Inhibiting protein phosphatase 2A increases the antitumor effect of protein arginine methyltransferase 5 inhibition in models of glioblastoma

Abstract

Background Despite multi-model therapy of maximal surgical resection, radiation, chemotherapy, and tumor treating fields, the median survival of Glioblastoma (GBM) patients is less than 15 months. Protein Arginine Methyltransferase 5 (PRMT5) catalyzes the symmetric di-methylation of arginine residues and is overexpressed in GBM. Inhibition of PRMT5 causes senescence in stem-like GBM tumor cells. LB100, a first-in-class small molecular inhibitor of Protein Phosphatase 2A (PP2A) can sensitize therapy-resistant tumor cells. Here, we tested the anti-GBM effect of concurrent PRMT5 and PP2A inhibition. Methods Patient-derived primary GBM neurospheres (GBMNS), transfected with PRMT5 target-specific siRNA were treated with LB100 and subjected to in vitro assays including PP2A activity and western blot. The intracranial mouse xenograft model was used to test the in vivo antitumor efficacy of combination treatment. Results We found that PRMT5-depletion increased PP2A activity in GBMNS. LB100 treatment significantly reduced the viability of PRMT5-depleted GBMNS compared to PRMT5 intact GBMNS. LB100 enhanced G1 cell cycle arrest induced by PRMT5-depletion. Combination therapy also increased the expression of phospho-MLKL. Necrostatin-1 rescued PRMT5-depleted cells from the cytotoxic effects of LB100, indicating that necroptosis caused the enhanced cytotoxicity of combination therapy. In the in vivo mouse tumor xenograft model, LB100 treatment combined with transient depletion of PRMT5 significantly decreased tumor size and prolonged survival, while LB100 treatment alone had no survival benefit. Conclusion Overall, combined PRMT5 and PP2A inhibition had significantly greater antitumor effects than PRMT5 inhibition alone.

PP2A and cancer epigenetics: a therapeutic opportunity waiting to happen

“…Therefore, PP2A-activating compounds may be effective in cancers with functional NDRG2 and may synergize with PRMT5 inhibiting compounds, while NDRG2 deletions may render this strategy ineffective. Conversely, in glioblastoma, PRMT5 exhibits a tumor suppressive role, while NDRG2 is reported to have oncogenic properties ( 122 , 123 ). Consistent with the findings, treatment of glioblastoma with PP2A-inhibitor LB100 has an anti-tumor effect in the context of PRMT5 regulation ( 122 ).…”

“…Conversely, in glioblastoma, PRMT5 exhibits a tumor suppressive role, while NDRG2 is reported to have oncogenic properties ( 122 , 123 ). Consistent with the findings, treatment of glioblastoma with PP2A-inhibitor LB100 has an anti-tumor effect in the context of PRMT5 regulation ( 122 ). Therefore, while the mechanism by which PP2A regulates PRMT5 is the similar, the biological outcome of this signaling is contradictory between these two cancers and supports the use of alternative therapeutic approaches.…”

“…Lung cancer, pancreatic cancer and leukemias have been reported to be sensitive to therapeutic re-activation of PP2A activity ( 169 , 171–173 ). On the other hand, cancers such as glioblastoma show therapeutic efficacy with PP2A catalytic inhibitors, such as LB100, particularly when used in combination with DNA damaging therapeutics ( 122 , 174–177 ). The combination of PP2A and epigenetic inhibitors are currently being explored and may prove effective in malignancies of the nervous system ( 122 , 144 ).…”

Review of PP2A Tumor Biology and Antitumor Effects of PP2A Inhibitor LB100 in the Nervous System

“…Most recently, our group studied the effects of combining LB100 with the knockdown of protein arginine methyltransferase 5 (PRMT5). For these experiments, patient-derived primary glioblastoma neurospheres (GBMNS) were initially transfected with PRMT5 siRNA ( Table 2 ) [ 100 ]. LB100 administration in conjunction with PRMT5 knockdown was then studied in the in vitro and in vivo intracranial mice xenografts models.…”

Review of PP2A Tumor Biology and Anti-Tumor Effects of PP2A Inhibitor LB100 in the Nervous System

“…Most recently, our group studied the effects of combining LB100 with the knockdown of protein arginine methyltransferase 5 (PRMT5). For these experiments, patient-derived primary glioblastoma neurospheres (GBMNS) were initially transfected with PRMT5 siRNA (Table 1) [94]. LB100 administration in conjunction with PRMT5 knockdown was then studied in the in vitro and in vivo intracranial mice xenografts models.…”