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Abstract

Colorectal cancer (CRC) is one of the main contributors to cancer-related fatality. Patient survival highly depends on tumour stage at diagnosis, as the 5-year overall survival rate drops drastically from ~64% for patients diagnosed in stage I–II to 12% for patients diagnosed in stage IV. Thus, ~90% of CRC related deaths are associated with metastatic disease that does not respond to currently available systemic treatments. Emerging evidence suggests that inter- and intra-tumour heterogeneity significantly contributes to cancer progression and therapy-resistance. In this PhD thesis, I focus on two main sources of intra-tumour heterogeneity: clonal dynamics and stem cell hierarchy. During CRC progression, the tumour ecosystem is subjected to continuous evolution. However, the patterns of clonal dynamics and their dependence on the surrounding environment are not yet well understood. To investigate this, I used an optical barcoding system (LeGO) in mouse tumour-derived organoids to longitudinally track individual clones in different environments. My findings revealed a key bottleneck both in vitro and in vivo, suggesting that clonal selection plays a major role during CRC progression. Moreover, my results indicate that clonal selection is highly influenced by the tumour microenvironment, which critically contributes to tumour heterogeneity and has implications for therapeutic intervention. Furthermore, CRC cells are hierarchically organized with cancer stem cells (CSCs) at the apex, initiating and fuelling tumour growth. LGR5 has been suggested as the marker for CSCs in CRC. However, a significant number of CRCs present none of few LGR5+ cells. LGR5+ CSCs become absent during metastasis initiation and therapy response. Instead, a foetal intestinal stem cell program is reactivated. In this thesis, I propose TROP2 as a marker for these foetal-like CSCs in CRC and functionally characterized them. My findings suggest a mutually exclusive distribution of LGR5+ and TROP2+ CSC populations in both, mouse and human CRC, implying that they represent distinct CSC populations. I demonstrated that while LGR5 marks CSCs in Apcmut CRCs, TROP2 marks CSCs in Apcwt (WNT-low, serrated) CRCs. Furthermore, TROP2 expression is associated with advanced tumour stages and poor prognosis. Here, I showed that, in CRC, TROP2 marks the cells located at the tumour invasive front as well as the metastasis-initiating cell. Additionally, TROP2 gain-of-function and loss-of-function experiments revealed its direct, but yet unknown, role in CRC progression. Finally, TROP2 represents a promising therapeutic target in CRC due to the potent therapeutic activity of the TROP2 antibody-drug conjugated, Sacituzumab Govitecan (SG). Thus, we have launched a multicentre randomized investigator-initiated trial (phase II/III; NCT06243393) to test SG as a 3rd line treatment versus standard-of-care in metastatic CRC patients.