←Within this dissertation the topic of virtual screening is discussed with regard to three different cancer targets and also a brief introduction of the tools used in virtual screening. In Chapter 1, the reader will be introduced to virtual screening and the programs that are used in virtual screening. In Chapter 2, the first of three projects are discussed. This project consists of the work that was done to find inhibitors of the P53 binding domain of MDMX. In this project the mobility of residues within the binding site of MDMX are discussed and the ways in which we attempted to model how drugs would bind two adjacent pockets within MDMX. In Chapter 3, the virtual screening and modeling work done for RING domain of MDM2 and MDMX is discussed. This work was done in conjunction with Moffitt Cancer Center in order to solve the 60 year old mystery of the mechanism of how thalidomide and possibly its analog lenalidomide caused children to be born limbless. Current thinking is that Cereblon through an unknown teratogenic mechanism activates an increase in FGF8. We suggest a mechanism that may happen in parallel that involves stabilization of MDM2 and the reduction of P63 levels. Chapter 4, the work that was done against the BH3 binding domain of MCL-1 is discussed in conjunction with collaboration with the Manetsch lab. In order to complete this screening the validation of IC50 values and then attempt to modify those products based upon the structure of MCL-1. Chapter 5 discusses the work done to find inhibitors of deoxycytidine kinase. All of these chapters taken together provide a brief overview of the computational work done produce inhibitors of Protein-Protein Interaction against three major cancer targets.

Solid lipid nanoparticles (SLNPs) and lipid-drug conjugates (LDCs) are two promising lipid nanoparticle (LNP) based drug delivery systems; this thesis explores new synthetic lipids that may circumvent the limitations of currently available components for LNPs with particular focus on the stability of LNP formulations. Neutral polyethylene glycol lipids (PEG-lipids) have been designed, synthesized, and characterized with ESI-MS, for stabilizing SLNPs containing dsDNA oligomer. 1st and 2nd generation PEG-lipids investigated the effects of serinol and iminodiacetic acid backbone structures, respectively, and aliphatic chain sequences within the lipid anchors on the stability of SLNPs. Assays were developed to analyze LNP stability in both PBS buffer and PBS buffer with 10 % serum at different incubation temperatures. The results indicate that the hydrocarbon branching sequence offer additional SLNP stability over straight chain isomers. LDC monomers were designed and synthesized to allow for the formulation of LDC nanocarriers for the thiopurine drugs. These hydrophobic LDC monomers were made by linking the polar thiopurine drug to a synthetic lipid. These synthetic lipids investigated branched and straight chain derivatives – the branched isomers once again demonstrated advantages in the stability of the LDCs. / Graduate

Computer-aided drug design is an exceptionally useful tool for screening a large number of potential drug molecules to evaluate their therapeutic potential. This technique is both effective and economical. Approximately 120 imidazole-containing molecules were computationally designed and evaluated using gas-phase and solution-phase simulations to assess their propensity for acting as a chelating agent with twenty-six biologically relevant cations. Of particular interest was their ability to chelate Zn2+ and Ca2+, which play a key role in the degradation of neurons following an ischemic stroke. The ultimate goal was to design a small molecule that could be administered before a medical procedure that featured stroke as a possible side effect. In the event that a stroke occurred, the destruction of neurons caused by release of excess Ca2+ and Zn2+ would be diminished and the patient would maintain motor and cognitive function. Promising in silico results were obtained.

Caseinolytic protease (ClpP) is a cylindrical protease forming the core of protein degradation machinery in eubacteria. ClpP is tightly regulated and is non-functional without a member of the Clp-ATPases. A new class of antibiotics, termed ADEPs, bind to ClpP and allow for activation without the Clp-ATPases; leading to cell death. A more efficient synthetic route to the ADEPs utilizing solid-phase peptide synthesis was investigated. A linear peptide was synthesized, however attempts to close the depsipeptidic macrocycle via macrolactonization failed. Further attempts of assembling a branched depsipeptide for ring closure via a macrolactamization resulted in products that were not stable to cleavage conditions. A group of molecules termed Activators of Self-Compartmentalizing Proteases (ACP) were identified through a screen for activity towards ClpP. Compound ACP1 was synthesized along with twelve analogs and their activity towards ClpP evaluated. The project resulted in a compound with a higher activity than its natural product counterpart.

One of the largest obstacles in drug development is to overcome solubility and bioavailability problems. Preformulation strategies such as nanoparticle formation are often employed but sometimes create new issues and are limited in their effectiveness and applications. Since the majority of drugs are marketed and sold as solid forms, drug delivery systems are not always desirable. This is where solid-state chemistry becomes important. Traditional solid-state chemistry approaches are often successful but are sometimes too restrictive and cannot be applied to certain compounds. Cocrystals have emerged as an alternative solid-state technique that can be applied to a broad range of compounds. However, the technology is still very new and its effectiveness in certain conditions had previously not been evaluated. The studies detailed herein investigated the ability of two different technology platforms for overcoming drug design challenges for two promising bioflavonoids: EGCg and quercetin. Studies have shown that EGCg might be useful for the treatment of Alzheimer's disease and other neurodegenerative diseases. Quercetin is being investigated for numerous bioactivities and is currently being marketed as an energy dietary supplement. Both of these bioflavonoids exhibit poor bioavailability and water solubilities that are at opposite ends of the spectrum. In the chapters to follow, nanoparticle technology was applied to EGCg and evaluated in cell models of AΒ production, a hallmark of Alzheimer's disease. Bioavailability improvements were also evaluated in rats. Additionally, new forms of both flavonoids were created using cocrystallization. These new cocrystals were characterized using powder and single crystal x-ray diffraction, differential scanning calorimetry, and thermogravimetric analysis. Solubility and bioavailability changes were also evaluated. These data have strong implications in drug development since they elucidated the strengths and weaknesses of two major technologies in compounds with different design challenges.

Despite over a century of advancement, tuberculosis remains a grave threat to world health. In particular, third world countries continue to struggle with the crushing weight of the disease. Furthermore, the emergence of drug resistance in TB strains poses a significant threat to the first world where incidence and mortality is low. The dwindling efficacy of current drug regimens necessitates research into new small molecules capable of arresting the growth and spread of TB. The capuramycin family of nucleoside antibiotics shows strong potential to become part of this new generation of anti-TB small molecules. Indeed, their ability to inhibit Translocase I, a key enzyme in the biosynthesis of bacterial cell walls, makes them exciting targets for medicinal chemistry efforts. The synthesis of the family focused on dividing the molecules into three congruent, synthetically separate parts: the variable amide linked tail, the hexauronic acid linker, and the uridine "head". Construction of the ubiquitous core structure comprised of the hexauronic acid and uridine would allow rapid diversification while the variable tail would allow SAR studies and development of novel new members of the family.

Advances in anti-cancer drug development have vastly improved cancer treatment strategies over the past few decades. Chemotherapeutic agents are now being replaced with targeted therapies that have much greater potency and far fewer unpleasant side effects. At the center of this, cell signaling pathways have been targeted as they moderate gene expression, control proliferation and are often dysregulated in cancer. The signal transducer and activator of transcription (STAT) proteins represent a family of cytoplasmic transcription factors that regulate a pleiotropic range of biological processes in response to extracellular signals. Of the seven mammalian members described to date, Stat3 has received particular attention, as it regulates the expression of genes involved in a variety of malignant processes including proliferation, survival, migration and drug resistance. Aberrant Stat3 activation has been observed in a number of human cancers, and its inhibition has shown promising anti-tumour activity in cancer cells with elevated Stat3 activity. Thus, Stat3 has emerged as a promising target for the development of cancer therapeutics. While Stat3 signaling can be inhibited by targeting upstream regulators of Stat3 activation (such as Janus kinase 2), direct inhibition of Stat3 protein may offer improved response, larger therapeutic windows for treatment and fewer side effects. The work presented within this thesis is focused on optimizing known Stat3 inhibitor S3I-201, a small molecule Stat3 SH2 domain binder that was discovered in 2007. We have performed an extensive structure activity relationship study that has produced some of the most potent Stat3 inhibitors in the scientific literature. These compounds showed high-affinity binding to Stat3’s SH2 domain, inhibited intracellular Stat3 phosphorylation and selectively induced apoptosis in a number of cancer cell lines. Lead agents further inhibited tumour growth in xenograft models of human malignancies and had favourable pharmacokinetic and toxicity profiles.

Advances in anti-cancer drug development have vastly improved cancer treatment strategies over the past few decades. Chemotherapeutic agents are now being replaced with targeted therapies that have much greater potency and far fewer unpleasant side effects. At the center of this, cell signaling pathways have been targeted as they moderate gene expression, control proliferation and are often dysregulated in cancer. The signal transducer and activator of transcription (STAT) proteins represent a family of cytoplasmic transcription factors that regulate a pleiotropic range of biological processes in response to extracellular signals. Of the seven mammalian members described to date, Stat3 has received particular attention, as it regulates the expression of genes involved in a variety of malignant processes including proliferation, survival, migration and drug resistance. Aberrant Stat3 activation has been observed in a number of human cancers, and its inhibition has shown promising anti-tumour activity in cancer cells with elevated Stat3 activity. Thus, Stat3 has emerged as a promising target for the development of cancer therapeutics. While Stat3 signaling can be inhibited by targeting upstream regulators of Stat3 activation (such as Janus kinase 2), direct inhibition of Stat3 protein may offer improved response, larger therapeutic windows for treatment and fewer side effects. The work presented within this thesis is focused on optimizing known Stat3 inhibitor S3I-201, a small molecule Stat3 SH2 domain binder that was discovered in 2007. We have performed an extensive structure activity relationship study that has produced some of the most potent Stat3 inhibitors in the scientific literature. These compounds showed high-affinity binding to Stat3’s SH2 domain, inhibited intracellular Stat3 phosphorylation and selectively induced apoptosis in a number of cancer cell lines. Lead agents further inhibited tumour growth in xenograft models of human malignancies and had favourable pharmacokinetic and toxicity profiles.

Aromatic stacked salt bridges are increasingly observed to play an important role in biology, suggesting that the two separate weak interactions cooperate with each other to mediate molecular recognition in a biological solution. In this thesis an in depth study was carried out in attempt to find the contribution of the guanidinium-carboxylate-aromatic triad in biological systems. Two different small molecule systems are used to carry out the study. From the results of the two chapters I proposed here that stacking aromatic ring enhances the salt bridge through desolvation effect. This hypothesis was also tested in a protein-protein interaction (Grb2 SH3 domain/SOS interaction). The most ideal peptide inhibitor cannot be obtained due to the synthetic difficulties. Limited result showed that increasing the hydrophobic area of the hot spot in this protein-protein interaction enhances the interaction. In researching the guanidinium-carboxylate-aromatic triad, we were inspired to study the pre-organization effect of 1,3,5-triethyl-2,4,6-trisubstituted benzene template. A computational and literature study done in this thesis showed that the installation of ethyl or methyl groups at 1,3,5 positions leads to consistent increases in binding affinity relative to unsubstituted hosts, but the amount of increase is non-trivial and varies with different substitutes. The installation of ethyl or methyl groups at 1,3,5 positions leads to consistent but relatively small increases in binding affinity relative to unsubstituted hosts. / Graduate

One of the objectives of the U.S. Food and Drug Administration is to protect the public health through post-marketing drug safety surveillance, also known as Pharmacovigilance. An inexpensive and efficient method to inspect post-marketing drug safety is to use data mining algorithms on electronic health records to discover associations between drugs and adverse events. The purpose of this study is two-fold. First, we review the methods and algorithms proposed in the literature for identifying association drug interactions to an adverse event and discuss their advantages and drawbacks. Second, we attempt to adapt some novel methods that have been used in comparable problems such as the genome-wide association studies and the market-basket problems. Most of the common methods in the drug-adverse event problem have univariate structure and thus are vulnerable to give false positive when certain drugs are usually co-prescribed. Therefore, we will study applicability of multivariate methods in the literature such as Logistic Regression and Regression-adjusted Gamma-Poisson Shrinkage Model for the association studies. We also adopted Random Forest and Monte Carlo Logic Regression from the genome-wide association study to our problem because of their ability to detect inherent interactions. We have built a computer program for the Regression-adjusted Gamma Poisson Shrinkage model, which was proposed by DuMouchel in 2013 but has not been made available in any public software package. A comparison study between popular methods and the proposed new methods is presented in this study.