Download Abstract - Iraqi Cultural Attache

Abstract
Modulation of DNA base excision repair (BER) has the potential to enhance
response to chemotherapy and improve outcomes in tumours such as
melanoma, glioma and pancreatic cancer. APE1, a critical protein in BER
that processes potentially cytotoxic abasic sites (AP sites), is a promising
new target in cancer. In the current study, my aim was to develop small
molecule inhibitors of APE1 for cancer therapy. An industry-standard high
throughput virtual screening strategy was adopted. The SYBYL8.0 (Tripos,
St Louis, MO, USA) molecular modelling software suite was used to build
inhibitor templates. Similarity searching strategies were then applied using
ROCS 2.3 (Open Eye Scientific, Santa Fe, NM, USA) to extract
pharmacophorically related subsets of compounds from a chemically diverse
database of 2.6 million compounds. The compounds in these subsets were
subjected to docking against the active site of the APE1 model, using the
genetic algorithm-based programme GOLD2.7 (CCDC, Cambridge, UK).
Predicted ligand poses were ranked on the basis of several scoring functions.
The top virtual hits with promising pharmaceutical properties underwent
detailed in vitro analyses using fluorescence-based APE1 AP-site cleavage
assays and counter screened using endonuclease IV AP-site cleavage assays,
fluorescence quenching assays and Whole cell extract AP-site cleavage
assays. Biochemical APE1 inhibitors were then subjected to detailed
cytotoxicity analyses. Several specific APE1 inhibitors were isolated by this
approach. The IC50 for APE1 inhibition ranged between 50 nM and 25 μM. I
also demonstrated that APE1 inhibitors lead to accumulation of AP sites in
genomic DNA and potentiated the cytotoxicity of alkylating agents in
melanoma, glioma and pancreatic cancer cell lines. I have
also shown that APE1 inhibitors induce delay in cell cycle progression and
caused delay in cancer cell growth. I also demonstrated that APE1
knockdown by shRNA results in decrease cancer cell growth and enhanced
cell killing by alkylating agent in Glioma cell line. This PhD project
provides evidence that APE1 is an emerging drug target and could have
therapeutic application in patients with melanoma, glioma and pancreatic
cancer.