JP7636788B2 - Method for synthesizing small molecule compound BRD0539 - Google Patents

Description

This invention relates to a novel method for synthesizing the small molecule compound BRD0539, an inhibitor of CRISPR-Cas9.

CRISPR-Cas9 was discovered in bacteria and archaea as an adaptive immune system of microorganisms that confers acquired immunity to foreign viruses and plasmids, and is an immune mechanism in which, when a virus invades a bacterium, the bacterium incorporates the viral gene into its own genes and memorizes it, and if the same virus invades again, it cuts and eliminates the gene (Non-Patent Document 1, etc.). By applying this, it is possible to recognize any base sequence in DNA and delete or replace that part, and it has attracted attention as a genome editing technology that is more accurate and simpler than conventional methods (Non-Patent Documents 2, 3, etc.).
Meanwhile, in 2019, a small molecule compound BRD0539 (Chemical 1), which acts as an inhibitor of CRISPR-Cas9, was discovered (Non-Patent Document 4). Using this inhibitor, BRD0539, it is possible to more precisely control the activity of CRISPR-Cas9.

The synthetic route for BRD0539 has already been reported (Non-Patent Document 5, Patent Document 1, etc.).
The outline of the first reported synthesis method of BRD0539 (Non-Patent Document 5) is shown in the following formula (Chemical Formula 4). That is, 4-bromoaniline is used as a starting material, and tricyclic compound 2 is synthesized by Povarov reaction, which is then converted to compound 3 by reduction. The conversion reactions after compound 3 (deprotection of Fmoc group, N-tosylation (sulfonamidation), Suzuki-Miyaura coupling) are carried out by solid-phase synthesis. In this synthesis method, the hydroxyl group is protected by being fixed to a polymer (Si) in the conversion after compound 3.

In a method reported later (Patent Document 1), as shown in the following formula (Chemical Formula 5), based on the above synthetic route, a compound corresponding to compound 3 is obtained as an N-Boc compound 3' (93% ee), and then a compound in which the hydroxy group is protected with a 3,3-dimethylbutyryl (DMB) group is used for subsequent transformations (removal of Boc, tosylation, Suzuki-Miyaura coupling, removal of DMB). In this synthetic method, the hydroxy group is protected with a DMB group and the subsequent transformations are carried out.

International Publication WO2020/068304

Science, 2007, 315, 1709-1712. Science, 2012, 337, 816-821. Nature Biotechnology, 2014, 32, 347-355. Cell, 2019, 177, 1067-1079.e19. ACS Comb. Sci. 2012, 14, 621-630. (Corrected in ACS Comb. Sci. 2014, 16, 46-46.)

Since the use of the CRISPR-Cas9 inhibitor BRD0539 allows for more precise control of CRISPR-Cas9 activity, if the synthesis of BRD0539 becomes easier and more readily available, this synthesis method will be highly useful.
However, all of the previously reported synthetic routes to BRD0539 (Non-Patent Document 5, Patent Document 1, etc.) were carried out by protecting the hydroxy group (corresponding to the hydroxy group of the hydroxymethyl group at position 4 of the final product, BRD0539) of the starting material (compound 3 in the above chemical formula (chemical formula 4)). The routes involved many complicated steps, and it was believed that improvements were required.
Therefore, an object of the present invention is to provide a synthetic route for BRD0539 that is more efficient than conventional synthetic routes.

まず、第１段階で、出発物質（化合物3）のアミノ基の保護基（ここではFmoc基）を脱保護した反応液に、アミン類（ピリジン）とトシルクロリド（TsCl）を加えてトシル化したところ（One-pot反応）、目的の化合物7を収率よく得ることに成功した（73%）。次に、第２段階の鈴木・宮浦カップリング反応においても、ヒドロキシ基を保護せずとも高収率（94%）で反応が進行し、目的のBRD0539を得ることができた。この合成経路は、化合物3から2段階で、BRD0539を収率69%で得ることができ、大変効率の良い合成法といえる。
即ち、本発明者は、出発物質である下記化合物Ａのヒドロキシ基を反応の全工程に渡って保護せずに、この反応を行うことができることを見出し、少ない工程で容易にBRD0539を合成することのできる経路を開発することに成功した。 As a result of extensive efforts, the present inventors have achieved a more efficient synthesis as shown in the following formula (Chemical Formula 6), which differs from conventional synthesis methods in that it does not require protection of the hydroxy group of the starting material (here, compound 3).

First, in the first step, the amino protecting group (Fmoc group in this case) of the starting material (compound 3) was deprotected, and the reaction solution was treated with amines (pyridine) and tosyl chloride (TsCl) to perform tosylation (one-pot reaction), successfully obtaining the target compound 7 in good yield (73%). Next, in the second step, the Suzuki-Miyaura coupling reaction, the reaction proceeded in high yield (94%) even without protecting the hydroxyl group, and the target BRD0539 was obtained. This synthetic route can obtain BRD0539 in 69% yield in two steps from compound 3, making it a very efficient synthetic method.
That is, the present inventors have discovered that this reaction can be carried out without protecting the hydroxy group of the starting material, Compound A, throughout all steps of the reaction, and have succeeded in developing a route by which BRD0539 can be easily synthesized in fewer steps.

で表される化合物BRD0539を合成する方法であって、
（ｉ）第１段階：下式（化２）に示すように、溶液中に下記化合物Ａを用意し、該化合物Ａのアミノ基の保護基Ｘを脱保護した後、アミン類の存在下で下記化合物Ａにｐ－トルエンスルホニルクロライド（ＴｓＣｌ）又はｐ－トルエンスルホン酸無水物を反応させて、化合物Ｂを合成する段階、及び

（式中、Ｘはアミノ基の保護基、Ｙはハロゲン原子又はトリフラート（ＴｆＯ）を表す。以下同じ。）
（ii）第２段階：下式（化３）に示すように、溶液中で、該化合物Ｂに２－フルオロフェニルボロン酸若しくはそのエステル又は２－フルオロフェニルトリフルオロボレート塩を反応させて、化合物BRD0539を合成する段階、

から成る方法である。 That is, the present invention provides a compound represented by the following formula (Chemical Formula 1):

A method for synthesizing a compound represented by the formula:
(i) First step: as shown in the following formula (Chemical Formula 2), a step of preparing the following compound A in a solution, deprotecting a protecting group X of an amino group of the compound A, and then reacting the following compound A with p-toluenesulfonyl chloride (TsCl) or p-toluenesulfonic anhydride in the presence of amines to synthesize a compound B;

(In the formula, X represents a protecting group for an amino group, and Y represents a halogen atom or triflate (TfO). The same applies below.)
(ii) second step: reacting compound B with 2-fluorophenylboronic acid or its ester, or 2-fluorophenyltrifluoroborate salt in a solution to synthesize compound BRD0539, as shown in the following formula (chemical formula 3);

The method consists of the following.

The present invention is a method for synthesizing the small molecule compound BRD0539, which consists of two steps as described below.

式中、Ｘはアミノ基の保護基を表す。 (i) First Stage:
(a) First, a solution of the following compound A is prepared. This compound A is represented by the following formula (Chemical Formula 7).

In the formula, X represents an amino-protecting group.

Examples of the protecting group for the amino group and the method for deprotecting the same are given below, however, the protecting group for the amino group of compound A of the present invention is not necessarily limited to these.
(a) 9-Fluorenylmethyloxycarbonyl (Fmoc) group - can be deprotected by treatment with amines such as tetrabutylammonium fluoride (TBAF) or piperidine or a base.
(b) tert-butoxycarbonyl (Boc) group - can be deprotected under strong acidic conditions such as trifluoroacetic acid or hydrochloric acid-ethyl acetate solution.
(c) Benzyloxycarbonyl (Cbz) group - can be removed by palladium-catalyzed hydrogenation or Birch reduction.
(d) 2-Nitrobenzenesulfonyl (Ns) group - can be deprotected by the action of a thiol under basic conditions.
(e) 2,2,2-trichloroethoxycarbonyl (Troc) group - can be deprotected by treatment with zinc powder and acetic acid, etc.
(f) Allyloxycarbonyl (Alloc) group: This can be deprotected by adding an amine in the presence of a palladium catalyst.
(g) Trifluoroacetyl (CF ₃ CO-) group -- can be removed by treatment with an aqueous solution of sodium hydroxide, etc.
(h) p-Toluenesulfonyl (Ts) group - can be removed by Birch reduction, etc.
(i) 2-(trimethylsilyl)ethoxycarbonyl (Teoc) group -- can be deprotected by treatment with a fluorine-containing compound (fluoride ion) such as tetrabutylammonium fluoride (TBAF).

Of these, 9-fluorenylmethyloxycarbonyl (Fmoc) group, tert-butoxycarbonyl (Boc) group, benzyloxycarbonyl (Cbz) group, and 2-nitrobenzenesulfonyl (nosyl) (Ns) group are preferred for use in the present invention.
In the above formula, Y represents a halogen atom or triflate (TfO).
As the solvent for the solution, an ether-based organic solvent such as THF or dioxane is preferable.
The concentration of Compound A in the reaction solution is preferably about 10 mM to 500 mM.

(b) Next, the protecting group X of the amino group of this compound A is deprotected. The appropriate deprotection method varies depending on the protecting group, and the method is described above. For example, when the Fmoc group used in the examples is deprotected using TBAF, the concentration of TBAF is about 0.01 mM to 1 M.
This deprotection is usually completed within 1 to several hours at a temperature of 0 to 60°C, but it is preferable to confirm that this deprotection has progressed before proceeding to the next reaction. Specifically, for example, the disappearance of the raw materials is confirmed by thin layer chromatography (TLC) using UV lamp irradiation or a color developer. It is preferable that this deprotection is completed to the extent that the raw materials are completely disappeared, but it is also acceptable if the deprotection is completed to the extent that most of the raw materials are disappeared.

(c) Next, p-toluenesulfonyl chloride (TsCl) or p-toluenesulfonic anhydride is added to the reaction solution of Compound A having the deprotected amino group in the presence of amines to perform tosylation (sulfonamidation).
Examples of the amines that can be used include pyridine, triethylamine, diisopropylethylamine, imidazole, 1,8-diazabicyclo[5.4.0]undec-7-ene (DBU), and 1,4-diazabicyclo[2.2.2]octane (DABCO).
This reaction is usually carried out at about 0 to 60°C, preferably at room temperature, and the reaction time is about several hours to several days. The pressure is generally normal pressure. Other preferable reaction conditions include anhydrous conditions (under an argon or nitrogen atmosphere, with a calcium chloride tube attached, etc.).
The concentrations of the amines, TsCl, and p-toluenesulfonic anhydride used in the reaction solution are each about 10 mM to 10 M, preferably about 10 mM to 500 mM.

As a result, compound B represented by the following formula is produced.

(ii) Second Stage:
In this step, compound B is reacted with 2-fluorophenylboronic acid or its ester, or 2-fluorophenyl trifluoroborate salt in a solution. This reaction is the so-called Suzuki-Miyaura coupling, and usually uses a palladium catalyst and a base. The reaction conditions can be the usual conditions for Suzuki-Miyaura coupling. Palladium catalysts include, but are not limited to, tetrakis(triphenylphosphine)palladium, tris(dibenzylideneacetone)dipalladium, palladium on carbon, palladium acetate, [1,1'-bis(diphenylphosphino)ferrocene]palladium dichloride, dichlorobis(tricyclohexylphosphine)palladium, and chloro(2-dicyclohexylphosphino-2',4',6'-triisopropyl-1,1'-biphenyl)[2-(2'-amino-1,1'-biphenyl)]palladium(II). Also, instead of the palladium catalyst, a nickel catalyst such as [1,1'-bis(diphenylphosphino)ferrocene]dichloronickel, nickel chloride, or bis(1,5-cyclooctadiene)nickel can be used. Examples of the base that can be used include, but are not limited to, sodium methoxide, sodium bicarbonate, potassium carbonate, sodium carbonate, sodium hydroxide, cesium fluoride, thallium hydroxide, and tripotassium phosphate hydrate. The concentrations of the base and catalyst in the reaction solution are each about 10 mM to 500 mM.
As the solvent, an ether-based organic solvent such as THF, as well as water can be used.

The concentration of compound B in the reaction solution is preferably about 10 mM to 500 mM.
As the reactant, 2-fluorophenylboronic acid or its ester, or 2-fluorophenyltrifluoroborate salt is used. Examples of the ester include pinacol ester (pin), trimethylene glycol ester, diisopropyl ester, catechol ester, neopentyl glycol ester, etc., and examples of the 2-fluorophenyltrifluoroborate salt include, but are not limited to, its potassium salt and its tetrabutylammonium salt. The concentration of these reactants in the reaction solution is about 10 mM to 500 mM.
This reaction is usually carried out at about 0 to 60°C, preferably at room temperature, and the reaction time is about several hours to several days. The pressure is generally normal pressure. Other preferable reaction conditions include anhydrous conditions (under an argon or nitrogen atmosphere, with a calcium chloride tube attached, etc.).
The result is the small molecule compound BRD0539.

These first and second steps may be carried out in any of the following ways.
(1) After extracting and purifying the compound B, which is the reaction product of the first step, the second step is carried out separately using this compound B.
(2) The second step is carried out without extracting the compound B, which is the reaction product of the first step.

The present invention will be illustrated by the following examples, but is not intended to limit the present invention.
In the following examples, melting points are uncorrected values measured on a Yanaco MP-500P. NMR spectra were measured on a JEOL ECX 400 spectrometer ( ^1H -NMR: 399.8 MHz, ^13C -NMR: 100.5 MHz). Reported chemical shifts for ^1H and ^13C NMR spectra were reported relative to the internal standard tetramethylsilane ( _δH = 0) or the solvent signal ( _{CDCl3 δC} = 77.0). IR spectra were measured on a Shimadzu Prestige-21 FT IR spectrometer. Optical rotations were measured on a Rudolph Research Analytical AUTOPOL 4T, and specific rotations were reported in units of ^10-1 deg ^cm2 g ^-1 . HRMS (high resolution mass spectra) were measured on a mass spectrometer (Agilent Technologies 6530C qTOF) in ESI positive mode. Flash column chromatography and silica gel column chromatography were all performed using Kanto Chemical silica gel 60N (spherical, neutral, 40-50 mm). Merck Kieselgel 60 F254 was used for thin-layer chromatography.

アルゴン雰囲気下、無水硫酸マグネシウム (0.312 g) に2-オキソ酢酸エチル（東京化成工業株式会社製） (47 %トルエン溶液, 1.37 g, 5.53 mmol) を加え0℃に冷却した。4-ブロモアニリン（東京化成工業株式会社製） (0.793 g, 4.61 mmol) の無水トルエン（富士フイルム和光純薬株式会社製）(9.5 mL) 溶液をシリンジポンプで30分かけて加え、0 ℃で1 時間攪拌した。反応液を吸引ろ過、減圧濃縮することで中間体イミン（化合物１）を得た。これにトルエンを加え留去した（共沸によって水を除去）後、続く反応を行った。アルゴン雰囲気下、化合物1を無水トルエン (27.3 mL) に溶解し、キラル尿素触媒 (R)-N-[(1R,2R)-2-(3-(3,5-ビス(トリフルオロメチル)フェニル)ウレイド)シクロヘキシル]-tert-ブチル-スルフィンアミド（Sigma-Aldrich社製） (0.0871 mg, 0.184 mmol) と粉末状の5Åモレキュラーシーブス (0.0792 g)、(9H-フルオレン-9-イル)メチル 2,3-ジヒドロ-1H-ピロール-1-カルボキシレート（自作）（1.65 g, 5.94 mmol) の無水トルエン (9.5 mL) 溶液を加え、室温で5分攪拌した。反応液を-65℃に冷却した後、無水のｐ－トルエンスルホン酸 (0.0158 g, 0.0917 mmol) の無水THF（富士フイルム和光純薬株式会社製） (200 μL) 溶液を少量ずつ加え-65℃で1時間攪拌した。反応液を減圧濃縮することにより反応を終了させ粗生成物を得た。粗生成物をフラッシュカラムクロマトグラフィー（ヘキサン : EtOAc = 1 : 1, シリカゲル 407 g）により精製し、化合物2 (1.78 g, 収率70 %) を得た。 Production Example 1
In this example, (3aR,4R,9bR)-1-((9H-fluoren-9-yl)methyl) 4-ethyl 8-bromo-3,3a,4,5-tetrahydro-1H-pyrrolo[3,2-c]quinoline-1,4(2H,9bH)-dicarboxylate (compound 2) was synthesized. The reaction scheme of this example is shown below.

Under an argon atmosphere, ethyl 2-oxoacetate (Tokyo Chemical Industry Co., Ltd.) (47% toluene solution, 1.37 g, 5.53 mmol) was added to anhydrous magnesium sulfate (0.312 g) and cooled to 0°C. A solution of 4-bromoaniline (Tokyo Chemical Industry Co., Ltd.) (0.793 g, 4.61 mmol) in anhydrous toluene (Fujifilm Wako Pure Chemical Industries, Ltd.) (9.5 mL) was added over 30 minutes using a syringe pump, and the mixture was stirred at 0°C for 1 hour. The reaction solution was suction filtered and concentrated under reduced pressure to obtain an intermediate imine (compound 1). Toluene was added and distilled off (water was removed by azeotropy), and the subsequent reaction was carried out. Under an argon atmosphere, compound 1 was dissolved in anhydrous toluene (27.3 mL), and the chiral urea catalyst (R)-N-[(1R,2R)-2-(3-(3,5-bis(trifluoromethyl)phenyl)ureido)cyclohexyl]-tert-butyl-sulfinamide (Sigma-Aldrich) (0.0871 mg, 0.184 mmol), powdered 5 Å molecular sieves (0.0792 g), and a solution of (9H-fluoren-9-yl)methyl 2,3-dihydro-1H-pyrrole-1-carboxylate (self-made) (1.65 g, 5.94 mmol) in anhydrous toluene (9.5 mL) were added, followed by stirring at room temperature for 5 min. After cooling the reaction solution to -65°C, a solution of anhydrous p-toluenesulfonic acid (0.0158 g, 0.0917 mmol) in anhydrous THF (FUJIFILM Wako Pure Chemical Industries, Ltd.) (200 μL) was added in small portions and stirred at -65°C for 1 hour. The reaction was terminated by concentrating the reaction solution under reduced pressure to obtain a crude product. The crude product was purified by flash column chromatography (hexane: EtOAc = 1:1, silica gel 407 g) to obtain compound 2 (1.78 g, 70% yield).

アルゴン雰囲気下、製造例１で得た化合物2 (2.12 g, 3.88 mmol) を無水THF（富士フイルム和光純薬株式会社製）(39 mL) に溶解し0℃に冷却した後、LiBH₄ in THF（関東化学株式会社製）(2.0 M, 2.9 mL, 5.8 mmol) を加え室温までゆっくりと温度を上げ、室温で3時間攪拌した。sat. NH₄Cl aq. (ca. 20 mL) を加え反応を終了させEtOAc (ca. 15 mL) で2回抽出した。抽出した有機層を飽和食塩水 (ca. 20 mL) で洗い、MgSO₄で乾燥させた後、吸引ろ過、減圧濃縮することで粗生成物を得た。粗生成物をフラッシュカラムクロマトグラフィー(ヘキサン : EtOAc = 1 : 1, シリカゲル 407 g) により精製し、化合物3 (1.19 g, 82 % e.e, 収率74 %) を得た。これらはベンゼン : CH₂Cl₂ = 9 : 1によって二回再結晶を行うことで99 % e.eの化合物3 (0.996 g) を得た。(Chiral OJ-H column, MeOH : EtOH = 1 : 1, 1.0 mL/min, 210 nm; 保持時間 : timemajor = 6.64 and timeminor = 5.91 min.)
化合物3: mp 107.8~110.1℃; [α]D20.0 +135.6°(c 0.980, CHCl₃); ¹H-NMR (399.8 MHz, CDCl₃): δ = 7.77 (d, 1 H, J = 7.3 Hz), 7.71 (d, 1 H, J = 1.8 Hz), 7.67-7.59 (m, 2 H), 7.47-7.37 (m, 3 H), 7.36-7.28 (m, 2 H), 7.10 (dd, 1 H, J = 2.3, 8.7 Hz), 6.40 (d, 1 H, J = 8.7 Hz), 5.25 (d, 0.7 H, J = 7.3 Hz), 5.09 (d, 0.3 H, J = 6.7 Hz), 4.57-4.46 (m, 1 H) 4.46-4.37 (m, 1 H), 4.37-4.18 (m, 2 H), 3.84-3.72 (m, 1 H), 3.71-3.50 (m, 2 H), 3.50-3.34 (m, 1 H) , 2.41-2.29 (m, 1 H), 2.01 (quin, 1 H, J = 11.1 Hz),1.94-1.75 (m, 2 H), 1.64 (s, 1 H) ppm; ¹³C-NMR (100.5 MHz, CDCl₃): δ = 156.6, 143.9,143.9, 142.1, 141.3, 132.6, 131.1, 130.9, 127.7, 127.3, 127.0, 125.0, 125.0, 124.8, 124.2, 119.9, 116.2, 110.1, 67.8, 67.5, 65.0, 56.4, 56.2, 52.7, 52.4, 47.3, 47.2, 45.1, 38.9, 38.4, 34.9, 22.6, 21.6; Anal. Calcd for C₂₇H₂₅BrN₂O₃: C, 64.16; H, 4.49; N, 5.54. HRMS (ESI+) Calcd for C₂₇H₂₅BrN₂O₃ [M+H]+: 505.1127, found: 505.1126. Production Example 2
In this preparation example, (3aR,4R,9bR)-(9H-fluoren-9-yl)methyl 8-bromo-4-(hydroxymethyl)-2,3,3a,4,5,9b-hexahydro-1H-pyrrolo[3,2-c]quinoline-1-carboxylate (compound 3) was synthesized from compound 2. The reaction scheme of this preparation example is shown below.

Under an argon atmosphere, compound 2 (2.12 g, 3.88 mmol) obtained in Preparation Example 1 was dissolved in anhydrous THF (FUJIFILM Wako Pure Chemical Co., Ltd.) (39 mL) and cooled to 0°C. _LiBH4 in THF (Kanto Chemical Co., Ltd.) (2.0 M, 2.9 mL, 5.8 mmol) was added, the temperature was slowly raised to room temperature, and the mixture was stirred at room temperature for 3 hours. The reaction was terminated by adding sat. _NH4Cl aq. (ca. 20 mL), and the mixture was extracted twice with EtOAc (ca. 15 mL). The extracted organic layer was washed with saturated saline (ca. 20 mL), dried over _MgSO4 , filtered by suction, and concentrated under reduced pressure to obtain a crude product. The crude product was purified by flash column chromatography (hexane: EtOAc = 1:1, silica gel 407 g) to obtain compound 3 (1.19 g, 82% ee, 74% yield). These were recrystallized twice from benzene:CH ₂ Cl ₂ = 9:1 to give compound 3 (0.996 g) with 99% ee. (Chiral OJ-H column, MeOH:EtOH = 1:1, 1.0 mL/min, 210 nm; retention time: timemajor = 6.64 and timeminor = 5.91 min.)
Compound 3: mp 107.8~110.1℃; [α]D20.0 +135.6°(c 0.980, CHCl ₃ ); ¹ H-NMR (399.8 MHz, CDCl ₃ ): δ = 7.77 (d, 1 H, J = 7.3 Hz), 7.71 (d, 1 H, J = 1.8 Hz), 7.67-7.59 (m, 2 H), 7.47-7.37 (m, 3 H), 7.36-7.28 (m, 2 H), 7.10 (dd, 1 H, J = 2.3, 8.7 Hz), 6.40 (d, 1 H, J = 8.7 Hz), 5.25 (d, 0.7 H, J = 7.3 Hz), 5.09 (d, 0.3 H, J = 6.7 Hz), 4.57-4.46 (m, 1 H) 4.46-4.37 (m, 1 H), 4.37-4.18 (m, 2 H), 3.84-3.72 (m, 1 H), 3.71-3.50 (m, 2 H), 3.50-3.34 13C ^- NMR (100.5 MHz _, CDCl 3 ): δ = 156.6, 143.9,143.9, 142.1, 141.3, 132.6, 131.1, 130.9, 127.7, 127.3, 127.0, 125.0, 125.0, 124.8, 124.2, 119.9, 116.2, 110.1, 67.8, 67.5, 65.0, 56.4, 56.2, 52.7, 52.4, 47.3, 47.2, 45.1, 38.9, 38.4, 34.9, 22.6, 21.6; Calcd for C ₂₇ H ₂₅ BrN ₂ O ₃ : C, 64.16; H, 4.49; N, 5.54. HRMS (ESI+) Calcd for C ₂₇ H ₂₅ BrN ₂ O ₃ [M+H]+: 505.1127, found: 505.1126.

Example 1
In this example, BRD0539 was synthesized in two steps from compound 3. The reaction scheme of this example is shown below.

Step 1: ((3aR,4R,9bR)-8-bromo-1-tosyl-2,3,3a,4,5,9b-hexahydro-1H-pyrrolo[3,2-c]quinolin-4-yl)methanol (compound 7) was synthesized from compound 3.
Under an argon atmosphere, compound 3 (1.08 g, 2.14 mmol) obtained in Production Example 2 was dissolved in anhydrous THF (manufactured by Fujifilm Wako Pure Chemical Industries, Ltd.) (79.3 mL), TBAF in THF (manufactured by Sigma-Aldrich Co., Ltd.) (1.0 M, 4.3 mL, 4.3 mmol) was added, and the mixture was stirred at room temperature for 2 hours. After confirming the disappearance of the raw materials by TLC, dehydrated pyridine (manufactured by Kanto Chemical Co., Ltd.) (13.7 mL) was added, and the mixture was cooled to 0°C, and TsCl (p-toluenesulfonyl chloride (also called tosyl chloride)/manufactured by Tokyo Chemical Industry Co., Ltd.) (0.612 g, 3.21 mmol) was added, and the mixture was stirred at room temperature for 3 hours. After that, TsCl (0.312 g, 1.63 mmol) was added and stirred for 15 hours, and then TsCl (0.502 g, 2.63 mmol) was added and stirred for 2 hours, after which water (ca. 100 mL) was added to stop the reaction, and after stirring for a while, the mixture was extracted three times with CH ₂ Cl ₂ (ca. 30 mL). The extracted organic layer was washed with saturated brine (ca. 100 mL), dried over MgSO ₄ , suction filtered, and concentrated under reduced pressure to obtain the crude product. This crude product was purified by flash column chromatography (hexane: EtOAc = 1:3, silica gel 407 g) to obtain compound 7 (0.683 g, 73% yield). A sample for analysis was obtained by recrystallization (EtOAc).
Compound 7: mp 207.2~208.5℃; [α]D20.0 +103°(c 1.07, CHCl ₃ ); ¹ H-NMR (CDCl ₃ /TMS): δ = 7.82 (s, 1 H), 7.78 (d, 2 H, J = 8.2 Hz), 7.36 (d, 2 H, J = 7.8 Hz), 7.12 (d, 1 H, J = 8.7 Hz), 6.39 (dd, 1 H, J = 1.1, 8.7 Hz), 5.00 (d, 1 H, J = 7.3 Hz), 4.19 (brs, 1 H), 3.63 (d, 1 H, J = 7.3 Hz), 3.55-3.44 (m, 2H), ^13C -NMR (CDCl ₃ /TMS): δ = 143.8, 143.2, 135.0, 133.4, 131.1, 129.9, 127.6 _, 122.4, 116.2, 110.5, 65.0, 58.9, 52.5 _, 47.2, _38.3 , 23.1, 21.6 _; Anal. 52.18; H, 4.89; N, 6.41. Found: C, 52.04; H, 4.82; N, 6.31 %.

Step 2: ((3aR,4R,9bR)-8-(2-fluorophenyl)-1-tosyl-2,3,3a,4,5,9b-hexahydro-1H-pyrrolo[3,2-c]quinolin-4-yl)methanol (BRD0539) was synthesized from compound 7.
Under an argon atmosphere, compound 7 (0.683 g, 1.56 mmol) obtained in the first step was dissolved in THF (52.0 mL) and distilled water (17.3 mL) was added. Then, 2-fluorophenylboronic acid (Tokyo Chemical Industry Co., Ltd.) (0.363 g, 2.34 mmol), K ₃ PO ₄ · nH ₂ O (tripotassium phosphate hydrate 76.7%, 1.12 g, 4.05 mmol), and palladium catalyst (chloro(2-dicyclohexylphosphino-2',4',6'-triisopropyl-1,1'-biphenyl) [2-(2'-amino-1,1'-biphenyl)] palladium (II) / Sigma-Aldrich X-Phos Pd-G2) (0.0614 g, 0.0781 mmol) were added and the mixture was stirred at room temperature for 3 hours. The reaction mixture was extracted three times with EtOAc (ca. 20 mL), and the organic layer was dried _{over Na2SO4} , filtered, and concentrated under reduced pressure to obtain the crude product. The crude product was purified by flash column chromatography (hexane: EtOAc = 1:2, silica gel 212 g) to give BRD0539 (0.661 g, 94% yield). An analytical sample was obtained by recrystallization (MeOH).
BRD0539: mp 101.2.0~104.4℃; [α]D20.0 105°(c 0.960, CHCl ₃ ); ¹ H-NMR (CDCl ₃ /TMS): δ = 7.90 (s, 1 H), 7.79 (d, 2 H, J = 7.8 Hz), 7.43 (t, 1 H, J = 7.8 Hz), 7.36-7.27 (m, 3 H), 7.25-7.04 (m, 3 H), 6.56 (d, 1 H, J = 8.2 Hz), 5.09 (d, 1 H, J = 7.3 Hz), 4.28 (brs, 1 H ), 3.62 (dd, 3 H, J =2.3, 9.2 Hz), 3.58-3.42 (m, 2 H), 3.42-3.25 (m, 2 H), 2.43 (s, 3 H), 1.98 (quin, 1 H, J = 10.6 Hz), 1.87-1.73 (m, 1 H), 1.71-1.57, (m, 2 H) ppm; ¹³ C-NMR (CDCl ₃ / TMS): δ = 159.7 (d, J = 246.4 Hz), 143.6, 142.9, 135.2, 131.3 (d, J = 1.9 Hz),130.4 (d, J = 3.8 Hz), 129.9, 129.0 (d, J = 4.8 Hz), 127.9 (d, J = 7.6 Hz), Anal. Calcd for C ₂₇ H ₂₅ BrN ₂ O ₃ : C, 66.35; H, 5.57; N, 6.19. Found: C, 66.34; H, 5.73; N, 6.15 %.

Claims (3)

The following formula (Chemical Formula 1)

A method for synthesizing a compound represented by the formula:
(i) First step: as shown in the following formula (Chemical Formula 2), a step of preparing the following compound A in a solution, deprotecting a protecting group X of an amino group of the compound A, and then reacting the following compound A with p-toluenesulfonyl chloride (TsCl) or p-toluenesulfonic anhydride in the presence of amines to synthesize a compound B;

(In the formula, X represents a protecting group for an amino group, and Y represents a halogen atom or triflate (TfO). The same applies hereinafter.)
(ii) second step: reacting compound B with 2-fluorophenylboronic acid or its ester, or 2-fluorophenyltrifluoroborate salt in a solution to synthesize compound BRD0539, as shown in the following formula (chemical formula 3);

The method comprising: 2. The method of claim 1, wherein the first and second steps are carried out in one of the following ways:
(1) After extracting and purifying the compound B, which is the reaction product of the first step, the second step is carried out separately using this compound B.
(2) The second step is carried out without extracting compound B, which is the reaction product of the first step. The method according to claim 1 or 2, wherein the protecting group X of the amino group is a 9-fluorenylmethyloxycarbonyl (Fmoc) group, a tert-butoxycarbonyl (Boc) group, a benzyloxycarbonyl (Cbz) group, or a 2-nitrobenzenesulfonyl (nosyl) (Ns) group.