Synthesis of 9-azasteroid partial structures via Birch reduction as key step

A high-energy intermediate model for the inhibition of the ergosterol biosynthesis suggests 9-azasteroids as potential antimycotics. Key step for the approach described in this work involves a Birch reduction of substituted quinoline structures. The diastereoselectivity of this reaction was studied. Subsequent functionalization to incorporate the lipophilic properties of the steroidal core afforded N -substituted perhydro-quinolinols as mimics of the AB-ring system of steroids.


Introduction
The ergosterol biosynthesis pathway represents an interesting target for the development of novel antimycotic agents. 1 During recent years, ∆ 8 -∆ 7 -isomerase was a key target in the development of bioactive agents. 2 This enzyme is responsible for the isomerization of fecosterol 1 to episterol 2. In particular perhydro-heterocyclic compounds with lipophilic chains such as phenpropimorph 3, fenpropidine 4, or tridemorph 5 were successfully established as commercial products (Scheme 1).In this context, we and others have started to investigate the potential of azasteroid partial structures 6 derived from the isoquinoline structural core as fungicides, which resemble the natural substrate of the enzymes involved to a greater extent. 3,4 H Another possible target to influence steroid biosynthesis is the cyclization reaction towards the steroidal ABCD ring system.Oxidation of squalene 7 to squalene-oxide 8 is followed by an electron cascade and cyclization reaction, which leads to the cationic species 9 after methyl group migration as a key branching point in the anabolism of steroids in fungi and plants: While the intermediate undergoes elimination to lanosterol 11 in fungi, the reactive compound cyclizes to cycloartenol 10 in plants (Scheme 2). 5 One traditional inhibition strategy for enzymes accommodating such cationic species as highenergy intermediates (HEIs) is the incorporation of nitrogen at the site of the positive charge.The generally accepted concept behind this approach is the hypothesis, that such amine species are protonated under physiological conditions and consequently exhibit higher affinity to the functional groups within the active site of the enzyme stabilizing the positive center. 6onsequently, azasteroids of the general type 12 incorporating key structural aspects of the steroidal ring system are potential inhibitors of proteins involved in this biosynthetic step.

Results and Discussion
Starting from the readily available 6-methoxyquinoline 13 reduction of the heteroaromatic core was performed using Raney-Ni to give 14a (R = H) in good yield (Scheme 3).Reductive alkylation with formaldehyde under hydrogenation conditions gave the N-methylated product 14b (R = Me).Both substrates were used in the subsequent one-pot Birch reaction (Scheme 3).The Birch reduction was performed according to an optimized protocol reported by us recently. 14This approach takes advantage of a one-pot comprehensive reduction of the heteroaromatic core avoiding isolation of the rather unstable intermediates 15 and 16.However, we were able to isolate compound 16b in a modified procedure to prove the reaction pathway outlined in Scheme 3.
The one-pot reaction is best carried out with lithium as electron source to give species 15 as an intermediate.In a one-pot procedure subsequent reduction to 16 was carried out using NaCNBH 3 at pH 4. As we had observed in our model study on dimethylanisidine 14 both enol ether 16 and ketal 17 can undergo further reduction leading to compound 18 as a by-product.
The NaCNBH 3 reaction was highly selective giving trans-fused products exclusively.Since we did not obtain crystals suitable for X-ray diffraction from derivatives of the liquid products 17 and 19 the stereochemistry of this step was assigned utilizing the crystalline material 18.Based on extensive NMR experiments and the structural data from the single crystal X-ray diffraction study the stereochemistry of all products resulting from the Birch reduction sequence was assigned.
Ketals 17a/b were obtained in excellent yields and deprotection of the carbonyl group was performed according to standard conditions to give compound 19.
Product 17a served as precursor for the introduction of the lipophilic substituent at the nitrogen atom of the ring system.Two targets of biological interest were prepared: A long carbohydrate chain was intended to mimic the carbocyclic structure of the steroid, and the 4tert.-butyl-benzylgroup represents a typical hydrophobic substituent in a variety of inhibitors of the ergosterol biosynthesis with a related mode of action.
At this stage reduction of the amide was carried out with RedAl ® to form compounds 21a/b.However, instead of proceeding via ketal cleavage and a subsequent second reduction step, the following shortcut in the total synthesis of the target products was developed: Deprotection of the ketal 20a/b according to the protocol developed on the model compound 9 gave ketones 22a/b.Finally, reduction of both the amide functionality and the ketone to the corresponding amino alcohols 23a/b was performed using RedAl ® in high to quantitative yields.This reagent turned out to be highly selective for the formation of an equatorial hydroxyl group.Structural assignment is based on studying the coupling system for the annelation site protons in compound 18 using 2-dimensional NMR.Assignment of the trans-configuration and equatorial position of the methoxy group was confirmed by X-ray diffraction (Figure 1 and experimental section).Typical shifts and coupling constants for this model system were applied in the structural assignment of the other perhydroquinolines.In summary, we developed a diastereoselective route to N-substituted perhydro-quinolinols as potential inhibitors for the ergosterol biosynthesis based on a Birch reduction protocol.The target compounds represent azasteroid partial structures of the parent AB ring system in ergosterol and incorporate lipophilic chains mimicking the hydrophobic properties of the steroidal core.

Experimental Section
General Procedures.Unless otherwise noted, chemicals were purchased from commercial suppliers and used without further purification.All solvents were distilled prior to use.Flash column chromatography was performed on silica gel 60 from Merck (40-63 µm).Kugelrohr distillation was carried out using a Büchi GKR-51 apparatus.Melting points were determined using a Kofler-type Leica Galen III micro hot stage microscope and are uncorrected.Elemental analyses were carried out in the Microanalytical Laboratory, University of Vienna.The NMR spectra were recorded from CDCl 3 solutions on a Bruker AC 200 (200 MHz) spectrometer and chemical shifts are reported in ppm using Me 4 Si as internal standard.
Lithium chips were added slowly, maintaining the temperature at -35±5°C.After the initially vigorous reaction had ceased the mixture was refluxed until the blue color disappeared.Ammonia and the organic solvents were evaporated by a stream of nitrogen at approx.50°C and the residue was treated with dry THF and MeOH.The resulting solution was cooled to -5±5°C and brought to pH 4 by addition of methanolic HCl using bromocresol green as indicator.During the addition of NaCNBH 3 pH = 4 was maintained by treatment with methanolic HCl.When the pH showed no further change solid sodium bicarbonate and some NaOH were added and the solvents evaporated.

trans-Decahydro-6,6-dimethoxy-1-methylquinoline (17b).
Precursor 14b (2.00 g, 11.28 mmol) was converted according to the general procedure in a mixture of 200 mL of liquid NH 3 and 50 mL of dry THF in the presence of 16 equiv.of dry methanol by treatment with 15 equiv. of lithium.The reaction was refluxed for an additional 10 min after disappearance of the blue color.After evaporation the residue was dissolved in 50 mL of THF and 20 mL of methanol and reduced with 1.00 equiv. of NaCNBH

General procedure for acylation of precursor 17a
A 10% solution of ketal 17a (1 equiv.)and dry triethylamine (1.2 equiv.) in dry diethyl ether was treated with a 10% solution of the corresponding acid chloride (1.1 equiv.) in dry diethyl ether and stirred overnight at room temperature.The reaction mixture was hydrolyzed with water, extracted with diethyl ether, dried over sodium sulfate, filtered, and concentrated.

trans-Octahydro-1-(1-oxohexadecyl)-quinolin-6(2H)-one (22a
). Ketal 20a (0.50 g, 1.14 mmol) was dissolved in a mixture of 10 mL acetone and 2 mL 2N HCl and refluxed overnight.Then the pH was adjusted to 7 by addition of sodium bicarbonate and the solvent was evaporated.The residue was treated with water and diethyl ether, the layers were separated and the aqueous layer was extracted with diethyl ether.The combined organic layers were dried over sodium sulfate and concentrated.The crude product was purified by flash column chromatography (silica gel) to give 0.33 g (74%) of 15a as yellow crystals.Mp.: 51-54°C. 1 H NMR (CDCl 3 ): 0.85-0.9(m, 3H, CH 3 ), 1.0-2.6 (m, 42H). 13C NMR (CDCl 3 ): 13.9 (q, C16'), 22.  structure was solved with direct methods using the program SHELXS86.Structure refinement on F 2 was carried out with the program SHELXL93. 18Non-hydrogen atoms were refined anisotropically.The N-bonded hydrogen atom was fully refined.C-bonded hydrogen atoms were inserted in idealized positions and were refined riding with their carrier atoms.Final refinement gave R 1 = 0.055/0.150for observed/all reflections and 115 parameters.CCDC 108184 contains the supplementary crystallographic data for this paper.These data can be obtained free of charge from The Cambridge Crystallographic Data Centre via www.ccdc.cam.ac.uk/data_request/cif.The structure determination showed that the methoxy group and the N-bonded H-atom adopt equatorial positions relative to the decahydroquinoline moiety (Fig. 1).The N-H group forms a weak hydrogen bond with the N atom of a neighboring molecule, N---N = 3.346(3) Å.This gives rise to continuous hydrogen bond chains …N-H---N-H---N-H… parallel to the b-axis.

Figure 1 .
Figure 1.Structure determination of compound 18 by X-ray diffraction with crystallographic atom numbering and 20% ellipsoids.

ARKIVOC 2005 (v) 83-95 ISSN 1424-6376 Page 89
17suspension of 14a (5.84 g, 35.78 mmol), 5% Pd/C (0.58 g), and 35% aqueous formaldehyde solution (30.7 g, 357.8 mmol) in methanol (250 mL) was charged into a Parr apparatus and hydrogenation was performed at a H 2 pressure of 6 bar overnight.The catalyst was separated by filtration through Celite and the remaining solution was concentrated to remove methanol.The residue was diluted with water and extracted with diethyl ether.The combined organic layers were dried over sodium sulfate, filtered, and concentrated.The crude product was purified by Kugelrohr distillation to give 5.10 g (92%) of 14b17as colorless oil which started to crystallize upon storage in a +4°C fridge.