Selective inhibition of the bromodomain and extra-terminal domain (BET) family of proteins is a promising method to regulate gene expression and disrupt pathways associated with diseases, including inflammation and certain cancers. Inhibitors that selectively target the D2 bromodomain within BET proteins have been shown to attenuate undesirable side effects in clinical settings compared to non-selective inhibitors. The need for D2 bromodomain selective inhibitors is an ongoing challenge in small-molecule drug discovery. Current non-selective inhibitors that target both the D1 and D2 bromodomains exhibit dose-limiting toxicity, minimizing their therapeutic potential. Recently, the Pomerantz lab developed inhibitors based on the 1,4-thiazepane scaffold identified from a small molecule screen of 3D-enriched fragments to target the D2 bromodomain in BET proteins. 1,4-thiazepanes exhibit high 3D character, which allows for potential conformational diversity and favorable interactions with the D2 bromodomain as it mimics the acetylated lysine on the histone tail. In this study, we synthesized various 1,4-thiazepanes, with different substituents on the benzene ring, through key reactions, including cyclization with α,β-unsaturated esters and 1,2-aminothiols to yield 1,4-thiazepanones, and subsequent reduction and acylation to yield 1,4-thiazepanes, which were characterized by 1H NMR spectroscopy. Competitive AlphaScreen assay was employed to evaluate the selectivity and binding affinity for D2 bromodomains. By varying substituents on the benzene rings, we can explore the electronic properties of the ring system to further optimize selectivity and affinity and introduce new vectors for the 1,4-thiazepanes to target D2 bromodomains.

Globally, bacterial infections are ranked as one of the leading causes of death, with lower respiratory infections ranking fourth among all causes of mortality. In response, clinical settings have implemented measures for the prevention and treatment of infections. Antibiotic treatments have traditionally targeted several proteins which include beta-lactamases, transpeptidases, ribosomal subunits, dihydropteroate and dihydrofolate synthetases, DNA supercoiling enzymes, penicillin-binding proteins, and RNA polymerase. However, bacteria rapidly develop resistance to these antibiotics. The ESKAPE pathogens (Enterococcus faecium, Staphylococcus aureus, Klebsiella pneumoniae, Acinetobacter baumannii, Pseudomonas aeruginosa, and Enterobacter spp.) are highly drug-resistant and account for more than 900,000 deaths worldwide each year. Thus, there is a need for the development of antibiotics that do not target canonical antibiotic targets. The GroEL/GroES chaperone is a protein found in all eubacteria and is necessary for cell viability under all conditions tested and, in every species tested. While current lead compounds inhibit cell viability in gram-positive ESKAPE bacteria, inhibition in gram-negative ESKAPE bacteria continues to be a challenge, due to the outer lipopolysaccharide layer and efflux pumps. Here we focus on the development of compounds that inhibit GroEL/GroES based on Entryway rules for gram-negative ESKAPE bacteria. To characterize the inhibition of GroEL/GroES, we utilize the protein substrate and an ATPase assay. By applying the Entryway rules to inhibitors of GroEL/GroES, greater inhibition and decreased cell viability of gram-negative ESKAPE bacteria will be sought. Compounds meeting both the criteria of accumulation and inhibition will be further validated as antibiotic candidates in clinical applications.

Tuberculosis is an airborne infectious disease caused by Mycobacterium tuberculosis which severely affects the lungs. It can be caught by breathing in the air that an infected person has contaminated through coughing, sneezing, or speaking. Mycobacterium tuberculosis is a pathogen that is estimated to latently infect about a quarter of the world’s population (Chin et al.,2023). Carbapenems can be described as parenteral bactericidal beta-lactam antibiotics that have an extremely broad spectrum (Werth, 2023). The goal of this summer’s research is to develop a carbapenem antibiotic that would be responsive against multi-drug resistant mycobacteria which is responsible for multi-drug resistant tuberculosis. The research focus is designing and modifying antibiotics that would be selective against 31 specific bacterial strains using chemical synthesis. Methodology: 1. Design and modify the antibiotics using chemical synthesis. 2. Purify and analyze them with Nuclear Magnetic Resonance spectroscopy to ensure their structure and purity. 3. Send the chemicals produced to Micromyx which will analyze its selectivity against 31 bacterial strains. 4. If the compound is found to be very active, it will be sent to microbiologists to test its structural biology against transpeptidases which are enzymes that catalyze the transfer of an amino acid residue or a peptide residue from one amino compound to another.