Neuroblastoma (NB), an embryonal tumor derived from the neural crest, originates from sympathetic nerve system, manifests as thoracic, paraspinous or abdominal tumors, and metastases to bone in high-risk cases. Although NB stands as the most common solid tumor in early childhood and accounts for about 15 % of total pediatric cancer death, there has been limited success in searching for novel therapeutic regimen for this lethal disease during the past two decades. Numerous epidemiological and clinical studies have pinpointed anaplastic lymphoma kinase (ALK) and MYCN as potent governors for NB malignant behavior. ALK and MYCN amplification and constitutive active mutations are common in high-risk NB patients. However, there is still lack of evidence showing that a small molecule compound could simultaneously inhibits ALK and MYCN and plays strong negative regulatory roles in NB. Here, we showed that 17-DMAG, a well-known HSP-90 inhibitor, significantly inhibits NB cell growth, arrests cell cycle, and strongly induces NB cell apoptosis. Interestingly, our data suggests that NB cells with both ALK and MYCN amplification/mutation are more sensitive to 17-DMAG treatment, while NB cells with only ALK or MYCN amplification are less sensitive and NB cells without ALK or MYCN amplification/mutation are least sensitive. Moreover, we also found that knocking down ALK and MYCN additively inhibits NB cell growth and that transduction of MYCN largely abolished the ALK-dependent NB cell growth, indicating that there is a cross-talk between MYCN and ALK signaling machinery. Our results provide proof-of-principle that 17-DMAG strongly inhibits NB cell growth by targeting both ALK and MYCN. Our findings might shed a light for further investigation of novel small molecule compound which is safe and exerts similar strong effects in vivo as novel approaches for management of high-risk NB. © 2013 International Society of Oncology and BioMarkers (ISOBM).