2327P - The impact of the microenvironment on the intratumoral cellular heterogeneity in colorectal cancer

Background

Colorectal cancer (CRC) is a major public health issue, as it is amongst the most common forms of cancer incidence and mortality worldwide. Lately, intratutmoral cellular heterogeneity has been highlighted for survival and drug response evaluation. In 2015, the consensus molecular subtype system (CMS) surfaced as a potential diagnostic and prognostic tool for CRC. The CMS classification is based on gene expression profiles and recognizes four subtypes. Whereas CMS2/3 is characterized by high Wnt signaling, and Myc activity, CMS4 is defined by its TGF-β signaling, epithelial-mesenchymal transition (EMT), fibroblast accumulation, and overall worst prognosis. I examined the influence of the tumoral microenvironment on the intratumoral cellular heterogeneity of CRC.

Methods

Patient-derived organoids (PDO) model intratumoral heterogeneity of CRC effectively. PDOs were cultured from CRC surgical biopsy samples, and grown alone or with fibroblasts, in Matrigel or in collagen-enriched gel, with growth medium enriched or depleted from amino acids or glucose. The accumulation of collagen in the tumors is negatively correlated with patient survival. The expression levels of markers for CMS2/3 and CMS4 were evaluated with RT-qPCR, immunohistochemistry, and flow cytometry.

Results

We observed a higher rate of CRC cell positivity for CMS2/3 and CMS4 markers, along with their double positivity when co-cultured with fibroblasts. Similarly, the mixed phenotype positivity increased for CRC cells cultured in amino acid or glucose-free conditions compared to standard growth medium. In collagen, only the CMS4 marker positivity was higher compared to those in Matrigel. Furthermore, cocultured organoids demonstrated an increased positivity for the EMT markers.

Conclusions

Altering the microenvironment of CRC significantly modifies the intratumoral cellular heterogeneity, as collagen and fibroblast enrichment shifts the organoids to a CMS4 phenotype with partial EMT activity, both of which are suggestive of worse prognosis.

Clinical trial identification

Editorial acknowledgement

Legal entity responsible for the study

The authors.

Funding

Hungarian National Research, Development and Innovation Office & Hungarian Ministry of Innovation and Technology.

Disclosure

All authors have declared no conflicts of interest.