High-risk neuroblastoma (NBL) is a leading cause of pediatric cancer death. The evolution of the tumor microenvironment under therapy remains obscure. In this study, we longitudinally profiled 22 high-risk NBL patients before and after induction chemotherapy through single-nucleus RNA and ATAC sequencing and whole-genome sequencing. We identified profound shifts in tumor and immune cell subpopulations and nominated enhancer-driven transcriptional regulators of NBL neoplastic states. Reduced proliferating and expanded metabolically active neoplastic states predicted a poorer diagnostic outcome, whereas expanded and more differentiated neuronal-like states were linked to a more favorable prognosis. Mesenchymal neoplastic cells increased and correlated with a poorer response to chemotherapy. Macrophages significantly expanded towards pro-angiogenic, immunosuppressive, and metabolic phenotypes. We identified paracrine signaling networks and established the HB-EGF/ERBB4 axis between macrophage and neoplastic subsets which promoted tumor growth through the induction of ERK signaling. These findings collectively reveal novel intrinsic and extrinsic regulators of therapy response in high-risk NBL.