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A Protocol for Rapid Post-mortem Cell Culture of Diffuse Intrinsic Pontine Glioma DIPG
Published on: March 7, 2017
PI3K/mTOR is a therapeutically targetable genetic dependency in diffuse intrinsic pontine glioma
Ryan J Duchatel1,2,3, Evangeline R Jackson1,2,3, Sarah G Parackal4,5
1Cancer Signalling Research Group, School of Biomedical Sciences and Pharmacy, College of Health, Medicine and Wellbeing, University of Newcastle, Callaghan, New South Wales, Australia.
Abstract:
Diffuse midline glioma (DMG), including tumors diagnosed in the brainstem (diffuse intrinsic pontine glioma; DIPG), are uniformly fatal brain tumors that lack effective treatment. Analysis of CRISPR/Cas9 loss-of-function gene deletion screens identified PIK3CA and MTOR as targetable molecular dependencies across patient derived models of DIPG, highlighting the therapeutic potential of the blood-brain barrier-penetrant PI3K/Akt/mTOR inhibitor, paxalisib. At the human-equivalent maximum tolerated dose, mice treated with paxalisib experienced systemic glucose feedback and increased insulin levels commensurate with patients using PI3K inhibitors. To exploit genetic dependence and overcome resistance while maintaining compliance and therapeutic benefit, we combined paxalisib with the antihyperglycemic drug metformin. Metformin restored glucose homeostasis and decreased phosphorylation of the insulin receptor in vivo, a common mechanism of PI3K-inhibitor resistance, extending survival of orthotopic models. DIPG models treated with paxalisib increased calcium-activated PKC signaling. The brain penetrant PKC inhibitor enzastaurin, in combination with paxalisib, synergistically extended the survival of multiple orthotopic patient-derived and immunocompetent syngeneic allograft models; benefits potentiated in combination with metformin and standard-of-care radiotherapy. Therapeutic adaptation was assessed using spatial transcriptomics and ATAC-Seq, identifying changes in myelination and tumor immune microenvironment crosstalk. Collectively, this study has identified what we believe to be a clinically relevant DIPG therapeutic combinational strategy.
Insights
Researchers explored new treatments for diffuse midline glioma (DMG), including diffuse intrinsic pontine glioma (DIPG). Combining paxalisib, metformin, and enzastaurin showed promising results in extending survival for these fatal brain tumors.
Area of Science:
- Neuro-oncology
- Cancer Therapeutics
- Molecular Biology
Background:
- Diffuse midline glioma (DMG), encompassing diffuse intrinsic pontine glioma (DIPG), represents a uniformly fatal brain tumor category with limited therapeutic options.
- CRISPR/Cas9 screens identified PIK3CA and MTOR as key molecular dependencies in DIPG patient-derived models.
- Paxalisib, a PI3K/Akt/mTOR inhibitor that penetrates the blood-brain barrier, emerged as a potential therapeutic agent.
Purpose of the Study:
- To identify and validate effective combination therapies for DIPG by targeting molecular dependencies and overcoming resistance mechanisms.
- To evaluate the efficacy of combining the PI3K/Akt/mTOR inhibitor paxalisib with metformin and enzastaurin, with or without radiotherapy, in preclinical DIPG models.
Main Methods:
- Utilized CRISPR/Cas9 loss-of-function screens to identify targetable genes in DIPG models.
- Administered paxalisib, metformin, and enzastaurin in various combinations to orthotopic patient-derived and syngeneic DIPG models in mice.
- Assessed therapeutic efficacy through survival studies and analyzed molecular adaptations using spatial transcriptomics and ATAC-Seq.
Main Results:
- Paxalisib treatment led to systemic glucose feedback and increased insulin levels, indicating PI3K inhibitor-associated metabolic changes.
- Combination therapy with paxalisib and metformin restored glucose homeostasis and reduced insulin receptor phosphorylation, overcoming a resistance mechanism and extending survival.
- The addition of enzastaurin synergistically enhanced survival, with further potentiation observed when combined with metformin and radiotherapy.
- Genomic analyses revealed alterations in myelination and tumor immune microenvironment crosstalk in response to treatment.
Conclusions:
- The combination of paxalisib, metformin, and enzastaurin represents a promising, clinically relevant therapeutic strategy for DIPG.
- Targeting PI3K/Akt/mTOR pathway dependencies, metabolic feedback loops, and PKC signaling offers a multi-pronged approach to combat DIPG.
- Further investigation into these combinatorial therapies is warranted to improve outcomes for patients with diffuse midline glioma.
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