PAX3-FOXO1 transgenic zebrafish models identify HES3 as a mediator of rhabdomyosarcoma tumorigenesis

Abstract

Alveolar rhabdomyosarcoma is a pediatric soft-tissue sarcoma caused by PAX3/7-FOXO1 fusion oncogenes and is characterized by impaired skeletal muscle development. We developed human PAX3-FOXO1 -driven zebrafish models of tumorigenesis and found that PAX3-FOXO1 exhibits discrete cell lineage susceptibility and transformation. Tumors developed by 1.6–19 months and were primitive neuroectodermal tumors or rhabdomyosarcoma. We applied this PAX3-FOXO1 transgenic zebrafish model to study how PAX3-FOXO1 leverages early developmental pathways for oncogenesis and found that her3 is a unique target. Ectopic expression of the her3 human ortholog, HES3, inhibits myogenesis in zebrafish and mammalian cells, recapitulating the arrested muscle development characteristic of rhabdomyosarcoma. In patients, HES3 is overexpressed in fusion-positive versus fusion-negative tumors. Finally, HES3 overexpression is associated with reduced survival in patients in the context of the fusion. Our novel zebrafish rhabdomyosarcoma model identifies a new PAX3-FOXO1 target, her3/HES3, that contributes to impaired myogenic differentiation and has prognostic significance in human disease.

CIC-DUX4 expression drives the development of small round cell sarcoma in transgenic zebrafish: a new model revealing a role for ETV4 in CIC-mediated sarcomagenesis

“…The tol2 kit β -actin promoter, 3' SV40 late poly A signal construct, and pDestTol2pA2 destination vector were used for construct generation and expression in zebrafish (31). The GFP viral 2A sequence was a kind gift from Steven Leach (32), and was previously sub-cloned into a middle entry Gateway expression system (33). Tol2 mRNA was synthesized from pCS2FA-transposase from Koichi Kawakami (34).…”

The PAXFOXO1s trigger fast trans-differentiation of chick embryonic neural cells into alveolar rhabdomyosarcoma with tissue invasive properties limited by S phase entry inhibition

“…In addition, our study supports the idea that the combination of genes induced by PAXFOXO1s is highly specific and reminiscent of that found in ARMS cells. Intriguingly, PAXFOXO1s activity can largely bypass the genomic constrain imposed by the origin of cell in which they act and can impose ARMS signature in various cell subtypes, including fibroblasts, mesenchymal stem cells or myoblasts, as well as various vertebrate species including chick, zebrafish, mouse and human [12,29,31,32](our data). This is further sustained by nonnegligible overlap of PAX bound CRMs identified in discrete ARMS cell lines [12].…”

“…Despite the apparently powerful activity of PAXFOXO1s, data from animal models have led to the conclusion that the fusion proteins are not efficient in triggering ARMS formation and spreading [24,[28][29][30][31]. In excess of 60 days are required for grafted PAX3FOXO1 expressing human myoblasts or mesenchymal stem cells to produce significant ARMS like growths in mice.…”

“…Importantly, these in vivo approaches have revealed several parameters enhancing PAXFOXO1 proteins oncogenicity. The random insertion of transgenes in zebrafish indicated that neural derived tissues are more prone than mesodermal derived tissues to produce tumours when exposed to PAX3FOXO1, highlighting the differential response of distinct cell lineages [29]. In addition, zebrafish and mouse models both indicate that a minimal level of PAX3FOXO1 needs to be reached to observe tumourigenesis [28,29,32].…”

“…In addition, zebrafish and mouse models both indicate that a minimal level of PAX3FOXO1 needs to be reached to observe tumourigenesis [28,29,32]. Finally, complementing PAXFOXO1s expression with genetic aberrations promoting cell cycle progression markedly accelerated and increased the frequency of tumour formation [28][29][30][31][32][33][34]. This was notably achieved by lowering the expression of p53 or the retinoblastoma protein, RB1; or conversely by ectopically elevating MYCN expression or RAS activity.…”