Gregory A. Ellis

Boronate-Mediated Biologic Delivery

Inefficient cellular delivery limits the landscape of macromolecular drugs. Boronic acids readily form boronate esters with the 1,2- and 1,3-diols of saccharides, such as those that coat the surface of mammalian cells. Here pendant boronic acids are shown to enhance the cytosolic delivery of a protein toxin. Thus, boronates are a noncationic carrier that can deliver a polar macromolecule into mammalian cells.

Structure and biosynthesis of the antibiotic bottromycin D.

Drug resistant infectious diseases are quickly becoming a global health crisis. While Streptomyces spp. have been a major source of antibiotics over the past 50 years, efficient methods are needed to identify new antibiotics and greatly improve the rate of discovery. LCMS-based metabolomics were applied to analyze extracts of 50 Streptomyes spp. Using this methodology, we discovered bottromycin D and used whole genome sequencing to determine its biosynthesis by a ribosomal pathway.

Solwaric acids A and B, antibacterial aromatic acids from a marine Solwaraspora sp.

Two novel trialkyl-substituted aromatic acids, solwaric acids A and B, were isolated from a marine Solwaraspora sp. cultivated from the ascidian Trididemnum orbiculatum. Solwaric acids A and B were isotopically labeled with U-13C glucose, and analysis of a 13C–13C COSY allowed for unambiguous determination of the location of the phenyl methyl group. The two novel compounds demonstrated antibacterial activity against methicillin-resistant Staphylococcus aureus (MRSA) and methicillin-sensitive Staphylococcus aureus (MSSA).

Investigation of Interspecies Interactions within Marine Micromonosporaceae Using an Improved Co-Culture Approach

With respect to bacterial natural products, a significant outcome of the genomic era was that the biosynthetic potential in many microorganisms surpassed the number of compounds isolated under standard laboratory growth conditions, particularly among certain members in the phylum Actinobacteria. Our group, as well as others, investigated interspecies interactions, via co-culture, as a technique to coax bacteria to produce novel natural products. While co-culture provides new opportunities, challenges exist and questions surrounding these methods remain unanswered. In marine bacteria, for example, how prevalent are interspecies interactions and how commonly do interactions result in novel natural products? In an attempt to begin to answer basic questions surrounding co-culture of marine microorganisms, we have tested both antibiotic activity-based and LC/MS-based methods to evaluate Micromonosporaceae secondary metabolite production in co-culture. Overall, our investigation of 65 Micromonosporaceae led to the identification of 12 Micromonosporaceae across three genera that produced unique metabolites in co-culture. Our results suggest that interspecies interactions were prevalent between marine Micromonosporaceae and marine mycolic acid-containing bacteria. Furthermore, our approach highlights a sensitive and rapid method for investigating interspecies interactions in search of novel antibiotics, secondary metabolites, and genes.

Total Syntheses of Proposed (±)-Trichodermatides B and C.

Total syntheses of putative (±)-trichodermatides B and C are described. These efficient syntheses feature the oxa-[3 + 3] annulation strategy, leading to B and C along with their respective C2-epimers. However, these synthetic samples are spectroscopically very different from the natural products. DFT calculations of C13 chemical shifts are conducted and the predicted values are in good agreement with those of synthetic samples, thereby questioning in the accuracy of structural assignments of trichodermatides B and C.

Potentiation of ribonuclease cytotoxicity by a poly(amidoamine) dendrimer

Variants of bovine pancreatic ribonuclease (RNase A) engineered to evade the endogenous ribonuclease inhibitor protein (RI) are toxic to human cancer cells. Increasing the basicity of these variants facilitates their entry into the cytosol and thus increases their cytotoxicity. The installation of additional positive charge also has the deleterious consequence of decreasing ribonucleolytic activity or conformational stability. Here, we report that the same benefit can be availed by co-treating cells with a cationic dendrimer. We find that adding the generation 2 poly(amidoamine) dendrimer in trans increases the cytotoxicity of RI-evasive RNase A variants without decreasing their activity or stability. The increased cytotoxicity is not due to increased RI-evasion or cellular internalization, but likely results from improved translocation into the cytosol after endocytosis. These data indicate that co-treatment with highly cationic molecules could enhance the efficacy of ribonucleases as chemotherapeutic agents.

Chemical Genomics, Structure Elucidation, and in Vivo Studies of the Marine-Derived Anticlostridial Ecteinamycin

A polyether antibiotic, ecteinamycin (1), was isolated from a marine Actinomadura sp., cultivated from the ascidian Ecteinascidia turbinata. 13C enrichment, high resolution NMR spectroscopy, and molecular modeling enabled elucidation of the structure of 1, which was validated on the basis of comparisons with its recently reported crystal structure. Importantly, ecteinamycin demonstrated potent activity against the toxigenic strain of Clostridium difficile NAP1/B1/027 (MIC = 59 ng/μL), as well as other toxigenic and nontoxigenic C. difficile isolates both in vitro and in vivo. Additionally, chemical genomics studies using Escherichia coli barcoded deletion mutants led to the identification of sensitive mutants such as trkA and kdpD involved in potassium cation transport and homeostasis supporting a mechanistic proposal that ecteinamycin acts as an ionophore antibiotic. This is the first antibacterial agent whose mechanism of action has been studied using E. coli chemical genomics. On the basis of these data, we propose ecteinamycin as an ionophore antibiotic that causes C. difficile detoxification and cell death via potassium transport dysregulation.

Brackish habitat dictates cultivable Actinobacterial diversity from marine sponges

Bacterial communities associated with marine invertebrates such as sponges and ascidians have demonstrated potential as sources of bio-medically relevant small molecules. Metagenomic analysis has shown that many of these invertebrates harbor populations of Actinobacteria, many of which are cultivable. While some populations within invertebrates are transmitted vertically, others are obtained from the environment. We hypothesized that cultivable diversity from sponges living in brackish mangrove habitats have associations with Actinobacterial populations that differ from those found in clear tropical waters. In this study, we analyzed the cultivable Actinobacterial populations from sponges found in these two distinct habitats with the aim of understanding the secondary metabolite potential. Importantly, we wanted to broadly evaluate the potential differences among these groups to guide future Actinobacterial collection strategies for the purposes of drug discovery.