Investigating the regulation of cancer cell death by NFE2L1 - Project Summary Resistance to therapy is responsible for 90% of cancer patient mortality. Our understanding of how cancer cells resist therapy-induced death is limited. My goal is to better understand the molecular mechanisms of cancer cell therapy resistance, with an eye toward improving therapy. Identifying and characterizing novel resistance mechanisms should facilitate the design of more effective cancer therapies. Immunotherapy and ionizing radiation are two common cancer therapies. These therapies kill cancer cells, in part, via ferroptosis: a non-apoptotic cell death mechanism characterized by lipid peroxidation. Sensitizing cancer cells to ferroptosis may improve the efficacy of these and other cancer therapies. To effectively sensitize cancer cells to ferroptosis, we must identify how cancer cells normally resist ferroptosis. However, the mechanisms by which cancer cells resist ferroptosis are poorly understood. I recently showed that the transcription factor nuclear factor erythroid 2 like 1 (NFE2L1, or NRF1) is required to protect cancer cells from ferroptosis. My overarching hypothesis is that NFE2L1 is central to an important pathway promoting ferroptosis resistance in cancer cells. The goal of this research, as part of my overall training plan, is to test this hypothesis and determine how NFE2L1 protects cancer cells from ferroptosis. Ultimately, this research has the potential to inform advances in cancer therapy. I propose two Specific Aims. In Aim 1, I will test whether certain post-translational modifications of NFE2L1 are required for ferroptosis resistance. These studies will be conducted using genetic and pharmacologic approaches to perturb NFE2L1 N-glycosylation and cleavage and assess the effects on ferroptosis sensitivity. To support the successful execution of this aim, I will be trained in glycobiology by my co-sponsor, Dr. Carolyn Bertozzi, who pioneered this field. In Aim 2, I will elucidate the role of nicotinamide N-methyltransferase (NNMT) in ferroptosis resistance. Preliminary data suggest that NNMT, which encodes an enzyme involved in nicotinamide adenine dinucleotide metabolism, is a direct target of NFE2L1. I hypothesize that NNMT regulates cancer cell ferroptosis sensitivity by altering redox homeostasis. Through my proposed experiments, I will determine whether NNMT is a direct target of NFE2L1. I will also assess ferroptosis sensitivity in cells in which NNMT is knocked out or overexpressed. Mass spectrometry will be used to measure the abundance of redox molecules in these cell lines to analyze the role of NNMT in ferroptosis-related redox regulation. I will be trained in mass spectrometry by my collaborator, Dr. Monther Abu-Remaileh, who is an expert in metabolomics. With the guidance of my expert team of mentors at Stanford University, my proposed experiments will elucidate this novel pathway promoting cancer cell ferroptosis evasion and identify possible strategies to minimize cancer therapy resistance.
