Faculty & Staff Directory
Franklin G. Berger
Director, Center for Colon Cancer Research
Department of Biology
University of South Carolina
|Office:||PSC 614||Phone Number:||Office: 803-777-1171; Lab: 803-777-7216; Office: 803-777-1231||Email:||firstname.lastname@example.org|
|Website:||Berger Lab/ Center for Colon Cancer Research|
Background: Colorectal cancer is the fourth-highest cancer in terms of incidence, and second-leading cause of cancer death. In the US, over 140,000 new cases of colorectal cancer are diagnosed each year, and nearly 50,000 patients die from the disease. Like most cancers, less-than-optimal efficacy of anti-neoplastic agents is a major barrier to the successful treatment of advanced colorectal cancer. The high genetic plasticity of neoplastic cells generally leads to rapid emergence of resistance to chemotherapeutic agents, reducing therapeutic effectiveness. For several years, we have been examining an important class of chemotherapeutic agents that target the pyrimidine biosynthetic enzyme thymidylate synthase (TYMS). TYMS catalyzes the reductive methylation of dUMP to form dTMP, making it indispensible for DNA synthesis during cell proliferation and DNA repair. Exposing cells to TYMS-directed antimetabolites, such as fluoropyrimidine analogs (e.g., 5-fluorouracil and 5-fluoro-2'- deoxyuridine), as well as antifolates (e.g., tomudex, AG337, and BW1843), results in the inhibition of TYMS, causing depletion of thymidylate pools, cessation of cell growth, and eventually, cell death. Thus, TS inhibitors have been useful in the treatment of a variety of neoplasms. However, a large fraction of patients are intrinsically resistant to TYMS inhibitors, and do not respond to these drugs; furthermore, those that do respond typically develop resistance, so that the drugs lose their efficacy.
Oxidative stress and cell death in response to TYMS inhibition: We have recently shown that apoptosis in response to TYMS inhibitors is due to oxidative stress brought on by increased concentrations of reactive oxygen species (ROS). This is a consequence of activation of the membrance-bound enzyme NADPH oxidase (NOX). NOX catalyzes transfer of an electron from NADPH to molecular oxygen to form superoxide, a reactrion that is essential for maximal response to TYMS inhibitors. The mechanism of NOX activation by TYMS-directed agents is not completely understood, but likely involves induction of expression of the NCF2 gene, which encodes the NOX regulatory subunit p67phox .
Role of the anti-oxidant genes in response to TYMS inhibitors: Recent gene expression profiling experiments revealed that cells mount a complex protective response to TYMS inhibitors, involving activation of genes that function in amelioration of oxidative and electrophilic stress. Of particular interest was the recognition that many of these genes are regulated by transcription factor NRF2, which is well-known to attenuate the concentration and impact of ROS. We have suggested that the expression and activation of NRF2 is a constraining factor in cell death following TYMS inhibition. Indeed, we have observed that down-regulation of NRF2 by specific siRNAs, as well as depletion of the transcription factor by CRISPR/Cas9-mediated gene editing, sensitizes cells to the cytotoxic effects of drug exposure. Our working model, depicted in the figure below, postulates that cellular response to TYMS inhibitors is determined by two counter-acting effects of drug exposure. In one, termed the death pathway, the inhibitors elicit DNA damage, NOX-dependent formation of superoxide, and oxidative stress, resulting in promotion of apoptosis (pathway (i.), red arrows). In the other, termed the survival pathway, NRF2 is activated, leading to induction of its downstream target genes, attenuation of oxidative stress, and resistance to apoptosis (pathway (ii.), green arrows). In the end, the relative activities of the death vs. survival pathways govern response to drug exposure. Predominance of pathway (i.) leads to drug sensitivity, while predominance of pathway (ii.) leads to resistance. Through an understanding of the role of NRF2 in response to TYMS-directed drugs, we hope to design strategies to reduce or eliminate the transcripion factor's promotion of survival, thereby increasing the therapeutic efficacy of drug treatment.
Ozer, U., Barbour, K.W., Clinton, S.A., and Berger, F.G. 2015. Oxidative stress and response to thymidylate synthase-targeted antimetabolites. Molecular Pharmacology 88: 970-981.