JPH01213292A - Anthracycline derivative having inhibition activity of reverse transcriptase of human immunological deficiency disease virus - Google Patents

Abstract

NEW MATERIAL:A compound (salt) expressed by formula I [R is H or 1-18C (substituted)alkyl, aralkyl, aryl or acyl]. EXAMPLE:14-O-(3,4-dimethoxybenzoyl)adriamycin. USE:Antivirus agent for human immunological deficiency disease virus (HIV). Antineoplastic agent. PREPARATION:For example, anhydrous methanol is added to a mixture of a sodium salt of a protocatechuic acid derivatives (e.g., 3,4-dimethoxybenzoic acid) expressed by formula II and hydrobromic salt of 14-bromodaunomycin and suspended. The suspended mixture is subsequently reacted at room temperature for two days and nights and furthermore at 40 deg.C for a day and night while stirred. After the conclusion of the reaction, the reaction solution is purified by using the preparative thin layer chromatography to provide a compound expressed by formula I in a red solid state.

Description

Detailed Description of the Invention (Field of Industrial Application) The present invention relates to a novel compound, 14-0-acyl adriamycin, which is an anthracycline derivative and inhibits the reverse transcriptase activity of the human immunodeficiency virus (CHIV). .

(Prior art and problems to be solved) HIV, which is the cause of acquired immunodeficiency syndrome (AIDS), is a retrovirus, and its reverse transcriptase is considered to be one of the targets for anti-HTV* quality search.

Adriamycin and other drugs are already known to inhibit the reverse transcriptase of avian leukemia virus, but they are highly toxic. Therefore, it is expected that derivatives of HIT with strong reverse transcriptase inhibitory activity and weak cytotoxicity will emerge.

(Means for Solving the Problems) The present inventors are conducting research to meet the above expectations. The present inventors have already prepared 5M of +4-0-(3-flufoxy-4-alkoxykmethyl)penzoyladriamycin, and these atrialysin derivatives are HI
It was discovered that V has strong reverse transcriptase inhibitory activity and weak cytotoxicity (Patent application No. 62-23G97 filed by the present applicant).
No. 2, filed on September 25, 1986). After further research, we discovered that the 14-hydroxyl group of adriamycin is an anthracycline derivative with strong catechol-type transcriptase mastication activity and weak cytotoxicity, as shown in the general formula below. It was completely exhausted.

Therefore, the gist of the present invention is to provide a novel compound having the following formula (I) (wherein R represents a hydrogen atom, or a substituted or unsubstituted linear chain having 1 to 18 carbon atoms or An object of the present invention is to provide an I4-0-acruadriamycin derivative represented by a branched alkyl group, an aralkyl group, an aryl group, or an acyl group, and an acid addition salt thereof.

In the compound of the present invention of general formula (I), when R is an alkyl group, an alkyl group having 1 to 1 g of carbon atoms,
Side light: methyl, ethyl, n-propyl, isopropyl,
n-butyl, isobutyl, tertiary butyl, n-hexyl, n-undecyl t+ is preferably an n-octadecyl group or the like. The case where R is a cycloalkyl group such as a cyclohexyl group is also included. Further, R is a benzyl group,
It can be an aralkyl group such as a phenethyl group, or an aryl group such as a phenyl group or a naphthyl group.

When R is an alkyl group, an aralkyl group, or an aryl group, it can also have a lower alkoxy group, a hydroxy group, a hexagonal group, etc. as a substituent thereon. moreover,
When R is an acyl group, it is preferably an alkanoyl group such as acetyl, propionyl, valeryl, or an aroyl group such as benzoyl group.

When the compound of the present invention of general formula (I) is an acid addition salt,
The acid to be added can be a pharmaceutically acceptable inorganic acid such as hydrochloric acid, sulfuric acid, phosphoric acid, or a pharmaceutically acceptable organic acid such as acetic acid, propionic acid, malic acid, methanesulfonic acid.

Next, examples of the novel anthracycline derivatives of the present invention represented by general formula (I) are summarized in Table 1. Further, the specific optical rotation and melting point of each compound example are also shown.

Next, the physicochemical and biological properties of the novel anthracycline derivative of general formula (I) obtained according to the present invention will be described.

Each of the above compounds of the present invention is a red powder or a crystalline powder, and mass spectrometry and R
The results of measuring the f value (silica gel thin layer chromatography) are shown in the third column after Examples 1 to 5 showing the production examples.
summarized in a table.

Next, evaluation tests for the HIT reverse transcriptase inhibitory activity and cytotoxicity of the novel anthracycline derivative of general formula (I) according to the present invention will be described below.

(f) HTV reverse transcriptase mastication activity was measured as follows. In other words, 9.5%/second HIV-infected cell culture supernatant
After treatment with I+ethylene glycol for 3 hours, centrifugation was performed to prepare 2,5×I OPFU/dOHI V lysate (
33.3% glycerol, 16°7 mmM) Lis-HC
l (pi(7,5), 533mM K(:, 1.0
．． 324 Triton x-1oo dissolved in 3.3mM dithiothreitol] to g411! T, ta. The reverse transcriptase activity of the utVfn solution prepared in this way was
DNA synthesis using globin mRNA as a template and oligo dT as a primer is [α P ] dATP
It was measured by O encroachment rate. That is, 1 μt of old V lysate was mixed with 9 at enzyme reaction solution (50 mM) +) uc/(pii 8.3), 70 mM KC.
I, IQ mMge/2.30 mM mercaptoethanol, 90 nM dNTPs.

3.3ttt/-oligo dT, JL4 mU/-
Human placental RNase inhibitor, 5μt/-β-globin mRNA, 0.14 mci/d, [α
P ] dATP), and after incubation at 37°C for 30 minutes, 10-chain trichloroacetic acid (TCA) was added to stop the reaction.

After the reaction is complete, the entire reaction solution is placed on a nitrocellulose filter to adsorb raw IIZ and DNA, and then filtered with 5% TCA and 20% TCA.
Wash with mM sodium pyrophosphate to remove unreacted [α-
32p) dATP was removed. The amount of raw EDNA on the nitrocellulose filter was calculated by measuring radioactivity.

The novel anthracycline conductor of the present invention was added to the enzyme reaction solution, and the results of IC5o'Ir measurement were compared with those of adriamycin (abbreviated as ADM) and are shown in Table 2 below.

On the other hand, cytotoxicity was determined by adding the novel anthracycline derivative of the present invention to 5 x 10 mouse lymphocytic leukemia cells P-388, stirring and culturing. The degree of cell proliferation after 48 hours was measured by the NTT method. The IC5° value of the novel anthracycline derivative of the present invention was determined by Atriamysia.
Table 2 shows a comparison with that of (ADM).

Furthermore, the HI of the novel anthracycline derivative of the present invention
The ratio of T reverse transcriptase inhibitory activity and cytotoxicity is shown in Table 2 in comparison with that of ADM.

As is clear from the results shown in Table 2 above, the compound of the present invention of general formula (I) exhibits HIV reverse transcriptase inhibiting activity, and therefore has HIM proliferation inhibiting activity, and also exhibits phosphorylation It has anti-cancer activity since it exhibits an activity of inhibiting the proliferation of leukemia cells P-3gg.

The production of the adriamycin derivative having the general formula (I) according to the present invention is carried out by adding the known 14-bromodaunomycin hydrobromide or 14-chloromeunomycin dove salt to the following general formula (1) in an organic solvent. metal salts of derotcatechuic acid derivatives (wherein R has the same meaning as above) (
sodium salt, potassium salt, etc.) or a substituted amine salt at room temperature or with heating. If necessary, chromatography is performed to separate and purify the 14-0-acyl adriamycin derivative i represented by general formula (I).

Examples of the present invention are shown below.

Example 1 Preparation of +4-0-(3,4-dimethoxybenzoyl)adriamycin 3.4-dimethoxybenzoic acid 40.8Ni-prepared sodium salt thereof and 14-bromodaunomycin hydrobromide 76.9q. 3.84 ml of anhydrous methanol was added and suspended. The mixture was stirred at room temperature for two days and nights, and then at 40°C for one day and night. Thin layer chromatography for preparing reaction solution [
The residue was purified using chloroform-methanol-concentrated aqueous ammonia (90:20:l) as a developing solvent to obtain 15.7sIF of the title compound as a red solid.

Example 2 Preparation of +4-0-(3,4-di-n-hexyloxybenzoyl)adriamycin 4 g of 3+4-di-n-bekyloxybenzoic acid, its sodium salt prepared from 0 Misaki and 14-bromodaunomycin bromide. Hydrogen salt 51.:29K.

2.6 mm of anhydrous methanol was added, and the mixture was stirred at room temperature all day and night. Next, 1.3-mL of chloroform was added, and the mixture was further stirred at room temperature all day and night. The reaction solution was purified by preparative thin layer chromatography [developing solvent: chloroform-methanol-concentrated aqueous ammonia (90:20:1)] to obtain 23.1 m9 of the title compound as a red solid.

Add 2.0 ml of this free base to 1.0 ml of chloroform. It was dissolved in 〇- and methanol 1,〇-, and 2.4 μL of 1.0N hydrochloric acid was added. This was concentrated under reduced pressure and the residue was thoroughly dried to obtain 2.1q of the hydrochloride of the title compound. Meanwhile, similarly dissolved free base 2. OwfK, 2.4 μt of 1.0N sulfuric acid was added. This was concentrated under reduced pressure and the residue was sufficiently dried to obtain 2.1 q of 172 sulfate of the title compound. Example 3 Production of +4-0-(3,4-di-undecyloxybenzoyl)adriamycin 3 .4-Di-undecyloxybenzoic acid 14.4 ml of the prepared tetramethylammonium salt thereof and 14-bromodaunomycin hydrobromide 29.0 q. was added 1.0 ml of anhydrous methanol and 1.04 ml of chloroform. They were added one after another and dissolved. After this was left at room temperature for a day and night, the precipitated insoluble matter was subdivided using ultrasonic waves. This reaction solution was purified by preparative thin layer chromatography [developing solvent: chloroform-methanol-concentrated aqueous ammonia (90:20:1)] to obtain 6.319 of the title compound as a red solid.

Dissolve 5,3 Tq of this free base in 1.3 d of chloroform and 4.0 ml of methanol, and add 5.4 μL of 1.0 N hydrochloric acid.
added. This was concentrated under reduced pressure and the residue was thoroughly dried to obtain 5.51 IP of the hydrochloride of the title compound.

Example 4 Preparation of +4-0-(3,4-di-octadecyloxybenzoyl)adriamycin 3. 4-di-octadecyloxybenzoic acid 37.3 ml of anhydrous methanol 2. 〇- and chloroform 2゜0- were added and dissolved by heating.

Then 14-promodaunomyquine hydrobromide 60.
0aF and sodium iodide 198IllF were added, followed by anhydrous dimethylformamide 4.0-. 40
The reaction solution was stirred at C for a day and night, concentrated under reduced pressure, and the resulting residue was extracted with 150% chloroform. This was washed with 20ml of water and then dried over anhydrous sodium sulfate. The residue obtained by concentrating this under reduced pressure was purified by preparative thin layer chromatography [developing solvent: chloroform-methanol-concentrated aqueous ammonia (90:20:l)] to obtain 3.2q of the title compound as a red solid. .

This free base 2. methanol to qq 2.9-11. 2.4μ of Q normal salt and 1.74μ of chloroform were sequentially added and dissolved by heating. This was concentrated under reduced pressure and the residue was thoroughly dried to obtain the title compound hydrochloride 3. Obtained Oq.

Example 5 Production of 14-o-(3,4-di-valeryloxybenzoyl)adriamycin
) 5 o.
5- and 00 form 1.0- were processed and dissolved. After standing overnight at room temperature, the reaction solution was purified by preparative thin layer chromatography [developing solvent: chloroform-methanol-concentrated aqueous ammonia (90:20:1)] to obtain 7.4q of the title compound as a red solid. Ta.

3.7 μL of methanol and 8.7 μL of 1.0N hydrochloric acid were added to 7.4119 of this free base and dissolved. This was concentrated under reduced pressure and the residue was thoroughly dried to obtain the title compound hydrochloride 7.
Obtained 7 capes.

Next, reference examples will be shown regarding several preparation examples of the compound of general formula (1) used in the method for producing the compound of the present invention.

Reference Example 1 Production of 3.4-di-n-hexyloxy7benzoic acid To derotcatechuic acid 200M9 and 60% oily sodium hydride 208TIIQ, anhydrous dimethylformamide 20
- was added, then 912 μt of hexyl bromide was added, and the mixture was stirred at room temperature all day and night. The reaction solution was added dropwise to a 5% aqueous potassium hydrogen sulfate solution, and extracted with chloroform. The chloroform layer was washed with water, dried over anhydrous sulfuric acid and concentrated under reduced pressure.

The obtained brown oil was subjected to silica gel column chromatography [developing solvent: hexane-chloroform (3:2)]
This was purified to obtain colorless oil 441M9. Tetrahydrofuran 22-1 + 096 sodium hydroxide aqueous solution 4.44 and methanol 11- were sequentially added and dissolved, and the mixture was left at room temperature for two days and nights. The reaction solution was concentrated under reduced pressure, and the resulting residue was extracted with 100% chloroform. This was washed successively with an aqueous solution of potassium hydrogen sulfate and water, dried over anhydrous sodium sulfate, concentrated under reduced pressure, and the resulting residue was subjected to silica gel column chromatography [Developing solvent: chloroform-ethanol (3
271 of the title compound as colorless plate-like crystals.
I got 4. Melting point: 124~I26'C6EIMS:
m/z 322 (M).

Reference Example 2 Production of benzhydryl derotcatechuic acid Ethyl acetate 2.5- was added to and dissolved in 50.0119 of derotcatechuic acid and 63.0119 of diphenyldiazomethane, and the mixture was allowed to stand at room temperature for 2 hours. Furthermore, 25.8119 g of diphenyldiazomethane was added, and the mixture was left to stand at room temperature for 2 hours.

The colorless residue obtained by concentrating the pale reddish-purple reaction solution under reduced pressure was purified by silica gel column chromatography [developing solvent: chloroform → chloroform-methanol (30:I)] to obtain the title compound 5114 as a colorless syrup. I got it. E
tMS; m/z 320+ (M).

Reference Example 3 Production of 3.4-di-undecyloxybenzoic acid Derotcatechuic acid benzhydryl ester 49.0 and 60% oily sodium hydride 13.5 were added and suspended in anhydrous dimethylformamide 4.9. .

Then, 77.8 μt of undecyl iodide was added, and the mixture was stirred at room temperature for 2 hours. The reaction solution was added dropwise to a 5% aqueous potassium hydrogen sulfate solution, and extracted with chloroform 5091. After drying the chloroform layer with anhydrous sodium sulfate,
It was concentrated under reduced pressure to remove only chloroform.

5.0-trifluoroacetic acid was added to the obtained dimethylformamide solution and left at room temperature for 30 minutes. The reaction solution was concentrated under reduced pressure, and the resulting residue was extracted with 50% chloroform. This was washed with a 2.5% aqueous potassium hydrogen sulfate solution, then dried over 3 um of anhydrous sulfuric acid, and concentrated under reduced pressure. The obtained residue was subjected to preparative thin layer chromatography [
Developing solvent: Chloroform-methanol < +0: +)
] to obtain 48.5 q'f of the title compound as a colorless solid. Melting point;'? 8~I Ol'c, EtMS ;m/
z 462 (M+).

Reference Example 4 Production of 3.4-dioctadecyloxybenzoic acid To 51.3 cm of benzhydryl derotcatechuic acid and 14.0 cm of 60% oily sodium hydride, 5.1 cm of anhydrous dimethylformamide was added and suspended.

Then add octadecyl iodide l 3 Array,
The mixture was stirred at room temperature for 2 hours. The reaction solution was added dropwise to a 2.54 aqueous potassium hydrogen sulfate solution 509Pt, and extracted with 50% chloroform. After drying the chloroform layer over anhydrous sodium sulfate, it was concentrated under reduced pressure to remove only chloroform.

5.0 dt of trifluoroacetic acid was added to the obtained dimethylformamide solution, and the mixture was left at room temperature for 6 minutes.

Next, 5.04 ml of chloroform was added and left at room temperature for 10 minutes. The reaction solution was concentrated under reduced pressure, and the resulting residue was extracted with 50% chloroform and washed with a 2.5% aqueous potassium hydrogen sulfate solution. Next, it was dried over anhydrous sodium sulfate, concentrated under reduced pressure, and the resulting residue was subjected to preparative thin layer chromatography [developing solvent: lyaloform-methanol (I0:
47.3119 of the title compound as a colorless solid purified by
I got it. Melting point; 94-II7°C0EIMS; m/z
658(M).

Reference Example 5 Production of 3.4-di-valeryloxi-7benzoic acid Anhydrous pyridine 2.0- was added to and dissolved in derotcatechuic acid benzhydryl ester 67.5, and valeryl chloride 55.5- was dissolved.
9TMi was added and left at room temperature overnight. 1 part water to the reaction solution
7 μL was added, concentrated under reduced pressure, and the resulting residue was extracted with chloroform. This was washed successively with a 5% aqueous potassium hydrogen sulfate solution, a saturated aqueous solution of sodium hydrogen carbonate, and water, and dried over anhydrous sulfuric acid.

Yellow oil obtained by concentration under reduced pressure'? 2. gqK) 1.0- of fluoroacetic acid was added and dissolved, and the mixture was left at room temperature for 10 minutes. This was concentrated under reduced pressure, and the resulting residue was azeotroped twice with toluene. It was purified by preparative thin layer chromatography [developing solvent: chloroform-methanol (g: +)] to obtain 24.8119 of the title compound as a colorless solid. Melting point;
97-103°C, EIMS; m/z 322 (M).

Procedure Nefu Seisho (self-motivated) February 1st, 1989

Claims (1)

[Claims] 1. The following general formula (I) ▲There are mathematical formulas, chemical formulas, tables, etc.▼(I) (In the formula, B represents a hydrogen atom or a substituted or unsubstituted carbon Anthracycline derivatives represented by 1 to 18 linear or branched alkyl groups, aralkyl groups, aryl groups, or acyl groups, or acid addition salts thereof. 2. The derivative or acid addition salt thereof according to claim 1, wherein in general formula (I), R is a methyl group. 3. The derivative or acid addition salt thereof according to claim 1, wherein in the general formula (I), R is a hexyl group. 4. The derivative or acid addition salt thereof according to claim 1, wherein in the general formula (I), R is an undecyl group. 5. The derivative or acid addition salt thereof according to claim 1, wherein in the general formula (I), R is an octadecyl group. 6. The derivative or acid addition salt thereof according to claim 1, wherein in the general formula (I), R is a valeryl group. 7. The derivative or acid addition salt thereof according to claim 1, wherein in the general formula (I), R is a hydrogen atom.