Development of zebrafish medulloblastoma-like PNET model by TALEN-mediated somatic gene inactivation.

Jaegal Shim,Jung-Hwa Choi,Hyena Kim,Cheol-Hee Kim,Ji Eun Lee,Dongwan Hong,Jeong-Soo Lee,Sunshin Kim,Jong Hwan Kim,Moon-Hak Park,Seon-Young Kim,Young-Ki Bae

doi:10.18632/oncotarget.19424

Abstract

Genetically engineered animal tumor models have traditionally been generated by the gain of single or multiple oncogenes or the loss of tumor suppressor genes; however, the development of live animal models has been difficult given that cancer phenotypes are generally induced by somatic mutation rather than by germline genetic inactivation. In this study, we developed somatically mutated tumor models using TALEN-mediated somatic gene inactivation of cdkn2a/b or rb1 tumor suppressor genes in zebrafish. One-cell stage injection of cdkn2a/b-TALEN mRNA resulted in malignant peripheral nerve sheath tumors with high frequency (about 39%) and early onset (about 35 weeks of age) in F0 tp53e7/e7 mutant zebrafish. Injection of rb1-TALEN mRNA also led to the formation of brain tumors at high frequency (58%, 31 weeks of age) in F0 tp53e7/e7 mutant zebrafish. Analysis of each tumor induced by somatic inactivation showed that the targeted genes had bi-allelic mutations. Tumors induced by rb1 somatic inactivation were characterized as medulloblastoma-like primitive neuroectodermal tumors based on incidence location, histopathological features, and immunohistochemical tests. In addition, 3′ mRNA Quanti-Seq analysis showed differential activation of genes involved in cell cycle, DNA replication, and protein synthesis; especially, genes involved in neuronal development were up-regulated.

Highlights

Medulloblastoma (MB) is the most common type of malignant solid tumors in the pediatric brain, which accounts for 20% of all pediatric tumors
Using RNA sequencing analysis together with histopathology and immunohistochemistry, we have demonstrated that brain tumors induced by rb1 somatic inactivation have a molecular feature of MB-like primitive neuroectodermal tumors (PNETs)
Somatic inactivation of cdkn2a/b gene by the injection of Transcription activatorlike effector nucleases (TALEN) mRNA leads to Malignant Peripheral Nerve Sheath Tumors (MPNSTs) in F0 founder tp53 e7/e7 mutant zebrafish

Summary

Introduction

Medulloblastoma (MB) is the most common type of malignant solid tumors in the pediatric brain, which accounts for 20% of all pediatric tumors. It occurs in the external granular layer (EGL) of the cerebellum and may undergo metastasis (reviewed in [1]). The WNT subgroup accounts for ~10% of all MB and usually has good prognosis; this subgroup involves mutations in genes including TP53, CTNNB1, AXIN1, SMARCA4, and CREBBP. The SHH subgroup accounts for up to 30% of MB and contrarily shows poor prognosis. Group 3 accounts for ~25% of all MB and carries no TP53 mutations while showing poor prognosis, immature histopathology, and highly metastatic property. Group 4 accounts for ~35% of all MB; similar to Group 3, Group 4 carries no TP53 mutations while mutations are often identified in genes including OTX2, N-MYC, FST, and CDK6

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Conclusion