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Figure 4 from Coordinated Targeting of S6K1/2 and AXL Disrupts Pyrimidine Biosynthesis in PTEN-Deficient Glioblastoma

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posted on 2024-08-23, 10:20 authored by Catherine A. Behrmann, Kelli N. Ennis, Pranjal Sarma, Collin Wetzel, Nicholas A. Clark, Kate M. Von Handorf, Subrahmanya Vallabhapurapu, Cristina Andreani, James Reigle, Pier Paolo Scaglioni, Jarek Meller, Maria F. Czyzyk-Krzeska, Ady Kendler, Xiaoyang Qi, Jann N. Sarkaria, Mario Medvedovic, Soma Sengupta, Biplab Dasgupta, David R. Plas

Targeting requirements in the S6K1/S6K2 network. A and B, S6 phosphorylation was reduced only upon genetic silencing of both S6K1 and S6K2 in LN229 GBM cells. C, Combined inactivation of S6K1 and S6K2 via sgRNA and siRNA was required to silence S6K signaling in PTEN-deficient GBM U87MG-GFP-Luc. D, DepMap correlation shows dependence on S6K1 and S6K2 when PTEN is mutated. E, PTEN inactivation caused an induction of S6K signaling seen as an increase in the phosphorylation of rpS6. This increase can only be abrogated by combination inactivation of both S6K1 (exon 5) and S6K2 (exon 9). F, sgNT and sgS6K1 GBM LN229 cells were treated with 10 μmol/L LY-2584702 or 10 μmol/L PF-4708671 for 3 hours. Inhibitor suppression of phospho-rpS6 was modestly improved by knockout of S6K1. G, sgNT and sgS6K2 GBM LN229 cells were treated as in F. Inhibitor suppression of phospho-rpS6 was significantly enhanced by S6K2 knockout. H, sgNT and sgS6K2 (exon 9) LN229 cells transfected with siNT or siPTEN for 72 hours were incubated with S6K1 inhibitors for 3 hours. PTEN inactivation induced phospho-rpS6, which was blocked by the combination of sgS6K2 and an S6K1 inhibitor.

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ARTICLE ABSTRACT

Intrinsic resistance to targeted therapeutics in PTEN-deficient glioblastoma (GBM) is mediated by redundant signaling networks that sustain critical metabolic functions. Here, we demonstrate that coordinated inhibition of the ribosomal protein S6 kinase 1 (S6K1) and the receptor tyrosine kinase AXL using LY-2584702 and BMS-777607 can overcome network redundancy to reduce GBM tumor growth. This combination of S6K1 and AXL inhibition suppressed glucose flux to pyrimidine biosynthesis. Genetic inactivation studies to map the signaling network indicated that both S6K1 and S6K2 transmit growth signals in PTEN-deficient GBM. Kinome-wide ATP binding analysis in inhibitor-treated cells revealed that LY-2584702 directly inhibited S6K1, and substrate phosphorylation studies showed that BMS-777607 inactivation of upstream AXL collaborated to reduce S6K2-mediated signal transduction. Thus, combination targeting of S6K1 and AXL provides a kinase-directed therapeutic approach that circumvents signal transduction redundancy to interrupt metabolic function and reduce growth of PTEN-deficient GBM. Therapy for glioblastoma would be advanced by incorporating molecularly targeted kinase-directed agents, similar to standard of care strategies in other tumor types. Here, we identify a kinase targeting approach to inhibit the metabolism and growth of glioblastoma.