Background: Tertiary lymphoid structures (TLSs) exert a crucial role in the tumor microenvironment (TME), impacting tumor development, immune escape, and drug resistance. Nonetheless, the heterogeneity of TLSs in colorectal cancer (CRC) and their impact on prognosis and treatment response remain unclear. Methods: The authors collected genome, transcriptome, clinicopathological information, and digital pathology images from multiple sources. An unsupervised clustering algorithm was implemented to determine diverse TLS patterns in CRC based on the expression levels of 39 TLS signature genes (TSGs). Comprehensive explorations of heterogeneity encompassing mutation landscape, TME, biological characteristics, response to immunotherapy, and drug resistance were conducted using multiomics data. TLSscore was then developed to quantitatively assess TLS patterns of individuals for further clinical applicability. Results: Three distinct TLS patterns were identified in CRC. Cluster 1 exhibited upregulation of proliferation-related pathways, high metabolic activity, and intermediate prognosis, while Cluster 2 displayed activation of stromal and carcinogenic pathways and a worse prognosis. Both Cluster 1 and Cluster 2 may potentially benefit from adjuvant chemotherapy. Cluster 3, characterized by the activation of immune regulation and activation pathways, demonstrated a favorable prognosis and enhanced responsiveness to immunotherapy. The authors subsequently employed a regularization algorithm to construct the TLSscore based on nine core genes. Patients with lower TLSscore trended to prolonged prognosis and a more prominent presence of TLSs, which may benefit from immunotherapy. Conversely, those with higher TLSscore exhibited increased benefits from adjuvant chemotherapy. Conclusions: The authors identified distinct TLS patterns in CRC and characterized their heterogeneity through multiomics analyses. The TLSscore held promise for guiding clinical decision-making and further advancing the field of personalized medicine in CRC.

Colorectal cancer (CRC) with KRAS mutation is prone to develop metachronous metastasis. However, effective measures for prediction and prevention of such clinical issue are currently unavailable. In this study, we confirmed that KRAS mutation is a risk factor for CRC metachronous metastasis through meta-analysis and public database analysis. Based on the ICGC-AGRO database, we have constructed a risk scoring model for predicting metachronous metastasis in KRAS-mutant CRC using machine learning, which was validated in public datasets. Furthermore, we discovered that KRAS inhibitors can effectively suppress the migration and invasion capabilities of high risk CRC cells by wound healing assay and transwell experiment, which was validated ex vivo and in vivo using organoids and splenic liver metastatic mouse model. Finally, RNA sequencing, RT-qPCR, and western blots have proven that KRAS inhibitors can suppress the epithelial-mesenchymal transition (EMT) and transforming growth factor beta (TGF-beta) signaling pathway in CRC. Collectively, our study suggests that KRAS inhibitors may inhibit the metastatic ability of CRC by suppressing EMT and the TGF-beta pathway, suggesting their potential for the prevention of metachronous metastasis in CRC with KRAS mutation.

Background: Colorectal cancer (CRC) is currently the third most common cancer worldwide, with high heterogeneity and poor prognosis. Gene expression-based molecular subtypes can effectively dissect tumor heterogeneity, but their clinical translation remains challenging. This study aims to conduct radiogenomic analysis regarding molecular subtypes and establish prognostic signatures for survival prediction of colorectal cancer. Methods: In this retrospective study involving 2948 CRC patients from 8 cohorts, we utilized a supervised deep learning framework to extract quantitative feature representations of molecular subtypes. Through correlation analysis, we selected key gene expression features related to these subtypes to establish a prognostic signature. A similar pipeline was applied to derive a non-invasive radiomic prognostic signature. Finally, we validated the prognostic value of both signatures in multiple cohorts and explored their biological interpretation. Results: We successfully established a molecular subtype-associated gene signature and a non-invasive radiogenomic signature. The gene signature classified patients into high-risk and low-risk groups with significantly different prognoses. The low-risk group had a better prognosis and showed a greater potential benefit from immunotherapy. Similarly, the radiogenomic signature exhibited characteristics related to molecular subtypes and comparable performance in prognostic prediction. Multivariate analysis confirmed the independent prognostic value of both signatures. In summary, this retrospective study demonstrates that our framework translates molecular subtypes into cost-effective biomarkers for risk stratification and treatment guidance.