Detection of EWS chimeric transcripts by nested RT-PCR to allow reinfusion of uncontaminated peripheral blood stem cells in high-risk Ewing's tumor in childhood

Abstract

BACKGROUND AND OBJECTIVE: Ewing's tumors (ET) are primary malignancies of bone and soft tissues characterized in at least 96% of cases by specific fusion transcripts originating from recurrent chromosomal translocations. Clinical protocols for high-risk metastatic ETs include high-dose radiation/chemotherapy followed by autologous peripheral blood stem cell (PBSC) reinfusion. We used nested reverse transcriptase polymerase chain reaction (RT-PCR) to search for the presence of ET-specific transcripts in PBSC collections from patients with high-risk ET in order to collect harvests free from neoplastic cells but still sufficient to obtain early stable engraftment. DESIGN AND METHODS: Thirty-seven harvest samples from 15 ET patients treated with mobilizing chemotherapy were analyzed. Nested RT-PCR was performed to detect ET-specific transcripts in RNA extracted from the PBSC collections. RESULTS: A total of 30 harvests was performed. On average, 2 harvests (range 1-4) were sufficient to collect the minimum required number of mononuclear cells (2.5