Total synthesis of clostrubin

Clostrubin is a potent antibiotic against methicillin- and vancomycin-resistant bacteria that was isolated from a strictly anaerobic bacterium Clostridium beijerinckii in 2014. This polyphenol possesses a fully substituted arene moiety on its pentacyclic scaffold, which poses a considerable challenge for chemical synthesis. Here we report the first total synthesis of clostrubin in nine steps (the longest linear sequence). A desymmetrization strategy is exploited based on the inherent structural feature of the natural product. Barton–Kellogg olefination forges the two segments together to form a tetrasubstituted alkene. A photo-induced 6π electrocyclization followed by spontaneous aromatization constructs the hexasubstituted B ring at a late stage. In total, 200 mg of clostrubin are delivered through this approach.

T he discovery of effective antibiotic agents is an urgent global demand for combating drug-resistant pathogenic bacterium strains 1,2 . Secondary metabolites that are produced by microbes as chemical defence have proven to be the most important source of such agents [3][4][5][6][7][8][9][10][11][12][13][14] . In May 2014, Hertweck and co-workers 15 reported the isolation of clostrubin (1, Fig. 1) from the strictly anaerobic bacterium C. beijerinckii. This compound exhibits remarkable potency against methicillinresistant Staphylococcus aureus and vancomycin-resistant enterocci, with minimum inhibitory concentrations of 0.12 and 0.97 mM, respectively. From a structural perspective, clostrubin (1) poses considerable synthetic challenge owing to the fused aromatic ring system and multisubstitution pattern. The potential of 1 as a lead compound for antibiotic development and its limited supply from natural sources stimulated us to launch a chemical synthesis programme immediately.
In this paper, we report the first total synthesis of clostrubin (1) from commercially available starting materials. This concise synthesis (nine-step for the longest linear sequence) benefits from the inherent structural symmetry of 1. An advanced olefin intermediate was constructed through Barton-Kellogg olefination, and a 6p electrocyclization promoted by ultraviolet light assembled the fully substituted B ring.
Synthesis of the two segments. The synthesis commenced with the preparation of diazoketone 3, as shown in Fig. 2. A sequence of double D-A reactions was carried out: 2,6-dibromo-1,4-benzoquinone 5, prepared in one step from commercially available 1,3,5-tribromophenol 58  silica gel may accelerate the hydrolysis of the silyl ether and thus led to a rapid aromatization along with release of HBr. Notably, when a single equivalent of 8 was used for the D-A reaction, a bromonaphthoquinone was readily prepared, presumably with the intermediacy of a mono cycloadduct 61 . The crude 9 was treated with K 2 CO 3 and MeI to give compound 11 (45% overall yield from 5). 11 underwent Clemmensen-type reduction in the presence of SnCl 2 and HCl to give the corresponding monoketone 62 , which instantaneously tautomerized to anthranol 12. Exposure of crude 12 to 1,8-diazabicyclo [5.4.0]undec-7-ene (DBU) and TsN 3 furnished diazoketone 3 in 89% overall yield. We observed unexpected reactivity of anthraquinone 11 (Fig. 3) during the above studies, which influenced the overall strategy of the synthesis. In theory, the C9 carbonyl of 11 should be sterically more hindered for nucleophilic attack due to two neighbouring methoxy groups. From an electronic perspective, this upper ketone could be considered as an equivalent of a double vinylogous carbonate than is also rather unreactive as an electrophile. To our surprise, we obtained hydrazone 13 in 61% yield when treating 11 with TsNHNH 2 ; the anticipated regioisomer was not detected. The structure of 13 was determined by X-ray crystallographic analysis. This observation interrupted our initial plan of exploiting the C10 tosylhydrazone of 11 as the potential precursor for the Barton-Kellogg olefination. We further examined other types of nucleophiles such as benzyl Grignard reagent for the addition reaction with 11. In this case, two regioisomeric alcohols 14 and 15 were isolated in 32% and 40% yields, respectively. Both structures were confirmed by nuclear Overhauser effect (NOE) studies. The enhanced reactivity of the C9 carbonyl may be attributable to inductive effects from the o-methoxy substitutents or relief of 1,3-allylic strain that occurs on nucleophilic additions. Thus, the strategy involving direct olefination of C10 carbonyl of anthraquinone 11 (for example, with functionalized benzylic metal species or phosphonate carbanion) had to be abandoned due to the poor regioselectivity.
We then focused on the synthesis of the thioester segment as the electron donor in the devised Barton-Kellogg olefination, as shown in Fig. 4. Aldehyde 16 was prepared in one step from commercially available 2-iodophenol 63 . Treatment with K 2 CO 3 and MeI gave methyl ether 17 (99% yield), which underwent MeMgBr addition followed by silylation to provide compound 18 (94% yield) in one pot. Hexamethylphosphoramide (HMPA) was found to be crucial to enhance the nucleophilicity of the magnesium alkoxide. 18 was subjected to the magnesiumhalogen exchange conditions (EtMgBr) to generate a functionalized Grignard reagent 45,47,[64][65][66] , which was quenched by CS 2 and MeI to give dithioester 19. It is noteworthy that lithium-halogen exchange did not lead to a satisfactory result. The desilylation took place spontaneously during acid workup to deliver alcohol 20 in 67% yield from 18. Oxidation of 20 with Dess-Martin periodinane (DMP) afforded ketone 21 (83% yield) without destroying the sulfur-containing functionalities, and the subsequent methanolysis furnished thioester 4 in 66% yield.
Completion of the synthesis. With both fragments in hand, we directed our attention to the construction of the aromatic B ring, as depicted in Fig. 5. It is well documented in the literature of Barton-Kellogg olefination that thioketones readily react with diazo compounds without promoters or catalysts [55][56][57] . After examination of the conventional conditions, we realized that the stabilized diazoketone 3 needed to be activated by Rh 2 (OAc) 4 to form the metal-carbenoid intermediate 52,54,[67][68][69][70] , which was further trapped by relatively unreactive thioester 4. The in situ to afford tetrasubstituted olefin 2 in 85% overall yield. We examined a series of conditions such as heating or FeCl 3 to promote the last C-C bond formation but only observed decomposition. Inspired by our synthesis of daphenylline 46 , we irradiated 2 with ultraviolet light (l ¼ 365 nm). To our delight, this symmetrical olefin underwent a 6p electrocyclization, presumably to provide pentacyclic intermediate 23, which was spontaneously oxidized under an air atmosphere during workup to furnish tetramethyl clostrubin 24 (55% yield from 2). Global deprotection of the methyl groups with aqueous HBr (48 wt%) gave clostrubin (1) with excellent efficiency. The spectra and physical properties of the synthetic 1 are consistent with those reported for the natural product. In total, 200 mg of 1 were obtained through the synthesis.

Discussion
In summary, we have accomplished the first total synthesis of clostrubin. The concise and efficient route took advantage of the 6p electrocyclization strategy as well as the inherent structural symmetry of the molecule. The synthesis provides a practical means to obtain this potent antibiotic for further investigations, considering the limited supply and difficult isolation of the naturally occurring sample.

Methods
General. All reactions were carried out under an argon atmosphere with dry solvents under anhydrous conditions, unless otherwise noted. Tetrahydrofuran was distilled immediately before use from sodium-benzophenone ketyl. Methylene chloride, N,N-dimethylformamide, triethylamine, N,N-diisopropylethylamine and chlorotrimethylsilane were distilled from calcium hydride and stored under an argon atmosphere. Methanol was distilled from magnesium and stored under an argon atmosphere. Reagents were purchased at the highest commercial quality and used without further purification, unless otherwise stated. Solvents for chromatography were used as supplied by Titan Chemical. Reactions were monitored by thin-layer chromatography carried out on S-2 0. 25