Biogenesis-Guided Synthesis and Structural Revision of Sarocladione Enabled by Ruthenium-Catalyzed Endoperoxide Fragmentation.

Sarocladione is the first 5,10:8,9-diseco-steroid with a 14-membered macrocyclic diketone framework to have been isolated from a natural source. Herein we report a biomimetic synthesis of sarocladione in only two or seven steps from inexpensive, commercially available ergosterol. The key feature of this synthesis was a novel ruthenium-catalyzed endoperoxide fragmentation, which transformed various saturated endoperoxides into olefinic diketones by cleavage of two C-C bonds. This synthesis allowed us to unambiguously determine the structure of sarocladione and provided experimental support for its revised biosynthetic origin. This work also vividly demonstrates that consideration of the biogenesis is a powerful tool for elucidating the structures of natural products.

Total Synthesis of Aplysiasecosterol A.

Aplysiasecosterol A (1) is a structurally unusual 9,11-secosteroid isolated from the sea hare Aplysia kurodai. We have accomplished the first and asymmetric total synthesis of 1 in a convergent fashion. The left-hand segment bearing three adjacent stereocenters was constructed through desymmetrizing reduction, ketalization, and radical cyclization. A strategy of asymmetric 2-bromoallylation followed by spontaneous desymmetrizing lactolization enabled a more expeditious access to this segment. The right-hand segment was prepared through two different approaches: one featuring Myers alkylation and Suzuki-Miyaura coupling and the other relying upon Aggarwal lithiation-borylation and Zweifel-Evans olefination. The two fragments were coupled by a Reformatsky type reaction. The three consecutive stereocenters embedded in the central domain of 1 were generated by an iron-mediated, hydrogen atom transfer based radical cyclization reaction.

Synthesis of Strophasterol†A Guided by a Proposed Biosynthesis and Innate Reactivity.

The synthesis of strophasterol A, a moderator of endoplasmatic reticulum (ER) stress in Alzheimer's disease, and the first member of a structurally unprecedented class of secosterols, was achieved through the implementation of a key step of its proposed biosynthesis and two C-H oxidations. Analysis of the innate reactivity of the intermediates enabled the identification of a novel way to prepare an α-chloro-γ-hydroxy-δ-keto enone, as well as its vinylogous α-ketol rearrangement to a δ-keto carboxylic acid.

Mimicking/extracting structure and functions of natural products: synthetic approaches that address unexplored needs in chemical biology.

Natural products are often attractive and challenging targets for synthetic chemists, and many have interesting biological activities. However, synthetic chemists need to be more than simply suppliers of compounds to biologists. Therefore, we have been seeking ways to actively apply organic synthetic methods to chemical biology studies of natural products and their activities. In this personal review, I would like to introduce our work on the development of new biologically active compounds inspired by, or extracted from, the structures of natural products, focusing on enhancement of functional activity and specificity and overcoming various drawbacks of the parent natural products.

Biomimetic Synthesis of the CDE Ring Moiety of Physalins, Complex 13,14-Secosteroids.

Physalins are a structurally complex family of 13,14-secosteroids isolated from the genus Physalis. We disclose a two-step construction of the CDE ring moiety of the physalins from a steroidal compound bearing 14-OH, 18-COOMe, and 17, 20-α-epoxide based on our biosynthetic proposal. C13-C14 bond cleavage by an alkoxy radical at C-14 and spontaneous epoxide ring opening gave a compound having a cyclononene and γ-lactone. Diastereoselective dihydroxylation of the resulting alkene with OsO4 provided the CDE ring moiety of physalin.

Synthesis of DFGH-Ring Derivatives of Physalins via One-Pot Construction of GH-Ring and Evaluation of Their NF-κB-Inhibitory Activity.

We designed and synthesized a series of derivatives containing the right-side DFGH-ring structure of physalin-type natural products, decorated with a hydrophobic substituent. The synthetic scheme utilizes a highly efficient, one-pot protocol for simultaneous construction of the GH-ring system, promoted by HF/pyridine. Among the compounds synthesized, 5d inhibited TNF-α-stimulated NF-κB activation with similar potency to physalin B.

Synthesis of the DFGH ring system of type B physalins: highly oxygenated, cage-shaped molecules.

Let the dominos fall: Synthesis of the complex DFGH ring system of the title compounds has been accomplished. The approach features simple treatment of the key intermediate with a Brønsted base to afford the tetracyclic cage-shaped target in one pot through a four-step domino transformation (see scheme; Mc = monochloromesylate, MOM = methoxymethyl).

Synthesis of 5,10-seco analogs of testosterone.

A number of 5,10-seco analogs of testosterone has been synthesized starting from products of the radical oxidation of 3beta,17beta-diacetoxy-5alpha-androstan-5alpha-ol. The obtained compounds possess a flexible 10-membered ring with substituents (O, -OH) at C-3 and C-5. Similar derivatives with an (E)- and (Z)-Delta(1(10))-double bond have been prepared also. X-ray analysis and a combination of NMR experiments have been used for their structure elucidation and conformation analysis.

Synthesis of cytotoxically active derivatives based on alkylated 2,3-seco-triterpenoids.

Extremely low content of biologically active triterpenoids with the fragmented or contracted ring A extractable from plants is the main disadvantage of their use in drug discovery and practical pharmacology. Development of new methods for synthesis of these compounds and their structural analogs from bioavailable triterpene precursors gives an opportunity to obtain promising agents for pharmacology with excellent yields. A new approach to synthesis of alkylated A-seco-triterpenoids, including the Beckmann fragmentation of 3-methyl-substituted allobetulin or betulinic acid methyl ester with 2-hydroxyimino group in the ring A was proposed. These compounds were used to prepare a series of 2,3-seco- and five-membered ring A lupane and oleanane derivatives, cytotoxicity of which was screened in vitro against the cancer (HEp-2, HCT 116, A549, RD TE32, MS) and non-cancerous (HEK 293) cell lines. Methyl 3-bromomethyl-1-cyano-3-oxo-2,3-seco-2-norlup-20(29)-en-30-al-28-oate was selected as the most active compound (IC50 3.4-10.4 µM for HEp-2, HCT 116, RD TE32, MS cells) capable of triggering caspase-8-mediated apoptosis in HCT 116 cells accompanied by typical apoptotic chromatin condensation, without any loss of mitochondrial membrane permeability.

A new type of steroids with a cyclobutane fragment in the AB-ring moiety.

The synthesis of a 5,10-seco steroid containing two double bonds in a AB-macrocycle as well as the preparation of a steroidal skeleton with a cyclobutane fragment is described. The structures of these compounds are different from those of natural steroids, but they are very similar with respect to conformation of the carbon skeleton.

Antihormonal potential of selected D-homo and D-seco estratriene derivatives.

Since many estrogen derivatives exhibit anti-hormone or enzyme inhibition potential, a large number of steroidal derivatives have been synthesised from appropriate precursors, in order to obtain potential therapeutics for the treatment of hormone-dependent cancers. In molecular docking studies, based on X-ray crystallographic analysis, selected D-homo and D-seco estratriene derivatives were predicted to bind strongly to estrogen receptor α (ERα), aromatase and 17,20 lyase, suggesting they could be good starting compounds for antihormonal studies. Test results in vivo suggest that these compounds do not possess estrogenic activity, while some of them showed weak anti-estrogenic properties. In vitro anti-aromatase and anti-lyase assays showed partial inhibition of these two enzymes, while some compounds activated aromatase. Aromatase activators are capable of promoting estrogen synthesis for treatment of pathological conditions caused by estrogen depletion, e.g. osteopenia or osteoporosis.

C-nor-9,11-secoestranes as modified estrogens and fertility regulation.

The synthesis of C-nor-9,11-secoestradiol (4) has been achieved from 17 beta-acetoxy-11-chloro-3-methoxy-C-nor-9,11-secoestra-1,3,5(10)-tr ien-9-one (1) through a sequence of reactions without affecting the stereochemistry of estradiol-17 beta. Removal of the 9-keto function of 1 by hydrogenolysis and its subsequent treatment with Na/NH3 gives C-nor-9,11-secoestradiol 3-(methyl ether) (3), which has been demethylated under alkaline conditions to furnish C-nor-9,11-secoestradiol (4). Pyridinium chlorochromate oxidation of 3 gives the corresponding 17-ketone 6. Jones' oxidation of 4 to the ketone 5 and reaction of 5 and 6 with lithium acetylide gives corresponding 17 alpha-ethynyl derivatives 7 and 8. Relative binding affinity to estradiol-17 beta receptors and uterotropic, antiuterotrophic, and antiimplantation activities of compounds 3-8 have been studied. The effect of conformational flexibility on ligand-receptor interaction of these compounds is discussed.

Studies in antifertility agents. 11. Secosteroids. 5. Synthesis of 9,11-secoestradiol.

9,11-Secoestradiol (9) and 11-hydroxy-9,11-secoestradiol (12) have been synthesized starting from 17-acetoxyestradiol 3-methyl ether (1) and found to possess significant antifertility activity in rats. 3-Methoxy-9,11-seco-9-oxo-17beta-acetoxyestra-1,3,5(10)-trien-11-oic acid (2), prepared by CrO3 oxidation of 1, on hydrogenolysis gave methyl 17beta-hydroxy-3-methoxy-9,11-secoestra-1,3,5(10)-triene-11-carboxylate (3). The 17-O-THP derivative of 3 was treated with LiAlH4 to give 17beta-(O-tetrahydropyranyl)-3-methoxy-11-hydroxy-9,11-secoestra-1,3,5(10)-triene (5). The 11-O-mesylate of 5 on LiAlH4 reduction followed by mild acid treatment and demethylation under alkaline conditions gave 9. LiAlH4 reduction of 3 gave 9,11-seco-11-hydroxyestradiol 3-methyl ether (11) which on demethylation gave 9,11-seco-11-hydroxyestradiol (12).

Studies in antifertility agents. 8. Seco steroids. 2. 5,6-Secoestradiol and some related compounds.

Three ring beta-secoestradiols, 2alpha,3beta- and 2beta,3beta-2-ethyl-3-(p-hydroxyphenyl)-6beta-methyl-trans-bicyclo[4.3.0]nonan-7beta-ols, have been synthesized and some of them shown to possess significant antiimplantation activity in rats.

17 alpha-O-(aminoalkyl)oxime derivatives of 3 beta,14 beta-dihydroxy-5 beta-androstane and 3 beta-hydroxy-14-oxoseco-D-5 beta-androstane as inhibitors of Na(+),K(+)-ATPase at the digitalis receptor.

The synthesis and binding affinities to the digitalis Na(+),K(+)-ATPase receptor of a series of 3 beta,14 beta-dihydroxy-5 beta-androstane and 3 beta-hydroxy-14-oxoseco-D-5 beta-androstane derivatives bearing a 17 alpha-(aminoalkoxy)imino chain are reported; some derivatives were also studied for their inotropic activity. Our recently proposed model of interaction of molecules with the digitalis receptor was used to design these compounds. On that basis, the possibility to design novel potent inhibitors of Na(+),K(+)-ATPase without being constrained by the stereochemistry of the classical digitalis skeleton in the D-ring region was predicted. The binding affinities of the most potent compounds in the two series, (EZ)-17 alpha-[2-[(2-aminoethoxy)imino]ethyl]-5 beta-androstane-3 beta,14 beta-diol (6f) and (EZ)-3 beta-hydroxy-17 alpha-[2-[(2-aminoethoxy)imino]ethyl]-14,15-seco-5 beta-androstan-14-one (24c) are higher than that of the potent natural compound digitoxigenin, despite the unusual alpha-exit of the substituent in position 17 of 6f or the disruption of the D-ring in 24c. These results further support the validity of our recently proposed model of binding at the digitalis receptor. Results of the inotropic tests on guinea pig atrium deserve further investigation on the pharmacological profile of these derivatives.

Synthesis, estrogen receptor binding, and tissue distribution of [18F]fluorodoisynolic acids.

Doisynolic acids, D-ring seco-steroids derived from alkaline fusion of estrones, are hormonal curiosities: Their binding affinity for the estrogen receptor is low (ca. 1-2% that of estradiol), but their in vivo potency is high and they have a long duration of action. To study the in vivo behavior of the doisynolic acids, we prepared fluorine-substituted analogs of both trans-doisynolic acid (with the natural 14 alpha-hydrogen configuration, trans-FDA) and the more active cis-doisynolic acid (with the unnatural 14 beta-hydrogen configuration, cis-FDA) from estrone and 14 beta-estrone, respectively. Modification of the D-ring haloform cleavage approach of Meyers allowed us to introduce fluorine (or fluorine-18) on the carbon atom derived from C-16 in the estrones. Fluorine substitution had little effect on the estrogen receptor binding affinity of the doisynolic acids. Tissue distribution of the fluorodoisynolic acids (trans-[18F]FDA and cis-[18F]FDA) was unusual and very different from that of typical, high-affinity ligands for the estrogen receptor. At 1-3 h in immature female rats, trans-[18F]FDA shows low and rather nonselective uptake in the principal estrogen target tissue (uterus) and slow clearance. By contrast, cis-[18F]FDA shows high uptake in nearly all tissues, with significant uterine uptake that continues to increase over the 1-6-h period. The uterine uptake of this isomer was blocked at the later times by a sufficiently high dose of unlabeled cis-FDA. After administration of the trans-[18F]FDA, a more polar metabolite slowly accumulates in the blood. The cis-[18F]FDA, however, showed no apparent metabolism, with 84% of the blood activity at 5 h assigned as the unmetabolized radioligand. After 5 h, only limited clearance from blood, liver, and kidneys has occurred. No metabolite from this isomer accumulates in the uterus. Although fluorodoisynolic acids will not be useful breast-tumor imaging agents, their behavior was found to be interesting as it deviates from that of other F-18 estrogens. Further long-term studies of cis-doisynolic acid, labeled with tritium, may be needed to explicate fully its unusual distribution properties and high in vivo activity.

The synthesis of functionalized 13,14-seco-steroids via Grob fragmentation.

A synthetic methodology for the synthesis of 13,14-seco-steroids with substituents at C-14 and C-17 is described. The approach involves Grob fragmentation of 14beta-hydroxy-17beta-tosylates, hydroboration-oxidation of the intermediate delta13(17)-olefin, and hydride reduction of the 14-ketone. An unambiguous structural assignment of (13R,14S,17S)-14,17-diacetoxy-3-methoxy-7alpha-methyl-13,14-secoestra-1,3,5(10)-triene was determined by X-ray analysis.

Synthesis of 13,14-secotestosterone derivatives.

A number of testosterone analogs with a 13,14-secosteroidal fragment have been prepared from (13S)-13-iodo-6beta-methoxy-3alpha, 5-cyclo-13,14-seco-5alpha-androstan-14,17-dione. The key steps involved stereoselective deiodination of the starting compound with triphenylphosphine and selective protection of the 17-keto group with trimethylsilylcyanide. Removal of iodine at C-13 proceeded with inversion of the configuration at C-13, which has been established by X-ray crystallography. 13,14-Secotestosterone analogues substituted and non-substituted at C-14 have been prepared. The obtained compounds containing flexible CD ring fragments are of great interest for comparative studies in biological tests together with testosterone and other steroids with a rigid tetracyclic skeleton.

Preparation of 2,3-seco-5 alpha-cholestane-2,3-diol and 4 alpha-methyl-2,3-seco-5 alpha-cholestane-2,3-diol and its reactions with o-nitrophenyl selenocyanate.

The reaction of 2,3-seco-5 alpha-cholestane-2,3-diol and 4 alpha-methyl-2,3-seco-5 alpha-cholestane-2,3-diol with o-nitrophenyl selenocyanate was studied. The diols were synthesized from cholesterol.

Studies of the synthesis of biomarkers. XI. Synthesis of 4,5-secocholestane and 4-methyl-4,5-secocholestane.

4,5-Secocholestane (1a) and 4-methyl-4,5-secocholestane (1b) were synthesized from cholesterol (2) in five and seven steps, respectively. The key intermediate, 5-oxo-4,5-secocholestan-4-al (7) was reduced by the Clemmensen method to afford 1a. Meanwhile, 7 underwent selective Wittig reaction, Clemmensen reduction, and hydrogenation to give another target molecule, 1b. The structure of an unknown biomarker was shown to be different from the proposed 1a by gas chromatographic and mass spectrometric comparison.