Appropriate high-dose chemotherapy (HDC) for high-risk neuroblastoma has not yet been established. In Japan, a unique HDC regimen (called double-conditioning regimen) comprising two cycles of total 800 mg/m2 of thiotepa and total 280 mg/m2 of melphalan is widely used. To re-evaluate the safety and the efficacy of this regimen for high-risk neuroblastoma, the medical records of 41 patients with high-risk neuroblastoma who underwent the double-conditioning regimen followed by autologous peripheral blood stem cell rescue between 2002 and 2012 were reviewed. MYCN-amplified high-risk neuroblastomas were observed in 23 patients. All patients underwent intensive multidrug induction chemotherapy, but none underwent anti-GD2 antibody immunotherapy. The primary tumor was resected at the adequate time point. The median follow-up duration for living patients was 9.2 years (range = 5.5–14.0 years). The 5-year event-free survival (EFS) and overall survival (OS) rates from treatment initiation were 41.5% ± 7.7% and 56.1% ± 7.8%, respectively. The 5-year EFS of MYCN-amplified high-risk neuroblastoma patients was 60.9% ± 10.2%, which was significantly superior compared to MYCN-non-amplified high-risk neuroblastoma patients (16.7% ± 8.8%; P < 0.001). MYCN amplification was the most favorable prognostic factor for EFS (hazard ratio = 0.29; 95% confidence interval = 0.12–0.66). Of the 41 patients, 3 died because of regimen-related toxicity (infection, n = 2; microangiopathy, n = 1). The double-conditioning regimen with thiotepa and melphalan is effective for high-risk neuroblastoma, especially in patients with MYCN amplification. However, the double-conditioning regimen is toxic and warrants special attention in clinical practice.