Abstracts E1-E11

Category E.  Chemical Biology and Drug Design

 

E1.  Synthesis of Electrophilic Sultams via Intramolecular C-Vinylation of a Triazole

Andie Jo Cassity, Jung Ho Jun, Nicole Marie Windmon, Naeem Asad, Kyu Ok Jeon, Anna J. Diepenbrock, and Paul R. Hanson

Department of Chemistry, University of Kansas, Lawrence KS, USA

The development of chemically unique electrophilic probes capable of modulating biological nucleophiles is the focal point of this project.  Toward this goal, an intramolecular C-vinylation method for the generation of electrophilic triazole-fused sultams is reported. This efficient method has enabled the synthesis of a small library of diverse sultams. The electrophilic character of these sultams has been studied using a variety of small nucleophiles, whereby scaffold reactivity screening, in combination with the reported method, is guiding efforts in second-generation probe design. 

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E2.  Computationally Designed Peptide Binds LPS

Jakki J. Deay1, Hui Sun Lee1,2, Pinakin R. Sukthankar1, Joanna S. Slusky1

1Department of Molecular Biosciences and Center for Computational Biology, University of Kansas, Lawrence, KS, USA;
2Department of Biological Sciences, Lehigh University, Bethlehem, PA, USA

Currently it takes days to determine if a patient has a bacterial infection, frequently leaving patients to be sent home with antibiotics before their test results are in. Over prescribing of unneeded antibiotics contributes to the growing epidemic of antibiotic resistant bacterium and could be avoided if there was a method to detect bacterial infections quickly. Lipopolysaccharides (LPS) are bacterial endotoxins unique to outer membranes of gram-negative bacteria and are an important target for quicker detection of pathogens. An LPS binding motif was identified from a crystal structure of an E. coli beta-barrel protein (FhuA) bound with LPS. As beta-barrel proteins are difficult to purify and manipulate, an alpha-helical peptide was computationally designed to mimic the LPS binding motif using preexisting helical templates and G-LoSA. Fluorescence anisotropy was used to probe the binding capabilities of the designed peptide with Escherichia coli, Salmonella enterica, Pseudomonas aeruginosa, and Klebsiella pneumoniae LPS. Polymyxin B is an antibiotic known to bind to LPS and was used as a positive control in the binding assay by labeling it two different fluorophores. The designed peptide was found to bind the four types of LPS with greater affinity than polymyxin B, with three of the four LPS samples binding to the peptide with approximately an order of magnitude greater affinity.

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E3.  An Efficient and Pot-economical Approach for the Syntheses of 13-Desmethyl-lyngbouilloside and Its Simplified Analogs

Arghya Ganguly, Salim Javed, Hannah Knapp, Gihan Disanayake,Dimuthu Vithanage and Paul R. Hanson*

Department of Chemistry, University of Kansas, Lawrence, KS, USA

Efforts towards an efficient, pot-economical asymmetric total synthesis of (–)-13-Desmethyl-lyngbouilloside, an unnatural analog of lyngbouilloside (anticancer activity) will be discussed. The key reactions involved in the syntheses of (–)-13-Desmethyl-lyngbouilloside are one-pot sequential RCM/CM/chemoselective hydrogenation, regio- and diastereoselective cuprate addition, a one-pot Pd-catalyzed reductive allylic transposition, Roskamp homologation and Boeckman acyl-ketene cyclization to form the macrocyclic core of the target molecule. This efficient, pot-economical and library amenable approach is further extended for the syntheses of diverse and structurally related analog library of (–)-lyngbouilloside and (+)-neopeltolide. An iterative regio- and diastereoselective cuprate addition and late stage alcohol functionalization followed by RCM/deprotection sequence enables the syntheses of a diverse range of sterically, electronically and stereochemically attenuated macrocycles containing carbon-, sulfur- and phosphorus warheads. These analogs will be submitted to our collaborators for biological screening.

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E4. Electrophilic Sultams: Diversity Oriented Synthesis to Develop Chemical Modifiers of Cysteine

Shazia Iqbal,a,b Maria A. Khan,a,b Qin Zang,a Elise Gao,a Joanna Loh,a Naeem Asad,a and Paul R. Hansona*

a Department of Chemistry, University of Kansas, Lawrence, KS, USA
b
H. E. J. Research Institute of Chemistry, International Center for Chemical and Biological Sciences, University of Karachi, Pakistan

Covalent modulators of cysteine residues have emerged as regulators of protein function, including ubiquitination, sumoylation, prenylation, farnesylation, phosphorylation and glycosylation to name a few.  In this regard, we report the synthesis of structurally diverse electrophilic sultam scaffolds as modulators of cysteine functionality. Employing 19F NMR analyses, their reactivity profiling with N-acetyl cysteine was examined through kinetic measurements. It was demonstrated that their reactivity with cysteine is highly dependant on the electronic effect, steric hindrance, as well as resident stereochemistry of the exo acetyl sultam scaffolds.

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E5. One-pot, sequential enyne ring-closing metathesis/Diels-Alder reaction for the synthesis of mono-, bi- and tricyclic P-heterocyclic compounds

Salim Javed, Dimuthu Vithanage, Arghya Ganguly, Gihan Dissanayake and Paul R. Hanson•

Department of Chemistry, University of Kansas, Lawrence, KS, USA

Development of an efficient, step and pot-economical approach for the synthesis of a library of bi- and tricyclic phosphorus heterocyles is reported. Phosphorus heterocycles have been shown to impart a wide variety of biological activities leading them to serve as novel pharmaceutical agents and biological probes. The key reactions involved in the synthesis of these small molecules include one-pot sequential, enyne ring-closing metathesis and Diels-Alder reaction (ERCM/DA). This  step economical protocol enables access to structurally and stereochemically diverse bi- and tricyclic phosphate and phosphoramidate analogs in fewer number of steps from easily accessible precursors.

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E6. Computational Chemical Biology Core: A Chemical Biology of Infectious Disease COBRE Core Laboratory

David K. Johnson1,2

1Computational Chemical Biology Core, University of Kansas, Lawrence, KS, USA;
2Molecular Graphics and Modelling Lab, University of Kansas, Lawrence, KS, USA

The University of Kansas Computational Chemical Biology Core (CCB) provides the computational resources and expertise to enhance the productivity of researchers studying infectious diseases. The CCB is able to provide or assist with virtual screening, protein-small molecule docking, binding site prediction, protein modeling and design, prediction of protein stability changes upon mutation, fragment based probe design, as well as preparation of presentation graphics. The core utilizes the KU Community Cluster at the Advanced Computing Facility for its high-performance computing needs. The KU Community Cluster offers 458 compute nodes with a total of 8,568 compute cores, including 17 nodes that offer GPU-accelerated computing. The CCB specializes in initial hit identification of non-traditional drug targets such as protein-protein or protein-RNA interfaces by offering high-throughput virtual screening via pocket optimization with exemplar screening at protein-protein interfaces and hotspot pharmacophore mimicry of protein-RNA interactions.

The CCB works in collaboration with the Molecular Graphics and Modeling Laboratory.

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E7. Novel Electrophilic Probes: Reactivity Profiling Studies of Electrophilic Sultams, Sulfonamides and Known Drugs

Jung Ho Jun, Andie Jo Cassity, Jay S. Jha, Collin D. Clay, and Paul R. Hanson*

Department of Chemistry, University of Kansas, Lawrence, KS, USA

Systematic studies of the role of thiol (or cysteine)-containing amino acids and peptides in many physiological processes have emerged. In particular, modification of thiol (or cysteine)-containing amino acids via Michael addition reactions have surfaced as modification of the ERK, Nrf2, and NF-kB biological pathways, as well as a potential cure for the parasitic disease known as Chagas disease. Thus, rapid, sensitive, and selective detection of regulatory thiols is of considerable importance and significant interest in the development of small-molecule electrophilic probes and drugs. Analysis of thiol reactivity under various conditions utilizing 19F NMR has allowed us to optimize thiol addition reactions to previously synthesized exo/endo vinyl sultams, vinyl β-keto sultam analogs of tetramic acids, as well as sulfonamides and known drugs, including antitumor and antifungal agents. Efficient catalytic reactions governed by arginine and histidine to increase thiol- and cysteine-Michael addition reactivity will also be discussed. For future work, all compounds will be screened to reveal biological activity and to provide a better understanding of biochemistry in health and disease.

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E8. Modeling Permeation of Aromatic Dipeptides across Lipid Bilayers

Brent L. Lee1, Krzysztof Kuczera2

1Department of Chemistry, University of Kansas, Lawrence, KS, USA;
2Departments of Chemistry and Molecular Biosciences, University of Kansas, Lawrence, KS, USA

The diffusion of small peptides across a lipid membrane was determined, characterized, and compared to experimental values by using molecular dynamics simulations.  Blocked phenylalanine, tyrosine, tryptophan, and wh5a were studied.  The lipid membrane was constructed by using 50 DOPC lipids surrounded by around 3000 water molecules.  The simulations were setup by using CHARMM and then run with the GROMACS program.  Potentials of mean force indicated preferential binding of the peptides to the lipid interface and large free energy barriers at the membrane center.  Peptide translational diffusion rates show small changes between solution, interface and membrane interior.  In contrast, the sidechain rotational correlation times show extremely large changes with membrane insertion, with values becoming 100 time greater in the head-group region and 10 times greater in the tail region, compared to solution. The peptides’ conformational freedom becomes systematically more restricted as they enter the membrane, sampling α, β and C7eq conformers in solution and only C7eq in the center.  Differences between system size, 40 or 50 lipids, and lipid type, DOPC or POPC, were also examined and indicate little or no change of properties.

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E9. Development of Mechanically Tunable Gelatin-Alginate Hydrogels to Promote Stem Cell Osteogenic Differentiation

Settimio Pacelli1, Helena K. Salt2, Cecilia Kurlbaum1, Madeline Fang1and Arghya Paul1

1Department of Chemical and Petroleum Engineering, Bioengineering Graduate Program, University of Kansas, Lawrence, KS, USA
2Department of Mechanical Engineering, School of Engineering, Undergraduate Program, University of Kansas, Lawrence, KS, USA

Surface properties of biocompatible scaffolds can be easily modified using bioactive molecules to control stem cell differentiation into the osteogenic lineage. An innovative solution is the use of a polydopamine coating as linking strategy to bind signal molecules capable of directing stem cell behavior. In this study, a gelatin-alginate interpenetrated network (IPN) was fabricated using different amounts of N-hydroxysuccinimide (NHS), and ethyl carbodiimide (EDC) as crosslinkers to produce tunable scaffolds for human adipose mesenchymal stem cell (hASCs) differentiation into the bone lineage. High and low crosslinked hydrogels were obtained and characterized by their ability to swell, degrade and resist different mechanical stimuli. In addition, the surface of the scaffolds was coated with polydopamine as a bonding layer for bioactive molecules to promote hASCs differentiation. Results showed a higher absorption of dexamethasone could be achieved using a polydopamine coating respect to the uncoated hydrogels. As expected, hASCs cultured on the scaffold coated with polydopamine and dexamethasone showed significantly higher differentiation markers such as alkaline phosphatase and higher calcium deposition when compared to the control groups. Due to these promising findings, this scaffold could be potentially used as an osteoinductive coating for biomedical implants to enhance bone regeneration.

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E10. The Synthetic Chemical Biology Core (SCB): A Resource for Research in Chemical Biology

Chamani T. Perera1, Digamber Rane1, Benjamin Neuenswander1, Blake R. Peterson1,2, Thomas E. Prisinzano1,2

1Department of Medicinal Chemistry, University of Kansas, Lawrence, KS, USA;
2School of Pharmacy, University of Kansas, Lawrence, KS, USA

The Synthetic Chemical Biology Core strives to provide comprehensive synthetic chemistry capabilities to investigators under one roof. The synthetic expertise of the core includes, but is not limited to, novel and commercially unavailable small molecules, fluorescent molecules and peptides. The core assists in identifying hits for medicinal chemistry optimization in infectious disease targets and provides synthesis capabilities for structure activity studies of said hits. The core staff will work with investigators to design and synthesis novel molecular probes to facilitate their research. SCB core additionally provides access to the model organism Danio rerio (zebrafish), and allows investigators to image embryonic and adult zebrafish treated with molecular probes for phenotypic drug discovery and other projects. SCB core encompasses the Purification and Analysis Laboratory (PAL) that provides purification, analysis and quality control of compounds via HPLC-MS. The core utilizes automated mass directed fractionation for purification in both reversed and normal phases (including chiral separations), and also provides relative purity analysis by UPLC coupled to a high-resolution mass spectrometer for structure confirmation.

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E11. Investigating “Stuffed” Domains of NRPS Assembly Lines: PchF and PchE of Pyochelin Biosynthesis

Trey A. Ronnebaum1, Geoff P. Horsman2, Squire J. Booker3,4, Thomas E. Prizinzano1,5, Audrey L. Lamb1,6

1Deparment of Chemistry, University of Kansas, Lawrence, KS, USA; 2Department of Chemistry and Biochemistry, Wilfrid Laurier University, Waterloo, ON, Canada; 3Department of Chemistry, Department of Biochemistry and Molecular Biology, Pennsylvania State University, State College, PA, USA; 4Howard Hughes Medical Institute, Chevy Chase, MD, USA; 5Department of Medicinal Chemistry, University of Kansas, Lawrence, KS, USA; 6Department of Molecular Biosciences, University of Kansas, Lawrence, KS, USA

Nonribosomal peptide synthetases (NRPSs) are one approach used by microbes to generate bioactive peptides. These bioactive peptides are not only used as secondary metabolites (toxins, pigments, siderophores – iron scavenging molecules), but have found their way into the clinic as antibiotics, anticancer drugs, and immunosuppressants. To elicit their unique bioactivity, these peptides must be tailored. Natural product chemists, metabolic engineers, and researchers in biochemistry and biotechnology work to exploit the biosynthesis of these secondary metabolites in order to generate new compounds for clinical use. The long term goal of this project is to understand the structure-function relationships of epimerases and methyltransferases that are incorporated into these assembly lines. Structural biology and mechanistic enzymology can provide novel insight and assist natural product investigations, protein engineering projects, antimicrobial development, and other therapeutic design.  Currently there is no adenylation-tailoring “stuffed” didomain NRPS structures, and limited biochemical characterization exists. This project concentrates on the adenylation-epimerase didomain of PchE and the adenylation-methyltransferase didomain of PchF in the biosynthetic pathway of the siderophore, pyochelin. Initial work includes substrate analogue synthesis and the establishment of adenylation, epimerization, and methyltransferase assays.

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