CLIAMS:1. A process for the synthesis of a compound of the following formula (I):

(I)
for which R1 is chosen from among the group constituted of a hydrogen atom and a C1-C6 alkyl group, linear or branched, and R2 is chosen from among the group constituted of a hydrogen atom, a C1-C6 alkyl group, linear or branched, and a C1-C6 alkoxy group, linear or branched, characterized in that it comprises the following successive steps:
(a) acetylation of a compound of formula (II)

(II)
for which R1 and R2 are defined as previously, to lead to a compound of following formula (III):

(III)
for which R1 and R2 are defined as previously ;
(b) subjecting the compound of formula (III) obtained in the previous step (a) to a Fries rearrangement to lead to a compound of following formula (IV):

(IV)
for which R1 and R2 are defined as previously ;
(c) cyclization, reduction and aromatization of the compound of formula (IV) obtained in the previous step (b) to lead to the formation of the compound of formula (I).

2. The process according to claim 1, characterized in that R1 represents a hydrogen atom.

3. The process according to claim 1 or 2, characterized in that R2 represents a methoxy group.

4. The process according to any one of claims 1 to 3, characterized in that acetylation step (a) is carried out with 2-chloroacetyl chloride in the presence of a base.

5. The process according to any one of claims 1 to 4, characterized in that Fries rearrangement step (b) is carried out at a temperature of between 90°C and 120°C.

6. The process according to any one of claims 1 to 5, characterized in that cyclization step (c) is carried out by heating in the presence of a base.

7. The process according to any one of claims 1 to 6, characterized in that the reducing step is carried out by an alkaline borohydride, particularly sodium borohydride.

8. The synthesis process according to any of the previous claims comprising the preliminary synthesis of a compound of formula (II) from a compound of the following formula (V):

(V)
for which R1 and R2 are defined as previously.

9. The synthesis process according to claim 8, characterized in that the process comprises the following successive steps for the synthesis of the compound of formula (II):
(a1) acetylation of a compound of formula (V) to lead to a compound of the following formula (VI):

(VI)
for which R1 and R2 are defined as previously ;
(a2) nitration of the compound of formula (VI) obtained in step (a1) to lead to a compound of the following formula (VII):

(VII)
for which R1 and R2 are defined as previously ;
(a3) hydrolyzing the compound of formula (VII) obtained in the previous step (a2) to lead to a compound of the following formula (VIII):

(VIII)
for which R1 and R2 are defined as previously ;
(a4) subjecting the compound of formula (VIII) obtained in the previous step (a3) to Knoevenagel condensation to lead to a compound of following formula (IX):

(IX)
for which R1 and R2 are defined as previously ;
(a5) cyclization of the compound of formula (IX) obtained in the previous step (a4) to obtain the compound of formula (II).

10. The synthesis process according to claim 9, characterized in that the step (a1) of acetylating the compound of formula (V) is carried out with acetic anhydride.

11. The synthesis process according to one of claims 9 or 10, characterized in that nitration step (a2) of the compound of formula (V) is carried out with concentrated nitric acid.

12. The synthesis process according to one of claims 9 or 10, characterized in that hydrolysis step (a3) of the compound of formula (VII) is carried out in basic medium.

13. The synthesis process according to any one of claims 9 to 11, characterized in that Knoevenagel condensation step (a4) is carried out by heating under acidic conditions.

14. The synthesis process according to any one of claims 9 to 12, characterized in that cyclization step (a5) of the compound of formula (IX) is carried out by copper coupling.
,TagSPECI:The invention relates to a process for the synthesis of a psoralen derivative substituted in position 4 and 8 from a 7-hydroxycoumarin derivative substituted in position 4 and 8.

Another object of the invention is a process for the synthesis of said 7-hydroxycoumarin derivative including the process above but preceded by a preliminary synthesis of the 7-hydroxycoumarin derivative substituted from a 4-hydroxybenzaldehyde derivative.

The invention applies to the field of extracorporeal photochemotherapy. Extracorporeal photochemotherapy is an autologous cell therapy technique that consists of obtaining a cell suspension rich in mononuclear cells (apheresis or constitution of a buffy coat in line), adding to it a photosensitizing agent such as 8-methoxypsoralen (8-MOP), and then exposing the suspension to UVA radiation. The cells thus modified are then reinjected into the patient.

Over the past few years, extracorporeal photochemotherapy has become an alternative treatment for various diseases, such as cutaneous T-cell lymphoma, autoimmune diseases, and the acute form and steroid-recalcitrant form of graft-versus-host disease.

The 8-MOP used for extracorporeal photochemotherapy is a molecule that is part of the furocoumarin or psoralen family. In particular, it is the active substance from the Ammi majus plant. 8-MOP can be prepared by extraction and isolation from such a plant using organic solvents. But, residual traces of these solvents, that may present a certain toxicity, are found in the final product.

Synthetic processes for obtaining furocoumarins also exist.

For example, document US 4,130,568 particularly proposes a process for preparing 8-methoxypsoralen from 6-allyl-7-hydroxy-8-methoxycoumarin. First, this starting product undergoes oxidative cyclization in the presence of a catalyst to form 4',5'-dihydro-furocoumarin, which is then processed with a strong acid to cleave the hydroxy group at position 5' and induce dehydrogenation to form 8-methoxypsoralen. The disadvantage of this synthesis is the difficulty obtaining the starting product.

Document US 4,147,703 also proposes a synthetic process for obtaining 8-methoxypsoralen from 6-hydroxy-7-methoxy-1-benzofuran-2(3H)one (A) by reacting it with hydrogen and a noble metal, (B) by reacting the product obtained in step (A) with a mixture of zinc cyanide and hydrochloric acid, (C) by reacting the product obtained in step (B) with dichlorodicyanoquinone in an inert solvent, (D) by reacting the product obtained in step (C) with ethyl cyanoacetate in a polar solvent, and (E) by decarboxylating the product obtained in step (D) to obtain 8-methoxypsoralen.

A process in three steps for preparing furocoumarins substituted in position 8 from 7-hydroxycoumarin is also known from document FR-2 404 641. The process consists of (a) reacting 7-hydroxycoumarin with a halogeno-acetal so as to form a coumarin acetal, (b) transforming the coumarin acetal into aldehyde by heating in a diluted solution, and (c) cyclizing the aldehyde in furocoumarin by heating in an alkaline solution. A global yield of 50% is claimed, which remains relatively low.

In the document Traven, V. F. (2000). Dihydrofurocoumarinones-new useful intermediates for substituted and condensed furocoumarins. Arkivoc, 4, 523-562, furocoumarinone derivatives are obtained from 7 chloroacetoxycoumarin derivatives by an unusual Fries rearrangement carried out at high temperature. A reduction/dehydration or acetylation step is then proposed to obtain psoralen derivatives. However, this synthesis produces many by-products.

Thus, these processes from the prior art provide a relatively low yield and often require steps that are laborious to implement, such as steps carried out at high temperatures.

In order to mitigate the disadvantages from the prior art, the invention proposes a synthesis process that is easily industrialized under good manufacturing practices, enabling a good yield but also a high purity of the final product to be obtained.

For this purpose, the invention proposes a process for the synthesis of a compound of the following formula (I):

(I)
for which R1 is chosen from among the group constituted of a hydrogen atom and a C1-C6 alkyl group, linear or branched, and R2 is chosen from among the group constituted of a hydrogen atom, a C1-C6 alkyl group, linear or branched, and a C1-C6 alkoxy group, linear or branched, comprising the following successive steps:
(a) acetylation of a compound of formula (II)

(II)
for which R1 and R2 are defined as previously, to lead to a compound of following formula (III):

(III)
for which R1 and R2 are defined as previously,
(b) subjecting the compound of formula (III) obtained in the previous step (a) to a Fries rearrangement to lead to a compound of following formula (IV):

(IV)
for which R1 and R2 are defined as previously;
(c) cyclization, reduction and aromatization of the compound of formula (IV) obtained in the previous step (b) to lead to the formation of the compound of formula (I).

Other objects and advantages will appear in the following description.

Figure 1 represents a diagram for the synthesis of a compound of formula (I) from a compound of formula (II).

Figure 2 represents a diagram for the synthesis of a compound of formula (II) from a compound of formula (V).

The invention relates to a process for the synthesis of a compound of the following formula (I):

(I)
for which R1 is chosen from among the group constituted of a hydrogen atom and a C1-C6 alkyl group, linear or branched, and R2 is chosen from among the group constituted of a hydrogen atom, a C1-C6 alkyl group, linear or branched, and a C1-C6 alkoxy group, linear or branched.

In particular, the compound of formula (I) is a psoralen derivative of formula (I) for which R1 is a hydrogen atom and R2 is a methoxy group. This derivative is 8-methoxypsoralen (8-MOP).

The process according to the invention comprises the following successive steps:
(a) acetylation of a compound of formula (II)

(II)
for which R1 and R2 are defined as previously, to lead to a compound of following formula (III):

(III)
for which R1 and R2 are defined as previously,
(b) subjecting the compound of formula (III) obtained in the previous step (a) to a Fries rearrangement to lead to a compound of following formula (IV):

(IV)
for which R1 and R2 are defined as previously;
(c) cyclization, reduction and aromatization of the compound of formula (IV) obtained in the previous step (b) to lead to the formation of the compound of formula (I).

In particular, the process applies to compounds of formula (I) for which R1 represents a hydrogen atom and/or R2 represents a methoxy group.

More specifically, this synthesis process is applied for the preparation of 8 MOP (compound of formula (I) for which R1 is a hydrogen atom and R2 is a methoxy group) from hydrangetin (compound of formula (II) for which R1 is a hydrogen atom and R2 is a methoxy group) that is commercially available.

The combination of hydrangetin as the starting product and the use of an acylation reaction and Fries rearrangement enables the 8-MOP compound to be obtained in only three steps, under industrializable conditions and by respecting good manufacturing practices (GMP).

Step (a) of the process according to the invention is an acetylation step performed by any known means. In particular, acetylation step (a) is carried out with 2-chloroacetyl chloride in the presence of a base.

This acetylation step is advantageously conducted at ambient temperature in order to prevent the formation of by-products.

It is understood, unless indicated otherwise, that the temperatures indicated in the present description are the temperatures of the reaction mixtures.

Fries rearrangement step (b) is carried out at a temperature of between 90°C and 120°C, particularly between 95°C and 110°C, for example 100°C.

According to the invention, this reaction temperature enables the compound of formula (IV) with a yield of over 80% to be selectively obtained from a compound of formula (III) substituted in position 8, for example by a methoxy group.

Heating to a higher temperature leads to the formation of degradation products and by-products.

The compound of formula (IV) is then cyclized, reduced and aromatized to lead to the final compound of formula (I).

Cyclization step (c) is carried out by heating in basic medium. In particular, cyclization is carried out by heating in the presence of an alkali carbonate, such as sodium carbonate.

In particular, the reduction step is carried out by an alkaline borohydride, particularly sodium borohydride.

Advantageously, the reaction medium is finally neutralized in acid medium, for example by sulfuric acid and enables the product to be aromatized.

Carrying out Fries rearrangement step (b) at the determined temperature controls the selective formation of the compound of formula (IV), and therefore controls the purity of the final product of formula (I), since the last cyclization/reduction/aromatization step (c) is also fully controlled.

So that the synthesis of the compound of formula (I) can respond to GMP requirements, finding a reliable source of the compound of starting formula (II) is necessary.

In the absence of a reliable source of starting compound of formula (II) and particularly a reliable source of hydrangetin, the applicant developed a process for the synthesis of the compound of formula (II) from a starting product that can be obtained from a reliable source.

Therefore, and according to a particular aspect of the invention, the compound of formula (II) used in previous step (a) is prepared by a synthesis process in only four steps.

According to this aspect, the process for the synthesis of the compound of formula (I) comprises the preliminary synthesis of a compound of formula (II) from a compound of the following formula (V):

(V)
for which R1 and R2 are defined as previously.

In particular, the compound of formula (V) is a 4-hydroxybenzaldehyde of formula (V) for which R1 is a hydrogen atom and R2 is a methoxy group, i.e., vanillin. Vanillin is a product that is easy to find commercially from reliable manufacturers.

More specifically, the process of the invention comprises the following successive steps for the synthesis of the compound of formula (II):
(a1) acetylation of a compound of formula (V) to lead to a compound of the following formula (VI):

(VI)
for which R1 and R2 are defined as previously;
(a2) nitration of the compound of formula (VI) obtained in step (a1) to lead to a compound of the following formula (VII):

(VII)
for which R1 and R2 are defined as previously;
(a3) hydrolyzing the compound of formula (VII) obtained in the previous step (a2) to lead to a compound of the following formula (VIII):

(VIII)
for which R1 and R2 are defined as previously;
(a4) subjecting the compound of formula (VIII) obtained in the previous step (a3) to Knoevenagel condensation to lead to a compound of following formula (IX):

(IX)
for which R1 and R2 are defined as previously;
(a5) cyclization of the compound of formula (IX) obtained in the previous step (a4) to obtain the compound of formula (II).

The acetylation step (a1) of the compound of formula (V) is carried out by known methods with acetic anhydride or acetyl chloride. In particular, acetylation is carried out with acetic anhydride.

This acetylation step (a1) is advantageously conducted at ambient temperature in order to prevent the formation of degradation products.

Then, the compound (VI) thus obtained in step (a1) undergoes nitration. In particular, nitration step (a2) of the compound of formula (V) is carried out with concentrated nitric acid, such as fuming nitric acid to lead to the compound of formula (VII).

This nitration step is carried out cold, particularly at a temperature of between 0 and 5°C.

Then, the acetyl group of the compound of formula (VII) is eliminated by hydrolysis. In particular, hydrolysis step (a3) of the compound of formula (VII) is carried out in basic medium.

For example, the compound of formula (VII) is treated with a strong base such as sodium hydroxide, and then is neutralized with a strong acid such as hydrochloric acid, to lead to the compound of formula (VIII).

The compound of formula (VIII) then undergoes Knoevenagel condensation, which is condensation between an aldehyde and an active methylene compound, such as malonic acid or a derivative thereof, to form a C-C bond.

In the process of the invention, Knoevenagel condensation step (a4) is carried out with malonic acid. The condensation is carried out by heating the reaction mixture in acid medium.

In order to prevent decarboxylation of the compound of formula (VIII), the Knoevenagel condensation is carried out at a temperature of between 50°C and 70°C, for a time of between 70 hours and 110 hours. Advantageously, condensation step (a4) is carried out at 60°C for 96 hours.

Lastly, step (a5) of cyclizing the compound of formula (IX) is performed by copper coupling. Cyclization is carried out at high temperature, particularly a temperature greater than 100°C, for example between 120°C and 150°C.

In a variation, cyclization step (a5) comprises reducing the nitro group of the compound of formula (IX) to facilitate cyclization to lead to the compound of formula (II).

Example 1: Synthesis of 8-MOP from hydrangetin

1st step: Acetylation (obtaining compound A)

Hydrangetin (1.8 g – 0.01 mole) is acetylated with 2 chloroacetyl chloride (1.35 g – 0.012 mole) by heating at 80°C in dimethylformamide in the presence of sodium carbonate. After vacuum filtration and evaporation, 7-(2’-chloroacetyloxy)-8-methoxycoumarin (compound A) is isolated by crystallization in ethyl acetate (yield 90 %).

2nd step: Fries rearrangement (obtaining compound B)

0.01 mole of 7-(2’-chloroacetyloxy)-8-methoxycoumarin (compound A) is directly heated in the presence of aluminum chloride (0.015 mole) at 100°C for 3 hours. After cooling and hydrolysis with diluted hydrochloric acid and ethyl acetate extraction, 6 (2’ chloroacetyl) 7 hydroxy 8 methoxycoumarin (compound B) is obtained after evaporation of the organic phase. The yield is 80%.

3rd step: Cyclization of the furan ring and reduction (obtaining 8-MOP)

6-(2’-chloroacetyl)-7-hydroxy-8-methoxycoumarin (1.5 g – 0.006 mole) is cyclized by heating in 20 ml of acetone in the presence of sodium carbonate (0.007 mole – 0.5 g), reducing by sodium borohydride (0.03 mole – 1.4 mg) and neutralizing in sulfuric acid medium. After vacuum filtration and evaporation and trituration in ethanol, 8-MOP is obtained (yield 80%). Lastly, the 8-MOP is recrystallized in ethanol.

The 1H NMR spectrum of 8-methoxysporalen was performed in deuterated chloroform, according to the method of the European Pharmacopoeia in effect 2.2.33. The assignment of resonance signals unambiguously confirms the structure of 8-methoxysporalen:

The 1H NMR spectrum of 8-methoxypsoralen was performed on a Brücker apparatus (300 MHz) and the signal assignments are reported in the table below:

 (ppm) / TMS multiplicity integration coupling constant assignment
4.1 singlet 3H OCH3
6.4 doublet 1H J = 9.5 Hz H-6
7.1 doublet 1H J = 2.1 Hz H-2
7.7 singlet 1H H-4
8.1 doublet 2H J = 9.5 Hz and J = 2.1 Hz H-5 and H-3

The FTIR infrared spectrum of 8-methoxypsoralen was performed according to the method of the European Pharmacopoeia in effect, 2.2.24, on a Perkin Elmer spectrophotometer, between 650 cm-1 and 4500 cm-1.

Interpretation of the infrared absorption bands is presented in the following table:

wave number  (cm-1) fonction vibration mode
3120, 3060 Aromatic CH elongation
2950 Aliphatic CH elongation
1705 C = O elongation
1620, 1590, 1550 CH = Aromatic CH elongation
1215, 1180 C-O-C elongation
870 furan ring elongation

Example 2: Synthesis of hydrangetin from vanillin

Step 1: Acetylation to obtain 4-acetoxy-3-methoxybenzaldehyde (compound 1)

28 ml (0.2 mole) of triethylamine and 220 mg of 4 dimethylaminopyridine are added in drops to a solution of vanillin (20.0 g – 0.171) and acetic anhydride (26 ml – 0.275 mole) in 100 ml of dichloromethane, using a magnetic stirrer, by maintaining a temperature of less than 25°C. This reaction mixture is stirred for 30 min. at ambient temperature.

The organic phase is successively washed with 30 ml of water, with 40 ml of an aqueous hydrochloric acid solution 20 % and lastly with 30 ml of a saturated aqueous solution of sodium chloride. After drying of the organic phase, the solution is vacuum evaporated. After cooling and adding cyclohexane, the product is isolated by vacuum filtration. The product is recrystallized in ethyl ether and 24.5 g of compound 1 is obtained (yield 75 %).

Step 2: Nitration to obtain 4-acetoxy-3-methoxy-2nitrobenzaldehyde (compound 2)

Small 12 g (0.06 mole) portions of compound 1 are slowly added to a fuming nitric acid solution (50 ml) cooled in an ice bath to 0°C. The reaction mixture is stirred for 1 hour at a temperature of less than 5°C.
The reaction mixture is then poured into the ice water and stirred again for 1 hour. The product is isolated by vacuum filtration and cyclohexane washing (yield 80 % - 11.5 g).

Step 3: Hydrolysis and obtaining 4-hydroxy-3-methoxy-2-nitrobenzaldehyde (compound 3)

10 g of compound 2 (0.042 mole) are added to a solution of 40 ml of sodium hydroxide 33 % (m/m). The reaction mixture is heated at reflux for 10 minutes.

The reaction mixture is diluted with 40 ml of water and acidified with concentrated hydrochloric acid (HCl 6N) to neutral pH. After cooling, the product is precipitated and compound 3 is isolated by vacuum filtration and by several washings with water (yield 85 % - 7.2 g of compound 3).

Step 4: Preparation of (4-hydroxy-3-methoxy-2-nitrobenzylidene)malonic acid (compound 4)

The mixture of compound 3 (9.8 g – 0.05 mole) and malonic acid (6.8 g – 0.065 mole) is heated in 15 ml of acetic acid at 60°C for 96 hours. After vacuum evaporation, the product is crystallized after cooling and addition of dichloromethane, and then isolated by vacuum filtration. 13.0 g of compound 4 is obtained (yield 90 %).

Step 5: Preparation of hydrangetin

A mixture of compound 4 (3.4 g – 0.014 mole) and copper powder (360 mg) is heated to 130°C-140°C in quinoline (50 ml) for 5 min. After cooling, 120 ml of water and then 60 ml of concentrated hydrochloric acid (HCl 10N) are added.

The aqueous phase is purified by extraction with 100 ml of dichloromethane to eliminate impurities, and then the product is extracted from the aqueous phase by a minimum of three extractions with 100 ml of ethyl acetate.

After drying of the organic phase, it is vacuum evaporated and the product is crystallized in ethyl acetate.

The product is finally recrystallized in ethyl acetate to obtain 2.1 g of hydrangetin (yield 78 %).

The 1H NMR spectrum of hydrangetin was performed in deuterated dimethyl sulfoxide, according to the method of the European Pharmacopoeia in effect 2.2.33. The assignment of resonance signals unambiguously confirms the structure of hydrangetin:

The 1H NMR spectrum of hydrangetin was performed on a Brücker apparatus (300 MHz) and the signal assignments are reported in the table below:

 (ppm) / TMS multiplicity integration assignment
3.6 singlet 3H OCH3
7.1 triplet 2H Aromatic H
7.3 doublet 1H Aromatic H
11.1 singlet 1H CH = O

The FTIR infrared spectrum of hydrangetin was performed according to the method of the European Pharmacopoeia in effect, 2.2.24, on a Perkin Elmer spectrophotometer, between 650 cm-1 and 4500 cm-1.

Interpretation of the infrared absorption bands is presented in the following table:

wave number  (cm-1) function vibration mode
3296 Phenolic OH elongation
3010 Aromatic CH elongation
1727 O-CO-CH3 elongation
1692 CH = CH elongation
1255 OCH3 elongation