CN111909204A - Tenofovir disopropiolate-based phosphoramidate compound and pharmaceutical composition and application thereof - Google Patents

Abstract

The invention discloses a tenofovir bisphenylpropionate phosphoramidate compound and a salt thereof, as well as a preparation method and a pharmaceutical composition containing the compound. Experiments have proved that the compound of the present invention has the activity of inhibiting HBV virus replication, and at the same time, the compound has the advantages of higher activity and larger development coefficient than the current AIDS drug tenofovir disoproxil fumarate (TAF). Experiments also prove that the compound of the present invention also has the activity of inhibiting the replication of HIV-1 virus, and can be used for the development of AIDS drugs or hepatitis B drugs.

Description

A kind of tenofovir disoproxil phosphoramidate compound and medicine combination thereof objects and uses

technical field

The invention relates to the technical field of medicine, in particular to a tenofovir disoproxil phosphoramidate compound, and a pharmaceutical composition containing the compound and its use.

Background technique

Hepatitis B, especially chronic hepatitis B, has become a global public health problem. According to statistics, more than 1 million people worldwide die of HBV infection and related diseases every year. But so far there is no ideal treatment and medicine for chronic hepatitis B. Therefore, how to effectively prevent and cure hepatitis B is a major problem that will be faced and need to be solved in the 21st century.

At present, TAF (Vilide) and ETV (Entecavir), a class of drugs for the treatment of hepatitis B in the Chinese market, have gradually shown their respective shortcomings since their listing.

In "2018 American Society of Liver Diseases Prevention, Diagnosis and Treatment of Chronic Hepatitis B", entecavir (ETV), tenofovir disoproxil fumarate (TDF), tenofovir alafenamide fumarate (TAF) are recommended As the best drug for the treatment of hepatitis B, the three have their own advantages and disadvantages:

Advantages of ETV: small side effects; Disadvantages: certain drug resistance, contraindicated in pregnant women;

Advantages of TDF: good antiviral effect, very few drug resistance; Disadvantages: certain nephrotoxicity, long-term application will have certain side effects on bones and kidneys;

Advantages of TAF: The antiviral effect of TAF is equivalent to that of TDF, and the laboratory safety parameters of kidney and bone are improved.

TAF is a new oral regimen for the treatment of hepatitis B. Although it cannot completely cure hepatitis B, its efficacy has been greatly improved, and the amount of hepatitis B virus can be well controlled. In terms of clinical effect, TAF and TDF have the same clinical effect in the treatment of hepatitis B, but the dosage of TAF is smaller, the toxicity is lower, and it will play a greater role in drug resistance and combination therapy. However, since TAF will lose a molecule of phenol after entering the human body, the TAF compound preparation has gradually shown toxic and side effects that cannot be ignored since it was launched: lactic acidosis and severe hepatomegaly combined with fatty liver.

Therefore, it is of great significance to develop new analogs of TAF that maintain minor changes in structure, maintain the excellent antiviral activity of TAF, and at the same time have better stability in plasma and oral bioavailability.

To sum up, although Gilead's TDF and TAF compound preparations still dominate the market for the current global AIDS/hepatitis B treatment drugs, our research on TNF-based AIDS/hepatitis B medicines is still of great significance. It is still of great value to further improve the bioavailability of the human body, increase the activity, reduce the toxic and side effects, and give full play to the efficacy of the treatment of hepatitis B and AIDS.

SUMMARY OF THE INVENTION

In the process of prodrug research on TAF analogs, it was unexpectedly found that after replacing the benzene ring chain of TAF with an amino NHR chain, the activity increased, and the biological activity selection coefficient SI also increased, which made the use of research samples. Compared with the dose of TAF, the TAF can be further reduced, that is, the toxicity of TAF can be further reduced, while the activity of TAF can be maintained, and the drug concentration in tissue cells, especially hepatocytes, is significantly increased compared with TAF. Therefore, the tenofovir disoproxil phosphoramidate compound of the present invention can significantly improve the therapeutic effect on hepatitis B and AIDS, and greatly reduce the nephrotoxicity and bone toxicity caused by TDF or TAF. Thus, the following invention has been proposed.

A tenofovir disoproxil phosphoramidate compound comprising:

a) Compound (Ia), the structure of which is shown below,

wherein R ₁ =R ₂ =C ₁ -C ₁₂ alkyl; or

b) A pharmaceutically acceptable salt prepared from compound (Ia) as described in a) with an acid.

Further, when the R ₁ and R ₂ groups are selected from isopropyl, ethyl, isobutyl, neopentyl, n-butyl, cyclohexyl, methyl or tert-butyl, the structural formula of the compound is respectively the following Structural formula:

Further, the pharmaceutically acceptable salts are oxalate, malonate, succinate, fumarate, aspartate, citrate, salicylate, hydrochloride, sulfate The molar ratio of , phosphate, acid and compound (Ia) ranges from (0.5 to 1):1.

Further, when the R ₁ and R ₂ groups are selected from isopropyl, ethyl, isobutyl, neopentyl, n-butyl, cyclohexyl, methyl or tert-butyl, the fumarate The structural formulas are as follows:

Further, the fumarate is prepared from fumaric acid and the compound in a molar ratio of 0.5:1.

In another aspect, the present invention also provides a pharmaceutical composition comprising a therapeutically effective amount of tenofovir disoproxil phosphoramidate compound, and at least one pharmaceutically acceptable carrier or excipient.

Further, the pharmaceutical composition further comprises a therapeutically effective amount of an additional therapeutic agent selected from the group consisting of compounds that inhibit HIV protease, HIV non-nucleoside inhibitors of reverse transcriptase, HIV nucleoside inhibitors of reverse transcriptase, HIV nucleotide inhibitors of reverse transcriptase, HIV integrase inhibitors, gp41 inhibitors, CXCR4 inhibitors, gp120 inhibitors, CCR5 inhibitors, viral capsid polymerization inhibitors, non-catalytic site HIV integrase site inhibitors at least one.

Further, the dosage form of the pharmaceutical composition is tablet or capsule.

The tenofovir disoproxil-based phosphoramidate compound of the present invention can be used for preparing medicines for preventing or treating viral diseases. A specific viral disease is HIV-1 infection or HBV infection or HIV-1 and HBV co-infection.

It has been determined that the compound of the present invention has excellent properties required to become a drug for the treatment of AIDS/hepatitis B, as follows:

In the in vitro anti-HBV activity screening, the IC ₅₀ of compound COP371371 was 6 times that of TAF (positive control), and the biological activity selection coefficient SI was 4.0 times that of TAF (positive control); the IC ₅₀ of compound COP374374 was TAF (positive control). ) 6 times, and the biological activity selection coefficient SI is 1.5 times that of TAF (positive control). It shows that the compound of the present invention has a higher activity of inhibiting virus replication than a class of anti-hepatitis B drugs TAF, and has a large selection coefficient of biological activity, and is expected to be a drug for the treatment of HBV infection. The compound of the present invention integrates various good properties such as high activity, low toxicity, high bioavailability, etc., and has the prospect of becoming a new generation of drugs for treating AIDS or treating hepatitis B.

Detailed ways

The following examples can make those skilled in the art understand the present invention more comprehensively, but do not limit the present invention in any way. The structures of all compounds were confirmed by ¹ H NMR or MS.

The raw materials used in the present invention: tenofovir, L-alanine ester hydrochloride and fumaric acid are all purchased from the market.

Example 1

9-[(R)-2-[[bis[[(S)-(1-n-butoxycarbonyl-2-phenyl)ethyl]amino]phosphinyl]methoxy]propyl]adenine Preparation of (COP374374).

Weigh commercially available tenofovir (COP00) 7.2g (25mmol), add it to a 250ml flask, then add 50g of thionyl chloride, heat up to 55°C, after 10 minutes, continue to heat up to 70°C and stir for 1 hour , distilled thionyl chloride under reduced pressure, added 100 ml of dry acetonitrile after the distillation was completed and refluxed at 85°C for 10 min, evaporated the acetonitrile to dryness under reduced pressure, cooled to room temperature, then added 50 ml of dry dichloromethane, and continued stirring for 0.5 h. 19.4 g (75 mmol, 3 eq) of L-phenylalanine n-butyl ester hydrochloride (AH374) was added, and after stirring for 10 min, about 20 ml of triethylamine was added dropwise, and the stirring was continued for 0.5 h.

After suction filtration, the filtrate was evaporated to dryness and then 100 ml of ethyl acetate was added. and washed twice with 100 g of 5% potassium bicarbonate aqueous solution and twice with 100 g of 20% aqueous sodium chloride solution, dried over anhydrous sodium sulfate, filtered with suction after drying, and the filtrate was evaporated to dryness. Ethyl acetate column chromatography obtained about 8 g of COP374374 (purity 98.2%).

¹ H NMR (400 MHz, CDCl ₃ ) δ, (ppm): 0.85-0.98 (6H, dt, 2×CH ₃ ), 1.02-1.15 (3H, d, CH ₃ ), 1.25-1.41 (4H, m, 2 ×CH ₂ ), 1.45-1.67(4H,m,2×CH ₂ ),2.73-2.98(5H,m,OCH and 2×CH ₂ ),3.01-3.54(4H,m,2×NH and OCH ₂ P ), 3.70-4.12(6H,m,2×NCH and 2×COOCH ₂ ),4.13-4.36(2H,m,NCH ₂ ),6.27(2H,s,NH ₂ ),7.00-7.38(10H,m, two hydrogens on the benzene ring), 7.88 (1H, s, H on the purine ring), 8.32 (1H, s, H on the purine ring).

ESI-MS:[M+H]+694.7

Example 2

9-[(R)-2-[[bis[[(S)-(1-n-butoxycarbonyl-2-phenyl)ethyl]amino]phosphinyl]methoxy]propyl]adenine Preparation of Fumarate (FCOP374374).

An equivalent amount of 9-[(R)-2-[[bis[[(S)-(1-n-butoxycarbonyl-2-phenyl)ethyl]amino]

Phosphosphinyl] methoxy] propyl] adenine (COP374374) and fumaric acid were dissolved in hot acetonitrile, refluxed and stirred for 2 hours, cooled and crystallized at room temperature, and the precipitated solid was filtered and washed with acetonitrile to obtain a white solid: 9-[(R)-2-[[bis[[(S)-(1-n-butoxycarbonyl-2-phenyl)ethyl]amino]phosphinyl]methoxy]propyl]adenine Fumarate (FCOP374374).

Example 3

In vitro determination of the anti-HBV virus activity of the compounds of the present invention

1. In vitro cell model: HepG 2.2.15 cells

2. Dot blot assay to determine the anti-HBV activity of compounds

2.1 HepG 2.2.15 cells ( ⁴ x 104 cells/well) were transferred to a 96-well plate and cultured overnight at 37°C, 5%.

2.2 On the second day, dilute the compounds and add different concentrations of compounds to the culture wells. The final concentration of DMSO in the culture medium was 1%. 1 μM entecavir (ETV) served as 100% Inhibition control; 1% DMSO served as 0% Inhibition control.

2.3 On the fifth day, replace with fresh culture medium containing compound.

2.4 On the eighth and ninth days, the culture medium in the culture wells was removed, and the cells were collected for dot hybridization.

3. Results: See Table 1.

Table 1: Cytotoxicity and extracellular anti-HBV activity of compounds (HepG2.2.15)

4. Experimental conclusion:

4.1 The positive compound TAF has the expected inhibitory effect on hepatitis B virus replication in HepG 2.2.15 cells, which proves that the experiment is effective and credible.

4.2 Anti-HBV activity. In the dot hybridization experiment, the activity of the two tested compounds in the table above was 6 times higher than that of TAF (positive control). And the development coefficient SI is higher than the positive control TAF inhibition rate.

4.3 This fully shows that the compound of the present invention has higher activity of inhibiting virus replication than the current anti-hepatitis B drug TAF, and is expected to become a new-generation drug for the treatment of HBV infection.

Example 4

Formulation and process

The pharmaceutical composition of the present invention can be made into tablets or capsules according to the general preparation method of oral pharmaceutical preparations, and the unit content of the tablet or capsule of the compound of the present invention at a dose of 5 mg is as follows:

Table II

name Proportion/mg Compounds of the present invention 2.50 lactose 1.65 starch 0.66 microcrystalline cellulose 0.08 Sodium starch glycolate 0.08 Magnesium stearate 0.03

.

Claims (10)

1. A tenofovir disoproxil phosphoramidate compound, comprising: a) the compound (Ia), the structure of which is shown below,

wherein R is₁＝R₂＝C₁-C₁₂An alkyl group; or

b) A pharmaceutically acceptable salt prepared from the compound (Ia) as described in a) and an acid.

2. The tenofovir disopropionate phosphoramidate compound of claim 1, wherein when R is₁、R₂When the group is selected from isopropyl, ethyl, isobutyl, neopentyl, n-butyl, cyclohexyl, methyl or tert-butyl, the structural formula of the compound is respectively as follows:

3. the tenofovir disopropionate phosphoramidate compound of claim 1, wherein the pharmaceutically acceptable salt is oxalate, malonate, succinate, fumarate, aspartate, citrate, salicylate, hydrochloride, sulfate or phosphate, and the molar ratio of acid to compound (Ia) is in the range of (0.5-1): 1.

4. The tenofovir disopropionate phosphoramidate compound of claim 3, wherein when R is₁、R₂When the group is selected from isopropyl, ethyl, isobutyl, neopentyl, n-butyl, cyclohexyl, methyl or tert-butyl, the structural formula of the fumarate is respectively as follows:

5. the tenofovir disopropionate phosphoramidate compound of claim 4, wherein the molar ratio of fumaric acid to compound (Ia) is 0.5: 1.

6. A pharmaceutical composition comprising a therapeutically effective amount of a tenofovir disopropionate phosphoramidate compound according to any of claims 1 to 5, and at least one pharmaceutically acceptable carrier or excipient.

7. The pharmaceutical composition of claim 6, wherein: the pharmaceutical composition further comprises a therapeutically effective amount of an additional therapeutic agent selected from at least one of a compound that inhibits HIV protease, an HIV non-nucleoside inhibitor of reverse transcriptase, an HIV nucleotide inhibitor of reverse transcriptase, an HIV integrase inhibitor, a gp41 inhibitor, a CXCR4 inhibitor, a gp120 inhibitor, a CCR5 inhibitor, a viral capsid polymerization inhibitor, a non-catalytic site HIV integrase site inhibitor.

8. The pharmaceutical composition of claim 6 or 7, wherein: the dosage form of the pharmaceutical composition is a tablet or a capsule.

9. Use of a tenofovir disoproxil phosphoramidate compound according to any of claims 1 to 5 in the manufacture of a medicament for the prevention or treatment of a viral disease.

10. The use according to claim 9, wherein the viral disease is HIV-1 infection or HBV infection or simultaneous infection of HIV-1 and HBV.