Graduation Semester and Year

2008

Language

English

Document Type

Dissertation

Degree Name

Doctor of Philosophy in Chemistry

Department

Chemistry and Biochemistry

First Advisor

Subhrangsu Mandal

Abstract

Most of biological catalysts including nucleases are made of active metal centers. Metal complexes that are able to interact with DNA and RNA mimicking these natural nucleases will be valuable as therapeutic agents that target nucleic acids. Metal complexes have been used for very long time to treat different kinds of illnesses including cancer. Platinum based compounds are among the most widely used anticancer agents in clinical use. The first chapter of this dissertation highlights the roles of metal ions and their complexes with regard to biochemical activities, as enzyme inhibitors, anticancer agents and metallo-nucleases. Moreover, we will discuss the problems associated with the use of anticancer drugs like cisplatin and the steps taken to alleviate that using targeted drug delivery system. Although several drugs are in use for treating various illnesses genetic diseases which are not treatable by conventional therapeutics have recently been a target by gene therapy. This chapter also discusses the types of gene delivery vehicles used and efforts made to optimize the process of transfection. In chapter II we discuss the DNA cleavage activities of Fe(III)-salen, its mechanism of action on DNA and sequence specificity. We have found that Fe(III)-salen cleaves DNA in the presence of DTT in a sequence neutral fashion by generating free radicals as indicated by DNA cleavage inhibition experiments. In vitro transcription assay performed on naked DNA template shows that Fe(III)-salen inhibits transcription. Further we have analyzed the effects of this metal complex on cultured HEK293 cells and investigated the ability of the complex to affect cell viability and tocause apoptosis (programmed cell death). The cell viability study undertaken shows that Fe(III)-salen is a potent cytotoxic agent with IC50 value of 2 µM. We have found that the metal complex has the ability to kill cells via apoptotic process as observed by DAPI staining of the nucleus of the cell. We have studied the mechanism of action of apoptosis and we were able to determine that the metal complex causes apoptosis through intrinsic mitochondrial pathway.In chapter III we have prepared a series of derivatives of Fe(III)-salen compounds. Initially, we synthesized several Fe(III)-salen derivatives by changing the bridging groups and introducing various hydroxy and methoxy functionalities into the salen moiety. We have investigated the effect bridging groups and substituents on the Fe(III)-salen mediated DNA cleavage properties. Our result showed that the DNA cleavage efficiency decreases as the bridging group is changed from ethylenediamine to ortho-pheynlenediamine to 2,3-diaminonaphthalene. Hydroxy-substituted Fe(III)-salnaphens were more efficient on inducing DNA cleavage compared to corresponding methoxy derivatives. The cytotoxicity analysis of Fe(III)-salen derivatives showed that several compound induce efficient cell death towards MCF7 and HEK293 cells. Most interestingly, however, Fe(III)-salen complexes with lower DNA cleavage activity exhibited higher cytotoxicity. These results demonstrated that in vitro DNA clevage properties of the metallo-salen are not essential for their cytotoxicity. Similarly, we have also investigated the effect of 4, 4΄-dialkoxy Fe(III)-salen complexes containing various long chain derivatives on HEK293 cells. Our result demonstrated that cytotoxicity of Fe(III)-salen lipid molecules is decreased with the increase in carbon chain length. In Chapter IV we investigated the self-assembly properties and DNA transfection abilities of our novel long chain containing 4, 4΄-dialkoxy Fe(III)-salen lipid complexes. Using dynamic light scattering, scanning electron microscopy and dye-encapsulation experiments, we demonstrated that Fe(III)-salen lipids self-assembled into hollow liposomal structures. We also showed that Fe(III)-salen liposomes form complexes with negatively charged DNA molecules forming lipid-DNA complexes. Moreover, One of the Fe(III)-salen liposome showed efficient DNA delivery (transfection) into cultured human cells.

Disciplines

Chemistry | Physical Sciences and Mathematics

Comments

Degree granted by The University of Texas at Arlington

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