BACKGROUNDColorectal cancer is a complex disease with high mortality rates. Over time, the treatment of metastatic colorectal cancer (mCRC) has gradually improved due to the development of modern chemotherapy and targeted therapy regimens. However, due to the inherent heterogeneity of this condition, identifying reliable predictive biomarkers for targeted therapies remains challenging. A recent promising classification system-the consensus molecular subtype (CMS) system-offers the potential to categorize mCRC patients based on their unique biological and molecular characteristics. Four distinct CMS categories have been defined: immune (CMS1), canonical (CMS2), metabolic (CMS3), and mesenchymal (CMS4). Nevertheless, there is currently no standardized protocol for accurately classifying patients into CMS categories. To address this challenge, reverse transcription polymerase chain reaction (RT-qPCR) and next-generation genomic sequencing (NGS) techniques may hold promise for precisely classifying mCRC patients into their CMSs. AIMTo investigate if mCRC patients can be classified into CMS categories using a standardized molecular biology workflow. METHODSThis observational study was conducted at the University of Chile Clinical Hospital and included patients with unresectable mCRC who were undergoing systemic treatment with chemotherapy and/or targeted therapy. Molecular biology techniques were employed to analyse primary tumour samples from these patients. RT-qPCR was utilized to assess the expression of genes associated with fibrosis (TGF-beta and beta-catenin) and cell growth pathways (c-MYC). NGS using a 25-gene panel (TumorSec) was performed to identify specific genomic mutations. The patients were then classified into one of the four CMS categories according to the clinical consensus of a Tumour Board. Informed consent was obtained from all the patients prior to their participation in this study. All techniques were conducted at University of Chile. RESULTSTwenty-six patients were studied with the techniques and then evaluated by the Tumour Board to determine the specific CMS. Among them, 23% (n = 6), 19% (n = 5), 31% (n = 8), and 19% (n = 5) were classified as CMS1, CMS2, CMS3, and CMS4, respectively. Additionally, 8% of patients (n = 2) could not be classified into any of the four CMS categories. The median overall survival of the total sample was 28 mo, and for CMS1, CMS2, CMS3 and CMS4 it was 11, 20, 30 and 45 mo respectively, with no statistically significant differences between groups. CONCLUSIONA molecular biology workflow and clinical consensus analysis can be used to accurately classify mCRC patients. This classification process, which divides patients into the four CMS categories, holds significant potential for improving research strategies and targeted therapies tailored to the specific characteristics of mCRC.