High-resolution approaches to dissect the role of subcellular redox circuits in vivo

Project Summary Metastasis is the leading cause of death for cancer patients. Oxidative stress, characterized by excessive exposure to reactive oxygen species (ROS), kills most metastasizing cancer cells. How highly-metastatic clones manage to overcome both cell-intrinsic and extrinsic oxidative insults to colonize distant organs is

poorly understood. The precise subcellular circuits enabling tumor adaptation to oxidative stress, and whether they could be exploited for therapy, have remained elusive. We hypothesize that organellar antioxidant pathways provide adaptive mechanisms essential for cancers to efficiently metastasize. A major impediment to

testing this hypothesis has been the lack of high-resolution and versatile tools to study ROS in vivo. To address this major technological gap and address which subcellular redox circuits are necessary or sufficient for highly metastatic tumors to progress, we will develop and apply tools with exquisite

spatiotemporal resolution in vivo. These tools include an optogenetic protein that produces localized ROS in tumor subcellular compartments, gene therapy strategies in mice that will pioneer the manipulation of tissue- extrinsic ROS in mouse tissues, and tumor organelle purification strategies coupled to mass spectrometry

analyses in primary and metastatic tumors. With these tools in hand, this proposal aims to answer three key questions: Can the subcellular burden of ROS be exploited to hinder metastasis? Are there specific organelle-based nodes that enhance tumor antioxidant capacity for metastasis? Are extracellular ROS in the colonized target organ a major metastasis limitation? By

integrating in vivo optogenetic modulation of ROS, high-resolution metabolomics, functional genetics and metastatic cancer models, our work will uncover targetable, ROS-mediated bottlenecks of metastasis at subcellular resolution. The tools and techniques developed in this proposal have the potential to revolutionize our ability to study ROS

and oxidative stress both in vitro and in vivo, and be broadly applied to any disease impacted by ROS and oxidative stress. As such, my lab’s work has the power to reshape our understanding of these processes not only in cancer, but across a wide range of diseases, which may pave the way for new therapeutic strategies

and improved patient outcomes.