Doctor of Philosophy (Ph.D.), Bowling Green State University, 2023, Biological Sciences

Globally abused phenethylamine drugs can cause “phenethylamine-induced-hyperthermia” (PIH), a serious health concern associated to sequela of complications including rhabdomyolysis, coagulopathy, kidney failure and death. In general, PIH is known to occur via activation of host sympathetic nervous system (SNS) leading to increased release of norepinephrine (NE) post phenethylamine exposure. NE then acts as signal to activate pathways which generate heat in various thermogenic tissues or prevent heat loss through vasoconstriction. Besides NE, recent research from our laboratory pointed towards potential involvement of gut bacteria in PIH, shown by altered PIH following gut microbial modulations such as via antibiotic treatment or fecal microbial transplantation (FMT). Here, we aimed to investigate the functional role of gut microbiome and bile acids as signal in PIH. We applied targeted LC-MS/MS, first time to our knowledge, to quantify serum concentrations of three important bacteria metabolized/associated bile acid species (BAs): cholic acid (CA), chenodeoxycholic acid (CDCA) and deoxycholic acid (DCA) prior to and post (hyperthermic dose of) 3,4-methylenedioxymethamphetamine (MDMA) (a phenethylamine) treatment. Male Sprague-Dawley rats were provided either sterile or antibiotic-water for five days prior to MDMA (20 mg/kg, sc)/saline administration. On day six BAs and body temperatures were measured at 0 minute before and 30 minutes and 60 minutes post MDMA/Saline challenge. We found all three focal BAs in serum to deplete significantly in MDMA treated rats compared to saline group at 60-minute, exactly the same time point when peak rise in core body temperature occurred, indicating contribution of these three BAs in PIH. While prior antibiotic cocktail (vancomycin, bacitracin and neomycin) treated groups had greatly reduced serum level of all three BAs and reversal of hyperthermia after MDMA. We also inferred metagenomic functions applying PICRUSt2 on 16S rRNA ge (open full item for complete abstract)

Committee: Christopher Ward Ph.D. (Committee Chair); Benjamin Ward Ph.D. (Other); Raymond Larsen Ph.D. (Committee Member); Vipaporn Phuntumart Ph.D. (Committee Member); Jon Sprague Ph. D. (Committee Member) Subjects: Biology

Doctor of Philosophy (Ph.D.), Bowling Green State University, 2024, Biological Sciences

Host responses to synthetic phenethylamines have been linked with alterations in the gut microbial profile. A novel gut commensal, Proteus mirabilis (P. mirabilis) strain Roy1, was discovered to be enriched in the cecal contents of a rat treated with the synthetic phenethylamine, 3,4-methylenedioxymethamphetamine (MDMA). The genomic DNA of this bacterium was subjected to Illumina sequencing, revealing a draft genome sequence of approximately 3.9 Mbps and comprises 3,602 coding sequences (CDSs). To identify functionally significant genes, a comparative genome analysis was conducted between P. mirabilis Roy1 and thirty different P. mirabilis genomes available in the NCBI database. The pan-genome comparisons indicated that P.mirabilis Roy1 shared approximately 75% of its core genome with the P. mirabilis referencestrains. Furthermore, the comparative analysis revealed unique genes contributing to its successful colonization and proliferation in the gastro-intestinal tract (GIT). Rats subjected to MDMA treatment exhibited diarrheal symptoms. The strain-specific genes detected in P. mirabilis Roy1 might have aided in bypassing the intestinal emptying process. The unique gene repertoire includes genes involved in inter/intraspecies competition, intestinal adherence, and pathogenicity. Interestingly, these gene sets closely resembled genes found in various Escherichia coli (E. coli) strains, suggesting that they may have been acquired from the rat's gut contents. The direct contributions of P. mirabilis Roy1 in driving phenethylamine-induced responses require further experimental validations. Phage therapy has emerged as a treatment option for bacterial infections. However, its efficacy against drug-induced responses has not been addressed. Therefore, phages specific to P. mirabilis strain Roy1 were isolated. For orally administered phages to be therapeutically effective, they need to endure the passage through gastric fluid, including surviving at low pH levels. Thus, aci (open full item for complete abstract)

Committee: Vipaporn Phuntumart Ph.D. (Committee Co-Chair); Raymond Larsen Ph.D. (Committee Co-Chair); Jon Sprague Ph.D. (Committee Member); Scott Rogers Ph.D. (Committee Member); Zhaohui Xu Ph.D. (Committee Member); Jeremy Oehrtman Ph.D. (Other) Subjects: Bioinformatics; Microbiology; Molecular Biology; Pharmacology

Doctor of Philosophy, University of Toledo, 2020, Medicinal Chemistry

Alzheimer's disease (AD) is a progressive, neurodegenerative disease resulting in cognitive decline, dementia, and eventually death. This work investigates furoxans (1,2,5 oxadiazole N-oxides) for their utility in the treatment of AD. Furoxans are thiol dependent nitric oxide (NO) mimetics, capable of releasing NO in a cellular environment. This work focuses on attenuated furoxans, designed to release NO in a slow, controlled manner to engage the NO/sGC pathway, resulting in in the phosphorylation of CREB, increasing the production of pro-survival gene products such as BDNF. IS-1-41 was identified as a hit compound, possessing promising biological activity such as: neuroprotection, procognitive effects, and a long systemic half-life. Efforts to explore attenuated furoxans resulted in a library of more than 60 novel attenuated furoxans. Several novel attenuated furoxans were found to be neuroprotective; however, when incubated with cysteine, none were found to breakdown to release NO despite evidence showing neuroprotection was NO/sGC dependent. We investigated the bioactivation of attenuated furoxans via selenocysteine and found that, while attenuated furoxans breakdown in the presence of selenocysteine, the resulting breakdown product has not released NO. This supports the hypothesis that attenuated furoxans interact with unknown protein target(s) to release NO, providing the observed neuroprotection, which can be further explored via photolytic target ID studies. After extensive pre-clinical development, four lead candidates, IS-1-41, AH-1-91, AH-2-36, and AH-2-87 were identified and are being investigated for their utility in the treatment of AD. Tangentially, work was undertaken to establish a platform to accomplish in vivo photoaffinity target ID studies to explore the mechanism of action of β-phenethylamines with a non-biased approach. Casper zebrafish were employed as the in vivo model, as casper zebrafish lack melanin and therefore cannot block t (open full item for complete abstract)

Committee: Isaac Schiefer (Committee Chair); William Messer (Committee Member); Zahoor Shah (Committee Member); Erin Prestwich (Committee Member); Wei Li (Committee Member) Subjects: Analytical Chemistry; Chemistry; Neurosciences; Pharmaceuticals; Pharmacy Sciences

PhD, University of Cincinnati, 2017, Arts and Sciences: Chemistry

A considerable number of dietary supplements suspected of containing phosphodiesterase-5 (PDE-5) inhibitors and substituted phenethylamines have been analyzed by the U.S. Food and Drug Administration. Often these samples are found to contain the active pharmaceutical ingredients (API) such as sildenafil or phentermine, and in many cases, products contain multiple PDE-5 inhibitors or substituted phenethylamines. In an analytical setting, it is important to confirm the presence of any API with two or more independent methods, and this requirement can often put undo strain on a laboratory. The development and use of methods that inherently contain two unique identification techniques is preferred, and the creation and validation of three of those methods is outlined here. First, direct deposit Fourier transform infrared detection and mass spectrometric detection (GC/FT-IR/MS) is used to identify PDE-5 inhibitors. Generally, GC/MS is not generally used for this category of drugs due to low volatility; PDE-5 inhibitors often co-elute and can produce non-specific electron ionization fragmentation patterns. In contrast, FT-IR has been proven to be more selective for identifying PDE-5 inhibitors, but is generally not as sensitive as spectrometric techniques. However, it has been shown that each technique can compensate for the other, which allows a wider range of usability. Using this combined technique can save time and resources while still delivering a high level of certainty in identification by providing results from two scientifically uncorrelated techniques. Multiple reference standards were utilized for method validation, including determination of linearity, dynamic range, and limit of detection. Second, a single HPLC-UV method has been developed for the determination of PDE-5 inhibitors and related analogs in pharmaceutical dosage forms and dietary supplement products. Using this protocol, 14 PDE-5 inhibitor compounds can be separated and determined in a single an (open full item for complete abstract)

Committee: Peng Zhang Ph.D. (Committee Chair); Anna Gudmundsdottir Ph.D. (Committee Member); Laura Sagle Ph.D. (Committee Member) Subjects: Chemistry