PYRAZOLOTETRACYCLINES

Abstract: Novel pyrazolotetracyclines are described, which have formula (I), in which R is hydrogen, a substituted or unsubstituted (C1-C6)alkyl, (C2-C6)alkenyl or (C2-C6)alkynyl group, a substituted or unsubstituted aryl or heteroaryl group, a substituted or unsubstituted cycloalkyl, cycloalkenyl or cycloalkadienyl group and R' represents an amino, dimethylamino, (C1-C3)alkylamino or formylamino group. These pyrazolotetracyclines are antibiotics useful for the treatment of infectious diseases.

Description: PYRAZOLOTETRACYCLINES The present invention concers novel tetracycline derivatives, more particularly novel pyrazolotetracyclines, a process for their preparation and pharmaceutical compositions containing them. Tetracyclines are among the most studied antibiotics since their discovery, at the end of years forty. Many books have been dedicated to the chemistry, to the activity of these products and, more particularly, to the relationship between their chemical structure and their antibiotic activity. Deep structure/activity studies on tetracyclines have led to conclude that the basic structure of the natural tetracycline of formula A which is a naphthacenecarboxamide derivative called, according to the Chemical Abstracts nomenclature (CAS Registry Number 60-54-8), [4S, (4α,4aα,5aα,6β,12aα) ]-4-(dimethylamino)-1,4,4a,5,5a,6, 11, 12a-octa hydro-3,6,10,12,12a-pentahydroxy-6-methyl-l,ll-dioxo-2-naphthacenecar boxamide, may be irreversibly, but in an appropriate manner, modified in the positions from 4a to 9 in order to obtain compounds still endowed with antibiotic activity, whilst any irreversible modification in the positions 1, 2, 3, 4 and from 10 to 12a involves a loss of the antibiotic activity. Therefore, these positions are considered as "inviolate" (Medical Research Series, Vol. 9, Lester A. Mitscher, The Chemistry of the Tetracycline Antibiotics, Marcel Dekker, Inc., 1978). Infact, Valcavi et al., Gazz. Chi . Ital., 1963, 93, 916-928, described certain pyrazolotetracyclines, i.e. compounds obtained by modification of the structure of tetracycline and chlorotetracycline on the positions 11 and 12 by the action of hydrazine and concluded that these compounds had lost their activity or were less active than the starting tetracyclines. it has now been found that pyrazolotetracyclines bearing an optionally substituted amino group in the 7-position of the tetracycline moiety are at least as active as the parent tetracyclines, and even more active. More particularly, it has been found that, notwithstanding the modification of the tetracycline structure on the positions 11 and 12, the novel compounds are endowed with the same properties of the parent tetracycline and are even more active on certain micro-organisms. Thus, it is an object of the present invention to provide novel pyrazolotetracyclines of formula (I) in which R is hydrogen, a substituted or unsubstituted (C.-C 8 )alkyl, (C 2 -C,)alkenyl or (C 2 -Cg)alkynyl group, a substituted or unsubstitu ted aryl or heteroaryl group, a substituted or unsubstituted cycloalkyl, cycloalkenyl or cycloalkadienyl group and R' represents a basic group, preferably an amino, dimethylamino, (C.-C- j )alkylamino or formyleunino group, and their acid addition salts, their solvates and their complexes. Among the salts, the solvates and the complexes, those which are pharmaceutically acceptable or which allow an easy separation of the pyrazolotetracycline involved are particularly preferred. The preferred salts are the hydrochloride, the hydrogenosulfate, the sulfate. The solvates can be those of the free base or of the salts, the hydrates being preferred. The complexes are generally those which allow an easy isolation of the pyrazolotetracycline, the complex with calcium chloride dihydrate being particularly preferred. A subclass of compounds according to the present invention includes pyrazolotetracyclines of formula I above, wherein R' is as defined above and R is hydrogen or a substituted or unsubstituted (C,-Cg)alkyl, (C 2 -Cg)alkenyl or (C 2 -Cg)alkynyl group; a cycloalkyl group of from 3 to 8 carbon atoms; a cycloalkenyl group of from 4 to 8 carbon atoms, such as cyclohexenyl, more particularly 1-cyclo hexenyl; a phenyl group which may be unsubstituted or substituted with one or more substituents selected from the group consisting of halogens, particularly fluoro, chloro and iodo substituents, ( c ι ~ C 3 )alkyl, (C.-C,)alkoxy, trifluoromethyl, pentafluoroethyl, amino, nitro, cyano, carboxy, methoxy- or ethoxycarbonyl, phenyl, chloro phenyl, bromophenyl, fluorophenyl, (C 1 ~C 3 )alkoxyphenyl; a naphthyl group which may be unsubstituted or substituted with one or two methoxy groups; a mono-, bi- or tricyclic heteroaryl group, preferably selected from the group consisting of lH-pyrrolyl, 1-(C 1 ~ C,)alkylpyrrolyl, furyl, thienyl, pyridyl, oxazolyl, lH-imidazolyl, 1-(C.-C-. ) alkylimidazolyl, thiazolyl, isothiazolyl, pyrimidyl, pyrazinyl, 1,3,4-thiadiazolyl, 1,2,4-thiadiazolyl, furazanyl, 1,3,5- triazinyl, 2-, 3-, 4-, 5-, 6- and 7-benzofuranyl or benzothienyl, 1H- and l-(C 1 -C 3 )alkyl-2-, 3-, 4-, 5-, 6- and 7-indolyl, 2-, 3-, 4-, 5-, 6-, 7- and 8-quinolyl, 1-, 3-, 4-, 5-, 6-, 7- and 8-isoquinolyl, 2- benzothiazolyl, quinoxalinyl, phthalazinyl, carbazolyl, as well as their salts, solvates and complexes, those which are pharmaceutically acceptable or allow an easy isolation of the pyrazolotetracycline being particularly preferred. Another advantageous subclass of compounds according to the present invention includes those of formula I wherein R is hydrogen or a group as defined above and R* is selected from the group consisting of formylamino, methylamino, isopropylamino and dimethyl amino, as well as their salts, solvates and complexes which are pharmaceutically acceptable or which allow an easy separation of the pyrazolotetracycline involved. Among these last compounds, that of formula I in which R is hydrogen and R' is dimethylamino (configuration of the 7-dimethylami no-6-demethyl-6-deoxytetracycline or minocycline) hereinbelow designated "pyrazolominocycline" as well as its salts, solvates and complexes which are pharmaceutically acceptable or which allow an easy isolation of the pyrazolotetracycline, for example the complex with calcium chloride dihydrate, are particularly preferred. Even in the case of the compounds of the present invention the description and the examples which follow will refer, in general, to pyrazolotetracyclines. Actually the correct nomenclature of these compounds is [9S(7aα,8aα,9α-12aα) ]2a,7,7a,8,8a,9,12, 12a-octahydro(lH) benzo[g]naphth[ 1, 2 , 3-cd] indazole and their structure (B) is as follows: According to another of its aspects, the present invention provides a process for the preparation of pyrazolotetracyclines of formula I and of their salts, solvates and complexes which comprises reacting the corresponding tetracycline of formula II wherein R' is as defined above, or a salt or solvate thereof, with a hydrazine of formula III R-NH-NH 2 (III) wherein R is as defined above and isolating the pyrazolotetracycline thus obtained in form of a base or of a complex thereof and, if necessary, transforming the free base in an acid addition salt thereof. The reaction of the tetracycline of formula II or of its salt or solvate with the hydrazine of formula III is easy and rapid. It occurs in an organic or aqueous-organic, preferably alcoholic or hydroalcoholic solvent, preferably in methanol. The reaction temperature is the room temperature (20÷30°C) when unsubstituted hydrazine (III, R = H) is used or a temperature of from 25 to 90°C when a substituted hydrazine is used to obtain a pyrazolotetracycline of formula I wherein R is other than hydrogen. The reaction is generally over after 1 to 5 hours and the compound I thus obtained is isolated according to the conventional procedures of the tetracyclines chemistry, for example by acidifying the reaction mixture and by forming a complex of the compound obtained therein with a calcium salt, preferably with calcium chloride dihydrate in an aqueous-acidic medium. The desired end product is then recovered from its complex by precipitation of the calcium .with an organic or inorganic acid capable of forming an insoluble calcium salt, separation of the insoluble calcium salt, correction of pH to about 8 and isolation of the pyrazolotetracycline in form of base. Generally, the recovery of the end product occurs by treatment with oxalic acid, elimination of the calcium oxalate and isolation of the pyrazolotetracycline by adjustment of the pH to alkaline values (7.5÷9.0). The isolation of the end product may also occur without using the calcium complex, by simple evaporation of the solvent and adjustment of the pH of the aqueous phase to about 8. The pyrazolotetracycline base may be converted to their acid addition salts by treatment with an inorganic or organic acid, for example with hydrochloric, hydrobromic, sulfuric, methanesulfonic, p- toluenesulfonic or 2-naphthalenesulfonic acid. The pyrazolotetracyclines of the present invention having the basic structure B can exist in three theoretical tautomeric forms I, I' and I" represented by the following structures: Studies carried out by the applicant have shown that only the compounds of formula I are pharmacologically active and lead to satisfactory results. The novel " tetracyclines of the present invention are useful as antibiotics which, in certain cases, exhibit better properties than those of the parent tetracyclines. More particularly, the M.I.C. of the compound of formula I (R = H, R' = dimethylamino) on Staphilococcus aureus ATCC 29737 is 0.4 mcg/ l, in respect of the value of 0.75 mcg/ml found for minocycline. Furthermore, these novel pyrazolotetracyclines possess a low toxicity, as the known, parent tetracyclines. For example, the DL_ 0 of pyrazolominocycline in rat is greater than 4000 g/Kg, in comparison with that of minocycline which, in the same conditions, is of 3300 mg/Kg. Therefore, the novel pyrazolotetracyclines of the present invention may be used as active ingredients of pharmaceutical compounds, in unit doses of from 50 to 500 mg of active ingredient to be administered one to four times daily. The novel pyrazolotetracyclines of the present invention may be employed for the preparation of pharmaceutical compositions for oral use, for example in gelatin capsules, tablets, granulates, as such or with the common pharmaceutical carriers. Such pharmaceutical compositions contain the pyrazolotetracyclines of the present invention in an amount equivalent to 50÷500 mg of anhydrous pyrazolo tetracycline, the preferred active ingredient being pyrazolo minocycline or one of its pharmaceutically acceptable salts or solvates. The following examples illustrate the invention without, however, limiting it. EXAMPLE 1 To a solution of 25 g of minocycline hydrochloride dihydrate in 500 ml of methanol, 10 ml of hydrazine hydrate are added, then the reaction mixture is let to stand 3 hours at 20÷25°C under stirring. Afterwards, the pH is adjusted to a value of about 1 by addition of 37% hydrochloric acid and the mixture is concentrated under vacuum. The residue is taken up with a solution of 4 g of calcium chloride dihydrate in 100 ml of water and 5 ml of 37% hydrochloric acid, then the pH of the solution is brought to 3.5 by addition of 15% sodium hydroxide. By filtering the obtained precipitate, washing it with water and drying under vacuum, 17 g of complex of pyrazolominocycline with calcium chloride dihydrate are obtained. Purity (HPLC) : 96.9%. Elementar analysis: Found C = 50.74%; H = = 5.24%; N = 12.50%; Cl = 9.76%; Ca = 2.04%. H j O (K.F.) = 6.33%. The complex thus obtained is suspended in a mixture ethanol/water containing oxalic acid, then the pH of the suspension is adjusted to a value of about 0.7. Dicalite (Dicalite Europa Sud S.p.A., Corsico, Milan, Italy) is then added, the pH is brought to 1.8 and, after a 10-minute stirring, the mixture is filtered in order to obtain a clear solution. To this solution there is added 15% sodium hydroxide to a pH of about 8, then the precipitate is recovered by filtration, washed with water and dried under vacuum to give 13 g of pyrazolominocycline base corresponding to the formula IV The product thus obtained is dissolved in ethanol and acidified with 37% hydrochloric acid to a pH of about 4, then the mixture is cooled, the solid is filtered, washed and dried under vacuum. Thus, there are obtained 7 g of l9S-(7aα. 8aα, 9α, 12aα) ]-6,9-(bis-dimethyl. amino)-2a,7,7a,8,8a,9, 12,12a-octahydro-3-10,12a-trihydroxy-12-oxo(IH) benzo[g]naphth[ 1,2,3-cd]indazole-ll-carboxamide hydrochloride (pyrazo lominocycline hydrochloride) . 1 H-NMR (D 2 0) 6 (p.p.m.): 1.21 (m, 2H) ; 2.10 (m, IH) ; 2.7÷3.0 (s+m, 9H); 3.30 (s, 6H) ; 3.63 (d, IH) ; 6.93 (d, J=8.8 Hz, IH); 7.41 (d, J=8.8 Hz, IH). 13 C-NMR (D 2 0+HC1) δ (p.p.m.): 27.5; 30.7; 36.7; 40.0; 43.0; 48.0; 73.3; 74.2; 103.3; 116.6; 116.7; 119.7; 121.1; 130.7; 133.8; 138.2; 142.3; 153.6; 172.0; 186.5; 191.3. Mass spectrum (FAB+): m/e 454 molecular ion (M+H) ' principal fragments : m/e 437 , 419 , 391 , 348 , 289 , 259 , 246 , 207 . EXAMPLE 2 To a solution of 20 g of minocycline hydrochloride dihydrate in 400 of methanol, 8 ml of hydrazine hydrate are added and the mixture is kept 3 hours at 20+25°C under stirring, then it is concentrated under vacuum. The residue is taken up with water, the pH of the solution is brought to 8 by addition of 15% sodium hydroxide, then the solid is filtered, washed with water and dried under vacuum. Thus, there is obtained 18 g of pyrazolominocycline base having a purity of 85.3% (HPLC) .

Claims: CLAIMS 1. Pyrazolotetracyclines of formula I in which R is hydrogen, a substituted or unsubstituted (C.-Cg)alkyl, (C 2 -Cg)alkenyl or (C 2 ~Cg)alkynyl group, a substituted or unsubstituted aryl or heteroaryl group, a substituted or unsubstituted cycloalkyl, cycloalkenyl or cycloalkadienyl group and R' represents a basic group and their acid addition salts, their solvates and their complexes. 2. Pyrazolotetracyclines according to claim 1 of formula I, wherein said basic group is selected among amino, dimethylamino, (C.- C 3 )alkylamino or formylamino. 3. Pyrazolotetracyclines according to claim 1 of formula I, wherein R' is as defined above and R is hydrogen or a substituted or unsubstituted (C 1 -C 8 )alkyl, (C 2 ~C 6 ) lkenyl or (C 2 -Cg)alkynyl group; a cycloalkyl group of from 3 to 8 carbon atoms; a cycloalkenyl group of from 4 to 8 carbon atoms; a phenyl group which may be unsubstituted or substituted with one or more substituents selected from the group consisting of halogens, particularly fluoro, chloro and iodo substituents, (C 1 -C- J )alkyl, (C 1 -C 3 )alkoxy, trifluoromethyl, penta fluoroethyl, amino, nitro, cyano, carboxy, methoxy- or ethoxy carbonyl, phenyl, chlorophenyl, bromophenyl, fluorophenyl, (C 1 ~ C 3 )alkoxyphenyl; a naphthyl group which may be unsubstituted or substituted with one or two methoxy groups; a mono-, bi- or tricyclic heteroaryl group, selected from the group consisting of lH-pyrrolyl, l-tC. -C 3 ) alkylpyrrolyl, furyl, thienyl, pyridyl, oxazolyl, 1H- imidazolyl, 1-(C.-C 3 ) lkylimidazolyl, thiazolyl, isothiazolyl, pyrimidyl, pyrazinyl, 1, 3, 4-thiadiazolyl, 1, 2 ,4-thiadiazolyl, furazanyl, 1,3,5-triazinyl, 2-, 3-, 4-, 5-, 6- and 7-benzofuranyl or benzothienyl, IH- or l-(C 1 -C 3 )alkyl-2-, 3-, 4-, 5-, 6-, 7-indolyl, 2-, 3-, 4-, 5-, 6-, 7- and 8-quinolyl, 1-, 3-, 4-, 5-, 6-, 7- and 8- isoquinolyl, 2-benzothiazoly1, quinoxalinyl, phthalaziny1, carbazolyl, as well as their salts, solvates and complexes. 4. Pyrazolotetracyclines according to claim 1 of formula I, wherein R is hydrogen or a group as defined in claim 3 and R" is selected from the group consisting of formylamino, methylamino, isopropylamino and dimethylamino, as well as their salts, solvates and complexes which are pharmaceutically acceptable or which allow an easy separation of said pyrazolotetracycline. 5. Pyrazolotetracyclines according to claim 1 of formula I, in which R is hydrogen and R' is dimethylamino and its salts, solvates and complexes. 6. [9S-(7aα,8aα,9α,12aα) ]-6,9-(bis-dimethylamino)-2a,7,7a,8,8a,9, 12,12a-octahydro-3,10,12a-trihydroxy-12-oxo(IH)benzo[g]naphth[1,2,3- cd]indazole-11-carboxamide of formula IV and its salts, solvates and complexes which are pharmaceutically acceptable or which allow an easy isolation of said [9S-(7aα,8aα,9α, 12aα) ]-6,9-(bis-dimethylamino)-2a,7,7a,8,8a,9,12,12a-octahydro-3,10, 12a-trihydroxy-12-oxo(lH)benzo[g]naphth[l,2,3-cd]indazole-ll-carboxa mide. 7. Complex with calcium chloride dihydrate of [9S-(7aα,8aα,9α, 12aα) ]-6,9-(bis-dimethylamino)-2a,7,7a,8,8a,9,12,12a-octahydro-3,10, 12a-trihydroxy-12-oxo(IH)benzo[g]naphth[1,2,3-cd]indazole-11-carboxa mide. 8. [9S-(7aα,8aα,9α,12aα) ]-6,9-(bis-dimethylamino)-2a,7,7a,8,8a,9, 12, 12a-octahydro-3,10,12a-trihydroxy-12-oxo(IH)benzo[g]naphth[1,2,3- cdJindazole-11-carboxamide hydrochloride. 9. A process for the preparation of pyrazolotetracyclines according to claim 1 which comprises reacting the corresponding tetracycline of formula II wherein R' is as defined in claim 1, or a salt or solvate thereof, with a hydrazine of formula III R-NH-NH, (III) wherein R is as claimed in claim 1. 10. A process according to claim 9 wherein the reaction is carried out in an alcoholic solvent at a temperature of from 20 to 90 β C in the presence of calcium chloride to give the pyrazolotetracycline in form of a complex. 11. A process according to claim 10 wherein the pyrazolotetra cycline complex is treated at pH 1.8 with an inorganic or organic acid capable of forming an insoluble calcium salt, the pH is adjusted to about 8 and the pyrazolotetracycline is isolated as a base. 12. A process according to claim 11, wherein said acid is oxalic acid. 13. Pharmaceutical composition for oral use containing an amount of from 50 to 500 mg of pyrazolotetracycline according to claim 1, as such or in admixture with a pharmaceutical carrier, per dosage unit. 14. A pharmaceutical composition according to claim 13 in which said pyrazolotetracycline is the [9S-(7aα,8act,9α,12aα) ]-6,9-(bis-dime thylamino)-2a,7,7a,8,8a,9,12,12a-octahydro-3, 10, 12a-trihydroxy-12-oxo (lH)benzo[g)naphth[1,2,3-cd]indazole-11-carboxamide or a pharmaceuti. cally acceptable salt or solvate or complex thereof. 15. A pharmaceutical composition according to claim 14 in which said pyrazolotetracycline is the [9S-(7aα,8aα,9α,12aα) ]-6,9-(bis-dime thylamino)-2a,7,7a,8,8a,9, 12, 12a-octahydro-3, 10,12a-trihydroxy-12-oxo (lH)benzo[g]naphth[1,2,3-cd]indazole-11-carboxamide hydrochloride.

Inventors: DA COL MARCO, DALL ASTA LEONE, RESTA IRENE, SEGHETTI ENNIO, DA COL, MARCO, DALL'ASTA, LEONE, RESTA, IRENE, SEGHETTI, ENNIO
Applicants: DA COL MARCO, DALL ASTA LEONE, OPOS BIOCHIMICA SRL, RESTA IRENE, SEGHETTI ENNIO, BIOCHIMICA OPOS S.P.A., DA COL, MARCO, DALL'ASTA, LEONE, RESTA, IRENE, SEGHETTI, ENNIO
Application year: 1996
Main classification: C07D231
IPC codes: A61K31, A61P31, C07D231