Atrt-09. combination of the pi3k inhibitor paxalisib with the nucleoside analog gemcitabine activates the integrated stress response and extends survival in orthotopic models of atypical teratoid/rhabdoid tumors

Abstract Relapsed atypical teratoid/rhabdoid tumors (AT/RT) are a deadly disease without a known cure. The Pacific Pediatric Neuro-Oncology Consortium (PNOC) is looking for effective combination therapies for an upcoming clinical trial to better treat relapsed AT/RT. We have previously identified that AT/RT are dependent on activation of mTORC1 and mTORC2 for growth and survival. Paxalisib is a highly brain penetrant PI3k inhibitor which acts upstream of mTOR to simultaneously inhibit mTORC1/2. In murine orthotopic xenograft models of AT/RT, Paxalisib significantly extends survival as a single agent therapy (CHLA-06: 40 to 54 days, p=0.001; BT12: 21 to 35 days, p=0.02). However, to improve the durability of this single agent therapy, we evaluated the efficacy of rational therapeutic partners. Analysis of RNAseq following mTOR inhibition (mTORi) showed an upregulation of genes associated with the integrated stress response (ISR) (ATF4, CHOP, PPP1R15A; t-test p<0.05). Gemcitabine is a pyrimidine nucleoside prodrug, which inhibits DNA synthesis and induces DNA damage, also leading to ISR activation. While the ISR can be protective, intense or prolonged activation leads to cell death. We therefore hypothesized that Paxalisib would combine with Gemcitabine to over-activate the ISR and drive AT/RT cell death. Through protein analyses, we demonstrate that combination therapy further activates the ISR compared to DMSO control (Western Blot: phospho-eIF2α, ATF4, and CHOP). Combination therapy also increases apoptosis (Western blot cPARP, MUSE Annexin V Assay) and synergizes to decrease AT/RT cell growth (SynergyFinder BLISS score CHLA-06: 16.8, BT12: 14.3, CHLA-266: 13.9). In mice bearing CHLA-06 orthotopic tumors, Paxalisib/Gemcitabine combination therapy slows tumor growth (Bioluminescent imaging) and extends median survival from 22 to 82.5 days (p<0.0001) compared to vehicle control treated mice. We aim to translate these results in this upcoming PNOC clinical trial to treat relapsed AT/RT.