Synthesis of the N-((1E)-alkenyl)-(2Z,4Z)-heptadienamide side chain of salicylihalamide A and apicularens A and B

[reaction: see text] The unstable N-((1E)-alkenyl)-(2Z,4Z)-heptadienamide side chain of salicylihalamide A (1) and apicularens A and B (3 and 4) has been prepared in one pot by the addition of (1Z,3Z)-hexadienylcuprate, prepared in situ from EtLi, CuBr.SMe2, and acetylene, to a (1 E)-alkenyl isocyanate.     

Total synthesis of (-)-salicylihalamide A

[see structure]. A 16-step synthesis of the novel cytotoxin salicylihalamide A (1E) has been achieved in 3.3% overall yield using ring closing metathesis to generate the macrolide and addition of (1Z,3Z)-hexadienylcuprate (2), which was generated in situ from ethylcuprate and acetylene, to alkenyl isocyanate 3 to form the side chain.     

Total synthesis of (-)-salicylihalamide

A concise total synthesis of the potent cytotoxic marine natural products salicylihalamide A and B (la, b) is reported. Key steps of our approach were the asymmetric hydrogenation reactions of beta-keto esters 18 and 32 catalyzed by [((S)-BINAP)Ru-Cl2]2. NEt3 and the cyclization of the macrolide core by ring closing olefin metathesis (RCM) using the "second-generation" ruthenium carbene complex 24 as the catalyst which bears an imidazol-2-ylidene ligand. The EIZ ratio obtained in this macrocyclization reaction was determined by the protecting groups at the remote phenolic OH group of the cyclization precursor. The elaboration of the resulting cycloalkene 37 into the final target involved a CrCl2-mediated synthesis of vinyliodide 49 which, after deprotection, did undergo a copper-catalyzed cross-coupling process with the (Z,Z)-configurated carboxamide 42 to form the labile enamide moiety of 1. Compound 42 was derived from a palladium-catalyzed Negishi coupling between butynylzinc chloride and 3-iodoacrylate 39 followed by a Lindlar reduction of enyne 40 thus obtained and a final aminolysis of the ester group.     

Stereochemical determination and complex biosynthetic assembly of etnangien, a highly potent RNA polymerase inhibitor from the myxobacterium Sorangium cellulosum

A potent novel analogue of the natural macrolide antibiotic etnangien, a structurally unique RNA polymerase inhibitor from myxobacteria, is reported. It may be readily obtained from fermentation broths of Sorangium cellulosum and shows high antibiotic activity, comparable to that of etnangien. However, it is much more readily available than the notoriously labile authentic natural product itself. Importantly, it is stable under neutral conditions, allowing for elaborate NMR measurements for assignment of the 12 hydroxyl- and methyl-bearing stereogenic centers. The full absolute and relative stereochemistries of these complex polyketides were determined by a combination of extensive high-field NMR studies, including J-based configuration analysis, molecular modeling, and synthetic derivatization in combination with an innovative method based on biosynthetic studies of this polyketide which is also presented here. A first look into the solution conformation and 3D structure of these promising macrolide antibiotics is reported. Finally, the complete biosynthetic gene cluster was analyzed in detail, revealing a highly unusual and complex trans-AT type polyketide biosynthesis, which does not follow colinearity rules, most likely performs programmed iteration as well as module skipping, and exhibits HMG-CoA box-directed methylation.     

Synthesis and evaluation of iron chelators with masked hydrophilic moieties

Synthetic iron chelators based on the natural siderophore ferrichrome have previously been shown to bind Fe(III) with high affinity (pKf > 27) and have shown no toxicity to mammalian cell cultures in vitro. A new class of lipophilic ferrichrome analogues carrying acetoxymethyl ester moieties has been synthesized. We have shown that these molecules penetrate rapidly through cell membranes and turn highly hydrophilic while inside the cells, upon esterase mediated hydrolysis of the lipophilic termini. The intracellular retention was visualized by labeling these analogues with a fluorescent naphthalic diimide probe. The prohydrophilic iron chelators have been shown to inhibit the metal-catalyzed intracellular formation of reactive oxygen species with high effectivity, and preliminary results suggest these molecules to be potent antimalarial agents.     

Palladium-catalyzed cross-coupling of aryl chlorides and triflates with sodium cyanate: a practical synthesis of unsymmetrical ureas

An efficient method for palladium-catalyzed cross-coupling of aryl chlorides and triflates with sodium cyanate is reported. The protocol allows for the synthesis of unsymmetrical N,N'-di- and N,N,N'-trisubstituted ureas in one pot and is tolerant of a wide range of functional groups. Insight into the mechanism of aryl isocyanate formation was gleaned through studies of the transmetalation and reductive elimination steps of the reaction, including the first demonstration of reductive elimination from an arylpalladium isocyanate complex to produce an aryl isocyanate.     

SAR Studies of the Leupyrrins: Design and Total Synthesis of Highly Potent Simplified Leupylogs

Leupyrrins are highly potent antifungal agents. A structure–activity-relationship study of natural and synthetic derivatives is reported which reveals important insights into the biological relevance of several structural subunits leading to the discovery of highly potent but drastically simplified leupylogs that incorporate a stable and readily available aromatic side chain. For their synthesis a concise strategy is described that enables a short and versatile access.     

Total synthesis of pinnatoxins A and G and revision of the mode of action of pinnatoxin A

Pinnatoxins belong to an emerging class of potent marine toxins of the cyclic imine group. Detailed studies of their biological effects have been impeded by unavailability of the complex natural product from natural sources. This work describes the development of a robust, scalable synthetic sequence relying on a convergent strategy that delivered a sufficient amount of the toxin for detailed biological studies and its commercialization for use by other research groups and regulatory agencies. A central transformation in the synthesis is the highly diastereoselective Ireland-Claisen rearrangement of a complex α,α-disubstituted allylic ester based on a unique mode for stereoselective enolization through a chirality match between the substrate and the lithium amide base. With synthetic pinnatoxin A, a detailed study has been performed that provides conclusive evidence for its mode of action as a potent inhibitor of nicotinic acetylcholine receptors selective for the human neuronal α7 subtype. The comprehensive electrophysiological, biochemical, and computational studies support the view that the spiroimine subunit of pinnatoxins is critical for blocking nicotinic acetylcholine receptor subtypes, as evidenced by analyzing the effect of a synthetic analogue of pinnatoxin A containing an open form of the imine ring. Our studies have paved the way for the production of certified standards to be used for mass-spectrometric determination of these toxins in marine matrices and for the development of tests to detect these toxins in contaminated shellfish.     

Looking glass inhibitors: L-DMDP, a more potent and specific inhibitor of alpha-glucosidases than the enantiomeric natural product DMDP

L-DMDP, prepared from D-gulonolactone, is a highly specific inhibitor of a number of plant and mammalian alpha-glucosidases [between 2 and 4 orders of magnitude more potent than the enantiomeric natural product DMDP] but is not an inhibitor of bacterial and yeast alpha-glucosidases. Additionally N-butyl-DMDP is a potent inhibitor of ceramide-specific glucosyltransferase but N-butyl-L-DMDP shows no inhibition.     

Structure-based design of potent non-peptide MDM2 inhibitors

A successful structure-based design of a class of non-peptide small-molecule MDM2 inhibitors targeting the p53-MDM2 protein-protein interaction is reported. The most potent compound 1d binds to MDM2 protein with a Ki value of 86 nM and is 18 times more potent than a natural p53 peptide (residues 16-27). Compound 1d is potent in inhibition of cell growth in LNCaP prostate cancer cells with wild-type p53 and shows only a weak activity in PC-3 prostate cancer cells with a deleted p53. Importantly, 1d has a minimal toxicity to normal prostate epithelial cells. Our studies provide a convincing example that structure-based strategy can be employed to design highly potent, non-peptide, cell-permeable, small-molecule inhibitors to target protein-protein interaction, which remains a very challenging area in chemical biology and drug design.     

Synthesis of the proposed structure of mucoxin via regio- and stereoselective tetrahydrofuran ring-forming strategies

An enantioselective total synthesis of the proposed structure of mucoxin (1) is described. Mucoxin, an annonaceous acetogenin isolated from bioactive leaf extracts of Rollinia mucosa, is the first acetogenin containing a hydroxylated trisubstituted tetrahydrofuran (THF) ring. This natural product is a highly potent and specific antitumor agent against MCF-7 (breast carcinoma) cell lines (ED50 = 3.7 x 10(-3) microg/mL compared to adriamycin, ED50 = 1.0 x 10(-2) microg/mL). The total synthesis described herein features two regio- and stereoselective THF ring-forming reactions. The 2,3,5-trisubstituted THF portion (C13-C17) was accessed using a highly regioselective cyclization of a methylene-interrupted epoxydiol, and the 2,5-disubstituted THF ring (C8-C12) was conveniently assembled via a 1,2-n-triol cyclization strategy. The spectral data of the synthetic material and two of its diastereomers did not match the reported data for the natural product. On the basis of detailed spectroscopic analysis of the synthesized molecule, we reason that the spectral discrepancies are due to stereochemical misassignment of the natural product.     

Structural rationale for the cross-resistance of tumor cells bearing the A399V variant of elongation factor eEF1A1 to the structurally unrelated didemnin B,ternatin, nannocystin A and ansatrienin B

At least four classes of structurally distinct natural products with potent antiproliferative activities target the translation elongation factor eEF1A1, which is best known as the G-protein that delivers amino acyl transfer RNAs (aa-tRNAs) to ribosomes during mRNA translation. We present molecular models in atomic detail that provide a common structural basis for the high-affinity binding of didemnin B, ternatin, ansatrienin B and nannocystin A to eEF1A1, as well as a rationale based on molecular dynamics results that accounts for the deleterious effect of replacing alanine 399 with valine. The proposed binding site, at the interface between domains I and III, is eminently hydrophobic and exists only in the GTP-bound conformation. Drug binding at this site is expected to disrupt neither loading of aa-tRNAs nor GTP hydrolysis but would give rise to stabilization of this particular conformational state, in consonance with reported experimental findings. The experimental solution of the three-dimensional structure of mammalian eEF1A1 has proved elusive so far and the highly homologous eEF1A2 from rabbit muscle has been crystallized and solved only as a homodimer in a GDP-bound conformation. Interestingly, in this dimeric structure the large interdomain cavity where the drugs studied are proposed to bind is occupied by a mostly hydrophobic α-helix from domain I of the same monomer. Since binding of this α-helix and any of these drugs to domain III of eEF1A(1/2) is, therefore, mutually exclusive and involves two distinct protein conformations, one intriguing possibility that emerges from our study is that the potent antiproliferative effect of these natural products may arise not only from inhibition of protein synthesis, which is the current dogma, but also from interference with some other non-canonical functions. From this standpoint, this type of drugs could be considered antagonists of eEF1A1/2 oligomerization, a hypothesis that opens up novel areas of research.     

Novel glutathione conjugates of phenyl isocyanate identified by ultra‐performance liquid chromatography/electrospray ionization mass spectrometry and nuclear magnetic resonance

Phenyl isocyanate is a highly reactive compound that is used as a reagent in organic synthesis and in the production of polyurethanes. The potential for extensive occupational exposure to this compound makes it important to elucidate its reactivity towards different nucleophiles and potential targets in the body. In vitro reactions between glutathione and phenyl isocyanate were studied. Three adducts of glutathione with phenyl isocyanate were identified using ultra‐performance liquid chromatography/electrospray ionization mass spectrometry and nuclear magnetic resonance (NMR). Mass spectrometric data for these adducts have not previously been reported. Nucleophilic attack on phenyl isocyanate occurred via either the cysteinyl thiol group or the glutamic acid α‐amino group of glutathione. In addition, a double adduct was formed by the reaction of both these moieties. NMR analysis confirmed the proposed structure of the double adduct, which has not previously been described. These results suggest that phenyl isocyanate may react with free cysteines, the α‐amino group and also with lysine residues whose side chain contains a primary amine. Copyright © 2014 John Wiley & Sons, Ltd.     

Total Synthesis and Conformational Study of Callyaerin A: Anti‐Tubercular Cyclic Peptide Bearing a Rare Rigidifying (Z)‐2,3‐ Diaminoacrylamide Moiety

The first synthesis of the anti‐TB cyclic peptide callyaerin A ( 1 ), containing a rare (Z)‐2,3‐diaminoacrylamide bridging motif, is reported. Fmoc‐formylglycine‐diethylacetal was used as a masked equivalent of formylglycine in the synthesis of the linear precursor to 1 . Intramolecular cyclization between the formylglycine residue and the N‐terminal amine in the linear peptide precursor afforded the macrocyclic natural product 1 . Synthetic 1 possessed potent anti‐TB activity (MIC₁₀₀=32 μm ) while its all‐amide congener was inactive. Variable‐temperature NMR studies of both the natural product and its all‐amide analogue revealed the extraordinary rigidity imposed by this diaminoacrylamide unit on peptide conformation. The work reported herein pinpoints the intrinsic role that the (Z)‐2,3‐diaminoacrylamide moiety confers on peptide bioactivity.     

Formal total synthesis of salicylihalamides A and B

An efficient synthesis of the macrolactone 3 of the salicylihalamides in 10 linear steps from alkene 6 is described. The key steps involved a Stille coupling between the chiral stannane 5 and benzyl bromide 4, which produced alkene 15 in good yield, and subsequent base-induced macrolactonization then gave compound 3. Macrolactone 3 was then converted into the known salicylihalamide A intermediate 18 in a three-step sequence. Compound 3 was also converted into another known salicylihalamide A and B intermediate 23 in a five-step sequence.     

Ring-closing metathesis: a powerful tool for the synthesis of simplified salicylihalamide-based V-ATPase inhibitors

Based on our synthetic strategy developed for the total synthesis of the macrocyclic salicylate natural product salicylihalamide, we describe herein the synthesis of a series of simplified salicylihalamide-based analogs. Alterations in the aromatic fragment, the macrolactone scaffold and side-chain were evaluated for in vitro inhibition of V-ATPase activity and human tumor cell growth.     

Nitrosyl isocyanate (ONNCO): gas-phase generation and a HeI photoelectron spectroscopy study

The nitrosyl isocyanate (ONNCO) has been generated from a heterogeneous reaction of gaseous nitrosyl chloride with silver isocyanate and studied for the first time in the gas phase. This structurally and energetically novel transient specimen is characterized by HeI photoelectron spectroscopy combined with DFT calculations. Both the calculations and the spectroscopic results suggest that the molecule adopts an open-chain trans structure. The observed decomposition products indicate the formation of ONNCO and further confirm the previously reported decomposition pathway.     

Synthesis and structure-activity relationship of N-arylrolipram derivatives as inhibitors of PDE4 isozymes

Structure activity studies of N-phenylrolipram derivatives have led to the identification of highly potent PDE4 inhibitors. The potential of these inhibitors for cellular activity was routinely assessed in an assay of fMLP induced oxidative burst in human eosinophils. Since first generation PDE4 inhibitors have been plagued with a number of unwanted side effects, parallel structure activity studies for competition with the [3H]-rolipram binding site in rat brain were performed. In this fashion 5-[4-(3-cyclopentyloxy-4-methoxyphenyl)-2-oxo-pyrrolidin-1-yl]-3-(3-methoxybenzyloxy)benzoic acid N',N'-dimethylhydrazide (22) was identified as a potent inhibitor of PDE4 which exhibits >1000 fold selectivity versus PDE3, and is a nanomolar inhibitor in all the cellular assays tested. Studies on the stereoselectivity of PDE4 inhibition of this class of rolipram based compounds revealed, that for example (S)-11 is a more potent inhibitor than (R)-11. This effect can also be observed in primary human cells where the (S)-enantiomer is about 10 fold more potent than the corresponding (R)-enantiomer.     

Telomestatin: formal total synthesis and cation-mediated interaction of its seco-derivatives with G-quadruplexes

The structurally unique natural product telomestatin incorporates seven oxazole rings and one sulfur-containing thiazoline in a macrocyclic arrangement. The compound is a potent inhibitor of the enzyme telomerase and therefore provides a structural framework for developing new potential therapeutic agents for cancer. An efficient formal total synthesis of telomestatin is reported in which the key steps are the use of dirhodium(II)-catalyzed reactions of diazocarbonyl compounds to generate six oxazole rings, demonstrating the power of rhodium carbene methodology in organic chemical synthesis. CD spectroscopy establishes that seco-derivatives of telomestatin are potent stabilizers of G-quadruplex structures derived from the human telomeric repeat sequence. Mass spectrometry studies, confirmed by molecular dynamics simulations, provide the first evidence that high affinity binding to terminal G-tetrads in both 1:1 and 2:1 ligand complexes is mediated through the macrocycle coordinating a monovalent cation, with selectivity for the antiparallel structure.     

Bent π-Conjugation within a Macrocycle: Asymmetric Total Syntheses of Spirohexenolides A and B

Macrocycles with bent π-conjugation motif are extremely rare in nature and synthetically daunting and anticancer haouamines and spirohexenolides were representative of such rare natural products with synthetically challenging bent π-conjugation within a macrocycle. While the total synthesis of haouamines has been elegantly achieved, spirohexenolides remains an unmet synthetic challenge due to the highly strained bent 1,3,5-triene conjugation within C15 macrocycle. Inspired by the chemical synthesis of cycloparaphenylenes (CPPs) and haouamines, herein we devise a synthetic strategy to overcome the highly strained bent 1,3,5-triene conjugation within the macrocycle and achieve the first, asymmetric total synthesis of spirohexenolides A (>20 mg) and B (>50 mg). Our synthesis features strategic design of ring-closing metathesis (RCM) macrocyclization followed by double dehydration to achieve the C15 macrocycle with the deformed nonplanar 1,3,5-triene conjugation. In addition, we have developed a new enantioselective construction of highly functionalized spirotetronate fragment (northeast moiety) through RCM and Ireland–Claisen rearrangement. Our in vitro bioassay studies reveal that both spirohexenolides are cytotoxic against a panel of human cancer cells with IC₅₀ 1.2–13.3 μM and spirohexenolide A is consistently more potent (up to 3 times) than spirohexenolide B, suggesting the importance of alcohol for their bioactivity and for medicinal chemistry development.