DE2135191B2 - 3 ', 4'-DIDESOXYKANAMYCIN B, METHOD OF MANUFACTURING IT AND MEDICINAL PRODUCT CONTAINING THIS PRODUCT - Google Patents

Description

20th

HO

2. Process for the preparation of 3 ', 4'-dideoxykanamycin B according to claim 1, characterized in that the amino groups of kanamycin B are reacted with ethyl chloroformate or salicylaldehyde in a manner known per se, then the 4 "and 6" hydroxyl groups with acetone or cyclohexanone dimethyl ketal, then the 2 "hydroxyl group is benzoylated in the usual way, from the 4", 6 "-Di-O-isopropylidene or 3 ', 4';4", 6 "-Di- O-cyclohexylidene derivative is split off by heating with aqueous acetic acid acetone or cyclohexanone, the 3 'and 4' hydroxyl groups are converted into 3 ', 4'-disulfonic acid ester groups in the usual way with methanesulfonic acid chloride, these are converted into the 3', by reaction with sodium iodide and zinc 4'-didehydro compound transferred, this is catalytically hydrogenated in a manner known per se with hydrogen in the presence of PtO ₂ and finally the remaining protective groups in a manner known per se with dilute acetic acid and then connect d removed barium hydroxide.

3. Medicines !, characterized by a content of a compound according to claim 1 and customary carriers and / or diluents and / or auxiliaries.

4 "

(Kanamycin: R = OH)
(Kanamycin B: R = NH ₂ )

On the basis of these findings, an attempt was first made to block the 3 'hydroxyl group and to synthesize the 3'-O-methylkanamycin. However, this derivative did not have any antibacterial properties Activity. Then, 3'-deoxykanamycin was synthesized and it was found that this derivative is active against the resistant bacteria described above. Hence became assumed that the replacement of the 3 'permanent hy-

Hydroxyl group inhibits the phosphorylation of kanamycin by resistenle bacteria, giving this antibiotic the ability to attack kanamycin-resistant bacteria.
The highly active 3 ', 4'-dideoxykanainv-

synthesized a B of the following formula:

55

The invention relates to 3 ', 4'-Dideoxykanamy-B, which has antibacterial effects and insiondere against infections of kanamycinresisten-ι organisms is effective, a method for his Position and medicinal products containing it. It rests on investigations into the mechanism r elucidated drug inactivation by resistant bacteria from patients and : due to the mechanism to new antibiotic: effective against the resistant bacteria : derivatives led.

HO

OH

The process for the preparation of 3.4'-dideoxy-

lcanamycin B is characterized in that: _n a conventional manner, the amino groups of kanamycin B with ethyl chloroformate and reacting salicylaldehyde, then the 4 "- and 6" ständigeii hydroxyl groups with acetone or Cyclohexanondimethylketal reacted, to this, the 2 'position hydroxyl group benzoylated in the usual way, from the 4 ", 6" -Di-O-isopropy-Hden- or 3 ', 4'; 4 ", 6" -Di-O-cyclohexylidene derivative obtained by heating with aqueous acetic acid, acetone or cyclohexanone split off, the 3 'and 4' hydroxyl groups are converted in the usual way with methanesulfonic acid chloride into 3 \ 4'-disulfonic acid ester groups, these are converted into the 3 \ 4'-didehydro compound in a manner known per se by reaction with sodium iodide and zinc Hydrogen catalytically hydrogenated in the presence of PtO ₂ and finally the remaining protective groups are removed in a manner known per se with dilute acetic acid and then barium hydroxide.

The synthetic route for the new 3 ', 4'-dideoxykanamycin B is briefly as follows:

Kanamycin B (free base) is used as the starting material. It is treated with the usual means

delt to the five free amino groups that are 4 ", 6" Hydroxyl groups and all or part of the 5- and 2 "positions and the 3 'and 4' positions Protect hydroxyl groups of the starting kanamycin B. It can e.g. B. the following kanamycin B derivatives are produced:

CH ₂ -NHCOOC ₂ H ₅ NHCOOC ₂ H ₅

H, C

OCO

HO

HO

CH ₂ N = CH

⁵ - ·· η · r F «isic acid treated with the 3 'and 4' positions

The compound of type I or Γ is then tssigbou ivate with _wabnger the following general

Hydroxyl groups are released. These products are then made into Denvai Forme! II transferred:

CH ₂ NHCOOC ₂ H ₅ (or N =

NHCOOC ₂ H ₅ (or N =

NHCOOC ₂ H ₅ (or N = OH

(O

NHCOOC ₂ H ₅ (or N =

O-CH

H, C

(II)

OCcT ^ OC ₂ H ₅ (or N = CH- ^ J

wherein the 3 'and 4' hydroxyl groups have been sulfonylated, ie W means a —SO ₂ CH, group.

If this disulfonylated derivative (II)

(W = SO ₂ CHj)

treated with sodium iodide and zinc dust in a suitable solvent, this is obtained 3 ', 4'-unsaturated derivative, which after reduction with hydrogen in a manner known per se in the presence of platinum oxide as a corresponding hydrogenation product, namely as one protected by amino and hydroxyl groups 3 ', 4' - dideoxykanamycin B is present. When using this derivative according to conventional measurement methods the Schute groups are split off, namely with dilute acetic acid and then Barium hydroxide, you get 3 ', 4'-dideoxykanamycin B.

The synthetically produced 3 ', 4'-dideoxykanamycin B inhibits the growth of different types of bacterial strains in the same way as

Kanamycin B, as can be seen from the table. It also inhibits the growth of Kanamycin B-resistant Staphylococci, the resistant Escherichia coli and the resistant Pseudomonas aeruginosa. 3'.4'-dideoxykanamycin B shows a therapeutic process

against infections caused by staphylococci. Klebsieila pneumoniae, Salmonella typhosa and Pseudomonas aeruginosa can be produced in mice. 3 ', 4'-Dideoxykanamycin B has low toxicity (LD ₅₀ 180 mg / kg mouse, Lv.) And shows a

high blood levels after the injection. It follows that the product according to the invention is a valuable one Relation to the treatment of different types of infections including the infections that through gram-negative and gram-positive resistant bacteria

series are generated is. From the table above it can be seen that the prepared according to the invention Product shows a strong effect in inhibiting pathogenic microorganisms.

Tabel

Antibacterial spectra of 3 ', 4'-dideoxykanamycin B and kanamycin B

Test organism ¹ ) ² ) Minimum inhibitory concentration Sample 2 (mcg / ml) 6 "-deoxy-
kanamycin 3,4'-dideoxykanamycin B 0.78 Kanamycin B Sample 1 1.56 1.56 Staphylococcus aureus FDA 209 P 1.25 1.56 0.78 1.56 Escherichia coli NIHJ 2.5 1.56 1.56 1.56 K-12CS-2 2.5 0.78 > 100 K-12 ML I629 ⁺ ■ 2.5 > 5 ()

1 35 191

irtselzung

; st organism ') ² l

Minsthcmmkon / cntralion (mcg mil

, 4-dideoxykanamycin B kanamycin B (»" -deoxy-

kanamycin

Sample!

Sample;

K-12 ML 163O ⁺
K-12 ML 1410
seudomonas aeruginosa A3
No. 11 ^{+ +}
No. 12 ^{+ +}
No. 39 ⁺⁺
No. 45 ^{+ +}
No. 67 ^{+ +}

'rotus rettgeri
GN 311 ^{+ +}
GN 466 ^{+ +}

3.12 > 50 > 1 () 0 1.56 0.78 1.56 3.12 50 100 3.12 > 50 > l () 0 6.25 50 100 6.25 > 50 > 100 0.78 25th KX) 6.25 > 50 > 100 12.5 3.12 3.12 3.12 1.56 3.12

') Agar nutrient medium. 37 C, 18 hours.

² ) The ^{strains marked with +} were artificially inoculated with the resistance factor (R-factor | in Escherichia coli K-12

obtained in vitro: the strains marked with * * are isolated from patients and resistant to many known antibiolics

Tribes; the rest are natural stems.

In the following examples, preferred methods of preparation of 3'.4'-dideoxykanamine B described in detail.

example 1

a) Penta-N-ethoxycarbonylkanamycin B

To a stirred mixture of 1.0 g of Kanamycin B (free base) and 0.9 g of anhydrous sodium carbonate in aqueous acetone (1: 1.20 ml) was added 1.05 ml of ethyl chloroformate and stirred for 5 hours at room temperature. The resulting precipitate was filtered off, washed with water and dried. 1.46 g of a solid substance were obtained. The melting point was 305 ° C. (decomposition), [n] ' _D ⁵ : + 88 ° (c = 0.5: dimethylformamide).

b) Penta-N-ethoxycarbonyl-3 ', 4'; 4 ".6" -d ^; -O-isopropylidene- canamycin B

13.7 g of the penta-N-ethoxycarbonylkanainycin B prepared according to Example la) were dissolved in 70 ml of dimethylformamide, and the solution was after addition of 18.8 g of 2,2-dimethoxypropane and 0.35 g of anhydrous p-toluenesulfonic acid for 1 hour heated to 65 ° C. The solution prepared in this way was concentrated to approximately 50 ml and, after the addition of 30 g of 2,2-dimethoxypropane, was again heated to 65 ° C. for 1 hour. Then 6 ml of triethylamine were added and the solution was poured into a stirred mixture of 500 ml of benzene and 500 ml of water poured in. A solid precipitate formed (solid A). After the solid A was filtered off, the benzene layer was allowed to stand. The resulting precipitates were filtered with benzene and washed, washed and dried. 3.5 g of a solid with a melting point of 236 to 237 ° C. [n] f: + 87 ° (c = 1; dimethylformamide) were obtained. The solid A obtained (8.1 g) turned out to be penta-N-ethoxycarbonyl-4 ", 6" -O-isopropylidenjo kanamycin B. It could be used in the same. In the same way as described above, the penta-N-ethoxycarbonyl-3 ', 4'; 4 ", 6" - di - O - isopropylidene-kanamycin B can be converted.

_1S c) 2 "-O-Benzoyl-penta-N-ethoxycarbonyl-

3 ', 4'; 4 ", 6" -di-O-isopropylidene-kanamycin B

3.24 g of the kanarnycin B derivative prepared according to Example I b) was dissolved in 48 ml of pyridine, and to the solution was added 2 g of benzoyl chloride.

The solution was allowed to stand at room temperature for 1 hour, the solution was evaporated and the residue was dissolved in chloroform. This solution was washed with water and dried over sodium sulfate. The solution was then concentrated to approximately 20 ml and after addition of n-hexane gave 3.35 g of a crystalline substance having a melting point of 205-209 ⁰ C. [n] S: + 114 ° (c = 1, dimethylformamide) .

d) 2 "-O-Benzoyl-penta-N-ethoxycarbonyl-

4 ", 6" -O-isopropylidene-kanamycin B

2.82 g of the kanamycin B derivative prepared according to Example 1 c) were dissolved in aqueous acetic acid (1: 3.40 ml) and the solution was heated to 95 ° C. for 30 minutes. The solution thus prepared was evaporated and the one obtained deacetonated product (2.58 g) dissolved in 15 ml of dimethylformamide. After adding 0.65 g of 2,2-dimethoxypropane and 55 mg of anhydrous p-toluenesulfonic acid, the solution was left to stand for 1 hour at room temperature. The solution was then poured into water and the resulting precipitates were filtered off, washed with water and treated with benzene. 2.01 g of a benzene-insoluble solid with a melting point of 285-287 ° C, [α] ?: + 105 ° (c = 1: dimethylformamide): NMTt spectrum (in dimethyl sulfoxide af,): τ 8. ^ 2 and 8.60 (each 3 proton singlet is isopropylidene pri>

tones), τ 1.5-2.8 (5-proton multiplet is assigned to benzoyl protons).

e) 2 "-O-Benzoyl-penta-N-ethoxycarbonyl-4" .6 "-O-isopropylidene-3 ', 4'-di-O-mesylkanamycin B.

1.18 g of the kanamycin B derivative prepared according to Example 1 d) were dissolved in 15 ml of pyridine, and the solution, after addition of 0.66 g of methanesulfonyl chloride (CH ₃ SO ₂ Cl), was left to stand for 1.5 hours at room temperature. The solution was evaporated to one third of the original volume and the solution thus prepared was poured into water. The resulting precipitates were filtered off and washed with water. 1.32 g of a solid with a melting point of 198 ° C. were obtained, [α]! ': + 107 ° (c = 1.5; dimethylformamide); NMR spectrum (in pyridine d ₅ ): τ 6.60 and 6.45 (each 3-proton singlet can be assigned to mesyl protons).

f) 2 "-O-benzoyl-3 ', 4'-dideoxy-3', 4'-didehydro-

penta-N-ethoxycarbonyI-4 ", 6" -0-isopropylidene-

kanamyein B

11 g of sodium iodide and 5 g of zinc dust were added to a solution consisting of 1.0 g of the kanamycin B derivative prepared according to Example 1 e) and 20 ml of dimethylformamide, and the mixture was heated to 95 ° C. for 1 hour with vigorous stirring. The mixture was then poured into a hot mixture of 100 ml of chloroform and 100 ml of water with stirring, and the isolated chloroform layer was washed with water, dried over sodium sulfate. After evaporation, 940 mg of a solid were obtained. The solid was treated with hot ethyl acetate and 310 mg of the insoluble solid was recrystallized from a mixture of methanol-chloroform-ethyl acetate. A substance was obtained with a melting point of 282-284 ⁰ C, [α] ?: + 36 ° (c = 0.4, dimethylformamide); NMR spectrum (in pyridine-d ₅ ): τ 4.06 (2 protons in a somewhat broadened singlet, to be assigned to the hydrogen in the 3 'and 4' positions).

g) 2 "-O-benzoyl-3 ', 4'-dideoxy-penta-

N-athoxycarbonyl ^ .o ^ O-isopropylidene-

kanamyein B

1.15 g of the Kanamycin B derivative prepared according to Example 10, ie the 2 "-O-benzoyl-3'.4'-dideoxy - 3 ', 4' - didehydro - pen ta - N - ethoxycarbonyl-4" , 6 "-O-isopropylidene-kanamycins B, which had been dissolved in a mixture of 45 ml of p-dioxane, 25 ml of methanol and 25 ml of water, was treated with hydrogen at 2 at pressure and in the presence of 0.3 g of platinum oxide, which had previously been activated, hydrogenated at room temperature for 5 hours. The solution thus prepared was filtered and evaporated. 1.10 g of a colorless powder with a melting point of 254-256 "C, [n] ?: +87" ( c = 0.2; dimethylformamide); NMR spectrum (in pyridine-d ₅ ): τ 7.8-8.5 (4 protons, broad signal, to be assigned to the hydrogen in the 3 ', 4 -position).

h) 2 "-O-benzoyl-3 ', 4'-dideoxy-penta-N-ethoxycarbonyl-kanamycin B.

980 mg of the kanamycin B derivative prepared according to Example 1 g) were dissolved in a mixture of 8 ml of acetic acid and 5 ml of water. The solution was heated for 5 minutes at 100 ⁰ C. The resulting precipitates were filtered and getrock net, to obtain 750 mg of a substance mi a melting point of 247-248 ° C, [- /] ?: + 89 (c = 2, 7; dimethylformamide). Found C: 52.4! H: 6.54; N: 7.48%. For C ₄₀ H ,,, N, O _1n C: 52.45; H: 6.71; N: 7.65%.

i) 3 ', 4'-Dideoxykanamycin B

150mg of the Kana produced according to example lh)

ίο mycin-B derivative and 2.3 g of barium hydroxide octahydrate were added to a mixture of 5 ml of water and 4 liters of p-dioxane. The mixture thus prepared was heated hanging on a boiling water bath for 8 hours. Carbon dioxide was then passed in and the resulting precipitates were centrifuged off. The supernatant layer was isolated and evaporated. The residue obtained in this way was dissolved in water and, after filtration, the solution was transferred to a column loaded with "Amberlite IRC-50" (H 'form) with 0.1-0.4N ammonium cyclrox.vd chromatographies '-Dideoxykanamycin B was concentrated. By adding acetone, 47 mc of a crystalline substance was obtained. [, <] ": +130 (c = 0.65: water) Found: C: 47.71; H: 8.38: N: 15.31% Calculated for C ₈ H ₃₇ N ₅ O ₈ : C: 47.88; H: 8.26; N: 15.51%.

Example 2 ^a > Penta-N-salicylidene-kanamycin B

To a suspension of 400 mg of Kanamycin B base in aqueous methanol (1: 8? 20 ml) were 520 mg of salicylaldehyde added, and the resulting Solution was poured into water. The precipitates were nitrated, dried and craved 750 mg penta-N-salicylidene-canamycin B.

b) 3'.4 '; 4 ", 6" -Di-O-cyclohexylidene-pcnta-N-salicyüdenkanamycin B 700 mg of the pcnta-N-sahcyhdenkanarr.ycins prepared according to Example 2a) B was dissolved in 12 ml of dry dimethylformamide and the solution was after adding 500 mg of cyclohexanone dimethyl alcohol and 30 mg of anhydrous p-toluenesulfonic acid for 1 hour

heated ^{to 50 c} C for a long time. Then a clot of the solvent was removed by evaporation in vacuo. The solution thus prepared was poured into water. The resulting nodules were filtered, dried and scraped

710 mg.

O 2 "-O-Benzoyl-3'.4 '; 4", 6 "-di-O-cyclohexylidene-penta-N-salicylidene-kanamycin B.

590 mg of the kanamycin B denvate prepared according to Example 2 b) were dissolved in 10 mf pyridine. and to the solution was added 120 mg of benzoyichloride added. The mixture was allowed to stand for 1 hour at room temperature, then the solution was in Poured water. The resulting precipitates were filtered off, dried, and were obtained a yield of 612 mg.

d) 2 "-O-Benzoyl-4", 6 "-O-cyc! ohexy1iden-penta-N-athoxycarbonylkanamycin B.

A mixture of 600 mg of the according to Example 2 el produced kanamycin B derivative and wäiiriccr Acetic acid (1: 3, 20 ml) was on for 30 minutes V5 t_ heated. The reaction mixture was

evaporated and the residue treated with ether. The ether-insoluble part was dissolved in aqueous acetone (1: 1, 15 ml). After addition of 400 mg of sodium carbonate, 270 mg of ethoxy carbonyl chloride were added to the solution added. It was then stirred for an additional 5 hours at room temperature. The resulting

Any precipitates were filtered off, washed with water and dried. You got 51! a solid substance, [u] i ': +98 (c = 1.5; prostitute formamide). This substance could be converted into 3'.4'-dideoxykanamycin B in a manner similar to that described in Examples 1 e) to 1 i)

Claims (1)

Patent claims: 1. 3 ', 4'-Dideoxykanamycin B of the formula
H ₂ NH ₂ C NH, It was found that drug-resistant Gram-negative bacteria from patients who carry the R-factor, resistant staphylococci and resistant Pseudomonas aeruginosa produce an enzyme, the kanamycin is inactivated by the phosphate residue of adenosine triphosphate on the 3 'hydroxyl group of the kanamycin is transmitted. CH ₂ NH ₂