Document Type

Dissertation

Date of Degree

Summer 2013

Degree Name

PhD (Doctor of Philosophy)

Degree In

Pharmacy

First Advisor

Mahfoud Assem

Abstract

Glioma is the most common and aggressive type of primary intracranial tumors. The poor prognosis of glioma patients has not changed since decades despite the advancements in diagnostic tools and treatment strategies. The inability to accurately predict the survival and response to anticancer therapy emerges from several factors including the high heterogeneity of the tumor and the inadequacy of the currently applied world-health organization (WHO) classification system. Both factors result in high variability in the clinical outcome due to variable sensitivity to treatment. Thus, molecular classification represents an important strategy for better categorization of glioma patients and for their stratification to anticancer therapy. Our high-throughput screening analysis for the identification of genetic aberrations in the glioma study population revealed high frequency of chromosomal instabilities in glioma specimens. This indicates that DNA-repair mechanisms are defective which may have contributed to gliomagenesis and progression.

Furthermore, DNA-repair represents an integral interplayer in the determination of glioma response to anti-neoplastic agents due to the fact that the majority of the currently applied agents possess their cytotoxicity via DNA-damaging actions. TDP1 has been implicated in the resistance to various types of anticancer agents in vitro, including radiation and topoisomerase poisons due to its ability to repair various types of DNA lesions. Moreover, it has been found to be overexpressed in different kinds of cancers, however, its relevance in glioma has not yet been studied.

In this work we show that TDP1 is overexpressed in patients with malignant glioma compared with non-tumor cases. An ascending increase of TDP1 protein expression with a correlation with glioma grade is evidenced in the astrocytic lineage and glioblastoma multiforme samples expressed the highest levels. Moreover, we show an association between high TDP1 transcript levels and the poor prognosis of glioma patients. These findings suggest that TDP1 plays an important role in gliomagenesis; however, the underlying molecular mechanisms need to be identified.

For an exploration of the predictive value of TDP1 in malignant glioma the correlation between TDP1 level and the sensitivity of malignant glioma cell lines to anticancer therapy has been investigated. We show that manipulating TDP1 level alone in malignant glioma cell lines is not sufficient to modulate their response to treatment. TDP1 overexpression or knockdown resulted in changes in the transcript levels of several DNA-repair genes including MGMT, topoisomerases and the base excision repair genes PARP-1, PNKP and XRCC1. This hindered the ability to characterize the role of TDP1 to modulate the in vitro sensitivity of malignant glioma cell lines to topoisomerase poisons and temozolomide. Nonetheless, this emphasizes the importance of the comprehensive role of several DNA-repair genes for a finalized DNA-repair process to determine the sensitivity of tumor cells to DNA-damaging anticancer agents.

Finally, we tested the ability of inhibiting TDP1 enzyme activity to potentiate or synergize the cytotoxicity of topoisomerase poisons using small-molecule ligands. We show that treatment of malignant glioma cell lines with a combinational therapy of a small-molecule TDP1 inhibitor and a topoisomerase poison enhances their sensitivity to the latter drug but with minimal efficacy.

As a conclusion, the characterization of TDP1 in glioma is a novel finding that can aid in enhancing the diagnosis and prognosis of patients. However, its role as a predictive biomarker for better stratification of patients to therapy needs further investigation.

Pages

xix, 145 pages

Bibliography

Includes bibliographical references (pages 133-145).

Copyright

Copyright 2013 Maha Al-Keilani

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