Lay Abstract
Desmoid-type fibromatosis (DF) is a rare benign tumor characterized by fibroblastic features and a variable and often unpredictable clinical course. Despite lacking metastatic potential, DF is locally invasive and with a high local recurrence rate. Desmoid-type fibromatosis is often associated with local and repeated injuries; it shows differences in growth rate, spontaneous regression, and stabilization of disease progression. It occurs both sporadically and in patients with familial adenomatous polyposis (FAP) and it is well known that alterations of the Wnt/b-catenin pathway can promote its onset. However, the wide variability of the growth rate, localization and aggressiveness of the tumor indicate that the desmoid tumor requires a favorable microenvironment to develop and grow. Different desmoid cell samples show a generally shared response to external microenvironmental stimuli, but with a considerable degree of variability and this could at least partially correlate to the clinical heterogeneous outcome of the desmoid tumors. We demonstrated that the inflammatory cytokine, TGF-b, plays a key role in the proliferation and myofibroblast differentiation of DF cells and it regulates cell-adhesion and cell-cell interactions in DF myofibroblast differentiation. However, primary cells from desmoid tumors are difficult to treat and investigate because of the limited biological material and for the variability between samples. For this reason, our aim is to create simple different desmoid-like cell models with single or few molecular alterations that ensure genomic and phenotypic stability and that allow us to perform extensive studies. We will use the CRISPR/Cas9 system to induce efficient, specific, stable and multiplexed gene-editing in the Wnt pathway genes to generate controlled cell models that could in part or more widely mimic DF cells behavior.
Scientific Abstract
Desmoid-type fibromatosis (DF) is a mesenchymal tumor unable to metastasize, but locally invasive and with a high-local recurrence rate (Alman et al., 1997; Lewis et al., 1999; Escobar et al., 2012). It occurs both sporadically and in patients with familial adenomatous polyposis (FAP) and it is well known that alterations of the Wnt/b-catenin pathway can promote its onset. The frequent association of this type of tumor with local and repeated injuries, the spontaneous regression and stabilization of disease progression make the clinical course unpredictable and the study of the shared molecular characteristics very puzzling. Different DF cell samples show a generally shared response to external factors, but with a considerable degree of variability and this could at least partially reflect the clinical heterogeneous outcome of the desmoid tumors, with cases of continuous growth, fluctuation, stabilization but also of regression of the tumor. The combination of the intrinsic biological characteristics of the desmoid tumor cells and the host microenvironment appear to be critical and to account for the diversity of clinical outcomes and the recurrence risk of desmoid tumors. One additional aspect of primary cultures from tumor biopsies is the concomitant presence of neoplastic and reactive tissues with a varying degree of the cell populations. Considering the high similarities between the neoplastic DF cells and normal activated fibroblasts it becomes even more difficult to isolate DF cells alone and select their peculiarities. For these reasons we opted to create a more controlled environment where to study the effects of isolated genetic changes in the Wnt pathway and the effect that these changes have on the response to external stimuli. More in particular, we decided to use the CRISPR/Cas9 system to induce efficient, specific, stable and multiplexed gene-editing in the Wnt pathway genes to generate controlled cell models that could in part or more widely mimic DF cells behavior. The project aims, at initially, developing stable cell lines with mutations in the APC and CTNNB genes and to extend the study by altering single Wnt pathway genes expression in these two cell lines together with the non-modified cells. The initial cell lines will be then characterized both by phenotype and genotype and in particular their response to external stimuli. Based on the results of these studies we will attempt to recreate the DF phenotype without altering the CTNNB or APC gene.
