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Background: In the last years, several efforts have been made to classify colorectal cancer (CRC) into well-defined molecular subgroups, representing the intrinsic inter-patient heterogeneity, known as Consensus Molecular Subtypes (CMSs). Methods: In this work, we performed a meta-analysis of CRC patients stratified into four CMSs. We identified a negative correlation between a high level of anaplastic lymphoma kinase (ALK) expression and relapse-free survival, exclusively in CMS1 subtype. Stemming from this observation, we tested cell lines, patient-derived organoids and mice with potent ALK inhibitors, already approved for clinical use. Results: ALK interception strongly inhibits cell proliferation already at nanomolar doses, specifically in CMS1 cell lines, while no effect was found in CMS2/3/4 groups. Furthermore, in vivo imaging identified a role for ALK in the dynamic formation of 3D tumor spheroids. Consistently, ALK appeares constitutively phosphorylated in CMS1, and it signals mainly through the AKT axis. Mechanistically, we found that CMS1 cells display several copies of ALKAL2 ligand and ALK-mRNAs, suggesting an autocrine loop mediated by ALKAL2 in the activation of ALK pathway, responsible for the invasive phenotype. Consequently, disruption of ALK axis mediates the pro-apoptotic action of CMS1 cell lines, both in 2D and 3D and enhanced cell-cell adhesion and e-cadherin organization. In agreement with all these findings, the ALK signature encompassing 65 genes statistically associated with worse relapse-free survival in CMS1 subtype. Finally, as a proof of concept, the efficacy of ALK inhibition was demonstrated in both patient-derived organoids and in tumor xenografts in vivo. Conclusions: Collectively, these findings suggest that ALK targeting may represent an attractive therapy for CRC, and CMS classification may provide a useful tool to identify patients who could benefit from this treatment. These findings offer rationale and pharmacological strategies for the treatment of CMS1 CRC. [ABSTRACT FROM AUTHOR]

Colorectal cancer (CRC) develops over several years, thus, early diagnosis and prevention are fundamental to reduce CRC burden. A new non-invasive blood test based on 4 mRNAs encompassing adhesion molecules such as CEACAM6, as well as a member of the collagen I superfamily, namely COL1A2, combined with LGALS4 and TSPAN8, was recently discovered and referred to as CELTIC panel [1]. The expression of the CELTiC panel was measured by quantitative PCR. The panel of putative biomarkers was subsequently evaluated in 101 subjects resulted positive to fecal immunochemical screening test (FIT) highlighting the potential to distinguish colonoscopy negative-FIT positive patients (NFIT, n=36), patients with low risk lesions (LR, n=36) and with high risk lesions or CRC (HR/CRC, n=92) [2]we searched for mRNA blood markers (CELTiC panel. In this study 174 healthy FIT negative subjects (FITN) were analysed [3] and compared to previous groups, evaluating also the influence of age and sex. Within the FITN group, CEACAM6 and COL1A2 display significantly lower expression in female than in male, confirming this sex difference of CEACAM6, a differentiation marker in normal colonocytes, also in the older groups (60-70 y.o.) and supporting current data on the importance of gender- and age-specific reference intervals for the early diagnosis of CRC [4,5]. The four genes showed significantly lower expression in FITN than in HR/CRC. Interestingly, TSPAN8 and COL1A2 were significantly lower expressed in FITN also than in NFIT and LR patients. TSPAN8, an integral membrane protein which upregulation promotes metastasis [6], was confirmed by logistic model as able to discriminate FITN from NFIT, LR and HR/CRC. Interestingly, also LGALS4, involved in cell-cell interaction and studied in CRC patients [7,8], was able to differentiate FITN from NFIT (false FIT positive). TSPAN8 and galectins have been described also in exosomes [9] suggesting the origin of the mRNAs of the CELTiC panel. Finally, the CELTIC panel showed good sensitivity (84-90%), specificity (76-81%) and AUC (0.87-0.89) to discriminate FITN from the other groups. The CELTiC panel was confirmed as a useful tool to diagnose CRC highlighting the importance of sex and age. A multicenter cross-sectional study will enroll 800 FIT positive subject at the Universities of Amsterdam and Bologna to better estimates the performances of the CELTiC panel and to further validate its robustness and usefulness in the early diagnosis of CRC and to exclude false FIT positive subjects. [ABSTRACT FROM AUTHOR]