[2S01a] 10. ASPIRE - Part 5

3:00 PM - 3:15 PM JST(6:00 AM - 6:15 AM UTC)

[2ASPR-12] Divergent signalosomes and a shared assembly mechanism define EML4-ALK variant oncogenicity

○Josephina Sampson¹ (1. University of Leeds (UK)/ University of Osaka (JP) (UK))

Keywords:

EML4-ALK fusions,Signalling,condensates,cancer,ALK TKI,Drug-resistance,Systems Biology

Background: The dysregulation of tyrosine kinase receptors (RTKs) promotes cellular transformation and non-small cell lung cancer (NSCLC). This can be caused by genetic aberrations such as point mutations, truncations and gene fusions. RTK translocations, especially EML4-ALK fusions, are primary drivers of NSCLC. Importantly, clinical outcomes of EML4-ALK NSCLC differ drastically depending on which variant is expressed. Variant 3 (V3) is notably more aggressive and resistant to ALK kinase inhibitors (KIs) than the canonical variant 1 (V1), through mechanisms that remain poorly understood. Although recent work highlighted molecular differences in the ability of the variants to form liquid-liquid phase separated (LLPS) structures, to respond to ALK kinase inhibitors, and to activate signaling pathways, we still don’t have a comprehensive list of the proteins that can be hijacked by EML4-ALK fusions. Purpose: The aim of our work is to define how EML4-ALK signalosomes drive variant-specific signalling in non-small cell lung cancer (NSCLC).
Methods: To achieve this we integrate proximity proteomics, phosphoproteomics and mathematical systems methods to elucidate the signalling cascades of EML4-ALK variants in NSCLC. We use super-resolution microscopy to examine the composition of EML4-ALK signalosomes and their importance in driving downstream signalling cascades.
Results: Our proteomic analyses revealed that while V1 recruits and activates canonical MAPK and PI3K effectors, V3 reconfigures cellular signaling through a non-canonical, inhibitor-resistant interferon response. We also identify a global rewiring of phospho-dependent signaling unique to V3, characterized by a massive expansion of the phosphotyrosine landscape and a simultaneous collapse of global pSer/pThr networks. Despite these divergent signaling architectures, we show that both variants require SHC1, GAB1 and the phosphatase and scaffold PTPN11 (SHP2) for the biogenesis of EML4-ALK signalosomes. Crucially, pharmacologic inhibition of PTPN11 disassembles these oncogenic hubs across variants, yet the V3-specific interferon response remains largely decoupled from this physical disruption.
Conclusion: Our work strongly deepens our understanding of EML4-ALK oncogenicity and of variant specific signaling rewiring. We show that V3 expressing cells generate a modified interferon response and immune state that is largely unresponsive to ALK targeted drug treatments. Our work also identifies PTPN11 as a critical, variant-agnostic vulnerability for potential therapeutic intervention in ALK-driven malignancies.