2 amino-3,4-dihydrcquinazoline derivatives and the use thereof as cathepsin D inhibitors
09663475 ยท 2017-05-30
Assignee
Inventors
Cpc classification
A61P29/00
HUMAN NECESSITIES
A61P17/02
HUMAN NECESSITIES
C07D403/06
CHEMISTRY; METALLURGY
A61P19/08
HUMAN NECESSITIES
A61K31/517
HUMAN NECESSITIES
A61K31/5377
HUMAN NECESSITIES
C07D413/06
CHEMISTRY; METALLURGY
C07C209/36
CHEMISTRY; METALLURGY
A61P35/00
HUMAN NECESSITIES
C07D239/95
CHEMISTRY; METALLURGY
A61P25/14
HUMAN NECESSITIES
A61K31/538
HUMAN NECESSITIES
C07C201/12
CHEMISTRY; METALLURGY
A61P25/28
HUMAN NECESSITIES
C07C213/02
CHEMISTRY; METALLURGY
C07C331/28
CHEMISTRY; METALLURGY
A61P37/06
HUMAN NECESSITIES
A61P15/00
HUMAN NECESSITIES
International classification
A61K31/517
HUMAN NECESSITIES
A61K31/538
HUMAN NECESSITIES
C07D239/95
CHEMISTRY; METALLURGY
C07C331/28
CHEMISTRY; METALLURGY
C07C213/02
CHEMISTRY; METALLURGY
C07C209/36
CHEMISTRY; METALLURGY
C07C201/12
CHEMISTRY; METALLURGY
C07D403/06
CHEMISTRY; METALLURGY
C07D413/06
CHEMISTRY; METALLURGY
Abstract
The present invention relates to compounds of the formula (I) and in particular to medicaments comprising at least one compound of the formula (I) for use in the treatment and/or prophylaxis of physiological and/or pathophysiological conditions in the triggering of which cathepsin D is involved, in particular for use in the treatment and/or prophylaxis of osteoarthritis, traumatic cartilage injuries, arthritis, pain, allodynia or hyperalgesia.
Claims
1. A compound of the formula I, ##STR00069## in which I.sup.1, I.sup.2, I.sup.3, independently of one another, denote CR.sup.1 or CT, X denotes H or NH.sub.2, Y denotes a cyclic alkylaryl group, characterised in that 1 or 2 aromatic rings Ar are condensed onto a cyclic alkyl having 5 or 6 C atoms, in which one or two CH.sub.2 groups may be replaced, independently of one another, by O, S, SO, SO.sub.2, NR, OCO, NRCONR, NRCO, NRSO.sub.2R, COO, CONR, and/or, in addition, 1-11 H atoms may be replaced by F and/or Cl, and which is unsubstituted or mono- or disubstituted by S, NR, O, R, T, OR, NRR, SOR, SO.sub.2R, SO.sub.2NRR, CN, COOR, CONRR, NRCOR, NRCONRR and/or NRSO.sub.2R, Ar denotes a phenyl or naphthyl, each of which is unsubstituted or mono-, di- tri- or tetrasubstituted by R.sup.1, or a mono- or bicyclic aromatic heterocycle having 1 to 4 N, O and/or S atoms, which is unsubstituted or mono-, di- or trisubstituted by R, S, NR and/or O, Q denotes CH.sub.2, CR.sup.1R.sup.2 or CO, T denotes a phenyl or naphthyl, each of which is unsubstituted or mono-, di- tri- or tetrasubstituted by R.sup.1, or a mono- or bicyclic saturated, unsaturated or aromatic heterocycle having 1 to 4 N, O and/or S atoms, which may be mono-, di- or trisubstituted by R, S, NR and/or O, R.sup.1, R.sup.2, independently of one another, denote H, OR, Hal, C(Hal).sub.3, NRR, SOR, SO.sub.2R, SO.sub.2NRR, CN, COOR, CONRR, NRCOR, NRCONRR, NRSO.sub.2R, a linear or branched alkyl having 1-10 C atoms, in which one, two or three CH.sub.2 groups may be replaced, independently of one another, by O, S, SO, SO.sub.2, NR, OCO, NRCONR, NRCO, NRSO.sub.2R, COO, CONR, NRCO, CC groups and/or by CHCH groups and/or, in addition, 1-20 H atoms may be replaced by F and/or Cl, and which is unsubstituted or mono-, di- or trisubstituted by S, NR, O, Hal, C(Hal).sub.3, OR, NRR, SO.sub.2R, SO.sub.2NRR, CN, CONRR, NRCOR and/or NRCONRR, or a cyclic alkyl having 3-7 C atoms, in which one, two or three CH.sub.2 groups may be replaced, independently of one another, by O, S, SO, SO.sub.2, NR, OCO, NRCONR, NRCO, NRSO.sub.2R, COO, CONR, NRCO and/or by CHCH groups and/or, in addition, 1-11 H atoms may be replaced by F and/or Cl, and which is unsubstituted or mono-, di- or trisubstituted by S, NR, O, Hal, OR, NRR, SO.sub.2R, SO.sub.2NRR, CN, CONRR, NRCOR, and/or NRCONRR, R, R, R, independently of one another, denote H, T, OH, Hal, C(Hal).sub.3, NH.sub.2, SO-alkyl, SO.sub.2-alkyl, SO2NH.sub.2, CN, COOH, CONH.sub.2, NHCO-alkyl, NHCONH.sub.2, NHSO.sub.2-alkyl and/or NHCO-alkyl, a linear or branched alkyl having 1-10 C atoms, in which one, two or three CH.sub.2 groups may be replaced, independently of one another, by O, S, SO, SO.sub.2, NH, NCH.sub.3, OCO, NHCONH, NHCO, NHSO.sub.2-alkyl-, COO, CONH, NCH.sub.3CO, CONCH.sub.3, CC groups and/or by CHCH groups and/or, in addition, 1-20 H atoms may be replaced by F and/or Cl, and which is unsubstituted or mono-, di- or trisubstituted by S, NR, O, Hal, C(Hal).sub.3, OH, NH.sub.2, SO.sub.2CH.sub.3, SO.sub.2NH.sub.2, CN, CONH.sub.2, NHCOCH.sub.3, and/or NHCONH.sub.2, or a cyclic alkyl having 3-7 C atoms, in which one, two or three CH.sub.2 groups may be replaced, independently of one another, by O, S, SO, SO.sub.2, NH, NCH.sub.3, OCO, NHCONH, NHCO, NHSO.sub.2-alkyl-, COO, CONH, NCH.sub.3CO, CONCH.sub.3 and/or by CHCH groups and/or, in addition, 1-11 H atoms may be replaced by F and/or Cl, and which is unsubstituted or mono-, di- or trisubstituted by S, NR, O, C(Hal).sub.3, OH, NH.sub.2, SO.sub.2CH.sub.3, SO.sub.2NH.sub.2, CN, CONH.sub.2, NHCOCH.sub.3, and/or NHCONH.sub.2, or R and R or R and R or R and R, if both are bonded to an N, may form a ring having 3-7 C atoms incorporating the N, in which one, two or three CH.sub.2 groups may be replaced, independently of one another, by O, S, SO, SO.sub.2, NH, N-alkyl, N-aryl, CHT-, CH(CH.sub.2T)-, OCO, NHCONH, NHCO, NHSO.sub.2, COO, CON alkyl- and/or by CHCH groups and/or, in addition, 1-11 H atoms may be replaced by F and/or Cl, characterised in that 1 or 2 aromatic rings Ar may be condensed onto this ring, and Hal, independently of one another, denotes F, Cl, Br or I, a physiologically acceptable salt of a compound of formula I, a solvate of a compound of formula I, a stereoisomer of a compound of formula I or a mixture thereof in all ratios.
2. A compound according to formula I of claim 1 in which Y is selected from the group consisting of the following radicals, which are unsubstituted or a mono- or disubstituted by S, NR, O, R, R.sup.1, T, OR, NRR, SOR, SO.sub.2R, SO.sub.2NRR, CN, COOR, CONRR, NRCOR, NRCONRR and/or NRSO.sub.2R: ##STR00070## Q denotes CH.sub.2 or CO and R.sup.1, independently of one another, denotes H, CF.sub.3, OR, Hal, CN, CONRR, a linear or branched alkyl having 1-10 C atoms or cyclic alkyl having 3-7 C atoms, in which one, two or three CH.sub.2 groups may be replaced, independently of one another, by O, CHCH groups and/or, in addition, 1-11 H atoms may be replaced by F and/or Cl, and which is unsubstituted or mono-, di- or trisubstituted by O, Hal, C(Hal).sub.3, OR, NRR, SO.sub.2R, SO.sub.2NRR, CN, CONRR, NRCOR and/or NRCONRR, and I.sup.1, I.sup.2, I.sup.3, X, Ar, T, R, R, R and Hal have the meanings indicated in claim 1, a physiologically acceptable salt thereof, a solvate thereof, a stereoisomer thereof or a mixture thereof in all ratios.
3. A compound according to formula I of claim 1 in which I.sup.1 denotes CH, I.sup.2 denotes CR.sup.1 or CT, I.sup.3 denotes CH or CCl, X denotes H, Y is selected from the group consisting of the following radicals, which are unsubstituted or a mono- or disubstituted by S, NR, O, R, R.sup.1, T, OR, NRR, SOR, SO.sub.2R, SO.sub.2NRR, CN, COOR, CONRR, NRCOR, NRCONRR and/or NRSO.sub.2R: ##STR00071## Q denotes CH.sub.2, R.sup.1, independently of one another, denotes H, CF.sub.3, OR, Hal, CN, CONRR, a linear or branched alkyl having 1-10 C atoms or cyclic alkyl having 3-7 C atoms, in which one, two or three CH.sub.2 groups may be replaced, independently of one another, by O, CHCH groups and/or, in addition, 1-11 H atoms may be replaced by F and/or Cl, and which is unsubstituted or mono-, di- or trisubstituted by O, Hal, C(Hal).sub.3, OR, NRR, SO.sub.2R, SO.sub.2NRR, CN, CONRR, NRCOR and/or NRCONRR, and Ar, T, R, R, R and Hal have the meanings indicated in claim 1, a physiologically acceptable salt thereof, a solvate thereof, a stereoisomer thereof or a mixture thereof in all ratios.
4. A compound according to formula I of claim 1 in which I.sup.1 denotes CH, I.sup.2 denotes CR.sup.1 or CT, I.sup.3 denotes CH or CCl, X denotes H, Y is selected from the group consisting of the following radicals, which are unsubstituted or a mono- or disubstituted by methoxyl: ##STR00072## Q denotes CH.sub.2, R.sup.1, independently of one another, denotes H, CF.sub.3, OR, Hal, CN, CONRR, a linear or branched alkyl having 1-10 C atoms or cyclic alkyl having 3-7 C atoms, in which one, two or three CH.sub.2 groups may be replaced, independently of one another, by O, CHCH groups and/or, in addition, 1-11 H atoms may be replaced by F and/or Cl, and which is unsubstituted or mono-, di- or trisubstituted by O, Hal, C(Hal).sub.3, OR, NRR, SO.sub.2R, SO.sub.2NRR, CN, CONRR, NRCOR and/or NRCONRR, and Ar, T, R, R, R and Hal have the meanings given in claim 1, a physiologically acceptable salt thereof, a solvate thereof, a stereoisomer thereof or a mixture thereof in all ratios.
5. A compound according to formula I of claim 1 in which R and R or R and R or R and R, if both are bonded to an N, form a ring having 3-7 C atoms incorporating the N, in which one, two or three CH.sub.2 groups may be replaced, independently of one another, by O, S, SO, SO.sub.2, NH, N-alkyl, N-aryl, CHT-, CH(CH.sub.2T)-, OCO, NHCONH, NHCO, NHSO.sub.2, COO, CON-alkyl- and/or by CHCH groups and/or, in addition, 1-11 H atoms may be replaced by F and/or Cl, characterised in that 1 or 2 aromatic rings Ar may be condensed onto this ring and I.sup.1, I.sup.2, I.sup.3, X, Y, Q, Ar, T, R.sup.1, R.sup.2 and Hal have the meanings given in claim 1, a physiologically acceptable salt thereof, a solvate thereof, a stereoisomer thereof or a mixture thereof in all ratios.
6. A compound according to formula I of claim 1 in which I.sup.1 denotes CH, I.sup.2 denotes CR.sup.1 or CT and R.sup.1 or T is selected from the group consisting of: H, CF.sub.3, CI, ethyl, propyl, phenyl, ##STR00073## I.sup.3 denotes CH or CCl, X denotes H, Y is selected from the group consisting of a following radical, which is unsubstituted or a mono- or disubstituted by methoxyl: ##STR00074## Q denotes CH.sub.2, a physiologically acceptable salt thereof, a solvate thereof, a stereoisomer thereof or a mixture thereof in all ratios.
7. A compound selected from the group consisting of: a) 3-indan-2-yl-3,4-dihydroquinazolin-2-ylamine b) 7-chloro-3-indan-2-yl-3,4-dihydroquinazolin-2-ylamine c) 5-chloro-3-indan-2-yl-3,4-dihydroquinazolin-2-ylamine d) 3-indan-2-yl-7-phenyl-3,4-dihydroquinazolin-2-ylamine e) 7-chloro-3-(1,2,3,4-tetrahydronaphthalen-1-yl)-3,4-dihydroquinazolin-2-ylamine f) 3-indan-2-yl-7-propyl-3,4-dihydroquinazolin-2-ylamine g) 7-chloro-3-(5,6-dimethoxyindan-2-yl)-3,4-dihydroquinazolin-2-ylamine h) 3-indan-2-yl-7-trifluoromethyl-3,4-dihydroquinazolin-2-ylamine i) 7-chloro-3-(4,5-dimethoxyindan-2-yl)-3,4-dihydroquinazolin-2-ylamine j) 7-chloro-3-(4-methoxyindan-2-yl)-3,4-dihydroquinazolin-2-ylamine k) 3-indan-1-yl-7-trifluoromethyl-3,4-dihydroquinazolin-2-ylamine l) 7-chloro-3-(5-methoxyindan-2-yl)-3,4-dihydroquinazolin-2-ylamine m) 3-(9H-fluoren-9-yl)-7-trifluoromethyl-3,4-dihydroquinazolin-2-ylamine n) 3-(5-methoxyindan-2-yl)-7-trifluoromethyl-3,4-dihydroquinazolin-2-ylamine o) 7-ethyl-3-(5-methoxyindan-2-yl)-3,4-dihydroquinazolin-2-ylamine p) 3-((S)-5-methoxyindan-2-yl)-7-trifluoromethyl-3,4-dihydroquinazolin-2-ylamine q) 3-((R)-5-methoxyindan-2-yl)-7-trifluoromethyl-3,4-dihydroquinazolin-2-ylamine r) 3-(1,2,3,4-tetrahydronaphthalen-2-yl)-7-trifluoromethyl-3,4-dihydroquinazolin-2-ylamine s) (2-amino-3-indan-2-yl-3,4-dihydroquinazolin-7-yl)-(2,3-dihydroindol-1-yl)-methanone t) (2-amino-3-indan-2-yl-3,4-dihydroquinazolin-7-yl)-(5-methoxy-1,3-dihydroisoindol-2-yl)methanone u) N,N-diethyl-2-amino-3-indan-2-yl-3,4-dihydroquinazoline-7-carboxamide v) (2-amino-3-indan-2-yl-3,4-dihydroquinazolin-7-yl)morpholin-4-ylmethanone w) 2-amino-3-indan-2-yl-7-trifluoromethyl-3H-quinazolin-4-one x) 2-hydrazino-3-indan-2-yl-7-trifluoromethyl-3H-quinazolin-4-one y) (2-amino-3-indan-2-yl-3,4-dihydroquinazolin-7-yl)-(2-benzylpyrrolidin-1-yl)methanone z) (2-amino-3-indan-2-yl-3,4-dihydroquinazolin-7-yl)-(2,3-dihydrobenzo-[1,4]oxazin-4-yl)methanone aa) 3-((1R,2S)-1-methoxyindan-2-yl)-7-trifluoromethyl-3,4-dihydro-1H-quinazolin-2-ylidenamine a physiologically acceptable salt thereof, a solvate thereof, a stereoisomer thereof or a mixture thereof in all ratios.
8. A process for the preparation of the compounds of the formula I of claim 1 in which X denotes H, Q denotes CH.sub.2 and and I.sup.1, I.sup.2, I.sup.3, Y, Ar, T, R.sup.1, R.sup.2, R, R, R and Hal have the meanings indicated in claim 1, characterised in that a compound of the formula II is converted into a compound of the formula Ill by reductive amination, a compound of the formula III is converted into a compound of the formula IV by hydrogenation in the presence of a catalyst, a compound of the formula IV is reacted with cyanogen bromide to give a compound of the formula V as the hydrobromide, and a compound of the formula V is converted into a compound of the formula I by treatment with a base. ##STR00075##
9. A process for the preparation of the compounds of the formula I of claim 1 in which X denotes H or NH.sub.2 and Q denotes CO and I.sup.1, I.sup.2, I.sup.3, Y, Ar, T, R.sup.1, R.sup.2, R, R, R and Hal have the meanings as in claim 1, characterised in that a compound of the formula VI is converted into a compound of the formula VII by reaction with thiophosgene, a compound of the formula VII is reacted with a suitable amine under basic conditions and optionally with addition of basic reagents to give a compound of the formula VIII, and a compound of the formula VIII is reacted with hydrazine to give a compound of the formula Ia or a compound of the formula I in which X denotes NH.sub.2 and Q denotes CO and I.sup.1, I.sup.2, I.sup.3, Y, Ar, T, R.sup.1, R.sup.2, R, R, R and Hal have the meanings as in claim 1, or a compound of the formula VIII is reacted with ammonia or hydroxylamine and optionally with use of tert-butyl hydroperoxide to give a compound of the formula Ib or a compound of the formula I in which X denotes H, Q denotes CO and and I.sup.1, I.sup.2, I.sup.3, Y, Ar, T, R.sup.1, R.sup.2, R, R, R and Hal have the meanings as in claim 1 ##STR00076##
10. A process for the preparation of the compounds of the formula I of claim 1, characterised in that a) the base of a compound of the formula I is converted into one of its salts by treatment with an acid, or b) an acid of a compound of the formula I is converted into one of its salts by treatment with a base.
11. A pharmaceutical preparation comprising at least one compound according to claim 1 a physiologically acceptable salt thereof, a solvate thereof, a stereoisomer thereof or a mixture thereof in all ratios in a dosage form.
12. Pharmaceutical preparation according to claim 11 comprising further excipients and/or adjuvants.
13. Pharmaceutical preparation comprising at least one compound according to claim 1, a physiologically acceptable salt thereof, a solvate thereof, a stereoisomer thereof or a mixture thereof in all ratios, and at least one further medicament active compound.
14. Process for the preparation of a pharmaceutical preparation, characterised in that a compound according to claim 1, a physiologically acceptable salt thereof, a solvate thereof, a stereoisomer thereof or a mixture thereof in all ratios, is brought into a suitable dosage form together with a solid, liquid or semi-liquid excipient or adjuvant.
15. Medicament comprising at least one compound according to claim 1, a physiologically acceptable salt thereof, a solvate thereof, a stereoisomer thereof or a mixture thereof in all ratios, in a form for use in the treatment of physiological and/or pathophysiological states, selected from the group consisting of osteoarthritis and traumatic cartilage injuries.
16. A method for the treatment of physiological and/or pathophysiological states selected from the group consisting of osteoarthritis and traumatic cartilage injuries comprising administering to a patient in need thereof a pharmaceutical preparation according to claim 11 by intra-articular administration.
17. Set (kit) consisting of separate packs of a) an effective amount of a compound according to claim 1, a physiologically acceptable salt thereof, a solvate thereof, a stereoisomer thereof or a mixture thereof in all ratios, and b) an effective amount of a further medicament active compound.
Description
EXAMPLE 1: ILLUSTRATIVE COMPOUNDS OF THE FORMULA I
(1) TABLE-US-00002 TABLE 2 The following compounds are in accordance with the invention Stability in plasma Cath D Ret. (h/r/m) or Com- IC.sub.50 time at pH 1.2 pound Structure [nM] [ min] Method M + H or pH 7.4 A1
and physiologically acceptable salts, derivatives, solvates, prodrugs and stereoisomers thereof, including mixtures thereof in all ratios.
(2) Stable: Recovery 75% after 4 h.
(3) In order to avoid any doubt, in all cases where the chemical name of a compound according to the invention and the depiction of the chemical structure of a compound according to the invention mistakenly do not agree, the compound according to the invention is defined unambiguously by the depiction of the chemical structure.
(4) The retention times were determined:
(5) Chromolith Speed Rod RP 18e 50-4.6 mm LCMS; polar.m, 2.4 ml/min, 220 nm, buffer A 0.05% of HCOOH/H2O, buffer B 0.04% of HCOOH/ACN, 0.0-2.8 min 4%-100% of buffer B; 2.8-3.3 min 100% of buffer B 3.3-3.4 min 100%-4% of buffer B or Chromolith Speed Rod RP 18e 50-4.6 mm LCMS; polar.m, 2.4 ml/min, 220 nm, buffer A 0.05% of HCOOH/H2O, buffer B 0.04% of HCOOH/ACN, 0.0-3.0 min 5%-100% of buffer B; 3.0-3.5 min 100% of buffer B
(6) TABLE-US-00003 TABLE 3 Compound NMR data peak lists A1 1H NMR (500 MHz, DMSO-d6) ppm = 7.29-7.22 (m, 2H), 7.20- 7.13 (m, 2H), 7.02-6.95 (m, 1H), 6.87-6.82 (m, 1H), 6.70- 6.60 (m, 2H), 6.11-5.77 (m, 2H), 4.89 (p, J = 8.0, 7.6, 1H), 4.07 (s, 2H), 3.15-3.04 (m, 4H). A2 1H NMR (500 MHz, DMSO-d6) ppm = 10.54 (s, 1H), 7.92 (s, 2H), 7.33-7.26 (m, 3H), 7.26-7.17 (m, 3H), 7.12-7.06 (m, 1H), 7.06-6.99 (m, 1H), 5.00 (p, J = 7.9, 1H), 4.49 (s, 2H), 3.27- 3.15 (m, 4H). A3 1H NMR (500 MHz, DMSO-d6) ppm = 9.37-8.92 (m, 1H), 8.34 (s, 1H), 7.31-7.26 (m, 2H), 7.23-7.18 (m, 2H), 7.16 (d, J = 8.1, 1H), 7.05 (dd, J = 8.1, 2.1, 1H), 6.96 (d, J = 2.1, 1H), 4.94 (p, J = 7.9, 1H), 4.41 (s, 2H), 3.24-3.14 (m, 4H). A4 1H NMR (500 MHz, DMSO-d6) ppm = 7.31-7.25 (m, 2H), 7.22- 7.16 (m, 2H), 7.01 (t, J = 8.0, 1H), 6.79-6.73 (m, 1H), 6.64-6.58 (m, 1H), 6.42 (d, J = 160.9, 2H), 4.94-4.86 (m, 1H), 4.15 (s, 2H), 3.21-3.03 (m, 4H). A5 1H NMR (400 MHz, DMSO-d6) ppm = 8.07 (s, 2H), 7.61 (d, J = 7.6, 2H), 7.49 (t, J = 7.5, 2H), 7.44-7.35 (m, 2H), 7.35-7.14 (m, 6H), 5.04 (p, J = 7.7, 1H), 4.54 (s, 2H), 3.25 (d, J = 7.8, 4H). A6 1H NMR (500 MHz, DMSO-d6) ppm = 7.19-7.07 (m, 4H), 6.76- 6.70 (m, 1H), 6.65-6.59 (m, 2H), 6.25 (s, 2H), 5.24-5.15 (m, 1H), 4.07 (d, J = 14.0, 1H), 3.76 (d, J = 14.1, 1H), 2.86-2.77 (m, 1H), 2.76-2.68 (m, 1H), 2.05-1.99 (m, 1H), 1.97-1.88 (m, 2H), 1.83-1.71 (m, 1H). A7 1H NMR (500 MHz, DMSO-d6) ppm = 7.28-7.22 (m, 2H), 7.20- 7.12 (m, 2H), 6.74 (d, J = 7.5, 1H), 6.52-6.48 (m, 1H), 6.48- 6.44 (m, 1H), 5.88 (s, 2H), 4.89 (p, J = 7.6, 1H), 4.03 (s, 2H), 3.15-3.02 (m, 4H), 2.40 (t, J = 7.5, 2H), 1.52 (h, J = 7.4, 2H), 0.87 (t, J = 7.3, 3H). A8 1H NMR (500 MHz, DMSO-d6) ppm = 7.03-6.99 (m, 1H), 6.87 (s, 2H), 6.86-6.82 (m, 2H), 6.81-6.73 (m, 2H), 4.94-4.87 (m, 1H), 4.20 (s, 2H), 3.72 (s, 6H), 3.11-3.00 (m, 4H). A9 1H NMR (500 MHz, DMSO-d6) ppm = 8.06 (s, 2H), 7.25 (d, J = 8.2, 1H), 7.15 (dd, J = 8.1, 2.0, 1H), 7.08 (d, J = 2.0, 1H), 6.89 (s, 2H), 4.99 (p, J = 7.7, 1H), 4.45 (s, 2H), 3.73 (s, 6H), 3.19- 3.07 (m, 4H). A10 1H NMR (400 MHz, DMSO-d6) ppm = 7.29-7.23 (m, 2H), 7.20- 7.14 (m, 2H), 7.05 (d, J = 7.6, 1H), 6.95 (dd, J = 7.9, 1.8, 1H), 6.80 (d, J = 1.8, 1H), 6.24 (s, 2H), 4.87 (p, J = 7.8, 1H), 4.18 (s, 2H), 3.10 (d, J = 7.8, 4H). A11 1H NMR (400 MHz, DMSO-d6) ppm = 6.95-6.82 (m, 3H), 6.67 (dd, J = 7.9, 2.2, 1H), 6.58 (d, J = 2.1, 1H), 6.14 (s, 2H), 4.90- 4.76 (m, 1H), 4.09 (s, 2H), 3.76 (s, 3H), 3.74 (s, 3H), 3.13 (dd, J = 16.4, 8.2, 1H), 3.00 (dd, J = 16.4, 7.4, 3H). A12 1H NMR (400 MHz, DMSO-d6) ppm = 7.16 (t, J = 7.8, 1H), 6.86 (t, J = 7.7, 2H), 6.79 (d, J = 8.2, 1H), 6.66 (dd, J = 7.9, 2.2, 1H), 6.58 (d, J = 2.2, 1H), 6.14 (s, 2H), 4.87 (p, J = 7.8, 1H), 4.07 (s, 2H), 3.78 (s, 3H), 3.16-3.01 (m, 3H), 2.90 (dd, J = 16.5, 6.9, 1H). A13 1H NMR (500 MHz, DMSO-d6) ppm = 7.34-7.19 (m, 3H), 7.15- 7.11 (m, 1H), 6.98-6.89 (m, 2H), 6.84 (d, J = 1.6, 1H), 6.37 (s, 2H), 5.61 (t, J = 7.9, 1H), 4.12-4.05 (m, 1H), 3.81-3.74 (m, 1H), 3.07-2.95 (m, 1H), 2.91-2.77 (m, 1H), 2.48-2.37 (m, 1H), 2.13-1.98 (m, 1H). A14 1H NMR (500 MHz, DMSO-d6) ppm = 7.14 (d, J = 8.3, 1H), 6.87 (d, J = 7.9, 1H), 6.84 (d, J = 2.4, 1H), 6.74 (dd, J = 8.2, 2.5, 1H), 6.67 (dd, J = 7.9, 2.2, 1H), 6.58 (d, J = 2.1, 1H), 6.22 (s, 2H), 4.85 (p, J = 7.7, 1H), 4.08 (s, 2H), 3.72 (s, 3H), 3.14-2.92 (m, 4H). A15 1H NMR (500 MHz, DMSO-d6) ppm = 7.94 (d, J = 7.6, 2H), 7.56- 7.44 (m, 4H), 7.42-7.33 (m, 2H), 6.97-6.76 (m, 4H), 6.27 (s, 1H), 3.53 (s, 2H), 2.53-2.51 (m, 1H). A16 1H NMR (400 MHz, DMSO-d6) ppm = 7.19-7.11 (m, 2H), 7.10 (s, 2H), 7.06 (d, J = 7.1, 1H), 6.93-6.82 (m, 2H), 6.75 (dd, J = 8.3, 2.5, 1H), 4.89 (p, J = 7.7, 1H), 4.25 (s, 2H), 3.73 (s, 3H), 3.13- 3.01 (m, 4H). A17 1H NMR (400 MHz, DMSO-d6) ppm = 7.14 (d, J = 8.3, 1H), 6.84 (d, J = 2.4, 1H), 6.78-6.69 (m, 2H), 6.52 (dd, J = 7.5, 1.8, 1H), 6.47 (d, J = 1.7, 1H), 5.84 (s, 2H), 4.88 (s, 1H), 4.02 (s, 2H), 3.72 (s, 3H), 3.13-2.91 (m, 4H), 2.45 (q, J = 7.5, 2H), 1.12 (t, J = 7.6, 3H). A18 1H NMR (400 MHz, DMSO-d6) ppm = 7.19-7.11 (m, 2H), 7.10 (s, 2H), 7.06 (d, J = 7.1, 1H), 6.93-6.82 (m, 2H), 6.75 (dd, J = 8.3, 2.5, 1H), 4.89 (p, J = 7.7, 1H), 4.25 (s, 2H), 3.73 (s, 3H), 3.13- 3.01 (m, 4H). A19 1H NMR (400 MHz, DMSO-d6) ppm = 7.19-7.11 (m, 2H), 7.10 (s, 2H), 7.06 (d, J = 7.1, 1H), 6.93-6.82 (m, 2H), 6.75 (dd, J = 8.3, 2.5, 1H), 4.89 (p, J = 7.7, 1H), 4.25 (s, 2H), 3.73 (s, 3H), 3.13- 3.01 (m, 4H). A20 1H NMR (400 MHz, DMSO-d6) ppm = 7.14-7.03 (m, 5H), 6.97 (dd, J = 7.9, 2.0, 1H), 6.80 (d, J = 1.8, 1H), 6.16 (s, 2H), 4.34 (q, J = 14.4, 2H), 4.25-4.11 (m, 1H), 3.14-2.75 (m, 4H), 2.08-1.84 (m, 2H). A21 1H NMR (500 MHz, DMSO-d6) ppm = 7.30-7.15 (m, 5H), 7.15- 7.08 (m, 2H), 7.04-6.95 (m, 2H), 6.86 (d, J = 7.6, 1H), 6.76 (s, 1H), 6.18 (s, 2H), 4.90 (p, J = 7.7, 1H), 4.18 (s, 2H), 3.98 (t, J = 8.3, 2H), 3.12 (d, J = 7.7, 4H), 3.06 (t, J = 8.3, 2H). A22 1H NMR (500 MHz, DMSO-d6) ppm = 7.30-7.14 (m, 5H), 6.98- 6.91 (m, 2H), 6.88-6.81 (m, 2H), 6.76 (d, J = 1.6, 1H), 6.05 (s, 2H), 4.88 (p, J = 7.8, 1H), 4.80-4.72 (m, 2H), 4.70-4.61 (m, 2H), 4.15 (s, 2H), 3.77-3.69 (m, 3H), 3.11 (d, J = 7.8, 4H). A23 1H NMR (500 MHz, DMSO-d6) ppm = 7.28-7.23 (m, 2H), 7.19- 7.14 (m, 2H), 6.89 (d, J = 7.5, 1H), 6.60 (dd, J = 7.5, 1.7, 1H), 6.52 (d, J = 1.6, 1H), 6.09 (s, 2H), 4.93-4.84 (m, 1H), 4.12 (s, 2H), 3.41-3.32 (m, 2H), 3.24-3.19 (m, 2H), 3.10 (d, J = 7.8, 4H), 1.15-0.98 (m, 6H). A24 1H NMR (500 MHz, DMSO-d6) ppm = 10.57 (s, 1H), 8.04 (s, 1H), 7.31-7.20 (m, 5H), 7.06 (dd, J = 7.7, 1.5, 1H), 6.98 (d, J = 1.5, 1H), 5.00 (p, J = 7.9, 1H), 4.53 (s, 2H), 3.30-3.15 (m, 6H), 2.53-2.51 (m, 2H), 1.32-0.89 (m, 6H). A25 1H NMR (500 MHz, DMSO-d6) ppm = 7.28-7.23 (m, 2H), 7.19- 7.14 (m, 2H), 6.91 (d, J = 7.5, 1H), 6.68 (dd, J = 7.5, 1.7, 1H), 6.59 (d, J = 1.6, 1H), 6.10 (s, 2H), 4.92-4.83 (m, 1H), 4.13 (s, 2H), 3.56 (s, 4H), 3.30 (s, 4H), 3.10 (d, J = 7.8, 4H). A26 1H NMR (500 MHz, DMSO-d6) ppm = 10.59 (s, 1H), 8.04 (s, 2H), 7.33-7.19 (m, 4H), 7.13 (dd, J = 7.7, 1.5, 1H), 7.04 (d, J = 1.5, 1H), 5.00 (p, J = 7.9, 1H), 4.53 (s, 2H), 3.66-3.52 (m, 8H), 3.29-3.15 (m, 4H). A27 1H NMR (400 MHz, DMSO-d6) ppm = 8.02 (d, J = 8.1, 1H), 7.45- 7.39 (m, 1H), 7.33 (dd, J = 8.4, 1.8, 1H), 7.29 (s, 2H), 7.27-7.21 (m, 2H), 7.21-7.12 (m, 2H), 5.33-5.19 (m, 1H), 3.60 (dd, J = 15.6, 7.8, 2H), 3.23 (dd, J = 15.6, 9.5, 2H).
EXAMPLE 2: PREPARATION OF THE COMPOUNDS FORMULA I ACCORDING TO THE INVENTION IN WHICH XH AND Q=CH2
(7) The claimed compounds of the formula I in which XH and Q=CH.sub.2 can be prepared, for example, by methods known to the person skilled in the art by the following synthesis sequences. The examples indicated describe the synthesis, but do not restrict this to the examples.
(8) Synthesis Sequence:
(9) ##STR00043##
(10) Starting from substituted ortho-nitrobenzaldehydes, a substituted ortho-nitrobenzylamine is prepared by reductive amination using a suitable amine and is converted into the corresponding aniline derivative by hydrogenation in the presence of a catalyst, for example Raney nickel. If the radical R contains functional groups which are reactive in the presence of a catalyst, for example Raney nickel, and hydrogen, reduction of these units may occur (for example alkenyl is converted into alkyl) and is part of the process. Cyclisation by reaction with cyanogen bromide at elevated temperatures of 10 C. to 80 C., preferably RT to 60 C., gives the guanidine derivatives according to the invention as hydrobromides. The free guanidine derivatives are obtained therefrom by treatment with base.
(11) This sequence may be followed by further steps, such as, for example, chiral separations, oxidations, reductions, metal-catalysed reactions, protecting-group removals, amide couplings, etc., without restricting the method to these reactions.
(12) The substituted ortho-nitrobenzaldehydes required as starting materials are either commercially available or can be prepared by corresponding methods, such as, for example, Suzuki reactions, hydrolyses, hydrogenations, amide couplings.
(13) A) Process for the Preparation of the Substituted Ortho-Nitrobenzaldehydes Having Amide Functions:
(14) ##STR00044##
(15) This process enables, for example, the preparation of the following (hitherto unknown) compounds:
(16) ##STR00045##
B) Process for the Preparation of the Ortho-Nitrobenzaldehydes Containing Aryl Radicals or Alkenyl Radicals by Suzuki Reaction:
(17) ##STR00046##
(18) Ortho-nitrobenzaldehydes containing aryl radicals or alkenyl radicals can be prepared in accordance with the above equation by Suzuki reaction with suitable boronic acids. Instead of the free boronic acids, the boronic acid esters can be employed with equal success. The yields on use of SPhos as ligand and potassium phosphate as base are typically between 60 and 99%. The temperatures of the reaction are between 10 C. and 80 C., preferably between RT and 60 C., particularly preferably between 30 C. and 50 C. This process enables, for example, the preparation of the following (hitherto unknown) compounds:
(19) ##STR00047## ##STR00048##
(20) The compounds prepared in this way are reacted further as follows for the preparation of the compounds of the formula I according to the invention:
(21) ##STR00049##
(22) A substituted ortho-nitrobenzylamine is prepared by reductive amination using a suitable amine and is converted into the corresponding aniline derivative by hydrogenation in the presence of a catalyst, for example Raney nickel. In this process, the double bond of the alkenyl group is also reduced at the same time.
(23) As above, the cyclisation by reaction with cyanogen bromide at elevated temperatures gives the guanidine derivatives according to the invention as hydrobromides, from which the free guanidine derivatives are liberated by treatment with base.
EXAMPLE 3: PREPARATION OF THE COMPOUNDS OF THE FORMULA I ACCORDING TO THE INVENTION IN WHICH XH OR XNH2 AND Q=CO
(24) The claimed compounds of the formula I in which XH (formula Ib) or XNH.sub.2 (formula Ia) and Q=CO can be prepared, for example, by methods known to the person skilled in the art, as described, for example, in Bio-organic Medicinal Chemistry (2007), 4009-4015. In a modification of this method, claimed compounds of the formula I in which XH (formula Ib) or X NH.sub.2 (formula Ia) and Q=CO can be prepared by the following synthesis sequences. The examples indicated describe the synthesis, but do not restrict this to the examples.
(25) Synthesis Sequence:
(26) ##STR00050##
(27) Starting from substituted ortho-aminobenzoic acid esters, the corresponding isocyanates are prepared by reaction with thiophosgene or similar reagents. The cyclisation to give corresponding cyclic thiourea derivatives is carried out by reaction with a suitable amine under basic conditions at temperatures of 40 C. to 100 C., preferably 60 C. to 90 C., particularly preferably 75 C. to 85 C., very particularly preferably at 80 C. In some cases, the addition of basic reagents, such as, for example, potassium tert-butoxide, proves favourable. The thiourea derivatives are converted into the target compounds according to the invention (XNH.sub.2) by further reaction with hydrazine at elevated temperatures of 80 C. to 200 C., preferably 100 C. to 150 C., particularly preferably 110 C. to 130 C., for example also in the microwave. By contrast, the target compounds according to the invention (XH) are obtained by reaction with ammonia or hydroxylamine. Performance of the reaction in the presence of tert-butyl hydroperoxide proves favourable here. This sequence may be followed by further steps, such as, for example, oxidations, reductions, metal-catalysed reactions, protecting-group removals, amide couplings, etc., without restricting the method to these reactions.
(28) The substituted ortho-aminobenzoic acid esters required as starting materials are either commercially available or are prepared, for example, from the corresponding ortho-aminobenzoic acids by esterification.
EXAMPLE 4: PREPARATION OF A21 (2-AMINO-3-INDAN-2-YL-3,4-DIHYDROQUINAZOLIN-7-YL)-(2,3-DIHYDROINDOL-1-YL)METHANONE
Step 1: 4-(2,3-Dihydroindole-1-carbonyl)-2-nitrobenzaldehyde
(29) ##STR00051##
(30) 4-Formyl-3-nitrobenzoic acid (650.00 mg; 3.33 mmol; 100.00 mol %) was dissolved in N,N-dimethylformamide (20.00 ml; 257.20 mmol; 7721.20 mol %), and INDOLINE (0.37 ml; 3.33 mmol; 100.00 mol %) and ethyldiisopropylamine (578.04 l; 3.33 mmol; 100.00 mol %) were added. The reaction mixture was cooled in an ice bath, Hatu C10H15N6O*F6P (1.39 g; 3.66 mmol; 110.00 mol %) was added with stirring, and stirring was continued at RT overnight. For work-up, the reaction mixture was poured into sat. sodium hydrogencarbonate solution with stirring, stirring was continued for 30 min, the crystals formed were filtered off with suction and rinsed with water. Recrystallisation from a little EA gave 430 mg of 4-(2,3-dihydroindole-1-carbonyl)-2-nitrobenzaldehyde as beige crystals. (Yield 42%, content >97%). MS-FAB (M+H.sup.+)=297.0 R.sub.f (polar method): 2.22 min (MS track).
(31) The following compounds can be prepared analogously to this step:
(32) ##STR00052##
Step 2: (2,3-Dihydroindol-1-yl)-[4-(indan-2-ylaminomethyl)-3-nitrophenyl]-methanone
(33) ##STR00053##
(34) 4-(2,3-Dihydroindole-1-carbonyl)-2-nitrobenzaldehyde (430.00 mg; 1.41 mmol; 100.00 mol %) was dissolved in 1,2-dichloroethane (10.00 ml; 126.31 mmol; 8963.18 mol %) with 2-aminoindane (262.78 mg; 1.97 mmol; 140.00 mol %), glacial acetic acid (81.41 l; 1.41 mmol; 100.00 mol %) and sodium triacetoxyborohydride, 95% (418.15 mg; 1.97 mmol; 140.00 mol %) were added, and the mixture was stirred at RT overnight. Sodium triacetoxyborohydride, 95% (40.00 mg; 0.19 mmol; 13.39 mol %) was again added, and the mixture was stirred at RT for 3 h. Water/dichloromethane was added to the reaction mixture, the aqueous phase was extracted 1 with dichloromethane, the org. phase was washed 1 with sat. NaCl solution, dried over sodium sulfate, filtered and evaporated. Purification of the crude product by column chromatography on a CombiflashRf unit (120 g RediSep silica column, 60 ml/min of heptane/ethyl acetate 5-100% of EA in 25 min) gave 330 mg of (2,3-dihydroindol-1-yl)-[4-(indan-2-ylaminomethyl)-3-nitrophenyl]-methanone as beige crystals.
(35) (Yield 56.6%, content 100%). MS-FAB (M+H+)=414.5 Rf (polar method): 1.80 min (MS track).
Step 3: [3-Amino-4-(indan-2-ylaminomethyl)phenyl]-(2,3-dihydroindol-1-yl)methanone
(36) ##STR00054##
(37) 330 mg of (2,3-dihydroindol-1-yl)-[4-(indan-2-ylaminomethyl)-3-nitrophenyl]-methanone were reduced in the presence of 300 mg of sponge nickel (water-wet) in 10 ml of THF at atmospheric pressure and room temperature using hydrogen overnight. Removal of the solvent gave 267 mg of [3-amino-4-(indan-2-ylaminomethyl)phenyl]-(2,3-dihydroindol-1-yl)methanone as wax-like solid.
(38) (Yield 71.7%, content 82.2%). MS-FAB (M+H+)=384.6 Rf (polar method): 1.74 min (MS track).
Step 4: [3-Amino-4-(indan-2-ylaminomethyl)phenyl]-(2,3-dihydroindol-1-yl)methanone
(39) Error! Objects Cannot be Created from Editing Field Codes.
(40) 3-Amino-4-(indan-2-ylaminomethyl)phenyl]-(2,3-dihydroindol-1-yl)methanone (267.00 mg; 0.70 mmol; 100.00 mol %) was dissolved in 1,4-dioxane (max. 0.005% of H2O) SeccoSolv (6.00 ml; 70.14 mmol; 10074.56 mol %), cyanogen bromide (81.12 mg; 0.77 mmol; 110.00 mol %) was added with stirring, and the mixture was stirred at 80 C. for 3 hours. The suspension was diluted with 1,4-dioxane and refluxed at 120 C. for a further 5 hours. Cyanogen bromide (20.00 mg; 0.19 mmol; 27.12 mol %) was again added, and the mixture was refluxed at 120 C. for 3 hours.
(41) The crystals which precipitated out on cooling were filtered off with suction, treated with 2 N NaOH and taken up in EA. The EA phase was washed 1 with sat. NaCl solution, dried over sodium sulfate, filtered, evaporated. Trituration of the residue with a little acetonitrile and removal of the solvent by suction filtration gave 72 mg of [3-amino-4-(indan-2-ylaminomethyl)phenyl]-(2,3-dihydroindol-1-yl)methanone as white crystals.
(42) (Yield 24.1%, content 95.2%). MS-FAB (M+H+)=409.5 Rf (polar method): 1.91 min (MS track).
(43) Compounds A1, A3, A4, A6, A8-A16, A20-23, A25, A26, A29-31 can be prepared by this method (data see Excel data sheet)
EXAMPLE 5: PREPARATION OF A9 (7-CHLORO-3-(5,6-DIMETHOXYINDAN-2-YL)-3,4-DIHYDROQUINAZOLIN-2-YLAMINE HYDROBROMIDE)
(44) ##STR00055##
(45) (2-Amino-4-chlorobenzyl)-(5,6-dimethoxyindan-2-yl)amine (580.00 mg; 1.74 mmol; 100.00 mol %) was dissolved in 10 ml of 1,4-dioxane, cyanogen bromide for synthesis (203.04 mg; 1.92 mmol; 110.00 mol %) was added with stirring, and the mixture was refluxed for 3 h. The crystals which precipitated out on cooling were filtered off with suction, rinsed with dioxane and dried in a lyophiliser (white crystals, content 100%). MS-FAB (M+H+)=358.1 Rf (polar method): 1.73 min (MS track).
(46) Compounds A2, A9, A24 and A26 were prepared by this method.
EXAMPLE 6: PREPARATION OF A7 (3-INDAN-2-YL-7-PROPYL-3,4-DIHYDROQUINAZOLIN-2-YLAMINE)
Step 1: 2-Nitro-4-propenylbenzaldehyde by Suzuki Reaction
(47) ##STR00056##
(48) 4-Chloro-2-nitrobenzaldehyde (3.200 g; 16.382 mmol; 100.00 mol %), trans-propenylboronic acid (1.610 g; 18.556 mmol; 113.27 mol %) and tripotassium phosphate monohydrate (11.913 g; 49.147 mmol; 300.00 mol %) (mortared) were suspended in 15 ml of tetrahydrofuran and 150.000 l of water in a 100 ml two-necked flask. The suspension was degassed in vacuo, blanketed with argon, and palladium(II) acetate (47% of Pd) for synthesis (18.390 mg; 0.082 mmol; 0.50 mol %) and 2-dicyclohexylphosphino-2,6-dimethoxybiphenyl (34.667 mg; 0.082 mmol; 0.50 mol %) were added in a counterstream of argon. The reaction mixture was stirred at 40 C. and under an argon atmosphere for a few hours. After the reaction, the mixture was filtered, and the filtrate was evaporated to dryness. Chromatographic purification of the residue on silica gel (eluent heptane/EA 11:1) gave 3.000 g of 2-nitro-4-propenyl-benzaldehyde (yield 95.8%, content 100%). MS-FAB (M+H.sup.+)=192.0 R.sub.f (polar method): 2.28 min (MS track).
(49) The following compounds can be prepared analogously to this step:
(50) ##STR00057## ##STR00058##
Step 2: Indan-2-yl-[2-nitro-4-((E)-propenyl)benzyl]amine
(51) ##STR00059##
(52) 2-Nitro-4-propenylbenzaldehyde (1.000 g; 0.005 mol; 100.00 mol %) and 2-aminoindane (0.949 ml; 0.007 mol; 140.00 mol %) were dissolved in 12 ml of 1,2-dichloroethane in a 100 ml one-necked flask, and 0.302 ml of acetic acid (glacial acetic acid) was added. 95% sodium triacetoxyborohydride (1.634 g; 0.007 mol; 140.00 mol %) was added with stirring, and the reaction mixture was stirred at RT overnight. After the reaction, aqueous, saturated NaHCO.sub.3 solution was added, and the mixture was diluted with DCM. The organic phase was separated off, dried over Na.sub.2SO.sub.4, and the solvent is removed in vacuo. Chromatographic purification of the residue on silica gel (eluent heptane/EA 3:1) gave 1.401 g of indan-2-yl-[2-nitro-4-((E)-propenyl)benzyl]amine (yield 86.8%, content 100%). MS-FAB (M+H.sup.+)=309.1 Rf (polar method): 1.83 min (MS track).
Step 3: (2-Amino-4-propylbenzyl)indan-2-ylamine
(53) ##STR00060##
(54) A solution of 1.200 g of indan-2-yl-[2-nitro-4-((E)-propenyl)benzyl]amine in 20 ml of THF was hydrogenated using hydrogen at RT and atmospheric pressure in the presence of 0.200 g of sponge nickel (water-wet) overnight. The solution was filtered off, and the solvent was removed, giving 1.001 g of (2-amino-4-propylbenzyl)indan-2-ylamine as colourless oil (yield: 90.7%, content 99.0%). MS-FAB (M+H.sup.+)=281.1 Rf (polar method): 1.82 min (MS track).
Step 4: Cyclisation to give 3-indan-2-yl-7-propyl-3,4-dihydrominazolin-2-ylamine
(55) ##STR00061##
(56) Cyanogen bromide (0.227 ml; 0.004 mol; 120.00 mol %), dissolved in dioxane, was added to a solution of 2-amino-4-propylbenzyl)indan-2-ylamine (1.000 g; 0.004 mol; 100.00 mol %) in 20 ml of 1,4-dioxane in a 100 ml two-necked flask with stirring, and the mixture was stirred at 80 C. for 4 hours. After the reaction, the reaction mixture was cooled in an ice bath, the precipitate formed was filtered off with suction and suspended in 2N NaOH solution. Suction filtration and drying gave 0.730 g of 3-indan-2-yl-7-propyl-3,4-dihydroquinazolin-2-ylamine as white solid (yield 67.7%, content 100%). MS-FAB (M+H.sup.+)=306.2 Rf (polar method): 1.96 min (MS track).
(57) Compounds A5, A7 and A17 can be prepared analogously to this example.
EXAMPLE 7: PREPARATION OF A28 (2-HYDRAZINO-3-INDAN-2-YL-7-TRIFLUOROMETHYL-3H-QUINAZOLIN-4-ONE)
Step 1: Methyl 2-isothiocyanato-4-trifluoromethylbenzoate
(58) ##STR00062##
(59) Thiophosgene (1.595 ml; 20.186 mmol; 200.00 mol %) was added dropwise to a mixture of methyl 2-amino-4-trifluoromethylbenzoate (2.212 g; 10.093 mmol; 100.00 mol %) in 20 ml of dichloromethane and 20 ml of NaHCO.sub.3 solution with ice-cooling. The reaction mixture was slowly warmed to 35 C., stirred for 4-5 hours, 1 further equivalent and about 10 ml of NaHCO.sub.3 solution were added, the mixture was stirred at 35 C. overnight, 1 equivalent of thiophosgene and 10 ml of NaHCO.sub.3 solution were again added, and the mixture was stirred for a further 2 hours. The phases were separated, and the aqueous phase was extracted three times with DCM. The combined organic phases were dried over Na.sub.2SO.sub.4, the solvent was removed. Chromatographic purification of the residue on silica gel (eluent heptane/EA 5:1) gave 2.40 g of methyl 2-isothiocyanato-4-trifluoromethylbenzoate as yellow solid (yield 91.1%, content 100%). MS-FAB (M31).sup.+=230.0 Rf (polar method): 2.71 min (MS track).
Step 2: 3-Indan-2-yl-2-thioxo-7-trifluoromethyl-2,3-dihydro-1H-quinazolin-4-one
(60) ##STR00063##
(61) Methyl 2-isothiocyanato-4-trifluoromethylbenzoate (2.397 g; 9.176 mmol; 100.00 mol %) was dissolved in N,N-dimethylformamide for synthesis (20.000 ml; 0.257 mol), 2-aminoindane (1.372 g; 10.094 mmol; 110.00 mol %) was added, and the mixture was stirred at 80 C. overnight. Addition of water resulted in a precipitate, which was filtered off with suction and, after drying, gave 3.30 g of 3-indan-2-yl-2-thioxo-7-trifluoromethyl-2,3-dihydro-1H-quinazolin-4-one as brownish solid (yield 99.2%, content 100%). MS-FAB (M+H.sup.+)=363.0 Rf (polar method): 2.74 min (MS track).
Step 3: 2-Hydrazino-3-indan-2-yl-7-trifluoromethyl-3H-quinazolin-4-one
(62) ##STR00064##
(63) A solution of 3-indan-2-yl-2-thioxo-7-trifluoromethyl-2,3-dihydro-1H-quinazolin-4-one (350.000 mg; 0.966 mmol; 100.00 mol %) and hydrazine (0.316 ml; 9.659 mmol; 1000.00 mol %) was stirred at 120 C. in the microwave for 45 min. 10 in tert-butanol. Water was added to the reaction mixture, and formed was filtered off with suction, washed with water and dried. Chromatographic purification (reversed phase, eluent 5%-65% of ACN, 20 min) gave, after drying, 43 mg of 4 2-hydrazino-3-indan-2-yl-7-trifluoromethyl-3H-quinazolin-4-one as white powder. (Yield 12.0%, content 97%). MS-FAB (M+H.sup.+)=361.1 Rf (polar method): 2.00 min (MS track).
EXAMPLE 8: PREPARATION OF 7-BROMO-3-INDAN-2-YL-2-THIOXO-2,3-DIHYDRO-1H-QUINAZOLIN-4-ONE
Step 1: Methyl 4-bromo-2-isothiocyanatobenzoate
(64) ##STR00065##
(65) Methyl 2-amino-4-bromobenzoate (5.000 g; 21.734 mmol; 100.00 mol %) was dissolved in 50 ml of dichloromethane, and 50 ml of NaHCO.sub.3 solution were added. Thiophosgene (3.435 ml; 43.467 mmol; 200.00 mol %) was added at 0 C. with stirring, the mixture was stirred for 20 min., then slowly warmed to the RT. The reaction mixture was stirred at 35 C. overnight, then 1 equivalent of thiophosgene was added, the mixture was stirred at 35 C. for 3 hours, a further 1 equivalent of thiophosgene and 10 ml of NaHCO3 solution were added, and the mixture was stirred again for a further 4.5 hours. After phase separation, the aqueous phase was extracted a further three times with DCM, the combined organic phases were washed with NaHCO.sub.3 and dried over MgSO4. Removal of the solvent and chromatographic purification on silica gel (eluent heptane/EA 3:1) gave 3.6 g of methyl 4-bromo-2-isothiocyanatobenzoate as white powder. (Yield 61.5%, content 100%). MS-FAB (M31).sup.+=241.8/239.8 Rf (polar method): 2.74 min (MS track).
Step 2: 7-Bromo-3-indan-2-yl-2-thioxo-2,3-dihydro-1H-quinazolin-4-one
(66) ##STR00066##
(67) A solution of methyl 4-bromo-2-isothiocyanatobenzoate (2.00 g; 7.35 mmol; 100.00 mol %) and 2-aminoindane (0.99 g; 7.35 mmol; 100.00 mol %) in 25 ml of DMF was stirred at 80 C. overnight. Potassium tert-butoxide for synthesis (0.42 g; 3.68 mmol; 50.00 mol %) was added to the reaction mixture, and stirring was continued at 80 C. until the reaction was complete. After cooling, the mixture was poured into water, and the solid formed was filtered off with suction. Drying gave 2.68 g of 7-bromo-3-indan-2-yl-2-thioxo-2,3-dihydro-1H-quinazolin-4-one (yield 97.7%, content 100%). MS-FAB (M30)=373.0/375.0 Rf (polar method): 2.69 min (MS track).
(68) This compound can be converted into the corresponding hydrazine derivative analogously to Example 6.
EXAMPLE 9: PREPARATION OF A27 (2-AMINO-3-INDAN-2-YL-7-TRIFLUOROMETHYL-3H-QUINAZOLIN-4-ONE)
(69) ##STR00067##
(70) A solution of 3-indan-2-yl-2-thioxo-7-trifluoromethyl-2,3-dihydro-1H-quinazolin-4-one (350.000 mg; 0.966 mmol; 100.00 mol %), hydroxylamine (50% SOLUTION IN WATER, 3.000 ml; 50.863 mmol; 5266.05 mol %) and tert-butyl hydroperoxide (3.233 g; 25.112 mmol; 2600.00 mol %) in 5 ml of 2-propanol was stirred at RT overnight. The mixture was poured into water, and the solid formed was filtered off with suction. Chromatographic purification on silica gel (eluent heptane/EA 3:2) gave 211 mg of 2-amino-3-indan-2-yl-7-trifluoromethyl-3H-quinazolin-4-one as yellow solid (yield 61.4%, content 97%). MS-FAB (M+H.sup.+)=346.0 Rf (polar method): 2.25 min (MS track).
EXAMPLE 10: PREPARATION OF A18 (3-((S)-5-METHOXYINDAN-2-YL)-7-TRIFLUOROMETHYL-3,4-DIHYDROQUINAZOLIN-2-YLAMINE) AND A19 (3-((R)-5-METHOXYINDAN-2-YL)-7-TRIFLUOROMETHYL-3,4-DIHYDROQUINAZOLIN-2-YLAMINE) BY CHIRAL SEPARATION
(71) ##STR00068##
(72) 80 mg of racemic 3-(5-methoxyindan-2-yl)-7-trifluoromethyl-3,4-dihydroquinazolin-2-ylamine were dissolved in 2 ml of methanol and separated into the corresponding enantiomers on an SFC unit (40 runs of 50 l). Stationary phase: ChiralCel OD-H, eluent CO2, methanol DEA 0.5 (30%). 31 mg of 3-((S)5-methoxyindan-2-yl)-7-trifluoromethyl-3,4-di hydroquinazolin-2-ylamine and 31 mg of 3-((R)5-methoxyindan-2-yl)-7-trifluoromethyl-3,4-dihydroquinazolin-2-ylamine were obtained.
(73) MS-FAB (M+H.sup.+)=362.1 Rf (polar method): 1.86 min (MS track). A18 Rf (ChiralCel OD-H, eluent CO2, methanol DEA 0.5 (30%): 3.90 min. A19 Rf (ChiralCel OD-H, eluent CO2, methanol DEA 0.5 (30%): 7.09 min. The absolute configuration of the enantiomers is not known and has been assigned arbitrarily.
(74) Abbreviations:
(75) DCM=dichloromethane
(76) DMA=dimethylacetamide
(77) DMF=dimethylformamide
(78) EA=ethyl acetate
(79) h=hours MTBE=methyl tert-butyl ether
(80) PE=petroleum ether
(81) RT=room temperature
(82) SPhos=2-dicyclohexylphosphino-2,6-dimethoxybiphenyl
(83) TFA=trifluoroacetic acid
EXAMPLE 11: IN-VITRO FLUORESCENCE ASSAY FOR IDENTIFICATION OF CATHEPSIN D INHIBITORS
(84) In order to identify modulators of cathepsin D activity, a continuous enzymatic test was carried out with a synthetic peptide which carries a fluorescent group (MCA=(7-methoxycoumarin-4-yl)acetyl) which is quenched by energy transfer from a Dpn (2,4 dinitrophenyl) group on the same molecule, in Greiner 384-well nb microtitre plates. Cleavage of the peptidic substrate by cathepsin D causes an increase in the fluorescence intensity. In order to determine the efficacy of substances, the time-dependent increase in the fluorescence intensity in the presence of the substance was compared with the time-dependent increase in fluorescence in the absence of substances. The reference substance used was pepstatin A (Sigma-Aldrich). The substrate used was MCA-GKPILFFRLK(Dnp)d-RNH.sub.2 (Enzo Life Sciences, Lrach). The enzyme employed was cathepsin D isolated from the human liver (Sigma-Aldrich) in a final concentration of 1.4 nM. The test was carried out in 100 mM sodium acetate buffer, 1.25% (v/v) of DMSO, 0.25% (w/v) of Chaps, pH 5.5. 2 l of each substance solution with serially diluted substance concentration were added to in each case 4 l of cathepsin D solution and incubated at room temperature for 10 min. The reaction was started by addition of 2 l of substrate solution (final concentration 5 M). After carrying out a starting-point fluorescence measurement (excitation wavelength 340 nm/emission wavelength 450 nm) using an Envision multilabel reader (Perkin Elmer), the reaction was incubated at room temperature for 60 min. The amount of peptide fragment cleaved off during the reaction time was subsequently measured by determination of the increase in the fluorescence intensity at 450 nm (excitation wavelength 340 nm).
(85) The IC.sub.50 values of the compounds according to the invention can be obtained from Table 2 from Example 1.
EXAMPLE 12: CARTILAGE EXPLANT ASSAY
(86) In order to investigate the effect of potential cathepsin D inhibitors on cartilage degradation, a pH-induced model based on bovine explants is used. The pH of the medium in which the explants are cultivated is matched here to the pathophysiological pH of an arthrotic knee. This pH is pH 5.5. In this ex vivo model, potential cathepsin D inhibitors are subsequently investigated for their action with respect to stopping of the cartilage degradation process. If the cartilage is destroyed, glycosaminoglycans (GAGs) are released into the cell culture supernatant. The amount of GAGs liberated can be determined quantitatively with the aid of DMMB (dimethylmethylene blue hydrochloride). If sulfated GAGs are detected using dimethylmethylene blue hydrochloride, the decrease in the absorption at 633 nm is utilised. Since work can also be carried out at very low GAG concentrations, a dye/GAG complex does not precipitate out even after extended incubation of DMMB with GAG, which sometimes happens after only a short time in other measurement methods. In order to determine the concentration, a calibration line is also recorded using chondroitin sulfate. The GAG values can be used to calculate an IC.sub.50 value, i.e. a concentration at which a substance exhibits 50% of its action.
(87) Solutions:
(88) Incubation Medium, pH 7.4:
(89) DMEM without FBS, addition of 1% of Pen/Strep and 30 g/ml of ascorbic acid, the medium is not stored.
(90) Incubation Medium, pH 5.5:
(91) DMEM without FBS, the pH is adjusted by addition of MES and monitored using a pH meter, addition of 1% of Pen/Strep and 30 g/ml of ascorbic acid.
(92) Solutions for the GAG Measurement:
(93) DMMB Colouring Solution (V=500 ml):
(94) Dissolve 8 mg of DMMB (dimethylmethylene blue) in 2.5 ml of ethanol+1 g of sodium formate+1 ml of formic acid, make up to 500 ml with bidistilled water.
(95) Incubation Medium:
(96) FBS (Medium without FBS)
(97) Chondroitin Sulfate Solutions (Standard Curve)
(98) Preparation of standard solutions with the following concentrations: 50 g/ml; 25 g/ml; 12.5 g/ml; 6.25 g/ml; 3.125 g/ml; 1.56 g/ml; 0.78 g/ml and a blank control of the medium. The preparation of the standard solution is carried out in the medium with which the experiment was also carried out.
(99) 1.) Procedure: pH-Induced Cartilage Degradation of Bovine Explants
(100) The bovine explants are firstly prepared. The induction of the cartilage degradation is carried out in 96-multiwell plates. One explant is cultivated per well. In each case, 200 l of DMEM (incubation medium pH 5.5) without FBS+30 g/ml of ascorbic acid are added. Thus negative control, explants (n=4) are incubated at pH 7.4 (without FBS). This control is not included in the calculation of the data, but instead ensures that the pH change has the desired effect on the liberation of GAG. At this point, the substances to be tested are added. No pre-incubation of the explants is carried out. The explants are cultivated with the corresponding substances for 3 days in the incubator at 37 C. and 7.5% CO.sub.2.
(101) 2.) Incubation Procedure
(102) In order to investigate the effect of cathepsin D inhibitors on the liberation of GAG (glycosaminoglycan), the substances are employed in the desired concentration and cultivated for 3 days. The compounds to be tested are tested in a first experiment in a concentration of 1 M and 1% of DMSO. Substances which have an effect of >50% on the liberation of GAG (this corresponds to <50% of the control in the Assay Explorer) are tested in the next experiment at 100 nM and 1% of DMSO. Substances which have an effect of >50% on the liberation of GAG under these conditions (this corresponds to <50% of the control in the Assay Explorer) are tested in a concentration/effect relationship. The compounds here are investigated in the following concentrations: 30 M, 10 M, 3 M, 1 M, 0.30 M, 0.10 M, 0.030 M, 0.01 M.
(103) The positive control used is pepstatin A with a concentration of 0.01 M. The assay window is defined by the control (pH 5.5), defined as 0% effect, and the control pH 5.5+0.01 M pepstatin A, defined as 100% effect. After incubation for 3 days, the cell culture supernatants are collected and stored at 20 C. or measured directly. The amount of liberated GAG is measured photometrically.
(104) The effect (1 value) of the respective substance in % based on the positive control (pH 5.5+0.010 pepstatin A) and the negative control (pH 5.5) is reported for concentrations of 1 M and 100 nM. The value represents the average of 4 replicants. In the determination of a concentration/effect relationship, an IC.sub.50 value is reported to the database (Assay Explorer).
(105) 4.) Measurement
(106) The cell culture supernatants (200 l) are either measured directly or stored at 20 C. In order to ensure an accurate determination of the concentration (g/ml of GAG in the supernatant) of GAG, the measurement values must be located in the linear region of the standard curve. In order to ensure this, various dilutions are routinely introduced (1/5, 1/10, 1/20, 1/40). The dilutions are prepared with medium and introduced automatically (Hamilton) into a 384-well plate (15 l). 60 l of DMMB solution are likewise added automatically (or using a multichannel pipette). A rapid colour reaction occurs, which is subsequently measured at 633 nm using a plate reader (for example Envision).
(107) Depending on the amount of sample present, at least one double determination is carried out.
(108) The data are provided by the MTP reader as csv or xls files and stored as raw data based on this format (xls) or used for the calculation of the percentage effect of the particular compound.
(109) 5.) Quality Controls
(110) As control for the induction of the pH-induced cartilage degradation, 4 explants are incubated at pH 7.4. This corresponds to the physiological pH of the cartilage, and no effect on the liberation of GAG is thus expected here. These GAG values (g/ml of supernatant) are thus always significantly lower than the GAG values for incucation at pH 5.5.
(111) A further control, which both serves for checking of the experiment, but is also important for the definition of the assay window, is the pepstatin control (pH 5.5+0.01 M pepstatin A). This substance non-specifically blocks the activity of most proteases and thus determines the maximum possible effect of a compound.
(112) 6.) Results
(113) All compounds measured exhibited an IC.sub.50 value of 10.sup.8 to 10.sup.10 M in the GAG assay. (1) Klompmakers, A. & Hendriks, T. (1986) Anal. Biochem. 153, 80-84, Spectrophotometric Determination of Sulfated Glycosaminoglycans. (2) Groves, P. J. et al. (1997) Anal. Biochem. 245, 247-248
(114) Polyvinyl alcohol-stabilised binding of sulfated GAGs to dimethylmethylene blue.
EXAMPLE 13: INVESTIGATION OF THE ANTI-HYPERALGESIC EFFECT IN ANIMALS
(115) In order to induce an inflammation reaction, a carrageenan solution (CAR, 1%, 50 l) was injected intra-articularly on one side into a rat knee joint. The uninjected side was used for control purposes. Six animals per group were used. The threshold was determined by means of a micrometer screw (medial-lateral on the knee joint), and the thermal hyperalgesia was determined by means of a directed infrared light source by the Hargreaves method (Hargreaves et al., 1988) on the sole of the foot. Since the site of inflammation (knee joint) is different from the site of measurement (paw sole), use is made here of the term secondary thermal hyperalgesia, the mechanism of which is of importance for the discovery of effective analgesics.
(116) Experimental description of thermal hyperalgesia (Hargreaves test): the experimental animal is placed in a plastic chamber on a quartz sheet. Before testing, the experimental animal is firstly given about 5-15 minutes time to familiarise itself with the environment. As soon as the experimental animal no longer moves so frequently after the familiarisation phase (end of the exploration phase), the infrared light source, whose focus is in the plane of the glass bottom, is positioned directly beneath the rear paw to be stimulated. An experiment run is then started by pressing the button: infrared light results in an increase in the skin temperature of the rear paw. The experiment is terminated either by the experimental animal raising the rear paw (as an expression of the pain threshold being reached) or by automatic switching-off of the infrared light source when a prespecified maximum temperature has been reached. Light reflected by the paw is recorded as long as the experimental animal sits still. Withdrawal of the paw interrupts this reflection, after which the infrared light source is switched off and the time from switching on to switching off is recorded. The instrument is calibrated in such a way that the infrared light source increases the skin temperature to about 45 degrees Celsius in 10 s (Hargreaves et al. 1988). An instrument produced by Ugo Basile for this purpose is used for the testing.
(117) CAR was purchased from Sigma-Aldrich. Administration of the specific cathepsin D inhibitors according to the invention was carried out intra-articularly 30 minutes before the CAR. Triamcinolone (TAC) in an amount of 10 g/joint was used as positive control, and the solvent (vehicle) was used as negative control. The hyperalgesia is quoted as the difference in the withdrawal times between the inflamed and non-inflamed paw.
(118) Result: TAC was capable of reducing the CAR-induced swelling, but the specific cathepsin D inhibitors according to the invention were not. In contrast, the specific cathepsin D inhibitors according to the invention were able to reduce the extent of thermal hyperalgesia as a function of the dose. Assessment: it has been shown that the compounds of the present invention exert an anti-hyperalgesic action. This can be postulated since the compounds exhibited no influence on inflammatory swelling and thus on the hyperalgesia trigger. It can thus be assumed that the compounds develop a pain-reducing action in humans.
EXAMPLE 14: STABILITY OF THE COMPOUNDS ACCORDING TO THE INVENTION IN BOVINE SYNOVIAL FLUID
1.) Extraction of Bovine Synovial Fluid
(119) In the preparation of bovine explants (for the diffusion chamber or other assays), either cow hoof (metacarpal joints) or cow knee is used. The synovial fluid can be obtained from both joints. To this end, the synovial fluid is carefully removed from the open joint using a 10 ml syringe and a cannula and transferred into prepared 2 ml Eppendorf vessels. The Eppendorf vessels are labelled depending on the animal (cow passport is available). It must be ensured here that blood does not enter the joint gap during preparation of the joints. If this is the case, the synovial fluid will become a reddish colour and must consequently be discarded. The synovial fluid is basically highly viscous and clear to yellowish in colour. The removal together with a macroscopic analysis of the synovial fluid is documented.
2.) Batch for Stability Testing of Substances in SF
(120) In order to check the stability of individual compounds, a pool of four different bovine synovial fluids is mixed. To this end, about 1 ml per SF is used. The mixture is prepared directly in a 5 ml glass vessel. The SFs are mixed thoroughly, but carefully. No air bubbles or foam should form. To this end, a vortex unit is used at the lowest speed. The compounds to be tested are tested in an initial concentration (unless required otherwise) of 1 M. After addition of the substance, the batch is again mixed thoroughly and carefully. For visual monitoring, all SF batches are photographed, and the pictures are filed in the eLabBio file for the corresponding experiment. Figure 1 shows photodocumentation of this type by way of example. The batches are incubated in the incubator for 48 h at 37 C. and 7.5% CO.sub.2.
3.) Sampling
(121) The sampling is carried out after the pre-agreed times (unless required otherwise, see below). 200 l of the SF are removed from the mixture per time and transferred directly into a 0.5 ml low-binding Eppendorf vessel. Low-binding Eppendorf vessels are used in order to minimise interaction of the substances with the plastic of the vessels. 200 l of acetonitrile have already been introduced into the Eppendorf vessel, so that a 1+1 mixture of the SF forms thereafter. This simplifies the subsequent analysis, but precipitation of protein may occur immediately after addition of the SF. This should be noted on the protocol. The 0 h sample is taken immediately after addition of the substance. This corresponds to the 100% value in the stability calculation. Ideally, the concentration employed should be retrieved here. The samples can be frozen at 20 C. 0 h 6 h 24 h 48 h
(122) The negative control used is SF without substance. The positive control used is SF with 1 M of substance. This corresponds to the 0 h value and thus 100% stability.
(123) The samples are stored in low-binding Eppendorf vessels at 20 C. The samples are subsequently measured quantitatively.
4.) Data Processing
(124) The concentrations measured (ng/ml) are plotted against the time in a graph (GraphPad Prism). The percentage stability of the substance is determined here. The 100% value used is the initial value in the SF at time 0 h. The data are stored in eLabBio under the respective experiment number and reported in the MSR database (as percent stability after the corresponding incubation times).
5.) Results
(125) All compounds measured remained stable.
EXAMPLE 15: IN-VITRO FLUORESCENCE ASSAY FOR IDENTIFICATION OF RENIN-INHIBITORY ACTIVITY
(126) In order to identify modulators of renin activity, a continuous enzymatic test was carried out with a synthetic peptide which carries a fluorescent group Edans (=(5-(aminoethyl)aminonaphthalenesulfonate) which is quenched by energy transfer from a Dabcyl (4-dimethylaminoazobenzene-4-carboxylate) group on the same molecule, in Greiner 384-well microtitre plates. Cleavage of the peptidic substrate by renin causes an increase in the fluorescence intensity. In order to determine the efficacy of substances, the time-dependent increase in the fluorescence intensity in the presence of the substance was compared with the time-dependent increase in fluorescence in the absence of substances. The reference substance used was renin inhibitor 2 (Z-Arg-Arg-Pro-Phe-His-Sta-Ile-His N-Boc-Lys methyl ester Z) (Sigma-Aldrich). The substrate used was renin FRET substrate I (DABCYL-g-Abu-Ile-His-Pro-Phe-His-Leu-Val-Ile-His-Thr-EDANS) (Anaspec, Fremont Calif.). The enzyme employed was recombinant human renin (Proteos, Kalamazoo, Mich.) in a final concentration of 10 nM. The test was carried out in 50 mM Mops buffer, 1.5% (v/v) of DMSO, 0.1% (w/v) of Igepal, pH 7.2, 0.5% (w/v) of BSA. 2 l of each substance solution with serially diluted substance concentration were added to in each case 4 l of renin solution and incubated at room temperature for 15 min. The reaction was started by addition of 4 l of substrate solution (final concentration 5 M). After carrying out a starting-point fluorescence measurement (excitation wavelength 340 nm/emission wavelength 495 nm) using an Envision multilabel reader (Perkin Elmer), the reaction was incubated at 37 C. for 60 min. The amount of peptide fragment cleaved off during the reaction time was subsequently measured by determination of the increase in the fluorescence intensity at 495 nm (excitation wavelength 340 nm).
(127) Result: all compounds measured have an IC.sub.50 of the renin selectivity of >301.1M.
EXAMPLE 16: INJECTION VIALS
(128) A solution of 100 g of a compound of the formula I and 5 g of disodium hydrogenphosphate in 3 l of bidistilled water is adjusted to pH 6.5 using 2 N hydrochloric acid, filtered under sterile conditions, transferred into injection vials, lyophilised under sterile conditions and sealed under sterile conditions. Each injection vial contains 5 mg of a compound of the formula I.
EXAMPLE 17: SOLUTION
(129) A solution is prepared from 1 g of a compound of the formula I, 9.38 g of NaH.sub.2PO.sub.4 2H.sub.2O, 28.48 g of Na.sub.2HPO.sub.4.12H.sub.2O and 0.1 g of benzalkonium chloride in 940 ml of bidistilled water. The pH is adjusted to 6.8, and the solution is made up to 1 l and sterilised by irradiation. This solution can be used in the form of eye drops.
EXAMPLE 18: OINTMENT
(130) 500 mg of a compound of the formula I are mixed with 99.5 g of Vaseline under aseptic conditions.
EXAMPLE 19: AMPOULES
(131) A solution of 1 kg of a compound of the formula I in 60 l of bidistilled water is filtered under sterile conditions, transferred into ampoules, lyophilised under sterile conditions and sealed under sterile conditions. Each ampoule contains 10 mg of a compound of the formula I.