The Impact of ETV6-NTRK3 Oncogenic Gene Fusions on Molecular and Signaling Pathway Alterations

SIMPLE SUMMARY: Gene fusions produce chimeric fusion proteins with unpredictable properties, many of which function as oncogenic drivers. ETV6-NTRK3 produces a constitutively active fusion kinase, which is not membrane bound like the endogenous NTRK3 kinase. Several studies have investigated the interactions of ETV6-NTRK3 over the years with few interactors being reported and some interactors reportedly being lost compared to the endogenous NTRK3. We utilize proximity labeling to produce the first proteomics level study on ETV6-NTRK3 close protein milieu and analyze the interactomes of all the four known ETV6-NTRK3 variants with functional kinase domains. Existing clinical literature shows that these four ETV6-NTRK3 variants occur at differing frequencies and originate in different tissues, with a “canonical” variant being more frequent and is also reported in other tissues. The ETV6-NTRK3 variants were found to have both similarities and differences in interactors, which might explain differences in reported frequencies and tissue specificities of the variants. ABSTRACT: Chromosomal translocations creating fusion genes are common cancer drivers. The oncogenic ETV6-NTRK3 (EN) gene fusion joins the sterile alpha domain of the ETV6 transcription factor with the tyrosine kinase domain of the neurotrophin-3 receptor NTRK3. Four EN variants with alternating break points have since been detected in a wide range of human cancers. To provide molecular level insight into EN oncogenesis, we employed a proximity labeling mass spectrometry approach to define the molecular context of the fusions. We identify in total 237 high-confidence interactors, which link EN fusions to several key signaling pathways, including ERBB, insulin and JAK/STAT. We then assessed the effects of EN variants on these pathways, and showed that the pan NTRK inhibitor Selitrectinib (LOXO-195) inhibits the oncogenic activity of EN2, the most common variant. This systems-level analysis defines the molecular framework in which EN oncofusions operate to promote cancer and provides some mechanisms for therapeutics.