Synthesis of a spiroacetal intermediate for the synthesis of the anti-Helicobacter pylori agent CJ-12, 954

A method has been established for the synthesis of the 5,5-spiroacetal moiety of the anti-Helicobacter pylori agents CJ-12,954 1 and CJ-13,014 2 in racemic form and as a mixture of stereoisomers. Retrosynthetically the key spiroacetal 9 is derived from the acyclic protected dihydroxyketone precursor 10 or 11 which in turn are available from enynone 25 or 26 . Enynones 25 and 26 were prepared via addition of the acetylide derived from acetylenes 13 or 14 to aldehyde 12 followed by oxidation of the resultant alcohols 23 and 24 respectively. Acetylenes 13 and 14 in turn were available via [2,3]-sigmatropic rearrangement of allyl propargyl ether 16 . Extension of this reaction in an asymmetric sense by use of a chiral base would have provided acetylene 14 in enantioenriched form, however, efforts towards this end were disappointing. Enynones 25 and 26 were converted to keto epoxides 10 and 11 respectively by treatment with meta -chloroperbenzoic acid followed by hydrogenation of the acetylene. Attempts to effect deprotection and cyclization of bis-silyl ethers 10 and 11 to spiroacetal 9 were complicated by the presence of the epoxide. In an alternative approach enynone 26 was converted to methyl acetal 29 upon treatment with camphorsulfonic acid in methanol. Subsequent epoxidation of the terminal alkene 29 afforded epoxide 30 which then underwent smooth hydrogenation and cyclization in situ to the desired spiroacetal 9 .


Introduction
Gastric and duodenal ulcers affect a significant portion of the human population worldwide.Initially they were thought to be caused by damage to stomach and duodenal tissue by digestive fluids (acid and pepsin).More recent studies have shown a relationship between the presence of the microaerophilic Gram-negative bacterium Helicobacter pylori, which appears to live beneath the mucus layer of the stomach, and gastric and duodenal ulcers. 1 Therapy to eliminate Helicobacter pylori from the gastroduodenal tract would remove the root cause of gastric and duodenal ulcers therefore antibiotics are prescribed for the treatment of Helicobacter pylori in addition to treatment aimed at decreasing the production of stomach acid. 2 In a screening program designed to discover such compounds from microbial secondary metabolites, seven new phthalide compounds 1-7 with anti-Helicobacter pylori activity were isolated from the basidiomycete Phanerochaete velutina CL6387. 3The two most potent compounds, CJ-12, 954 1 and CJ-13, 014 2 have MICs of 5 ng/ml establishing that the presence of a spiro acetal moiety in addition to a phthalide unit enhances biological activity.The phthalide compounds 1-7 were specific for Helicobacter pylori in that they did not show antibacterial activities when tested against a panel of other microorganisms.This observed specific activity against Helicobacter pylori suggests that phthalides 1-7 may exhibit less side effects caused by disturbance of the normal gastro-intestinal microbial flora and may not induce drug resistance of non-target micro organisms.
The phthalides 1-7 are related to spirolaxine and sporotriacale 4 which have also been reported to have cholesterol lowering activity. 5Spirolaxine has helicobactericidal activity similar to 1 and 2. The rare 5,5-spiroacetal found in CJ-12, 954 1 and CJ-13,014 2 is also present in an insect pheromone 6 and in a constituent of hop oil. 7We herein report a synthesis of the 5,5spiroacetal moiety of the anti-Helicobacter pylori agents CJ-12,954 and 1 and CJ-13,014 2.

Results and Discussion
The stereochemistry of the stereogenic centre on the phthalide ring in both CJ-12,954 1 and CJ-13,014 2 has not been defined to date.CJ-12,954 1 and CJ-13,014 2 both exhibit 2S,7S stereochemistry in the spiroacetal ring and only differ in the configuration of the spirocentre.The focus of this work was on the development of a synthetic route to the spiroacetal moiety of CJ-12, 954 1 and CJ-13,014 2 as a mixture of stereoisomers using a synthesis that could be adapted at a later stage to control the stereochemistry at C-2 and C-7 of the spiroacetal ring system.
The key step in the retro synthesis adopted for the synthesis of CJ-12,954 1 and CJ-13,014 2 (Scheme 1) hinges on the addition of the Grignard reagent derived from bromide 8 to spiro acetal epoxide 9 followed by deoxygenation of the resultant secondary alcohol.The acyclic protected dihydroxy ketone precursor 10 or 11 to spiro acetal epoxide 9 is then constructed from addition of the acetylide of acetylene 13 or 14 to aldehyde 12.

Scheme 1
The acetylenes 13 and 14 were prepared using via [2,3]-sigmatropic rearrangement of allyl propargyl ether 16 (Scheme 2).Propargyl alcohol was converted to its C-trimethylsilyl derivative 15 8 via formation of the dianion using two equivalents of butyllithium followed by quenching with trimethylsilyl chloride.Subsequent allylation using conditions reported in the literature 9 (ethylmagnesium bromide and HMPA) afforded allyl propargyl ether 16.Unfortunately attempts to carry out this reaction using alternative reagents such as sodium hydride / allyl bromide, butyllithium / allyl bromide, ethyl bromide / allyl bromide were unsuccessful hence the use of the highly toxic agent HMPA could not be avoided.Treatment of allyl propargyl ether 16 with butyllithium at -78 o C for 10 min.effected smooth [2,3]-sigmatropic rearrangement to alcohol 17. 10 Subsequent O-silylation with tert-butyldimethylsilyl chloride or tert-butyldiphenylsilyl chloride afforded silyl ethers 18 and 19 respectively, which then underwent C-desilylation upon treatment with sodium methoxide to afford acetylenes 13 and 14.

Scheme 3
With quantities of aldehyde 12 and acetylenes 13 and 14 in hand, our attention next focused on their union via formation of the lithium acetylide (Scheme 4).Treatment of acetylene 13 in which the hydroxyl group is protected as robust tert-butyldiphenylsilyl ether with butyllithium in THF at -78 o C for 1h Followed by treatment with aldehyde 12 failed to generate the desired alcohol 23.In all cases unreacted acetylene 13 was recovered from the reaction.
In order to test whether butyllithium was in fact removing the acetylene proton prior to the addition of aldehyde 12, a series of quenches with deuterium oxide were performed.As a result of these studies, it was established that both butyllithium and tert-butyllithium were insufficiently basic to remove the acetylenic proton.Use of Schlosser's base 17 (a mixture of butyllithium and potassium tert-butoxide) effected deprotonation under these conditions and resulted in 100% incorporation of deuterium.Disappointingly, application of these conditions to the addition of the acetylide of acetylene 13 to aldehyde 12 only afforded the desired alcohol 23 # in 28% yield.
Attempts to optimise the use of Schlosser's base to effect this reaction proved uneventful.The principal side reactions observed were desilylation of acetylene 13 and aldehyde 12.Our attention then turned to the use of butyllithium with TMEDA as a cosolvent.The efficacy of the butyllithium / TMEDA system was first demonstrated by a deuterium quench and ultimately the coupling of acetylene 13 with aldehyde 12 to afford alcohol 23 proceeded in 85% yield.
Use of a tert-butyldiphenylsilyl ether to protect the hydroxyl group in acetylene 13 proved problematic in the subsequent deprotection step (vide infra).Thus, a parallel study was also undertaken using acetylene 14 which bears a more labile tert-butyldimethylsilyl ether.

Scheme 4
In this latter case the addition of the acetylide of acetylene 14 to aldehyde 12 was only achieved by adding the acetylide to a solution of the aldehyde (reverse addition) affording alcohol 24 in a modest 52% yield.
Alcohols 23 and 24 underwent smooth oxidation in good yield to ketones 25 and 26 using Dess-Martin periodinane or tetrapropylammonium perruthenate and N-methylmorpholine-Noxide respectively.Subsequent selective epoxidation of the terminal alkenes 25 and 26 using meta-chloroperbenzoic acid buffered with sodium acetate effected smooth conversion to epoxy ynones 10 and 11 respectively.Finally removal of the triple bond by hydrogenation over palladium on charcoal afforded the key saturated keto epoxides 10 and 11 which were precursors to the desired 5,5-spiroacetal 9.
At this stage it was envisaged that deprotection of both the silyl ethers in keto epoxides 10 and 11 would liberate a diol which would immediately undergo cyclization to the desired spiroacetal 9. Unfortunately initial attempts to effect this transformation using silyl ether 10 which bears a robust tert-butyldiphenylsilyl ether at C-8, were unproductive, hence the analogous bis-silyl ether 11 was prepared which bore a more labile tert-butyldimethylsilyl ether at C-8. Unfortunately attempts to deprotect bis-silyl ether 11 using a variety of conditions such as tetrabutylammonium fluoride, HF/pyridine and pyridinium p-toluenesulfonate in dichloromethane or methanol afforded complex mixtures for which the 1 H n.m.r.spectra of the crude product mixture clearly indicated that the epoxide had undergone reaction.
A solution to this problem was found by forming the two five membered rings of 5,5spiroacetal 9 sequentially (Scheme 5).Treatment of bis-silyl ether 26 with a catalytic quantity of camphorsulfonic acid in methanol afforded methyl acetal 29 in 97% yield which then underwent selective epoxidation of the terminal alkene using meta-chloroperbenzoic acid buffered with sodium acetate affording epoxide 30 in 89% yield.The long reaction times required to effect this epoxidation and the epoxidation of alkenes 25 and 26 above, was somewhat surprising, nevertheless epoxidation of alkene 29 in the presence of the alkyne did proceed selectively.Finally hydrogenation of alkyne over palladium on charcoal in the presence of sodium bicarbonate as buffer afforded the key 5,5-spiroacetal 9 in 63% yield.The modest yield for this step was attributed to the volatility of this compound.

Scheme 5
Having successfully prepared spiroacetal 9, albeit as a mixture of all possible stereoisomers, it was next decided to extend the synthetic methodology developed herein to execute a stereocontrolled synthesis of spiroacetal 9.In order to prepare anti-Helicobacter pylori agents CJ-12,954 1 and CJ-13,014 2 with the required 2S, 7S configuration of the 5,5-spiroacetal ring system, a synthesis of spiroacetals 9a or 9b which have the correct 2S,7R stereochemistry must be achieved.The stereochemistry of the spirocentre or the epoxide does not need to be controlled as the stereocentre in the epoxide is removed in a subsequent step and it was anticipated that two stereoisomers of the 5,5-spiroacetal, which differ only in the configuration of the spirocentre, would always be formed in the final spirocyclization step.
Applying the synthetic methodology reported herein to the synthesis of spiroacetal 9 with the correct 2S, 7R stereochemistry requires the synthesis of (S)-acetylene 14 and (S)-aldehyde 12. (S)-Aldehyde 12 is available from (S)-1,4-pentanediol which in turn is readily prepared from ethyl 4-oxopentanoate by enantioselective reduction of the ketone with bakers' yeast 18 followed by reduction of the ester using lithium aluminium hydride.
The successful completion of the synthesis of spiroacetal epoxide 9, albeit as a mixture of stereoisomers, constitutes a synthesis of the spiroacetal moiety of CJ-12, 954 1 and CJ-13, 014 2. It now remains to append the phthalide fragment 8 to this spiroacetal fragment.The inability to effect the key [2,3]-sigmatropic rearrangement of allyl propargyl ether 16 to alcohol 17 in an asymmetric fashion has necessitated that future work also be directed towards a synthesis of this 5,5-spiroacetal 9 using methodology that enables control of the stereochemistry at C-2 and C-7 of the spiroacetal ring system.

Experimental Section
General Procedures.Melting points were determined using a Kofler hot-stage apparatus and are uncorrected.Infrared spectra were recorded with a Perkin-Elmer 1600 series Fourier-transform infrared spectrometer as thin films between sodium chloride plates. 1 H and 13 C n.m.r spectra were obtained using either at Bruker AC200 spectrometer or a Bruker DRX-400 spectrometer.Both 1 H and 13 C n.m.r spectra were interpreted with the aid COSY, HETCOR and DEPT 135 experiments and are reported downfield from tetramethylsilane as standard.High-resolution mass spectra were recorded using a VG7070 spectrometer operating with an ionisation potential of 70 eV at a nominal resolution of 5000 or 10000 as appropriate.Major fragments are given as percentages of the base peak and are assigned where possible.Tetrahydrofuran and diethyl ether were dried using sodium/benzophenone and distilled prior to use.Flash chromatography was performed using Merck Kieselgel 60 or Riedel-de-Haen Kieselgel S silica gel (both 230-400 mesh) with the indicated solvents.Compounds were visualized under ultraviolet light or by staining with iodine or vanillin in methanolic sulfuric acid.

4-(tert-Butyldimethylsilyloxy)pent-1-yl acetate (21).
To a stirred solution of alcohol 20 (1.15 g, 7.9 mmol), imidazole (0.72 g, 10.6 mmol) and 4-dimethylaminopyridine (117 mg, 0.96 mmol) in DMF (20 mL) at room temperature was added tert-butyldimethylsilyl chloride (1.44 g, 9.6 mmol) in small portions.After 4 h the reaction was quenched by the addition of saturated ammonium chloride (15 mL) and water (10 mL) then the mixture was diluted with diethyl ether (30 mL).The organic phase was separated, washed with water (2 x 10 mL), brine (5 mL) then dried over MgSO 4 .Evaporation of the organic extract in vacuo followed by flash chromatography of the residue using 20% diethyl ether -hexane as eluent afforded the title compound 21 14 (1.93 g, 95%) as a light yellow oil for which the 1 H NMR data was in good agreement with the literature. 14