title: Acquired genetic alterations in tumor cells dictate the development of high-risk neuroblastoma and clinical outcomes.
abstract: BACKGROUND
  Determining the driving factors and molecular flow-through that define the switch from favorable to aggressive high-risk disease is critical to the betterment of neuroblastoma cure.
  
  
  METHODS
  In this study, we examined the cytogenetic and tumorigenic physiognomies of distinct population of metastatic site- derived aggressive cells (MSDACs) from high-risk tumors, and showed the influence of acquired genetic rearrangements on poor patient outcomes.
  
  
  RESULTS
  Karyotyping in SH-SY5Y and MSDACs revealed trisomy of 1q, with additional non-random chromosomal rearrangements on 1q32, 8p23, 9q34, 15q24, 22q13 (additions), and 7q32 (deletion). Array CGH analysis of individual clones of MSDACs revealed genetic alterations in chromosomes 1, 7, 8, and 22, corresponding to a gain in the copy numbers of LOC100288142, CD1C, CFHR3, FOXP2, MDFIC, RALYL, CSMD3, SAMD12-AS1, and MAL2, and a loss in ADAM5, LOC400927, APOBEC3B, RPL3, MGAT3, SLC25A17, EP300, L3MBTL2, SERHL, POLDIP3, A4GALT, and TTLL1. QPCR analysis and immunoblotting showed a definite association between DNA-copy number changes and matching transcriptional/translational expression in clones of MSDACs. Further, MSDACs exert a stem-like phenotype. Under serum-free conditions, MSDACs demonstrated profound tumorosphere formation ex vivo. Moreover, MSDACs exhibited high tumorigenic capacity in vivo and prompted aggressive metastatic disease. Tissue microarray analysis coupled with automated IHC revealed significant association of RALYL to the tumor grade in a cohort of 25 neuroblastoma patients. Clinical outcome association analysis showed a strong correlation between the expression of CFHR3, CSMD3, MDFIC, FOXP2, RALYL, POLDIP3, SLC25A17, SERHL, MGAT3, TTLL1, or LOC400927 and overall and relapse-free survival in patients with neuroblastoma.
  
  
  CONCLUSIONS
  Together, these data highlight the ongoing acquired genetic rearrangements in undifferentiated tumor-forming neural crest cells, and suggest that these alterations could switch favorable neuroblastoma to high-risk aggressive disease, promoting poor clinical outcomes.
authors.affiliation: Department of Radiation Oncology, University of Oklahoma Health Sciences Science Center, 940 Stanton L. Young Blvd., BMSB 737, Oklahoma City, OK, 73104, USA. Faizan-Khan@ouhsc.edu.
authors.forename: Faizan H
authors.initials: FH
authors.lastname: Khan
authors.affiliation: Department of Radiation Oncology, University of Oklahoma Health Sciences Science Center, 940 Stanton L. Young Blvd., BMSB 737, Oklahoma City, OK, 73104, USA. Vijayabaskar-Pandian@ouhsc.edu.
authors.forename: Vijayabaskar
authors.initials: V
authors.lastname: Pandian
authors.affiliation: Department of Radiation Oncology, University of Oklahoma Health Sciences Science Center, 940 Stanton L. Young Blvd., BMSB 737, Oklahoma City, OK, 73104, USA. Satish-Ramraj@ouhsc.edu.
authors.forename: Satishkumar
authors.initials: S
authors.lastname: Ramraj
authors.affiliation: Department of Pathology, University of Texas Health Sciences Center, San Antonio, TX, USA. natarajan@uthscsa.edu.
authors.forename: Mohan
authors.initials: M
authors.lastname: Natarajan
authors.affiliation: Stephenson Cancer Center, Oklahoma City, OK, USA. Sheeja-Aravindan@ouhsc.edu.
authors.forename: Sheeja
authors.initials: S
authors.lastname: Aravindan
authors.affiliation: Department of Radiation Oncology, University of Oklahoma Health Sciences Science Center, 940 Stanton L. Young Blvd., BMSB 737, Oklahoma City, OK, 73104, USA. Terence-Herman@ouhsc.edu.|Stephenson Cancer Center, Oklahoma City, OK, USA. Terence-Herman@ouhsc.edu.
authors.forename: Terence S
authors.initials: TS
authors.lastname: Herman
authors.affiliation: Department of Radiation Oncology, University of Oklahoma Health Sciences Science Center, 940 Stanton L. Young Blvd., BMSB 737, Oklahoma City, OK, 73104, USA. naravind@ouhsc.edu.
authors.forename: Natarajan
authors.initials: N
authors.lastname: Aravindan
mesh_terms: D000818:Animals; D045744:Cell Line, Tumor; D002869:Chromosome Aberrations; D002871:Chromosome Banding; D055028:Comparative Genomic Hybridization; D056915:DNA Copy Number Variations; D004195:Disease Models, Animal; D018450:Disease Progression; D056726:Genetic Association Studies; D064593:Heterografts; D006801:Humans; D009362:Neoplasm Metastasis; D009367:Neoplasm Staging; D014411:Neoplastic Stem Cells / Q000378:metabolism / Q000473:pathology; D009447:Neuroblastoma / Q000235:genetics* / Q000401:mortality* / Q000473:pathology; D011379:Prognosis; D000090822:Ribosomal Protein L3