Activation of ALK Signaling Pathway Induces DNA Damage Response in Colorectal Cancer

In recent years, there have been several attempts to classify colorectal cancer (CRC) into well-defined molecular subgroups that reflect the inherent heterogeneity between patients. Consensus molecular subtypes (CMS) classification successfully achieves this purpose and provides an important tool for personalized medicine. We previously identified an inverse association between high levels of anaplastic lymphoma kinase (ALK) expression and recurrence-free survival, only in the CMS1 subtype, confirmed by a wide array of in vitro and in vivo assays. To support the hypothesis of ALK having a strong implication in CRC patient tumor initiation, we overexpressed ALK receptor in a normal human colon epithelial cell line (NCM460). ALK overexpression was confirmed both in 2D and in 3D spheroid models, cultured in 3D. The role of ALK pathway activation in colonocytes was then investigated by examining its effects on proliferation, survival, migration, and invasion through various 2D and 3D in vitro assays. Immunohistochemistry analysis revealed an increase in the proliferation marker KI67, and a decrease in the E-cadherin (E-CAD) expression in ALK-overexpressing spheroids compared to the control ones suggesting a correlation with tumor metastasis, progression, and invasion. Moreover, we identified a strong correlation between high level of expression of ALK and the downregulation of MSH2, a DNA Mismatch Repair (MMR) protein associated to microsatellite instability, which is typically found in CMS1 patients. Finally, the mass density, size, and weight of in vitro NCM 460 spheroids were analyzed with an emerging microfluidic technology provided by CellDynamics isrl company. Overall, these results suggest that ALK overexpression itself is sufficient to induce aggressive features in normal colonocytes, further supporting the hypothesis that ALK may be an attractive target for CMS1 colorectal cancer therapy.