HDAC6 inhibitors, with improved solubility and their uses

Abstract

The present invention relates to small molecule compounds and their use as HDAC inhibitors and their use in the treatment of various diseases, such as cancer. The present invention further relates to methods for improvement of solubility by introducing basic substituents which offer the opportunity to create pharmaceutically acceptable salts. Moreover, it comprises methods of synthesizing the compounds and methods of treatment.

Claims

1. A compound having the general formula I
Y—[CH.sub.2].sub.p-Q-H-L-(HAm)  (formula I) wherein H is ##STR00126## wherein L is a linker having the formula ##STR00127## wherein (HAm) is hydroxamic acid with the formula ##STR00128## or a carbamate-protected hydroxyamic acid; wherein A is CH.sub.2, C═O or C═S; B is CH.sub.2, C═O or C═S; n is 0 or 1; p is 0 to 6; Q is selected from —CH.sub.2—, O, NH, alkylamino, an ester, and an amide-group; R.sub.1 to R.sub.3 are each independently selected from the group consisting of hydrogen, branched or unbranched alkyl selected from C.sub.1 to C.sub.6, and aryl; X is a branched or unbranched alkyl selected from C.sub.1 to C.sub.6; Y is selected from the group consisting of amino; cyclic alkylamino; dialkylamino; cyclic diaminoalkyl; heterocyclic alkylamino; amino acid substituents connected either via their α-amino group or their carboxy group; and pharmaceutically acceptable salts thereof; Z is carbon, nitrogen or oxygen; with the proviso that when Z is nitrogen, only one of R2 and R3 is present, and when Z is oxygen, neither of R2 and R3 is present; and wherein the symbol ##STR00129## represents a five or six membered aromatic or heteroaromatic ring system; and wherein Q is connected to said head group H at said ##STR00130## or a pharmaceutically acceptable salt thereof.

2. The compound of claim 1, wherein at least one of A and B is C═O.

3. The compound according to claim 1, wherein n is 0.

4. The compound according to claim 1, wherein Q is O and p is 2.

5. The compound according to claim 1, wherein Y is selected from the group consisting of amino, cyclic alkylamino, dialkylamino, cyclic diaminoalkyl, and heterocyclic alkylamino.

6. The compound according to claim 1, wherein the hydroxyamic acid (HAm) is protected by a carbamate, such that the hydroxamic acid is represented by the formula ##STR00131##

7. The compound according to claim 1, wherein said compound is selected from the group consisting of: ##STR00132## ##STR00133##

8. The compound according to claim 1, wherein said compound is selected from ##STR00134##

9. A pharmaceutical composition comprising (a) at least one compound, or a pharmaceutically acceptable salt thereof, according to claim 1, (b) optionally, one or more further agent(s) or drug(s), and (c) optionally, one or more pharmaceutically acceptable excipient(s) and/or carrier(s).

10. A method for treating acute myeloid leukemia (AML); wherein said method comprises administering, to a subject in need of such treatment, a compound of claim 1.

11. The method according to claim 10, wherein such treatment is in combination with one or more further agent(s) or drug(s) selected from tyrosine kinase inhibitor(s) and proteasome inhibitor(s), and/or wherein such treatment is in combination with a therapy comprising sensitizing AML cells.

12. The compound, according to claim 1, wherein Y is selected from 1-methylpiperazinyl and morpholinyl.

13. The pharmaceutical composition, according to claim 9, further comprising a tyrosine kinase inhibitor and/or a proteasome inhibitor.

14. The method according to claim 10, wherein the therapy comprising sensitizing AML cells is a radiation therapy.

Description

BRIEF DESCRIPTION OF THE DRAWINGS

(1) FIG. 1. Structure of HDAC inhibitors (A) Shown are the structures of the prior art HDAC inhibitor stubacin (1), ST80 (2), tubastatin A (3) and of the pan-HDACi trichostatin A (4). (B) Overview: general structure of the novel HDAC6 inhibitors of the invention. (C) Shown are the structures of four specific exemplary HDAC6 inhibitors of the invention, in the form of their corresponding HCl-salts; they may however also be used in the form of their free base: 4-(2-((11-(4-(Hydroxycarbamoyl)benzyl)-4-methyl-3,5-dioxo-1,3,4,5,6,11-hexahydro-2,6-methano[1,3]diazocino[5,6-b]indol-8-yl)oxy)ethyl)morpholin-4-ium chloride (21); 4-(2-((7-(4-(Hydroxycarbamoyl)benzyl)-2-methyl-1,3-dioxo-2,3,5,6,7,11c-hexahydro-1H-imidazo[1′,5′:1,2]pyrido[4,3-b]indol-10-yl)oxy)ethyl)morpholin-4-ium chloride (53a): 4-(2-((8-(4-(Hydroxycarbamoyl)benzyl)-2-methyl-1,4-dioxo-1,2,3,4,6,7,8,12c-octahydropyrazino[1′,2′:1,2]pyrido[4,3-b]indol-11-yl)oxy)ethyl)morpholin-4-ium chloride (53b); and 4-(2-((11-(4-(Hydroxycarbamoyl)benzyl)-2-methyl-1,3-dioxo-2,3,5,6,11,11 b-hexahydro-1H-imidazo[1′,5′:1,2]pyrido[3,4-b]indol-8-yl)oxy)ethyl)morpholin-4-ium chloride (65).

(2) Also shown are three compounds which may serve as core structures for further attachment of a Y—(CH.sub.2).sub.p-Q- part, namely: N-Hydroxy-4-((2-methyl-1,3-dioxo-2,3,5,6-tetrahydro-1H-imidazo[1′,5′:1,2]pyrido[4,3-b]indol-7(11cH)-yl)methyl)benzamide (31a), N-hydroxy-4-((2-methyl-1,4-dioxo-1,3,4,6,7,12c-hexahydropyrazino[1′,2′:1,2]pyrido[4,3-b]indol-8(2H)-yl)methyl)benzamide (31b); and N-Hydroxy-4-((2-methyl-1,3-dioxo-2,3,5,6-tetrahydro-1H-imidazo[1′,5′:1,2]pyrido[3,4-b]indol-11(11bH)-yl)methyl)benzamide (41).

(3) FIG. 2: MV4-11 cells were treated for 24 h with 200 nM Marbostat-100 or MS-275 (5 μM) and compared with different new HDAC6is. The cells were analysed by Western blot. The results shown are obtained from at least two independent experiments.

EXAMPLES

(4) Materials and Methods:

(5) Fluorescence HDAC Assay

(6) The IC.sub.50 values for the various inhibitors against the isoenzymes of HDAC were measured by Reaction Biology Corporation (Malvern, Pa., USA) (Buffer, Kalin et al. 2010): For the enzyme inhibition assay, human recombinant protein material was used (human HDAC2 (CAT #: HDAC2), human HDAC6 (CAT #: HDAC6) or human HDAC8 (CAT #: HDAC8)). A 10-point dose response curve was prepared using a 3-fold dilution series (1:3), started at a concentration of 1 μM in DMSO for the synthesized drugs and 10 μM in DMSO for the reference substance Trichostatin A231 (TSA, for HDAC 2, 6 and 8). Reaction time was 90 minutes at 37° C. in a volume of 50 μL in a 96 well plate. Reaction buffer: 50 mM Tris-HCl, pH=8.0, 137 mM NaCl, 2.7 mM KCl, and 1 mM MgCl.sub.2 freshly added: 1 mg/ml bovine serum albumin. The fluorescence was measured using a fluoroscope Ascent™ FL fluorometer. The percent enzyme activity was calculated relative to the DMSO control. IC.sub.50 values were calculated using GraphPad Prism (GraphPad Prism 4.0 Software, San Diego, USA) based on a sigmoidal dose-response equation.

(7) Purity Controls and Determination of the Enantiomeric Excess by Analytical HPLC

(8) were performed with a Waters GmbH (Eschborn, Germany) system, equipped with a Waters 1525 Binary HPLC pump, a Waters 2707 autosampler and a Waters 2998 UV-Vis detector. The column used was a Hibar 125-4, Purospher rp 18e, 3 μm from Merck KGaA, the evaluation was carried out with the software Empower 3, 2010 (Waters GmbH, Eschborn, Germany).

(9) By determining the percentage of area peaks at 220 nm, the purity of the samples could be determined. The column temperature was always 10° C. for compounds measured with this instrument.

(10) Method A

(11) TABLE-US-00001 Time Flow rate [min] [mL/min] % A % B 1. — 0.80 80.0 20.0 2. 30.0 0.80 20.0 80.0 3. 31.0 0.80 5.0 95.0 4. 40.0 0.80 5.0 95.0 5. 41.0 0.80 80.0 20.0 6. 50.0 0.80 80.0 20.0

(12) Method B

(13) TABLE-US-00002 Time Flow rate [min] [mL/min] % A % B 1. — 0.80 95.0 5.0 2. 30.0 0.80 20.0 80.0 3. 31.0 0.80 2.0 98.0 4. 40.0 0.80 2.0 98.0 5. 41.0 0.80 95.0 5.0 6. 50.0 0.80 95.0 5.0

(14) Eluent A: 0.1% formic acid+water, eluent B: 0.1% formic acid+acetonitrile. Injection volume: 10 [μL]. The detection was carried out at a wavelength of 220 nm.

Example 1: Synthesis of Compound 21 According to Scheme 1A

(15) ##STR00048## ##STR00049## ##STR00050##

tert-Butyl 4-((8-hydroxy-4-methyl-3.5-dioxo-3.4.5.6-tetrahydro-2.6-methano[1.3]diazocino[5.6-b]indol-11(1H)-yl)methyl)benzoate (15)

(16) ##STR00051##

(17) Preparation according to literature (Mahboobi, Sellmer et al. 2016) as follows: tert-Butyl 4-((8-(benzyloxy)-4-methyl-3.5-dioxo-3.4.5.6-tetrahydro-2.6-methano[1.3]diazocino[5.6-b]indol-11(1H)-yl)methyl)benzoate (14) 0.52 g (0.94 mmol) and 0.22 g Pd (10% Pd) were dissolved in tetrahydrofuran (65 mL).

(18) The mixture was stirred under hydrogen atmosphere at room temperature and continued until completion of the reaction was observed by TLC (CH.sub.2Cl.sub.2:MeOH (10:1)). The product was collected by filtration over Na.sub.2SO.sub.4 and the solvent was removed under reduced pressure. The product was obtained as a colorless solid. Yield 0.42 g (0.91 mmol; 97%) Colorless crystals; mp: 201.2-205.1° C. IR (KBr): 2969, 2933, 1727, 1715, 1684, 1663 cm.sup.−1; .sup.1H NMR (300 MHz, DMSO): δ 8.89 (s, 1H), 7.81 (d, J=8.3 Hz, 2H), 7.16 (t, J=8.2 Hz, 3H), 6.86 (d, J=2.3 Hz, 1H), 6.58 (dd, J=8.8, 2.3 Hz, 1H), 5.34 (s, 2H), 4.78 (d, J=16.5 Hz, 1H), 4.49 (d, J=16.4 Hz, 1H), 3.86 (d, J=12.9 Hz, 1H), 3.75 (s, 1H), 3.42 (dd, 1H), 2.89 (s, 3H), 1.51 (s, 9H). ESI-MS m/z (%): 406.14 [MH.sup.+-C.sub.4H.sub.8] (100), 462.20 [MH.sup.+] (16.77), 479.23 [MNH.sub.4+] (39.69), 945.38 [MNa.sup.+] (4.89). Anal. calcd for C.sub.26H.sub.27N.sub.3O.sub.5: C, 67.66; H, 5.90; N, 9.10; found: C, 67.41; H, 5.98; N, 8.84.

tert-Butyl 4-((4-methyl-8-(2-morpholinoethoxy)-3,5-dioxo-3,4,5,6 tetrahydro-2,6-methano[1,3]diazocino[5,6-b]indol-11(1H)-yl)methyl)benzoate (17)

(19) ##STR00052##

(20) A stirred mixture of 15 (0.28 g, 0.61 mmol), 4-(2-chloroethyl)morpholine hydrochloride (16) (0.15 g, 0.81 mmol) and K.sub.2CO.sub.3 (0.42 g, 3.0 mmol) in 2-butanone (30.0 mL) was heated till reflux for 4 d.

(21) The mixture was cooled to room temperature, the solid filtered off and the solvent removed under reduced pressure. After purification by cc (SiO.sub.2; CH.sub.2Cl.sub.2, MeOH 10:1) and removal of the solvent under reduced pressure the product (0.24 g, 0.42 mmol, 68%) was obtained as a colorless solid. mp.: 204.7-208.0° C.; IR (KBr): 2857, 1714, 1687 cm.sup.−1; .sup.1H NMR (300 MHz, CDCl.sub.3) δ 7.91 (d, J=8.3 Hz, 2H), 7.15 (d, J=2.3 Hz, 1H), 7.09 (d, J=9.0 Hz, 1H), 7.03 (d, J=8.3 Hz, 2H), 6.83 (dd, J=8.9, 2.4 Hz, 1H), 5.34-5.22 (m, 1H), 5.09 (d, J=17.0 Hz, 1H), 4.84 (d, J=16.4 Hz, 1H), 4.25 (d, J=15.7 Hz, 3H), 3.89 (d, J=13.3 Hz, 1H), 3.83 (d, J=8.0 Hz, 5H), 3.32 (dd, J=13.1, 2.1 Hz, 1H), 3.06 (s, 3H), 2.93 (s, 2H), 2.72 (s, 4H), 1.56 (s, 9H). ESI-MS m/z (%): 575.29 [MH.sup.+] (100), 1171.55 [2MNa.sup.+] (0.3). Anal. calcd for C.sub.32H.sub.38N.sub.4O.sub.6×0.25 H.sub.2O: C, 66.36; H, 6.70; N, 9.67; found: C, 65.98; H, 6.58; N, 9.43.

4-(2-((11-(4-Carboxybenzyl)-4-methyl-3,5-dioxo-1,3,4,5,6,11-hexahydro-2,6-methano[1,3]diazocino[5,6-b]indol-8-yl)oxy)ethyl)morpholin-4-ium 2,2,2-trifluoroacetate (18)

(22) ##STR00053##

(23) tert-Butyl 4-((4-methyl-8-(2-morpholinoethoxy)-3,5-dioxo-3,4,5,6-tetrahydro-2,6-methano[1,3]diazocino[5,6-b]indol-11(1H)-yl)methyl)benzoate (17) (0.82 g, 1.43 mmol) was dissolved in CH.sub.2Cl.sub.2 (25 mL) and trifluoro acetic acid (10.0 mL) was added. The mixture was stirred at room temperature (2 h) and the solvent and excess of trifluoro acetic acid removed under reduced pressure.

(24) Yield 0.90 g (1.42 mmol, 99%) slightly yellow crystals; mp: 182.9-184.0° C. IR (KBr): 3441, 1642 cm.sup.−1; .sup.1H NMR (300 MHz, DMSO) δ 12.98 (s, 1H), 9.95 (s, 1H), 7.85 (d, J=8.3 Hz, 2H), 7.36 (d, J=8.9 Hz, 1H), 7.13 (d, J=8.3 Hz, 2H), 7.10 (d, J=2.4 Hz, 1H), 6.85 (dd, J=8.9, 2.4 Hz, 1H), 5.43 (s, 2H), 4.82 (d, J=16.6 Hz, 1H), 4.54 (d, J=16.5 Hz, 1H), 4.34 (s, 2H), 3.99 (d, J=11.3 Hz, 2H), 3.91 (d, J=12.7 Hz, 1H), 3.83 (s, 1H), 3.71 (t, J=11.9 Hz, 2H), 3.64-3.49 (m, 4H), 3.45 (d, J=11.3 Hz, 1H), 3.23 (s, 2H), 2.90 (s, 3H). ESI-MS m/z (%): 519.23 [MH.sup.+] (100), 1059.42 [2MNa.sup.+] (0.03). Anal. calcd for C.sub.30H.sub.31F.sub.3N.sub.4O.sub.8: C, 56.96; H, 4.94; N, 8.86; found: C, 56.88; H, 5.05; N, 8.56.

4-((4-Methyl-8-(2-morpholinoethoxy)-3,5-dioxo-3,4,5,6-tetrahydro-2,6-methano[1,3]diazocino[5,6-b]indol-11(1B)-yl)methyl)-N—((tetrahydro-2H-pyran-2-yl)oxy)benzamide (20)

(25) ##STR00054##

(26) A mixture of 4-(2-((11-(4-carboxybenzyl)-4-methyl-3,5 dioxo-1,3,4,5,6,11-hexahydro-2,6-methano[1,3]diazocino[5,6-b]indol-8-yl)oxy)ethyl)morpholin-4-ium 2,2,2-trifluoroacetate (18) (0.63 g; 1.00 mmol), BOP (0.53 g, 1.20 mmol) diisopropylethlamine (0.52 mL, 3.00 mmol) and O-(tetrahydro-2H-pyran-2-yl)hydroxylamine (19) in THF was stirred at room temperature overnight. The mixture was poured into water, extracted with ethyl acetate (3×50 mL), the combined organic layers dried (Na.sub.2SO.sub.4), the solvent removed and the product purified by cc (SiO.sub.2, CH.sub.2Cl.sub.2, MeOH 20:1). Yield 0.66 g (1.00 mmol, 99%) colorless foam. mp.: 135.7-138.0° C.; IR (KBr): 3433, 1729, 1684 cm.sup.−1; .sup.1H NMR (300 MHz, DMSO) δ 11.60 (s, 1H), 7.67 (d, J=8.3 Hz, 2H), 7.29 (d, J=8.9 Hz, 1H), 7.12 (d, J=8.3 Hz, 2H), 7.00 (d, J=2.4 Hz, 1H), 6.75 (dd, J=8.9, 2.4 Hz, 1H), 5.37 (s, 2H), 4.95 (s, 1H), 4.82 (d, J=16.6 Hz, 1H), 4.53 (d, J=16.4 Hz, 1H), 4.13-3.98 (m, 3H), 3.89 (d, J=12.8 Hz, 1H), 3.84 (s, 1H), 3.59 (d, J=4.6 Hz, 3H), 3.57 (s, 2H), 3.53-3.39 (m, 2H), 2.89 (s, 3H), 2.70 (t, J=5.7 Hz, 2H), 2.54 (s, 2H), 1.61 (d, J=49.1 Hz, 6H), 1.17 (t, J=7.1 Hz, 1H); ESI-MS m/z (%): 618.29 [MH.sup.+] (100).

4-(2-((11-(4-(Hydroxycarbamoyl)benzyl)-4-methyl-3,5-dioxo-1,3,4,5,6,11-hexahydro-2,6-methano[1,3]diazocino[5,6-b]indol-8-yl)oxy)ethyl)morpholin-4-ium chloride (21)

(27) ##STR00055##

(28) To a stirred solution of 4-((4-methyl-8-(2-morpholinoethoxy)-3,5-dioxo-3,4,5,6-tetrahydro-2,6-methano[1,3]diazocino[5,6-b]indol-11(1H)-yl)methyl)-N—((tetrahydro-2H-pyran-2-yl)oxy)benzamide (20) (1.15 g, 1.86 mmol) in CH.sub.2Cl.sub.2 (40.0 mL) HCl in iso-propanol (1.50 mL, 5-6N) was added dropwise. After 2 h the resulting solid was allowed to precipitate, the solvent decanted and the solid dissolved in the necessary amount of MeOH. The solution obtained was added dropwise to a mixture of light petrol/Et.sub.2O whilst stirring, the precipitating solid removed by filtration and the slightly wet solid died in vacuo. Yield 0.80 g (1.40 mmol, 75%) pale yellow solid. mp: 158.4-162.0° C. IR (KBr): 3433, 1671, 1468 cm.sup.−1; .sup.1H NMR (300 MHz, DMSO) δ 11.19 (s, 1H), 10.96 (s, 1H), 9.53-8.49 (m, 1H), 7.66 (d, J=8.3 Hz, 2H), 7.35 (d, J=8.9 Hz, 1H), 7.10 (d, J=8.3 Hz, 2H), 7.07 (d, J=2.4 Hz, 1H), 6.83 (dd, J=8.9, 2.4 Hz, 1H), 5.41 (d, J=18.6 Hz, 2H), 4.84 (d, J=16.6 Hz, 1H), 4.56 (d, J=16.6 Hz, 1H), 4.39 (d, J=4.6 Hz, 2H), 3.97 (d, J=12.4 Hz, 2H), 3.90 (d, J=12.9 Hz, 1H), 3.78 (d, J=11.4 Hz, 1H), 3.54 (d, J=8.0 Hz, 3H), 3.48 (s, 2H), 3.41 (d, J=3.1 Hz, 4H), 2.90 (s, 3H). ESI-MS m/z (%): 534.24 [MH.sup.+] (100). Anal. calcd for C.sub.28H.sub.32ClN.sub.5O.sub.6+1.75 H.sub.2O: C, 55.90; H, 5.95; N, 11.64; found: C, 55.99; H, 5.84; N, 11.26.

(29) ##STR00056## ##STR00057## ##STR00058##

(30) Synthesis of the S-Enantiomer:

Ethyl (S)-2-(5-(benzyloxy)-1H-indole-3-yl)-3-nitropropanoate ((S)-7a)

(31) ##STR00059##

(32) From 5-(benzyloxy)-1H-indol (6) (2.79 g, 12.50 mmol) as described by Sellmer et al. (Sellmer, Stangl et al. 2018). The product was purified by cc (SiO.sub.2; CH.sub.2Cl.sub.2, n-hexane, EtOAc 1:5:1). Yield 3.00 g (8.15 mmol, 65%) yellow oil. IR (ATR, attenuated total reflection): 1726; 1552 cm.sup.−1. .sup.1H NMR (300 MHz, DMSO-d.sub.6) δ 11.07 (s, 1H), 7.49 (d, J=6.9 Hz, 2H), 7.44-7.32 (m, 3H), 7.28 (m, 3H), 6.85 (dd, J=8.8, 2.4 Hz, 1H), 5.24 (dd, J=15.0, 10.7 Hz, 1H), 5.11 (s, 2H), 4.90 (dd, J=15.1, 4.8 Hz, 1H), 4.67 (dd, J=10.6, 4.7 Hz, 1H), 4.21-3.95 (m, 2H), 1.12 (t, J=7.1 Hz, 3H). ee determination: HPLC with a Chiralcel OD-H column, hexane/2-propanol (90:10), flow rate=0.6 mL/min, 220 fn. t.sub.R=134.5 min(major), 152.5, min (minor), ≥99% ee, [α].sup.20.sub.259 +134.093 (c 0.1; MeOH). HRMS (ESI-MS) m/z: calcd: 369.1445 [MH.sup.+], found: 369.1445 [MH.sup.+]. RP (reversed phase)-HPLC (220 nm, Method A): 100.0%, t.sub.R=21.6 min.

Ethyl (S)-3-amino-2-(5-(benzyloxy)-1H-indole-3-yl)propanoate hydrochloride ((S)-8a)

(33) ##STR00060##

(34) From ethyl (S)-2-(5-(benzyl oxy)-1H-indole-3-yl)-3-nitropropanoate ((S)-7a) (4.90 g, 13.31 mmol) as described by Sellmer et al. (Sellmer, Stangl et al. 2018). Yield 4.12 g (10.99 mmol; 83%) colorless foam. IR (ATR): 3400; 2908; 1721 cm.sup.−1. .sup.1H NMR (300 MHz, DMSO-d.sub.6) δ 11.10 (d, J=2.1 Hz, 1H), 8.11 (m, 3H), 7.48 (d, J=6.9 Hz, 2H), 7.44-7.28 (m, 4H), 7.24 (dd, J=14.5, 2.4 Hz, 2H), 6.86 (dd, J=8.8, 2.3 Hz, 1H), 5.10 (s, 2H), 4.25 (t, 0.1=7.2 Hz, 1H), 4.09 (q, J=10.8, 7.1 Hz, 2H), 3.52-3.40 (m, 1H), 3.19-3.06 (m, 1H), 1.13 (t, J=7.1 Hz, 3H). [α].sup.20.sub.589 −69.3 (c 0.1; MeOH). HRMS (C.sub.20H.sub.22N.sub.2O.sub.3, ESI-MS) m/z: calcd.: 339.1703 [MH.sup.+], found.: 339.1703 [MH.sup.+]. RP-HPLC (220 nm, Method A): 99.3%, t.sub.R=7.2 min.

Ethyl (S)-6-(benzyloxy)-2,3,4,9-tetrahydro-1H-pyrido[3,4-b]indole-4-carboxylate hydrochloride ((S)-9a)

(35) ##STR00061##

(36) From ethyl (S)-3-amino-2-(5-(benzyloxy)-1H-indol-3-yl)propanoate hydrochlorid ((S)-8a) (4.12 g, 10.99 mmol) as by Sellmer et al. (Sellmer, Stangl et al. 2018). Yield 2.82 g (7.29 mmol, 66%). mp.: 262.7° C. IR (KBr): 2922; 2797; 1719 cm.sup.−1. .sup.1H NMR (400 MHz, DMSO-d.sub.6) δ 11.19 (s, 1H), 10.10 (s, 1H), 9.08 (s, 1H), 7.47-7.41 (m, 2H), 7.41-7.35 (m, 2H), 7.34-7.27 (m, 2H), 7.12 (d, J=2.3 Hz, 1H), 6.85 (dd, J=8.8, 2.4 Hz, 1H), 5.07 (q, J=12.0 Hz, 2H), 4.37-4.25 (m, 2H), 4.16 (d, J=2.6 Hz, 1H), 4.15-4.04 (m, 2H), 3.67 (d, J=9.7 Hz, 1H), 3.49 (d, J=12.0 Hz, 1H), 1.19 (t, J=7.1 Hz, 3H). ee was determination: HPLC with a Chiralcel OD-H column, hexane/2-propanol (90:10), flow rate=0.6 mL/min, 220 nm. t.sub.R=83.3 min (major), 110.2 min (minor), ≥96% ee, [α].sup.20.sub.589 −126.5 (c 0.1; MeOH). HRMS (C.sub.21H.sub.22N.sub.2O.sub.3, ESI-MS) m/z: calcd.: 351,1703 [MH.sup.+], found: 351,1705 [MH.sup.+]. RP-HPLC (220 nm, Method A): 99.4%, t.sub.R=8.0 min.

(S)-6-(Benzyloxy)-N-methyl-2,3,4,9-tetrahydro-1H-pyrido[3,4-b]indole-4-carboxamide hydrochloride ((S)-67a)

(37) ##STR00062##

(38) Ethyl (S)-6-(benzyloxy)-2,3,4,9-tetrahydro-1H-pyrido[3,4-b]indol-4-carboxylat hydrochlorid ((S)-9a) (0.40 g, 1.03 mmol) was dissolved in a mixture of DCM/MeOH (1:1; 10 mL), DMSO (8 drops), DMF (6.9 mL) and a methanolic methylamine solution (30%, 19 mL) at 0° C. After adding of a catalytic amount of NaCN (0.304 g) it was stirred for 6 d. Purification by cc (SiO.sub.2; CH.sub.2Cl.sub.2, MeOH, NH.sub.3 conz 50:10:0.1) (dry load technique) yielded 0.254 g (0.68 mmol; 66%) colorless crystals. mp.: 178.4° C. IR (KBr): 2940, 1637 cm.sup.−1. .sup.1H NMR (300 MHz, DMSO-d.sub.6) δ 10.65 (s, 1H), 7.92 (m, 1H), 7.46 (d, J=7.0 Hz, 2H), 7.42-7.31 (m, 3H), 7.17 (d, 0.1=8.7 Hz, 1H), 6.99 (d, J=2.3 Hz, 1H), 6.73 (dd, J=8.7, 2.4 Hz, 1H), 5.04 (s, 2H), 3.91-3.75 (m, 2H), 3.44 (d, J=4.2 Hz, 1H), 3.11 (dd, J=13.0, 4.1 Hz, 1H), 2.94 (dd, J=12.9, 4.8 Hz, 1H), 2.58 (d, J=4.5 Hz, 3H). [α].sup.20.sub.589 −35.6 (c 0.1; MeOH). HRMS (C.sub.20H.sub.21N.sub.3O.sub.2, ESI-MS) m/z: calcd.: 336.1707[MH.sup.+], found: 336.1705 [MH.sup.+]. RP-HPLC (220 nm, Method A): 98.7%, t.sub.R=6.2 min.

(6S)-8-(Benzyloxy)-4-methyl-6,11-dihydro-2,6-methano[1,3]diazocino[5,6-b]indole-3,5(1H,4H)-dione ((S)-12)

(39) ##STR00063##

(40) From 6-(benzyloxy)-N-methyl-2,3,4,9-tetrahydro-1H-pyrido[3,4-b]indole-4-carboxamide ((S)-67a) after release of the base with dilut. NH.sub.3-Lösung (0.163 g, 0.44 mmol) following Sellmer et al. (Sellmer, Stangl et al. 2018). Purification by cc (SiO.sub.2; EE) (dry load technique) yielded 0.128 g (0.35 mmol; 80%) colorless crystals. mp.: 273.6° C. IR (KBr): 3281; 1730; 1673 cm.sup.−1. .sup.1H NMR (400 MHz, CDCl.sub.3) δ 8.03 (s, 1H, NH (Indole)), 7.50-7.45 (m, 2H, Ar—H), 7.42-7.30 (m, 3H, Ar—H), 7.23-7.19 (m, 2H, Ar—H), 6.92 (dd, J=8.8, 2.4 Hz, 1H, Ar—H), 5.10 (d, J=3.2 Hz, 2H, OCH.sub.2), 4.87 (d, J=16.3 Hz, 1H, NCH.sub.AH.sub.BC), 4.50 (d, J=16.3 Hz, 1H, NCH.sub.AH.sub.BC), 3.90 (d, J=13.1 Hz, 1H, NCH.sub.AH.sub.BCH), 3.83 (br s, 1H, NCH.sub.2CH), 3.37 (dd, J=13.1, 2.2 Hz, 1H, NCH.sub.AH.sub.BCH), 3.07 (s, 3H, NCH.sub.3).

(41) .sup.13C NMR (101 MHz, CDCl.sub.3) δ 173.23 (quart.CO), 161.49 (quart.CO), 154.03 (quart. Ar—C), 137.40 (quart. Ar—C), 132.48 (quart. Ar-{right arrow over (C)}), 130.90 (quart. Ar—C), 128.53 (+, Ar—CH), 127.85 (+, Ar—CH), 127.69 (+, (Ar—CH).sub.2), 126.61 (quart. Ar—C), 113.63 (+, Ar—CH), 112.04 (+, Ar—CH), 106.47 (quart. Ar—C), 101.44 (+, Ar—CH), 70.83 (−, OCH.sub.2), 50.23 (−, NCH.sub.2C), 47.58 (−, NCH.sub.2CH), 36.82 (+, Ar—CH), 30.95 (+, NCH.sub.2CH), 27.89 (+, NCH.sub.3). [α].sup.20.sub.589 −27.4 (c 0.1; MeOH). HRMS (C.sub.21H.sub.19N.sub.3O.sub.3, ESI-MS) m/z: calcd.: 362.1499 [MH.sup.+], found: 362.1498 [MH.sup.+]. RP-HPLC (220 nm, Method A): 99.5%, t.sub.R=18.2 min.

tert-Butyl 4-(((6S)-8-(benzyloxy)-4-methyl-3,5-dioxo-3,4,5,6-tetrahydro-2,6-methano[1,3]diazocino[5,6-b]indole-11(1H)-yl)methyl)benzoate ((S)-14)

(42) ##STR00064##

(43) A stirred mixture of 8-(benzyloxy)-4-methyl-6,11-dihydro-2,6-methano[1,3]diazocino[5,6-b]indole-3,5(1H,4H)-dione ((S)-12) (0.127 g; 0.35 mmol), tert-butyl 4-(bromomethyl)benzoate (13) (0.103 g, 0.38 mmol) und K.sub.2CO.sub.3 (0.243 g, 1.76 mmol) in 2-butanone (5.20 mL), was heated to 80° C. for 16 h. The mixture was cooled to rt, the solid filtered off and the solvent removed i.vac. After purification by cc (SiO.sub.2; CH.sub.2Cl.sub.2, EtOAc 3:1) and removal of the solvent i.vac. the product (0.16 g, 0.29 mmol, 83%), yellow oil. IR (ATR): 1710, 1685 cm.sup.−1. .sup.1H NMR (300 MHz, CDCl.sub.3) δ 7.99 (dd, J=8.5, 2.3 Hz, 1H), 7.92 (d, J=8.3 Hz, 1H), 7.51-7.46 (m, 2H), 7.43-7.32 (m, 4H), 7.07 (m, 3H), 6.91 (dd, J=8.9, 2.4 Hz, 1H), 5.30 (d, J=17.0 Hz, 1H), 5.15 (m, 1H), 5.11 (d, J=2.7 Hz, 2H), 4.85 (d, J=16.4 Hz, 1H), 4.26 (d, J=16.4 Hz, 1H), 3.90 (d, J=13.0 Hz, 2H), 3.40-3.27 (m, 1H), 3.07 (s, 3H), 1.57 (s, 9H). [α].sup.20.sub.589 −24.0 (c 0.1; MeOH). HRMS (C.sub.33H.sub.33N.sub.3O.sub.5, ESI-MS) m/z: calcd.: 574.2312 [MNa.sup.+], found: 574.2317 [MNa.sup.+]. RP-HPLC (220 nm, Method A): 99.0%, t.sub.R=7.0 min.

tert-Butyl 4-(((6S)-8-hydroxy-4-methyl-3,5-dioxo-3,4,5,6-tetrahydro-2,6-methano[1,3]diazocino[5,6-b]indole-11(1H)-yl)methyl)benzoate ((S)-15)

(44) ##STR00065##

(45) As described by Mahboobi et al. (Mahboobi, Teller et al. 2002) to a solution of tert-butyl 4-((8-(benzyloxy)-4-methyl-3,5-dioxo-3,4,5,6-tetrahydro-2,6-methano[1,3]diazocino[5,6-b]indole-11(1H)-yl)methyl) benzoate ((S)-14) (0.155 g, 0.28 mmol) in THF/MeOH (2:1) Pd/C 10% (0.049 g) and ammonium formiate (0.29 g, 4.6 mmol) were added, and the mixture was stirred for 20 min at 75° C. After filtration of Pd/C, purification cc (CH.sub.2Cl.sub.2, EtOAc (3:1)) yielded (0.100 g, 0.22 mmol, 79%) colorless crystals. mp.: 206.3° C. IR (KBr): 3416, 1715, 1663 cm.sup.−1. .sup.1H NMR (400 MHz, DMSO-d.sub.6) δ 8.86 (s, 1H), 7.79 (d, J=8.3 Hz, 2H), 7.14 (m, 3H), 6.85 (d, J=2.3 Hz, 1H), 6.57 (dd, J=8.8, 2.3 Hz, 1H), 5.33 (s, 2H), 4.76 (d, J=16.5 Hz, 1H), 4.48 (d, J=16.4 Hz, 1H), 3.85 (d, J=12.6 Hz, 1H), 3.74 (br s, 1H), 3.40 (dd, J=13.2, 2.2 Hz, 1H), 2.88 (s, 3H), 1.50 (s, 9H). [α].sup.20.sub.589 89.5 (c 0.1; MeOH). HRMS (C.sub.26H.sub.27N.sub.3O.sub.5, ESI-MS) m/z: calcd.: 484.1843 [MNa.sup.+], found:484.1841 [MNa.sup.+]. Anal. (C.sub.26H.sub.27N.sub.3O.sub.5) calcd: C, 67.66; H, 5.90; N, 9.10; found: C, 67.48; H, 6.06; N, 8.88.

tert-Butyl 4-(((6S)-4-methyl-8-(2-morpholinoethoxy)-3,5-dioxo-3,4,5,6-tetrahydro-2,6-methano[1,3]diazocino[5,6-b]indole-11(1H)-yl)methyl)benzoate

C.SUB.32.H.SUB.3.N.SUB.4.O.SUB.6 .(MW=574.68 g/mol) ((S)-17)

(46) ##STR00066##

(47) A mixture of (S)-15 (0.082 g, 0.18 mmol), 4-(2-chloroethyl)morpholine hydrochloride (16) (0.044 g, 0.24 mmol) und K.sub.2CO.sub.3 (0.123 g, 0.89 mmol) in 2-butanone (8.8 mL) was heated to 70° C. for 16 h. Purification by cc (SiO.sub.2; CH.sub.2Cl.sub.2, EtOAc, 3.1) (dry load technique) yielded 0.067 g (0.12 mmol; 67%) colorless yellow oil. IR (ATR): 1712, 1688 cm.sup.−1. .sup.1H NMR (400 MHz, CDCl.sub.3) δ 7.91 (d, J=8.3 Hz, 2H, Ar—H), 7.15 (d, J=2.4 Hz, 1H, Ar—H),7.09 (d, J=8.9 Hz, 1H, Ar—H), 7.03 (d, J=8.3 Hz, 2H, Ar—H), 6.83 (dd, J=8.9, 2.4 Hz, 1H, Ar—H), 5.33-5.24 (m, 1H, NCH.sub.AH.sub.BAr), 5.09 (d, J=17.0 Hz, 1H, NCH.sub.AH.sub.BAr), 4.84 (d, J=16.4 Hz, 1H, NCH.sub.AH.sub.BC), 4.29-4.21 (m, 3H, NCH.sub.AH.sub.BC, OCH.sub.2CH.sub.2), 3.89 (d, J=13.1 Hz, 1H, NCH.sub.AH.sub.BCH), 3.85 (br s, 1H, NCH.sub.2CH), 3.82 (s, J=32.6 Hz, 4H, O(CH.sub.2).sub.2CH.sub.2N-morpholine), 3.32 (dd, J=13.1, 2.2 Hz, 1H, NCH.sub.AH.sub.BCH), 3.06 (s, 3H, NCH.sub.3), 2.95 (m, 2H, OCH.sub.2CH.sub.2), 2.75 (s, 4H, O(CH.sub.2).sub.2(CH.sub.2).sub.2N-morpholine), 1.56 (s, 9H, OC(CH.sub.3).sub.3.

(48) .sup.13C NMR (101 MHz, CDCl.sub.3) δ 173.13 (quart.CO), 165.17 (quart.CO), 161.26 (quart.CO), 152.68 (quart.O—Ar—C), 141.02 (quart. Ar—C), 133.80 (quart. Ar—C), 131.90 (quart. Ar—C), 131.77 (quart. Ar—C), 130.20 (+, (Ar—CH).sub.2), 126.49 (quart. Ar—C), 126.07 (+, (Ar—CH).sub.2), 113.22 (+, Ar—CH), 110.44 (+, Ar—CH), 105.97 (quart. Ar—C), 101.48 (+, Ar—CH), 81.27 (quart.OC(CH.sub.3).sub.3), 72.83 (−, O(CH.sub.2).sub.2CH.sub.2N-morpholine), 65.46 (−, OCH.sub.2CH.sub.2), 57.68 (−, OCH.sub.2CH.sub.2), 53.91 (−, O(CH.sub.2).sub.2(CH.sub.2).sub.2N-morpholine), 49.46 (−, NCH.sub.2C), 47.37 (−, NCH.sub.2CH),47.12 (−, NCH.sub.2Ar), 36.87 (+, NCH.sub.2CH), 28.36 (+, OC(CH.sub.3).sub.3), 27.91 (+, NCH.sub.3). [α].sup.20.sub.589 42.1 (c 0.1; MeOH). HRMS (C.sub.32H.sub.38N.sub.4O.sub.6, ESI-MS) m/z: calcd.: 575.2864[MH.sup.+], found: 575.2874 [MH.sup.+]. RP-HPLC (220 nm, Method A): 99.7%, t.sub.R=11.4 min.

4-(2-(((6S S)-11-(4-carboxybenzyl)-4-methyl-3,5-dioxo-1,3,4,5,6,11-hexahydro-2,6-methano[1,3]diazocino[5,6-b]indole-8-yl)oxy)ethyl)morpholin-4-ium 2,2,2-trifluoroacetate ((S)-18)

(49) ##STR00067##

(50) To a solution of tert-butyl 4-((4-methyl-8-(2-morpholinoethoxy)-3,5-dioxo-3,4,5,6-tetrahydro-2,6-methano[1,3]diazocino[5,6-b]indole-11(1H)-yl)methyl)benzoate ((S)-17) (0.161 g; 0.28 mmol) in CH.sub.2Cl.sub.2 (4.90 mL), CF.sub.3COOH (1.97 mL) was added and the mixture was stirred for 20 min. at r.t. After solvent removal a slight brown oil was obtained (0.237 g, 0.37 mmol; 99%). IR (ATR): 1671; 1167 cm.sup.−1. .sup.1H NMR (400 MHz, DMSO-d.sub.6) δ 9.99 (s, 1H), 7.85 (d, J=8.3 Hz, 2H), 7.36 (d, J=8.9 Hz, 1H), 7.13 (d, J=8.3 Hz, 2H), 7.10 (d, J=2.3 Hz, 1H), 6.85 (dd, J=8.9, 2.4 Hz, 1H), 5.42 (s, 2H), 4.82 (d, J=16.6 Hz, 1H), 4.54 (d, J=16.6 Hz, 1H), 4.38-4.27 (m, 4H), 4.04-3.94 (m, 3H), 3.91 (d, J=12.8 Hz, 2H), 3.83 (s, 2H), 3.47-3.42 (m, 4H), 2.90 (s, 3H). .sup.19F NMR (377 MHz, DMSO-d.sub.6) δ −73.70 (s). [α].sup.20.sub.589 10.7 (c 0.1; MeOH). HRMS (C.sub.28H.sub.30N.sub.4O.sub.6, ESI-MS) m/z: calcd.: 519.2238[MH.sup.+], found: 519.2238 [MH.sup.+]. RP-HPLC (220 nm, Method B): 100.0%, t.sub.R=10.5 min.

4-(((6S)-4-Methyl-8-(2-morpholinoethoxy)-3,5-dioxo-3,4,5,6-tetrahydro-2,6-methano[1,3]diazocino[5,6-b]indole-11(1H)-yl)methyl)-N—((tetrahydro-2H-pyran-2-yl)oxy)benzamide ((S)-20)

(51) ##STR00068##

(52) From 4-((4-methyl-8-(2-morpholinoethoxy)-3,5-dioxo-3,4,5,6-tetrahydro-2,6-methano[1,3]diazocino[5,6-b]indole-1(1H)-yl)methyl)benzoic acid ((S)-18) (0.237 g, 0.46 mmol) as described by Sellmer et al. (Sellmer, Stangl et al. 2018). Purification by cc (SiO.sub.2; CH.sub.2Cl.sub.2, MeOH, 10:1) (dry load technique) yielded 0.131 g (0.21 mmol; 77%) yellow oil. IR (ATR): 1728; 1638 cm.sup.−1. .sup.1H NMR (400 MHz, DMSO-d.sub.6) δ 11.57 (s, 1H), 7.66 (d, J=8.2 Hz, 2H), 7.27 (d, J=8.9 Hz, 1H), 7.11 (d, J=8.2 Hz, 2H), 7.00 (d, J=2.3 Hz, 1H), 6.74 (dd, J=8.9, 2.4 Hz, 1H), 5.36 (s, 2H), 5.11-5.07 (m, 1H), 4.98-4.93 (m, 1H), 4.81 (d, J=16.6 Hz, 1H), 4.52 (d, J=16.6 Hz, 1H), 4.11-4.06 (m, 2H), 3.88 (d, J=12.6 Hz, 1H), 3.82 (br s, 1H), 3.60-3.57 (m, 4H), 3.56-3.52 (m, 2H), 2.88 (s, 3H), 2.77-2.71 (m, 2H), 2.40 (m, 2H), 2.32 (m, 1H), 1.70-1.55 (m, 6H). ee determination by HPLC with a Phenomenex Lux Cellulose-2 column, methanol/2-propanol (90:10), flow rate=1.0 ml/min, 220 nm. t.sub.R=21.4-26.8 min (major), 38.7-43.6 min (minor), 94% ee, [α].sup.20.sub.589 32.0 (c 0.1; MeOH). HRMS (C.sub.33H.sub.39N.sub.5O.sub.7, ESI-MS) m/z: calcd: 618.2922[MH.sup.+], found: 618.2928 [MH.sup.+]. RP-HPLC (220 nm, Method B): 97.7%, t.sub.R=11.2 min.

N-Hydroxy-4-(((6S)-4-methyl-8-(2-morpholinoethoxy)-3,5-dioxo-3,4,5,6-tetrahydro-2,6-methano[1,3]diazocino[5,6-b]indole-11(1H)-yl)methyl)benzamide ((S)-21)

(53) ##STR00069##

(54) 4-((4-Methyl-8-(2-morpholinoethoxy)-3,5-dioxo-3,4,5,6-tetrahydro-2,6-methano[1,3]diazocino[5,6-b]indol-11(1H)-yl)-N—((tetrahydro-2H-pyran-2-yl)oxy)benzamid ((S)-20) (0.060 g, 0.10 mmol) was dissolved in CH.sub.2Cl.sub.2 (2.09 mL), 6N HCl.sub.iprop (0.08 mL) (pH=1) was added and stirred for 2 h at r.t. (dc-control: CH.sub.2Cl.sub.2/MeOH (10:1)). The solvent was removed i.vac., the remaining solid washed with CH.sub.2Cl.sub.2 and dissolved again in MeOH (1.0 mL). The desired product was obtained by crystallization from PE/EE (1:1; 30 mL). Yield (0.048 g, 0.08 mmol, 80%). mp.: 175.7° C. IR (KBr): 3416, 1671, 1614 cm.sup.−1. 1H NMR (400 MHz, DMSO-d.sub.6) δ 11.18 (s, 1H, NHOH), 9.01 (s, 1H, NHOH), 7.65 (d, J=8.3 Hz, 2H, Ar—H), 7.33 (d, J=8.9 Hz, 1H, Ar—H), 7.09 (d, J=8.3 Hz, 2H, Ar—H), 7.05 (d, J=2.4 Hz, 1H, Ar—H), 6.82 (dd, J=8.9, 2.4 Hz, 1H, Ar—H), 5.36 (s, 2H, NCH.sub.2Ar), 4.82 (d, J=16.6 Hz, 1H, NCH.sub.AH.sub.BC), 4.55 (d, J=16.5 Hz, 1H, NCH.sub.AH.sub.BC), 4.45-4.33 (m, 2H, OCH.sub.2CH.sub.2), 3.95-3.80 (m, 6H, OCH.sub.2CH.sub.2N-morpholine, NCH.sub.AH.sub.BCH, OCH.sub.2CH.sub.2N-morpholine, NCH.sub.2CH), 3.55-3.42 (m, 5H, OCH.sub.2CH.sub.2N, OCH.sub.2CH.sub.2N-morpholine, NCH.sub.AH.sub.BCH), 3.20 (s, 2H, OCH.sub.2CH.sub.2N- morpholine), 2.89 (s, 3H, NCH.sub.3). .sup.13C NMR (101 MHz, DMSO) δ 173.58 (quart.CO), 164.26 (quart.CO), 161.31 (quart.CO), 152.68 (quart.O—Ar—C), 141.20 (quart. Ar—C), 135.49 (quart. Ar—C), 132.37 (quart. Ar—C), 131.99 (quart. Ar—C), 127.72 (−, Ar—CH).sub.2), 127.02 (−, (Ar—CH).sub.2), 126.61 (quart. Ar—C), 112.22 (−, Ar—CH), 111.75 (−, Ar—CH), 105.50 (quart. Ar—C), 102.20 (−, Ar—CH), 63.63 (+, O(CH.sub.2).sub.2CH.sub.2N-morpholine), 63.38 (+, OCH.sub.2CH.sub.2), 55.58 (+, OCH.sub.2CH.sub.2), 52.14 (+, O(CH.sub.2).sub.2(CH.sub.2).sub.2N-morpholine), 49.44 (+, NCH.sub.2C), 46.69 (+, NCH.sub.2CH), 46.56 (+, NCH.sub.2Ar), 36.49 (−, NCH.sub.2CH), 27.88 (−, NCH.sub.3). [α].sup.20.sub.589 14.3 (c 0.1; H.sub.2O). HRMS (C.sub.28H.sub.31N.sub.5O.sub.6, ESI-MS) m/z: calcd: 534.2347 [MH.sup.+], found: 534.2350 [MH.sup.+]. RP-HPLC (220 nm, Method B): 96.0%, t.sub.R=8.5 min.

(55) Synthesis of the R-Enantiomer

Ethyl (R)-2-(5-(benzyloxy)-1H-indole-3-yl)-3-nitropropanoate ((R)-7a)

(56) ##STR00070##

(57) Preparation as described for the S-enantiomer (S)-7a by use of 5-(benzyloxy)-1H-indole (2.90 g, 12.99 mmol). Purification by cc (SiO.sub.2; CH.sub.2Cl.sub.2, n-hexane, EtOAc 1:5:1). Yield 3.30 g (8.96 mmol, 69%) brown oil. IR (ATR): 1726; 1552 cm.sup.−1. .sup.1H NMR (300 MHz, CDCl.sub.3) δ 8.14 (s, 1H), 7.52-7.47 (m, 2H), 7.44-7.27 (m, 4H), 7.17 (d, J=2.3 Hz, 1H), 7.11 (d, J=2.6 Hz, 1H), 6.98 (dd, J=8.8, 2.4 Hz, 1H), 5.21-5.15 (m, 1H), 5.12 (s, 2H), 4.67 (dd, J=9.7, 4.9 Hz, 1H), 4.59 (dd, J=14.1, 4.9 Hz, 1H), 4.31-4.06 (m, 2H), 1.23 (t, J=9.8, 4.5 Hz, 3H). ee determination by HPLC with a Chiralcel OD-H column, hexane/2-propanol (90:10), flow rate=0.6 mL/min, 220 nm. t.sub.R=145.6 min (major), 139.2 min (minor), ≥99% ee, [α].sup.20.sub.589 −89.5 (c 0.1; MeOH). HRMS (ESI-MS) m/z: calcd: 369.1445 [MH.sup.+], found: 369.1445 [MH.sup.+]. RP-HPLC (220 nm, Method A): 100.0%, t.sub.R=21.7 min.

Ethyl (R)-3-amino-2-(5-(benzyloxy)-1H-indole-3-yl)propanoate hydrochloride ((R)-8a)

(58) ##STR00071##

(59) Preparation as described for the S-enantiomer (S)-8a by use of (R)-ethyl 2-(1H-indole-3-yl)-3-nitropropanoate ((R)-7a) (4.71 g, 12.79 mmol). Yield (3.66 g, 9.76 mmol; 76%) colorless crystals. mp.: 46.7° C. IR (KBr): 3414, 2906, 1720, 1484 cm.sup.−1. .sup.1H NMR (300 MHz, DMSO-d.sub.6) δ 11.10 (d, J=2.2 Hz, 1H), 8.12 (s, 2H), 7.52-7.29 (m, J=12.7, 9.7, 7.8, 2.5 Hz, 6H), 7.26 (d, J=2.5 Hz, 1H), 7.22 (d, J=2.3 Hz, 1H), 6.86 (dd, J=8.8, 2.4 Hz, 1H), 5.10 (s, 2H), 4.29-4.21 (m, 1H), 4.09 (q, J=10.8, 7.1 Hz, 2H), 3.60 (dd, J=8.7, 4.6 Hz, 1H), 3.19-3.07 (m, 1H), 1.13 (t, J=7.1 Hz, 3H). [α].sup.2.sub.589 77.5 (c 0.1; MeOH). HRMS (C.sub.20H.sub.22N.sub.2O.sub.3. ESI-MS) m/z: calcd.: 361.1523 [MNa.sup.+], found: 361.1519 [MNa.sup.+]. RP-HPLC (220 nm, Method A): 100.0%, t.sub.R=21.7 min.

Ethyl (R)-6-(benzyloxy)-2,3,4,9-tetrahydro-1H-pyrido[3,4-b]indole-4-carboxylate hydrochloride ((R)-9a)

(60) ##STR00072##

(61) Preparation as described for the S-enantiomer (S)-9a by use of ethyl (R)-3-amino-2-(5-(benzyloxy)-1H-indole-3-yl)propanoate hydrochloride ((R)-8a) (3.06 g, 8.16 mmol). Yield (2.20 g, 5.69 mmol, 70%) colorless crystals. mp.: 266.4-267.0° C. IR (KBr): 2921; 2793; 1719 cm.sup.−1. .sup.1H NMR (400 MHz, DMSO-d.sub.6) δ 11.20 (s, 1H), 10.20 (s, 1H), 9.12 (s, J=430.8 Hz, 1H), 7.47-7.26 (m, 6H), 7.12 (d, J=2.4 Hz, 1H), 6.85 (dd, J=8.8, 2.4 Hz, 1H), 5.07 (q, J=12.1 Hz, 2H), 4.38-4.21 (m, 2H), 4.19-4.15 (m, 1H), 4.15-4.03 (m, 2H), 3.58 (m, 2H), 1.20 (t, J=7.1 Hz, 3H). ee determination by HPLC with a Chiralcel OD-H column, hexane/2-propanol (90:10), flow rate=0.6 mL/min, 220 nm. t.sub.R=107.9 min (major), 87.4 min (minor), ≥99% ee, [α].sup.20.sub.589 128.8 (c 0.1; MeOH). HRMS (C.sub.21H.sub.22N.sub.2O.sub.3, ESI-MS) m/z: calcd: 351.1703[MH.sup.+], found: 351.1704 [MH.sup.+]. RP-HPLC (220 nm, Method A): 99.7%, t.sub.R=8.0 min.

(R)-6-(Benzyloxy)-N-methyl-2,3,4,9-tetrahydro-1H-pyrido[3,4-b]indole-4-carboxamide hydrochloride ((R)-67a)

(62) ##STR00073##

(63) Preparation as described for the S-enantiomer (S)-67a by use of ethyl (R)-6-(benzyloxy)-2,3,4,9-tetrahydro-1H-pyrido[3,4-b]indol-4-carboxylat hydrochlorid ((R)-9a) (0.40 g, 1.03 mmol. Yield (0.135 g, 0.36 mmol; 35%) colorless crystals. mp.: 156.7° C. IR (KBr): 2933; 1643 cm.sup.−1. .sup.1H NMR (400 MHz, DMSO-d.sub.6) δ 10.65 (s, 1H), 7.92 (m, 1H), 7.46-7.27 (m, 5H), 7.16 (d, J=8.7 Hz, 1H), 7.00 (d, J=2.3 Hz, 1H), 6.73 (dd, J=8.7, 2.4 Hz, 1H), 5.03 (d, J=4.2 Hz, 2H), 3.91-3.78 (m, 2H), 3.49-3.42 (m, 1H), 3.13 (dd, J=12.9, 4.1 Hz, 1H), 2.96 (dd, J=12.9, 4.8 Hz, 1H), 2.57 (d, J=4.6 Hz, 3H). [ ].sup.20.sub.589 36.9 (c 0.1; MeOH). HRMS (C.sub.20H.sub.21N.sub.3O.sub.2, ESI-MS) m/z: calcd.: 336.1707 [MH.sup.+], found: 336.1706 [MH.sup.+]. RP-HPLC (220 nm, Method A): 99.2%, t.sub.R=6.2 min.

(6R)-8-(Benzyloxy)-4-methyl-6,11-dihydro-2,6-methano[1,3]diazocino[5,6-b]indole-3,5(1H,4H)-dione ((R)-12)

(64) ##STR00074##

(65) Preparation as described for the S-enantiomer (S)-12 by use of (R)-6-(Benzyloxy)-N-methyl-2,3,4,9-tetrahydro-1H-pyrido[3,4-b]indole-4-carboxamide hydrochloride ((R)-67a) (0.136 g, 0.37 mmol). Yield (0.094 g; 0.26 mmol, 70%) beige crystals. mp.: 236.1-236.4° C. IR (KBr): 2926, 1729, 1672 cm.sup.−1. .sup.1H NMR (400 MHz, CDCl.sub.3) δ 8.03 (s, 1H, NH (Indol)), 7.51-7.47 (m, 2H, Ar—H), 7.42-7.29 (m, 3H, Ar—H), 7.23-7.19 (m, 2H, Ar—H), 6.92 (dd, J=8.8, 2.4 Hz, 1H, Ar—H), 5.14-5.05 (m, 2H, NCH.sub.2Ar (Benzyl)), 4.87 (d, J=16.3 Hz, 1H, NCH.sub.AH.sub.BC), 4.50 (d, J=16.3 Hz, 1H, NCH.sub.AH.sub.BC), 3.90 (d, J=13.1 Hz, 1H, NCH.sub.AH.sub.BCH), 3.83 (s, 1H, NCH.sub.AH.sub.BCH), 3.37 (dd, J=13.1, 2.2 Hz, 1H, NCH.sub.AH.sub.BCH), 3.07 (s, 3H, NCH.sub.3).

(66) .sup.13C NMR (101 MHz, CDCl.sub.3) δ 173.25 (quart.CO), 160.96 (quart.CO), 154.05 (quart. Ar—C), 137.42 (quart. Ar—C), 132.49 (quart. Ar—C), 130.92 (quart. Ar—C), 128.55 (+, (Ar—CH).sub.2), 127.87 (+, (Ar—CH).sub.2), 127.71 (+, Ar—CH), 126.62 (quart. Ar—C), 113.65 (+, Ar—CH), 112.06 (+, Ar—CH), 106.49 (quart. Ar—C), 101.46 (+, Ar—CH), 70.85 (−, NCH.sub.2Ar (Benzyl)), 50.25 (−, NCH.sub.2C), 47.60 (−, NCH.sub.2CH), 36.84 (+, NCH.sub.2CH), 27.91 (+, NCH.sub.3). [α].sup.20.sub.589 30.4 (c 0.1; MeOH). HRMS (C.sub.21H.sub.19N.sub.3O.sub.3, ESI-MS) m/z: calcd: 362.1499 [MH.sup.+], found: 362.1500 [MH.sup.+]. RP-HPLC (220 nm, Method A): 99.5%, t.sub.R=18.2 min.

tert-Butyl 4-(((6R)-8-(benzyloxy)-4-methyl-3,5-dioxo-3,4,5,6-tetrahydro-2,6-methano[1,3]diazocino[5,6-b]indole-11(1H)-yl)methyl)benzoate ((R)-14)

(67) ##STR00075##

(68) Preparation as described for the S-enantiomer (S)-14 by use of (6R)-8-(benzyloxy)-4-methyl-6,11-dihydro-2,6-methano[1,3]diazocino[5,6-b]indole-3,5(1H,4H)-dione ((R)-12) (0.071 g, 0.20 mmol). Yield (0.128 g, 0.23 mmol, 87%) yellow oil. IR (KBr): 1710; 1684 cm.sup.−1. .sup.1H NMR (400 MHz, DMSO-d.sub.6) δ 7.88 (d, J=8.3 Hz, 1H), 7.80 (d, J=8.3 Hz, 2H), 7.46 (d, J=7.6 Hz, 3H), 7.37 (d, J=7.6 Hz, 1H), 7.30-7.28 (m, 1H), 7.13 (d, J=8.3 Hz, 2H), 7.09 (d, J=2.4 Hz, 1H), 6.82 (dd, J=8.9, 2.4 Hz, 1H), 5.39 (s, 2H), 5.11-5.03 (m, 2H), 4.79 (d, J=16.6 Hz, 1H), 4.50 (d, J=16.5 Hz, 1H), 3.87 (d, J=12.8 Hz, 1H), 3.81 (br s, 1H), 3.42 (dd, J=13.1, 2.0 Hz, 1H), 2.89 (s, 3H), 1.50 (s, 9H). [α].sup.20.sub.589 25.0 (c 0.1; MeOH). HRMS (C.sub.33H.sub.33N.sub.3O.sub.5, ESI-MS) m/z: calcd: 552.2493 [MH.sup.+], found: 5522491 [MH.sup.+]. RP-HPLC (220 nm, Method A): 98.7%, t.sub.R=7.0 min.

tert-Butyl 4-(((6R)-8-hydroxy-4-methyl-3,5-dioxo-3,4,5,6-tetrahydro-2,6-methano[1,3]diazocino[5,6-b]indole-11(1B)-yl)methyl)benzoate ((R)-15)

(69) ##STR00076##

(70) Preparation as described above for the S-Enantiomer (S)-15 by use of tert-Butyl 4-(((6R)-8-(benzyloxy)-4-methyl-3,5-dioxo-3,4,5,6-tetrahydro-2,6-methano[1,3]diazocino[5,6-b]indole-1(1H)-yl)methyl)benzoate ((R)-14) (0.138 g, 0.25 mmol). Yield (0.068 g, 0.15 mmol, 60%) colorless crystals. mp.: 105.9° C. IR (KBr): 2927, 1713, 1665 cm.sup.−1. .sup.1H NMR (400 MHz, DMSO-d.sub.6) δ 8.86 (s, 1H), 7.80 (d, J=8.3 Hz, 2H), 7.21-7.10 (m, 3H), 6.85 (d, J=2.2 Hz, 1H), 6.57 (dd, J=8.8, 2.3 Hz, 1H), 5.35 (d, J=14.8 Hz, 2H), 4.76 (d, J=16.5 Hz, 1H), 4.48 (d, J=16.4 Hz, 1H), 3.85 (d, J=12.7 Hz, 1H), 3.75 (d, J=11.6 Hz, 1H), 3.46-3.34 (m, 1H), 2.88 (s, 3H), 1.50 (s, 9H). [α].sup.20.sub.589 −96.8 (c 0.1; MeOH). MS(C.sub.26H.sub.27N.sub.3O.sub.5, LC-MS, ESI) m/z (%): 462 [MH.sup.+] (26), 406 [MH.sup.+—C.sub.4H.sub.8] (100), 479 [MNH.sub.4.sup.+](83). Anal. (C.sub.26H.sub.27N.sub.3O.sub.5) calcd.: C, 67.66; H, 5.90; N, 9.10; found: C, 67.26; H, 6.04; N, 8,74.

tert-Butyl 4-(((6R)-4-methyl-8-(2-morpholinoethoxy)-3,5-dioxo-3,4,5,6-tetrahydro-2,6-methano[1,3]diazocino[5,6-b]indole-11(1H)-yl)methyl)benzoate ((R)-17)

(71) ##STR00077##

(72) Preparation as described for the S-enantiomer (S)-17 by use of tert Butyl 4-(((6R)-8-hydroxy-4-methyl-3,5-dioxo-3,4,5,6-tetrahydro-2,6 methano[1,3]diazocino[5,6-b]indole-11(1H)-yl)methyl)benzoate ((R)-15) (0.157 g, 0.34 mmol). The product was purified by cc (SiO.sub.2; CH.sub.2Cl.sub.2, EtOAc 3:1) and (SiO.sub.2; CH.sub.2Cl.sub.2, MeOH 10:1) (dry-load-technique). Yield 0.161 g (0.28 mmol, 82%) yellow oil. IR (KBr): 2927; 1712; 1688 cm.sup.−1. .sup.1H NMR (400 MHz, CDCl.sub.3) δ 7.91 (d, J=8.3 Hz, 2H, Ar—H), 7.15 (d, J=2.4 Hz, 1H, Ar—H), 7.09 (d, J=8.9 Hz, 1H, Ar—H), 7.03 (d, J=8.3 Hz, 2H, Ar—H), 6.83 (dd, J=8.9, 2.4 Hz, 1H, Ar—H), 5.33-5.24 (m, 1H, NCH.sub.AH.sub.BAr), 5.09 (d, J=17.0 Hz, 1H, NCH.sub.AH.sub.BAr), 4.84 (d, J=16.4 Hz, 1H, NCH.sub.AH.sub.BC), 4.32-4.17 (m, 3H, NCH.sub.AH.sub.BC, OCH.sub.2CH.sub.2), 3.89 (d, J=13.1 Hz, 1H, NCH.sub.AH.sub.BCH), 3.85 (br s, 1H, NCH.sub.2CH), 3.85-3.77 (m, 4H, O(CH.sub.2).sub.2CH.sub.2N-morpholine), 3.32 (dd, J=13.1, 2.2 Hz, 1H, NCH.sub.AH.sub.BCH), 3.06 (s, 3H, NCH.sub.3), 2.95 (s, 2H, OCH.sub.2CH.sub.2), 2.75 (br s, 4H, O(CH.sub.2).sub.2(CH.sub.2).sub.2N-morpholine), 1.56 (s, 9H, OC(CH.sub.3).sub.3).

(73) .sup.13C NMR (101 MHz, CDCl.sub.3) δ 173.12 (quart.CO), 165.16 (quart.CO), 161.25 (quart.CO), 153.65 (quart.O—Ar—C), 141.01 (quart. Ar—C), 133.83 (quart. Ar—C), 131.93 (quart. Ar—C), 131.79 (quart. Ar—C), 130.20 (+, (Ar—CH).sub.2), 126.49 (quart. Ar—C), 126.07 (+, (Ar—CH).sub.2), 113.15 (+, Ar—CH), 110.46 (+, Ar—CH), 105.98 (quart. Ar—C), 101.53 (+, Ar—CH), 81.27 (quart.OC(CH.sub.3).sub.3), 66.42 (−, O(CH.sub.2).sub.2CH.sub.2N-morpholine), 65.87 (−, OCH.sub.2CH.sub.2), 57.61 (−, OCH.sub.2CH.sub.2), 53.83 (−, O(CH.sub.2).sub.2(CH.sub.2).sub.2N-morpholine), 49.46 (−, NCH.sub.2C),), 47.36 (−, NCH.sub.2CH), 47.13 (−, NCH.sub.2Ar), 36.86 (+, NCH.sub.2CH), 28.16 (+, OC(CH.sub.3).sub.3), 27.91 (+, NCH.sub.3). [α].sup.20.sub.589 −46.8 (c 0.1; MeOH). HRMS (C.sub.32H.sub.38N.sub.4O.sub.6, ESI-MS) m/z: calcd.: 575.2864 [MH.sup.+], found: 575.2866 [MH.sup.+]. RP-HPLC (220 nm, Method A): 98.5%, t.sub.R=11.4 min.

4-(2-(((6R)-11-(4-Carboxybenzyl)-4-methyl-3,5-dioxo-1,3,4,5,6,11-hexahydro-2,6-methano[1,3]diazocino[5,6-b]indole-8-yl)oxy)ethyl)morpholine-4-ium 2,2,2-trifluoroacetate ((R)-18)

(74) ##STR00078##

(75) Preparation as described for the S-enantiomer (S)-18 by use of tert-butyl 4-(((6R)-4-methyl-8-(2-morpholinoethoxy)-3,5-dioxo-3,4,5,6-tetrahydro-2,6-methano[1,3]diazocino[5,6-b]indole-11(1H)-yl)methyl)benzoate ((R)-17) (0.033 g, 0.06 mmol). Yield (0.05 g, 0.08 mmol; 99%) brown oil. IR (ATR): 2925; 1723; 1684 cm.sup.−1. .sup.1H NMR (400 MHz, DMSO-d.sub.6) δ 9.93 (s, 1H), 7.84 (d, J=8.3 Hz, 2H), 7.34 (d, J=8.9 Hz, 1H), 7.10 (m, 3H), 6.83 (dd, J=8.9, 2.4 Hz, 1H), 5.41 (s, 2H), 4.81 (d, J=16.6 Hz, 1H), 4.53 (d, J=16.6 Hz, 1H), 4.00-3.96 (m, 3H), 3.89 (d, J=12.6 Hz, 3H), 3.82 (s, 2H), 3.74-3.65 (m, 3H), 3.58 (m, 2H), 3.50 (m, 1H), 3.44 (d, J=11.1 Hz, 1H), 2.88 (s, 3H). .sup.19F NMR (377 MHz, DMSO-d.sub.6) δ −73.70 (s). [α].sup.20.sub.589 −11.7 (c 0.1; MeOH). HRMS (C.sub.28H.sub.30N.sub.4O.sub.6. ESI-MS) m/z: calcd: 519.2238 [MH.sup.+], found: 519.2244 [MH.sup.+]. RP-HPLC (220 nm, Method B): 100.0%, t.sub.R=10.5 min.

4-(((6R)-4-Methyl-8-(2-morpholinoethoxy)-3,5-dioxo-3,4,5,6-tetrahydro-2,6-methano[1,3]diazocino[5,6-b]indole-11(1B)-yl)methyl)-N—((tetrahydro-2H-pyran-2-yl)oxy)benzamide ((R)-20)

(76) ##STR00079##

(77) Preparation as described for the S-enantiomer (S)-20 by use of 4-(2-(((6R)-11-(4-carboxybenzyl)-4-methyl-3,5-dioxo-1,3,4,5,6,11-hexahydro-2,6-methano[1,3]diazocino[5,6-b]indole-8-yl)oxy)ethyl)morpholine-4-ium 2,2,2-trifluoroacetate ((R)-18) (0.063 g, 0.10 mmol). Yield (0.042 g, 0.07 mmol, 70%) yellow oil. IR(ATR): 1727; 1683 cm.sup.−1. .sup.1H NMR (400 MHz, DMSO-d.sub.6) δ 11.56 (s, 1H), 8.15 (s, 1H), 7.65 (d, J=8.2 Hz, 2H), 7.28 (d, J=8.9 Hz, 1H), 7.10 (d, J=8.3 Hz, 2H), 7.01 (m, 1H), 6.76 (d, J=8.7 Hz, 1H), 5.37 (d, J=17.7 Hz, 2H), 4.93 (m, 1H), 4.80 (d, J=16.7 Hz, 1H), 4.51 (d, J=16.6 Hz, 1H), 4.13 (m, 2H), 4.00 (m, 1H), 3.87 (d, J=12.7 Hz, 1H), 3.81 (m, 1H), 3.63-3.56 (m, 4H), 3.49 (m, 1H), 3.42 (d, J=13.4 Hz, 1H), 3.16-3.08 (m, 4H), 2.87 (s, 3H), 2.77-2.66 (m, 2H), 1.67-1.51 (m, 6H). ee determination by HPLC with a Phenomenex Lux Cellulose-2 column, methanol/2-propanol (90:10), flow rate=1.0 mL/min, 220 nm. t.sub.R=33.8-37.0 min (major), 18.7-23.4 min (minor), 94% ee, [α].sup.20.sub.589 −32.0 (c 0.1; MeOH). HRMS (C.sub.33H.sub.39N.sub.5O.sub.7, ESI-MS) m/z: calcd: 618.2922 [MH.sup.+], found: 618.2928 [MH.sup.+]. RP-HPLC (220 nm, Method B): 97.8%, t.sub.R=11.4 min.

N-Hydroxy-4-(((6R)-4-methyl-8-(2-morpholinoethoxy)-3,5-dioxo-3,4,5,6-tetrahydro-2,6-methano[1,3]diazocino[5,6-b]indole-11(1B)-yl)methyl)benzamide ((R)-21)

(78) ##STR00080##

(79) Preparation as described for the S-enantiomer (S)-21 by use of 4-(((6R)-4-Methyl-8-(2-morpholinoethoxy)-3,5-dioxo-3,4,5,6-tetrahydro-2,6-methano[1,3]diazocino[5,6-b]indol-11(1H)-yl)methyl)-N—((tetrahydro-2H-pyran-2-yl)oxy)benzamid ((R)-20) (0.044 g, 0.07 mmol). Yield (0.029 g, 0.05 mmol, 71%) colorless crystals. mp.: 280.8° C. IR (KBr): 3415; 1724; 1679 cm.sup.−1. .sup.1H NMR (400 MHz, DMSO-d.sub.6) δ 11.18 (s, 1H, NHOH), 9.00 (s, 1H, NHOH), 7.65 (d, J=8.3 Hz, 2H, Ar-H), 7.33 (d, J=7.8 Hz, 1H, Ar—H), 7.09 (d, J=8.3 Hz, 2H, Ar—H), 7.05 (d, J=2.4 Hz, 1H, Ar—H), 6.82 (dd, J=8.9, 2.4 Hz, 1H, Ar—H), 5.36 (s, 2H, NCH.sub.2Ar), 4.82 (d, J=16.6 Hz, 1H, NCH.sub.AH.sub.BC), 4.54 (dd, J=16.2, 8.7 Hz, 1H, NCH.sub.AH.sub.BC), 4.44-4.33 (m, 2H, OCH.sub.2CH.sub.2), 3.96-3.79 (m, 6H, OCH.sub.2CH.sub.2N-morpholine, NCH.sub.AH.sub.BCH, OCH.sub.2CH.sub.2N-morpholine, NCH.sub.2CH), 3.55-3.42 (m, 5H, OCH.sub.2CH.sub.2N, OCH.sub.2CH.sub.2N-morpholine, NCH.sub.AH.sub.BCH), 3.20 (s, J=16.3 Hz, 2H, OCH.sub.2CH.sub.2N-morpholine), 2.89 (s, 3H, NCH.sub.3). .sup.13C NMR (101 MHz, DMSO) δ 173.58 (quart.CO), 164.16 (quart.CO), 161.31 (quart.CO), 152.68 (quart.O—Ar—C), 141.27 (quart. Ar—C), 135.62 (quart. Ar—C), 132.33 (quart. Ar—C) 131.99 (quart. Ar—C) 127.71 (−, Ar—CH).sub.2), 127.02 (−, Ar—CH).sub.2), 126.61 (quart. Ar—C), 112.22 (−, Ar—CH), 111.75 (−, Ar—CH), 105.50 (quart. Ar—C), 102.12 (−, Ar—CH), 63.63 (+, O(CH.sub.2).sub.2CH.sub.2N-morpholine), 63.32 (+, OCH.sub.2CH.sub.2), 55.59 (+, OCH.sub.2CH.sub.2), 52.14 (+, O(CH.sub.2).sub.2(CH.sub.2).sub.2N-morpholine), 49.44 (+, NCH.sub.2C), 46.67 (+, NCH.sub.2CH), 46.57 (+, NCH.sub.2Ar), 36.41 (−, NCH.sub.2CH), 27.88 (−, NCH.sub.3). [α].sup.20.sub.589 −14.0 (c 0.1; H.sub.2O). HRMS (C.sub.28H.sub.31N.sub.5O.sub.6, ESI-MS) m/z: calcd: 534.2347 [MH.sup.+], found: 534.2354 [MH.sup.+]. RP-HPLC (220 nm, Method B): 95.0%, t.sub.R=8.5 min.

Example 2: Synthesis of Compounds 31a and 31b

(80) ##STR00081## ##STR00082## ##STR00083##

(81) To obtain a compound 53a with core structure 31a, which possess an additional solubility enhancing substructure, this part can be introduced as shown in scheme 2b by use of 5-benzyloxy-indole-2-carbaldehyde 42 instead of 22.

(82) ##STR00084## ##STR00085## ##STR00086##

(83) ##STR00087## ##STR00088##

(84) To obtain a compound 53b with core structure 31b, which possess an additional solubility enhancing substructure, this part can be introduced as shown in scheme 3b by use of 27 instead of 22.

(85) ##STR00089## ##STR00090##

tert-Butyl 4-((2-formyl-1H-indol-1-yl)methyl)benzoate (23)

(86) ##STR00091##

(87) Under nitrogen a solution of the indole-2-carbaldehyde (18.5 g; 127.4 mmol) in DMF (300 mL) was cooled to 0° C. After addition of NaH (1.1 equ; 60% in paraffin) the mixture was stirred for 10 min. The alkylating agent, tert-butyl 4-(bromomethyl)benzoate (1.1 equ.) was added and stirring at rt continued until completion of the reaction (TLC). The mixture was poured into water. The crude product was isolated by extraction of the aqueous phase with Et.sub.2O (4×250 mL). Silica gel chromatography (CH.sub.2Cl.sub.2/light petrol 1:1) afforded the desired product. Yield 27.16 g (80.98 mmol, 64%) yellow crystals after cc (CH.sub.2Cl.sub.2) from CH.sub.2Cl.sub.2/light petrol; mp.: 131.7-133.1° C., IR (KBr): 1709, 1671 cm.sup.−1; .sup.1H NMR (300 MHz, CDCl.sub.3): δ 9.90 (s, 1H), 7.92-7.84 (m, 2H), 7.78 (d, J=8.1 Hz, 1H), 7.42-7.29 (m, 3H), 7.20 (ddd, J=8.0, 6.6, 1.3 Hz, 1H), 7.09 (d, J=8.4 Hz, 2H), 5.88 (s, 2H), 1.54 (s, 9H); HRMS (ESI-MS) m/z: calc.: 336.1594 [MH.sup.+], found: 336.1603 [MH.sup.+], Anal. calcd. for C.sub.21H.sub.21NO.sub.3: C, 75.20; H, 6.31; N, 4.18; found C, 75.07; H, 6.33; N, 4.09.

(E)-tert-Butyl 4-((2-(2-nitrovinyl)-1H-indol-1-yl)methyl)benzoate (24)

(88) ##STR00092##

(89) (E)-tert-butyl 4-((2-(2-nitrovinyl)-1H-indol-1-yl)methyl)benzoate (24) was prepared according to a modified literature procedure (Yang, Li et al. 2016) as follows: A solution of tert-butyl 4-((2-formyl-1H-indol-1-yl)methyl)benzoate (23) (24.55 g; 73.20 mmol) and ammonium acetate (2.82 g, 36.6 mmol) in nitromethane (250 mL) was heated at reflux for 10 h under nitrogen. After removal of half of the solvent the mixture was cooled, the crystalline precipitating product removed by filtration and crystallized from ethanol. Yield 17.50 g (46.2 mmol, 63%) yellow crystals from ethanol; mp.: 196.3-199.5° C.; IR (KBr): 1704, 1632 cm.sup.−1; .sup.1H NMR (300 MHz, CDCl.sub.3): δ 8.03 (d, J=13.3 Hz, 1H), 7.92 (d, J=8.3 Hz, 2H), 7.70 (d, J=8.0 Hz, 1H), 7.57 (d, J=13.4 Hz, 1H), 7.37-7.28 (m, 2H), 7.19 (td, J=6.2, 1.8 Hz, 2H), 7.04 (d, J=8.4 Hz, 2H), 5.53 (s, 2H), 1.56 (s, 9H). HRMS (ESI-MS) m/z: calcd.: 379.1652 [MH.sup.+], found: 379.1657 [MH.sup.+]. Anal. calcd. for C.sub.22H.sub.22N.sub.2O.sub.4: C, 69.83; H, 5.86; N, 7.40, found: C, 69.81; H, 5.92; N, 7.42.

tert-Butyl 4-((2-(2-nitroethyl)-1H-indol-1-yl)methyl)benzoate (25)

(90) ##STR00093##

(91) To a solution of (E)-tert-butyl 4-((2-(2-nitrovinyl)-1H-indol-1-yl)methyl)benzoate (24) (15.0 g; 39.7 mmol) in CHCl.sub.3 (750 mL) and .sup.ipropanol (75 mL) silica gel (75 g) and NaBH.sub.4 (47.6 mmol; 1.80 g) were added and the mixture stirred over night at room temperature. Water (100 mL) was added dropwise while stirring, the mixture filtered over a pad of celite, the organic layer dried (Na.sub.2SO.sub.4) and purified by cc (SiO.sub.2; CH.sub.2Cl.sub.2). Yield 11.18 g (29.4 mmol, 74%) yellow crystals from CH.sub.2Cl.sub.2; mp.: 150.4-152.2° C.; IR (KBr): 1704, 1555 cm.sup.−1; .sup.1H NMR (300 MHz, CDCl.sub.3) δ 7.94-7.87 (m, 2H), 7.60 (dd, J=6.5, 1.7 Hz, 1H), 7.23-7.09 (m, 3H), 6.98 (d, J=8.3 Hz, 2H), 6.41 (s, 1H), 5.40 (s, 2H), 4.63 (t, J=7.3 Hz, 2H), 3.38 (t, J=7.3 Hz, 2H), 1.56 (s, 9H). HRMS (ESI-MS) m/z: calcd.: 381.1809 [MH.sup.+], found: 381.1814 [MH.sup.+]; Anal. calcd. for C.sub.22H.sub.24N.sub.2O.sub.4: C, 69.46; H, 6.36; N, 7.36; found: C, 69.29; H, 6.33; N, 7.19.

tert-Butyl 4-((2-(2-aminoethyl)-1H-indol-1-yl)methyl)benzoate hydrochloride (26)

(92) ##STR00094##

(93) To a stirred solution of tert-butyl 4-((2-(2-nitroethyl)-1H-indol-1-yl)methyl)benzoate (25) (10.80 g; 28.4 mmol) in HOAc (108 mL) zinc dust (170 mmol; 11.13 g) was added in small portions at 20° C. After 4 h, ice was added (250 g) and the mixture alkalized with aqueous ammonia (25%) till pH=14. The mixture was filtered, the aqueous layer extracted with ethyl acetate (3×100 mL), the combined organic layers dried (Na.sub.2SO.sub.4) and the solvent removed under reduced pressure. The remaining solid was dissolved in THF (10 mL), the solution cooled to 0° C. and HCl (5-6N in .sup.ipropanol) was added dropwise till pH=2. Et.sub.2O was added whilst stirring, the precipitating hydrochloride filtered off and washed with Et.sub.2O. Yield 9.60 g (24.8 mmol, 84%) colorless crystals; mp.: 208.9-210.2° C.; IR (KBr): 3446, 1718, 1506 cm.sup.−1; .sup.1H NMR (300 MHz, DMSO) δ 7.86 (s, 2H), 7.82 (d, J=8.3 Hz, 2H), 7.54 (d, J=6.7 Hz, 1H), 7.35 (d, J=7.7 Hz, 1H), 7.10-6.99 (m, 4H), 6.46 (s, 1H), 5.54 (s, 2H), 3.08 (d, J=7.3 Hz, 2H), 2.99 (d, J=7.4 Hz, 2H), 1.51 (s, 9H). HRMS (ESI-MS) m/z: calcd.: 351.2067[MH.sup.+], found: 351.2075[MH.sup.+]; Anal. calcd. for C.sub.22H.sub.27ClNO.sub.2: C, 68.29; H, 7.03; N, 7.24; found: C, 68.08; H, 7.04; N, 7.07.

Ethyl 5-(4-(tert-butoxycarbonyl)benzyl)-2,3,4,5-tetrahydro-1H-pyrido[4,3-b]indole-1-carboxylate (27)

(94) ##STR00095##

(95) To a stirred solution of tert-butyl 4-((2-(2-aminoethyl)-1H-indol-1-yl)methyl)benzoate hydrochloride (26) (9.00 g; 23.3 mmol) in MeOH (200 mL) ethyl glyoxalate (5.56 mL; 50% in toluene) and silica gel (18.0 g) were added and the mixture stirred for 1 h. The solvents were removed under reduced pressure and the product purified by cc (SiO.sub.2; CH.sub.2Cl.sub.2, MeOH, NH.sub.3 (25%), 10:1:0.1) (dry load method). Yield 9.90 g (22.8 mmol, 98%) yellow foam; IR (KBr): 3049, 2931, 1733, 1712 cm.sup.−1; .sup.1H NMR (300 MHz, CDCl.sub.3): δ 7.89 (d, J=8.3 Hz, 2H), 7.79-7.72 (m, 1H), 7.21-7.08 (m, 3H), 7.03 (d, J=8.3 Hz, 2H), 5.33 (d, J=17.6 Hz, 1H), 5.26 (d, J=17.4 Hz, 1H), 4.94 (s, 1H), 4.35-4.10 (m, 2H), 3.49 (ddd, J=13.0, 9.1, 4.7 Hz, 1H), 3.24 (ddd, J=12.5, 5.4, 4.0 Hz, 1H), 2.81-2.53 (m, 1H), 2.38 (s, 1H), 1.56 (s, 9H), 1.32 (t, J=7.1 Hz, 3H). HRMS (ESI-MS) m/z: calcd.: 435.2278 [MH.sup.+], found: 435.2285 [MH.sup.+]; Anal. calcd. for C.sub.26H.sub.30N.sub.2O.sub.4+0.5 H.sub.2O: C, 70.41; H, 7.04; N, 6.32, found: C, 70.34; H, 6.79; N, 6.24.

tert-Butyl 4-((2-methyl-1,3-dioxo-2,3,5,6-tetrahydro-1H-imidazo[1′,5′:1,2]pyrido[4,3-b]indol-7(11cH)-yl)methyl)benzoate (28a)

(96) ##STR00096##

(97) Ethyl 5-(4-(tert-butoxycarbonyl)benzyl)-2,3,4,5-tetrahydro-1H-pyrido[4,3-b]indole-1-carboxylate (27) (3.00 g; 6.90 mmol) was dissolved in acetonitrile (15.0 mL). Diisopropylethylamine (3.0 mL) was added to the stirred mixture. After addition of N-succinimidyl-N-methylcarbamate (1.31 g; 7.60 mmol) stirring was continued for 16 h at rt. The mixture was poured into water and the crude product was extracted with EtOAc (3×50 mL). The combined organic layers were dried (Na.sub.2SO.sub.4) and evaporated. Yield 1.44 g (3.23 mmol, 47%) colorless foam after cc (CH.sub.2Cl.sub.2, ethyl acetate 10:1). .sup.1H NMR (300 MHz, CDCl.sub.3): δ 8.12-8.01 (m, 1H), 7.84 (d, J=8.3 Hz, 2H), 7.19-7.08 (m, 3H), 6.96 (d, J=8.3 Hz, 2H), 5.34 (t, J=1.8 Hz, 1H), 5.24 (s, 2H), 4.48 (dd, J=13.7, 6.0 Hz, 1H), 3.20-3.05 (m, 1H), 2.97 (s, 3H), 2.88-2.71 (m, 1H), 2.56 (dd, J=16.0, 4.7 Hz, 1H), 1.51 (s, 9H).

4-((2-Methyl-1,3-dioxo-2,3,5,6-tetrahydro-1H-imidazo[1′,5′:1,2]pyrido[4,3-b]indol-7(11cH)-yl)methyl)benzoic acid (29a)

(98) ##STR00097##

(99) tert-Butyl 4-((2-methyl-1,3-dioxo-2,3,5,6-tetrahydro-1H-imidazo[1′,5′:1,2]pyrido[4,3-b]indol-7(11cH)-yl)methyl)benzoate (28a) (1.00 g; 2.24 mmol) was dissolved in trifluoro acetic acid (10.0 mL) and the mixture stirred for 15 min. at room temperature. The solution was added to water (100 mL), the precipitating product collected by filtration and dried in vacuo. mp.: 267.2-269.6° C.; IR (KBr): 1717, 1678 cm.sup.−1; .sup.1H NMR (300 MHz, DMSO) δ 12.91 (s, 1H), 7.94-7.89 (m, 1H), 7.86 (d, J=8.2 Hz, 2H), 7.43 (d, J=7.1 Hz, 1H), 7.15 (d, J=8.2 Hz, 2H), 7.13-7.05 (m, 2H), 5.52 (s, 1H), 5.46 (s, 2H), 4.33 (dd, J=13.6, 4.9 Hz, 1H), 3.28-3.15 (m, 2H), 2.86 (s, 3H), 2.78 (s, 1H). HRMS (ESI-MS) m/z: calcd.: 390.1361 [MH.sup.+], found: 390.1360[MH.sup.+]; Anal. calcd. for C.sub.22H.sub.19N.sub.3O.sub.4+0.5 H.sub.2O: C, 66.32; H, 5.06; N, 10.55; found: C, 66.01; H, 5.09; N, 10.19.

4-((2-Methyl-1,3-dioxo-2,3,5,6-tetrahydro-1H-imidazo[1′,5′:1,2]pyrido[4,3-b]indol-7(11cH)-yl)methyl)-N—((tetrahydro-2H-pyran-2-yl)oxy)benzamide (30a)

(100) ##STR00098##

(101) 4-((2-Methyl-1,3-dioxo-2,3,5,6-tetrahydro-1H-imidazo[1′,5′:1,2]pyrido[4,3-b]indol-7(11cH)-yl)methyl)benzoic acid (29a) (0.78 g, 2.00 mmol) was dissolved in DMF (15.0 mL) and BOP (1.2 equ.), EtN(iProp).sub.2 (0.78 mL) and O-(tetrahydro-2H-pyran-2-yl)hydroxylamine (3.0 equ.) were added. The solution was stirred over night at room temperature, poured into water and extracted with ethyl acetate (3×50 mL). CC (SiO.sub.2, CH.sub.2Cl.sub.2, MeOH (10:1) and removal of the solvent under reduced pressure yielded the product as colorless foam; mp.: 184.7-186.9° C.; .sup.1H NMR (300 MHz, DMSO) δ 11.58 (s, 1H), 7.91 (dd, J=6.6, 2.1 Hz, 1H), 7.67 (d, J=8.2 Hz, 2H), 7.43 (d, J=7.2 Hz, 1H), 7.16-7.04 (m, 4H), 5.52 (s, 1H), 5.44 (s, 2H), 4.95 (s, 1H), 4.33 (dd, J=13.5, 5.1 Hz, 1H), 3.49 (d, J=11.3 Hz, 1H), 3.28-3.14 (m, 1H), 2.87 (s, 3H), 2.76 (d, J=17.0 Hz, 2H), 2.09 (s, 1H), 1.61 (d, J=49.2 Hz, 6H). HRMS (ESI-MS) m/z: calcd.: 489.2132 [MH.sup.+], found: 489.2133 [MH.sup.+].

N-Hydroxy-4-((2-methyl-1,3-dioxo-2,3,5,6-tetrahydro-1H-imidazo[1′,5′:1,2]pyrido[4,3-b]indol-7(11cH)-yl)methyl)benzamide (31a)

(102) ##STR00099##

(103) mp.: 248.1-251.7° C.; IR (KBr): 3251, 1769, 1690, 1650 cm.sup.−1; .sup.1H NMR (300 MHz, DMSO) δ 11.14 (s, 1H), 9.02 (d, J=1.7 Hz, 1H), 7.91 (dd, J=6.7, 2.1 Hz, 1H), 7.65 (d, J=8.2 Hz, 2H), 7.44 (dd, J=6.8, 1.7 Hz, 1H), 7.16-7.09 (m, 3H), 7.08-7.03 (m, 1H), 5.52 (s, 1H), 5.42 (s, 2H), 4.33 (dd, J=13.7, 4.9 Hz, 1H), 3.29-3.14 (m, 1H), 2.87 (s, 3H), 2.77 (d, J=16.5 Hz, 2H). HRMS (ESI-MS) m/z: calcd.: 405.1557 [MH.sup.+], found: 405.1558 [MH.sup.+]; Anal. calc. for C.sub.22H.sub.20N.sub.4O.sub.4+0.5 H.sub.2O: C, 63.91; H, 5.12; N, 13.55; found: C, 64.10; H, 5.13; N, 13.19.

tert-Butyl 4-((2-methyl-1,4-dioxo-1,3,4,6,7,12c-hexahydropyrazino[1′,2′:1,2]pyrido[4,3-b]indol-8(2H)-yl)methyl)benzoate (28b)

(104) ##STR00100##

(105) Ethyl 5-(4-(tert-butoxycarbonyl)benzyl)-2,3,4,5-tetrahydro-1H-pyrido[4,3-b]indole-1-carboxylate (27) (3.55 g, 8.16 mmol) was dissolved in 50 mL of dichloromethane and cooled to −50° C. 0.68 mL (8.57 mmol) of chloroacetyl chloride and 1.55 mL (8.98 mmol) of diisopropylethylamine were added. The solution was brought to 0° C. within 1 h and the solvent was removed at this temperature under reduced pressure.

(106) 30 mL methylamine (40% in methanol) was added and the solution was stirred overnight at room temperature. 80 mL of ethyl acetate and 40 mL of water were added, the organic layer was separated, dried over Na.sub.2SO.sub.4 and the solvent removed.

(107) Yield 2.40 g; 5.22 mmol (64%) yellow foam after silica gel chromatography with 1. ethyl acetate, 2. ethyl acetate/acetonitrile 1:1. mp.: 121.1-124.0° C.; IR (KBr): 1712, 1669 cm.sup.−1; .sup.1H NMR (300 MHz, DMSO) δ 7.91 (d, J=7.4 Hz, 1H), 7.84-7.78 (m, 2H), 7.37 (d, J=7.9 Hz, 1H), 7.12 (d, J=8.3 Hz, 2H), 7.09-6.94 (m, 2H), 5.59 (s, 1H), 5.44 (t, J=10.9 Hz, 2H), 4.63 (d, J=12.1 Hz, 1H), 4.16 (d, J=17.6 Hz, 1H), 3.96 (d, J=17.8 Hz, 1H), 2.99 (dd, J=12.5, 7.8 Hz, 1H), 2.90 (s, 3H), 2.78 (d, J=5.2 Hz, 2H), 1.50 (s, 9H). HRMS (ESI-MS) m/z: calcd.: 482.2050 [MNa.sup.+], found: 482.2055[MNa.sup.+]; Anal. calcd. for C.sub.27H.sub.29N.sub.3O.sub.4+1 H.sub.2O: C, 67.91; H, 6.54; N, 8.80; found: C, 67.86; H, 6.22; N, 8.64.

4-((2-Methyl-1,4-dioxo-1,3,4,6,7,12c-hexahydropyrazino[1′,2′:1,2]pyrido[4,3-b]indol-8(2H)-yl)methyl)benzoic acid (29b)

(108) ##STR00101##

(109) tert-Butyl 4-((2-methyl-1,4-dioxo-1,3,4,6,7,12c-hexahydropyrazino[1′,2′:1,2]pyrido[4,3-b]indol-8(2H)-yl)methyl)benzoate (28b) (0.46 g, 1.00 mmol) was dissolved in 20 mL of dichloromethane. After addition of 10 mL trifluoroacetic acid stirring at rt was continued for 16 h. The solution was diluted with 40 mL of dichloromethane and the organic solution was washed with water. Removal of the solvent left a light brown solid. Yield 0.40 g (quant.). mp.: 283.3-287.4° C.; IR (KBr): 1700, 1669 cm.sup.−1; .sup.1H NMR (300 MHz, DMSO) δ 12.75 (s, 1H), 7.91 (d, J=7.9 Hz, 1H), 7.86 (d, J=8.2 Hz, 2H), 7.38 (d, J=7.9 Hz, 1H), 7.12 (d, J=8.2 Hz, 2H), 7.03 (dt, J=22.5, 7.3 Hz, 2H), 5.59 (s, 1H), 5.55-5.38 (m, 2H), 4.63 (d, J=12.1 Hz, 1H), 4.16 (d, J=17.7 Hz, 1H), 3.96 (d, J=17.7 Hz, 1H), 3.06-2.94 (m, 1H), 2.90 (s, 3H), 2.80 (s, 2H). HRMS (ESI-MS) m/z: calcd.: 402.1452 [MH].sup.−, found: 402.1452 [MH].sup.−.

4-((2-Methyl-1,4-dioxo-1,3,4,6,7,12c-hexahydropyrazino[1′,2′:1,2]pyrido[4,3-b6]indol-8(2H)-yl)methyl)-N—((tetrahydro-2H-pyran-2-yl)oxy)benzamide (30b)

(110) ##STR00102##

(111) 4-((2-Methyl-1,4-dioxo-1,3,4,6,7,12c-hexahydropyrazino[1′,2′:1,2]pyrido[4,3-b]indol-8(2H)-yl)methyl)benzoic acid (29b) (0.31 g, 0.77 mmol) was dissolved in 60 mL of tetrahydrofuran. 0.41 g (0.92 mmol) BOP, 0.52 mL 83.00 mmol) diisopropylethylamine and 0.23 g (2.00 mmol) NH.sub.2OTHP were added. The mixture was stirred at room temperature for 3.5 h. TLC-control: SiO.sub.2; dichloromethane/methanol 10:1. The solvent was removed under reduced pressure and the residue was dissolved in 100 mL of ethyl acetate and 50 mL of water. The organic layer was separated and the volume of the solvent was reduced until crystallization occurs. The crystals were filtered of, washed with diethylether and dried.

(112) Yield 0.22 g; 0.44 mmol (57%) colorless crystals. mp.: 188.7-192.8° C.; IR (KBr): 1675, 1653 cm.sup.−1; .sup.1H NMR (300 MHz, DMSO) δ 11.58 (s, 1H), 7.91 (d, J=7.7 Hz, 1H), 7.67 (d, J=8.1 Hz, 2H), 7.38 (d, J=8.0 Hz, 1H), 7.11 (d, J=8.2 Hz, 2H), 7.07-6.95 (m, 2H), 5.59 (s, 1H), 5.54-5.34 (m, 2H), 4.95 (s, 1H), 4.63 (d, J=12.2 Hz, 1H), 4.16 (d, J=17.7 Hz, 1H), 4.03 (d, J=7.2 Hz, 1H), 3.96 (d, J=17.6 Hz, 1H), 3.49 (d, J=10.7 Hz, 1H), 3.06-2.94 (m, 1H), 2.89 (s, 3H), 2.81 (s, 2H), 1.61 (d, J=49.1 Hz, 6H). HRMS (ESI-MS) m/z: calcd.: 503.2289 [MH.sup.+], found: 503.2289 [MH.sup.+]; Anal. calcd. for C.sub.28H.sub.30N.sub.4O.sub.5+0.75 H.sub.2O: C, 65.17; H, 6.15; N, 10.86; found: C, 65.11; H, 5.95; N, 10.87.

N-hydroxy-4-((2-methyl-1,4-dioxo-1,3,4,6,7,12c-hexahydropyrazino[1′,2′:1,2]pyrido[4,3-b]indol-8(2H)-yl)methyl)benzamide (31b)

(113) ##STR00103##

(114) 4-((2-Methyl-1,4-dioxo-1,3,4,6,7,12c-hexahydropyrazino[1′,2′:1,2]pyrido[4,3-b]indol-8(2H)-yl)methyl)-N—((tetrahydro-2H-pyran-2-yl)oxy)benzamide (30b) (0.18 g, 0.36 mmol) was dissolved in a mixture of 20 mL methanol, 20 mL tetrahydrofuran and 20 mL of dichloromethane. After addition of 0.2 mL of 6 N HCl in isopropanol the mixture was stirred for 45 min. at room temperature. TLC-control: SiO.sub.2; dichloromethane/methanol 10:1. The solvent was removed, ethanol was added and the mixture heated to 70° C. for 15 min. After cooling the product was filtered of and dried.

(115) Yield 0.12 g, 0.29 mmol (80%) light beige powder. mp.: 204.1-205.6° C.; IR (KBr): 1677, 1653 cm.sup.−1; .sup.1H NMR (300 MHz, DMSO) δ 11.16 (s, 1H), 9.02 (s, 1H), 7.90 (d, J=7.7 Hz, 1H), 7.66 (d, J=8.2 Hz, 2H), 7.39 (d, J=8.0 Hz, 1H), 7.08 (d, J=8.2 Hz, 3H), 7.00 (dd, J=14.7, 7.4 Hz, 1H), 5.59 (s, 1H), 5.51-5.34 (m, 2H), 4.64 (d, J=12.2 Hz, 11), 4.16 (d, J=17.7 Hz, 1H), 3.95 (d, J=17.7 Hz, 1H), 3.00 (dd, J=12.1, 7.8 Hz, 1H), 2.89 (s, 3H), 2.82 (s, 2H). HRMS (ESI-MS) m/z: calcd.: 419.1714 [MH.sup.+], found: 419.1715 [MH.sup.+].

Example 3: Synthesis of Compound 41

(116) ##STR00104## ##STR00105## ##STR00106##

(117) To obtain a compound 65 with core structure 41, which possess an additional solubility enhancing substructure, this part can be introduced as shown in scheme 4b by use of 5-benzyloxy-indole-3-carbaldehyde 54 instead of 32.

(118) ##STR00107## ##STR00108## ##STR00109##

tert-Butyl 4-((3-formyl-1H-indol-1-yl)methyl)benzoate (33)

(119) ##STR00110##

(120) Preparation from 1H-indole-3-carbaldehyde (32) (9.60 g, 66.2 mmol) and tert-butyl 4-(bromomethyl)benzoate (18.0 g, 66.4 mmol) as described for 23.

(121) Yield 18.2 g (52 mmol, 78%) colorless solid. mp.: 112.7-114.9° C.; IR(KBr): 1704, 1664 cm.sup.−1; .sup.1H NMR (300 MHz, CDCl.sub.3) δ 10.02 (s, 1H), 8.34 (dd, J=6.2, 2.0 Hz, 1H), 7.96 (d, J=8.3 Hz, 2H), 7.74 (s, 1H), 7.36-7.26 (m, 3H), 7.20 (d, J=8.3 Hz, 2H), 5.42 (s, 2H), 1.57 (s, 9H).

(E)-tert-Butyl 4-((3-(2-nitrovinyl)-1H-indol-1-yl)methyl)benzoate (34)

(122) ##STR00111##

(123) Preparation from tert-Butyl 4-((3-formyl-1H-indol-1-yl)methyl)benzoate (33) (10.0 g, 29.8 mmol) as described for 24. Yield 11.17 g (29.5 mmol, 99%) yellow crystals. mp.: 144.2-146.7° C.; IR (KBr): 1714, 1619, 1320 cm.sup.−1; .sup.1H NMR (300 MHz, CDCl.sub.3) δ 8.26 (d, J=13.5 Hz, 1H), 7.96 (d, J=8.4 Hz, 2H), 7.83-7.78 (m, 1H), 7.76 (s, 1H), 7.58 (s, 1H), 7.38-7.28 (m, 3H), 7.18 (d, J=8.4 Hz, 2H), 5.41 (s, 2H), 1.57 (s, 9H).

tert-Butyl 4-((3-(2-nitroethyl)-1H-indol-1-yl)methyl)benzoate (35)

(124) ##STR00112##

(125) Preparation from (E)-tert-Butyl 4-((3-(2-nitrovinyl)-1H-indol-1-yl)methyl)benzoate (34) (10.5 g, 27.7 mmol) as described for 25.

(126) Yield 6.30 g (16.6 mmol, 59%) light yellow oil after cc (SiO.sub.2, CH.sub.2Cl.sub.2). IR (KBr): 1707, 1549, 1311 cm.sup.−1; .sup.1H NMR (300 MHz, CDCl.sub.3) δ 7.95-7.88 (m, 2H), 7.61-7.56 (m, 1H), 7.23-7.12 (m, 3H), 7.10 (d, J=8.4 Hz, 2H), 6.99 (s, 1H), 5.32 (s, 2H), 4.67 (t, J=7.2 Hz, 2H), 3.50 (t, J=7.1 Hz, 2H), 1.56 (s, 9H).

tert-Butyl 4-((3-(2-aminoethyl)-1H-indol-1-yl)methyl)benzoate hydrochloride (36)

(127) ##STR00113##

(128) Preparation from tert-Butyl 4-((3-(2-nitroethyl)-1H-indol-1-yl)methyl)benzoate (35) (6.0 g, 15.8 mmol) as described for 26. Yield 3.60 g (10.3 mml, 65%) colorless crystals. mp.: 196.9-198.5° C.; IR (KBr): 1714, 1297 cm.sup.−1; .sup.1H NMR (300 MHz, CDCl.sub.3) δ 8.27 (s, 2H), 7.88 (d, J=8.3 Hz, 2H), 7.61 (d, J=6.8 Hz, 1H), 7.15-7.02 (m, 6H), 5.26 (s, 2H), 3.23 (s, 4H), 1.54 (s, 9H).

Ethyl 9-(4-(tert-butoxycarbonyl)benzyl)-2,3,4,9-tetrahydro-1H-pyrido[3,4-b]indole-1-carboxylate hydrochloride (37)

(129) ##STR00114##

(130) Preparation from tert-Butyl 4-((3-(2-aminoethyl)-1H-indol-1-yl)methyl)benzoate hydrochloride (36) (3.50 g, 9.07 mmol) as described for 27 without purification by cc.

(131) Yield 2.45 g (5.10 mmol, 56%) colorless crystals from a saturated solution in MeOH by addition of Et.sub.2O. mp.: 211.2-214.9° C.; IR (KBr): 1747, 1714, 1253 cm.sup.−1; .sup.1H NMR (300 MHz, DMF) δ 10.11 (s, 2H), 7.80 (d, J=8.3 Hz, 2H), 7.57 (d, J=6.9 Hz, 1H), 7.17 (dd, J=15.8, 6.4 Hz, 2H), 7.12-7.07 (m, 3H), 5.81 (s, 1H), 5.63 (q, J=17.5 Hz, 2H), 4.17 (tt, J=14.3, 7.1 Hz, 1H), 4.02-3.86 (m, 1H), 3.72 (d, J=12.8 Hz, 1H), 3.53 (s, 1H), 3.04 (d, J=4.8 Hz, 2H), 1.51 (s, 9H), 1.08 (t, J=7.1 Hz, 3H).

tert-Butyl 4-((2-methyl-1,3-dioxo-2,3,5,6-tetrahydro-1H-imidazo[1′,5′:1,2]pyrido[3,4-b]indol-11(11bH)-yl)methyl)benzoate (38)

(132) ##STR00115##

(133) Preparation from ethyl 9-(4-(tert-butoxycarbonyl)benzyl)-2,3,4,9-tetrahydro-1H-pyrido[3,4-b]indole-1-carboxylate hydrochloride (37) (2.45 g, 5.10 mmol) as described for 28a.

(134) Yield 2.05 g (4.60 mmol, 90%) colorless foam after cc (SiO.sub.2, ethyl acetate). mp.: 120.3-123.0° C.; IR (KBr): 1714, 1461 cm.sup.−1; .sup.1H NMR (300 MHz, CDCl.sub.3) δ 7.88 (d, J=8.4 Hz, 2H), 7.54 (d, J=7.5 Hz, 1H), 7.25-7.11 (m, 3H), 6.99 (d, J=8.4 Hz, 2H), 6.16 (d, J=17.3 Hz, 1H), 5.64 (d, J=17.2 Hz, 1H), 5.10 (s, 1H), 4.57-4.45 (m, 1H), 3.03 (s, 3H), 3.01 (d, J=7.9 Hz, 1H), 2.97-2.89 (m, 1H), 2.89-2.81 (m, 1H), 1.55 (s, 9H).

4-((2-Methyl-1,3-dioxo-2,3,5,6-tetrahydro-1H-imidazo[1′,5′:1,2]pyrido[3,4-b]indol-11(11bH)-yl)methyl)-N—((tetrahydro-2H-pyran-2-yl)oxy)benzamide (40)

(135) ##STR00116##

(136) Preparation from 4-((2-methyl-1,3-dioxo-2,3,5,6-tetrahydro-1H-imidazo[1′,5′:1,2]pyrido[3,4-b]indol-11(11bH)-yl)methyl)benzoic acid (39) (1.62 g, 4.16 mmol) as described for 30a.

(137) The crude 4-((2-methyl-1,3-dioxo-2,3,5,6-tetrahydro-1H-imidazo[1′,5′:1,2]pyrido[3,4-b]indol-11(11 bH)-yl)methyl)benzoic acid (39) used therefore was obtained in quantitative yield from tert-butyl 4-((2-methyl-1,3-dioxo-2,3,5,6-tetrahydro-1H-imidazo[1′,5′:1,2]pyrido[3,4-b]indol-11(11 bH)-yl)methyl)benzoate (38) following the procedure described for 29a und used without purification.

(138) Yield 0.86 g (1.76 mmol, 42%) colorless foam. mp.: 151.2-154.0° C.; IR (KBr): 2946, 1713 cm.sup.−1; .sup.1H NMR (300 MHz, CDCl.sub.3) δ 7.65 (d, J=8.3 Hz, 2H), 7.55 (d, J=7.6 Hz, 1H), 7.21 (d, J=3.6 Hz, 2H), 7.19-7.11 (m, 1H), 7.02 (d, J=7.9 Hz, 2H), 6.13 (d, J=17.2 Hz, 1H), 5.64 (d, J=17.1 Hz, 1H), 5.08 (d, J=16.0 Hz, 2H), 4.52 (d, J=9.1 Hz, 1H), 4.02-3.88 (m, 1H), 3.63 (d, J=11.0 Hz, 1H), 3.03 (d, J=3.8 Hz, 3H), 3.00 (s, 1H), 2.90 (dd, J=23.1, 12.0 Hz, 2H), 1.73 (d, J=76.3 Hz, 6H).

N-Hydroxy-4-((2-methyl-1,3-dioxo-2,3,5,6-tetrahydro-1H-imidazo[1′,5′:1,2]pyrido[3,4-b]indol-11(11bH)-yl)methyl)benzamide (41)

(139) ##STR00117##

(140) Preparation from 4-((2-Methyl-1,3-dioxo-2,3,5,6-tetrahydro-1H-imidazo[1′,5′:1,2]pyrido[3,4-b]indol-11(11 bH)-yl)methyl)-N—((tetrahydro-2H-pyran-2-yl)oxy)benzamide (40) (0.50 g, 1.02 mmol) as described for 31a.

(141) Yield 225 mg (0.56 mmol, 55%). mp.: 134.5-136.9° C.; IR (KBr): 1710, 1462 cm.sup.−1; .sup.1H NMR (300 MHz, DMSO) δ 11.15 (s, 1H), 9.03 (s, 1H), 7.64 (d, J=8.2 Hz, 2H), 7.52 (d, J=7.1 Hz, 1H), 7.27 (d, J=7.7 Hz, 1H), 7.12 (d, J=6.0 Hz, 1H), 7.06 (d, J=8.1 Hz, 3H), 6.00 (d, J=17.2 Hz, 1H), 5.79-5.61 (m, 2H), 4.32 (dd, J=13.3, 4.8 Hz, 1H), 3.17-3.05 (m, 1H), 2.90 (s, 3H), 2.85-2.72 (m, 2H).

Example 4: Biological Evaluation

(142) Enzymatic Inhibitory Activities on HDACs 2, 6 and 8

(143) HDAC enzyme inhibition assays were conducted by Reaction Biology Corporation (Malvern, Pa., USA) using a ten point dose response curve with half-log serial dilutions, fluorogenic peptides at 50 μM as enzymatic substrates. Substrate for HDAC 2 and 6: fluorogenic peptide from p53 residues 379-382 (RHKK(Ac)AMC). Substrate for HDAC-8: fluorogenic peptide from p53 residues 379-382 (RHK(Ac)K(Ac)AMC). The investigated compounds inhibited HDAC6 selectively, on the basis of the enzymatic inhibitory study.

(144) TABLE-US-00003 TABLE 1 Inhibition of HDAC-subtypes HDAC2, HDAC6 and HDAC8 (IC.sub.50-values [nM]).sup.a. Compound Cpd. No. HDAC2 HDAC6 HDAC8 21  3 135 ± 885  2.41 ± 0.76   121 ± 10 (R)-21 13 000 7.05 1 970 0 (S)-21  1 795 3.02 364 31a  9 580 ± 380  8.99 ± 1.11 1 095 ± 75 31b  5 195 ± 565 16.45 ± 0.75 251.5 ± 32.5 41 410 000 ± 700  30.7 ± 5.8   83 ± 6 .sup.aCompounds were tested in a 10-dose IC.sub.50 mode with 3-fold serial dilution starting from 100 μM solutions for HDAC2 and HDAC8, respectively starting from 1 μM solutions for HDAC6. IC.sub.50 values were extracted by curve-fitting the dose/response slopes. TSA was used as an internal standard. ND: Not Determined. Assays were performed by Reaction Biology Corporation, USA. Determination in duplicate ± SD.

(145) Cellular Data.

(146) Since acetylated α-tubulin (ac-Tub) accumulates when HDAC6 is inhibited, its extent of accumulation is a valuable surrogate parameter for inhibitory efficiency. MS-275 only blocks the nuclear enzymes HDAC1, 2 and 3, which leads to hyperacetylation of histones without affecting the acetylation status of tubulin and was therefore used as negative control. Due to the poor potency of MS-275 it was used in a concentration of 5 μM. The absence of acetylated histone H.sub.3 (ac-H.sub.3) revealed that our inhibitors are selective and have no effect on nuclear deacetylases. HSP90 served as loading control. HDAC6 blots underlined that the target enzyme stays intact although being affected by our compounds (FIG. 2).

(147) Cell Culture and Western Blot

(148) MV4-11 (human, biphenotypic B myelomonocytic leukemia, American Type Culture Collection, ATCC Accession No.: CRL-9591) cells were used for the cellular test system. The cells were cultured in Rosewell Park Memorial Institute (RPMI) 1640 medium (Biochrom, Berlin, Germany, CAT #: F1215) supplemented with 10% heat inactivated fetal bovine serum (FBS Superior, Biochrom, CAT #: S0615), 2 mM glutamine (Biochrom, CAT #: K0283) and 1% penicillin and streptomycin (Biochrom, CAT #: A2212). The cells were cultured at 37° C. in a humidified atmosphere enriched with 5% CO.sub.2. Etinostat (MS-275, 29) was purchased from Biotrend (Cologne, Germany). Chemicals were blotting or cell culture grade and purchased from commercial suppliers (Carl Roth, Germany: Dimethylsulfoxide (DMSO), NaCl, Tris-(hydroxymethyl)-aminoethan (TRIS), glycerin, ethylenediaminetetraacetic acid (EDTA), bovine serum albumin (IgG free), sodium dodecylsulfate (SDS), ammonium persulfate (APS), glycine, dry milk, Tween 20; Sigma Aldrich, Germany: Nonidet P 40 substitute, N,N;N′,N′-Tetraethylethylendiamine (TEMED), protease inhibitor cocktail tablets cOmplete™ (CAT #: 04693116001), phosphatase inhibitor cocktail 2 (CAT #: P5726), Sample buffer according to Lämmli (CAT #: 11337), Dulbecco's phosphate buffered saline (PBS)) or purchased as indicated. Exclusively ultrapure water was used throughout the assays (Astacus Membrane Pure, MembraPure GmbH, Bodenheim, Germany).

(149) The assay was carried out as a modification of Buchwald et al. (Buchwald, Pietschmann, et al. 2010; Buchwald, Pietschmann, et al. 2013; Beyer, Kiweler, et. al. 2017). In brief, the cells were counted using a Neubauer counting chamber and 1,000,000 cells per 5 mL were seeded in every well of a 6 well plate. The cells were allowed to adapt for 2 h and then stimulated with the respective compound at the indicated final concentration (5 nM to 800 nM). The DMSO control concentration corresponded to the maximum concentration of DMSO used in the assay. 10 mM stock solutions in DMSO were stored at −80° C. and diluted 1:100 or 1:1000 with PBS immediately before the stimulation. MS-275 stock solution was stored at a concentration of 5 mM.

(150) The cells were collected on ice after the indicated time, centrifuged at 1000×g at 4° C., the supernatant was removed and cell pellets were washed with cold PBS buffer. After centrifugation at 14200×g at 4° C. and removal of the supernatant, cells were lysed with 120 μL NET-N buffer (100 mM NaCl, 1 mM EDTA, 10 mM TRIS-HCl (pH 8.0), 0.5% Nonidet P-40 (NP-40), 10% glycerin, protease inhibitor cocktail tablets cOmplete™ (1 per 10 mL of buffer), phosphatase inhibitor cocktail 2 (100 μL per 10 mL of buffer, added freshly)) for 1 h on ice. In the case that cells were not analyzed immediately after harvesting, cell pellets were flash frozen in liquid nitrogen (−196° C.), stored at −80° C. and lysed right after thawing. In the next step, the protein concentration was determined by Bradford assay[78] for all lysates. Sample buffer according to Lämmli was added in a proportion of 1:1 (v/v) and the lysates were heated to 95° C. for 5 minutes. The proteins (40 μg) were separated by SDS-polyacrylamide gel electrophoresis (SDS-PAGE) and blotted with a Mini-Protean Tetra Handcast System (BioRad, Munich, Germany). A 10% polyacrylamide (Rotiphorese NF-Acrylamide/Bis Solution 30%, Carl Roth, Germany) gel was used to analyze HDAC6, acetylated α-tubulin and a 12.5% polyacrylamide gel was used for ac-H.sub.3. HSP90 served as loading control. The proteins were blotted on an Immobilon-P Transfer Membrane (Merck Millipore, Darmstadt, Germany, CAT #: IPVH00010). The membranes were washed three times with TRIS buffered saline containing Tween 20 (TBST-T; 20 mM Tris, 140 mM NaCl, 0.05% Tween 20) for at least 5 minutes. After blocking with 5% dry milk in TBS-T for 1 h under shaking, the membranes were washed again as mentioned before. The membranes were incubated with the primary antibodies diluted 1:1000 in 2% dry milk in TBS-T overnight: HDAC6 (160 kDa, monoclonal rabbit antibody, D2E5, Cell signaling, Germany, CAT #: 7558S), HSP90 (90 kDa, monoclonal mouse antibody, AC88, Enzo Life Science, Germany, CAT #: ADI-SPA-830), acetylated α-tubulin (55 kDa, monoclonal mouse antibody, Sigma Aldrich, Germany, CAT #: T7451), acetylated Histone H.sub.3 (15 kDa, monoclonal rabbit antibody, Merck Millipore, CAT #: 06-599). Membranes were washed again with TBS-T and then incubated for 2 h with the secondary antibody diluted 1:10000 in 2% dry milk in TBS-T. HDAC6 and ac-H.sub.3 membranes were incubated with Licor IRDye 800 CW donkey fluorescence anti-rabbit IgG (H+L) antibody (LiCor Biosciences, Germany, CAT #: 92532213). HSP90 and α-tubulin were incubated with Licor IRDye 800 CW donkey fluorescence anti-mouse IgG (H+L) antibody (CAT #: 92532212). Finally, the membranes were washed three times and were imaged with a LICOR Odyssey Imager (LI-COR, Lincoln, USA) and measured with ImageStudioLite (Version 5.2.5).

Example 5: Enhancement of Solubility

(151) TABLE-US-00004 TABLE 2 Comparison of the solubility of compounds with and without a solubility improving Y—(CH.sub.2).sub.p—Q— Part Compound Y—(CH.sub.2).sub.p—Q— Part Solubility in H.sub.2O + 200-1000 g/L − <1 g/L

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