De novo Induction of Genetically Engineered Brain Tumors in Mice Using Plasmid DNA

Abstract

Spontaneous mouse models of cancer show promise to more accurately recapitulate human disease and predict clinical efficacy. Transgenic mice or viral vectors have been required to generate spontaneous models of glioma, a lethal brain tumor, because nonviral gene transfer is typically transient. To overcome this constraint, we used the Sleeping Beauty transposable element to achieve chromosomal integration of human oncogenes into endogenous brain cells of immunocompetent mice. Genetically engineered, spontaneous brain tumors were induced with plasmid DNA in a matter of weeks in three separate mouse strains. The phenotype of tumors was influenced by the combination of oncogenes delivered, resembling human astrocytoma or glioblastoma in the majority of cases. At least five different genes can be cotransfected simultaneously including reporters, allowing measurement of tumor viability by in vivo imaging. This model can accelerate brain tumor research in a variety of ways such as generation of ''humanized'' models for high throughput drug screening and candidate gene validation with exceptional speed and flexibility. [Cancer Res 2009;69(2):431-9]

Lentiviral-Induced High-Grade Gliomas in Rats: The Effects of PDGFB, HRAS-G12V, AKT, and IDH1-R132H

“…In order to reach these practical goals for de novo glioma modeling in mice, genetic alterations decreasing the efficacy of the TP53 and RB tumor suppressor genes have been necessary [16,17,23,25,27,28]. In rats, our data confirm the results of Assanah et al [18], suggesting that the need for p53 inactivation in glioma may be speciesdependent, and that the manipulation of the RTK/RAS/ PI(3)K signaling pathway is sufficient to create gliomas in rats.…”

“…For example, the Sleeping Beauty Transposon system has been used to stably incorporate oncogenes encoded on nonviral plasmids into the brains of newborn mice [16]. Other systems previously used to create endogenous gliomas include the RCAS/tv-a system in mice [17], retroviral vectors in rats and mice [18], and lentiviral vectors in mice [19].…”

Oncogenic ras-driven cancer cell vesiculation leads to emission of double-stranded DNA capable of interacting with target cells

“…The following cell lines were used: IEC-18 -non-tumourigenic, immortalized rat intestinal epithelial cell line; RAS-3 -tumourigenic variant of IEC-18 cells transfected with the V12 mutant c-H-ras human oncogene; Spontaneous primitive glioblastomalike brain tumour (PBT) cell lines -generated by integrating N-ras and SV40LT expression vectors into neonatal mouse brain using the sleeping beauty transposase, as described [22]. PBTs were generated in either wild type C57BL6 mice or in their strains harbouring null mutation of the acidic sphingomyelinase gene (ASMase-/-), or in heterozygous littermates (ASMase+/-), from which cell lines were established; RAT-1 -immortalized, non-tumourigenic rat fibroblasts.…”

“…To explore the mechanisms of vesicular DNA emission, we generated primitive brain tumour (PBT) cell lines harbouring N-ras and SV40LT oncogenes in the wild type or ASMase-deficient background [22]. These cells are deficient for both Rb/TP53 activity (due to SV40LT expression [27]) and ASMase (due to targeted insertion of neomycin expressing cassette into the gene [28]), the changes affecting both known vesiculation pathways of exosomal and ectosomal biogenesis, respectively [24,26].…”

The CXCL12/CXCR4 pathway or the autocrine proliferative loop of the glioblastoma stem cells

“…Indeed, the authors induced brain tumors in a model recently described and based on a plasmid which allow a knock-in of immature brain cells using a Sleeping Beauty transposase (9). The inserted genes are the simian virus 40 large T antigen (SV40-LgT) and a constitutively active human NRAS oncogene (NRAS).…”