Chemical Synthesis and Antigenicity Evaluation of an Aminoglycoside Trisaccharide Repeating Unit of Pseudomonas aeruginosa Serotype O5 O-Antigen Containing a Rare Dimeric-ManpN3NA.

Pseudomonas aeruginosa bacteria are becoming increasingly resistant against multiple antibiotics. Therefore, the development of vaccines to prevent infections with these bacteria is an urgent medical need. While the immunological activity of lipopolysaccharide O-antigens in P. aeruginosa is well-known, the specific protective epitopes remain unidentified. Herein, we present the first chemical synthesis of highly functionalized aminoglycoside trisaccharide 1 and its acetamido derivative 2 found in the P. aeruginosa serotype O5 O-antigen. The synthesis of the trisaccharide targets is based on balancing the reactivity of disaccharide acceptors and monosaccharide donors. Glycosylations were analyzed by quantifying the reactivity of the hydroxyl group of the disaccharide acceptor using the orbital-weighted Fukui function and dual descriptor. The stereoselective formation of 1,2-cis-α-fucosylamine linkages was achieved through a combination of remote acyl participation and reagent modulation. The simultaneous SN2 substitution of azide groups at C2' and C2″ enabled the efficient synthesis of 1,2-cis-ß-linkages for both 2,3-diamino-D-mannuronic acids. Through a strategic orthogonal modification, the five amino groups on target trisaccharide 1 were equipped with a rare acetamidino (Am) and four acetyl (Ac) groups. Glycan microarray analyses of sera from patients infected with P. aeruginosa indicated that trisaccharides 1 and 2 are key antigenic epitopes of the serotype O5 O-antigen. The acetamidino group is not an essential determinant of antibody binding. The ß-D-ManpNAc3NAcA residue is a key motif for the antigenicity of serotype O5 O-antigen. These findings serve as a foundation for the development of glycoconjugate vaccines targeting P. aeruginosa serotype O5.

New Strategies in the Efficient Total Syntheses of Polycyclic Natural Products.

Polycyclic natural products are an inexhaustible source of medicinal agents, and their complex molecular architecture renders challenging synthetic targets where innovative and effective approaches for their rapid construction are urgently required. The total synthesis of polycyclic natural products has witnessed exponential progression along with the emergence of new synthetic strategies and concepts, such as sequential C-H functionalizations, radical-based transformations, and functional group pairing strategies. Our group exerts continued interest in the construction of bioactive and structurally complex natural products as well as evaluation of the mode of action of these molecules. In this Account, we will showcase how these new synthetic strategies are employed and guide our total synthesis endeavors.During the last two decades, a series of remarkable advances in C-H functionalization have led to the emergence of many new approaches to directly functionalize C-H bonds into useful functional groups. These selective transformations have provided a great opportunity for the step- and atom-economical construction of key fragments in complex molecule synthesis. We recently furnished the total syntheses for polycyclic natural products: incarviatone A, chrysomycin A, polycarcin V, and gilvocarcin V by employing a multiple C-H bond functionalization strategy. The polysubstituted benzene or naphthalene skeleton was constructed through sequential and site-selective C-H functionalizations from readily available simple starting materials, which reduced the number of steps and streamlined synthesis.Recently, we have also completed the total syntheses for a number of skeletally diverse tetracyclic Isodon diterpenoids inspired by their biogenesis and radical-based retrosynthetic disconnections. Radical transformations are strategically and tactically utilized in our syntheses, and radical-based reactions, including organo-SOMO catalysis, Birch reduction, regioselective 1,6-dienyne reductive cyclization, visible-light-mediated Schenck ene reaction, and photoradical-mediated late-stage skeletal rearrangement, play significant roles in our synthetic endeavors. Protecting-group-free and scalable syntheses are also built into our work to achieve the "ideal" synthesis. Furthermore, our synthetic work reveals that late-stage skeletal rearrangement through a photo radical process is possible in a biological setting in complement with nature's carbocation chemistry in complex natural product biosynthesis.Lycopodium alkaloids are a large family of structurally unique polycyclic natural products with impressive biological activities. Owing to their fascinating polycyclic architectures and diverse biological activities, these alkaloids have continued to serve as targets as well as inspirations for the synthetic community for decades. To access these bioactive natural products or natural product-like molecules for biological exploration and drug discovery, we applied a novel functional group pairing strategy to furnish the total syntheses for several Lycopodium alkaloids and obtained numerous skeletally diverse compounds with structural complexity comparable to natural products.

Genetic Engineering in Combination with Semi-Synthesis Leads to a New Route for Gram-Scale Production of the Immunosuppressive Natural Product Brasilicardin†A.

Brasilicardin A (1) consists of an unusual anti/syn/anti-perhydrophenanthrene skeleton with a carbohydrate side chain and an amino acid moiety. It exhibits potent immunosuppressive activity, yet its mode of action differs from standard drugs that are currently in use. Further pre-clinical evaluation of this promising, biologically active natural product is hampered by restricted access to the ready material, as its synthesis requires both a low-yielding fermentation process using a pathogenic organism and an elaborate, multi-step total synthesis. Our semi-synthetic approach included a) the heterologous expression of the brasilicardin A gene cluster in different non-pathogenic bacterial strains producing brasilicardin A aglycone (5) in excellent yield and b) the chemical transformation of the aglycone 5 into the trifluoroacetic acid salt of brasilicardin A (1 a) via a short and straightforward five-steps synthetic route. Additionally, we report the first preclinical data for brasilicardin A.

Design, Multigram Synthesis, and in Vitro and in Vivo Evaluation of Propylamycin: A Semisynthetic 4,5-Deoxystreptamine Class Aminoglycoside for the Treatment of Drug-Resistant Enterobacteriaceae and Other Gram-Negative Pathogens.

Infectious diseases due to multidrug-resistant pathogens, particularly carbapenem-resistant Enterobacteriaceae (CREs), present a major and growing threat to human health and society, providing an urgent need for the development of improved potent antibiotics for their treatment. We describe the design and development of a new class of aminoglycoside antibiotics culminating in the discovery of propylamycin. Propylamycin is a 4'-deoxy-4'-alkyl paromomycin whose alkyl substituent conveys excellent activity against a broad spectrum of ESKAPE pathogens and other Gram-negative infections, including CREs, in the presence of numerous common resistance determinants, be they aminoglycoside modifying enzymes or rRNA methyl transferases. Importantly, propylamycin is demonstrated not to be susceptible to the action of the ArmA resistance determinant whose presence severely compromises the action of plazomicin and all other 4,6-disubstituted 2-deoxystreptamine aminoglycosides. The lack of susceptibility to ArmA, which is frequently encoded on the same plasmid as carbapenemase genes, ensures that propylamycin will not suffer from problems of cross-resistance when used in combination with carbapenems. Cell-free translation assays, quantitative ribosome footprinting, and X-ray crystallography support a model in which propylamycin functions by interference with bacterial protein synthesis. Cell-free translation assays with humanized bacterial ribosomes were used to optimize the selectivity of propylamycin, resulting in reduced ototoxicity in guinea pigs. In mouse thigh and septicemia models of Escherichia coli, propylamycin shows excellent efficacy, which is better than paromomycin. Overall, a simple novel deoxy alkyl modification of a readily available aminoglycoside antibiotic increases the inherent antibacterial activity, effectively combats multiple mechanisms of aminoglycoside resistance, and minimizes one of the major side effects of aminoglycoside therapy.

Chemical synthesis of marine saponins.

Covering: 1989-2017 Saponins are characteristic metabolites of starfish and sea cucumbers, and occasionally are also found in sponges, soft coral, and small fish. These steroid or triterpenoid glycosides often show remarkable biological and pharmacological activities, such as antifungal, antifouling, shark repellent, antitumor and anti-inflammatory activities. Over one thousand marine saponins have been characterized; the majority of them can be categorized into three major structural types, i.e., asterosaponins, polyhydroxysteroid glycosides, and holostane glycosides. Thus far, only 12 marine saponins have been synthesized; those representing the major types were successfully synthesized recently. The syntheses involve preparation of the aglycones from the terrestrial steroid or triterpene materials, installation of the carbohydrate units, and manipulation of the protecting groups. Herein, we provide a comprehensive review on these syntheses.

Diverse Synthesis of Natural Trehalosamines and Synthetic 1,1'-Disaccharide Aminoglycosides.

A general strategy for the diverse synthesis of ten disaccharide aminoglycosides, including natural 2-trehalosamine (1), 3-trehalosamine (2), 4-trehalosamine (3), and neotrehalosyl 3,3'-diamine (8) and synthetic aminoglycosides 4-7, 9, and 10, has been developed. The aminoglycoside compounds feature different anomeric configurations and numbers of amino groups. The key step for the synthesis was the glycosylation coupling of a stereodirecting donor with a configuration-stable TMS glycoside acceptor. Either the donor or acceptor could be substituted with an azido group. The aminoglycosides prepared in the present study were characterized by 1D and 2D NMR spectroscopy.

Biosynthetic and Synthetic Strategies for Assembling Capuramycin-Type Antituberculosis Antibiotics.

Mycobacterium tuberculosis (Mtb) has recently surpassed HIV/AIDS as the leading cause of death by a single infectious agent. The standard therapeutic regimen against tuberculosis (TB) remains a long, expensive process involving a multidrug regimen, and the prominence of multidrug-resistant (MDR), extensively drug-resistant (XDR), and totally drug-resistant (TDR) strains continues to impede treatment success. An underexplored class of natural products-the capuramycin-type nucleoside antibiotics-have been shown to have potent anti-TB activity by inhibiting bacterial translocase I, a ubiquitous and essential enzyme that functions in peptidoglycan biosynthesis. The present review discusses current literature concerning the biosynthesis and chemical synthesis of capuramycin and analogs, seeking to highlight the potential of the capuramycin scaffold as a favorable anti-TB therapeutic that warrants further development.

Fight Against Antimicrobial Resistance: We Always Need New Antibacterials but for Right Bacteria.

Antimicrobial resistance in bacteria is frightening, especially resistance in Gram-negative Bacteria (GNB). In 2017, the World Health Organization (WHO) published a list of 12 bacteria that represent a threat to human health, and among these, a majority of GNB. Antibiotic resistance is a complex and relatively old phenomenon that is the consequence of several factors. The first factor is the vertiginous drop in research and development of new antibacterials. In fact, many companies simply stop this R&D activity. The finding is simple: there are enough antibiotics to treat the different types of infection that clinicians face. The second factor is the appearance and spread of resistant or even multidrug-resistant bacteria. For a long time, this situation remained rather confidential, almost anecdotal. It was not until the end of the 1980s that awareness emerged. It was the time of Vancomycin-Resistance Enterococci (VRE), and the threat of Vancomycin-Resistant MRSA (Methicillin-Resistant Staphylococcus aureus). After this, there has been renewed interest but only in anti-Gram positive antibacterials. Today, the threat is GNB, and we have no new molecules with innovative mechanism of action to fight effectively against these bugs. However, the war against antimicrobial resistance is not lost. We must continue the fight, which requires a better knowledge of the mechanisms of action of anti-infectious agents and concomitantly the mechanisms of resistance of infectious agents.

Total Synthesis of a Densely Functionalized Plesiomonas shigelloides Serotype 51 Aminoglycoside Trisaccharide Antigen.

Plesiomonas shigelloides, a pathogen responsible for frequent outbreaks of severe travelers' diarrhea, causes grave extraintestinal infections. Sepsis and meningitis due to P. shigelloides are associated with a high mortality rate as antibiotic resistance increases and vaccines are not available. Carbohydrate antigens expressed by pathogens are often structurally unique and are targets for developing vaccines and diagnostics. Here, we report a total synthesis of the highly functionalized trisaccharide repeating unit 2 from P. shigelloides serotype 51 from three monosaccharides. A judicious choice of building blocks and reaction conditions allowed for the four amino groups adorning the sugar rings to be installed with two N-acetyl (Ac) groups, rare acetamidino (Am), and d-3-hydroxybutyryl (Hb) groups. The strategy for the differentiation of amino groups in trisaccharide 2 will serve well for the syntheses of other complex glycans.

Differential Effects of Linkers on the Activity of Amphiphilic Tobramycin Antifungals.

As the threat associated with fungal infections continues to rise and the availability of antifungal drugs remains a concern, it becomes obvious that the need to bolster the antifungal armamentarium is urgent. Building from our previous findings of tobramycin (TOB) derivatives with antifungal activity, we further investigate the effects of various linkers on the biological activity of these aminoglycosides. Herein, we analyze how thioether, sulfone, triazole, amide, and ether functionalities affect the antifungal activity of alkylated TOB derivatives against 22 Candida, Cryptococcus, and Aspergillus species. We also evaluate their impact on the hemolysis of murine erythrocytes and the cytotoxicity against mammalian cell lines. While the triazole linker appears to confer optimal activity overall, all of the linkers incorporated into the TOB derivatives resulted in compounds that are very effective against the Cryptococcus neoformans species, with MIC values ranging from 0.48 to 3.9 µg/mL.

Asymmetric Total Synthesis of Brasilicardins.

Brasilicardins, bacterial diterpenoid natural products that display highly potent immunosuppressive activity, are promising immunosuppressant drug candidates. Structurally, they can be described as hybrids of terpenoids, amino acids, and saccharides, and share a characteristic highly strained anti-syn-anti-fused perhydrophenanthrene terpenoid scaffold (ABC-ring system) with two quaternary asymmetric carbon atoms. A unified and stereoselective total synthesis of all four brasilicardins has been designed based on the strategic use of an intramolecular conjugate addition. The ABC-ring system was initially constructed with high stereocontrol by novel intramolecular conjugate additions of Weinreb amides and in situ generated (Z)-vinyl copper species. The late-stage common intermediate was subjected to stereoselective installation of the amino acid component, followed by introduction of the saccharide unit via glycosylation to accomplish the total synthesis of brasilicardins A-D. Our synthesis offers opportunities to synthesize various brasilicardin analogues for biological and pharmacological investigations.

Streamlined Total Synthesis of Trioxacarcins and Its Application to the Design, Synthesis, and Biological Evaluation of Analogues Thereof. Discovery of Simpler Designed and Potent Trioxacarcin Analogues.

A streamlined total synthesis of the naturally occurring antitumor agents trioxacarcins is described, along with its application to the construction of a series of designed analogues of these complex natural products. Biological evaluation of the synthesized compounds revealed a number of highly potent, and yet structurally simpler, compounds that are effective against certain cancer cell lines, including a drug-resistant line. A novel one-step synthesis of anthraquinones and chloro anthraquinones from simple ketone precursors and phenylselenyl chloride is also described. The reported work, featuring novel chemistry and cascade reactions, has potential applications in cancer therapy, including targeted approaches as in antibody-drug conjugates.

Total Synthesis of Trioxacarcins DC-45-A1, A, D, C, and C7″-epi-C and Full Structural Assignment of Trioxacarcin C.

Trioxacarcins DC-45-A2, DC-45-A1, A, D, C7″-epi-C, and C have been synthesized through stereoselective strategies involving BF3·Et2O-catalyzed ketone-epoxide opening and gold-catalyzed glycosylation reactions, and the full structural assignment of trioxacacin C was deciphered via the syntheses of both of its C7″ epimers. The gathered knowledge sets the foundation for the design, synthesis, and biological evalution of analogues of these natural products as potential payloads for antibody-drug conjugates and other delivery systems for targeted and personalized cancer chemotherapy.

Multivalency in the recognition and antagonism of a HIV TAR RNA-TAT assembly using an aminoglycoside benzimidazole scaffold.

Recognition of RNA by high-affinity binding small molecules is crucial for expanding existing approaches in RNA recognition, and for the development of novel RNA binding drugs. A novel neomycin dimer benzimidazole conjugate 5 (DPA 83) was synthesized by conjugating a neomycin-dimer with a benzimidazole alkyne using click chemistry to target multiple binding sites on HIV TAR RNA. Ligand 5 significantly enhances the thermal stability of HIV TAR RNA and interacts stoichiometrically with HIV TAR RNA with a low nanomolar affinity. 5 displayed enhanced binding compared to its individual building blocks including the neomycin dimer azide and benzimidazole alkyne. In essence, a high affinity multivalent ligand was designed and synthesized to target HIV TAR RNA.

N-glycosyl-N-hydroxysulfamides as potent inhibitors of Brucella suis carbonic anhydrases.

We investigated a series of N-hydroxysulfamides obtained by Ferrier sulfamidoglycosylation for the inhibition of two bacterial carbonic anhydrases (CAs, EC 4.2.1.1) present in the pathogen Brucella suis. bsCA I was moderately inhibited by these compounds with inhibition constants ranging between 522 and 958 nM and no notable differences of activity between the acetylated or the corresponding deacetylated derivatives. The compounds incorporating two trans-acetates and the corresponding deprotected ones were the most effective inhibitors in the series. bsCA II was better inhibited, with inhibition constants ranging between 59.8 and 799 nM. The acetylated derivatives were generally better bsCA II inhibitors compared to the corresponding deacetylated compounds. Although these compounds were not highly isoform-selective CA inhibitors (CAIs) for the bacterial over the human CA isoforms, some of them possess inhibition profiles that make them interesting leads for obtaining better and more isoform-selective CAIs targeting bacterial enzymes.

Enantioselective synthesis of putative lipiarmycin aglycon related to fidaxomicin/tiacumicin†B.

An enantioselective synthesis of a putative lipiarmycin aglycon was accomplished and features: 1) Brown's enantioselective alkoxyallylboration and allylation of aldehydes, 2) chain elongation by iterative Horner-Wadsworth-Emmons olefination, 3) Evans' aldol reaction and 4) an ene-diene ring-closing metathesis. A neighboring-group-assisted chemoselective reductive desilylation was uncovered in this study and was instrumental to the realization of the present synthesis.

Total synthesis of the protected aglycon of fidaxomicin (tiacumicin†B, lipiarmycin†A3).

Fidaxomicin, also known as tiacumicin B or lipiarmycin A3, is a novel macrocyclic antibiotic that is used in hospitals for the treatment of Clostridium difficile infections. This natural product has also been shown to have excellent bactericidal activity against multidrug-resistant Mycobacterium tuberculosis. In spite of its attractive biological activity, no total synthesis has been reported to date. The enantioselective synthesis of the central 18-membered macrolactone is reported herein. The key reactions include ring-closing metathesis between a terminal olefin and a dienoate moiety for macrocyclization, a vinylogous Mukaiyama aldol reaction, and a Stille coupling reaction of sterically demanding substrates. The retrosynthesis involves three medium-sized fragments, thus leading to a flexible yet convergent synthetic route.

Total synthesis of trioxacarcin DC-45-A2.

An enantioselective total synthesis of trioxacarcin DC-45-A2 (1) featuring a novel Lewis acid-induced cascade rearrangement of epoxyketone 6 to forge the polyoxygenated 2,7-dioxabicyclo[2.2.1]heptane core of the molecule is described.

Total synthesis of the tiacumicin†B (lipiarmycin A3/fidaxomicin) aglycone.

Tiacumicin B (lipiarmycin A3, fidaxomicin) is an atypical macrolide antibiotic which is used for the treatment of Clostridium difficile infections. Tiacumicin B is also a potent inhibitor of Mycobacterium tuberculosis, but due to its limited oral bioavailability is unsuitable for systemic therapy. To provide a basis for structure-activity studies that might eventually lead to improved variants of tiacumicin B, we have developed an efficient approach to the synthesis of the tiacumicin B aglycone. The synthesis features a high-yielding intramolecular Suzuki cross-coupling reaction to effect macrocyclic ring closure. Key steps in the synthesis of the macrocyclization precursor were a highly selective, one-pot Corey-Peterson olefination and an ene-diene cross-metathesis reaction. Depending on the reaction conditions, the final deprotection delivered either the fully deprotected tiacumicin B aglycone or partially protected versions thereof.

Di-N-Methylation of Anti-Gram-Positive Aminoglycoside-Derived Membrane Disruptors Improves Antimicrobial Potency and Broadens Spectrum to Gram-Negative Bacteria.

The effect of di-N-methylation of bacterial membrane disruptors derived from aminoglycosides (AGs) on antimicrobial activity is reported. Di-N-methylation of cationic amphiphiles derived from several diversely structured AGs resulted in a significant increase in hydrophobicity compared to the parent compounds that improved their interactions with membrane lipids. The modification led to an enhancement in antibacterial activity and a broader antimicrobial spectrum. While the parent compounds were either modestly active or inactive against Gram-negative pathogens, the corresponding di-N-methylated compounds were potent against the tested Gram-negative as well as Gram-positive bacterial strains. The reported modification offers a robust strategy for the development of broad-spectrum membrane-disrupting antibiotics for topical use.