Enantioselective Total Synthesis of Cannogenol-3-O-α-L-Rhamnoside via Sequential Cu(II)-Catalyzed Michael Addition/Intramolecular Aldol Cyclization Reactions

Abstract

A concise and scalable enantioselective total synthesis of the natural cardenolides cannogenol and cannogenol-3-O-α-L-rhamnoside has been achieved in 19 linear steps. The synthesis features a Cu(II)-catalyzed enantioselective and diastereoselective Michael reaction/tandem aldol cyclization and a one-pot reduction/transposition, which resulted in a rapid (6 linear steps) assembly of a functionalized intermediate containing C19 oxygenation that could be elabo-rated to cardenolide cannogenol. In addition, a strategy for achieving regio- and stereoselective glycosylation at the C3 position of synthetic cannogenol was developed and applied to the preparation of cannogenol-3-O-α-L-rhamnoside.

Graphical abstract

Cardiotonic steroids represent a large and important family of natural steroids that exhibit a wide spectrum of biological activities. The majority of these compounds are potent Na⁺/K⁺ ATPase inhibitors, a feature that makes them of great value for the treatment of various heart-associated diseases.¹ Cardiotonic steroids are secreted endogenously and used for the regulation of essential life processes such as renal sodium pressure, sodium transport, arterial pressure, cell growth differentiation, apoptosis, fibrosis, immunity, carbohydrate metabolism and various central nervous functions.² The unique biological properties often coupled with challenging structural features prompted numerous semi-synthetic and synthetic efforts toward the preparation of various cardiotonic steroids and their analogs.³ These studies have resulted in many creative solutions that have enabled medicinal chemistry studies on many natural cardiotonic steroids and their analogs. These advances, however, are difficult to translate into the synthesis of highly oxygenated cardiotonic steroids, in particular, those carrying O-atoms at the C1, C11, C12 and C19 positions. Not surprisingly, many of these challenging targets have only been synthesized in the past decade.⁴

Our group has long-standing interests in developing concise synthetic approaches to cardiotonic steroids.⁵ Recently, we described a concise approach to cardenolide core that is based on sequential enantioselective Cu(II)-catalyzed Michael/intramolecular aldol cyclization reactions (Figure, 1B).^5a This strategy enabled rapid enantioselective formation of cardiotonic steroid skeleta (i.e. 6) featuring the oxidized C14 and C19 positions, but lacking the required oxygenation at the C3 position. Based on these studies, a new approach that was based on diastereoselective Michael reaction/intramolecular aldol cyclization of the functionalized β-ketoester 7 and enone 8 was developed.^5c The resultant annulated product could be rapidly advanced to the fully functionalized steroidal core 9. This steroidal intermediate was subsequently elaborated to the natural cardenolides 19-hydroxysarmentogenin and the trewianin aglycone bearing oxygenation at the C3, C11, C14 and C19 positions. These results suggest that Cu(II)-catalyzed Michael addition/Aldol cascade represents a powerful and straightforward method for achieving fast and selective syntheses of cardiotonic steroids with various oxidation patterns. To further demonstrate the generality and utility of this approach, our group targeted steroids possessing the C19 hydroxylation, but lacking the C11 hydroxyl group. This oxidation pattern is commonly found in various cardenolides including cannogenol (2), corotoxigenin, securigenol, pachygenin, and strophanthidin derivatives and their bufadienolide counterparts such as 19-hydroxybufalin (3). It should be noted that applying the previously developed synthetic approaches⁴ to such steroids (including our diastereoselective approach^5b depicted in Figure 1B) would require incorporating a strategy for a selective removal of the C11 alcohol in the presence of the C3, C14, C17 and C19 oxygenation as well as other functionalities. This task might be operationally challenging and would result in a significant increase in the overall number of steps. To circumvent these problems, we pursued a more direct approach that is based on our enantioselective Cu(II)-catalyzed Michael reaction/aldolization of 7 and 5 resulting in steroid 10 (Figure 1C). This is the first example of using this strategy in the synthesis of natural cardenolides, and the formation of 10 is accomplished on decagram scale with excellent overall yield and selectivities.

Figure 1.

Prior and current synthetic approaches.

Steroid 10 was then subjected to epimerization/reductive transposition sequence to provide intermediate 11 lacking the C11 oxygenation. This expedient synthesis of 11 was accomplished in only 6 linear steps, 24% yield and 92% ee on a 3 gram scale. This key intermediate may be converted to a various C19 oxygenated cardiotonic steroids, which was demonstrated by synthesizing cannogenol (2), a natural cardenolide with anticancer properties.⁶ Finally, this manuscript describes a strategy to accomplish a regioselective introduction of sugar at the less reactive C3 position of 2 to provide natural cardiac glycoside cannogenol-3-O-α-L-rhamnoside (1),⁷ an effective nanomolar growth inhibitor of several cancer cell lines.^{6a, 8} Our studies commenced with generating sufficient quantities of known precursors 5 and 7 (Scheme 1). Thus, β-ketoester 7 was prepared in three steps from 3-ethoxy-2-cyclohexenone (120 gram scale) and enone 5 was synthesized in two steps from 2-methyl-1,3-cyclopentanedione (60 gram scale). These substrates were subjected to asymmetric Michael reaction^5a using 2,2′-(cyclopropane-1,1-diyl)bis(4-phenyl-4,5-dihydrooxazole) ligand complexed with Cu(SbF₆)₂ as the catalyst (10 mol%) under neat conditions.⁹ Gratifyingly, this transformation proceeded efficiently, and Michael adduct 12 was consistently generated on 7.2 gram scale in 92% yield, >20:1 dr and 92% ee. This compound was then subjected to aldolization with p-TSA, which resulted in steroid 10 with unnatural α-C13/C14 configuration (57%, >20:1 dr, 14 gram scale). Our prior studies on a related system containing C11 oxygenation^5c indicated.

Scheme 1.

Synthesis of intermediate 11.

that such unnatural steroids could be epimerized under basic conditions to form the more stable natural β-configuration at the C13 and C14 stereosenters. Indeed, subjecting 10 to NaHMDS resulted in formation of the desired diastereomer 14 through the intermediacy of retro-aldol product 13 (64%, >20:1 dr; 7.3 g scale). Intermediate 14 was then treated with DIBAL-H (3 gram scale), which led to 15, which was stable at room temperature for up to 2 h. The tetraol 15 was not isolated, but rather the excess of DIBAL-H was quenched, and the crude mixture was heated at reflux in formic acid and 8:2 water:THF to accomplish transposition/vinyl chloride hydrolysis to produce a mixture of C17 epimers in 71% yield, 8:1 dr. This mixture was purified by SiO₂ column chromatography to afford α-C17 diastereomer 11 in 63% yield. This transformation allowed us to remove the unwanted oxygenation at C7 and introduced important oxygenation at C3 in a single step to set the proper steroidal framework mimicking the natural product. Thus, the key intermediate 11 was synthesized as a single diastereomer in 6 linear steps (9 overall steps) from commercially available building blocks in 31% overall yield and 92% ee.

With the key intermediate 11 in hand, we turned our attention to installation of the stereocenter at C5 and the butenolide ring at C17. Steroid 11 was subjected to hydrogenation with Pd/C in the presence of 1% potassium hydroxide in methanol (Scheme 2). This C19 alcohol-directed hydrogenation resulted in the exclusive formation of the β-C5 center. The subsequent global oxidation of the crude material using Dess-Martin periodinane (DMP) followed by the selective reduction of the C19 aldehyde and C3 ketone in the presence of the C17 ketone by a bulky K-selectride yielded 16 in 62% yield over three steps after a single purification at the end of third step. The iodination of 16 under Barton protocol¹⁰ produced vinyl iodide 17 in 77% yield. Compound 17 was benzoylated to provide monobenzoate 18b (90% yield) along with the crystalline bis-benzoate 18a in 8% yield, the absolute and the relative configuration of which was confirmed by x-ray crystallographic analysis.

Scheme 2.

Synthesis of intermediate 17 and Confirmation of its Absolute and Relative Configuration

As the conditons for the deprotection of the C19 benzoate were not compatible with the butenolide functionality, 17 was subjected to TBS protection (Scheme 3) resulting in intermediate 19 in 74%.

Scheme 3.

Synthesis of cannogenol from 17

This protection was necessary as the direct Stille coupling of 17 and 20 was sluggish and resulted in lower yields. In contrast, the Stille coupling of 19 and commercially available stannylated butenolide 20 proceeded efficiently and resulted in steroid 21 in 82% yield. This product was elaborated to cannogenol (2) via 3-step sequence in 69% yield.^4f,4h This sequence involved TMS protection of the C14 tertiary alcohol, hydrogenation of the Δ¹⁶-olefin, and global removal of the silyl protecting groups. The TMS protection of the C14 tertiary alcohol was required to achieve a steric bias for the subsequent hydrogenation to give exclusive β-C17, and the attempts to accomplish this protection prior to the Stille coupling reaction resulted in poor yields of the cross-coupled product.

Next, we investigated the possibility of adopting this route to the synthesis of cannogenol-3-O-α-L-rhamnoside (1). Our initial studies commenced with attempts to accomplish C3-selective glycosylation of triol 16 as a model substrate (Scheme 4). Attempts to glycosylate 16 with trichloroacetimidate 22 available in 3 steps from L-rhamnose^4b,4c,11 resulted in the equimolar mixture of products 23 and 24 formed in ~70% conversion. This result indicated that the C19 hydroxyl is more reactive than the secondary C3 alcohol.

Scheme 4.

Initial Glycosylation Studies

Anticipating similar reactivity trends for cannogenol (2), our following efforts were focused on selectively protecting the C19 position of 2 (Scheme 5). Due to its higher reactivity, the C19 hydroxyl could be selectively protected as a p-bromobenzoate (27) or an acetate (28). These derivatives were glycosylated with trichloroacetimidate 22 using TMSOTf as a promoter to afford protected α-glycosides 27 (36% yield) and 28 (56% yield). These glycosylated steroids were subjected to ester deprotection under basic conditions; however, this step represented a significant challenge. The use of various basic conditions for ester hydrolysis (i.e. Na₂CO₃, K₂CO₃, NaOMe, LiOH or NH₃) resulted in the deprotection of the benzoyl esters on rhamnose moiety. However, the deprotection of the C19 ester was slow and proceeded with the formation of multiple side-products due to concomitant opening and isomerization of butenolide. To address this problem, more labile C19 methoxyacetate 29 was prepared in 99% yield. This compound was subjected to TfOH-catalyzed glycosylation with 22, which resulted in α-rhamnoside 30 in 81% yield (>20:1 dr). Compound 30 was subjected to ammonia (50% solution in methanol) to provide the desired cannogenol-3-O-α-L-rhamnoside (1) in 69% yield. The ¹H, ¹³C NMR data and optical rotation data ([α]_D²⁰ = −10.8, c = 0.147 in MeOH, reported⁷ [α]_D²⁰ = −15.5, c = 0.55 in MeOH) of the synthetic 1 were in good agreement with the corresponding data obtained for the natural sample of 1.^6a,7,8

Scheme 5.

Synthesis of Cannogenol-3-O-α-L-rhamnoside (1) from cannogenol (2)

In conclusion, a new asymmetric approach to cardenolides carrying the C19 oxygenation has been developed and applied to the synthesis of the natural products cannogenol (2) and cannogenol-3-O-α-L-rhamnoside (1) possessing anticancer activity. This approach features 3 g scale enantioselective synthesis of the functionalized cardenolide core in 6 linear steps, 24% yield and 92% ee involving enantioselective Michael/tandem Aldol addition sequence established by our group earlier. This key intermediate could be elaborated to cardenolide cannogenol (2) in 9 steps, 27% yield. Cannogenol (2) contains multiple hydroxyl functionalities, and a strategy for the selective introduction of sugar moieties at the C3 position of 2 was developed and successfully applied to the formation of cannogenol-3-O-α-L-rhamnoside (1). The herein described strategies will expedite medicinal chemistry studies on C19-hydroxylated cardenolides related to cannogenol, and the exploration of 1, 2 and related analogs is the subject of ongoing investigation by our group.