An Oral Formulation of YK-4-279: Preclinical Efficacy and Acquired Resistance Patterns in Ewing Sarcoma

Abstract

Ewing sarcoma is a transcription factor-mediated pediatric bone tumor caused by a chromosomal translocation of the EWSR1 gene and one of several genes in the ETS family of transcription factors, typically FLI1 or ERG. Full activity of the resulting oncogenic fusion protein occurs only after binding RNA helicase A (RHA), and novel biologically targeted small molecules designed to interfere with that interaction have shown early promise in the preclinical setting. Herein, we demonstrate marked preclinical antineoplastic activity of an orally bioavailable formulation of YK-4-279 and identify mechanisms of acquired chemotherapy resistance that may be exploited to induce collateral sensitivity. Daily enteral administration of YK-4-279 led to significant delay in Ewing sarcoma tumor growth within a murine model. In advance of anticipated early-phase human clinical trials, we investigated both de novo and acquired mechanism(s) by which Ewing sarcoma cells evade YK-4-279-mediated cell death. Drug-resistant clones, formed by chronic in vitro exposure to steadily increased levels of YK-4-279, overexpressed c-Kit, cyclin D1, pStat3(Y705), and PKC isoforms. Interestingly, cross-resistance to imatinib and enzastaurin (selective inhibitors of c-Kit and PKC-b, respectively), was observed and the use of YK-4-279 with enzastaurin in vitro led to marked drug synergy, suggesting a potential role for combination therapies in the future. By advancing an oral formulation of YK-4-279 and identifying prominent mechanisms of resistance, this preclinical research takes us one step closer to a shared goal of curing adolescents and young adults afflicted by Ewing sarcoma.

The biology of DHX9 and its potential as a therapeutic target

“…A small molecule (YK-4-279) blocking the interaction between DHX9 and EWS-FLI1 was found to induce apoptosis in ESFT cells and inhibit tumour growth in xenograph models [197]. YK-4-279 activity has been optimized through pharmacokinetic studies and an orally available formulation has been developed, thus making it a promising candidate for clinical development [198, 199]. …”

Potential approaches to the treatment of Ewing's sarcoma

“…There are several approaches to disrupt the RHA binding to EWS-FLI1, thus reducing the EWS-FLI1 activity and tumorigenesis. Small molecule YK-4-279, which has been shown to interrupt the binding of EWS-FLI1 to RHA, induces apoptosis in ES cells and reduces growth in ES xenografts [62–64](Figure 3). …”

Recent advances in targeted therapy for Ewing sarcoma

“…YK-4-279 drug-resistant clones demonstrated enhanced proliferative rate over their parental cell lines and overexpressed c-Kit, cyclin D1, pStat3 (Y705) protein, and PKC isoforms β and δ. In contrast, pro-apoptotic proteins (such as Bim, Bax, Bid, and Bak) were significantly downregulated 26 . In addition, YK-4-279 drug-resistant cells displayed significant cross-resistance to both the PKC inhibitor enzastaurin and the US Food and Drug Administration-approved c-Kit inhibitor imatinib.…”

“…Regardless of the route of YK-4-279 administration, sustained complete responses were not documented across a complete cohort of treated animals 22, 24, 26 . As such, Lamhamedi-Cherradi et al recently investigated both de novo and acquired mechanism(s) in which Ewing sarcoma cells evade YK-4-279-mediated apoptosis 26 . YK-4-279 drug-resistant clones demonstrated enhanced proliferative rate over their parental cell lines and overexpressed c-Kit, cyclin D1, pStat3 (Y705) protein, and PKC isoforms β and δ.…”