Embryonal rhabdomyosarcoma (ERMS) is a devastating malignancy of muscle that is diagnosed in hundreds of children and adults annually in the United States. Survival rates are less than 30% in patients with unresectable, metastatic, or relapsed RMS, with continued tumor growth and metastasis being initiated by a subset of cells called tumor propagating cells (TPCs). Yet, to date, targeted approaches to kill TPCs or to differentiate them into non-proliferative, differentiated ERMS cell types have not been developed. The long- term goal of our work is to uncover therapeutically relevant pathways that curb ERMS growth by killing or differentiating the TPCs. The overall objective of this application is to determine the extent to which the NOTCH1 pathway regulates self-renewal within TPCs and metastasis of ERMS. Our central hypothesis is that NOTCH1 pathway activation supports ERMS growth by specifically inducing self-renewal while also elevating the proclivity for metastasis. Our preliminary data indicate a prominent role for NOTCH1 in regulating TPC self- renewal in both zebrafish and human ERMS. Importantly, NOTCH1 pathway inactivation also led to a dramatic tumor shrinkage of human ERMS xenografts, while NOTCH1 pathway activation induced metastatic progression in our zebrafish tumor model. The rationale underlying our research is that the NOTCH1 pathway is active in 60% of human ERMS, is linked with a poor outcome, and is required for continued tumor growth in vivo, suggesting that targeting this pathway would benefit a large fraction of high-risk patients.
Aim 1 will dynamically visualize the effects of NOTCH1 pathway activation on ERMS self-renewal and metastasis in a fluorescent-transgenic zebrafish model that accurately recapitulates the molecular and histopathogenesis of the human disease. These experiments are designed to establish that NOTCH1 increases self-renewal and elevates metastatic progression.
Aim 2 will elucidate the molecular mechanisms regulated by intracellular NOTCH1 (ICN1) in human ERMS, by testing whether the ICN1/SNAIL1 axis enhances human ERMS self- renewal by suppressing terminal differentiation through MEF2C and whether ICN1 is stabilized intracellularly by loss of FBXW7, a ubiquitin-ligase known to degrade intracellular NOTCH1 and which is mutationally inactivated in a subset of human ERMS.
Aim 3 will assess NOTCH1 antibody inhibitors for their preclinical efficacy in patient derived xenografts of human ERMS. Our work will uncover the molecular pathways by which ICN1 drives ERMS growth, self-renewal and metastasis. Such insights will provide new biomarkers for assessing drug effects on TPCs and will likely identify novel drug targets beyond NOTCH1 for the treatment of ERMS. Our work is predicted to have a large positive translational impact, as it will directly address the feasibility and likely benefit of using new NOTCH1 blocking antibody to treat human ERMS. These findings will be relevant to applying these same antibody therapies to a wide range of NOTCH1-dependent human tumors.

Public Health Relevance

The NOTCH1 pathway is active in a majority of human rhabdomyosarcoma and has potent roles in elevating self-renewal, growth, and likely metastasis in the embryonal RMS subtype (ERMS). Here, we will uncover the molecular mechanisms by which the NOTCH1 pathway regulates ERMS growth. We will also directly assess the therapeutic benefit of NOTCH1 inhibitory antibodies in preclinical patient-derived xenograft models of ERMS.

National Institute of Health (NIH)
National Cancer Institute (NCI)
Research Project (R01)
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Cancer Molecular Pathobiology Study Section (CAMP)
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Xu, Wanping
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Massachusetts General Hospital
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