Abstract 3956: Combination of SPARC and radiation suppresses HSP27 and induces p21(CIP1/WAF1) in neuroblastoma tumor cells

Neuroblastoma (NBL) is the third most common malignancy in children and accounts for more than 15% of cancer-related deaths in children in the US. Approximately 50% of patients present with advanced-stage and/or high risk disease. Despite significant intensification of conventional chemotherapy regimens, the long-term survival rates for these children remain less than 40%. A somewhat improved outcome has been obtained with multimodality therapeutic approaches including chemotherapy, surgery, autologous bone marrow transplantation, radiation therapy and use of biological agents. However, long-term survival rates for children with high-risk disease remain poor and survivors experience significant immediate and late toxicities which further limits conventional chemotherapy dose intensification. Novel biological therapies are therefore much needed. Secreted protein, acidic and rich in cysteine (SPARC), also known as osteonectin or BM-40, belongs to the matricellular family of secreted proteins, which also includes thrombospondin 1 and 2, osteopontin, and tenascins C and X. The biological functions of SPARC are known to be variable in human cancers. In neuroblastoma however, SPARC expression has been found to be associated with impaired tumor growth and angiogenesis. Our previous studies showed that overexpression of SPARC induced autophagy-mediated apoptosis in neuroblastoma cells. In the present study we have attempted to decipher the mechanisms of this apoptotic induction. We used two neuroblastoma cell lines SK-N-BE2 (p53mt) and NB1691 (p53wt) and transfected them with plasmid overexpressing Myc-DDK-tagged SPARC with and without ionizing radiation. We observed that overexpression of SPARC accompanied with ionizing radiation (5Gy) suppressed HSP27 in NB1691 (5-fold) and SK-N-BE(2) (2-fold) cells. Further we also observed that expression of p21(CIP1/WAF1) was increased in SPARC overexpressed radiated (5Gy) cells; 4-fold in NB1691 cells and 0.5-fold SK-N-BE(2) cells. Levels of cyclin B were also found to be increased in both SK-N-BE2 (3-fold) and NB1691 (5-fold) after exposure to radiation. However SPARC overexpression in radiated cells caused a decrease in levels of Cyclin B in NB1691 (-3 fold) and SK-N-BE(2) (-2 fold) cells. Cell cycle analysis revealed that radiation induced G2-phase arrest in both NB1691 and SK-N-BE(2) cells. This G2 phase arrest was followed by decreased mitochondrial membrane potential (Δψ) in SPARC overexpressed cells when compared to controls. We further observed that suppression of native SPARC by siRNA in both SK-N-BE2 and NB1691 cells altered their sensitivity to radiation as determined by MTT assay. Our results demonstrate that SPARC expression regulates radiation sensitivity in NB1691 (p53wt) and to a lesser extent in SK-N-BE2 (p53mt) cells via modulation of the heat shock protein HSP27 and the DNA damage response molecule p21(CIP1/WAF1). Citation Format: Christopher S. Gondi, Smita Tanpure, Reuben Antony, Karen S. Fernandez, Meena Gujrati, Julian Lin. Combination of SPARC and radiation suppresses HSP27 and induces p21(CIP1/WAF1) in neuroblastoma tumor cells. [abstract]. In: Proceedings of the 105th Annual Meeting of the American Association for Cancer Research; 2014 Apr 5-9; San Diego, CA. Philadelphia (PA): AACR; Cancer Res 2014;74(19 Suppl):Abstract nr 3956. doi:10.1158/1538-7445.AM2014-3956