Generation of model mice for brain tumors using conditional knockout technique

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The purpose of this research is generating model mice for brain tumors, using conditional knockout technique to destroy the functions of tumor-suppressor genes selectively in brain cells. We produced five lines of genetically modified mice, by use of the homologous recombination vector DNAs and mouse ES cell line RENKA. The five lines included 2 kinds of "flox" mice in which frame-shift exon of tumor-suppressor gene Ptc or Nf1 was flanked with recombination target sequence loxP, and 2 kinds of "knock-in" mice in which Cre recombinase was introduced in the translation initiation site of Nestin or Tubb3 gene, marker genes for neural cells. We also produced Nf1-flox; P53-null mice in which a tumor suppressor gene P53 located nearly in Nf1 gene was destroyed. Crossbreeding of the "flox" mice and "knock-in" mice successfully generated the mice in which tumor suppressor gene was destroyed selectively in brain cells by the Cre-loxP recombination system.Among the mice obtained, brain tumors which arose from the cells in cerebellar cortex were observed with frequency of 50% (2/4)in Ptc-flox;Tubb3-Cre hetero mice before 5 months of age. The tumor cells consisted of undifferentiated cells which hardly expressed mature marker proteins and resembled those of human medulloblastomas in histology. On the other hand, no brain tumor was observed in Nf1-flox;Neatin-Cre hetero nor Nf1-flox;P53-null;Neatin-Cre hetero mice within 6 months of age, suggesting the necessity of generating homo knockout mice for the Nf1 and P53 gene.These results indicate that mutation of Ptc gene is one of the causative events of medulloblastoma, and, therefore, we were able to contribute to clarification of etiology of the medulloblastoma. In addition, the Ptc-flox;Tubb3-Cre hetero mice may work as model mice of medulloblastoma which is useful to develop new therapeutic drugs or protocols.