Abstract
Neuroblastoma is the most common extracranial solid tumor occurring in early childhood and has a survival rate less than 40% for the most aggressive subset. We have recently reported several ...common genetic variations associated with this human cancer (NEJM 2008, Nat Genet 2008, Nature 2009). Here, we describe an integrative analysis of matched germline and somatic tumor data identifying NME7 (non-metastatic cells 7) as both a predisposition gene and candidate oncogene in neuroblastoma. We first performed a genome-wide association study (GWAS) of copy number variations (CNVs) in a total of 1,440 Caucasian neuroblastoma patients and 4,436 healthy Caucasian controls genotyped at ∼550,000 single nucleotide polymorphisms (SNPs). In addition to confirming the 1q21.1 CNV association we previously reported (Nature 2009), we identified a new under-represented deletion associated with neuroblastoma at chromosome 1q24.2 within the non-metastatic cells 7 (NME7) gene (PCMH = 2.22 × 10−15; OR = 0.55, 95% CI: 0.43-0.62). To evaluate whether somatically acquired alterations of NME7 in tumor DNA may also be important to tumorigenesis, we analyzed tumor DNA copy number in 591 primary tumors and matched mRNA expression in a subset of 100 samples. We observed somatic gain of the NME7 locus in 24% of neuroblastomas (PGISTIC = 3.3 × 10−9). Tumor acquired somatic gain correlated with increased NME7 mRNA expression (P = 0.007), and Western blot analysis of six neuroblastoma cell lines confirmed a strict correlation between mRNA and protein levels (R2 = 0.90). To investigate the functional significance of NME7, we transiently transfected siRNA targeting NME7 in a panel of eight neuroblastoma cell lines and monitored cell growth using the RTces system. We observed significant growth inhibition (G.I.) with siRNA targeting NME7 compared to transfection with non-targeting siRNA control in all neuroblastoma cell lines (G.I.range = 23.0 − 55.0%; Prange = < 0.0001 − 0.03). Percent growth inhibition was significantly higher in lines harboring somatic gain of the locus (P = 0.05; G.I.median = 44%). Importantly, no difference in cell growth was observed in control RPE1 (non-neuroblastoma) cells after transfection with siRNA targeting NME7 (P = 0.44). Finally, we performed a chamber-based in vitro cell migration assay to determine if NME7 may also affect cell motility in neuroblastoma. We observed a significant decrease in cell migration in the neuroblastoma cell line IMR5 after siRNA silencing of NME7 (P = 0.02), while no difference in cell migration was observed in RPE1 cells (P = 0.43). Together these data demonstrate the utility of combining germline and somatic data in assessing GWAS signals in cancer, and identify NME7 as both a predisposition locus and candidate oncogene in neuroblastoma. Ongoing efforts are focused on expanding our in vitro cell migration assay to additional cell lines and determining if NME7 influences colony formation in soft agar.
Citation Format: {Authors}. {Abstract title} abstract. In: Proceedings of the 101st Annual Meeting of the American Association for Cancer Research; 2010 Apr 17-21; Washington, DC. Philadelphia (PA): AACR; Cancer Res 2010;70(8 Suppl):Abstract nr 3866.
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