title: Common genetic variants in NEFL influence gene expression and neuroblastoma risk.
abstract: The genetic etiology of sporadic neuroblastoma is still largely obscure. In a genome-wide association study, we identified single-nucleotide polymorphisms (SNP) associated with neuroblastoma at the CASC15, BARD1, LMO1, DUSP12, HSD17B12, HACE1, and LIN28B gene loci, but these explain only a small fraction of neuroblastoma heritability. Other neuroblastoma susceptibility genes are likely hidden among signals discarded by the multiple testing corrections. In this study, we evaluated eight additional genes selected as candidates for further study based on proven involvement in neuroblastoma differentiation. SNPs at these candidate genes were tested for association with disease susceptibility in 2,101 cases and 4,202 controls, with the associations found replicated in an independent cohort of 459 cases and 809 controls. Replicated associations were further studied for cis-effect using gene expression, transient overexpression, silencing, and cellular differentiation assays. The neurofilament gene NEFL harbored three SNPs associated with neuroblastoma (rs11994014: Pcombined = 0.0050; OR, 0.88; rs2979704: Pcombined = 0.0072; OR, 0.87; rs1059111: Pcombined = 0.0049; OR, 0.86). The protective allele of rs1059111 correlated with increased NEFL expression. Biologic investigations showed that ectopic overexpression of NEFL inhibited cell growth specifically in neuroblastoma cells carrying the protective allele. NEFL overexpression also enhanced differentiation and impaired the proliferation and anchorage-independent growth of cells with protective allele and basal NEFL expression, while impairing invasiveness and proliferation of cells homozygous for the risk genotype. Clinically, high levels of NEFL expression in primary neuroblastoma specimens were associated with better overall survival (P = 0.03; HR, 0.68). Our results show that common variants of NEFL influence neuroblastoma susceptibility and they establish that NEFL expression influences disease initiation and progression.
authors.affiliation: Università degli Studi di Napoli Federico II, Dipartimento di Medicina Molecolare e Biotecnologie Mediche, Naples, Italy. CEINGE Biotecnolgie Avanzate, Naples, Italy. mario.capasso@unina.it.
authors.forename: Mario
authors.initials: M
authors.lastname: Capasso
authors.affiliation: Division of Oncology, Children's Hospital of Philadelphia, Philadelphia, Pennsylvania. Center for Childhood Cancer Research, Children's Hospital of Philadelphia, Philadelphia, Pennsylvania. Department of Pediatrics, Perelman School of Medicine, University of Pennsylvania, Philadelphia, Pennsylvania. Genomics and Computational Biology, Biomedical Graduate Studies, Perelman School of Medicine, University of Pennsylvania, Philadelphia, Pennsylvania.
authors.forename: Sharon
authors.initials: S
authors.lastname: Diskin
authors.affiliation: Università degli Studi di Napoli Federico II, Dipartimento di Medicina Molecolare e Biotecnologie Mediche, Naples, Italy. CEINGE Biotecnolgie Avanzate, Naples, Italy.
authors.forename: Flora
authors.initials: F
authors.lastname: Cimmino
authors.affiliation: Università degli Studi di Napoli Federico II, Dipartimento di Medicina Molecolare e Biotecnologie Mediche, Naples, Italy. CEINGE Biotecnolgie Avanzate, Naples, Italy.
authors.forename: Giovanni
authors.initials: G
authors.lastname: Acierno
authors.affiliation: Università degli Studi di Napoli Federico II, Dipartimento di Medicina Molecolare e Biotecnologie Mediche, Naples, Italy.
authors.forename: Francesca
authors.initials: F
authors.lastname: Totaro
authors.affiliation: Università degli Studi di Napoli Federico II, Dipartimento di Medicina Molecolare e Biotecnologie Mediche, Naples, Italy.
authors.forename: Giuseppe
authors.initials: G
authors.lastname: Petrosino
authors.affiliation: CEINGE Biotecnolgie Avanzate, Naples, Italy.
authors.forename: Lucia
authors.initials: L
authors.lastname: Pezone
authors.affiliation: Division of Oncology, Children's Hospital of Philadelphia, Philadelphia, Pennsylvania. Center for Childhood Cancer Research, Children's Hospital of Philadelphia, Philadelphia, Pennsylvania.
authors.forename: Maura
authors.initials: M
authors.lastname: Diamond
authors.affiliation: Abramson Family Cancer Research Institute, Perelman School of Medicine at the University of Pennsylvania, Philadelphia, Pennsylvania. Genomics and Computational Biology, Biomedical Graduate Studies, Perelman School of Medicine, University of Pennsylvania, Philadelphia, Pennsylvania.
authors.forename: Lee
authors.initials: L
authors.lastname: McDaniel
authors.affiliation: Department of Pediatrics, Perelman School of Medicine, University of Pennsylvania, Philadelphia, Pennsylvania. The Center for Applied Genomics, Children's Hospital of Philadelphia, Philadelphia, Pennsylvania. Division of Genetics, Children's Hospital of Philadelphia, Philadelphia, Pennsylvania.
authors.forename: Hakon
authors.initials: H
authors.lastname: Hakonarson
authors.affiliation: Università degli Studi di Napoli Federico II, Dipartimento di Medicina Molecolare e Biotecnologie Mediche, Naples, Italy. CEINGE Biotecnolgie Avanzate, Naples, Italy.
authors.forename: Achille
authors.initials: A
authors.lastname: Iolascon
authors.affiliation: Department of Pediatrics, Perelman School of Medicine, University of Pennsylvania, Philadelphia, Pennsylvania. Division of Genetics, Children's Hospital of Philadelphia, Philadelphia, Pennsylvania. Department of Molecular Medicine, University of Rome "La Sapienza," Rome, Italy. Department of Biostatistics and Epidemiology, Perelman School of Medicine, University of Pennsylvania, Philadelphia, Pennsylvania.
authors.forename: Marcella
authors.initials: M
authors.lastname: Devoto
authors.affiliation: Division of Oncology, Children's Hospital of Philadelphia, Philadelphia, Pennsylvania. Center for Childhood Cancer Research, Children's Hospital of Philadelphia, Philadelphia, Pennsylvania. Department of Pediatrics, Perelman School of Medicine, University of Pennsylvania, Philadelphia, Pennsylvania. Abramson Family Cancer Research Institute, Perelman School of Medicine at the University of Pennsylvania, Philadelphia, Pennsylvania. Genomics and Computational Biology, Biomedical Graduate Studies, Perelman School of Medicine, University of Pennsylvania, Philadelphia, Pennsylvania.
authors.forename: John M
authors.initials: JM
authors.lastname: Maris
mesh_terms: D000483:Alleles; D016022:Case-Control Studies; D002454:Cell Differentiation / Q000235:genetics; D002460:Cell Line; D015870:Gene Expression; D020022:Genetic Predisposition to Disease; D014644:Genetic Variation; D055106:Genome-Wide Association Study / Q000379:methods; D005838:Genotype; D057809:HEK293 Cells; D006801:Humans; D009447:Neuroblastoma / Q000235:genetics* / Q000378:metabolism; D016900:Neurofilament Proteins / Q000096:biosynthesis / Q000235:genetics*; D020641:Polymorphism, Single Nucleotide; D012306:Risk
chemical_list: D016900:Neurofilament Proteins; C044882:neurofilament protein L