The MYCN proto-oncogene is amplified in a number of advanced-stage human tumors, such as neuroblastomas. Similar to other members of the MYC family of oncoproteins, MYCN (also known as N-Myc) is a transcription factor, and its stability and activity are tightly controlled by ubiquitination-dependent proteasome degradation. Although numerous studies have demonstrated that N-Myc is a driver of neuroblastoma tumorigenesis, therapies that directly suppress N-Myc activity in human tumors are limited. Here we have identified ubiquitin-specific protease 7 (USP7; also known as HAUSP) as a regulator of N-Myc function in neuroblastoma. HAUSP interacts with N-Myc, and HAUSP expression induces deubiquitination and subsequent stabilization of N-Myc. Conversely, RNA interference (RNAi)-mediated knockdown of USP7 in neuroblastoma cancer cell lines, or genetic ablation of Usp7 in the mouse brain, destabilizes N-Myc, which leads to inhibition of N-Myc function. Notably, HAUSP is more abundant in patients with neuroblastoma who have poorer prognosis, and HAUSP expression substantially correlates with N-Myc transcriptional activity. Furthermore, small-molecule inhibitors of HAUSP's deubiquitinase activity markedly suppress the growth of MYCN-amplified human neuroblastoma cell lines in xenograft mouse models. Taken together, our findings demonstrate a crucial role of HAUSP in regulating N-Myc function in vivo and suggest that HAUSP inhibition is a potential therapy for MYCN-amplified tumors.