Generation of Cancerous Neural Stem Cells Forming Glial Tumor by Oncogenic Stimulation

Abstract

Neural stem cells in the brain have been shown to be ‘cells of origin’ of certain brain cancers, most notably astrocytomas and medulloblastoma. In particular, in a mouse model, the targeting of genetic modifications for astrocytoma-relevant tumor suppressors to neural stem cells causes malignant astrocytoma to arise, thereby suggesting that astrocytoma is derived from neural stem cells. However, it remains to be determined whether this important finding is reproducible in humans. Herein, we generated cancerous neural stem cells by introducing a set of oncogenes to human fetal neural stem cells (hfNSCs). Serial genetic modification with v-myc for immortalization and consequent H-Ras for oncogenic stimulation with viral gene delivery proved sufficient to induce the transformation of hfNSCs. The resultant F3.Ras cells evidenced a variety of the hallmarks of brain cancer stem cells and most importantly were tumorigenic, forming brain cancers consisting of both a large number of differentiated and a very few undifferentiated populations of cells in an in vivo mouse model. On the contrary, oligodendrocytes derived from the v-myc expressing parent neural stem cells were not transformed by H-Ras, which suggests that neural stem cells may be more susceptible to cancerous transformation by a combination of oncogenes. We also determined that v-myc expressing fetal neural stem cells were defective in p53 response upon the introduction of H-Ras; this finding suggests that an insufficient p53-dependent tumor suppressive mechanism would be associated with high oncogenic susceptibility to H-Ras introduction.

Role of MEK partner-1 in cancer stemness through MEK/ERK pathway in cancerous neural stem cells, expressing EGFRviii

“…Doxycycline (Dox)-inducible EGFRviii was stably established in F3 cells to generate Dox-inducible F3.EGFRviii cells, as described previously [ 10 ]. As shown in Fig.…”

“…F3 cells were maintained as previously described [ 6 , 10 ]. F3.EGFRviii cells were derived from F3 cells and U87 and 293 T cells were purchased from Korean cell line bank ( cellbank.snu.ac.kr /).…”

“…Recently, it has been demonstrated that neural stem cells (NSCs) undergoing neoplastic transformation are responsible for glioma formation [ 7 – 9 ]. NSCs are more susceptible to oncogenic transformation than differentiated glial cells [ 7 , 10 ]. For example, immortalized human fetal NSCs established by the introduction of v-myc (HB.F3 cells) [ 11 ], which are widely used as a model system to examine the possible therapeutic effects of genetically modified NSCs or glial cells [ 12 – 14 ], undergo oncogenic transformation by H-Ras (forming cancerous neural stem cells (CNSCS): F3.Ras cells) [ 10 ] but not Akt [ 15 ].…”

“…For example, immortalized human fetal NSCs established by the introduction of v-myc (HB.F3 cells) [ 11 ], which are widely used as a model system to examine the possible therapeutic effects of genetically modified NSCs or glial cells [ 12 – 14 ], undergo oncogenic transformation by H-Ras (forming cancerous neural stem cells (CNSCS): F3.Ras cells) [ 10 ] but not Akt [ 15 ]. In particular, oligodendrocytes derived from F3 cells are resistant to oncogenic transformation by H-Ras [ 10 ], implying that NSCs are more susceptible to oncogenic transformation than glial cells (e.g., astrocytes and oligodendrocytes). Importantly, F3.Ras cells have similar molecular properties as GSCs [ 6 ], implying that common mechanisms control neural cancer stemness in both GSCs and CNSCs.…”

Genetic modification of human adipose-derived stem cells for promoting wound healing

“…Cells were passaged every 4 days. Retrovirus encoding v-myc gene (courteously provided by Dr. Seung U. Kim, ChungAng University, Korea) and CA-AKT1 genes (courteously provided by Dr. Paul Shapiro, University of Maryland, Baltimore, MD, USA) was produced from 293GPG cells and infected into parental cells with 4 mg/ml polybrene as described elsewhere [22]. CM was prepared from culturing 1 Â 10 6 hASCs or genetically modified hASCs in 10 ml serum-free alpha-MEM media at 37 8C for 3 days and then filtered through a 0.22-mm microfilter (Millipore, Bedford, MA, USA).…”