ALK mutation dynamics and clonal evolution in a neuroblastoma model exhibiting two ALK mutations.

Abstract

The ALK gene is a major oncogene of neuroblastoma cases exhibiting ALK activating mutations. Here, we characterized two neuroblastoma cell lines established from a stage 4 patient at diagnosis either from the primary tumor (PT) or from the bone marrow (BM). Both cell lines exhibited similar genomic profiles. All cells in the BM-derived cell line exhibited an ALK F1174L mutation, whereas this mutation was present in only 5% of the cells in the earliest passages of the PT-derived cell line. The BM-derived cell line presented with a higher proliferation rate in vitro and injections in Nude mice resulted in tumor formation only for the BM-derived cell line. Next, we observed that the F1174L mutation frequency in the PT-derived cell line increased with successive passages. Further Whole Exome Sequencing revealed a second ALK mutation, L1196M, in this cell line. Digital droplet PCR documented that the allele fractions of both mutations changed upon passages, and that the F1174L mutation reached 50% in late passages, indicating clonal evolution. In vitro treatment of the PT-derived cell line exhibiting the F1174L and L1196M mutations with the alectinib inhibitor resulted in an enrichment of the L1196M mutation. Using xenografts, we documented a better efficacy of alectinib compared to crizotinib on tumor growth and an enrichment of the L1196M mutation at the end of both treatments. Finally, single-cell RNA-seq analysis was consistent with both mutations resulting in ALK activation. Altogether, this study provides novel insights into ALK mutation dynamics in a neuroblastoma model harbouring two ALK mutations.

• Authors: Simon Durand, Cécile Pierre-Eugène, Olivier Mirabeau, Caroline Louis-Brennetot, Valérie Combaret, Léo Colmet-Daage, Orphée Blanchard, Angela Bellini, Estelle Daudigeos-Dubus, Virginie Raynal, Gudrun Schleiermacher, Sylvain Baulande, Olivier Delattre, Isabelle Janoueix-Lerosey
• Journal: Oncotarget (Oncotarget) Vol. 10 Issue 48 Pg. 4937-4950 (Aug 13 2019) ISSN: 1949-2553 [Electronic] United States
• PMID: 31452835 (Publication Type: Journal Article)