEA is 1:(1.0-1.2):(1.3-2.2):(1.5-3.5), and preferably the molar ratio of the compound represented by Formula 2 to the compound represented by Formula 3, HATU, and DIPEA is 1:1.02:1.5:2.0. Thus, the efficiency of preparing the compound represented by Formula 4 by this method can be further improved.

[0028] According to an embodiment of the present invention, in step (2), the volume ratio of petroleum ether to ethyl acetate in the petroleum ether / ethyl acetate mixed solvent is (3-6):1, and preferably the volume ratio of petroleum ether / ethyl acetate mixed solvent is 4:1.

[0029] According to an embodiment of the present invention, in step (2), the reaction solution is preferably stirred and reacted for 6.5 hours after the reaction solution is naturally heated to room temperature.

[0030] According to a specific embodiment of the present invention, in step (2), the following steps are included: at 0°C, the compound represented by formula 2 (6.5 g, 34.90 mmol) and the compound represented by formula 3 (6.18 g, 35.60 mmol) are added to DMF (65 mL), stirred, and then HATU (19.90 g, 52.35 mmol) and DIPEA (9.03 g, 69.80 mmol) are added. The reaction solution is naturally warmed to room temperature and stirred for reaction for 6.5 hours. After the reaction is completed, 65 mL of water is added to the reaction solution for washing, and then ethyl acetate (2x65 mL) is added for extraction. After the organic phases are combined, 65 mL of saturated brine is added for washing, and the organic phases are dried over anhydrous sodium sulfate. The filtrate is concentrated under reduced pressure to evaporate the solvent, and the concentrate is purified by silica gel column chromatography using a petroleum ether / ethyl acetate mixed solvent with a volume ratio of 4:1 to obtain a compound represented by formula 4, with an amount of 9.14 g and a yield of 76.6%.

[0031] According to an embodiment of the present invention, in step (3), the contacting method of the compound represented by Formula 4 and LiOH is not particularly limited. Thus, the efficiency of the contact reaction between the compound represented by Formula 4 and LiOH can be improved, the reaction speed can be accelerated, and the efficiency of preparing the compound represented by Formula 5 by using this method can be further improved.

[0032] According to an embodiment of the present invention, in step (3), the following steps are included: in a reaction container, THF and H2O are mixed in a volume ratio of 1:1, and then the compound shown in Formula 4 is added, the temperature is cooled to 0°C to 5°C with stirring, LiOH is added, and the reaction solution is naturally heated to room temperature, and then stirred for reaction for 3 hours to 4 hours. TLC shows that the compound shown in Formula 1 is completely reacted, and the reaction solution is concentrated under reduced pressure to remove THF. The remaining liquid is added with ether to extract the water layer, the water layer is cooled to 0°C, and HCl solution is added to adjust the pH value to pH 3. Ethyl acetate is added to the acidified aqueous phase for extraction, and the organic phases are combined, washed with saturated brine, dried over anhydrous sodium sulfate, and the filtrate is concentrated to dryness under reduced pressure to obtain the compound shown in Formula 5.

[0033] According to an embodiment of the present invention, in step (3), the molar ratio of the compound represented by Formula 4 to LiOH is 1:(1.5-3.5), preferably the molar ratio of the compound represented by Formula 4 to LiOH is 1:2.2. Thus, the efficiency of preparing the compound represented by Formula 5 by this method can be further improved.

[0034] According to an embodiment of the present invention, in step (3), the stirring reaction time is preferably 3.5 hours.

[0035] According to a specific embodiment of the present invention, in step (3), the following steps are included: in a reaction container, THF (400 mL) and H2O (400 mL) are mixed, and then the compound shown in Formula 4 (80.0 g, 234.1 mmol) is added, and the temperature is cooled to 0°C to 5°C with stirring, and LiOH (12.33 g, 515.0 mmol) is added. After the reaction solution is naturally warmed to room temperature, the reaction is stirred for 3.5 hours. TLC shows that the compound shown in Formula 1 is completely reacted, and the reaction solution is concentrated under reduced pressure to remove THF. Ether (2x400 mL) is added to the residue to extract the water layer, and the water layer is cooled to 0°C, and 3M HCl solution is added to adjust the pH value to pH 3. Ethyl acetate (2x400 mL) is added to the acidified aqueous phase for extraction. After the organic phases are combined, 400 mL of saturated brine is added for washing, and the filtrate is dried over anhydrous sodium sulfate. The filtrate is concentrated to dryness under reduced pressure to obtain the compound shown in Formula 5, with an amount of 64.52 g and a yield of 84.1%.

[0036] According to an embodiment of the present invention, in step (4), the contacting method of the compound represented by Formula 5 with the compound represented by Formula 6, HATU, and DIPEA is not particularly limited. Thus, the efficiency of the contact reaction between the compound represented by Formula 5 and the compound represented by Formula 6, HATU, and DIPEA can be improved, the reaction speed can be accelerated, and the efficiency of preparing the compound represented by Formula I by using this method can be further improved.

[0037] According to an embodiment of the present invention, in step (4), the following steps are included: adding the compound of formula 5 and the compound of formula 6 to DMF at 0°C, stirring, then adding HATU and DIPEA, stirring and reacting the reaction solution after the reaction solution naturally rises to room temperature for 6 to 7.5 hours, adding water to the reaction solution for washing, then adding ethyl acetate for extraction, adding saturated brine to the organic phase after merging, washing, drying over anhydrous sodium sulfate, concentrating the filtrate under reduced pressure to evaporate the solvent, and purifying the concentrate by silica gel column chromatography with a petroleum ether / ethyl acetate mixed solvent to obtain the compound of formula I. Thus, the efficiency of the contact reaction between the compound of formula 5 and the compound of formula 6, HATU, and DIPEA can be improved, the reaction speed can be accelerated, and the efficiency of preparing the compound of formula I by using this method can be further improved.

[0038] According to an embodiment of the present invention, in step (4), the molar ratio of the compound represented by Formula 5 to the compound represented by Formula 6, HATU, and DIPEA is 1:(1.0-1.2):(1.3-2.5):(1.6-3.0), and preferably the molar ratio of the compound represented by Formula 5 to the compound represented by Formula 6, HATU, and DIPEA is 1:1.05:1.6:2.2. Thus, the efficiency of preparing the compound represented by Formula I by this method can be further improved.

[0039] According to an embodiment of the present invention, in step (4), the volume ratio of petroleum ether to ethyl acetate in the petroleum ether / ethyl acetate mixed solvent is (3-6):1, and preferably the volume ratio of petroleum ether / ethyl acetate mixed solvent is 4:1.

[0040] According to an embodiment of the present invention, in step (4), the reaction solution is preferably stirred and reacted for 6.5 hours after the reaction solution is naturally heated to room temperature.

[0041] According to a specific embodiment of the present invention, in step (4), the following steps are included: at 0°C, the compound represented by formula 5 (5.50 g, 16.78 mmol) and the compound represented by formula 6 (3.46 g, 17.62 mmol) are added to DMF (60 mL), stirred, and then HATU (10.21 g, 26.85 mmol) and DIPEA (4.77 g, 36.92 mmol) are added. The reaction solution is naturally warmed to room temperature and stirred for reaction for 6.5 hours. After the reaction is completed, 60 mL of water is added to the reaction solution for washing, and then ethyl acetate (2x60 mL) is added for extraction. After the organic phases are combined, 60 mL of saturated brine is added for washing, and the organic phases are dried over anhydrous sodium sulfate. The filtrate is concentrated under reduced pressure to evaporate the solvent, and the concentrate is purified by silica gel column chromatography using a petroleum ether / ethyl acetate mixed solvent with a volume ratio of 4:1 to obtain a compound represented by formula 7, with an amount of 7.14 g, a yield of 84.0%, a purity of 99.6% (HPLC), and an optical purity of 99.4% (chiral HPLC).

[0042] According to a specific embodiment of the present invention, the synthesis route of the compound Tovorafenib represented by Formula I can be as follows:

[0043]

[0044] Compared with the prior art, the pharmaceutical synthesis process of Tovorafenib described in the present invention has at least the following beneficial effects:

[0045] 1. Compared with the prior art, the method described in the present invention uses a prochiral ketone compound (compound shown in Formula 1) as a raw material, isopropylamine as an amine donor, water as a solvent, DMSO as a cosolvent, transaminase (ATASK-000250) as a biocatalyst, and PLA (pyridoxal phosphate) as a coenzyme regeneration system to reduce and aminize the ketone in one step to form an optically pure compound shown in Formula 2 (a chiral α-heteroamine compound), and then the compound shown in Formula 2 is subjected to a condensation reaction with the compound shown in Formula 3 to obtain a compound shown in Formula 4, and the compound shown in Formula 4 is then hydrolyzed under an alkaline environment to obtain a corresponding carboxylic acid compound (compound shown in Formula 5), ​​and then the compound shown in Formula 5 is subjected to a condensation reaction with the compound shown in Formula 6 to obtain the product compound shown in Formula I, Tovorafenib.

[0046] 2. Compared with the prior art methods, the present invention has the following significant advantages: (1) The most difficult step in the synthesis of Tovorafenib is to construct chiral α-heteroamines. The commonly used method is to synthesize such chiral (hetero) aromatic amines through asymmetric hydroaminoalkylation of olefins and asymmetric hydrogenation of the formed N-containing intermediates (such as amides, amides or imines), such as CN111032047A. The chemical catalysts for catalyzing such reactions are mostly transition metals, which are expensive, highly polluting, and sensitive to water and air. In addition, the coordination effect of heteroatoms and the easy degradation of heteroaromatic rings lead to cumbersome steps in the synthesis process, harsh reaction conditions, difficult product purification, and low yield. The synthesis process of the present invention relates to a method for preparing chiral α-heteroamines, which utilizes a biocatalyst amine dehydrogenase to directly reduce and aminize a potential chiral ketone into an optically pure chiral α-heteroamine in one step under mild conditions. The process overcomes the shortcomings of existing catalytic technologies, has a simple process route, mild reaction conditions, high yield, and has the advantages of green synthesis such as step economy and atom economy, as well as good stereoselectivity (ee value up to 99%). (2) The synthesis route mostly uses a method of condensing amino groups with carboxyl groups in HATU and an alkaline environment, which is simple to operate, has a high reaction yield, and is suitable for mass production.

[0047] 3. In summary, this route has the advantages of low raw material cost, high yield, simple purification, etc. It is easy to industrialize and is very suitable for large-scale industrial production. DETAILED DESCRIPTION

[0048] The embodiments of the present invention are described in detail below. The embodiments described below are exemplary and are only used to explain the present invention, and should not be construed as limiting the present invention. If no specific techniques or conditions are specified in the embodiments, the techniques or conditions described in the literature in this area or the product specifications are used. The reagents or instruments used that do not specify the manufacturer are all conventional products that can be obtained commercially.

[0049] Example 1 Synthesis of the compound represented by formula 2

[0050] The compound of formula 1 (10.0 g, 54.0 mmol) was dissolved in DMSO (100 mL) and placed in a 5 L reaction bottle. A phosphate buffer solution (2106 mL, 0.25 mM pyridoxal phosphate, 1 M isopropylamine) with a pH of 7.5 and a concentration of 0.1 M and a transaminase (1.5 g, purchased from Codex Amine Transaminase Screening Kit ATASK-000250) were added. The reaction was shaken at 30 ° C and 250 rpm for 48 hours. 10 M of The reaction was stopped by stirring with NaOH solution (1736 mL), and the reaction solution was extracted with ethyl acetate (2×2 L). The mixed solution was filtered and the organic layer was separated, washed with saturated brine and dried over anhydrous sodium sulfate. The filtrate was concentrated under reduced pressure to evaporate the solvent. The concentrate was purified by silica gel column chromatography using a mixed solvent of petroleum ether / ethyl acetate in a volume ratio of 10:1 to obtain the compound of formula 2, with an amount of 7.40 g, a yield of 73.6%, a purity of 99.5% (HPLC), and an optical purity of 99.3% (chiral HPLC).

[0051] LC-MS (APCI): m / z = 187.1 (M+1) + .

[0052] 1 H NMR (400MHz, CDCl3) δ8.32(d,J=2.3Hz,1H),4.44(td,J=6.7,1.9Hz,1H),3.92(s,3H),1.92(s,2H),1.57(dd,J=6.8,1.9Hz,3H).

[0053] Example 2 Synthesis of the compound represented by formula 4

[0054] At 0°C, the compound represented by formula 2 (6.5 g, 34.90 mmol) and the compound represented by formula 3 (6.18 g, 35.60 mmol) were added to DMF (65 mL) and stirred. HATU (19.90 g, 52.35 mmol) and DIPEA (9.03 g, 69.80 mmol) were added. The reaction solution was naturally warmed to room temperature and stirred for 6.5 hours. After the reaction was completed, 65 mL of water was added to the reaction solution for washing, and ethyl acetate (2x65 mL) was added for extraction. The organic phases were combined and washed with 65 mL of saturated brine, dried over anhydrous sodium sulfate, and the filtrate was concentrated under reduced pressure to evaporate the solvent. The concentrate was purified by silica gel column chromatography using a petroleum ether / ethyl acetate mixed solvent with a volume ratio of 4:1 to obtain the compound represented by formula 4, with an amount of 9.14 g and a yield of 76.6%.

[0055] LC-MS (APCI): m / z = 342.0 (M+1) + .

[0056] Example 3 Synthesis of the compound represented by formula 4

[0057] At 0°C, the compound represented by formula 2 (6.5 g, 34.90 mmol) and the compound represented by formula 3 (6.06 g, 34.90 mmol) were added to DMF (65 mL) and stirred. HATU (17.25 g, 45.37 mmol) and DIPEA (6.77 g, 52.38 mmol) were added. The reaction solution was naturally warmed to room temperature and stirred for 6 hours. After the reaction was completed, 65 mL of water was added to the reaction solution for washing, and ethyl acetate (2x65 mL) was added for extraction. The organic phases were combined and washed with 65 mL of saturated brine, dried over anhydrous sodium sulfate, and the filtrate was concentrated under reduced pressure to evaporate the solvent. The concentrate was purified by silica gel column chromatography using a petroleum ether / ethyl acetate mixed solvent with a volume ratio of 3:1 to obtain the compound represented by formula 4, with an amount of 8.90 g and a yield of 74.6%.

[0058] Example 4 Synthesis of the compound represented by formula 4

[0059] At 0°C, the compound represented by formula 2 (6.5 g, 34.90 mmol) and the compound represented by formula 3 (7.27 g, 41.88 mmol) were added to DMF (80 mL) and stirred, and HATU (29.19 g, 76.78 mmol) and DIPEA (15.79 g, 122.15 mmol) were added. The reaction solution was naturally warmed to room temperature and stirred for 8 hours. After the reaction was completed, 80 mL of water was added to the reaction solution for washing, and then ethyl acetate (2x80 mL) was added for extraction. The organic phases were combined and washed with 80 mL of saturated brine, dried over anhydrous sodium sulfate, and the filtrate was concentrated under reduced pressure to evaporate the solvent. The concentrate was purified by silica gel column chromatography using a petroleum ether / ethyl acetate mixed solvent with a volume ratio of 6:1 to obtain the compound represented by formula 4, with an amount of 9.08 g and a yield of 76.1%.

[0060] Comparative Example 1 Synthesis of the Compound Represented by Formula 4

[0061] At 0°C, the compound represented by formula 2 (6.5 g, 34.90 mmol) and the compound represented by formula 3 (7.87 g, 45.37 mmol) were added to DMF (65 mL) and stirred. HATU (17.25 g, 45.37 mmol) and DIPEA (5.86 g, 45.37 mmol) were added. The reaction solution was naturally warmed to room temperature and stirred for 9 hours. After the reaction was completed, 65 mL of water was added to the reaction solution for washing, and ethyl acetate (2x65 mL) was added for extraction. The organic phases were combined and washed with 65 mL of saturated brine, dried over anhydrous sodium sulfate, and the filtrate was concentrated under reduced pressure to evaporate the solvent. The concentrate was purified by silica gel column chromatography using a petroleum ether / ethyl acetate mixed solvent with a volume ratio of 4:1 to obtain the compound represented by formula 4, with an amount of 8.85 g and a yield of 74.2%.

[0062] It can be seen that changing the molar ratio of the compound represented by Formula 2 to the compound represented by Formula 3, HATU, and DIPEA to 1:1.3:1.3:1.3 and adjusting the stirring reaction to 9 hours did not further improve the efficiency (yield) of preparing the compound represented by Formula 4.

[0063] Comparative Example 2 Synthesis of the Compound Represented by Formula 4

[0064] At 0°C, the compound represented by formula 2 (6.5 g, 34.90 mmol) and the compound represented by formula 3 (5.81 g, 33.50 mmol) were added to DMF (100 mL) and stirred. HATU (33.18 g, 87.25 mmol) and DIPEA (18.04 g, 139.6 mmol) were added. The reaction solution was naturally warmed to room temperature and stirred for reaction for 5.5 hours. After the reaction was completed, 100 mL of water was added to the reaction solution for washing, and then ethyl acetate (2x100 mL) was added for extraction. The organic phases were combined and washed with 100 mL of saturated brine, dried over anhydrous sodium sulfate, and the filtrate was concentrated under reduced pressure to evaporate the solvent. The concentrate was purified by silica gel column chromatography using a petroleum ether / ethyl acetate mixed solvent with a volume ratio of 4:1 to obtain the compound represented by formula 4, with an amount of 8.36 g and a yield of 73.0%.

[0065] It can be seen that changing the molar ratio of the compound represented by Formula 2 to the compound represented by Formula 3, HATU, and DIPEA to 1:0.96:2.5:4.0 and adjusting the stirring reaction to 5.5 hours did not further improve the efficiency (yield) of preparing the compound represented by Formula 4.

[0066] Example 5 Synthesis of the compound represented by formula 5

[0067] In a reaction vessel, THF (400 mL) and H2O (400 mL) were mixed, and then the compound shown in Formula 4 (80.0 g, 234.1 mmol) was added, and the temperature was cooled to 0°C to 5°C with stirring, and LiOH (12.33 g, 515.0 mmol) was added. After the reaction solution naturally warmed to room temperature, the reaction was stirred for 3.5 hours. TLC showed that the compound shown in Formula 1 was completely reacted. The reaction solution was concentrated under reduced pressure to remove THF, and ether (2x400 mL) was added to the residue to extract the water layer. The water layer was cooled to 0°C, and 3M HCl solution was added to adjust the pH value to pH 3. Ethyl acetate (2x400 mL) was added to the acidified aqueous phase for extraction. After the organic phases were combined, 400 mL of saturated brine was added for washing, dried over anhydrous sodium sulfate, and the filtrate was concentrated to dryness under reduced pressure to obtain the compound shown in Formula 5, with an amount of 64.52 g and a yield of 84.1%.

[0068] LC-MS (APCI): m / z = 328.0 (M+1) + .

[0069] Example 6 Synthesis of the compound represented by formula 5

[0070] In a reaction vessel, THF (400 mL) and H2O (400 mL) were mixed, and then the compound shown in Formula 4 (80.0 g, 234.1 mmol) was added, and the temperature was cooled to 0°C to 5°C with stirring, and LiOH (8.41 g, 351.15 mmol) was added. After the reaction solution naturally warmed to room temperature, the reaction was stirred for 3 hours. TLC showed that the compound shown in Formula 1 was completely reacted. The reaction solution was concentrated under reduced pressure to remove THF, and ether (2x400 mL) was added to the residue to extract the water layer. The water layer was cooled to 0°C, and 3M HCl solution was added to adjust the pH value to pH 3. Ethyl acetate (2x400 mL) was added to the acidified aqueous phase for extraction. After the organic phases were combined, 400 mL of saturated brine was added for washing, dried over anhydrous sodium sulfate, and the filtrate was concentrated to dryness under reduced pressure to obtain the compound shown in Formula 5, with an amount of 62.91 g and a yield of 82.0%.

[0071] Example 7 Synthesis of the compound represented by formula 5

[0072] In a reaction vessel, THF (400 mL) and H2O (400 mL) were mixed, and then the compound represented by Formula 4 (80.0 g, 234.1 mmol) was added, and the temperature was cooled to 0°C to 5°C with stirring, and LiOH (19.62 g, 819.35 mmol) was added. After the reaction solution naturally warmed to room temperature, the reaction was stirred for 4 hours. TLC showed that the compound represented by Formula 1 was completely reacted. The reaction solution was concentrated under reduced pressure to remove THF, and ether (2x400 mL) was added to the residue to extract the water layer. The water layer was cooled to 0°C, and 3M HCl solution was added to adjust the pH value to pH 3. Ethyl acetate (2x400 mL) was added to the acidified aqueous phase for extraction. After the organic phases were combined, 400 mL of saturated brine was added for washing, dried over anhydrous sodium sulfate, and the filtrate was concentrated to dryness under reduced pressure to obtain the compound represented by Formula 5, with an amount of 63.99 g and a yield of 83.4%.

[0073] Example 8 Synthesis of Tovorafenib, a compound represented by Formula I

[0074] At 0°C, the compound represented by formula 5 (5.50 g, 16.78 mmol) and the compound represented by formula 6 (3.46 g, 17.62 mmol) were added to DMF (60 mL) and stirred. HATU (10.21 g, 26.85 mmol) and DIPEA (4.77 g, 36.92 mmol) were added. The reaction solution was naturally warmed to room temperature and stirred for 6.5 hours. After the reaction was completed, 60 mL of water was added to the reaction solution for washing, and then ethyl acetate (2x60 mL) was added for extraction. The organic phases were combined and washed with 60 mL of saturated brine, dried over anhydrous sodium sulfate, and the filtrate was concentrated under reduced pressure to evaporate the solvent. The concentrate was purified by silica gel column chromatography using a petroleum ether / ethyl acetate mixed solvent with a volume ratio of 4:1 to obtain Tovorafenib, a compound represented by formula I, with an amount of 7.14 g, a yield of 84.0%, a purity of 99.6% (HPLC), and an optical purity of 99.4% (chiral HPLC).

[0075] LC-MS (APCI): m / z = 505.6 (M+1) + .

[0076] 1 H NMR(400MHz,DMSO-d6)d ppm 11.88(s,1H)9.63(d,J=7.82Hz,1H)8.79(s,1H)8.68(s,1H)8.59(s,1H)8.38(s,1H)7.86(b r.s.,1H)7.48(br.s.,1H)5.37(tdd,J=7.27,7.27,7.21,7.09Hz,1H)1.61(d,J=6.85Hz,3H)

[0077] Example 9 Synthesis of the compound Tovorafenib represented by formula I

[0078] At 0°C, the compound represented by formula 5 (5.5 g, 16.78 mmol) and the compound represented by formula 6 (3.3 g, 16.78 mmol) were added to DMF (60 mL) and stirred. HATU (8.29 g, 21.81 mmol) and DIPEA (3.47 g, 26.85 mmol) were added. The reaction solution was naturally warmed to room temperature and stirred for 6 hours. After the reaction was completed, 60 mL of water was added to the reaction solution for washing, and then ethyl acetate (2x60 mL) was added for extraction. The organic phases were combined and washed with 60 mL of saturated brine, dried over anhydrous sodium sulfate, and the filtrate was concentrated under reduced pressure to evaporate the solvent. The concentrate was purified by silica gel column chromatography using a mixed solvent of petroleum ether / ethyl acetate with a volume ratio of 3:1 to obtain Tovorafenib, a compound represented by formula I, with an amount of 6.92 g, a yield of 81.5%, a purity of 99.5% (HPLC), and an optical purity of 99.2% (chiral HPLC).

[0079] Example 10 Synthesis of Tovorafenib, a compound represented by Formula I

[0080] At 0°C, the compound represented by formula 5 (5.5 g, 16.78 mmol) and the compound represented by formula 6 (3.96 g, 20.136 mmol) were added to DMF (80 mL) and stirred. HATU (15.95 g, 41.95 mmol) and DIPEA (6.51 g, 50.37 mmol) were added. The reaction solution was naturally warmed to room temperature and stirred for 7.5 hours. After the reaction was completed, 80 mL of water was added to the reaction solution for washing, and then ethyl acetate (2x80 mL) was added for extraction. The organic phases were combined and washed with 80 mL of saturated brine, dried over anhydrous sodium sulfate, and the filtrate was concentrated under reduced pressure to evaporate the solvent. The concentrate was purified by silica gel column chromatography using a mixed solvent of petroleum ether / ethyl acetate with a volume ratio of 6:1 to obtain Tovorafenib, a compound represented by formula I, with an amount of 7.03 g, a yield of 82.8%, a purity of 99.4% (HPLC), and an optical purity of 99.5% (chiral HPLC).

[0081] Comparative Example 3 Synthesis of the Compound Tovorafenib Represented by Formula I

[0082] At 0°C, the compound represented by formula 5 (5.5 g, 16.78 mmol) and the compound represented by formula 6 (3.23 g, 16.44 mmol) were added to DMF (60 mL) and stirred. HATU (7.66 g, 20.136 mmol) and DIPEA (2.82 g, 21.81 mmol) were added. The reaction solution was naturally warmed to room temperature and stirred for 6 hours. After the reaction was completed, 80 mL of water was added to the reaction solution for washing, and then ethyl acetate (2x80 mL) was added for extraction. The organic phases were combined and washed with 80 mL of saturated brine, dried over anhydrous sodium sulfate, and the filtrate was concentrated under reduced pressure to evaporate the solvent. The concentrate was purified by silica gel column chromatography using a petroleum ether / ethyl acetate mixed solvent with a volume ratio of 4:1 to obtain Tovorafenib, a compound represented by formula I, with an amount of 6.71 g, a yield of 80.6%, a purity of 98.6% (HPLC), and an optical purity of 97.8% (chiral HPLC).

[0083] It can be seen that changing the molar ratio of the compound represented by Formula 5 to the compound represented by Formula 6, HATU, and DIPEA to 1:0.98:1.2:1.3 does not further improve the yield of the compound represented by Formula I, but the purity of the compound represented by Formula I is reduced.

[0084] Comparative Example 4 Synthesis of the Compound Tovorafenib Represented by Formula I

[0085] At 0°C, the compound represented by formula 5 (5.5 g, 16.78 mmol) and the compound represented by formula 6 (4.62 g, 23.49 mmol) were added to DMF (100 mL) and stirred, and HATU (19.14 g, 50.34 mmol) and DIPEA (7.59 g, 58.73 mmol) were added. The reaction solution was naturally warmed to room temperature and stirred for 8 hours. After the reaction was completed, 80 mL of water was added to the reaction solution for washing, and then ethyl acetate (2x80 mL) was added for extraction. The organic phases were combined and washed with 80 mL of saturated brine, dried over anhydrous sodium sulfate, and the filtrate was concentrated under reduced pressure to evaporate the solvent. The concentrate was purified by silica gel column chromatography using a mixed solvent of petroleum ether / ethyl acetate with a volume ratio of 4:1 to obtain Tovorafenib, a compound represented by formula I, with an amount of 6.89 g, a yield of 81.1%, a purity of 99.1% (HPLC), and an optical purity of 98.9% (chiral HPLC).

[0086] It can be seen that changing the molar ratio of the compound represented by Formula 5 to the compound represented by Formula 6, HATU, and DIPEA to 1:1.4:3.0:3.5 and stirring the reaction for 8 hours did not further improve the yield of the compound represented by Formula I, but the purity of the compound represented by Formula I was reduced.

[0087] In the description of this specification, the description with reference to the terms "one embodiment", "some embodiments", "example", "specific example", or "some examples" etc. means that the specific features, structures, materials or characteristics described in conjunction with the embodiment or example are included in at least one embodiment or example of the present invention. In this specification, the schematic representations of the above terms do not necessarily refer to the same embodiment or example. Moreover, the specific features, structures, materials or characteristics described may be combined in any one or more embodiments or examples in a suitable manner. In addition, those skilled in the art may combine and combine the different embodiments or examples described in this specification and the features of the different embodiments or examples, without contradiction.

[0088] Although the embodiments of the present invention have been shown and described above, it is to be understood that the above embodiments are exemplary and are not to be construed as limitations of the present invention. A person skilled in the art may change, modify, replace and vary the above embodiments within the scope of the present invention.

Claims

1. A pharmaceutical synthesis process of Tovorafenib, characterized in that: include: (1) contacting the compound represented by Formula 1 with a transaminase to obtain the compound represented by Formula 2; (2) contacting the compound represented by Formula 2 with the compound represented by Formula 3, HATU, and DIPEA to obtain the compound represented by Formula 4; (3) contacting the compound represented by Formula 4 with LiOH to obtain the compound represented by Formula 5; (4) contacting the compound represented by Formula 5 with the compound represented by Formula 6, HATU and DIPEA to obtain the compound represented by Formula I, Tovorafenib, 2. The method according to claim 1, characterized in that In step (1), the following steps are included: dissolving the compound shown in formula 1 in DMSO, placing the mixture in a reaction bottle, adding a phosphate buffer having a pH of 7.5 and a concentration of 0.1 M and a transaminase, shaking the mixture at 30° C. and 250 rpm for 48 hours, adding a 10 M NaOH solution to the reaction solution in an ice-water bath and stirring to stop the reaction, extracting the reaction solution with ethyl acetate, filtering the mixed solution and separating the organic layer, washing the mixture with saturated brine, and drying the mixture with anhydrous sodium sulfate, concentrating the filtrate under reduced pressure to evaporate the solvent, and purifying the concentrate by silica gel column chromatography with a petroleum ether / ethyl acetate mixed solvent having a volume ratio of 10:1 to obtain the compound shown in formula 2.

3. The method according to claim 2, characterized in that In step (1), the transaminase is purchased from Codex Amine Transaminase Screening Kit ATASK-000250; Optionally, in step (1), the phosphate buffer consists of 0.25 mM pyridoxal phosphate and 1 M isopropylamine; Optionally, in step (1), the weight ratio (g / g) of the compound represented by formula 1 to the transaminase is 1:0.

15.

4. The method according to claim 1, characterized in that: In step (2), the following steps are included: adding the compound represented by formula 2 and the compound represented by formula 3 to DMF at 0°C, stirring, then adding HATU and DIPEA, and stirring the reaction solution after the reaction solution is naturally heated to room temperature for 6 to 8 hours. After the reaction is completed, water is added to the reaction solution for washing, and then ethyl acetate is added for extraction. After the organic phases are combined, saturated brine is added for washing, and the organic phases are dried over anhydrous sodium sulfate. The filtrate is concentrated under reduced pressure to evaporate the solvent, and the concentrate is purified by silica gel column chromatography using a petroleum ether / ethyl acetate mixed solvent to obtain a compound represented by formula 4.

5. The method according to claim 4, characterized in that In step (2), the molar ratio of the compound represented by formula 2 to the compound represented by formula 3, HATU, and DIPEA is 1:(1.0-1.2):(1.3-2.2):(1.5-3.5), preferably the molar ratio of the compound represented by formula 2 to the compound represented by formula 3, HATU, and DIPEA is 1:1.02:1.5:2.0; Optionally, in step (2), the volume ratio of petroleum ether to ethyl acetate in the petroleum ether / ethyl acetate mixed solvent is (3-6):1, preferably the volume ratio of petroleum ether / ethyl acetate mixed solvent is 4:1; Optionally, in step (2), the reaction solution is preferably allowed to naturally warm to room temperature and then stirred for reaction for 6.5 hours.

6. The method according to claim 1, characterized in that In step (3), the following steps are included: THF and H2O are mixed in a volume ratio of 1:1, and then the compound shown in formula 4 is added, the temperature is cooled to 0°C to 5°C with stirring, LiOH is added, the reaction solution is naturally warmed to room temperature, and the reaction is stirred for 3 hours to 4 hours. TLC shows that the compound shown in formula 1 is completely reacted, the reaction solution is concentrated under reduced pressure, THF is removed, and the remaining liquid is added with ether to extract the water layer, the water layer is cooled to 0°C, HCl solution is added to adjust the pH value to pH 3, ethyl acetate is added to the acidified aqueous phase for extraction, the organic phases are combined, saturated brine is added for washing, anhydrous sodium sulfate is dried, and the filtrate is concentrated to dryness under reduced pressure to obtain the compound shown in formula 5.

7. The method according to claim 6, characterized in that In step (3), the molar ratio of the compound represented by formula 4 to LiOH is 1:(1.5-3.5), preferably the molar ratio of the compound represented by formula 4 to LiOH is 1:2.2; Optionally, in step (3), the stirring reaction time is preferably 3.5 hours.

8. The method according to claim 1, characterized in that In step (4), the following steps are included: adding the compound represented by formula 5 and the compound represented by formula 6 to DMF at 0°C, stirring, then adding HATU and DIPEA, and stirring the reaction solution after the reaction solution is naturally warmed to room temperature for 6 to 7.5 hours, adding water to the reaction solution for washing, and then adding ethyl acetate for extraction, combining the organic phases, adding saturated brine for washing, and drying over anhydrous sodium sulfate. The filtrate is concentrated under reduced pressure to evaporate the solvent, and the concentrate is purified by silica gel column chromatography using a petroleum ether / ethyl acetate mixed solvent to obtain the compound represented by formula I.

9. The method according to claim 8, characterized in that In step (4), the molar ratio of the compound represented by Formula 5 to the compound represented by Formula 6, HATU, and DIPEA is 1:(1.0-1.2):(1.3-2.5):(1.6-3.0), preferably the molar ratio of the compound represented by Formula 5 to the compound represented by Formula 6, HATU, and DIPEA is 1:1.05:1.6:2.2; Optionally, in step (4), the volume ratio of petroleum ether to ethyl acetate in the petroleum ether / ethyl acetate mixed solvent is (3-6):1, preferably the volume ratio of petroleum ether / ethyl acetate mixed solvent is 4:1; Optionally, in step (4), the reaction solution is preferably allowed to naturally warm to room temperature and then stirred for 6.5 hours.

10. The method according to claim 1, characterized in that In step (1), the following steps are included: dissolving the compound represented by formula 1 (10.0 g, 54.0 mmol) in DMSO (100 mL), placing the mixture in a 5 L reaction bottle, adding phosphate buffer (2106 mL, 0.25 mM pyridoxal phosphate, 1 M isopropylamine) with a pH of 7.5 and a concentration of 0.1 M and transaminase (1.5 g, purchased from Codex Amine Transaminase Screening Kit ATASK-000250), shaking the mixture at 30° C. and 250 rpm for 48 hours, and placing the reaction solution in an ice water bath. 10M NaOH solution (1736mL) was added under stirring to stop the reaction, the reaction solution was extracted with ethyl acetate (2×2L), the mixed solution was filtered and the organic layer was separated, saturated brine was added for washing, and dried over anhydrous sodium sulfate, the filtrate was concentrated under reduced pressure to evaporate the solvent, and the concentrate was purified by silica gel column chromatography with a mixed solvent of petroleum ether / ethyl acetate in a volume ratio of 10:1 to obtain the compound shown in formula 2, with an amount of 7.4g, a yield of 73.6%, a purity of 99.5% (HPLC), and an optical purity of 99.3% (chiral HPLC); In step (2), the following steps are included: at 0°C, the compound represented by formula 2 (6.5 g, 34.90 mmol) and the compound represented by formula 3 (6.18 g, 35.60 mmol) are added to DMF (65 mL), stirred, and then HATU (19.90 g, 52.35 mmol) and DIPEA (9.03 g, 69.80 mmol) are added. The reaction solution is naturally warmed to room temperature and stirred for reaction for 6.5 hours. After the reaction is completed, 65 mL of water is added to the reaction solution for washing, and then ethyl acetate (2x65 mL) is added for extraction. After the organic phases are combined, 65 mL of saturated brine is added for washing, and the organic phases are dried over anhydrous sodium sulfate. The filtrate is concentrated under reduced pressure to evaporate the solvent, and the concentrate is purified by silica gel column chromatography using a petroleum ether / ethyl acetate mixed solvent with a volume ratio of 4:1 to obtain the compound represented by formula 4, with an amount of 9.14 g and a yield of 76.6%; In step (3), the following steps are included: in a reaction container, THF (400 mL) and H2O (400 mL) are mixed, and then the compound shown in formula 4 (80.0 g, 234.1 mmol) is added, the temperature is cooled to 0°C to 5°C with stirring, LiOH (12.33 g, 515.0 mmol) is added, the reaction solution is naturally warmed to room temperature, and the reaction is stirred for 3.5 hours. TLC shows that the compound shown in formula 1 is completely reacted, the reaction solution is concentrated under reduced pressure, THF is removed, ether (2x400 mL) is added to the residue to extract the water layer, the water layer is cooled to 0°C, 3M HCl solution is added to adjust the pH value to pH 3, ethyl acetate (2x400 mL) is added to the acidified aqueous phase for extraction, the organic phases are combined, 400 mL of saturated brine is added for washing, anhydrous sodium sulfate is dried, and the filtrate is concentrated to dryness under reduced pressure to obtain the compound shown in formula 5, with an amount of 64.52 g and a yield of 84.1%; In step (4), the following steps are included: adding the compound represented by formula 5 (5.50 g, 16.78 mmol) and the compound represented by formula 6 (3.46 g, 17.62 mmol) to DMF (60 mL) at 0°C, stirring, then adding HATU (10.21 g, 26.85 mmol) and DIPEA (4.77 g, 36.92 mmol), the reaction solution is naturally warmed to room temperature and stirred for reaction for 6.5 hours, after the reaction is completed, 60 mL of water is added to the reaction solution for washing, and then ethyl acetate (2x60 mL) is added for extraction, the organic phases are combined and then 60 mL of saturated brine is added for washing, and dried over anhydrous sodium sulfate, the filtrate is concentrated under reduced pressure to evaporate the solvent, and the concentrate is purified by silica gel column chromatography using a petroleum ether / ethyl acetate mixed solvent with a volume ratio of 4:1 to obtain the compound represented by formula 7, with an amount of 7.14 g, a yield of 84.0%, a purity of 99.6% (HPLC), and an optical purity of 99.4% (chiral HPLC).

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