Squalene accumulation in cholesterol auxotrophic lymphomas prevents oxidative cell death

Abstract

Cholesterol is essential for cells to grow and proliferate. While normal mammalian cells meet their need for cholesterol through its uptake or de novo synthesis 1 , the extent to which cancer cells rely on each of these pathways remains poorly understood. Here, using a competitive proliferation assay on a pooled collection of DNA-barcoded cell lines, we identified a subset that is auxotrophic for cholesterol and thus highly dependent on its uptake. Metabolic gene expression analysis pinpointed loss of squalene monooxygenase (SQLE) expression as a cause of the cholesterol auxotrophy, particularly in ALK+ anaplastic large cell lymphoma (ALCL) cell lines and primary tumors. SQLE catalyzes the oxidation of squalene to 2,3-oxidosqualene in the cholesterol synthesis pathway and its loss results in accumulation of the upstream metabolite squalene, which is normally undetectable. In ALK+ ALCLs, squalene alters the cellular lipid profile and protects cancer cells from ferroptotic cell death, providing a growth advantage under conditions of oxidative stress and in tumor xenografts. Finally, a CRISPR-based genetic screen identified cholesterol uptake by the low-density lipoprotein receptor (LDLR) as essential for the growth of ALCL cells in culture and as patient-derived xenografts. This work reveals that the cholesterol auxotrophy of ALCLs is a targetable liability, and, more broadly, that systematic approaches are useful for identifying nutrient dependencies unique to individual cancer types.

A key mammalian cholesterol synthesis enzyme, squalene monooxygenase, is allosterically stabilized by its substrate

“…This work provides mechanistic insights into the regulation of human SM, an essential rate-limiting enzyme of cholesterol synthesis that is implicated in disease (32)(33)(34)(35). The substrate of SM, squalene, is known to bind to the catalytic domain of SM.…”

GPX4 at the Crossroads of Lipid Homeostasis and Ferroptosis

“…Higher cholesterol levels may expand the pool of oxidizable lipids and contribute to lipid peroxidation during ferroptosis. Of note, in ALK + large cell lymphomas, the accumulation of squalene due to the loss of squalene monooxygenase expression inhibits lipid peroxidation and the onset of ferroptosis . It is therefore possible that different MVA intermediates and products (e.g., cholesterol) will have cell type‐specific roles in modulating ferroptosis sensitivity.…”

Targeting Scavenger Receptor Type B1 In Cholesterol-Addicted Lymphomas Abolishes Glutathione Peroxidase 4 and Results in Ferroptosis

“…Furthermore, our western blot data show a profound reduction in GPX4 which suggest post-translational mechanisms that would further reduce GPX4. Certainly, in this regard, there is precedent in the literature for lipid metabolism, in general, and intermediate metabolites in cholesterol biosynthesis, in particular, involved in regulating GPX4 stability (16). Interestingly, inhibiting de novo cholesterol biosynthesis using statins did not reduce GPX4 expression or induce ferroptosis (40), suggesting that manipulation of de novo cholesterol biosynthesis is unable to replicate the effects of HDL NP treatment.…”

“…Recently, a number of cell lines were found to be auxotrophic for cholesterol, including the cell lines SR-786 (ALK + ALCL), SUDHL1 (ALK + ALCL), and U937 (isolated from histiocytic lymphoma, but of myeloid lineage), among others (16). The ALK + ALCL cells were identified based upon reduced viability when cultured in lipoprotein deficient serum, and the cell death phenotype was rescued by addition of cholesterol-rich low-density lipoprotein (LDL) or free cholesterol.…”

“…Data suggest a link between cellular cholesterol uptake and ferroptosis. For instance, removal of lipoproteins from media used to culture ALK + anaplastic large T cell lymphoma (ALK + ALCL) cell lines (SR-786, SUDHL1) and a histiocytic lymphoma cell line (U937) induced ferroptosis (16). The data focused on the role of cholesterol-rich low-density lipoprotein (LDL) binding to the LDL receptor (LDLR) as responsible for cholesterol delivery to the malignant cells.…”