In metastatic colorectal cancer (mCRC), resistance to EGFR-targeted therapy is associated with MAPK pathway activating mutations. As a result, mCRC patients with such mutations, like oncogenic RAS, are currently excluded from anti-EGFR targeted therapy. However, an increasing amount of evidence supports the notion that not all RAS oncogenes are equal, but impose isoform- and mutation-specific phenotypes in a context-dependent manner. Di-verging phenotypes are likely to exist for most types of activating mutations. Unfortunately, these concepts are challenging to support with clinical evidence, as ideal stratification of patient cohorts per mutation type would severely dilute the desired patient numbers to draw significant conclusions.
In this thesis we have investigated the similarities and discrepancies between various onco-genic MAPK pathway mutations (RASGAP deficiencies, BRAF mutants and fusions, and RAS oncogenes) that are prevalent in CRC progression and therapy resistance with the aim to improve future patient stratification for treatment regimens. We used patient-derived or-ganoids from CRC samples that maintain the histopathological features of the native tumor, including high concordance of somatic mutations, transcriptome and drug response bet-ween matched primary tumors and derived organoids cultures. Subsequently, patient-derived CRC organoids were genetically modified using CRISPR/Cas9 technology to introduce oncogenic mutations of interest in an identical genetic background for the investigation of mutation-specific effects on tumor growth and therapy resistance.