SU797584A3 - Method of preparing n-substituted bleomycin derivatives - Google Patents

Description

one

The invention relates to a process for the preparation of novel N-substituted bleomycin derivatives having valuable pharmacological properties.

The purpose of the invention is to obtain new useful compounds that expand the means of action on a living organism.

The goal is achieved by synthesizing indicated compounds based on the known nucleophilic substitution reaction of amines Cll.

A method for the preparation of N-sa substituted bleomycin derivatives is proposed. of general formula I

n

Well if, “/ and

- / b / -% D OH N o ij

---- v

ABOUT

0.; - n1

 he N

N. where H is a group of the formula -COR, where R is hydrogen, -alkyl, halo-substituted C -C-alkyl, croaonoyl, phenyl, p-nitrofenyl, benzyl, p-nitrobeneyl, phenoxymethyl, cyanamyl, naphthyl, nicotinoyl, isonicotinoyl, furyl or thienyl, a group of the formula -CO- (CH2) l, -GOOH, where n 2-7; - yo-yn-eeoon, where K is hydrogen or methyl; -CH-CHrj-CH (CH,) - COOH or -CO-CH "-C (gSNp) -CO. oi p where R is —C — C) alkyl, allyl, benzyl, or p-nitrobeneyl; a group of the formula —SOjR, where, —C, 2 is alkyl; a group of the formula where Z. is hydrogen, methyl, a chlorine atom or a nitro group. or naphthyl; a group of the formula - CONHR, where.-alkyl cyclohexyl, phenyl, p-tolyl or naph. til, group of formula

About MHt

"/

HI

with a reagent selected from the group consisting of compounds of the formula R-Y, where R is as defined above;

Y is carboxyl, an acid halide group; ester group, group of formulas -CO-0-COR or - C O Mgr; compounds of formula

do.

(

about . do

where n - 2-7,

compounds of formula

to -eo.

dHz-CoMH (eH,),) isiHi

where. has the values given

above,

compounds of formula

K-C- do.

-ABOUT,

55Cn-CO where R has the values given above, of the formula

Ji2Ci d- (io 1 IHg-CO

compounds of formula

-CO-K (J,

R where R is hydrogen, C -Cg-alkyl, a group of the formula - {CH, j), NZ Z, where Z- and Z are hydrogen or methyl, a group of the formula where Z. is hydrogen, methyl, a nitro group or methoxy group, a group of the formula where 2. - a hydrogen or chlorine atom or a nitro group, styryl or naphthylmethyl, a group of the formula </ BR> where R is a hydrogen or nitro group, consisting in the fact that 3- (4-aminobutyl) -aminopropylamino bleomycin of the general formula II is reacted where R has the meanings given Compounds of the formula R-X, where R has the meanings given above, X is a halogen atom. compounds of the formula -X, where R and X have the meanings given above, compounds of the formula where R has the meanings given above, compounds of the formula R &apos; -di - d-OR n where R has the meanings given

/ "N

f R-dO - 3 I

B-rio ,,

g

0

KO-CO-

(eng) „0

 --yo

f

H-CJOv. I / 0

Cng-yo

cJHt (i-cioI

dHrcfo

dH-CoJ c

k

dH- do-

 - dov

I 0

I

dH-eo

Table

d - {(iHt) rt-dooH i

d- dHj-dH-dooH li11

BLM-NH-d-dHj-dH-doOH

BLM-NH-d-dH cJn-dooH.

0

BLM-NH-d-dH-d dooH HI, ft -C-Cij-alkyl; or a compound of the formula wherein R is hydrogen or a nitro group, in an amount of 1-10 moles of the said reagent per -1 mole of bleomycin at 10-65 ° C, in the presence of a solvent. An example of a reaction for the preparation of N-substituted derivatives of blemycin Ag in accordance with the invention can be represented as shown in Table. one.

I m

BLM -NH- do

dOOH

to

R -NCO

R5- e - OK and NH

In the above formulas, BLM means a residue of bleomycin without its terminal amino side group, Rg-R is as defined, R is lower alkyl, substituted or unsubstituted phenyl.

Reagents (representatives of reagents) are also presented, except when R corresponds to the formula R-Y.

Of these models, reaction 1a represents a method for directly N-acylating bleomycin AB with a carboxylic acid in the presence of carbodiimide. Models Ib-lg represent the M-acylation of bleomycin Ag with an acid halide, carboxylic acid ester, carboxylic acid azide, carboxylic anhydride, acelimidazole, and mixed anhydride, obtained by reacting the alkoxyhexylformate with the carboxylic acid. Lh-11 models are methods for the N-acylation of bleomycin Aj by intramolecular (internal) diacid anhydride. : Compounds that can be used as reagents in accordance with the invention are: formic acid, acetic acid, propionic acid, butyric acid, valeric acid, caprylic acid, lauric acid, crotonic acid, benzoic acid, phenylacetic acid , 2-naphthoic acid, p-nitrobenzoic acid, acetyl chloride, propionyl chloride, chloroacetyl chloride, chloropropionyl chloride, bromoacetyl chloride, benzoyl chloride, p-nitrobenzoyl chloride, phenylacetyl chloride, phenoxyacetyl chloride, et octacinate, ethyl caprylate, benzoyl azide, propionic acid azide, butyric acid azide, acetic anhydride, propionic anhydride, butyric anhydride, octanoic anhydride, lauric anhydride, crotonic anhydride, phenylacetic anhydride, phenoxyus anhydride, acetic anhydride, crotonic anhydride, phenylacetic anhydride, phenoxysuccinus anhydride, acetic anhydride, crotonic anhydride, phenylacetic anhydride, acetic anhydride, acetic anhydride, acetic anhydride, acetic anhydride one

BLM-NH-SO -RBLM-NH-d-NH-B I

BLM-NH-t-B NH

RID, anhydride, and language; anhydride, cork anhydride, azelaic anhydride, phthalic anhydride, 3-nitrophthalic anhydride, methyl chloride, methyliodide, ethyl bromide, butyl bromide, allyl bromide, 1-nitrobenzyl chloride chloride, benzyl bromide, n-nitrobenzyl bromide, methylsulfonyl chloride, etilsulfonilhlorid, benzenesulfonyl, p-toluenesulfonyl, p-chlorobenzenesulfonyl chloride, p-nitrobenzenesulfonyl chloride, f} -naftalinosulfonilhlorid, isocyanate, etnlizotsianat, phenylisocyanate, tsyklogeksilizotsianat, p-metilfenilizotsianat -naftilizotsianat c, p -naftilizotsianat, ethyl formate hydrochloride, methyl acetamide hydrochloride, methylpropionimide hydrochloride, methyl isobutyroimide hydrochloride, methylvaleroimide hydrochloride, methylpelargonimide hydrochloride aydrohidori phenylacetamide, methyl hydrochloride. m-toluimide ester, p-toluimide methyl ester hydrochloride, p-chlorophenylacetamide methyl ester hydrochloride, cinnamic acid imide methyl ester hydrochloride, Oi-naphthylacetimide methyl ester hydrochloride.

The proposed method is carried out in a solvent selected from the group including water, an alcohol, for example, methanol and ethanol, a ketone, for example acetone, and mixtures thereof. The solvent is selected in accordance with the reagent used. When alkyl halides are used as reagents, for example. ethylnodide, acyl galgrundum, alkylsulfonyl halide, the reaction proceeds quickly and loading of the excess component leads not only to the target monosubstitution of the amino group in the side chain of bleomycin Ag, but also to the substitution of the intra-ring amino group and to the substitution (introduction of two substituents) to the terminated groups, the endgroup of amino groups and the endgroup of groups containing groups of endgroups of groups of endgroup groups amino groups in the terminal side chain). Therefore, it is advisable to introduce these components in a ratio of 1-1.5 mol per 1 mole of bleomycin Ag. However, less reactive reagents, which do not cause side reactions, can be introduced in an amount of up to 10 moles per 1 mole of ble omicin. The reaction carried out in accordance with the invention can be regarded as the nucleophilic displacement reaction of the terminal amino group of bleomycin Ag and it is expediently carried out in an alkaline medium. Pyridine, bicarbonate, sodium, triethylamine and the like are used for this purpose. When using imidates as a reagent, it is advisable to conduct the reaction at a pH of 8.5-9.5 by adding a stronger base, triethylamine. However, prolonged reaction at pH 10 or wine should be avoided, taking into account the stability of bleomycin A5 The high molecular weight substituted derivatives, for example lauroyl bleomycin AS, are poorly soluble in water. Therefore, in such cases, the reaction liquid (mass, mixture) should be subjected to purification by august chromatography, directly filtered through a gel of an organic solvent, for example, through a Zephshexdex Z-H-20, but the process of precipitation is unnecessary. In tab. 2 summarizes the properties of the N-substituted derivatives of bleomycin Ag, which are obtained by using the hydrochloride method, obtained.

rH

in um

Vd

-ABOUT

ABOUT -

r

n rЧ

vo r-tn

n

 CN

GN N

in mid

CM

VO r VO

O

“O

0

o

CO

 (N

CO CM

n (N GO SL H CTi GM tN M M

about

(U

2§

/

about

about

L) a

-U "

five

5 (d n

0)

s

X

I

about

I

10 The M-e-substituted derivatives of bleomycin Ac obtained by the proposed method do not have a clear melting point, a decomposition point or a boiling point. They are soluble in water and methanol, form salts with acids and careless (complex) compounds with copper, give a positive reaction to Pauli, Ehrlich, Dragendorff and permanganate; on ferric chloride. N-substituted derivatives of bleomycin Ag find in the infrared spectrum region an absorption zone at the following waves, cm: 1040-1075, 1710-1735, 1625-1680 and 2910-2940. Example 1. Preparation of N-chloroacetyl bleomycin Ag. 1 g of bleomycin Ag hydrochloride is dissolved in a mixture of 13 l of water and 20 ml of acetone. 520 mg of sodium bicarbonate and 330 mg of chloroacetyl chloride are added to the solution with stirring. The mixture is left to stand for 1 hour and 45 minutes. The pH of the mixture is adjusted to 4.5 with hydrochloric acid. The acetone is distilled off in vacuo. The residue is diluted with water and passed through a column with a suspension of 110 ml of CM-Sephadex C-25 in a 0.02 M aqueous solution of ammonium chlorin to adsorb the desired product. The adsorbed product was eluted successively with 230 ml of a 0.1% aqueous solution of ammonium chloride and 580 M of a 0.2 M aqueous solution of ammonium chloride. Collect fractions with the desired product and desalted with Amberlite CC-50. The eluate is evaporated to dryness and 439.6 mg of N-chloroacetyl bleomycin Ac are obtained. Yield 41.7%. Similarly, from p-nitrobenzoyl chloride and poluchat 4- {p-nitrobenzoyl) -bleomycin AZ. Example 2. Obtaining N-octanoylbleomycin Ag. 100 mg of bleomycin Ag are dissolved in a mixture of 2 ml of ethanol and 0.5 ml of water. 50 mg of ethylcaprylate L is introduced into the solution and the mixture is kept at 37 ° C for 16 hours. At the end of the reaction, the solvent is distilled off under vacuum. 10 ml of water are added to the residue. The mixture is extracted with 10 ml of ether. The aqueous layer is passed through a column with a suspension of 20 ml of CM-Sephadex C-25 in a 0.02 M visible ammonium chloride solution in order to adsorb the desired product. The adsorbed product is eluted sequentially with 120 ml of a 0.1 M aqueous solution of ammonium chloride and 44 ml of 0.3 M ammonium chloride solution. Collect fractions containing the desired product, and desalted with amberlite CC-50. The eluate is evaporated to dryness, to obtain 28.2 mg of N-octanoylbleomycin Ag. Yield 5.5%. Example 3. Obtaining N-nopionylbleomycin A5. 100 mg of bleomycin A are dissolved in a mixture of 2 ml of acetone and 0.5 ml of water. 1 ml of an acetone solution containing 10 mg of propionic azide obtained from propionic acid hydrazide is added to the solution. The mixture is left to stand for 26 hours. Upon completion of the reaction, the acetone is distilled off under vacuum. 10 ml of water are added to the residue. The mixture is passed through a column with a suspension of 20 ml of CM-Sephadex C-25 in a 0.02 M aqueous solution of ammonium chloride to adsorb the desired product. The adsorbed product was eluted sequentially with a 0.1 M aqueous solution of ammonium chloride and a 0.2 M aqueous solution of ammonium chloride. Collect fractions containing the desired product, and desalted with amberlite. The resulting eluate was evaporated to dryness and 21.4 mg of N-propionyl bleomycin Hell was obtained. Yield 20.6%. Example 4. Obtaining N-acetylbleomycin Ag. 1 g of bleomycin A5 hydrochloride is dissolved in 50 ml of methanol. 0.78 ml of pyridine and 0.58 ml of acetic anhydride are added to the solution with stirring. The mixture is left for 5 hours at room temperature. Then the reaction mixture is evaporated under vacuum. The concentrate is dissolved in a small amount of water. The solution is passed through a column with a suspension of 110 ml of CM-Sephadex C-25 in a 0.02 M aqueous solution of ammonium chloride to adsorb the desired product. The blue absorption band (containing the desired product) was eluted with a 2 M aqueous solution of ammonium chloride. Collect the eluate fractions containing the desired product and pass through a column of 70 ml of a suspension of activated carbon in water to absorb the desired product. The adsorbed product is washed with water and eluted with acetone (0.02 N). hydrochloric acid (1: 1) by volume. The eluate is evaporated to dryness and 552.3 mg of M-acetyl bleomycin Ag hydrochloride are obtained. Yield 53.6%. The compounds listed in Table 2 are prepared analogously. 3

m y s c; you fd

tn

a: o

ABOUT

0)

e; about

about

l 1L

X

S

X

i§x Example 5. Preparation of N-butyrylbleomycin Ag., Prepare a solution of 100 mg of bleomycin Ag in a mixture of 3 ml of acetone and 2 ml of water. 12 mg of hydrochloride 1-ethyl-3-dimethylaminopropylcarbodiimide and 8 mg of butyric acid are added to the solution. The pH of the mixture is adjusted to 4.7 and the mixture is left to stand for 16 hours. The reaction mixture is evaporated to one third of the original volume under vacuum. To the concentrate is poured io ml of water. The mixture is passed through a column with a suspension of 20 ml of CM-Sefavdeks C-25 in a 0.02 M aqueous solution of ammonium chloride. The adsorbed product was eluted with a 0.1 M aqueous solution of ammonium chloride. Collect fractions containing the batch product and pass through a column with 20 ml Amberlite CC-50 YH-type) to adsorb the desired product. The product I adsorbed is successively 100 ml of water, 100 ml of 0.5% acetic acid and 100 ml of water, and then eluted with an aceto system of 0.02 n. hydrochloric acid (1: 1 by volume J. The eluate is evaporated to dryness and 32 mg of N-butyrylbleomycin A are obtained.

N-Allylbleomycin AS

N-Ethylbleomycin AS

Continued table.

1.6 25 0.1- 70

0.25 (34.0)

25 0.1-840, 25 (41.5) Example b. Preparation of N- (n-nitrobeneyl) -bleomycin A. 1 g of bleomycin At hydrochloride At is dissolved in a mixture of 20 ml of water and 60 ml of ethanol, 0.56 ml of triethylmino and 600 mg of p-nitrobenzyl bromide are added to the solution with stirring. The mixture is left for 35 hours at room temperature. The reaction liquid is evaporated and the inorganic material is filtered off. One stripped off liquid is passed through a column with a suspension of 150 ml of Sephadex C-25 in water to adsorb the target product, 35 ml of the initial eluate fractions are set aside and 27 ml of the last part of the fractions are passed through a column of 50 ml of CM-Sephadex C-25 to adsorb the target product. The adsorbed product was eluted in succession with O, 1 M aqueous solution of ammonium chloride and 0.25 M aqueous solution of ammonium chloride. Collect fractions of the eluate with the desired product and treat with activated charcoal. The resulting solution was evaporated and 120 ml of M- (p-nitrobeneyl) -bleomycin A hydrochloride were obtained. Yield 11%. The compounds obtained in Table 2 are obtained analogously. 4. Table 4

Example 7. Getting N- (Methylsulfonyl) -bleomycin Ag.

100 mg of bleomycin Ag hydrochloride are dissolved in 4 ml of acetone and 8 ml of distilled water. 160 mg of sodium bicarbonate and 104 mg of methanesulfonyl chloride are added to the solution, dropwise, with stirring. The mixture was allowed to sit at 19 hours at room temperature. . The reaction mixture is neutralized and passed through a column with 5 ml of Amberlite IRA C00 (C 1 -type) in distilled water to adsorb the desired product. The adsorbed product is washed with 100 ml of distilled water. Collect the eluate and prokunnoi liquid. The mixture is passed through a column with a suspension of 20 ml of Sephadex C-25 in 0.02 M aqueous

N-Benzenesulfo 200 Nilbleomycin Hell

N- (p-Toluenesulfonkl) -gly6mycin Ag 200

N- (p-Chlorobenzenesulfonyl) -blueN-Benzene sulfo20 nilbleomycin AS

N- (P-TOLUOLSULFO-gg

Nile) -bleomycin A5

N- (p-Chlorobenzene 40 sulfonyl) -bleomycin A5

N - (- Naphthalene-Q-fonil) bleomycin A-s

ammonium chloride solution. The adsorbed product is eluted sequentially with 200 ml of a 0.1 M aqueous solution of ammonium chloride and 78 ml of a 0.2 M aqueous solution of ammonium chloride e, the eluate fractions containing the desired product are collected and passed through a layer of 5 ml of Amberlite CC-50 (H-type) . The resin bed is washed successively with 25 ml of water, 25 ml of 0.5% acetic acid and 25 ml of water, the target product is eluted with an acetone system — 0.02 n. hydrochloric acid (1: 1 by volume). The eluate was evaporated to dryness and, from 16.2 mg of N- (methylsulfonyl) -bleomycin Ag, hydrochloride was obtained. Yield 15.3%.

The compounds listed in Table 2 are prepared analogously. five.

Table 5

3

34.6

four

four

00gs11

4 SNS

81.7

 ii

Ojcjl

45 4

Continued table. five

27.6

0.130 (12.6) 0.2

19.5

od20 (3.8)

0.3

75.9

0.140 (35.8) 0.1

47.1

0.150 (20.7) 0.2

Example 8. Getting M- (ethylcarbamoyl) -bleomycin Ag.

200 mg of bleomycin Ag hydrochloride is dissolved in 10 ml of water. 60 mg of sodium bicarbonate are added to the solution and 0.1 ml of ethyl isocyan a is added dropwise with stirring. The mixture was incubated for 4 hours and 45 minutes at room temperature, filtered, the filter was adjusted to 4. The filtrate was passed through a column of 30 ml CM-Sephadex C-25 to adsorb the desired product - AD

N- (Phenylcarbamoyl) -bleomyin and A

500

N- (Cyclohexylcarbamoyl) -bleomia2ao zing A g

N- (Phenylcarbamoyl) -bleomycin Hell

N- (Cyclohexylcarbamoyl) -bleomycin p and Mp, 9. Preparation of M -. (Acetimidoyl) - bleomycin Ag.

Dissolve 600 mg of bleomycin A5 hydrochloride in 15 ml of water. 90 mg of methyl acetimidate hydrochloride are added to the solution. The pH of the mixture was adjusted to 8.8 by addition of triethylamine. The mixture is stirred for 4 hours at room temperature and then evaporated to one third of the original volume. The pH of the concentrate is adjusted to 7. The reaction liquid is passed through a column with 60 ml of CM-Sephadex C-25. The adsorbed product is eluted sequentially with a 0.1 M aqueous solution of ammonium chloride and a 0.25 M aqueous solution of ammonium chloride. Fractions of the eluate with the desired product are collected and treated with activated charcoal. The resulting solution was evaporated to give 100 mg of H- (ethylcarbamoyl) -bleomycin iHCl hydrochloride. Output yield 28.6%.

Similarly, compounds are prepared

presented in table. 6

Table b

--t-Q 0.08

20

-daO

0.05

100

Continued table. b

0.150

80 (15) 0.5

0.130

60 0.25

The target product is eluted with 417 ml of a 0.3 M aqueous solution of ammonium chloride and 320 ml of the second half of the eluate is passed through a column with 40 ml of activated carbon. . The coal layer is washed with water and eluted with acetone system -0.02 n. hydrochloric acid (1: 1 by volume). Eluate

5 is evaporated and 410 mg of N- (methylacetimidoyl) -bleomycin Ag. Hydrochloride are obtained. Yield 65%.

Similarly, compounds obtained in Table 2 are prepared. 7

1Л 1Л Gg- -

1L tN

O (N

CN

,

-and

W

:I

sh

CN

about

: ё

oh oh

I

S

five

S

OH

AS

sha

e; s

(OS

ha about - p

i

00

N

in

g

P

about r

vo

1L

ate h from co-v 3 -f

143in -1L

L

 Sh

1L N

(N

rsi gn tS

about um

about

(N

Yu

O (N

ky

4D

vj

About Ti

Ti

about

H

(N

1L

um

M

g

r "

-with

M CC

 -and-5 W

one

K rj

 oh oh

Yu

 2

iJ xj.3)

 s

0W

. I

1T "

I and

V,

oh oh

about

oh oh

oh oh

oh oh

oh

(N

ABOUT)

I

I in

S d; S Well

with;

with; to about

with S

X PG

S f t S

about

a) S OK

h

S

C

0) S

S 40

to

I

one

2

about 2

t

i0)

Sho o)

S

a

sss; S

 well

0u a:

;. h:

n with: h to

ё-§ VI

S nI

0d

S d

)

0) S R

I

vi

S

S

1S

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5. About

with about

with

with about

rg - O) - S

(C

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I with:

1s;

(

Mr. S

Claims (1)

1. Preparative Organic, M.-L., Himi, 1964, p. 396406. where X has the values given above, compounds of the formula where R has the values given above, and X represents a halogen atom, compounds of the formula j-X where k3 and X have the values given above. compounds of the formula RT-NCO, where R has the meanings given above, compounds of the formula I and NH where R has the meanings given above, and, Cs-alkyl, or C-C-. no formula