Therapeutic targeting of ATR in alveolar rhabdomyosarcoma

Abstract

Despite advances in multi-modal treatment approaches, clinical outcomes of patients suffering from PAX3-FOXO1 fusion oncogene-expressing alveolar rhabdomyosarcoma (ARMS) remain dismal. Here we show that PAX3-FOXO1-expressing ARMS cells are sensitive to pharmacological ataxia telangiectasia and Rad3 related protein (ATR) inhibition. Expression of PAX3-FOXO1 in muscle progenitor cells is not only sufficient to increase sensitivity to ATR inhibition, but PAX3-FOXO1-expressing rhabdomyosarcoma cells also exhibit increased sensitivity to structurally diverse inhibitors of ATR. Mechanistically, ATR inhibition leads to replication stress exacerbation, decreased BRCA1 phosphorylation and reduced homologous recombination-mediated DNA repair pathway activity. Consequently, ATR inhibitor treatment increases sensitivity of ARMS cells to PARP1 inhibition in vitro, and combined treatment with ATR and PARP1 inhibitors induces complete regression of primary patient-derived ARMS xenografts in vivo. Lastly, a genome-wide CRISPR activation screen (CRISPRa) in combination with transcriptional analyses of ATR inhibitor resistant ARMS cells identifies the RAS-MAPK pathway and its targets, the FOS gene family, as inducers of resistance to ATR inhibition. Our findings provide a rationale for upcoming biomarker-driven clinical trials of ATR inhibitors in patients suffering from ARMS.

Elimusertib outperforms standard of care chemotherapy in preclinical patient-derived pediatric solid tumor models

“…We assessed the presence of frequent genetic alterations in pediatric tumors (46) as well as markers that cause genetic vulnerability to ATR inhibition (22,25,27,28,47,48) in our cell lines using publicly available datasets (49). In line with previous reports (28), the presence of MYCN amplifications, both on ecDNA or as homogenously staining regions (50,51), in NB cell lines, expression of fusion oncoproteins such as EWS-FLI1 or PAX3-FOXO1 (25,29) and TP53 deficiency (22) were associated with higher elimusertib sensitivity (Fig. 1m).…”

“…3a-af). Consistent with our previous work using AZD6738 (29) mice harboring PDX derived from ARMS showed the most pronounced response, with only one out of the seven ARMS PDX models classified as progressive disease after elimusertib treatment (Extended Data Fig. 3a-g) ERMS (Extended Data Fig.…”

“…1f). In line with previous reports testing other ATR inhibitors (24,26,29), cell lines derived from Ewing sarcoma, MYCN -amplified neuroblastoma and alveolar rhabdomyosarcoma were (significantly) more sensitive to ATR inhibition than control cell lines, suggesting a therapeutic window may exist for elimusertib in these pediatric solid tumors.…”

“…Notably, we observe that elimusertib outperformed most SoC agents in most entities, particularly in ARMS. This is in line with our previous reports describing the exquisite sensitivity of ARMS cells to ATR inhibition, which at least in part seem due to PAX3-FOXO1-induced replication stress (29). We propose that based on both our previous and current studies on ATR inhibitors, patients suffering from ARMS should be designated as a high-priority patient group in which ATR inhibitors should be tested clinically.…”

“…We and others have previously shown that diverse ATR inhibitors exhibit preclinical activity against a subset of ARMS, rhabdoid tumors, OS, EWS, MYCN -amplified neuroblastomas and medulloblastomas (24-26,28,29,61), but most of these studies only tested a small number of preclinical models and used ATR inhibitors that are currently not being clinically developed for the use in pediatric patients. In line with our results, the anti-tumor activity of different clinical-stage ATR inhibitors as monotherapy and in combination with other agents has been widely recognized in cancers in adults (21,22,26,38,48,59,62-64).…”