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DeWitt, John

Neuroscience Graduate Program

2012

PhD

Neuroblastoma is the most common pediatric cancer in infants, arising from the sympathoadrenallineage of the neural crest. Despite recent advances in other pediatric cancers, long term survival in high risk cases ofneuroblastoma remains below 40%. Therefore, to develop successful therapeutics targeting high risk tumors, further research into the mechanisms involved in high risk tumor formation is necessary. Prognosis in neuroblastoma is determined by a number of factors, including certain genetic and biological variables. The genetic variable correlated most with high risk disease is amplification of the MYCN gene, which is present in ~25% of tumors. Additionally, ~70% of these MYCN-amplified tumors express the neurotrophin receptor TrkB, and its ligand, brain-derived neurotrophic factor (BDNF), with concurrent expression of these proteins correlated with high risk disease independent of MYCN-amplification status. To better understand factors influencing MYCN-amplified tumor cell phenotype, and the role of TrkB signaling in high risk neuroblastoma, the experiments in this dissertation examined growth factor effects on MYCN-amplified tumor cells from the TH-MYCN mouse model ofneuroblastoma, as well as the creation, and expression of a constitutively active TrkB receptor in a neural crest derived cell line.
In addition to being significantly correlated with a poor prognosis in neuroblastoma, the presence of activated TrkB signaling promotes a more aggressive phenotype in established neuroblastoma cell lines. However, whether TrkB signaling is sufficient to transform neural crest derived cells had not been established. To determine the role of TrkB signaling in malignant transformation, the two immunoglobulin-like (rg) ligand binding domains were removed from a full length rat TrkB receptor. Expression of this receptor, termed [delta]IgTrkB, leads to elevated phosphorylated Erk levels in the absence of ligand, indicating the receptor is constitutively active. When expressed in the neural crest derived cell line NCM-1, constitutive TrkB signaling confers a highly transfonned phenotype characterized by enhanced proliferation, anchorage-independent cell growth, anoikis resistance, and matrix invasion. Furthermore, [delta]IgTrkB NCM-1 cells upregulate transcripts for a number of cancer promoting genes, in addition to the poor prognosis marker MYCN. In vivo, [delta]IgTrkB NCM-1 cells form highly aggressive tumors, requiring euthanasia ofmice by 15 days following injection, while wild type cells fail to grow. Thus, TrkB signaling is sufficient to transfonn cells derived from the neural crest.