A Genomic and Epigenetic Comparative Study of Low-grade Biphenotypic Sinonasal Sarcoma with Metachronous and Synchronous High-grade Rhabdomyosarcomatous Transformation

Background: Biphenotypic sinonasal sarcoma (BSNS) is usually histologically distinguishable in its conventional form based on morphology and ancillary studies. However, a subset of BSNS loses characteristic histologic features over time and instead undergoes high-grade rhabdomyosarcomatous transformation (HGRT). The fact that the driver PAX3 fusion of BSNS overlaps with that of other tumors, e.g., alveolar rhabdomyosarcoma, further complicates this diagnostic dilemma. DNA methylation profiling has emerged as a helpful tool in classifying poorly differentiated malignancies. We sought to investigate whether conventional BSNS and HGRT retain the same epigenetic signature over time. Materials and Methods: Five pure BSNS samples from the initial diagnostic specimens and 4 HGRT components (3 from the long-term recurrence and 1 synchronous) were collected. An immunohistochemical panel of smooth muscle, skeletal muscle, and neural markers was performed and scored as positive or negative. Molecular analyses included RNA-based (4 cases) and DNA-based (2 cases) next-generation sequencing. DNA methylation profiling using the Illumina Infinium EPICv2 microarray was performed, and the methylomes from paired BSNS and HGRT components were compared with a control cohort of BSNS and various high-grade sarcomas. Results: The tumors arose from 3 females and one male with an age range of 38–88 years (median 70.5 years old). The time interval between initial BSNS diagnosis to HGRT was between 0 and 14 years (median 8 years). The BSNS components were diffusely positive for S-100 and SMA (4/4), while the HGRT components were positive for desmin (4/4) myogenin (3/4) and Myo D1 (1/1). By RNA sequencing, 2 showed PAX3::FOXO1 fusion and 2 showed PAX3::MAML3 fusion. Compared to the low-grade BSNS, the HGRT from the same patients acquired additional secondary genomic alterations and significantly more frequent chromosomal arm-level copy number variations. DNA methylation profiling demonstrated that in 3 patients, both low-grade BSNS and HGRT converged with conventional BSNS. In one patient with PAX3::FOXO1 fusion, the HGRT from a late recurrence 14 years since initial diagnosis overlapped epigenetically with embryonal rhabdomyosarcoma. Conclusion: The epigenetic signature of BSNS is usually preserved in cases that underwent HGRT, even after many years from the initial onset of BSNS. However, in rare instances, the HGRT component may display epigenetic divergence over time.