Title Targeting PI 3 K in personalized treatment of BRAF-mutated pediatric low-grade gliomas

Purpose: Alteration of the BRAF/MEK/MAPK pathway is the hallmark of pediatric low-grade gliomas (PLGGs), and mTOR activation has been documented in the majority of these tumors. We investigated combinations of MEK1/2, BRAF and mTOR inhibitors in gliomas carrying specific genetic alterations of the MAPK pathway. ExperimentalDesign:Weusedhumanglioma lines containing BRAF (adult high-grade: AM-38, DBTRG, PLGG: BT40), or wild-type BRAF (pediatric high-grade: SF188, SF9427, SF8628) and isogenic systems of KIAA1549:BRAF-expressing NIH/3T3 cells and BRAF-expressing murine brain cells. Signaling inhibitors included everolimus (mTOR), PLX4720 (BRAF), and AZD6244 (MEK1/2). Proliferation was determined using ATP-based assays. In vivo inhibitor activities were assessed in the BT40 PLGG xenograft model. Results: In BRAF cells, the three possible doublet combinations of AZD6244, everolimus, and PLX4720 exhibited significantly greater effects on cell viability. In BRAF cells, everolimus þ AZD6244 was superior compared with respective monotherapies. Similar results were found using isogenic murine cells. In KIAA1549:BRAF cells, MEK1/2 inhibition reduced cell viability and S-phase content, effects that were modestly augmented by mTOR inhibition. In vivo experiments in the BRAF pediatric xenograft model BT40 showed the greatest survival advantage in mice treated with AZD6244 þ PLX4720 (P < 0.01). Conclusions: In BRAF tumors, combination of AZD6244 þ PLX4720 is superior to monotherapy and to other combinatorial approaches. In BRAF pediatric gliomas, everolimus þ AZD6244 is superior to either agent alone. KIAA1549:BRAFexpressing tumors display marked sensitivity to MEK1/2 inhibition. Application of these results to PLGG treatment must be exercised with caution because the dearth of PLGG models necessitated only a single patient-derived PLGG (BT40) in this study. Clin Cancer Res; 22(21); 5312–21. 2016 AACR.

[1]  T. Golub,et al.  Expression profiles of 151 pediatric low-grade gliomas reveal molecular differences associated with location and histological subtype. , 2015, Neuro-oncology.

[2]  K. Kurian,et al.  Current Understanding of BRAF Alterations in Diagnosis, Prognosis, and Therapeutic Targeting in Pediatric Low-Grade Gliomas , 2015, Front. Oncol..

[3]  Daniel J. Guillaume,et al.  Pilomyxoid astrocytoma treated successfully with vemurafenib , 2014, Pediatric blood & cancer.

[4]  David T. W. Jones,et al.  Phase II study of sorafenib in children with recurrent or progressive low-grade astrocytomas. , 2014, Neuro-oncology.

[5]  P. Varlet,et al.  Vemurafenib in pediatric patients with BRAFV600E mutated high‐grade gliomas , 2014, Pediatric blood & cancer.

[6]  A. Gajjar,et al.  Complete clinical regression of a BRAF V600E-mutant pediatric glioblastoma multiforme after BRAF inhibitor therapy , 2014, BMC Cancer.

[7]  M. Berger,et al.  Targeting Wee1 for the treatment of pediatric high-grade gliomas. , 2014, Neuro-oncology.

[8]  A. Resnick,et al.  Paradoxical activation and RAF inhibitor resistance of BRAF protein kinase fusions characterizing pediatric astrocytomas , 2013, Proceedings of the National Academy of Sciences.

[9]  D. Gutmann,et al.  Pediatric glioma-associated KIAA1549:BRAF expression regulates neuroglial cell growth in a cell type-specific and mTOR-dependent manner. , 2012, Genes & development.

[10]  K. Flaherty,et al.  Combined BRAF and MEK inhibition in melanoma with BRAF V600 mutations. , 2012, The New England journal of medicine.

[11]  M. Berger,et al.  PTEN promoter methylation and activation of the PI3K/Akt/mTOR pathway in pediatric gliomas and influence on clinical outcome. , 2012, Neuro-oncology.

[12]  Rebecca A. Ihrie,et al.  Cooperative interactions of BRAFV600E kinase and CDKN2A locus deficiency in pediatric malignant astrocytoma as a basis for rational therapy , 2012, Proceedings of the National Academy of Sciences.

[13]  S. Pfister,et al.  Functional characterization of a BRAF insertion mutant associated with pilocytic astrocytoma , 2011, International journal of cancer.

[14]  Tom Misteli,et al.  RAF inhibitor resistance is mediated by dimerization of aberrantly spliced BRAF(V600E) , 2011, Nature.

[15]  Noemi Andor,et al.  Asymmetry-defective oligodendrocyte progenitors are glioma precursors. , 2011, Cancer cell.

[16]  N. Alon,et al.  BRAF-KIAA1549 Fusion Predicts Better Clinical Outcome in Pediatric Low-Grade Astrocytoma , 2011, Clinical Cancer Research.

[17]  A. Iafrate,et al.  BRAF V600E Mutations Are Common in Pleomorphic Xanthoastrocytoma: Diagnostic and Therapeutic Implications , 2011, PloS one.

[18]  Kirsten Schmieder,et al.  Analysis of BRAF V600E mutation in 1,320 nervous system tumors reveals high mutation frequencies in pleomorphic xanthoastrocytoma, ganglioglioma and extra-cerebellar pilocytic astrocytoma , 2011, Acta Neuropathologica.

[19]  S. Nelson,et al.  Melanomas acquire resistance to B-RAF(V600E) inhibition by RTK or N-RAS upregulation , 2010, Nature.

[20]  E. Peng,et al.  Rapamycin regulates Akt and ERK phosphorylation through mTORC1 and mTORC2 signaling pathways , 2010, Molecular carcinogenesis.

[21]  M. Belvin,et al.  RAF inhibitors prime wild-type RAF to activate the MAPK pathway and enhance growth , 2010, Nature.

[22]  Chao Zhang,et al.  RAF inhibitors transactivate RAF dimers and ERK signaling in cells with wild-type BRAF , 2010, Nature.

[23]  J. Reis-Filho,et al.  Kinase-Dead BRAF and Oncogenic RAS Cooperate to Drive Tumor Progression through CRAF , 2010, Cell.

[24]  Hanlee P. Ji,et al.  Oncogenic BRAF mutation with CDKN2A inactivation is characteristic of a subset of pediatric malignant astrocytomas. , 2010, Cancer research.

[25]  Keith L. Ligon,et al.  Profiling Critical Cancer Gene Mutations in Clinical Tumor Samples , 2009, PloS one.

[26]  T. Shaikh,et al.  Duplication of 7q34 in Pediatric Low‐Grade Astrocytomas Detected by High‐Density Single‐Nucleotide Polymorphism‐Based Genotype Arrays Results in a Novel BRAF Fusion Gene , 2009, Brain pathology.

[27]  P. Pandolfi,et al.  Inhibition of mTORC1 leads to MAPK pathway activation through a PI3K-dependent feedback loop in human cancer. , 2008, The Journal of clinical investigation.

[28]  G. Reifenberger,et al.  BRAF gene duplication constitutes a mechanism of MAPK pathway activation in low-grade astrocytomas. , 2008, The Journal of clinical investigation.

[29]  Abraham Nudelman,et al.  In vivo efficacy of a novel histone deacetylase inhibitor in combination with radiation for the treatment of gliomas. , 2007, Neuro-oncology.

[30]  Todd R. Golub,et al.  BRAF mutation predicts sensitivity to MEK inhibition , 2006, Nature.

[31]  D. Gutmann,et al.  Proteomic analysis reveals hyperactivation of the mammalian target of rapamycin pathway in neurofibromatosis 1-associated human and mouse brain tumors. , 2005, Cancer research.

[32]  A. Nicholson,et al.  Mutations of the BRAF gene in human cancer , 2002, Nature.

[33]  G. Reifenberger,et al.  The WHO Classification of Tumors of the Nervous System , 2002, Journal of neuropathology and experimental neurology.

[34]  J. Blair,et al.  The role of surgery in optic pathway/hypothalamic gliomas in children. , 2014, Journal of neurosurgery. Pediatrics.

[35]  I. Pollack,et al.  Association of molecular alterations, including BRAF, with biology and outcome in pilocytic astrocytomas , 2009, Acta Neuropathologica.

[36]  T. Chou,et al.  Quantitative analysis of dose-effect relationships: the combined effects of multiple drugs or enzyme inhibitors. , 1984, Advances in enzyme regulation.