Evaluating neurite outgrowth and signal integration in response to NGF and GDNF in neuroblastoma cell lines

Neuroblastoma is a cancerous tumor that develops from cells of the neural crest. Unfortunately, the highly migratory nature of these cells causes approximately 70% of neuroblastomas to have already metastasized by the time they are diagnosed. Primarily affecting infants and children, neuroblastoma accounts for 15% of pediatric cancer fatalities. This illustrates the need for a better understanding of the mechanism by which neural cells transform into cancer. We hypothesize that neural crest cells’ failure to differentiate into nerve cells is an important step leading to neuroblastoma. The roles of receptor tyrosine kinases (RTKs) in governing cellular processes like differentiation are extensive and dynamic. Defects in RTK pathways have been linked to various cancers including neuroblastoma. While identifying each pathway involved in differentiation is important, it is as important to understand how the pathways interact; this has posed a challenge to researchers. To model RTK pathway associations, Dr. Mark Grimes has pioneered computer methods that incorporate bioinformatics and pattern recognition analysis to neuroblastoma cell line phosphoproteomic data sets. Along with finding 31 of the 58 RTKs in the human genome present and activated in neuroblastoma cells, his mapping of associations between tyrosine kinase signaling pathways has identified several networks of RTKs that represent functional cohorts. In my study, I focused on two RTKs that cause differentiation in neuroblastoma cells that were from within the same cluster or cohort: TrkA and RET. TrkA and RET were activated with their respective ligands, NGF and GDNF, both individually and together, to analyze the effects of co-stimulation. Quantitative analysis of neurite outgrowth and activation of downstream effectors derived from western blots was used to determine if these two pathways act synergistically. Improving the current understanding of interactions among RTK pathways will help us to better understand cell differentiation and how it goes awry in neuroblastoma