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

Background and Objectives. Ewing's tumors (ET) are primary malignancies of bone and soft tissues characterized in at least 96% of cases by specific fu sion transcripts originating from recurrent chromosomal translocations. Cli nical protocols for high-risk metastatic: ETs include high-dose radiation/c hemotherapy followed by autologous peripheral blood stem cell (PBSC) reinfu sion. We used nested reverse transcriptase polymerase chain reaction (RI-PC R) to search for the presence of ET-specific transcripts in PBSC collection s from patients with high-risk ET in order to collect harvests free from ne oplastic cells but still sufficient to obtain early stable engraftment. Design and Methods. Thirty-seven harvest samples from 15 ET patients treate d with mobilizing chemotherapy were analyzed. Nested RT-PCR was performed t o detect ET-specific transcripts In RNA extracted from the PBSC collections . Results. A total of 30 harvests was performed. On average, 2 harvests (rang e 1-4) were sufficient to collect the minimum required number of mononuclea r cells (2.5x10(6)/kg). Nested RT-PCR revealed neoplastic cells in 4/30 (13 %) harvests, which were derived from 3/15 patients (20%). After further cyt oreductlve/mobilizing chemotherapy, a total of 7 additional harvests taken from these 3 patients were all free from neoplastic cells. Interpretation and Conclusions. PBSC collections from ET patients undergoin g autologous stem cell transplantation are at risk of neoplastic contaminat ion. Additional harvests following a further cycle of cytoreductlve/mobiliz ing therapy may he sufficient to obtain non-contaminated material for reinf usion. (C) 1999, Fenata Storti Foundation.