The aggressive colorectal cancer subtype marker HTR2B has a dual role depending on the tumor microenvironment.

Colorectal cancer (CRC) is the second-third most frequent cancer type in the Western countries, with significant inter- and intra-patient cellular heterogeneity. The transcriptomics-based CRC classification identified a cluster of patients with epithelial-mesenchymal transition (EMT), the accumulation of fibroblasts, and dismal prognosis. Although the serotonin receptor HTR2B is thought to be one of the epithelial markers of this aggressive CMS4 subgroup, its precise role in CRC tumorigenesis is still largely unknown. By using patient-derived organoids (PDO), we prove the heterogeneous expression of this receptor that is under the control of the mTOR pathway. Unfavorable conditions, applying the clinically used 5-fluorouracil, co-culturing with fibroblasts, and collagen-I accumulation in the extracellular matrix (ECM) increased the number of HTR2B + cells. Interestingly, whereas stimulating HTR2B had no effect on PDOs under normal conditions, it reduced cellular survival under unfavorable conditions when carbohydrates were lacking. In contrast, serotonin and an HTR2B agonist induced invasion of HTR2B + tumor cells only in collagen-I, representing a permissive ECM for cellular migration. CRC cells positive for HTR2B also expressed the NOTCH3 receptor and NOTCH target genes at a higher intensity than cells with low HTR2B levels. Similar to HTR2B + organoids, NOTCH3 + PDOs had increased EMT markers and HTR2B levels. Furthermore, blocking NOTCH activity inhibited the serotonin-induced invasion. Collectively, our results show that the serotonin receptor HTR2B marks a CRC cell population with increased proliferative capacity, invasive potential, and partial EMT, and they highlight the critical role of ECM in shaping the effect of serotonin in CRC.