A Systems Analysis of the Oxidative Stress Response in Breast Epithelia and Triple-Negative Breast Cancer

In order to survive, cells must sense and respond to countless internal and external stimuli. Some stimuli will induce stress, which triggers variable responses depending on the type, severity and duration of stress. For cells living in an aerobic environment, reactive oxygen species (ROS) are a continual source of stress that must be dealt with. ROS are reactive derivatives of oxygen produced by normal cellular metabolism and in response to various microenvironmental factors. Depending on concentration and context, ROS can promote cell signaling and proliferation or cause oxidative damage to biomolecules and apoptosis or senescence. To deal with these pervasive, potentially harmful species, cells are equipped with several pathways to sense and detoxify ROS.
In this dissertation, we use systems biology approaches to interrogate a stress-associated transcriptional state in breast mammary epithelial cells. Using several bioinformatic and analytical approaches, we identify the antioxidant transcription factor NRF2 as a candidate regulator of the group of transcripts. From there, we use molecular and cellular biology techniques, image processing, and computational modeling to decipher the broader NRF2 network responsible for oxidative stress handling. We uncover that NRF2 is activated together with another stress-responsive pathway, the p53 pathway, in single cells to mount a coordinated response to oxidative stress in 3D spheroid culture. NRF2–p53 coordination is retained in normal primary breast tissue and hormone-negative DCIS. However, the two pathways are largely uncoupled in triple-negative breast cancers, where p53 is usually mutated.
We then develop a computational systems model of NRF2–p53 signaling in response to transient perturbations in oxidative stress. Using the model, we robustly test hypotheses regarding NRF2–p53 network architecture and coordination during different stages of breast cancer. The integrated NRF2–p53 model predicts variable extents of uncoupling among TNBCs lines, and high uncoupling coincides with the most-severe 3D growth alterations upon NRF2 knockdown, suggesting a reduced tolerance for oxidative stress.
While previous research has focused on NRF2’s direct interaction with mutated TNBC tumor suppressors, our work describes an important systems-level role for wild-type NRF2 and p53 in oxidative-stress tolerance of normal breast–mammary epithelia and hormone-negative premalignancies.