Abstract B67: Targeting glutamine metabolism in neuroblastoma and Ewing's sarcoma

Therapeutic targeting of aberrant cancer metabolism is currently an area of great interest in pediatric oncology. Previous reports have established that oncogenic Myc alters cellular metabolism resulting in cells dependent on exogenous glutamine. Therefore, we chose to target glutamine metabolism in the pediatric cancers Neuroblastoma and Ewing's Sarcoma. Neuroblastoma (NB) is a pediatric malignancy of the developing sympathetic nervous system, and approximately 20% of patients have amplified MYCN and very poor prognosis. Ewing's Sarcoma (EWS) is an aggressive tumor of the bone and soft tissue characterized by chromosome translocations resulting in EWSR1/ETS family fusion proteins. Approximately 85% of EWS cases contain an EWSR1/FLI fusion protein that targets c-Myc for overexpression and potentially cooperates with c-Myc in cell transformation. We chose to use the glutamine antagonist DON (6-diazo-5-oxo-norleucine) as a tool to examine the effect of inhibiting glutamine metabolism in NB and EWS cells. DON was first explored as a cancer chemotherapeutic in the 1950s and 1960s, but severe nausea in patients following treatment limited its use as a cancer therapeutic. DON is a competitive inhibitor which irreversibly alkylates glutamine-utilizing enzymes, including those involved in nucleotide biosynthesis and glutaminolysis, two important Myc-regulated metabolic pathways in cancer. DON effectively inhibited a panel of six NB and three EWS cell lines, while having minimal effect on BJ fibroblast control cells. In addition, we found a strong correlation between sensitivity to glutamine addiction and susceptibility to DON (R2 = 0.73). Next, we used human tumor xenografts in nude mice to examine the effect of inhibiting glutamine metabolism in the context of an established tumor. Following injection of neuroblastoma (SK-N-AS, SK-N-BE2) or Ewing's Sarcoma (SK-N-MC, SK-ES-1) cells, tumors were allowed to reach 200mm3 in size before being randomized into treatment groups. Mice were then treated with 100mg/kg DON or water control twice per week. DON treatment significantly inhibited both tumor growth and BrdU incorporation in all NB and EWS tumors tested, suggesting that targeting glutamine metabolism could be an effective therapeutic approach. Interestingly, although DON treatment was strongly cytostatic in all cell lines tested, only in the SK-N-BE2 cells did we detect increased cleavage of caspase-3 as a marker for apoptosis. These results suggest that perhaps c