Inhibition of PERK mediated UPR acts as a switch for reversal of residual senescence and as senolytic therapy in glioblastoma.

Background: Glioblastoma due to recurrence is clinically challenging with 10-15months overall survival. Previously we showed therapy induced senescence (TIS) in glioblastoma reverses causing recurrence. Here, we aim to delineate TIS reversal mechanism for potential therapeutic intervention to prevent GBM recurrence.

Methods: Residual senescent (RS) and End of Residual Senescence (ERS) cells were captured from GBM patient-derived primary-cultures and cell lines mimicking clinical scenario. RNA-sequencing, transcript/protein validations, knock-down/inhibitor studies, ChIP RT-PCR, biochemical assays and IHCs were performed for mechanistics of TIS reversal. In vivo validations were conducted in GBM orthotopic mouse model.

Results: Transcriptome analysis showed co-expression of ER stress-UPR and senescence associated secretory phenotype (SASP) with TIS induction and reversal. Robust SASP production and secretion by RS cells could induce senescence, ROS, DNA damage and ER stress in paracrine fashion independent of radiation. Neutralization of most significantly enriched cytokine from RS-secretome IL1β, suppressed SASP and delayed senescence reversal. Mechanistically, with SASP and massive protein accumulation in Endoplasmic reticulum, RS cells displayed stressed ER morphology, upregulated ER stress markers and PERK pathway activation via peIF2α-ATF4-CHOP which was spontaneously resolved in ERS. ChIP RT-PCR showed CHOP occupancy at CXCL8/IL8, CDKN1A/p21 and BCL2L1/BCLXL aiding survival. PERK knockdown/inhibition with GSK2606414 in combination with radiation led to sustained ER stress and senescence without SASP. PERKi in RS functioned as senolytic via apoptosis and prevented recurrence in vitro and in vivo ameliorating overall survival.

Conclusion: We demonstrate that PERK mediated UPR regulates senescence reversal and its inhibition can be exploited as potential seno-therapeutic option in glioblastoma.