Synthesis of cis -enamide macrocycles via ring-closing metathesis

Herein we report the synthesis of 13, 14-and 15-membered cyclic lactams, using Grubbs ’ RCM method in 25, 52 and 46% yields respectively. The cis -enamide functional group was successfully introduced into these cyclic lactams by syn sulfoxide elimination The synthetic cyclic lactams resemble natural cyclic peptides. Our synthetic methodology provides a simple route to making medium-sized cyclic lactams that could be used as models to mimic the β -turn in natural proteins, an important marker in understanding their biological activity


Introduction
Over the past five decades, about 500 cyclopeptides have been isolated from over 120 species.2][3][4][5][6] We are interested in the chemistry of the cyclic lactams with an enamide (styrylamide) functionality in their cyclic structure, e.g.zizyphine A, 7 or a secondary alcohol group, which probably is the precursor to the styrylamide through dehydration, as in pandamine (Figure 1). 85][16][17][18] Many of the acyclic enamides isolated from marine sources as ascidians, [19][20][21][22][23] microorganisms, [24][25][26] fungal and higher plants [27][28][29][30][31] exhibit a variety of pharmacological activities. 25,26,32,33Especially the macrolides salicylihalamides A and B, 34,35 apicularen A and B 36 and lobatamide A-F 37,38 isolated from marine sponge Haliclona sp., Chondromyces sp.(myxobacteria) and tunicates, respectively.Salicylihalamides A, B and lobatamides are highly potent cytotoxic macrolides, incorporating salicylic acid, a 12-membered lactone ring and an enamide side-chain (trans A, cis B) (Figure 2).© ARKAT USA, Inc Salicylihalamide A and lobatamide A display potent cytotoxicity in the NCI 60-cell line human tumor assay, and it is suggested that these compounds may act by a novel mechanism of action. 39Recently, it has been reported that salicylate enamide macrolides selectively inhibit the Vo sector of the V-ATPase through Nacyliminium ion generated from N-acyl enamines under acidic conditions. 39Figure 4 shows how the acyliminium is captured by a nucleophilic amino acid side-chain, and subsequently undergoes irreversible hydrolysis.A study on the structure-activity relationships (SAR) of (-)-salicylihalamide A reported by Barbander and co-workers, revealed that the characteristic N-acyl enamine functionality is essential for the biological activity. 40,41,42This mechanistic route via the enamide ATPase protein interaction, its biological implications, and its stereochemistry attracted our attention and prompted us to develop a simple and efficient synthetic route to the acyclic enamide tuberine and its analogs, in order to study in detail their biological properties. 14 Biologically active macrocylic lactams and enamides have been reported, isolated from natural sources.The cryptophycins are a family of highly cytotoxic, cyclic depsipeptides and are among the the most potent agents, and their binding to tubulin is very strong and poorly reversible.Many cryptophycins have been isolated fron Cyanobacterium nostoc sp. 43Cryptophycin 52 (Figure 4) was produced by total chemical synthesis and it destroys microtubules in vascular smooth muscle cells.Ixabepilone is a semisynthetic epothilone B analogue, which acts like Paclitaxel as a microtubule stabilizing agent.It is used in the treatment of aggresive metastatic or locally advanced breast cancer. 44Tanespimycin is a derivative of the antibiotic geldanamycin that is being studied in the treatment of cancer, specifically in young patients with certain types of leukemia or solid tumors, especially kidney tumors.It works by inhibiting Hsp90, which is expressed in those tumors, and it belongs to the family of drugs called antitumor antibiotics. 45Finally, sagamilactam is a polyunsaturated and polyoxygenated macrocyclic lactam containing diene, triene and tetraene conjugated olefins and a decalin moiety which has antitrypanosomal activity.6][17][18] This method was successfully employed in the synthesis of some naturally occurring N-acylenamides. 16In most cases when the N-acyl is a formyl group, the Z-N-alkenylformamides were the major product.The naturally occurring tuberine also exhibited a similar Z:E ratio.In addition, 1 H NMR signals for the two stereoisomers due to the conformers arising from restricted amide bond rotation were encountered, which led us to calculate a rotational barrier of 19 kcal/mol using 2D EXY NMR.Since many of the naturally occurring enamides also have a cis-enamide, it was of interest to study in depth the physical properties of cis-enamides.The cis-N-(β-2,6difluorostyryl)formamide was selected as a model.The NMR studies revealed a rotational barrier for the cisenamide with the same magnitude as the trans-enamide.However, the X-ray structure of the trans-trans-N-(β-4-methoxystyryl)formamide (tuberine) showed that the benzene π-plane was coplanar to the π-enamide bond (Figure 5a), while in the cis-enamide N-(β-2,6-difluorostyryl)formamide (Figure 5b) the styrylamide π-plane of the double bond was at 48 degrees with respect to the benzene ring π-plane.These structures agree well with computational models.A similar conformational behavior in the molecular modelling study of sanjonine G1 was observed by Zhu et al., where the σ bond in the macrocyle lies out of the plane defined by the aromatic ring.

OMe
This subtle difference in the orientation of the double bond in the trans and cis enamide function and the biological significance prompted us to synthesize and study the biological properties of some model 13-, 14and 15-membered cyclic enamides and explore the relationship between stereochemistry and biological properties.Our goal in this project was to attempt and introduce the cis enamide functionality into a cyclic 13-, 14-and 15-sized lactams, using a syn-sulfoxide elimination methodology, and to study these compounds as models for the natural cyclopeptides.

Results and Discussion
Our synthetic strategy for the high membered macrocycles is shown in Scheme 1.It started with the alkylation of an appropriately substituted hydroxybenzaldehyde 1 with allyl bromide under basic conditions.The allyloxybenzaldehydes 2 were subjected to an addition of nitromethane, followed by a subsequent Michael addition of thiophenol to nitrostyrenes 3 to give nitro sulfides 4. The nitro group was reduced to amino giving compounds 5, and these were then derivatized with a suitable n-alkenyl carboxylic acid to give the amides 6-8 with a diene functionality, which then were ring closed by the RCM method using a Grubbs-Hoveyda first generation catalyst to give the macrocyclic compounds 9-11 in very good yields.The sulfide group was oxidized to sulfoxide and the sulfoxides subjected to thermal elimination to give the cis-enamides 12-14 in good yields.This methodology is straightforward compared to the selenocyanate oxidative elimination reported independently by Schmidt and Joullie in the synthesis of the cyclopeptide alkaloids franguline A and nummularine F, respectively. 47Our initial goal was to find a suitable methodology to construct the required ring system, followed by a syn elimination of the sulfoxide to generate the enamide functional group.][50][51][52][53][54][55][56]

Conclusions
We have synthesized 13-, 14-and 15-membered cis-enamide macrocycles using Grubbs RCM method in 25, 52 and 46% overall yields, respectively.The cis-enamide functional group was successfully introduced into these cyclic lactams by a syn sulfoxide elimination.This synthetic methodology provides a simple route to making medium-sized cyclic lactams.][59][60]

Experimental Section
General.All reagents were purchased in the highest quality available and were used without further purification. 1H and 13 C NMR spectra were recorded using a Varian 500 MHz Spectrometer in CDCl3 with TMS as internal standard and 200 MHz.The chemical shifts are expresed in ppm and the coupling constants (J) in Hertz.Electronic impact mass spectra were obtained by direct insertion in Agilent 5975C mass spectrometer.Infrared spectra were recorded on a Spectrum FT-IR 1600 spectrophotometer.Melting points were determined on a Fisher-Johns melting point apparatus and were corrected.
2,4-Dibromo-5-hydroxybenzaldehyde (1b).To a solution of 3-hydroxybenzaldehyde 1a (40.9 mmol, 5.0 g) in CHCl3 (100 mL) was added bromine (81.7 mmol, 4.2 mL).The resulting solution was stirred at rt for 3 d.The excess bromine was removed with a saturated solution of sodium thiosulfate (20 mL), the organic phase was washed with water and dried over anhydrous Na2SO4.Removal of solvent gave a solid which was recrystallized from acetic acid to give 1b (10.

2-(3-Allyloxy-4,6-dibromophenyl)-2-phensulfanylethanamine (5b).
To a solution of compound 4b (0.5 g, 1.0 mmol) in dry MeOH (1.0 mL) was added a 0.1 M solution of SmI2 (100 mL, 10.0 mmol) in THF under Ar atmosphere at rt.The mixture was stirred for 24 h at the same temperature.After this time, it was added dropwise to an aqueous solution of NaOH until the blue colored suspension turned colorless.The resulting suspension was then extracted with CH2Cl2 (3×10 mL).The combined organic extracts were dried with MgSO4, filtered and the solvent was evaporated to afford the crude amine as a clear yellow liquid (0.  (31).