Novel Spiroannulated 3-Benzofuranones. Synthesis and Inhibition of the Human Peptidyl Prolyl cis/trans Isomerase Pin1

The novel 3H-spiro[1-benzofuran-2-cyclopentan]-3-one skeleton has been made accessible by different routes. One- and two-step protocols lead to tricyclic and tetracyclic benzofuranones 2 and 3, respectively. A four-step synthesis to spirocompound 4 is described. The novel spirocyclic benzofuranones display modest to no inhibition of the human peptidyl prolyl cis/trans isomerase Pin1.


Introduction
Spirocyclic benzofuranones have attracted considerable attention with regard to their pharmaceutical activity and the various approaches directed towards their synthesis [1]. Aside from griseofulvin, known as an orally active antimycotic drug [2], synthetic and naturally occurring [3] compounds, which feature the spirobenzofuranone moiety, have been found to display inter alia antiinflammatory [4] and herbicidal activity [5] or act as aromatase inhibitors [6]. We have recently shown that synthetic spirocyclic 2-benzofuranones 1, involving a lactone skeleton, and aryl indanyl ketones are efficient inhibitors of the human peptidyl prolyl cis/trans isomerase Pin1 [7]. In this article, we describe the synthesis of 3-benzofuranones 2-4, the first representatives of the novel 3H-spiro-[1benzofuran-2-cyclopentan]-3-one skeleton and a screening of their inhibition of Pin1.

Results and Discussion
Commercially available coumaranone 5a and the methoxy-substituted derivative 5b [8] served as the starting materials for the preparation of spirocyclic benzofuranones 2 and 3, as shown in Scheme 2.
Scheme 2. Synthesis of spiroannulated 3-benzofuranones 2 and 3. THF Thus, deprotonation of the ketones 5a,b with lithium diisopropylamide and subsequent treatment with allyl bromide led to the dienes 6a,b. Although the protocol aimed at a double allylation, neither the base nor the electrophile were used in a 2:1 molar ratio with respect to the ketone because these conditions led mainly to the formation of aldol products and other side reactions that obviously consumed the electrophile. Although the allylic alkylation protocol was not thoroughly optimized, it was observed that the additive 1,3-dimethyltetrahydro-2(1H)pyrimidone (DMPU) was crucial, because, in its absence, the reaction failed completely. Ring closing metathesis [9] permitted conversion of the dienes 6a,b into the spirocyclic ketones 2a,b. A one-step procedure led to the formation of spirocycles 3a,b when the coumaranones 5a,b were treated with two equivalents of lithium diisopropylamide and α,α'-dibromoxylene. Again, the yields were moderate due to various side reactions, wherein condensation and subsequent reduction-oxidation processes seem to take place prior to the desired reaction with the electrophile. Again, the protocol was not optimized.
Benzofuranone 4, isomeric to 3b and featuring an angular arrangement of the spirocyclic indane moiety was synthesized in a three-step protocol, outlined in Scheme 3. Thus, 2-bromo-1,4dimethoxybenzene (7a) was submitted to a bromine/lithium exchange to give lithioarene 7b and subsequently allowed to react with the lithium salt 8b generated from indanecarboxylic acid (8a). The ketone 9 thus obtained was selectively deprotected by treatment with aluminum chloride cleaving the methoxy group ortho to the carbonyl group.  The conversion of ortho-hydroxy aryl ketones to 3-benzofuranones [10] was applied in the final step to the spiroannulated compound 4. Thus, phenolic ketone 10 was treated with ethyl bromomalonate in the presence of potassium carbonate and butanone, leading to 3-benzofuranone 4 in 36% yield. As a mechanistic explanation of this transformation, it is assumed that ethyl bromomalonate acts as an electrophilic brominating agent [10,11] that brings about the introduction of the α-bromo substituent into the ketone 10. Under the basic conditions, a final intramolecular nucleophilic substitution leads to a ring closure. It deserves to be mentioned that this protocol has neither been applied to the generation of quaternary carbon centers nor to the preparation of spirocyclic compounds. Although the yield is moderate and the product is accompanied by unidentified phenolic compounds, the latter can be easily separated by column chromatography. Thus, the protocol provides a ready access to the hitherto unknown spiroannulated system 4.

Conclusions
Benzofuranones 2-4 featuring a novel spirocyclic skeleton have been synthesized in one-to threestep procedures. They were tested as inhibitors of a peptidyl prolyl cis/trans isomerase. This study shows, however, that the activity of the spiroannulated 3-benzofuranone derivatives 2-4 is distinctly lower than that obtained with the lactone-type spirocompounds 1 on the one hand [7a] and aryl indanyl ketones [7b, 13] on the other hand.
General procedure for the double allylation of benzofuranones 5 (G. P. 1): A 250-mL Schlenk-flask was equipped with a magnetic stirrer, a thermocouple and a septum and was connected to a combined argon/vacuum line. The air in the flask was replaced by argon. Diisopropylamine (1.49 g, 2.07 mL, 14.7 mmol) and dry THF (52 mL) were injected, and the mixture was stirred at -78°C. A solution of n-butyllithium (9.75 mL, 15.6 mmol) in hexane was added at such a rate that the temperature did not exceed -70°C. Stirring was continued at 0°C for 30 min.
In a 50 mL Schlenk-flask, benzofuranone 5 (12.3 mmol) was dissolved in dry THF (20 mL) under argon. This solution was added through a cannula to the mixture of lithium diisopropylamide, prepared as described above, at -78°C. Stirring was continued at the same temperature for 2 h. Freshly distilled allyl bromide (1.73 g, 1.24 mL, 14.3 mmol) and 1,3-dimethyltetrahydro-2(1H)pyrimidone (DMPU) (1.89 g, 1.78 mL, 14.7 mmol) were injected. The mixture was allowed to warm up to room temperature overnight and stirred for 48 h at this temperature. The solution was washed with a saturated aqueous solution (50 mL) of NH 4 Cl. The aqueous solution was extracted with chloroform (3 times 50 mL). The organic layers were combined and dried with Na 2 SO 4 . The solvent was removed in a rotary evaporator and the oily residue was purified by column chromatography on silica gel. According to this procedure the following products were obtained:

General procedure for the preparation of spiroannulated benzofuranones 2 by ring-closing metathesis (G. P. 2):
A mixture of benzylidene-bis-(tricyclohexylphosphino)-dichlororuthenium ("Grubbs catalyst I") (0.016 g, 0.02 mmol) in dry dichloromethane (50 mL) was stirred under argon in a 100-mL flask equipped with a magnetic stirrer, a septum and a connection to the combined argon/vacuum line. At room temperature, diallyl compound 6 (4.0 mmol), dissolved in 6 mL of dry dichloromethane, was added and the mixture was stirred at room temperature for 19 h. The solvent was removed in a rotary evaporator and the residue was purified by column chromatography. According to this procedure the following products were obtained:

General procedure for the preparation of spiroannulated benzofuranones 3 (G. P. 3):
According to G. P. 1, benzofuranones 5 were deprotonated with lithium diisopropylamide. Then, the solution was treated with 1 equivalent (relative to 5) of α,α'-dibromo-o-xylene, dissolved in THF, and 1 equivalent of DMPU. The mixture was warmed up to room temperature. After work up according to G. P. 1, the residue was purified by column chromatography. According to this procedure the following compounds were obtained:   Under nitrogen, a solution of 7a (6.015 g, 27.27 mmol) in dry diethyl ether (60 mL) was stirred at -78°C in a two-necked flask, equipped with a magnetic stirrer, a septum and a connection to the nitrogen/vacuum line. A solution of n-butyllithium (19.8 mL, 31.6 mmol) was added slowly by syringe, and stirring was continued at the same temperature for 30 min. In a second flask, 1indanecarboxylic acid (8a, 2.25 g, 14.0 mmol) was dissolved in dry diethyl ether (30 mL), and treated at -78°C under stirring with a solution of n-butyllithium (8.6 mL, 13.8 mmol). Stirring was continued at 20°C for 30 min. The solution of the first flask was added through a cannula, and the mixture was allowed to reach room temperature overnight. Water (20 mL) was added, followed by 2 N hydrochloric acid (3 mL) and the aqueous layer was extracted three times with diethyl ether. The combined organic layers were dried with magnesium sulfate, the solvent was removed in a rotary evaporator and the residue was purified by column chromatography to give colorless, oily 9 (1,68 g, 55%); R f = 0.36 (chloroform); 1