Integrating next-generation sequencing into the routine neuroblastoma workflow identifies actionable genomic alterations and reduces low-yield karyotype testing.

Neuroblastoma (NB) is a genetically complex entity. Alterations at both the gene and chromosomal levels have been identified in NB, some with prognostic and therapeutic implications. More recently, ALK mutations and amplifications have been identified and may carry therapeutic implications with the advent of ALK inhibitors. Given the increasing number and complexity of these alterations, genetic and molecular testing must be performed using multiple modalities. We analyze the role of adding next generation sequencing (NGS) to our NB workflow. We examined 22 cases which all underwent NGS. A subset also had available karyotype, MYCN FISH, and chromosomal microarray (CMA) results. In cases with segmental chromosomal aberrations (SCA) and MYCN amplification identified by CMA and/or FISH, NGS was a reliable detection method. NGS also detected mutations in ALK, HRAS, CDKN2A, and ATM. Two cases were also found to have high tumor mutational burden (TMB). Karyotyping was conclusive in 11 of 22 cases, none of which revealed chromosomal abnormalities. The incorporation of NGS not only detects mutations and high TMB, which may have therapeutic significance, but also reliably detects SCAs. However, CMA remains an important complement providing a global view of copy number alterations and loss of heterozygosity. We also found that karyotype has limited utility in the setting of NB. Based on this data, we developed an institutional workflow for NB, including MYCN FISH, CMA, and NGS.