Natural products and small molecules play a major role in drug development. However, using natural products as a source of medicine comes with many challenges, such as lack of natural abundance and difficulty in isolation. Consequently, synthetic organic chemistry is a solution in order to access these compounds in usable quantities. However, synthetic chemisty comes with its own challenges such as efficiency, chemoselectivity, stereoselectivity and enantioselectivity. Therefore, synthetic tools that addresses these challenges are required solve these limitations. This thesis discusses new methodologies using strained carbocycles (cyclopropanes and cyclopropenes) as the reactive subunit for the construction of different carbocyclic and heterocyclic compounds. The homo-Nazarov cyclization of alkenyl and heteroaryl cyclopropyl ketones was used in order to construct cyclohexenones, cyclohexenols, heteroaryl ring-fused cyclohexenones, dihydrofurans, furans and furanones in a mild and efficient manner. Benzofused heteroaromatic compounds were achieved via the Lewis acid-catalyzed cycloisomerization of cyclopropene-3,3-dicarbonyls and furan-3-carboxylates. These heteroaromatic compounds can be applied to medicinal chemistry and material science.

Recombinant antibodies and antibody fragments have become powerful tools for therapy, in vivo and in vitro diagnostics, and laboratory research. However, the production of antibody fragments in high yield for preclinical and clinical trials can be a serious bottleneck in drug discovery. This dissertation describes the development of novel screening systems for isolating antibody fragments and alternatively, E. coli genes, that facilitate expression in E. coli. In the first part of this work, we have developed a screening system for isolating Fab mutants exhibiting 4~5 fold higher expression level at 37oC compared to the parental Fab, by utilizing the APEx 2-hybrid system and multi-color FACS as a screening tool. In the APEx 2-hybrid system, the bacterial periplasm constitutes the milieu for the association of membrane-anchored bait protein and solubly expressed, epitope-tagged prey protein. Upon disruption of the outer membrane, only prey proteins that bind to the bait remain cell-associated and are detected by flow cytometry using fluorescently labeled anti-epitope antibodies. In the second part of this work we developed a new strategy to engineer scFv that can be expressed in soluble and active form in the absence of disulfide bonds. This was achieved using a strain incapable of forming disulfide bonds in proteins expressed in its periplasm in combination with the APEx 2-hybrid system. The selected clones exhibited higher solubility, activity, and stability than that of the wild type scFv in the reducing condition of the cytoplasm. Finally, we sought to isolate E. coli gene fragments that can enhance IgG production in the periplasm of E. coli by a newly developed screening system based on soluble expression of IgG and E. coli genomic fragments. The isolated gene fragments, which are located between moeA and iaaA in the E. coli chromosome, improved the total expression of polypeptides of IgG and also assembly of IgG. / text

The escalation with overdose control (EWOC) method is a popular modelbased dose finding design for phase I clinical trials. Dose finding for combined drugs has grown rapidly in oncology drug development. A two-dimensional EWOC design is proposed for dose finding with two agents in combination based on a four-parameter logistic regression model. During trial conduct, the posterior distribution of the maximum tolerated dose (MTD) combination is updated continuously in order to find the appropriate dose combination for each cohort of patients. The probability that the next dose combination exceeds the MTD combination can be controlled by a feasibility bound, which is based on a prespecified quantile for the MTD distribution such as to reduce the possibility of over-dosing. Dose escalation, de-escalation or staying at the same doses is determined by searching the MTD combination along rows and columns in a two-drug combination matrix. Simulation studies are conducted to examine the performance of the design under various practical scenarios, and illustrate it with a trial example. / published_or_final_version / Statistics and Actuarial Science / Master / Master of Philosophy

Most antibiotics, including the drugs currently used for treating tuberculosis (TB), were first discovered as molecules that inhibit bacterial growth in laboratory culture conditions and later translated to infection models and clinical use. Mycobacterium tuberculosis (Mtb) has evolved specifically to survive in its human host, and it is in this infectious context that new drugs need to work. The host environment is characterized by a multitude of antimicrobial defenses induced by the immune system, and we can leverage these defenses to kill Mtb in vivo. Mtb employs a diverse set of responses to survive host defenses. By blocking these responses, we can make Mtb more susceptible to host immunity, turning these previously impotent defenses into effective strategies of immune control.

A challenge in clinical medicine is that of correct diagnosis of disease. Medical researchers invest considerable time and effort to enhance accurate disease diagnosis. Diagnostic tests are important components in modern medical practice. The receiver operating characteristic (ROC) is a commonly used statistical tool for describing the discriminatory accuracy and performance of a diagnostic test. A popular summary index of discriminatory accuracy is the area under ROC curve (AUC). In the era of high-dimensional data, scientists are evaluating hundreds to multiple thousands of biomarkers simultaneously. A critical challenge is the combination of these markers into models that give insight into disease. In infectious disease, markers are often evaluated in the host as well as in the microorganism or virus causing infection, adding more complexity to the analysis. In addition to providing an improved understanding of factors associated with infection and disease development, combinations of relevant markers is important to diagnose and treat disease. Taken together, this presents many novel and major challenges to, and extends the role of, the statistical analyst. In this thesis, we will address the problem of how to select from multiple markers using existing methods. Logistic regression models offer a simple method for combining markers. We applied resampling methods (e.g., Cross-Validation and bootstrap) to adjust for overfitting associated with model selection. We simulated several multivariate models to evaluate the performance of the resampling approaches in this setting. We applied the methods to data collected from a study of tuberculosis immune reconstitution inflammatory syndrome (TB-IRIS) in Cape Town. Baseline levels of five biomarkers were evaluated and we used this dataset to evaluate whether a combination of these biomarkers could accurately discriminate between Tuberculosis Immune Reconstitution Inflammatory Syndrome (TB-IRIS) and non TB-IRIS patients, applying AUC analysis and resampling methods. / Thesis (M.Sc.)-University of KwaZulu-Natal, Pietermaritzburg, 2012.

Polymorphism and crystallinity are now recognised as important issues in drug development. This is shown by the increased amount of research in this area over recent years. In pharmaceutical development milling is an important unit operation for size reduction to improve powder handling, processing and dissolution rate. The aim of this thesis was to investigate the effect of ball milling (and cryo-milling) on the solid state properties, including amorphous drug formation, of pharmaceutical solids. Milling was carried out using an oscillatory ball mill (Mixer Mill MM301, Retsch GmbH & Co., Germany). In cryo-milling the milling jars were immersed in liquid nitrogen for three min before milling. XRPD was used as the main technique to evaluate the milled samples. Ranitidine hydrochloride (RAN) and indomethacin (INDO) were the model drugs used in this study. It was found that upon milling, RAN form 1 converts to RAN form 2 via an amorphous phase. A faster amorphization rate was observed when the crystalline samples were cryo-milled. Amorphous ranitidine hydrochloride was characterized to have a glass transition (T[g]) range of 13 to 30 �C and a crystallization exotherm (T[c]) between 30 and 65 �C. Conversion was found to occur faster when the temperature of the solid powder was greater than the T[c]. Under various storage conditions, similarly, crystallization of the amorphous phase mainly led to RAN form 2. However, some form 1 and amorphous phase was also detected on the XRPD diffractograms. Using partial least squares regression, the amount of solid form components in the ternary RAN mixtures were successfully quantified. RAN form 2 did not convert to form 1 under any milling (including cryo-milling) or storage conditions used in this study. Overall, RAN form 2 was found to be the thermodynamically stable form and the two (RAN) polymorphs are likely to be a monotropic pair. In a co-milling study of INDO and RAN, the two crystalline drugs were successfully converted into a single amorphous phase after 60 min of co-milling in a cold room (4 �C). The T[g] range (26-44 �C) was also characterized for these mixtures. DRIFTS spectra of the co-milled amorphous samples indicated an interaction had occurred between the carboxylic acid carbonyl (HO-C=O) and benzonyl amide (NC=O) of the INDO molecule with the aci-nitro (C=NO₂) of RAN. Depending on the ratio of INDO to RAN, in general, the amorphous mixtures were stable at 4 �C after 30 days of storage. Crystallization was faster when the binary mixtures were stored at higher temperature or contained higher amounts of RAN in the mixture. Although XRPD and DRIFTS suggested an interaction between the two drugs, co-crystal formation was not observed between INDO and RAN. Ball milling can be used to produce amorphous drug. The rate and extent of amorphization is dependent on the milling conditions. A faster rate of amorphization was observed when the crystalline drugs were cryo-milled. Amorphous drug formation can be made either alone or in combination with another crystalline drug. Amorphization could offer a significant improvement on the dissolution profile and the bioavailability of the poorly water soluble drug - indomethacin. Furthermore, ball milling can also be used to produce a homogenous mix between solids. The �goodmix� effect can be used for seed-induced crystallization or, when the XRPD or Raman data were combined with partial least squares regression, to create a reliable calibration model for quantitative analysis.

Thesis (M.P.H.)--University of Hong Kong, 2007.

Thesis (Ph.D.)--Baylor University, 2008. / Includes bibliographical references (p. 112-117).

Thesis (M. P. H.)--University of Hong Kong, 2007. / Also available in print.

Thesis (Ph. D.)--University of Texas at Austin, 2008. / Vita. Includes bibliographical references.