Elucidating the survival mechanisms of rhabdomyosarcoma

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PROJECT SUMMARY/ABSTRACT Rhabdomyosarcoma (RMS) is the most common soft tissue cancers affecting children. Despite aggressive combinatorial therapy, nearly 30% of children diagnosed with RMS will succumb to either metastasis or recurrence. While RMS can occur anywhere in the body, all cases are defined by the expression of skeletal muscle markers. Skeletal muscle differentiation is controlled by a family of four DNA binding myogenic regulatory factors - MyoD, Myf5, myogenin and MRF4. The prevailing belief is that RMS tumor cells are unable to complete differentiation due to the dysfunction of these myogenic factors. Our laboratory previously showed that NF-κB contributes to RMS by maintaining tumor cells in an undifferentiated state. Interestingly, even though NF-κB is widely known to function in cancer as a survival factor, RMS appears to be unique in not needing NF-κB to overcome stress-induced apoptosis. Instead, we find that MyoD is able to compensate for NF-κB. One mechanism by which MyoD promotes survival is by keeping RMS cells in a partially differentiated state. Another mechanism, which we elucidated through a series of transcriptomic and bioinformatic analyses, indicates that MyoD is capable of suppressing apoptotic genes. Unique to this latter mechanism is that suppression requires promoter methylation. Based on these data, I hypothesize that MyoD functions in RMS by acting as an oncogene to promote cell survival, which occurs by 1) maintaining RMS cells in a partially differentiated state and 2) suppressing apoptotic genes through the regulation of DNMTs and promoter hypermethylation. The second part of this hypothesis will be explored in this proposal through two aims. In Aim 1, I plan to characterize the pro- apoptotic genes suppressed by MyoD and their relevance in RMS. I will accomplish this through gain and loss of function studies for each gene and observing the death response phenotype. I will also explore the methylation of the gene promoters in response to MyoD expression or treatment with compounds that regulate methylation. In Aim 2, I will perform preclinical studies to test the efficacy of demethylation agents as a therapeutic strategy in RMS. Both in vitro and xenograft tumor studies will be utilized, using demethylation agents alone or in combination with RMS standard of care drugs. The impact of my findings might not only demonstrate a novel role of MyoD in RMS, but might also advance novel therapeutic strategy for the treatment of RMS patients.