Neuroblastoma (NB) is a common childhood cancer, accounting for 40% of cancer diagnoses in infants under one year of age. Despite its prevalence, the exact origin of NB remains incompletely understood. However, it is believed to arise from the aberrant differentiation of neural crest cells (NCCs), a multipotent, stem-cell-like population with limited self-renewal capacity. During development NC- derived cells respond to precise developmental cues, navigate specific developmental trajectories, overcome developmental bottlenecks and ultimately commit to defined cell fates. This process is tightly regulated by transcription factors (TFs), with microRNAs (miRNAs) acting as essential post-transcriptional regulators that modulate TF output. In turn, TFs often control the transcription of miRNAs, creating intricate feedback loops that fine-tune gene expression programs essential for cellular development and differentiation. In NB, this regulatory balance is disrupted, with aberrant miRNA expression likely leading to the loss of normal TF-mediated control over neural crest cell maturation. Using extensive preliminary data from an in vitro differentiation model, our lab has identified a critical developmental "bottleneck" in the differentiation of NCCs to mature sympathetic neurons. We have pinpointed a series of candidate TFs and miRNAs that are regulated immediately before, during, and after this bottleneck, suggesting they may be crucial in controlling the developmental trajectory of these cells. While some of these candidates are established players in neuronal development, others are novel and uncharacterized. Importantly, several of these TFs and miRNAs show altered expression in NB patient cohorts, although their cancer-promoting roles are yet to be reported. By experimentally modifying miRNA and TFs levels, we aim to observe and analyse the resultant cellular responses in NB. Ultimately, this research seeks to confirm the significant role of miRNAs and TFs in the development of sympathetic nervous system and evaluate their prospects as biomarkers or therapeutic agents in the fight against NB.