Polypharmacologic phosphoinositide modulation by FTY720 triggers endomembrane trafficking collapse and metabolic starvation in cancer cells.

Phosphoinositides coordinate membrane trafficking and bioenergetic homeostasis, and many tumors rely on elevated phosphoinositide flux to sustain growth. Therapeutic development has largely pursued single-phosphoinositide kinase inhibition, whereas polypharmacologic strategies that perturb the broader network remain underexplored. FTY720 (fingolimod), a clinically approved sphingosine-1-phosphate receptor modulator, shows anti-tumor activity at micromolar concentrations, but its non-canonical mechanisms remain incompletely defined. Building on our work with the structurally related compound KRP203, we show that high-dose FTY720 produces isozyme-divergent modulation across phosphoinositide kinases and biases PIKFYVE activity toward phosphatidylinositol, a pattern we term ASURA (Asymmetric Simultaneous Uncoupling of Related Activities). FTY720 induces vacuolization and endomembrane remodeling in cancer cells, and suppresses macropinocytic ruffling as demonstrated by tracer uptake and scanning ion conductance microscopy analyses. Quantitative metabolomics revealed depletion of intracellular amino acids and ribonucleoside triphosphates, coupled with reduced glycolysis. Concurrently, FTY720 induced extensive rewiring of the hexosamine pathway, nitrogen metabolism, and tricarboxylic acid (TCA)-cycle anaplerosis, along with redox signatures indicating oxidative stress despite a nutrient-replete medium. The metabolites depleted by FTY720 showed extensive, directionally concordant overlap with those depleted by PTEN induction. Patient-derived glioblastoma (GBM) neurospheres were sensitive to FTY720, and co-treatment with a PI3Kα-selective inhibitor augmented growth suppression in U87MG cells. Together, these data support a model in which ASURA-dose FTY720 disrupts phosphoinositide-regulated trafficking and nutrient access, imposing intracellular nutrient stress that culminates in tumor-cell death.