Low Frequency ALK Hotspots Mutations In Neuroblastoma Tumours Detected By Ultra-deep Sequencing: Implications For ALK Inhibitor Treatment.

The ALK tyrosine kinase receptor is oncogenically activated in neuroblastoma. Whereas numerous ALK fusion genes have been reported in different malignancies, in neuroblastoma ALK is mainly activated through point mutations. Three hotspot residues (F1174, F1245, and R1275) account for 85% of mutant ALK seen in neuroblastoma. In a cohort of 105 Swedish neuroblastoma cases of all stages, these hotspot regions were re-sequenced (>5000X). ALK mutations were detected in 16 of 105 patients (range of variant allele fraction: 2.7-60%). Mutations at the F1174 and F1245 hotspot were observed in eleven and three cases respectively. ALK mutations were also detected at the I1171 and L1240 codons in one tumor each. No mutations were detected at R1275. Sanger sequencing could confirm ALK status for all mutated samples with variant allele fraction above 15%. Four of the samples with subclonal ALK mutation fraction below this would have gone undetected relying on Sanger sequencing only. No distinct mutation spectrum in relation to neuroblastoma tumours genomic subtypes could be detected although there was a paucity of ALK mutations among 11q-deleted tumors. As ALK mutations status opens up an excellent opportunity for application of small molecule inhibitors targeting ALK, early and sensitive detection of ALK alterations is clinically important considering its potential role in tumour progression.