Abstract 987: Proteomics identifies differences in the biological phenotype of tumor deposits and lymph node metastases in colorectal cancer
Authors: Brouwer Nelleke P. , Webbink Loth , Van Lent-Van Vliet Shannon , Jansen Pascal W. , Simmer Femke , Tauriello Daniele V. , Vermeulen Michiel , Nagtegaal Iris D. 
Published: June 2022

Background: Distant metastatic disease remains the main cause of colorectal cancer (CRC) mortality, but our knowledge on how cancer cells travel from the primary tumor to a secondary site is limited. Lymph node metastases (LNM) play a central role in the staging of CRC patients since they are currently viewed as the gateway to distant metastases. However, in recent years additional forms of locoregional spread with varying prognostic impact have been identified, with tumor deposits (TD) having significantly more impact than LNM. The biological explanation for the difference in prognostic impact between LNM and TD is lacking, while this is essential for a more comprehensive understanding of the role of these biomarkers CRC spread. Therefore, in this study, shotgun proteomics were used to compare the biological phenotype of TD and LNM.

Methods: From 12 CRC patients, one TD and one LNM were included for paired protein analyses. Proteins were isolated from formalin-fixed paraffin-embedded tissue after which a filter aided sample preparation method was performed. Then, the samples were analyzed by liquid-chromatography tandem mass-spectrometry, and after data handling and filtering, 5578 differentially expressed proteins were used for downstream analyses. Differences in expression were visualized using heatmaps and volcano plots, and enrichment analyses as well as gene set enrichment was performed. All analyses were performed using R version 4.1.2.

Results: Among the main findings was that the proteins ASPN, MRC2, SPON1, MXRA5, BGN, LUM, and PLOD2 were overexpressed in TD. These proteins play a role in stimulating pro-tumorigenic processes such as migration, invasion, and epithelial to mesenchymal transition. For the enrichment analyses all proteins with a Fold change of less than -1 and more than 1 were included. Enrichment analyses using the KEGG pathways showed more immune activity in the LNM, with upregulated B-cell and T-cell signaling (p&lt;0.001), while TD showed more interaction with the extracellular matrix in the form of proteoglycans, focal adhesion, and ECM-receptor interaction (p&lt;0.01). In addition, when hallmarks from the Molecular Signatures Database (MSigDB) were used for the enrichment analyses, TD showed the hallmark of epithelial mesenchymal transition (p &lt;0.005).

Conclusions: This study shows that TD have a more aggressive and invasive phenotype compared to LNM on protein level. TD had a higher expression of proteins that promote pro-tumorigenic processes and a more extensive interaction with the extracellular matrix. Furthermore, the hallmark of epithelial mesenchymal transition is enriched in TD compared to LNM. These findings form a possible explanation for the strong prognostic impact of TD, and give insight into the heterogeneity of different modes of locoregional spread in CRC.

Citation Format: Nelleke P. Brouwer, Loth Webbink, Shannon Van Lent-Van Vliet, Pascal W. Jansen, Femke Simmer, Daniele V. Tauriello, Michiel Vermeulen, Iris D. Nagtegaal. Proteomics identifies differences in the biological phenotype of tumor deposits and lymph node metastases in colorectal cancer [abstract]. In: Proceedings of the American Association for Cancer Research Annual Meeting 2022; 2022 Apr 8-13. Philadelphia (PA): AACR; Cancer Res 2022;82(12_Suppl):Abstract nr 987.