High Expression of the PAX3-FKHR Oncoprotein Is Required to Promote Tumorigenesis of Human Myoblasts

Abstract

PAX3-FKHR is a fusion oncoprotein generated by the 2;13 chromosomal translocation in alveolar rhabdomyosarcoma (ARMS), a cancer associated with the skeletal muscle lineage. Previous studies determined that high-level PAX3-FKHR expression is a consistent feature in ARMS tumors. To investigate the relationship between expression and phenotype in human myogenic cells, PAX3-FKHR was introduced into immortalized human myoblasts to produce a low overall PAX3-FKHR expression level. Although PAX3-FKHR alone failed to exert transforming activity, a combination of PAX3-FKHR and MYCN induced transforming activity in cell culture assays. Furthermore, myoblasts expressing PAX3-FKHR with or without MYCN formed tumors in SCID mice. These tumors demonstrated invasive features and expressed myogenic markers, consistent with rhabdomyosarcoma. Comparisons of tumor and parental cells revealed that only a subset of parental cells developed into tumors and that tumor cells expressed high PAX3-FKHR levels compared with transduced parental cells. Subcloning of parental PAX3-FKHR/MYCN-transduced myoblasts identified rare high PAX3-FKHR-expressing subclones with high transforming and tumorigenic activity; however, most subclones expressed low PAX3-FKHR and showed neither transforming nor tumorigenic activity. Finally, RNA interference experiments in myoblast-derived tumor and ARMS cells revealed that high PAX3-FKHR expression plays a crucial role in regulating proliferation, transformation, and differentiation. These findings support the premise that high PAX3-FKHR-expressing cells are selected during tumorigenesis.

Antitumor Activity of Sustained N-Myc Reduction in Rhabdomyosarcomas and Transcriptional Block by Antigene Therapy

“…The correlation of N-Myc expression with clinical outcome in cases with alveolar histology is consistent with fusion gene-positive cases, which express N-Myc highly, generally having a poorer outcome than fusion gene-negative alveolar cases (6,7,9). Seventy-seven percent of fusion-positive samples were immunohisto- Both N-Myc and the fusion proteins are known to play pivotal and co-operative roles in the biology of ARMS (7,10,11,30,33,34). An association of N-Myc expression with fusion gene positivity is consistent with previous studies that have shown that MYCN is a direct downstream target of the PAX3-FOXO1 fusion protein (7,10,29,30).…”

Identification of target genes of PAX3-FOXO1 in alveolar rhabdomyosarcoma

“…However, in addition to these effects, PAX3-FOXO1 can also cause growth suppression and cell death in other settings (20,21,38). These paradoxical features of PAX3-FOXO1 being both oncogenic and growth-suppressive were demonstrated in previous studies with immortalized murine fibroblasts (20,21) and human myoblasts (38).…”

“…However, in addition to these effects, PAX3-FOXO1 can also cause growth suppression and cell death in other settings (20,21,38). These paradoxical features of PAX3-FOXO1 being both oncogenic and growth-suppressive were demonstrated in previous studies with immortalized murine fibroblasts (20,21) and human myoblasts (38). We identified and validated that tumor-suppressor genes, GREM1 (39) and DAPK1 (40), are upregulated at both RNA and protein levels in PAX3-FOXO1-positive ARMS tumors and PAX3-FOXO1-ER inducible cell culture systems.…”

PAX3-FKHR Regulates the Expression of Pleiotrophin to Mediate Motility in Alveolar Rhabdomyosarcoma Cells

“…The unique expression, function, and subcellular location of PAX3-FKHR contribute to its oncogenic behavior by modifying cell growth, differentiation, and migration [1]. Various evidence suggests that PAX3-FKHR regulates cell migration, contributing to the high propensity of ARMS to metastasize [2–4]. …”