Milea Timbergen

44 A major challenge remains to study DTF in the preclinical setting. This is partly due to the rarity of the tumour type, but also to the limited availability of pure DTF cell lines and other cell and animal models 3 . Culturing a fresh DTF resection specimen, inevitably leads to an overgrowth of WT fibroblasts and concomitant loss of DTF tumour cells. Separating tumour cells from their surrounding stromal cells remains challenging and time consuming. Even if a “pure” DTF cell line is obtained, DTF tumour cells often reach senescence after several passages. To our knowledge, no studies investigating the effect of immortalization protocols on primary DTF cells have been published yet. Experimenting with primary cultures consisting of both stromal cells and tumour cells is an alternative but has its drawbacks. Additionally, representative cellular and animal models of DTF (e.g., organoids or mice expressing mutated CTNNB1 ) in relevant tissues are difficult to generate and expensive to maintain. The existing Apc + /Apc 1638N mouse model has already been proven as a useful model for FAP-associated DTF and is often used as a tumour model for non-FAP related DTF (Table 1) 61, 146 . A mouse DTF model based on specific β-catenin mutations is, to our knowledge, currently not available. A recently developed genetically engineered Xenopus tropicalis model harbouring a mutated APC, may yield another DTF tumour model that can be exploited as a platform to define novel therapeutic targets and preclinical validation studies 147 . Well-defined preclinical models are as necessary as well-annotated large series of DTF tumour samples, to better understand DTF biology and to provide experimental support and rationale for translational research investigating the inhibition of signalling pathways in DTF. Additionally, signalling pathways are often seen as separate entities, however, in reality cross-talk occurs between different pathways. The precise interactions between different signalling- and biochemical pathways is complex and still poorly understood. Aberrant signalling of one pathway can often be corrected via compensatory mechanisms in another pathway 148 . In DTF tumours and cell lines, the Wnt, Notch and Hh pathways have been shown to be involved in cross-talk, implicating optimal therapeutic efficacy, is reached when all interacting pathways are inhibited in a combinatorial approach 149 . Future studies should not focus on individual signalling cascades but rather on the simultaneous inhibition of multiple pathways. Furthermore, clinical studies to evaluate the efficacy of systemic and targeted treatments without any randomization procedures remain challenging in current clinical practice as they are often difficult to interpret. Due to the unpredictable growth behaviour of DTF with reports of spontaneous regression without treatment and stable disease, it is difficult to 2