Abstract embargoed

Jordhan D. Booth and Karina Kapusta

Department of Chemistry and Physics, Tougaloo College, Tougaloo, MS

Introduction/Background. Bioimaging is pivotal in medical diagnosis, treatment, and early detection of diseases, including cancer. Fluorescent organic near-infrared (NIR) dyes are commonly used in bioimaging, valued for their brightness, deep tissue penetration, reduced autofluorescence, and relatively low toxicity. One such promising compound, indolizine squaraine dye (SO3SQ), exhibited high fluorescence in fetal bovine serum but disappointingly diminished effectiveness in whole blood. Intriguingly, the addition of certain ionic liquids improved its effectiveness significantly.

Hypothesis/Goal of Study. Optimizing SO3SQ for biomedical applications seemed a complex task, demanding considerable time and resources if approached solely through experimental testing. To bypass these challenges, we applied comprehensive computational chemistry techniques, such as molecular mechanics and dynamics, to unearth the mechanisms controlling SO3SQ's behavior in a biological environment.

Methods and Results. The investigation began with the use of molecular docking and mechanics to build complexes of the dye with abundant plasma proteins, such as albumin. Molecular dynamics simulations were then utilized to assess their stability, revealing a distinctly higher affinity of the dye to human serum albumin compared to bovine serum. This prediction, later confirmed experimentally, set the stage for more detailed exploration. Further, we investigated the influence of ionic liquids on the dye-albumin complex's strength and morphology. These computational techniques shed light on how fluorescence improved in the presence of certain ionic liquids.

Discussion/Conclusions. As a result of this broad investigation not only computational tools helped to explain the nature of the fluorescence activity of dye/albumin/ionic liquid complexes in various alternations, but also some of the obtained information became a driving force for more extensive research related to the biomedical application of studied dye, such as albumin, alpha-tubulin, and blood detection. Thus, this work is presented as an example of an in silico-driven comprehensive investigation of macromolecular complexes for biomedical research.

Citation/Acknowledgements. This work was supported by the Mississippi INBRE, funded by an Institutional Development Award (IDeA) from the National Institute of General Medical Sciences of the National Institutes of Health under grant number P20GM103476. The content is solely the responsibility of the authors and does not necessarily represent the official views of the National Institutes of General Medical Sciences or the National Institutes of Health. Additionally, the author wants to thank the National Science Foundation and MS EPSCOR: Center for Emergent Molecular Optoelectronics (CEMOs) [Award#: OIA – 1757220] for collaboration.

Irosha N. Nawarathne1, Lola Beeser1, Braden Glenn1, Rachel Tyler1, Nikkolette Perkins1, Isabella Beasley1, Daniel Armstrong1, Wyatt Treadway1, Clara Nikkel1, Jake Smith1, Amir Mortazavi2, Samir Jenkins2, Ruud Dings2, Marissa Fullerton3, and Daniel Voth3

1Department of Chemistry, Lyon College, Batesville, AR, 2Department of Microbiology and Immunology, University of Arkansas for Medical Sciences, Little Rock, AR, 3Department of Radiation Oncology, Winthrop P. Rockefeller Cancer Institute, University of Arkansas for Medical Sciences, Little Rock, AR

Introduction/Background. Naturally occurring and synthetically derived naphthoquinones and their derivatives possess a wide range of biological activities such as antimicrobial, anticancer, anti-inflammatory, antiproliferative properties. Naphthoquinone scaffold is present in some of the widely used antibacterials and antineoplastics such as doxorubicin, rifamycins including rifampin, streptonigrin, napabucasin, and mytomycin. While the mode of antibacterial action of naphthoquinone scaffold varies from one set of derivatives to the other, the scaffold is known to selectively target cancer cells acting as ROS-inducing agents and inhibit enzymes such as STAT3, DNA topoisomerase I and II, and AKT kinases.

Hypothesis/Goal of Study. Although the natural product-inspired naphthoquinone scaffold is an evolutionary selected and biologically pre-validated starting point, the ROS generation with naphthoquinones represents a challenge in developing clinically relevant agents; therefore, any modification to the naphthoquinone core has to be designed to modulate redox properties, by adding highly electronegative atoms such as nitrogen and oxygen, to guarantee new lead structures. Furthermore, triazoles are known to mimic different functional groups, resulting optimal bioisosteres for the synthesis of new biologically active molecules.

Methods and Results. Therefore, we conducted a methodical investigation of N and O incorporated simple aminonaphthoquinones by sequentially coupling the ubiquitous click chemistry with Michael addition reaction of naphthoquinones, following a comprehensive evaluation of anti-cancer, toxicity, and redox properties of the derivatives to contribute to the advancement of cancer therapeutics. Moreover, the sequential coupling of complex rifamycin chemistry with simple click chemistry has shed some light to the advancement of rifamycin derivatives with azido, alkyne, and triazole functionalities at naphthalenic C8. The novel rifamycin derivatives demonstrate promising antimicrobial properties against prevalent rifamycin-resistant tuberculosis (RR-TB) and other deadly diseases caused by both Gram+ and Gram- pathogens with persistently growing multidrug-resistance while contributing to medicinal chemistry in a synthetic point of view that has been neglected despite the undeniable advantages of naphthoquinone derivatives as pharmaceuticals.

Discussion/Conclusions. In this presentation, how our innovative approach led into novel naphthoquinone derivatives with promising biological activities will be discussed.

Citation/Acknowledgements. This project was supported by the Arkansas INBRE program, with a grant from the National Institute of General Medical Sciences, (NIGMS), P20 GM103429 from the National Institutes of Health.

Gerard Rowe and Kenneth Roberts
Department of Chemistry and Physics, University of South Carolina Aiken, Aiken, SC

Introduction/Background. The non-heme iron enzyme 2,4’-dihydroxyacetophenone dioxygenase (DAD) catalyzes the conversion of its substrate DHA into the products 4’-hydroxybenzoic acid and formic acid.  Unlike other well-characterized ring-cleaving catechol dioxygenases, this enzyme cleaves a non-aromatic carbon-carbon bond external to the aromatic ring. 

Hypothesis/Goal of Study. In this study, MD and QM/MM studies were carried out on the DHA-loaded active site to determine key amino acid residues and to shed light on the mechanism of action. In particular, we consider the possibility that Tyr94 might act as a ligand for the iron cofactor and plays an important role in deprotonation and subsequent rebound during the reaction as well as the role of a potential water channel on the backside of the iron-binding site.

Methods and Results. The protein structue was solvated with VMD and equilibrated with the CHARMM force field in NAMD.  QM/MM calculations were carried out using Orca at the B97-3C level of theory.

Discussion/Conclusions. The results of the QM/MM calculations suggest that DHA acts as a redox non-innocent ligand prior to O2 binding to the iron center and that the resting state of the enzyme includes an iron-bound hydroxide and a deprotonated tyrosine ligand, both of which are involved in proton transfer events during the catalytic cycle.  During the course of these calculations, several amino acids were identified as candidates for site-directed mutagenesis experiments.

Citation/Acknowledgements. The authors wish to thank University of South Carolina Research Computing, USC Aiken's Summer Scholar Institute, USC Magellan Undergraduate Research Program, and SC INBRE for funding and resources.