Discovery and investigation of an ATG4B-IMPDH2 axis in breast cancer cells

Autophagy is an intracellular catabolic process that plays both pro- and anti-tumorigenic roles depending on cancer stage and context. ATG4B is a core cysteine protease in autophagy and has been associated with modulating the responses of breast cancer cells to nutrient deprivation and targeted treatment. To date, the molecular mechanisms underlying ATG4B in nutrient stress and treatment response in breast cancers remain poorly defined. To this end, I identified a new protein-protein interaction between ATG4B and IMPDH2, a metabolic enzyme involved in de novo GTP biosynthesis. A key amide donor in the de novo biosynthesis of purines, like GTP, is glutamine, a critical amino acid for cancer cell growth. I discovered that ATG4B is important for assembly of IMPDH2 into ring-and-rod structures under glutamine deprivation, suggesting that ATG4B may be important in regulating cellular GTP. I discovered that genetic loss of ATG4B is associated with an enrichment in purine nucleotides and metabolites involved in the purine salvage pathway, an alternate pathway for GTP production, suggesting an increased utilization of the purine salvage pathway. To investigate potential cellular consequences of this metabolic shift, I examined growth of parental and ATG4B knockout (KO) breast cancer cells and found that ATG4B KO significantly impaired growth under glutamine deprivation. Inhibition of IMPDH had a modest effect on growth of glutamine-deprived ATG4B KO cells, suggesting a reliance on IMPDH-independent pathways for growth. Depletion of exogenous purines resulted in a significant impairment in ability of ATG4B KO cells to grow in glutamine-depleted media, supporting a reliance on the purine salvage pathway for growth. I then investigated the potential therapeutic applications of the ATG4B-IMPDH2 axis in breast cancer cells and found that pharmacological inhibition of ATG4 and IMPDH results in a modest but synergistic reduction in growth. Collectively, my research has identified a new protein-protein interaction between ATG4B and IMPDH2, and a novel metabolic reprogram wherein cells may increase utilization of the purine salvage pathway for growth in glutamine-deprived conditions in breast cancer cells.