Methylation profiling reveals novel molecular classes of rhabdomyosarcoma

Abstract

Rhabdomyosarcomas (RMS) represent a family of aggressive soft tissue sarcomas that present in both children and adults. Pathologic risk stratification for RMS has been based on histologic subtype, with poor outcomes observed in alveolar rhabdomyosarcoma (ARMS) and the adult-type pleomorphic rhabdomyosarcoma (PRMS) compared to embryonal rhabdomyosarcoma (ERMS). Genomic sequencing studies have expanded the spectrum of RMS, with several new molecularly defined entities, including fusion-driven spindle cell/sclerosing rhabdomyosarcoma (SC/SRMS) and MYOD1-mutant SC/SRMS. Comprehensive genomic analysis has previously defined the mutational and copy number spectrum for the more common ERMS and ARMS and revealed corresponding methylation signatures. Comparatively, less is known about epigenetic correlates for the rare SC/SRMS or PRMS histologic subtypes. Herein, we present exome and RNA sequencing, copy number analysis, and methylation profiling of the largest cohort of molecularly characterized RMS samples to date. In addition to ARMS and ERMS, we identify two novel methylation subtypes, one having SC/SRMS histology and defined by MYOD1 p. L122R mutations and the other matching adult-type PRMS. Selected tumors from adolescent patients grouped with the PRMS methylation class, expanding the age range of these rare tumors. Limited follow-up data suggest that pediatric tumors with MYOD1-mutations are associated with an aggressive clinical course.

Cell-Free DNA as a Diagnostic and Prognostic Biomarker in Pediatric Rhabdomyosarcoma

“…Using data from Clay et al, 33 Koelsche et al, 34 and Seki et al, 20 the mean beta value of RASSF1A -M was 0.550 (range, 0.032-0.933; Data Supplement). We next examined the presence of RASSF1A-M in plasma using ddPCR.…”

“…One of the limitations of this study was the absence of paired primary tumor samples. However, the presence of RASSF1A -M, as extracted from data published by several groups, 20 , 33 , 34 indicated the potential to detect RASSF1A -M in primary tumors, with admittedly a large variation in the level of RASSF1A- M. Still, for the patients in our cohort who were RASSF1A -M‒negative, on the basis of cfDNA obtained at diagnosis, we were unable to determine whether this was due to absence of RASSF1A methylation or no detectable ctDNA. This is underlined by the 18 samples testing negative for RASSF1A -M, in which ctDNA was detected by cfRRBS and/or shWGS.…”

“…In all ddPCR assays, total cfDNA was determined using the reference gene ACTB . Since there was no matched tumor material available, we used data on RASSF1A -M in rhabdomyosarcoma tumors from published data sets from Clay et al, 33 Koelsche et al, 34 and specifically requested data from Seki et al 20 Data from Clay and Koelsche were analyzed in R2. 35 We focused on hypermethylation of the promotor region of RASSF1A as this is typically hypermethylated in cancer.…”

Complex Elucidation of Cells-of-Origin in Pediatric Soft Tissue Sarcoma: From Concepts to Real Life, Hide-and-Seek through Epigenetic and Transcriptional Reprogramming

“…DNA methylation profiles of STS was proven useful to improve lineage classification and to reveal methylome patterns that were specific of tumor types or stages [ 3 , 28 ]. These results allowed the distinction among subtypes of RMS [ 133 ], angiosarcoma [ 134 ] or small blue round cell sarcomas [ 135 ] for example. In addition, specific methylation profiles correlated with diverse clinical outcomes in dedifferentiated liposarcoma [ 96 ].…”

“…Thus, manipulation of the expression level of the PAX3-FOXO1 transgene by using the histone deacetylase inhibitor entinostat potentiates the effect of actinomycin D, only when this fusion gene is expressed in Pax7+ satellite cells in the postnatal period [ 88 ]. If this statement needs to be tempered considering the important epigenetic reorganization undergone by tumor cells during the transformation and escape oncogenic process, which reduces the impact of the original cellular context, the approaches of personalized medicine should probably be rethought beyond the framework of genetics alone [ 133 , 143 , 144 ].…”