Oral Presentation Advances in Neuroblastoma Research Congress 2016

Identification of germline mutations in 776 children with neuroblastoma (#112)

Douglas R. Stewart 1 , Perry Evans 2 , Trevor Pugh 3 , Hongling Liao 1 , Karina Conkrite 2 , Rajesh Patidar 1 , Kristina A. Cole 2 , Neil Menghani 2 , Lance Farra 2 , Shile Zhang 1 , Sivasish Sindiri 1 , Young Song 1 , Xinyu Wen 1 , Jaime Guidry Auvil 1 , Daniela S. Gerhard 1 , John M. Maris 2 , Javed Khan 1 , Jun Wei 1 , Sharon J. Diskin 2
  1. National Cancer Institute, Bethesda, MD, USA
  2. Children's Hospital of Philadelphia, Philadelphia, PA, United States
  3. Princess Margaret Cancer Centre, Toronto, Canada

BACKGROUND. Genetic susceptibility to neuroblastoma is complex, and the contribution of rare germline mutations beyond ALK and PHOX2B is not known. Here, we report on our efforts to identify rare deleterious germline variants influencing tumorigenesis in sporadic neuroblastoma (NB). 

METHODS. We conducted Complete Genomics whole-genome sequencing (106 high-risk tumor-germline) and Illumina exome sequencing (222 high-risk tumor-germline; 52 overlapping with whole-genome). A custom-capture 500-gene panel was sequenced in a representative validation cohort of 500 NB patients (germline-only) and compared to 1,001 cancer-free controls sequenced/analyzed by the same platform/pipeline. Rare variants were annotated using the database for non-synonymous functional predictions (dbNSFP), aggregated, and tested for enrichment compared to Exome Aggregation Consortium (ExAC). Indels were called by bam2MPG and Platypus in the 222 exome cohort and in 1000 Genomes as a control. We initially focused on known cancer predisposition genes and genes underlying syndromes associated with NB.

RESULTS. SNV calls showed high concordance in overlapping exome and whole-genome samples (average 90.4% per sample), and 119/119 (100%) of SNVs tested from non-overlapping cases verified by Sanger sequencing. We observed 131 potentially deleterious rare variants/mutations in known cancer predisposition genes that were either loss of function, or had a CADD score > 20 and predicted to be damaging by multiple algorithms. Considering these, 25/276 cases (9%) harbored known pathogenic mutations, many present in genes previously reported to be mutated in NB: ALK (n=2), APC (n=3), BARD1 (n=3), BRCA1 (n=1), BRCA2 (n=2), CHEK2 (n=3), PALB2 (n=2), PHOX2B (n=1), PINK1 (n=1), NF1 (n=1), SDHB (n=1) and TP53 (n=1). In addition, we identified pathogenic variants in genes underlying syndromes associated with NB, including HRAS (n=1/276, Costello) and EZH2 (1/276, Weaver). 

CONCLUSION. In this case series, we conservatively estimate ~10% of children with NB harbor a known pathogenic germline mutation in a cancer susceptibility gene. Validation sequencing is underway in the 500-sample cohort and evaluation of additional mutations, including non-coding, is ongoing.