Raloxifene prevents stress granule dissolution, impairs translational control and promotes cell death during hypoxia...

Glioblastoma (GBM) is the most common primary malignant brain tumor, and it has a uniformly poor prognosis. Hypoxia is a feature of the GBM microenvironment, and previous work has shown that cancer cells residing in hypoxic regions resist treatment. Hypoxia can trigger the formation of stress granules (SGs), sites of mRNA triage that promote cell survival. A screen of 1120 FDA-approved drugs identified 129 candidates that delayed the dissolution of hypoxia-induced SGs following a return to normoxia. Amongst these candidates, the selective estrogen receptor modulator (SERM) raloxifene delayed SG dissolution in a dose-dependent manner. SG dissolution typically occurs by 15 min post-hypoxia, however pre-treatment of immortalized U251 and U3024 primary GBM cells with raloxifene prevented SG dissolution for up to 2 h. During this raloxifene-induced delay in SG dissolution, translational silencing was sustained, eIF2α remained phosphorylated and mTOR remained inactive. Despite its well-described role as a SERM, raloxifene-mediated delay in SG dissolution was unaffected by co-administration of β-estradiol, nor did β-estradiol alone have any effect on SGs. Importantly, the combination of raloxifene and hypoxia resulted in increased numbers of late apoptotic/necrotic cells. Raloxifene and hypoxia also demonstrated a block in late autophagy similar to the known autophagy inhibitor chloroquine (CQ). Genetic disruption of the SG-nucleating proteins G3BP1 and G3BP2 revealed that G3BP1 is required to sustain the raloxifene-mediated delay in SG dissolution. Together, these findings indicate that modulating the stress response can be used to exploit the hypoxic niche of GBM tumors, causing cell death by disrupting pro-survival stress responses and control of protein synthesis.