Oncogenic KRAS promotes malignant brain tumors in zebrafish.

Background: Zebrafish have been used as a vertebrate model to study human cancers such as melanoma, rhabdomyosarcoma, liver cancer, and leukemia as well as for high-throughput screening of small molecules of therapeutic value. However, they are just emerging as a model for human brain tumors, which are among the most devastating and difficult to treat. In this study, we evaluated zebrafish as a brain tumor model by overexpressing a human version of oncogenic KRAS (KRAS(G12V)).

Methods: Using zebrafish cytokeratin 5 (krt5) and glial fibrillary acidic protein (gfap) gene promoters, we activated Ras signaling in the zebrafish central nervous system (CNS) through transient and stable transgenic overexpression. Immunohistochemical analyses were performed to identify activated pathways in the resulting brain tumors. The effects of the MEK inhibitor U0126 on oncogenic KRAS were evaluated.

Results: We demonstrated that transient transgenic expression of KRAS(G12V) in putative neural stem and/or progenitor cells induced brain tumorigenesis. When expressed under the control of the krt5 gene promoter, KRAS(G12V) induced brain tumors in ventricular zones (VZ) at low frequency. The majority of other tumors were composed mostly of spindle and epithelioid cells, reminiscent of malignant peripheral nerve sheath tumors (MPNSTs). In contrast, when expressed under the control of the gfap gene promoter, KRAS(G12V) induced brain tumors in both VZs and brain parenchyma at higher frequency. Immunohistochemical analyses indicated prominent activation of the canonical RAS-RAF-ERK pathway, variable activation of the mTOR pathway, but no activation of the PI3K-AKT pathway. In a krt5-derived stable and inducible transgenic line, expression of oncogenic KRAS resulted in skin hyperplasia, and the MEK inhibitor U0126 effectively suppressed this pro-proliferative effects. In a gfap-derived stable and inducible line, expression of oncogenic KRAS led to significantly increased mitotic index in the spinal cord.

Conclusions: Our studies demonstrate that zebrafish could be explored to study cellular origins and molecular mechanisms of brain tumorigenesis and could also be used as a platform for studying human oncogene function and for discovering oncogenic RAS inhibitors.