The role of LEF1 and WNT signaling in growth and differentiation of rhabdomyosarcoma
von Julia Dräger
Datum der mündl. Prüfung:2017-02-02
Erschienen:2017-02-16
Betreuer:Prof. Dr. Heidi Hahn
Gutachter:Prof. Dr. Heidi Hahn
Gutachter:Prof. Dr. Holger Bastians
Zum Verlinken/Zitieren:
http://dx.doi.org/10.53846/goediss-6134
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Name:Dissertation Julia Dräger.pdf
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Zusammenfassung
Englisch
Rhabdomyosarcoma (RMS) is the most common type of soft tissue sarcoma in children and shows characteristics of skeletal muscle differentiation. RMS occur in two main histological subtypes in children: alveolar RMS (ARMS) and embryonal RMS (ERMS), which are associated with distinct genetic and molecular alterations. Despite more intensive therapies during the last decades, patients with metastatic RMS have a very poor prognosis. Thus, it is of uttermost importance to increase the knowledge of the basic biology of RMS and to develop new treatment strategies in order to improve the outcome of the disease. WNT signaling plays an important role in muscle development and differentiation and also contributes to a variety of human diseases including cancer. However, only few data on the role of WNT signaling in RMS have been published and are restricted to β-catenin dependent (canonical) WNT signaling. These data mainly support a tumor suppressive role of canonical WNT signaling in RMS besides promoting myogenic differentiation. Our findings now demonstrate that the prominent transcription factor of canonical WNT signaling LEF1/LEF1 can be highly expressed in primary human ARMS and ERMS samples. However, the common absence of nuclear β-catenin and downregulation of AXIN2 in these RMS samples indicate that canonical WNT signaling is not active and probably inhibited in RMS. Furthermore, our in vivo studies reveal that Wnt3a-driven canonical Wnt/β-catenin signaling does not play a prominent role in RMS pathogenesis in mice. To study the role of LEF1 in RMS in more depth, a stable LEF1 knockdown (LEF1 KD) in the two human ARMS cell lines Rh41 and RMS-13 and in the human ERMS cell line TE671 was established. Our data demonstrate that dependent on the cellular context, LEF1 reduces the aggressiveness of RMS cells. Thus, LEF1 can induce pro-apoptotic signals and can suppress proliferation, migration and invasiveness – especially in the ARMS cell line RMS-13 – both in vitro and in vivo. This less aggressive phenotype is associated with reduced c-MYC and TCFs expression. Furthermore, LEF1 can induce myodifferentiation of RMS-13 cells. Importantly, this seems not to involve canonical β-catenin driven WNT signaling activity. Indeed, despite an upregulation of AXIN2, Wnt3a stimulation does not induce SuperTOPFlash (TOP) reporter activity or nuclear translocation of β-catenin in ARMS cell lines. Together these data indicate that LEF1 has a tumor suppressive function and can induce myodifferentiation in a subset of RMS. This may also involve TCF factors whereas β-catenin activity plays a subordinate role. Interestingly, WNT5A a major player of β-catenin independent (non-canonical) WNT signaling is also overexpressed on protein and on transcript level in primary human ARMS and ERMS samples. Moreover, LEF1 mRNA levels tend to be positively correlated with that of WNT5A in primary human ARMS samples. This is similar to the ARMS cell line RMS-13, in which LEF1/LEF1 expression correlates with that of WNT5A/WNT5A. These data indicate that besides LEF1 also WNT5A might be involved in the pathogenesis and aggressiveness of ARMS. Furthermore, in RMS-13 cells, WNT5A expression is accompanied by activation of PI3K/AKT signaling and LEF1 expression is positively regulated by PI3K. We here propose a new model, in which LEF1 and WNT5A establish a positive feedback loop that involves activation of PI3K signaling and counteracts the aggressiveness of a subset of ARMS, which correlates with downregulation of c-MYC. Recently, activation of β-catenin driven WNT signaling has been proposed as a new treatment option for RMS. However, this may be only of benefit for specific subtypes of RMS, but not for those, in which tumor progression and myogenic differentiation is regulated via LEF1/WNT5A interactions in a non-canonical manner. Together, these data show that further investigations are needed to identify the specific RMS subtypes that may respond to activation of either canonical or LEF1/WNT5A-mediated non-canonical WNT signaling.
Keywords: Rhabdomyosarcoma; WNT signaling; LEF1/TCF; WNT5A signaling; PI3K/AKT signaling
