IT8349082A1 - AMINOGLUCOSIDE DERIVATIVES, THEIR PREPARATION AND THEIR USE AS MEDICINES. - Google Patents

Description

DESCRIPTION of the industrial invention entitled:

"Aminoglycoside derivatives, their preparation and use as medicines".

SUMMARY OF THE INVENTION

The invention relates to compounds of formula I

,'6

6'CHR5

? 0 (I)

3 *\j 2 ;;1 H 1 ;im ;;wherein - 1 ;;R-j represents hydrogen or a group of formula ;6"CH?0H ;;?0 ITHCH ? HCH HCH ;H C OH ;;(? ) (Ila) (Ib) (Ile) ;;Rg represents a hydroxy or amino group, ;R^ and R^ each represent hydrogen or a hydroxy group, ;Rg represents an amino, methylamino, dimethylamino group or, when R2 represents an amino group, also a hydroxy group, and ;Rg represents hydrogen or a methyl group. ;These compounds can be used in therapeutics as antibiotics. ;;The present invention relates to aminoglycoside derivatives, their preparation and their use as medicines. ;The invention relates more particularly to the compounds of formula I ;;6'CHR5 ;3 2\NH2 (I) ;R-> 1 H 1 ;CH ;;wherein ;R.j represents hydrogen or a group of the formula ;6 "CH-OH ;-o r HCH HCH HCH ;;H C ;;(II) (H a) (H b) (I e) R2 represents a hydroxy or amino group, ;Rg and R^ each represent hydrogen or a hydroxy group, ;R^ represents an amino, methylamino, dimethylamino group or, when R? represents an amino group, also a hydroxy group, and ;Rg represents hydrogen or a methyl group, ;in the form of a free base or in the form of an acid addition salt. ;According to the process of the invention, to prepare the compounds of the invention ;;a) a compound of the formula IIIa or IIIb is reduced ;?. ;CHR ;;H?X (11la) ;;CHR ;;(IHb) ;;where X represents a group susceptible to elimination and R,I to Rb, have the meanings already indicated, and the amino and/or hydroxo groups may be protected, and, if desired, the protecting groups of the protected groups still present in the compound thus obtained are eliminated, or ;b) to prepare a compound in which Rg represents a methylamino or dimethylamino group, a corresponding compound of formula I in which Rg represents an amino group is subjected to a mono- or dimethylation or a corresponding compound in which Rg represents a methylamino group is subjected to a monomethylation, other amino groups possibly present being able to be protected, and, if desired, the protecting groups of the protected amino groups in the compound thus obtained are eliminated, and the compounds of formula I thus obtained are recovered in the form of free base ? in the form of an acid addition salt. ;Procedure a) can be carried out in the usual manner for such reductions known for the hydrogenolytic cleavage of an aziridine ring, for example, using Raney nickel and hydrogen in an inert solvent such as an alcohol, e.g., methanol. It may be advantageous to operate under elevated pressure. ;? Process b) can be carried out using usual methylating agents. For example, for the introduction of a single methyl group into a compound containing a ?'-?? group, this is first reacted selectively with a carbonic acid derivative of the formula Y-CO0Rg where Y represents a reactive group and Rg means an alkyl or aralkyl group, to prepare the corresponding NHCOORg derivative which is then reduced in the usual manner, e.g., with a complex metal hydride, e.g., LiAlH^. The introduction of two methyl groups into compounds containing a α-α group can, for example, be carried out by protecting all the amino groups except the one at the 6-position, introducing methyl groups in the usual way, for example by reacting with formaldehyde in the presence of a reducing agent or NaH2O2PO4, and then removing the protecting groups from the final product. The introduction of a further methyl group into a α-methylamino compound can be carried out in a similar manner. Suitable protecting groups include, for example, benzyloxycarbonyl, tert-butoxycarbonyl, or trifluoromethylcarbonyl groups. These groups can be introduced and removed in the usual way, for example, in a manner analogous to the methods described in the following examples. Examples of X groups that can be removed include mesyloxy, tosyloxy, trifluoromethanesulfonioxy, and chlorine, bromine, or iodine. ;The compounds of formula I can be converted in the usual way into their acid addition salts and vice versa. ;The starting products of formula IIIa can be obtained by introducing in the usual way an X group into a compound of formula VI ;;% ;CHR, NH_ (VI) ;NO. ;;/?> ;H2OH ;;where R-| and Rg have the meanings already indicated ;;The starting products of formula IIIb can be obtained by cyclizing a compound of formula IV ?CH?R ;;(IV) CH20H ;;where R-j and Rg have the meanings already indicated. ;The amino and hydroxy groups can be protected except for the amino group in position 1 in formula IV. ;The cyclization is carried out in the usual manner or using triphenylphosphane and an azodicarboxylic acid ester, for example, azodicarboxylic acid diisopropyl ester, in an inert solvent such as a chlorinated hydrocarbon, for example, chloroform. ;The compounds of formulas IIIa and IIIb are new and are also part of the present invention. ;The compounds of formulas IV and VI and the methods for their preparation are described in European patent application No. 0072351. ;The starting products may be used in the form of mixtures such as those obtained when the preparation is carried out according to fermentation methods. The reaction of these compounds leads to corresponding mixtures of intermediates or final products. Such mixtures may be separated into their individual components at any stage of the preparation, but this is preferably done with the unprotected final products of formula I. The methods used for the separation are usual methods as described in U.S. Patent No. 3984 393 or by K. Byrne and co-workers in J. Chromatogr. 131, 191 (1977). ;The compounds of formula I possess interesting chemotherapeutic properties, particularly antimicrobial, as shown by serial dilution tests carried out in vitro and by tests carried out in vivo on mice using various strains of bacteria, such as for example Staph. aureus, Staph. epidermidis, Pseudomonas aeruginosa, E. coli, Proteus vulgaris, Proteus rairabilis, Proteus morganii, Enterobacter cloacae, Klebsiella pneumoniae and Serratia marcescens. This activity has been observed in vitro at concentrations between about 0.05 and 50 µg/ml and in vivo at concentrations between about 0.4 and 100 mg/kg of body weight of the animal. They are particularly effective against aminoglycoside-resistant strains and therefore possess a broad spectrum of activity. ;;Due to these properties, the compounds of formula I can be used therapeutically as chemotherapeutic agents, especially as antibiotics with antibacterial activity. ;For therapeutic use, an appropriate daily dose is between about 0.1 and 2 g of active substance, which can be administered in divided doses 2 to 4 times a day in the form of unit doses each containing about 25 to 1500 mg of the compound, or in a prolonged-acting form. ;The compounds of formula I can be used in the form of the free base or in the form of chemotherapeutically acceptable acid addition salts, for example in the form of pentahydrochloride or sulfate. These salts exert the same order of activity as the corresponding free bases. The compounds of formula I may be used in conjunction with chemotherapeutically acceptable diluents and vehicles and, optionally, with other excipients and may be administered in the form of tablets, capsules or injectable preparations. Such pharmaceutical compositions are also part of the present invention. The invention also encompasses the compounds of formula I in the form of the free base or in the form of a chemotherapeutically acceptable acid addition salt for use as chemotherapeutic agents, especially as antibiotics with antibacterial activity. The preferred meanings of the substituents are as follows: ;R-j = a) a group of formula IIa ;b) a group of formula II ;= a) a hydroxy or amino group ;b) an amino group ;;R3+R4 = a} hydrogen ;b) a hydroxy group ;= a) an amino, methylamino, dimethylamino group b) an amino group ;R, = a) hydrogen or a methyl group ;b) hydrogen; ;as well as combinations of these meanings. ;;A particular group of compounds of the invention is that in which ;Ri represents a group of formula IIa, ;R2 represents an amino group, ;R^ e represent hydrogen and ;R^ represents hydrogen or a methyl group and ;Rj represents an amino, methylamino or dimethylamino group. Another interesting group includes compounds where R1 represents a group of formula II, R2 represents a hydroxy or amino group, R3 e represent a hydroxy group, Rg represents an amino group, and Rg represents hydrogen. Another interesting group includes compounds where R1 represents a group of formula II or IIa, R2 represents a hydroxy or amino group, R3 e represent hydrogen or a hydroxy group, Rg represents an amino, methylamino, or dimethylamino group, and Rg represents hydrogen or a methyl group. A particularly preferred compound is 1-C-methylaminogentamicin C-1 in the form of the free base or in the form of an acid addition salt, preferably the pentahydrochloride. The invention also comprises a medicament containing, as the active ingredient, a compound of formula I in the form of a free base or in the form of a chemotherapeutically acceptable acid addition salt. The following examples illustrate the present invention without limiting its scope in any way. Temperatures are all given in degrees Celsius, Example 1; 1-C-methylgentamicin C,, pentahydrochloride [method a)]; 1 g of 3,2l,6J,3"-tetra-N-tert.-butoxycarbonyl-1-desamino-1-nitro-1-C-methanesulfonyloxymethylgentamicin C2 (compound of formula IIIa) is dissolved in 30 ml of methanol and reduced with hydrogen in the presence of a Raney nickel suspension at 4 atmospheres. After removing the catalyst by filtration, an amorphous foam is obtained which is taken up in chloroform, shaken three times with dilute ammonia and washed until neutral. After eliminating the solvent in a rotary evaporator, a residue is obtained which dissolves in 10 ml of trifluoroacetic acid and after 7 minutes is diluted with 200 ml of diethyl ether. The residue is filtered by suction. The precipitate formed is dissolved in methanol and transformed into pentahydrochloride on IRA 401 S (in the form of chloride). Rf = 0.17 (in a mixture of dichloromethane, methanol and 25% ammonia in the ratio of 70:26:9) NMR spectrum: 1.29 (d, J = 7, 3H); 1.38 (s, 3H); 1.52 (s, 3H); 2.44 (dd, J] = 13, J2 = 3.6, 1H); 2.96 (s, 3H); 5.10 (d, J = 3.6, 1H); 5.86 (d, J = 3.6, 1H). Example 2: 1-C-methylgentamicin, pentahydrochloride [method a)] Dissolve 0.5 g of 1-C-hydroxymethyl-S^ '^ ?.S"-tetra-N-tert.-butoxycarbonyl gentamicin and 0.26 g of triphenylphosphane in 10 ml of anhydrous chloroform, add 0.2 g of diisopropyl ester of azodicarboxylic acid and heat the mixture under reflux for 5 minutes. The residue obtained after evaporation of the solvent can be purified by chromatography on silica gel using, as eluent, a mixture of dichloromethane and methanol (?5:1) or can be used directly without purification. The residue is taken up in 10 ml of trifluoroacetic acid and after 7 minutes it is diluted with 200 ml of diethyl ether. After silica gel chromatography of the filtered precipitate (eluent: dichloromethane/methanol/30% ammonia in the ratio of 15:4:1), fractions containing the desired product are obtained, which are concentrated, dissolved in 1N methanolic hydrochloric acid and precipitated with diethyl ether. A white amorphous powder is thus obtained. Rf = 0.6 (in a mixture of chloroform, methanol and 33% ammonia in the ratio of 1:1:1, lower phase). NMR spectrum: 1.32 (d, 3 = 7, 3H); 1.38 (s, 3H); 1.53 (s, 3H); 2.43 (dd, J = 13, J2 = 4.2, 1H); 2.76 (s, 3H); 2.95 (s, 3H); 5.13 (d, J = 3.8 IH); 5.87 (d, J = 3.7, IH). ;Example 3: 1-C-methyl1gentamicin C-| pentahydrochloride Obtained in a manner analogous to Example 1 or 2. ;NMR spectrum; 1.37 (s., 3H); 1.53 (s, 3H); 2.44 (dd, ;13, J2 = 4, .IH); 2.95 (.s, 3H); .5.13 (_d, O = 3.6, IH); ;5.84, (d, J = 3.5, IH). ;Example 4; 1-C-methyl-6-N-methyl-1-genfra?riiciW C-j ?, pentachlorohydrate [method"b)]; 220 mg of 1-C-methyl-6'-N-benzil-oxycarbonylgentamicin C are dissolved in 50 ml of anhydrous tetrahydrofuran, 500 mg of lithium aluminum hydride are added and the mixture is heated for 3 hours under reflux. 10 ml of methanol is then carefully added and the mixture is acidified with 1 ml of acetic acid and filtered through celite. The organic phase is evaporated and chromatographed on silica gel using, as eluent, a mixture of chloroform, methanol and 30% ammonia in the ratio of 15:4:1. The fractions having an Rf value of 0.21 are collected. in a mixture of chloroform, methanol and ammonia at 30% (70:21:9), dissolve in 4 ml of 1N hydrochloric acid and precipitate by adding 100 ml of diethyl ether. A white powder is thus obtained. ;NMR spectrum: 1.36 (s, 3H); 1.52 (s, 3H); 2.45 (dd, J] = 12.5 Hz, J2 = 4 Hz, 1H); 2.76 (s, 3H); 2.94 (s, 3H); 5.12 (d, J = 3.6 Hz, 1H); 5.85 (d, J = 3.6 Hz, 1H). ;Necessary starting products can be prepared as follows. ;A) 3,21,61,3"-tetra-N-tert,-butoxycarbonyl-1-desamino-1-nitro-1-C-methanesulfonyloxymethylgentamicin ;a} 3,2?,61,3 ?-tetra-N-tert,-butoxycarbonyl-1-desamino-1-nitrogentamicin ;Heated to 75? A mixture of 50 g of potassium permanganate and 125 g of potassium dihydrogenphosphate in 0.5 litres of water is prepared, and 25 g of 3,21,61,3"-tetra-N-tert.-butoxycarbonylgentamicin C? in solution in 0.5 litres of acetone are added at once. The mixture is heated for 3 minutes under reflux and poured into ice-cold water containing 150 g of sodium hydrogen sulphite. The manganese dioxide is filtered off and the solution is extracted with chloroform. After drying, the organic phase is concentrated in a rotary evaporator and the residue is chromatographed on silica gel using, as eluent, a mixture of ethyl acetate and hexane in the ratio of 3:2. A colourless foam is thus obtained. Rf = 0.17 (in a mixture of dichloromethane and methanol in the ratio 20:1). ;b) 3,2',6',3"-tetra-N-tert.-butoxycarbonyl-1-desamino-1-nitro-1-C-hydroxymethylgentamicin C, ;Dissolve 0.64 g of 3,21,61,3"-tetra-N-tert.-tMrtoxicocarbonyl-1-desamino-1-nitrogentamicin C? in 5 ml of methanol and, after cooling to -7?, react with 0.3 ml of triethylamine. After 5 minutes, the solution is poured into 10 ml of a 37? solution of formaldehyde and brought to room temperature. After dilution with water and acidification with 0.1 N hydrochloric acid, the solution is extracted with dichloromethane and the organic phase is concentrated in a rotary evaporator. The substance having an Rf = 0.56 (in a mixture of dichloromethane and methanol in the ratio 10:1) is isolated by chromatography on silica gel using, as eluent, a mixture of dichloromethane and methanol in the ratio 100:2-4). ;c) 3,2',6*,311-tetra;-N-tert.-butoxycarbonyl-1-desamino-1-nitro-1-C-methanesulfonyloxymethyl-1gentamicin Cg

Dissolve 1.82 g of 3,2',6',311-tetra-N-tert.-butoxycarbonyl-1-desamino-1-nitro-1-C-hydroxymethylgentamicin and 0.6 ml of diisopropylethylamine in 60 ml of dichloromethane, add 0.16 ml of methanesulfonyl chloride, and heat the mixture under reflux for 1 hour. The residue obtained after evaporation is chromatographed on silica gel with chloroform as the eluent. An amorphous foam is obtained.

Rf = 0.43 (in a mixture of dichloromethane, methanol and 33% ammonia in the ratio of 9:1:0.2).

B)_^1-C-hydroxymethy1-3,21,61,3"-tetra-N-tert.-butoxycarboni1geritamycin C-j

a) 3,2',61,311 tetra-N-1erz,-butoxycarboni1-1~desamino-1-nitrogentami'Cin'a'C^

To a solution of 17 g of 3f2'?,6'',3"-tetra"N' tert.-butoxycarbonyl 1gentamicin C-| and 3.4' g of 3-tert.-butyl 1-4-hydroxy-5-methylenephenyl sulfide in 750 ml of 1,2-dichloroethane, 34 g of 3-chloroperbenzoic acid in solid form are added, while boiling. After 30 minutes, the solution is concentrated in a rotary evaporator and the residue is stirred with 20% KHSO4, saturated NaHCO2 and water and chromatographed on silica gel using, as eluent, a mixture of ethyl acetate and hexane in the ratio 3:2. S? isolates the substance having an Rf value = 0.74 (in a mixture of dichloromethane and methanol in the ratio of 9:1). b) 1-C-hydroxymethyl-3,2',6',3"-tetra-N-tert.-butoxycarbonyl 1gentamicin C-|

Dissolve 0.8 g of 3,2',6',3"-tetra-N-tert.-butoxycarbonyl-1-desamino-1-nitrogentamicin in 20 ml of chloroform, react with 10 g of paraformaldehyde and 1 ml of triethylamine and heat for 30 minutes under reflux. The resulting mixture is filtered by suction and the mother liquor is chromatographed on silica gel using, as eluent, a mixture of ethyl acetate and hexane in the ratio 2:1. Rf = 0.62 (in a mixture of dichloromethane and methanol in the ratio 9:1). The resulting product is reduced with hydrogen under 4 atmospheres using a suspension of Raney nickel in 30 ml of methanol. After removing the catalyst by filtration, an amorphous foam is obtained which is used as is in the next reaction.

C) 1-C-hydroxymeti1-3,21,61,.?'11-tetra-W-te'rz.-butoxycarboni1gentamicikna C^

? compound is obtained in a similar way to example B)b) passing through 3,2',6',3"-tetra-N-tert.? butoxycarbonyl1-1-desammino-1-nitrogentamicin C-ja [Rf = 0.16 in a mixture of dichloromethane and methanol in the ratio 20:1]?

Rf value = 0.5 (in a mixture of dichloromethane and methanol in the ratio 9:1) before hydrogenation with Raney nickel.

D) 1-C-methyl-G'-N-benzyloxycarbonylgentamicin C-|

740 mg of 1-C-methylgentamienin C-ja is dissolved in 30 ml of methanol and a solution of 500 mg of N-benzyloxycarbonyloxy 5-norbornene-2,3-dicarboxymide in 5 ml of methanol and 5 ml of chloroform is added dropwise at -15°C. After one hour, the mixture is raised to room temperature. After another two hours, the mixture is concentrated in a rotary evaporator and the residue is chromatographed on silica gel using, as the eluent, a mixture of chloroform, methanol and 30% ammonia in the ratio of 15:4:1. A product with an Rf value of 0.66 is obtained (in a mixture of chloroform, methanol and 30% ammonia in the ratio of 70:21:9).

E) 1-N,1-C-methy1engentamicin C-j

Dissolve 0.5 g of 1-C-hydroxymethyl-3,2?,6''3"-tetra-N-tert.-butoxycarbonyl gentamicin and 0.269 g of triphenylphosphane in 10 ml of anhydrous chloroform, add 0.2 g of di-isopropyl ester of the acid

azodicarboxylic acid and heat everything for 5 minutes

under reflux. After evaporation of the solvent, we obtain

a residue that is chromatographed on silica gel (eluent: dichloromethane/methanol in a ratio of 15:1).

thus obtaining a product with an Rf value = 0.49 (in

a mixture of dichloromethane and methanol in the ratio of 9:1). Dissolve 0.2 g of 1-N,1-C-methylene-3,2',6',3"-tetra-N-tert.-butoxycarbonyl gentamicin C-|a in 5 ml of trifluoroacetic acid and after 5 minutes dilute with

100 ml of diethyl ether. The residue is chromatographed.

on silica gel using, as eluent, a mixture

of dichloromethane, methanol and ammonia at 30% in the ratio of 15:4:1), which gives an amorphous powder.

Rf = 0.3 (in a mixture of chloroform, methanol and

of 33% ammonia in the ratio of 1:1:1, lower phase,).

NMR spectrum: 1.2 (s, 3H); 1.37 (dd, j = 13.8, 02 = 4.5, IH); 1.56 (s, IH); 2.01 (s, IH); 2.51 (s, 3H); 4.91 (d, 0 = 3.6,IH); 5.23 (d, J = 3.6, IH).

F) 1-N,1-C-methyTengentamicin C-j

Obtained in a similar way to example E) passing through 11 1-N,1-C-methylen-3,2',6?,311~tetra-N~tert.-butoxycarbonyl 1geritamicha; Rf = 0.47 (in a mixture of dichloromethane and methanol 9:1).

Title compound:

Rf = 0.36 (in a mixture of chloroform, methanol, and 33% ammonia in a ratio of 1:1:1, lower phase). NMR spectrum: 1.25 (s, 3H); 1.27 (d, J = 7, 3H); 1.39 (dd, J1 = 13.5, J2 = 4.6, 1H); 1.59 (s, 1H); 2.01 (s, 1H); 2.67 (s, 3H); 2.69 (s, 3H); 4.96 (d, J = 3.6, 1H); 5.33 (d, J = 3.6, 1H).

Claims (9)

Claims 1) Compounds of formula I 6'CHRc 3 2\ | 2 (I) R, 1 H 1 in which R-j represents hydrogen or a group of formula 6"CH20H ^0 NH TilHCH NHCH HCH H C (II) (Ila) (H b) Ile R^ represents a hydroxy or amino group, R^ and R^ each represent hydrogen or a hydroxy group, R,- represents an amino, methylamino, dimethylamino group or, when representing an amino group, equally a hydroxy group, and Rc represents hydrogen or a methyl group, b in the form of a free base or in the form of an acid addition salt. 2) L1 1-Ometi 1gentamicin C^, in the form of free base or in the form of an acid addition salt. 3) The compounds according to claim 1 or 2, characterised in that they are in the form of pentahydrochloride. 4) A process for the preparation of the compounds of formula I specified in claim 1, characterized in that a) a compound of formula 111a or 111b is reduced 76 CKR, NO, (U la) H?X 1 0 (Illb) wherein X represents a group susceptible to elimination and R-| to Rg have the meanings indicated in claim 1, and the amino and/or hydroxy groups may be protected, and, if desired, the protecting groups of the protected groups still present in the compound thus obtained are eliminated, or b) to prepare a compound in which Rg represents a methylamino or dimethylamino group, a corresponding compound of formula I in which Rg represents an amino group is subjected to a mono- or dimethylation or a corresponding compound of formula I in which Rg represents a methylamino group is subjected to an inonomethylation, other amino groups possibly present being protected, and, if desired, the protecting groups of the protected amino groups in the compound thus obtained are eliminated, and the compounds of formula I thus obtained are recovered in the form of the free base or in the form of an acid addition salt. 5) The compounds specified in claim 1 or 2, in the form of the free base or in the form of a chemotherapeutically acceptable acid addition salt, for use as chemotherapeutic agents. 6) The compounds specified in claim 1 or 2, in the form of the free base or in the form of a chemotherapeutically acceptable acid addition salt, for use as antibiotics with antibacterial activity. 7) A medicament, characterised in that it contains, as an active ingredient, a compound specified in claim 1 or 2, in the form of a free base or in the form of a chemotherapeutically acceptable acid addition salt. 8) A pharmaceutical composition, characterized in that it contains a compound specified in claim 1 or 2, in the form of a free base or in the form of a chemotherapeutically acceptable acid addition salt, in association with chemotherapeutically acceptable diluents or vehicles. 9). The compounds of formula ttl.a and 111b 0 OR1 CHR (111b) OR wherein the symbols R-j to Rg have the meanings set forth in claim 1 and X represents a group susceptible to being eliminated.