FI57596B - FOERFARANDE FOR FRAMSTAELLNING AV 3 ', 4'-DIDEOXIKANAMYCIN B SOM AER ANTIBACTERIAL OR SPECIFICALLY ACTIVE MOT INFECTOR ORGANISM RESISTANT ORGANISM - Google Patents

Description

Ι, ΓΊΓ "! R_, ANNOUNCEMENT rnr λ / 4 & A ^ (11) UTLÄGGNINGSSKRIFT 57596 q45) ·; - n: · --- o ^ ^ (51) Kv.lk.3 / lirt.CI.3 C 07 H 15 / 22 SUOMI — FINLAND pi) ρμι # «ιηιιμι« ι »-ριμπ» λ ^ ι 17 ^ / 71 (22) HtktmlspUvf - Araeknlngtd ·· l8.06.71 ^^ (23) Alkupllvi — GIMfh «tad« f l8.06.71 (41) Tullut JulklMluI - Bllvlt offamllg 30.01.72

And the Board of Registration (44) NlhtMkilpwon j. month] outside »l« * n date.-

PktMlt · och rtgiltantyrilNn AimMcm utlagd och utl. * KrHton publlcond 30.05. oO

(32) (33) (31) P) ry4 «* y« uoHteui — Begird priorities 29.07.70 ll.05.7i Japan-Japan (JP) 65760/70, 30871/71 (71) Zaidan Hojin Biseibutsu Kagaku Kenkyu Kai, Well. 14-23, 3-chome, Kami-Osaki, ”” Shingawa-ku, Tokyo, Japan-Japan (JP) (72) Sumio Umezava, Tokyo, Hamao Umezawa, Tokyo, Tsutomu Tsuchiya, Yokohama-shi, Kanagawa-ken, Japan-Japan (JP) (74) Forssen & Salomaa Oy (54) Method for the preparation of 3 ', 4'-dideoxycanamycin B which is effective against infections caused by bacteria and in particular kanamycin-resistant organisms - Förfarande för framställning av 3', U'-dideoxikanamycin B som This invention relates to a process for the preparation of 3 ', 4'-dideoxycanamycin B, which agent is antibacterial and in particular acts on infections caused by kanamycin-resistant organisms.

The said compound is represented by formula 6 '

w H2NH2C

ίχ.

H0H2C I J

HO3> 1 | ___ k "

OH

2 57596

The preparation method is as follows: the starting material, kanamycin B, is converted into a kanamycin B derivative of general formula 1 or I 'in which five amino groups, 4 ", 6M-hydroxyl groups and 3', 4'- and 2" -hydroxyl groups are protected.

CH.NHCOR

A K ° / NHCOR

°% ~ ί \ n H NHCOR ϋ / M, \ OH Γ (I> OH2C / <JÖK * oz ch2n-chr 'ΧΙ3 · *, on \ n-chr' N-CHR * υ ^ / \ / (I ' ) oh2c / (t \ / '\ | \ N-CHR' / K "o Y_j £« oz R ethoxy R '* O-hydroxyphenyl X group CHg ch3 ^ C ^

or a group (CHjJj C

3 57596 Υ means group

CV

or group C

The 3 'and 4'-hydroxyl groups of Z * benzoyl are liberated by cleavage of a ketone from a compound of formula I or 1', followed by conversion of the compound thus obtained with methanesulphonic acid chloride to a compound of formula II: CH2NHCOR (or N-CHR ') jfs (ow λ NHCOR (or N-CHR ') 3S-1 n / \ NHCOR (or N-CHR') NHCOR U \ 1 / \ (or N-CHR ') ^ \ j \ oZ * λ OH2C / (II) 1 oz

NHCOR

w (N-CHR ') wherein W - methylsulfonyl and R, R', Y and Z have the same meaning as in formulas I or I '.

Derivative II (H - SO 2 CH 3) is then treated by the method described by T-itson et al. (Carbohydrate Research, Vol. 1, pp. 338-340, 1965), te. with iodide ion and a reducing metal powder, e.g. zinc dust, in a suitable solvent to form a derivative having a double bond between the 3 'and 4' carbon atoms. After hydrogenation of the unsaturated group, the protecting groups are deprotected in the usual manner to give the desired final product, 3 ', 4'-dideoxycanamycin B.

Hamao Umezawa et al. (Science 157, (1967), p. 1559) found that 57596 patients

A

drug-resistant Gram-negative bacteria, resistant staphylococci, and resistant Pseudomonas aeruginosa comprising factor R derived from kanamycin produce enzymes that inactivate kanamycin by transferring phosphate from the ATP to the 3′-hydroxyl group in kanamycin. Kanamycin has the structure of formula III.

6 'ch2nr2 jv H0 \? - 12 ^ NH2 ~ R ° \ a (C λ1 (III) 6 »

HOH 2 C

ie Η ° λΤτ ~ ^ | 2 "

OH

kanamycin A: R - OH kanamycin B: R - NH2 Based on these findings, the inventors initially attempted to protect the 3'-hydroxyl group by synthesizing 3 * -O-methylanamycin. However, the compound had no antibacterial effect. In the next step, 3'-deoxycanamycin was synthesized and found to be effective against the above-mentioned resistant bacteria. Therefore, it was considered possible that replacing the hydroxyl group phosphorylated by resistant bacteria with hydrogen would give the antibiotic the property of acting on resistant bacteria. The inventors applied the principle to kanamycin B and prepared a highly active compound, 3 ', Α'-dideoxycanamycin B.

The synthesized compound 3 ', 4'-dideoxycanamycin B inhibits the growth of several types of bacterial strains in the same manner as kanamycin B, as shown in Table I, and further inhibits the growth of kanamycin and kanamycin B resistant staphylococci, Escherichia coli and Pseudomonas aeruginosa. 3 ', Α'-dideoxycanamycin B shows a therapeutic effect against staphylococci,

Infections with Klebsiella pneumoniae, Salmonella typhosa and Pseudomonas aeruginosa in mice. 3 ', 4'-dideoxycanamycin B has low toxicity (LDjq 180 mg / kg mouse in vivo) and shows high post-injection blood levels, thus indicating that the derivative is a valuable chemotherapeutic agent in the treatment of several types of infections caused by both gram-negative and gram-positive resistant bacteria.

As can be seen from the following table, it has a strong inhibitory effect on microorganisms, including pathogenic microorganisms.

- TABLE 1 Antibacterial spectra of 3 ', 4'-dideoxycanamycin B and kanamycin B "" --—-; -

Experiment Minimum contraceptive concentration ^ ug / ml 3 ', 4'-dideoxycanamycin B Kanamycin B Sample 1 Sample 2

Staphylococcus aureaus FDA 209 P 1.25 0.78 0.78

Escherichia coli NIHJ 2.5 1.56 1.56 K-12 CS-2 2.5 1.56 0.78 K-12 ML 1629 2.5 1.56 50 K-12 ML 1630 2.5 3.12 50 K-12 ML 1410 1.56 0.78

Pseudomonas aeruginosa A3 2.5 3.12 50

Well. 11 3.12 50 ^ No. 12 6.25 50

Well. 39 6.25 50

Well. 45 1.25 0.78 25 ~ No. 67 6.25 50

Proteus rettgeri GN 311 12.5 3.12 GN 466 2.5 3.12 1.56 x ^ Medium agar, 37 ° C, 18 hours

The invention is explained below by giving examples of the preparation of 3 *, 4'-dideoxycanamycin B.

6 57596

Example 1

a) Penta-N-ethoxycarbonylcanamycin B

To a mixture of kanamycin B free base (1.0 g) and anhydrous sodium carbonate (0.9 g) in aqueous acetone (1: 1, 20 mL) was added ethoxycarbonyl chloride (1.05 mL) followed by stirring was continued for 5 h at room temperature. The resulting precipitate was filtered, washed with water and dried to give a substance with a melting point of 305 ° C (decomposed), m + 88 ° (c = 0.5; dimethylformamide).

b) Penta-N-ethoxycarbonyl-3 ', 4'; 4 ", 6" -di-O-isopropylidenecanamycin B (formula I; R OC 2, X - Y - CH (CH 3) 2, Z = H)

Penta-N-ethoxycarbonylcannamycin B (13.7 g) prepared by the method described in a) was dissolved in dimethylformamide (70 ml), followed by the addition of 2,2-dimethoxypropane (18.8 g) and anhydrous p-toluenesulfonic acid. (0.35 g) and heated for one hour at 65 ° C. The resulting solution was concentrated to 50 ml, and after the addition of 2,2-dimethoxypropane (30 g), the solution was heated again at 65 ° C for 1 hour. Triethylamine (6 ml) was added and the solution was poured into a mixture of benzene (500 ml) and water (500 ml), which was stirred to give a precipitate (substance A). After removal of substance A by filtration, the benzene layer was allowed to stand. The resulting precipitate was filtered, washed with benzene and water and dried to a solid (3.5 g), m.p. 236-237 ° C; mp <87 + 87 ° (c = 1; dimethylformamide).

The obtained substance A (8.1 g) turned out to be Penta-N-ethoxycarbonyl-4 ", 6" -O-isopropylidene-kanamycin B and the compound could be converted by the treatment described above into Penta-N-ethoxycarbonyl-3 *, 4 '; 4 ", 6" -di-O-isopropylidenecanamycin B.

c) 2'-O-benzoyl-Penta-N-ethoxycarbonyl-3 ', 4'; 4 ", 6" -di-O-isopropylidene-kanamycin B (formula I, R * OC 2 H 2, X = Y * C (CH 3) 2, Z * COCl 2 H 2)

The kanamycin B derivative prepared according to b) (3.24 g) was dissolved in pyridine (48 ml), followed by the addition of benzoyl chloride (2 g). After standing for one hour at room temperature, the solution was evaporated and the residue was dissolved in chloroform. The solution was washed with water, dried over sodium sulfate and concentrated to 20 mL. Addition of n-hexane formed 2 "-O-benzoyl-Penta-N-ethoxycarbonyl-3 ', 4'; 4", 6 "-di-O-isopropylidene-1-kanamycin B crystals (3.35 g) having melting point 205-209 ° C; / ° </ ρ * 7 57596 + 114 ° (c 1; dimethylformamide).

d) 2 "-O-Benzoylpenta-N-ethoxycarbonyl-4", 6 "-O-isopropylidenecanamycin B

The kanamycin B derivative (2.82 g) prepared by the method described in c) was dissolved in a mixture of water and acetic acid (1: 3.40 ml), and the solution was heated at 95 ° C for 30 minutes. The resulting solution was evaporated, and the resulting deacetonated product (2.58 g) was dissolved in dimethylformamide (15 ml), 2,2-dimethoxypropane (0.65 g) and anhydrous p-toluenesulfonic acid (55 mg) were added, and the solution was allowed to stand at room temperature. temperature for one hour. The solution was poured into water, the precipitate obtained was filtered, washed with water and treated with benzene to give a benzene-insoluble solid, 2 "-O-benzoylpenta-N-ethoxycarbonyl-4", 6 "-O-isopropylidenecanamycin B, 2.01 g mp 285-287 ° C, 21 ° / + 4 ° + 105 ° (c 1, dimethylformamide) NMR spectrum (dimethylsulfoxide-d 6): δ 8.72 and 8.60 (each 3 proton singlet due to isopropylidene protons), 1y 1.5-2.8 (5-proton multiplet due to benzoyl protons).

e) 2M-O-Benzoylpenta-N-ethoxycarbonyl-4 ", 6" -O-isopropylidene-3,4,4-di-O-mesylcanamycin B (in the general formula HR »0C2H3, Y * W - SO2CH3, Z = COCgH5 and Z '-H).

The kanamycin B derivative prepared by the method of d) (1.18 g) was dissolved in pyridine (15 ml), methylsulfonyl chloride (0-38Ο201, 0.66 g) was added to the solution, and it was allowed to stand at room temperature for 1.5 hours.

The solution was concentrated to 1/3 of its original volume and poured into water. The resulting precipitate was filtered and washed with water to give a solid (1.32 g), m.p. 198 ° C, + 107 ° (c - 1.5, dimethylformamide); NMR spectrum (pyridine-d 6): 7 * -6.60 and 6.45 (each 3-proton singlet is due to mesyl protons).

f) 2'-O-benzoyl-3 ', 4'-dideoxy-3', 4'-didehydro-Penta-N-ethoxycarbonyl-4 ', 6 "-O-isopropylidenecanamycin B.

To a solution of kanamycin B derivative (1.0 g) prepared by the method of d) in dimethylformamide (20 ml) were added sodium iodide (11 g) and zinc dust (5 g), and the mixture was heated at 95 ° C with vigorous stirring for 1 hour.

The mixture was poured into a hot mixture of chloroform (100 ml) and water (100 ml) with stirring, and the separated chloroform layer was washed with water, dried over sodium sulfate and evaporated to dryness to give a solid (940 mg). This was treated with hot ethyl acetate, and the solvent-insoluble solid (310 mg) was recrystallized from methanol-chloroform-ethyl acetate-22 °, m.p. 282-284 ° C, λmax +36 (c = 0.4, dimethylformainide); NMR spectrum (pyridine-d 6): "Ir 4.06 (2-proton somewhat broadened singlet, H-3 'and 4').

g) 2H-O-Benzoyl-3 ', 4'-dideoxypenta-N-ethoxycarbonyl-4 ", 6" -O-isopropylidene-kanamycin B.

A solution of kanamycin B derivative (1.15 g) prepared according to f) in a mixture of p-dioxane (45 ml), methanol (25 ml) and water (25 ml) was hydrogenated with hydrogen at 2 atm under platinum oxide (0.3 g). which was preactivated before use, in the presence of w at room temperature for 5 hours. The resulting solution was filtered and evaporated to dryness to give 1.10 g of a colorless powder, m.p. 254-256 ° C, mp + 87 ° (c * 0.2, dimethylformamide); NMR spectrum (pyridine-d 6): -

Ty 7.8-8.5 (4-proton broad singlet H-3 ', 4').

h) 2 "-O-Benzoyl-3 ', 4'-dideoxypenta-N-ethoxycarbonylcanamycin B

The kanamycin B derivative prepared by the method of g (980 mg) was dissolved in a mixture of acetic acid (8 ml) and water (5 ml), and the solution was heated at 100 ° C for 5 minutes. The resulting precipitate was filtered off and dried, 750 mg, m.p. 247-248 ° C, / + 89 ° (c = 2.7, dimethylformamide). Found: C, 52.41; H, 6.54; N, 7.48%. Calculated from the formula ^ o ^ 61 ^ 5 ^ 10: ^ * ^ 5; 6.71; N, 7.65%.

i) 3 ', 4'-dideoxycanamycin B

Kanamycin B derivative (150 mg) and barium hydroxide octahydrate (2.3 g) prepared by the method described in h) were added to a mixture of water (5 ml) and p-dioxane (4 ml), and the resulting mixture was heated in a boiling water bath for 8 hours. Carbon dioxide was introduced into the mixture, and the resulting precipitate was removed by centrifugation. The supernatant was removed and evaporated to dryness. The resulting residue was dissolved in water, and after filtration, the solution was chromatographed on an "Amberlite IRC-50" column (H + form) with 0.1-0.4 ammonium hydroxide. The fraction containing 3 ', 4'-dideoxy-kanamycin B was concentrated. Addition of acetone gave 47 mg of crystals, m.p. + 130 DEG (c 0.65, water). Found: C, 47.71; H 8.38; N, 15.31%. Calculated for C 18 H 18 N 2 O 3 O 2 • 47.88; H, 826; N, 15.51%.

Example 2 a) Synthesis of Penta-N-salicylidenecamamycin B.

9 57596

To a suspension of kanamycin B base (400 mg) in aqueous methanol (1: 8, 20 mL) was added salicylaldehyde (520 mg), and the resulting solution was poured into water. The precipitate was filtered off and dried, yielding 750 mg.

b) 3 ', 4'; 4 ", 6" -di-O-cyclohexylidene penta-N-salicylidenecanamycin B (in formula 1 'R - CgH2OH), X = Y = (CH2C, Z - H)

Penta-N-salicylidenecanamycin B (700 mg) prepared by the method of a) was dissolved in dry dimethylformamide (12 ml), and a solution of cyclohexanone dimethyl ketal (500 mg) and anhydrous p-toluenesulfonic acid (30 mg) first was heated for 1 hour. At 50 ° C, after which a small portion of the solvent was removed by evaporation in vacuo. The resulting solution was poured into water. The precipitate formed was filtered off and dried, yielding 710 mg.

c) 2 "-O-benzoyl-3 ', 4'; 4", 6 "-di-O-cyclohexylidene penta-N-salicylidene-kanamycin B (in formula 1 'R' C 1 H 2 H 2 (OH), XY = ( CI (CH 2 Cl 2, CH 2 Cl 2).

The kanamycin B derivative obtained by the method of b) (590 mg) was dissolved in pyridine (10 ml), and benzoyl chloride (120 mg) was added to the solution. After standing at room temperature for one hour, the solution was poured into water. The resulting precipitate was filtered and dried to give 612 mg.

d) 2'-O-benzoyl-4 ", 6" -O-cyclohexylidene penta-N-ethoxycarbonylanamycin B.

The kanamycin B derivative (600 mg) obtained by the method described in c) in aqueous acetic acid (1: 3, 20 ml) was heated at 95 ° C for 30 minutes. The reaction mixture was evaporated to dryness and the residue was treated with ether. The ether-insoluble portion was dissolved in aqueous acetone (1: 1, 15 ml), anhydrous sodium carbonate (400 mg) was added to the solution, then ethoxycarbonyl chloride (270 mg) was added, and stirring was continued at room temperature for 5 hours.

The resulting precipitate was filtered, washed with water and dried to give a solid (515 mg), mp + 98 ° (C = 1.5, dimethylformamide). The material could be converted to 3 ', 4'-dideoxycanamycin B by a series of reactions similar to those described in Example 1, steps e-i.

Claims (1)

Process for the preparation of 3 ', 4'-dideoxycanamycin B having the formula 6' CH2NH2 Γχ r X * tNH2 H2O2OH, which is effective against infections caused by bacterial and in particular kanamycin-resistant organisms, characterized in that kanamycin is protected in a manner known per se. Amino groups of 6 'ch2nh2 *' \ 0H, / t? '™ 2 ^ 0 \ / cH) l hoh2c 3> 6 oi' * '\ ηκ2 Λ / ", Η- OH n with the aid of chloroformic acid ethyl ester or salicylaldehyde, which then the 3 'and 4' and 4 "and 6" hydroxyl groups are protected with acetone or cyclohexanone dimethyl ketal, then the 2 "hydroxyl group is benzoylated in the usual manner, cleaved from the resulting 3 ', 4'; 4", 6 "-di-O- from an isopropylidene or 3 ', 4': 4 ", 6" -di-O-cyclohexylidene derivative by heating acetone or cyclohexanone with aqueous acetic acid, the 3'- and 4'-hydroxyl groups are converted in the usual manner into methanesulfonic acid chloride by these 3', 4'-disulfon natriumjodi din and zinc to give the 3 ', 4'-didehydro compound, these are catalytically hydrogenated in a manner known per se with hydrogen in the presence of wt PtO 4 and finally the remaining protecting groups are removed in a manner known per se, first with dilute acetic acid and then with barium hydroxide. 57596 12 For the preparation of 3 ', 4'-dideoxycanamycin B is also antibacterial and specific Active substance for infection with oramamycin-resistant organisms and in the form of 6' ch2nh2 HOH0C 51 "76 H0 V-I2 * · OH amino acids B 6 'ch2nh2 5' /) —— o _ 4 '| (oh nh2 81.2 0 6 "Vj- \ e HOH2C 1 J 5 / ~ 0 o' 4M (nh2} / OH