ABSTRACT
The role of PPM1D mutations in de novo gliomagenesis has not been systematically explored. Here we analyze whole genome sequences of 170 pediatric high-grade gliomas and find that truncating mutations in PPM1D that increase the stability of its phosphatase are clonal driver events in 11% of Diffuse Midline Gliomas (DMGs) and are enriched in primary pontine tumors. Through the development of DMG mouse models, we show that PPM1D mutations potentiate gliomagenesis and that PPM1D phosphatase activity is required for in vivo oncogenesis. Finally, we apply integrative phosphoproteomic and functional genomics assays and find that oncogenic effects of PPM1D truncation converge on regulators of cell cycle, DNA damage response, and p53 pathways, revealing therapeutic vulnerabilities including MDM2 inhibition.
Subject(s)
Glioma/genetics , Mutation , Oncogenes/genetics , Protein Phosphatase 2C/genetics , Adolescent , Adult , Animals , Brain Stem Neoplasms/genetics , Carcinogenesis/genetics , Cell Cycle , Child , Child, Preschool , DNA Damage , Disease Models, Animal , Female , HEK293 Cells , Humans , Infant , Male , Mice , Proto-Oncogene Proteins c-mdm2 , Transcriptome , Tumor Suppressor Protein p53/genetics , Young AdultABSTRACT
Low-dose methotrexate (MTX) is a first-line therapy for the treatment of arthritis. However, there is considerable interindividual variability in MTX exposure following standard dosing. Polymorphisms in SLCO1B1 significantly effect MTX clearance, altering therapeutic response. One decreased function variant, rs4149056 (c.521T>C, Val174Ala), slows MTX clearance and in vitro uptake of MTX. This phenotype was recapitulated in a mouse model using a knockout (KO) of the murine orthologue, Slco1b2. Our objective was to investigate the impact of this phenotype on the pharmacokinetics and therapeutic outcomes of low-dose MTX in a murine model of collagen-induced arthritis (CIA). We evaluated response to MTX in mice with CIA using wildtype (WT), heterozygous, and KO Slco1b2 mice on a DBA1/J background. Arthritis was macroscopically evaluated daily to quantify disease progression. Mice received 2 mg/kg or a pharmacogenetically guided MTX dose subcutaneously 3 times a week for 2 weeks. MTX concentrations were collected at the end of the study and exposure (day*µM) was estimated using a two-compartment model. Mice displayed a seven-fold range in MTX exposure and revealed a significant exposure-response relationship (p = 0.0027). KO mice receiving the 2 mg/kg dosing regimen had 2.3-fold greater exposure to MTX (p < 0.0001) and a 66% reduction in overall disease progression (p = 0.011) compared to WT mice. However, exposure and response were equivalent when pharmacogenetically guided dosing was used. These studies demonstrate that an exposure-response relationship exists for MTX and that Slco1b2 genotype affects MTX exposure and therapeutic response. Such evidence supports the use of SLCO1B1-pharmacogenetic dosing of low-dose MTX for patients with arthritis.
Subject(s)
Antirheumatic Agents/administration & dosage , Arthritis/drug therapy , Liver-Specific Organic Anion Transporter 1 , Methotrexate/administration & dosage , Mice, Knockout , Pharmacogenomic Testing , Animals , Antirheumatic Agents/pharmacology , Collagen/drug effects , Collagen/metabolism , Genotype , Humans , Male , Methotrexate/pharmacology , Mice , PharmacogeneticsABSTRACT
BACKGROUND: Diffuse intrinsic pontine gliomas (DIPGs) are highly lethal childhood brain tumors. Their unique genetic makeup, pathological heterogeneity, and brainstem location all present challenges to treatment. Developing mouse models that accurately reflect each of these distinct features will be critical to advance our understanding of DIPG development, progression, and therapeutic resistance. The aims of this study were to generate new mouse models of DIPG and characterize the role of specific oncogenic combinations in DIPG pathogenesis. METHODS: We used in utero electroporation (IUE) to transfect neural stem cells in the developing brainstem with PiggyBac DNA transposon plasmids. Combinations of platelet-derived growth factor B (PDGFB), PdgfraD842V, or PdgfraWT, combined with dominant negative Trp53 (DNp53) and H3.3K27M expression, induced fully penetrant brainstem gliomas. RESULTS: IUE enabled the targeted transfection of brainstem neural stem cells. PDGFB + DNp53 + H3.3K27M induced the rapid development of grade IV gliomas. PdgfraD842V + DNp53 + H3.3K27M produced slower forming grade III gliomas. PdgfraWT + DNp53 + H3.3K27M produced high- and low-grade gliomas with extended latencies. PDGFB, PdgfraD842V, and PdgfraWT DIPG models display unique histopathological and molecular features found in human DIPGs. H3.3K27M induced both overlapping and unique gene expression changes in PDGFB and PdgfraD842V tumors. Paracrine effects of PDGFB promote disruption of pericyte-endothelial interactions and angiogenesis in PDGFB DIPG mouse models. CONCLUSION: Brainstem-targeted IUE provides a rapid and flexible system to generate diverse DIPG mouse models. Using IUE to investigate mutation and pathohistological heterogeneity of DIPG will provide a valuable tool for future genetic and preclinical studies.
