SALICYL FUMARATE DERIVATIVE AND ITS APPLICATION IN THE TREATMENT OF PARKINSON'S DISEASE AND OTHER NEURODEGENERATIVE DISEASES

Abstract

A salicyl fumarate derivative, and its general structural formula (A) is:

##STR00001##

Where the structural formula (A), R.sub.1 is one of H.sup.+, Na.sup.+, K.sup.+ or NH4.sup.+. R.sub.2 is one of fumaric acid ester products. It has the general structural formula of the combination of salicylic acid and fumaric acid through esterification reaction. This category of compounds possesses good effects in treatment of neurodegenerative diseases.

Claims

1. A salicyl fumarate derivative wherein: its general structural formula (A) is: ##STR00013## within the structural formula (A), R.sub.1 is one of H.sup.+, Na.sup.+, K.sup.+ or NH.sup.4+, R.sub.2 is one of fumaric acid ester products.

2. The salicyl fumarate derivative according to claim 1, wherein: the esterification of fumaric acid is one of: mono-methyl fumarate, mono-ethyl fumarate, mono-propyl fumarate, mono-isopropyl fumarate, mono-butyl fumarate, or mono-tertbutyl fumarate.

3. The salicyl fumarate derivative according to claim 2, wherein: the salicyl fumarate derivative is methyl salicyl fumarate, its structural formula is: ##STR00014##

4. The salicyl fumarate derivative according to claim 2, wherein: the salicyl fumarate derivative is ethyl salicyl fumarate, its structural formula is: ##STR00015##

5. A production method for the salicyl fumarate derivative according to claim 1, wherein: 1) an esterification reaction of tert-butyl alcohol and salicylic acid is carried out, protecting the carboxyl group of salicylic acid, obtaining intermediate product I; 2) an esterification of intermediate product I and the fumaric acid derivative is carried out, obtaining intermediate product II; and 3) a tert-butanol in intermediate product II is removed under acidic condition, obtaining target product III; wherein the chemical reaction formula is as follows: ##STR00016##

6. A method comprising applying the salicyl fumarate derivative according to claim 1 to drug production for the treatment of Parkinson's Disease.

7. A method comprising applying the salicyl fumarate derivative according to claim 2 to drug production for the treatment of Parkinson's Disease.

8. A method comprising applying the salicyl fumarate derivative according to claim 3 to drug production for the treatment of Parkinson's Disease.

9. A method comprising applying the salicyl fumaratc derivative according to claim 4 to drug production for the treatment of Parkinson's Disease.

10. A composition including a compound having a structural formula of: ##STR00017##

11. A composition including a compound having a structural formula of: ##STR00018##

Description

DESCRIPTION OF ATTACHED FIGURES

[0026] FIG. 1 MR spectrogram of Compound A (methyl salicyl fumarate);

[0027] FIG. 2 MR spectrogram of Compound B (ethyl salicyl fumarate);

[0028] FIG. 3 Dose response protection and safety study of MSF and ESF on MPTP-induced model of dopaminergic neuronal damage in mice;

[0029] FIG. 4 Protective effects of MSF and ESF on dopaminergic neurons in MPTP-induced model of chronic dopaminergic neuronal damage in mice;

[0030] FIG. 5 Effects of MSF and ESF on the concentrations of MPTP metabolites MPP+;

[0031] FIG. 6 HPLC Chromatogram of MSF in mouse blood sample;

[0032] FIG. 7 HPLC Chromatogram of ESF in mouse blood sample;

[0033] FIG. 8 Mouse blood drug concentrations with MSF and ESF dosage of 100 mg/kg;

[0034] FIG. 9 Mouse liver drug concentrations with MSF and ESF dosage of 100 mg/kg;

[0035] FIG. 10 Mouse brain drug concentrations with MSF and ESF dosage of 100 mg/kg;

[0036] FIG. 11 Under different MSF and ESF dosages, the drug concentrations in mouse striatum;

[0037] FIG. 12 The protective effect on dopaminergic neurons in Nrf2 knockout mice with MPTP-induced dopaminergic neuronal damage;

[0038] FIG. 13 Effects of MSF and ESF on intracellular glutathione concentrations in in vitro experiments;

[0039] In above figures, FIGS. 1 and 2 are NMR spectrograms generated by instrument work station; they were originally in color. After they were adjusted to black and white there were some text-and-image overlaps. This specific information has been documented in the Implementation Example 1 in the next section;

[0040] Chinese to English translations for FIGS. 3, 4, 5, 12, and 13;

[0041] Chinese to English translations for FIGS. 6 and 7;

[0042] Chinese to English translations for FIGS. 8, 9, and 10.

Specific Implementation Patterns Implementation Example 1 Synthesis of Salicyl Fumarate Derivatives

[0043] 1. Protecting the Carboxyl Group on the Salicylic Acid

[0044] Dissolve salicylic acid (45 g, 795.9 nmol, 1 equivalent weight) in dimethylformamide (DMF) (450 ml), under 0° C. (ice water bath) condition, add N′N-carbonyldiimidazole (63.5 g). At room temperature mix for 1 h, and at the same time slowly drip DBU (58.5 ml) and tert-butanol (63 ml). Then at room temperature mix for 2 h. After the reaction is done use LC-MS to detect (product spectrogram is illustrated in attached FIG. 1). Put reactants into water (500 ml), and use ethyl acetate to extract 3 times (3×800 ml). Combine with organic layers, then rinse 3 times (3×800 ml) with water; then rinse 1 time with saline (1000 ml); then dry with anhydrous Na.sub.2SO.sub.4; then filter; then concentrate the filtrate until no organic solvent residue is left. Purify the produced concentrate by silica gel column chromatography (petroleum ether:ethyl acetate=100:1 as mobile phase), obtaining the product (product 52 g, yield 82.5%). Product is light-yellow and oil-like (named: chemical compound A3). The chemical reaction formula is as follows:

##STR00007##

[0045] 2. Esterification of Monomethyl Fumarate with Compound A3 (Compound A4)

[0046] Add DCM (100 ml) to monomethyl fumarate (10.4 g), then add the mixed solution of HATU (45.6 g) and DIEA (31 g). The reaction mixture is stirred for 30 minutes, then compound A3 (10 g) is added to the reaction mixture at room temperature and stirred for 16 hours at room temperature, LC-MS shows the reaction is complete. The reaction mixture is quenched with water (200 mL) and extracted with DCM (2×200 mL). Wash the combined organic layer with salt water (300 mL), then dry with anhydrous Na.sub.2SO.sub.4; then filter; then concentrate the filtrate until no organic solvent residue is left. Purify the produced concentrate by silica gel column chromatography (petroleum ether:ethyl acetate=30:1 as mobile phase), obtaining the product (product 12.6 g, yield 79.9 g). Product is light-yellow and oil-like (named: chemical compound A4). The chemical reaction formula is as follows:

##STR00008##

[0047] 3. Preparation of Methyl Salicyl Fumarate (Compound A)

[0048] Under 0° C. (ice water bath) conditions, add compound A4 (5.6 g) to mixed solution of DCM (100 ml), 2N HCl and Et.sub.2O (25 ml). The mixture reacts by stirring at room temperature overnight, TCL shows the reaction is complete. Remove the reaction solvent, add water (100 ml), then extract 3 times with DCM (3×150 ml). Combine organic layers, then rinse with salt water; dry with anhydrous Na.sub.2SO.sub.4. The product is purified by silica gel column chromatography (petroleum ether:ethyl acetate=2:1), and crude product is obtained. Mix the crude product with n-hexane (crystallization) overnight, filter to obtain pure product A (product 4.5 g, yield 98.4%); analyze with MR and MS assays, pure product A is identified as methyl salicyl fumarate. The reaction formula is as follows:

##STR00009##

[0049] 4. Esterification of Mono-Ethyl Fumarate with Compound A3 (Compound B2)

[0050] Add mono-ethyl fumarate (10.4 g) to DCM (100 ml), then add HATU (45.6 g) and DIEA (31 g) to mixed solution. The reaction mixture is stirred for 30 minutes, then compound A3 (10 g) is added to the reaction mixture at room temperature and stirred for 16 hours at room temperature, LC-MS shows the reaction is complete. The reaction mixture is quenched with water (200 mL) and extracted with DCM (2×200 mL). Wash the combined organic layer with salt water (300 mL), then dry with anhydrous Na.sub.2SO.sub.4; then filter; then concentrate the filtrate until no organic solvent residue is left. Purify the produced concentrate by silica gel column chromatography (petroleum ether:ethyl acetate=30:1 as mobile phase), obtaining the product (product 12.8 g, yield 79.9%). Product is light-yellow and oil-like (named: chemical compound B2). The chemical reaction formula is as follows:

##STR00010##

[0051] 5. Preparation of Ethyl Salicyl Fumarate (Compound B)

[0052] Under 0° C. (ice water bath) conditions, add compound B2 (5.6 g) to mixed solution of DCM (100 ml), 2N HCl and Et.sub.2O (25 ml). The mixture reacts by stirring at room temperature overnight. TCL shows the reaction is complete. Remove the reaction solvent, add water (100 ml), then extract 3 times with DCM (3×150 ml). Combine organic layers, then rinse with salt water; dry with anhydrous Na.sub.2SO.sub.4. The product is purified by silica gel column chromatography (petroleum ether:ethyl acetate=2:1), and crude product is obtained. Mix the crude product with n-hexane (crystallization) overnight, filter to obtain pure product A (product 4.0 g, yield 86.5%); analyze with MR and MS assays, pure product B is identified as ethyl salicyl fumarate. The reaction formula is as follows:

##STR00011##

[0053] 6. Analyzing Compound a and Compound B, the Specific Data are Below:

[0054] MR Spectrometry Assay

[0055] Compound A: .sup.1H NMR (400 MHz, DMSO): δ13.18 (s, 1H), 7.98˜7.96 (m, 1H), 7.71˜7.67 (m, 1H), 7.46˜7.42 (m, 1H), 7.32˜7.30 (m, 1H), 6.99˜6.98 (m, 2H), 3.80 (s, 3H); (NMR chromatogram shown in FIG. 1). MR spectrometry verifies compound A as methyl salicyl fumarate.

[0056] Compound B: .sup.1H NMR (400 MHz, DMSO): δ13.16 (s, 1H), 7.98˜7.96 (m, 1H), 7.70˜7.67 (m, 1H), 7.45˜7.41 (m, 1H), 7.31˜7.29 (m, 1H), 6.97˜6.96 (m, 2H), 4.27˜4.22 (m, 2H), 1.29˜1.26 (m, 3H); (NMR chromatogram shown in FIG. 2). MR spectrometry verifies compound B as ethyl salicyl fumarate.

[0057] 7. The applicants used the same fore-mentioned synthesis routes to perform esterification of salicylic acid and fumarate esters. They obtained propyl salicyl fumarate (compound structural formula shown as C), isopropyl salicyl fumarate (compound structural formula shown as D), and butyl salicyl fumarate (compound structural formula shown as E) respectively. Considering the pharmacological activities of propyl salicyl fumarate, isopropyl salicyl fumarate, and butyl salicyl fumarate were lower than those of methyl salicyl fumarate and ethyl salicyl fumarate through preliminary experiments, the pharmacological studies of these three compounds will not be mentioned in this application.

[0058] However, from the synthesis routes of the 5 compounds, methyl salicyl fumarate, ethyl salicyl fumarate, propyl salicyl fumarate, isopropyl salicyl fumarate, and butyl salicyl fumarate, it can be known that if the carboxyl group on the salicylic acid can be effectively protected, salicyl fumarate derivatives can be synthesized. Therefore it is verified that the synthesis routes provided by the applicants are generally applicable for the effective synthesis of salicyl fumarate derivatives.

##STR00012##

Implementation Example 2 the Protective Role of MSF and ESF on Dopaminergic Neurons

[0059] Using MSF and ESF to conduct therapeutic studies with widely accepted MPTP-induced dopaminergic neuron-damaged PD mouse models. Dimethyl fumarate (DMF) was used as positive control drug, 6 dose groups were used for DME. For the experimental groups of MSF and ESF, 10 dose groups were used for each drug. 10 mice in each group. Both the experimental group and the control group used intraperitoneal administration. The control group was treated with DMF. The experimental groups were treated with different doses of ESF and MSF (as shown in FIG. 3). Statistically significant protective effects of dopamine neurons were found, and the dose-effect relationship was observed. The lowest dosage for protective effect (10 mg/kg/d), the optimal dosage (100-200 mg/kg/d), and the dosage where toxic effects began to appear (>500 mg/kg/d) were demonstrated. Compared to the results of DMF, MSF and ESF had more significant protective effects. Furthermore, the dosage where DMF showed protective effect (50 mg/kg/d) was higher than MSF and ESF, and the dosage where DMF showed toxic effect (>200 mg) was lower than MSF and ESF. Therefore, MSF and ESF have more effective and safer therapeutic dose windows (as shown in FIG. 3).

[0060] MSF and ESF (200 mg/kg/d) also showed significant dopamine neuron protection in models of chronic MPTP-induced dopamine neuron damage in mice, and the protection of dopamine neurons was close to a blank control level (as shown in FIG. 4).

[0061] The concentration of MPTP metabolite MPP+ in brain tissue was determined by HPLC. For the MPTP injected mouse model, the ESF (500 mg/kg) administered group and the blank group had the same MPP+ content in the striatum, excluding the possibility that MSF and ESF had an effect on the uptake and metabolism of MPTP in vivo and in the brain, confirming their neuroprotective effects (as shown in FIG. 5).

[0062] A reliable, high-performance liquid chromatography MSF and ESF analysis method (as shown in FIGS. 6 and 7) was established to determine the level of MSF and ESF in peripheral blood, liver, and brain tissue, demonstrating that MSF and ESF can enter the brain through the blood-brain barrier after intraperitoneal injection in mice. Furthermore, the molecular structure of MSF and ESF remains unchanged in the brain (as shown in FIG. 8, FIG. 9, FIG. 10, and FIG. 11), and it is inferred that MSF and ESF molecules produce neuroprotective pharmacological activities directly through their original molecule structures. Unlike the positive control drug DMF which worked differently. DMF needs to entering the body and quickly metabolizing into a form of single methyl metabolite monomethyl fumarate (MMF) which is the active form to produce pharmacological activity.

[0063] MSF and ESF have conventional fumaric acid (fumarate) activation target Nrf2 similar to the positive control drug DMF, which results in activating the mechanism of intracellular antioxidative stress. By performing MPTP experiments on transgenic mice with the Nrf2 gene knocked out, DMF lost the dopaminergic neuron protective effect due to the loss of its effective target, while MSF and ESF still show significant protective effects in this model (as shown in FIG. 12); thus confirming that MSF and ESF, in addition to acting on Nrf2 target, also act on another neuroprotective target. Inferring from the inclusion of salicyl in their molecule structure, this new target may be associated with anti-neuroinflammation.

[0064] The results of in vitro cell experiments showed that positive control drug DMF had the effect of depleting glutathione (GSH, endogenous antioxidant polypeptide), which may be the cause of its obvious toxicity and side effects. MSF and ESF have no effect on the concentration of glutathione in the cell (as shown in FIG. 13).