Paradoxical activation and RAF inhibitor resistance of BRAF protein kinase fusions characterizing pediatric astrocytomas.

Abstract

Astrocytomas are the most common type of brain tumors in children. Activated BRAF protein kinase mutations are characteristic of pediatric astrocytomas with KIAA1549-BRAF fusion genes typifying low-grade astrocytomas and (V600E)BRAF alterations characterizing distinct or higher-grade tumors. Recently, BRAF-targeted therapies, such as vemurafenib, have shown great promise in treating V600E-dependent melanomas. Like (V600E)BRAF, BRAF fusion kinases activate MAPK signaling and are sufficient for malignant transformation; however, here we characterized the distinct mechanisms of action of KIAA1549-BRAF and its differential responsiveness to PLX4720, a first-generation BRAF inhibitor and research analog of vemurafenib. We found that in cells expressing KIAA1549-BRAF, the fusion kinase functions as a homodimer that is resistant to PLX4720 and accordingly is associated with CRAF-independent paradoxical activation of MAPK signaling. Mutagenesis studies demonstrated that KIAA1549-BRAF fusion-mediated signaling is diminished with disruption of the BRAF kinase dimer interface. In addition, the KIAA1549-BRAF fusion displays increased binding affinity to kinase suppressor of RAS (KSR), an RAF relative recently demonstrated to facilitate MEK phosphorylation by BRAF. Despite its resistance to PLX4720, the KIAA1549-BRAF fusion is responsive to a second-generation selective BRAF inhibitor that, unlike vemurafenib, does not induce activation of wild-type BRAF. Our data support the development of targeted treatment paradigms for BRAF-altered pediatric astrocytomas and also demonstrate that therapies must be tailored to the specific mutational context and distinct mechanisms of action of the mutant kinase.

• Authors: Angela J Sievert, Shih-Shan Lang, Katie L Boucher, Peter J Madsen, Erin Slaunwhite, Namrata Choudhari, Meghan Kellet, Phillip B Storm, Adam C Resnick
• Journal: Proceedings of the National Academy of Sciences of the United States of America (Proc Natl Acad Sci U S A) Vol. 110 Issue 15 Pg. 5957-62 (Apr 09 2013) ISSN: 1091-6490 [Electronic] United States
• PMID: 23533272 (Publication Type: Journal Article, Research Support, N.I.H., Extramural, Research Support, Non-U.S. Gov't)

Chemical References

• BRAF-KIAA1549 fusion protein, human
• Enzyme Inhibitors
• Indoles
• Oncogene Proteins, Fusion
• PLX 4720
• Sulfonamides
• Vemurafenib
• BRAF protein, human
• Proto-Oncogene Proteins B-raf

Topics

• Animals
• Astrocytoma (metabolism)
• Cell Line, Tumor
• Cell Transformation, Neoplastic
• Child
• Dimerization
• Enzyme Inhibitors (pharmacology)
• Genetic Vectors
• HEK293 Cells
• Humans
• Indoles (pharmacology)
• Mice
• Mice, Inbred BALB C
• Mutation
• NIH 3T3 Cells
• Neoplasm Transplantation
• Oncogene Proteins, Fusion (metabolism)
• Phenotype
• Protein Interaction Mapping
• Protein Structure, Tertiary
• Proto-Oncogene Proteins B-raf (antagonists & inhibitors, metabolism)
• Signal Transduction
• Sulfonamides (pharmacology)
• Vemurafenib