Title

IN VIVO AND IN VITRO STUDIES OF POLYAMIDES THAT ARE ACTIVE ANTIVIRAL AGENTS AGAINST HPV16

Document Type

Dissertation

Degree

Doctor of Philosophy

Major

Chemistry

Date of Defense

9-5-2016

Graduate Advisor

James K. Bashkin, DPhil

Committee

Dr. Mindy Steiniger

Keith J. Stine

Chung F. Wong

Abstract

Long-term, persistent infection with high-risk strains of human papillomavirus (HPV) is the precursor of most cervical cancers and an increasing number of the head and neck cancers. While HPV vaccines can protect patients under twenty-five years old from possible infections, no HPV antiviral drugs are available for the definitive treatment of preexisting or future viral infections. To treat current or future HPV infections, drugs that selectively block virus-specific processes, but do not damage the host cells, are needed. The compounds developed in our group are unique Imidazole-pyrrole polyamides, analogs of natural products Distamycin A and Netropsin that function by interfering with natural virus-host interactions. Within the overall program of my group, my work focuses on three specific projects. The first project (Chapter 2) includes bioavailability LC-MS/MS analysis study of one of the leading compounds in plasma and whole blood. The second project includes the biophysical study of interactions between two polyamides, called TMG Asymmetric Hairpin Polyamides (TMG-AHP), and an essential viral DNA segment. The aim of this study was to determine where and how strongly the compounds bind to the viral DNA. In this work, two methods were used: the quantitative deoxyribonuclease (DNase I) footprinting method and the affinity cleavage assay (AC). The results are presented in Chapters 3 and 4. The remarkable findings resulted in binding location maps that help us to better understand the mechanism of action of AHPs and to address the question: what is the primary reason for neutralizing a virus by our polyamides. Based on the knowledge gathered from the results of the first project, a complex study (RNA-Seq) concerned with genome expression was performed. The aim of the third project is to find the common features and differences in the polyamides’ mechanism of action by quantifying messenger RNA (mRNA), after treatment of HPV infected skin cells with 8 different polyamides. In parallel with the analysis of human and viral transcriptome, a separate study concerned with the differential expression profile of seven DDR genes that are components of the homologous recombination (HR) pathway have been studied by RT-qPCR method. The results are presented in Chapter 5 and 6. Data gathered from the third project help us further understand the mechanism of polyamide action.

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