Date of Award

2008

Degree Type

Dissertation

Department

Chemistry

First Advisor

Xu, Yan

Subject Headings

Lymphocytic leukemia -- Chemotherapy, Fludarabine, Base excision repair, Fludarabine, Methoxyamine, Chronic lymphocytic leukemia

Abstract

Nucleotide analogs (e.g. fludarabine) are antimetabolites used in the treatment of a wide variety of hematological malignancies and solid tumors. Upon being metabolized to their active triphosphate form, these agents are incorporated into DNA. Among other molecular targets, their incorporation may lead to activation of base excision repair (BER) pathway. The molecular mechanism of BER recognition and repair of the incorporated fludarabine has not yet been elucidated. The main focus of this research was to study the involvement of BER pathway in the response to fludarabine induced DNA damage. Also, the possibility of enhancing antineoplastic activity of fludarabine by inhibition of BER (e.g. methoxyamine) was investigated. Firstly, capacity of uracil DNA glycosylase to recognize and excise incorporated fludarabine was established. Secondly, the formation of abasic (AP) sites after fludarabine treatment was confirmed in different human cancer cell lines. These results demonstrated that incorporated fludarabine, acting as an abnormal base in DNA, initiates BER. The possibility to enhance fludarabine-induced damage by inhibiting BER was then considered. Exposure of cells to fludarabine and methoxyamine (MX) combined regimens caused increased apoptosis, clonogenic death, upregulation of some key BER proteins, enhanced DNA strand breaks. It also enhanced anti-tumor effects in human xenografts. This response of cells to fludarabine plus MX was due to MX binding to the ara-AP sites formed by fludarabine, thus turning the repairable DNA damage into lethal lesions. In addition, mitochondrial DNA was found to be targeted by fludarabine and fludarabine plus MX. Apoptotic signaling from nuclear and mitochondrial DNA damage triggered mitochondrial mediated cell death during BER disruption by MX. The modulation of fludarabine cytotoxicity by manipulating BER via MX was analyzed in a similar series of experiments using primary lymphocytes obtained from CLL patients. MX enhancement of activity of fludarabine was confirmed the r

Download

Included in

Chemistry Commons

COinS