Total Synthesis of the Reported Structure of Neaumycin B

Abstract

The stereoselective total synthesis of structure 1 assigned to the macrolide natural product neaumycin B is reported in a 2.3% overall yield on 90 mg scale. The synthesis features a gram-scale nickel-catalyzed reductive cross-coupling/spiroketalization tactic to construct the spiroketal core of neaumycin B. The stereostructures of the C3–C6, C8–C14, and C20–C41 segments of synthetic neaumycin B were unambiguously verified by X-ray crystallography.

The first congener of neaumycin was isolated in 2012 by Shen et al. from the soil actinomycete Streptomyces sp. NEAU-x21.¹ The structure of neaumycin was then substantially revised in 2015, with the isolation of neaumycin A and the congener neaumycin B, albeit without stereochemistry.² In 2018, Jenson and Fenical et al. isolated a substance from a marine microbial Micromonospora sp. (strain CNY-010) from the surface of the tropical brown alga Stypopodium zonale collected in the Bahamas Islands.³ A combination of genomic data and 2D NMR studies led to the assignment of neaumycin B to be 1. Preliminary in vitro study of neaumycin B against several cancer cell lines displayed significant potency (LD₅₀: 5.6 × 10⁻⁵ μg/mL), in particular with selectivity toward U87 human glioblastoma, which is among the most malignant types of gliomas.⁴ The bioactivity of neaumycin B holds promise as a lead structure for drug design. The development of a total synthesis of neaumycin B (1) would thus be of significance and, as such, has drawn considerable interest.^5a–c Herein we report the first total synthesis of the reported structure of neaumycin B (1).

From the retrosynthetic perspective (Scheme 1), we envisioned that neaumycin B (1) could be dissected into a linear southern (C1–C17) hemisphere (2) and a spirocyclic northern hemisphere (C18–C41) (3), which in turn could be united via Stille coupling⁶ and macrolactonization, to complete, upon deprotection, neaumycin B (1).

Scheme 1.

Structure and Retrosynthetic Analysis of Neaumycin B (1)

The linear southern hemisphere 2 (Scheme 1) was envisioned to arise from an asymmetric 1,2-addition of southwestern (7) and southeastern (8) fragments, while the spiroketal core of the northern hemisphere (3) could be constructed from the linear ketone precursor 4 (Scheme 1), which in turn could be dissected into the northwestern (5) and northeastern (6) fragments.

Our synthesis began with the northwestern fragment 5 (Scheme 2), the C21–C29 segment of neaumycin B. Union of epoxide (+)-9 (prepared in two steps from a known compound; see SI for details) and dithiane 10^7b proceeded smoothly via a Brook rearrangement/epoxide ring opening sequence^7a–c to yield adduct 11, which underwent benzyl protection, hydrolysis of trimethylsilyl ether, and Cu/TEMPO-catalyzed aerobic oxidation⁸ to aldehyde 12 (see X-ray), gratifyingly with no oxidation of sulfur or epimerization at the α position of the carbonyl. A Felkin–Anh selective aldol reaction of the enolate derived from methyl acetate with 12 then delivered the desired syn adduct 13 with good diastereoselectivity (15:1). Dithioacetal hydrolysis⁹ and Evans–Saksena reduction¹⁰ followed by 1,3-diol protection led to compound 15, with the desired anti configuration. Saponification of the methyl ester and subsequent thioesterification completed synthesis of northwestern fragment 5 on a decagram scale.

Scheme 2.

Synthesis of the Northwestern Fragment (5)

We next turned to northeastern (C30–C41) fragment 6 (Scheme 3). Regioselective opening of epoxy alcohol (−)-16 (91% ee, readily prepared from (E)-2-hexene-1-ol via Sharpless epoxidation¹¹) with trimethylaluminum¹² led almost exclusively to 1,2-diol 17 (>20:1 1,2-diol/1,3-diol) as indicated by the crude ¹H NMR (see Figure S27), which upon biphasic periodate cleavage and dibromo-olefination¹³ furnished dibromo-alkene 18 on a 38 g scale. Exposure of 18 to n-BuLi and capture of the lithium alkynylide with formaldehyde delivered propargylic alcohol 19. Trans-hydrosilylation directed by the hydroxyl group employing ruthenium catalysis^14a–d furnished alcohol 20, with both good regio-, Z/E selectivity and yield. Oxidation of the resulting allylic hydroxyl with MnO₂ then afforded quantitatively aldehyde 21, which underwent an Evans aldol¹⁵ reaction with 22 to form 23; subsequent transamidation yielded Weinreb amide 24, with the desired syn configuration (82%, 2 steps). Monoaddition of allylmagnesium bromide to amide 24 then yielded the β-hydroxyl ketone, which was followed by Narasaka–Prasad reduction^16a–c (Et₂BOMe, NaBH₄) and acetal protection to access 25. Hydroboration/oxidation, followed by an Appel reaction,¹⁷ completed the northeastern fragment 6.

Scheme 3.

Synthesis of the Northeastern Fragment (6)

With northwestern and northeastern fragments 5 and 6 in hand, we set out to examine suitable methods for their union (Scheme 4). Although neither organolithium nor Grignard chemistry successfully delivered the desired coupled ketone in satisfactory yield, fragment coupling was achieved via the recently developed nickel-catalyzed reductive cross-coupling protocol¹⁸ to afford ketone 4 in good yield (76%) on a gramscale. Ketone 4 was then exposed to p-toluenesulfonic acid in methanol to achieve deprotection/spiroketalization and to furnish 26 and the epimer 26′, as a 1:0.8 mixture, which were separable by chromatography. Pleasingly, exposure of the pure undesired epimer 26′ to the same acidic condition reestablished the equilibrium, thereby permitting harvest of 26 upon each chromatography separation/re-equilibration to achieve 26 in 67% overall yield. A key NOE correlation between H-33 and H-28 (Scheme 4, see Figure S53) verified the stereogenicity of 26.

Scheme 4.

Fragment Union/Spiroketalization

To achieve 3,4-dimethoxybenzyl protection at the C27-OH (Scheme 5), 26 was treated first with catalytic tetra-n-butylammonium fluoride (TBAF)¹⁹ to affect closure of the five-membered siloxane ring across C35–C37 and leave the C27-OH exposed for selective DMB protection (see Supporting Information for details). Subsequent exposure of the siloxane to methyllithium then unmasked the C35-OH to give 27, which now serves as the directing group for the future stereoselective epoxidation.

Scheme 5.

Synthesis of the Northern Hemisphere (3a)

With the thus locked conformation at the C35–C36 bond in 27, due to A^1,3 strain invoked by the preinstalled C37 silyl group,¹⁴ epoxidation of the C36–C37 olefin proceeded with exclusive syn-selectivity with vanadyl catalysis²⁰ to furnish 28. Desilylation with TBAF²¹ and methylation of C35-OH then gave 30, followed by hydrogenolysis of the C21 and C23 benzyl ethers to give diol 31. Chemoselective oxidation of the C21 primary alcohol next delivered β-hydroxy aldehyde 32, which upon treatment with the Wittig phosphoranylidene reagent,^22a,b yielded olefin 33 with excellent E selectivity. Pleasingly, compound 33 was crystalline, thus enabling unambiguous verification of the stereostructure spanning the C20–C41 segment by X-ray crystallography (see Scheme 5).

Next, triethyl silyl protection of the C23-OH in 33, reduction of ester, and MnO₂ oxidation furnished enal 34, which was treated with 2,3-dichloro-5,6-dicyano-1,4-benzoquinone (DDQ) to affect deprotection of the C27-OH to give 35. Finally, Takai–Utimoto olefination^23a,b of aldehyde 35 employing diiodo(tributylstannyl)methane led to 3a in good yield, but as an unacceptable 4:1 mixture of (E,E)/(Z,E) isomers.

We therefore revised our synthetic route to a second-generation northern hemisphere (3b, see Scheme 6). β-Hydroxy aldehyde 32 was first protected with triethylsilyl, and a Takai–Utimoto reaction^23a,b was carried out. The steric bulk at the α-carbon next to the carbonyl permitted Takai olefination with CrCl₂/Bu₃SnCHI₂ to proceed with excellent E-selectivity (>20:1), leading to the corresponding vinylstannane.^23a,b Iodination then afforded E-iodide 37 in 78% yield over two steps. DDQ removal of the 3,4-dimethoxybenzyl group then gave 38. Next Stille cross-coupling^24c between alkenyl iodide 38 and germylstannane 39^24a–c afforded dienyl germane 40, with no isomerization of the diene. Iodination of germane 40 with N-iodosuccinimide then provided access to the second-generation northern hemisphere (3b) with excellent stereospecificity.

Scheme 6.

Synthesis of the Second-Generation Northern Hemisphere (3b)

Turning to the linear southwestern and southeastern fragments (Scheme 7a and b), synthesis of alkyne 7 began from known aldehyde 41 (prepared from d-xylose in five steps^25a,b). Marshall asymmetric propargylation²⁶ delivered the anti-configured homopropargylic alcohol 42 in good yield, but with modest 4:1 diastereoselectivity (Scheme 7). Adduct 42 was then treated with TBAF to afford diol 43 with excellent purity after chromatography. To gain proof of the stereostructure of 7, PMP acetalization of diol 43 furnished crystalline 44, which upon X-ray crystallography analysis verified the structure of the southwestern segment (C8–C14). Silyl ether protection then completed the southwestern fragment (7).

Scheme 7.

Synthesis of Southwestern (7) and Southeastern (8) Fragments

Synthesis of the southeastern fragment (8, Scheme 7b) began with β-keto ester 45. Catalytic asymmetric dynamic kinetic resolution²⁷ successfully delivered alcohol 46, in good yield and excellent enantio- and diastereoselectivity. Tert-Butyldimethylsilyl (TBS) protection and diisobutylaluminum (DIBAL) reduction of the ester then afforded compound 47, which upon 4-methoxybenzyl (PMB) protection of the primary alcohol followed by removal of the TMS group under mild basic conditions then revealed terminal alkyne 48. Semihydrogenation, hydroboration/oxidation, and Cu/(2,2,6,6-tetramethylpiperidin-1-yl)oxyl (TEMPO)-catalyzed aerobic oxidation8 completed the Southeastern fragment 8.

Having prepared alkyne 7 and aldehyde 8 (Scheme 8) on a multigram scale, Carreira asymmetric alkynylation²⁸ cleanly afforded adduct 51 with excellent yield and diastereoselectivity. Subsequent trans-reduction of the triple bond with conventional aluminum hydride reagents (LiAlH₄, Red-Al), however, failed to give the desired allylic alcohol in appreciable yield. We therefore turned to a Ru-catalyzed hydrostannation/destannation tactic,^29a,b which gratifyingly led to the desired trans-reduction in good yield (72%). Stereochemical assessment of the C7-hydroxyl of 53 was achieved via Mosher ester analysis³⁰ (see Table S27). Methylation then led to compound 54, which, after exposure to 50% aqueous trifluoroacetic acid (TFA), cleanly furnished triol 55 as the major product. Selective tosylation and epoxide ring closure, followed by reinstallation of the silyl protecting group, then gave 56 in high overall yield (92%, three steps), which was subjected to BF₃-assisted nucleophilic ring opening with lithium trimethylsilyl (TMS) acetylide to furnish alcohol 57 in near quantitative yield. Methylation employing Meerwein’s reagent gave 58, which was followed by global silyl group removal with TBAF and the hydroxyl groups reprotected with triethylsilyl, which at the end of our synthesis proved much easier to remove. The PMB protecting group in 59 was next removed, and in turn Dess–Martin oxidation³¹ of the resulting alcohol followed by Horner–Wadsworth–Emmons (HWE) olefination³² led to 61. Finally, bromination of the terminal alkyne and hydrostannation³³ permitted regio- and stereoselective formation of the vinylstannane, completing the synthesis of the linear southern hemisphere 2 on a 1.26 g scale.

Scheme 8.

Synthesis of the Southern Hemisphere (2)

With both the northern (3b) and southern (2) hemispheres successfully prepared (Scheme 9), a Stille union reaction⁶ united 2 and 3. To our delight, the desired (E,E,E)-1,3,5-triene 62 was formed in excellent yield on an appreciable 200 mg scale with no isomerization of double bonds! The methyl ester was next hydrolyzed to afford seco-acid 63 via transesterification with trimethyltin hydroxide.³⁴ Macrolactonization, employing Mukaiyama’s conditions,³⁵ afforded the silylprotected macrocycle 64 in good yield.³⁶ Finally, global removal of triethylsilyl groups gratifyingly proceeded cleanly under mild conditions (TBAF, HOAc, and 0 °C) to complete the synthesis of the reported structure of neaumycin B (1) isolated as a white powder. Notably, 1 was prepared on a 90 mg scale in a single batch, with a 2.3% overall yield.

Scheme 9.

End Game for the Proposed Structure of Neaumycin B (1)

Unfortunately, the ¹H NMR spectra of synthetic 1 displayed significant deviations from that of the spectra reported by Fenical et al.³ In addition, 1 displayed no activity against glioblastoma cells (see Figure S5). Importantly, the stereo-chemical assignments of synthetic 1 were derived from X-ray crystallography analysis of each fragment (Figure 1). That is, crystal structures of compounds 33, 44, and 50 (see SI for details) confirmed the stereochemistry that spans C20–C41, C8–C14, and C3–C6 of synthetic neaumycin B (see Figure 1), respectively. The stereogenicity at C7 was also confirmed by Mosher ester analysis (see Table S27 for details). Based on the evidence reported here, we are confident that the synthetic neaumycin B (1) prepared here matches the structure reported by Fenical et al.³

Figure 1.

Crystal Structures of Each Fragment.