A virus-directed enzyme prodrug therapy approach to purging neuroblastoma cells from hematopoietic cells using adenovirus encoding rabbit carboxylesterase and CPT-11

Tumor cells that contaminate hematopoietic cell preparations contribute to the relapse of neuroblastoma patients who receive autologous stem cell resc ue as a component of therapy. Therefore, effective purging methods are need ed. This study details in vitro experiments to develop a viral-directed enz yme prodrug purging method that specifically targets neuroblastoma cells. T he approach uses an adenovirus to deliver the cDNA encoding a rabbit liver carboxylesterase that efficiently activates the prodrug irinotecan,7-ethyl- 10-[4-(1-piperidino)-1-piperidino]carbonyloxycamptothecin (CPT-11), The dat a show that an adenoviral multiplicity of infection of 50 transduces 100% o f cultured neuroblastoma cells and primary tumor cells, irrespective of the level of tumor cell line contamination, Exposure of neuroblastoma cell lin es or of mixtures of these cell lines with CD34(+) cells at a ratio of 10:9 0 to replication-deficient AdRSVrCE for 24 h and subsequent exposure of cel ls to 1-5 ELM CPT-11 for 4 h increased the toxicity of CPT-11 to three neur oblastoma cell lines (SJNB-1, NB-1691, and SK-N-SH) from similar to 20-50-f old and eradicated their clonogenic potential, Also, after "purging," RNA f or neuroblastoma cell markers (tyrosine hydroxylase, synaptophysin, and N-M YC) was undetectable by reverse transcription-PCR, In contrast, the purging protocol did not affect the number or type of colonies formed by CD34(+) c ells in an in vitro progenitor cell assay, No bystander effect on CD34(+) c ells was observed. The method described is being investigated for its poten tial clinical utility, particularly its efficacy for use with patients havi ng relatively high tumor burdens, because no published methods have been sh own to be efficacious when the tumor burden exceeds 1%.