Abstract A123: Reversal of glucocorticoid resistance in pediatric acute lymphoblastic leukemia is dependent on restoring BIM expression

Abstract

Abstract Introduction: Acute lymphoblastic leukemia (ALL) is the most common pediatric malignancy. Glucocorticoids (e.g. dexamethasone, prednisolone) form a critical component of chemotherapy regimens for pediatric ALL and initial resistance to glucocorticoid therapy is predictive of poor outcome. Acquired resistance to glucocorticoids is common at relapse and is disparate compared with other chemotherapeutic drugs used in the treatment of pediatric ALL. Glucocorticoids induce apoptosis in lymphoid cells which is mediated through the glucocorticoid receptor (GR). Resistance to glucocorticoids in pediatric ALL is associated with upregulation of the oncogene C-MYC and failure to induce the proapoptotic gene BIM. The purpose of this study was to identify small molecules that are able to reverse glucocorticoid resistance in pediatric ALL patient-derived xenografts (PDXs). Methods: A 40,000 compound high-throughput screening (HTS) campaign was carried out to identify compounds that potentiated the cytotoxicity of dexamethasone against a pediatric ALL PDX derived from a patient with aggressive and fatal relapse. Cytotoxicity assays were carried out by alamarBlue assay. Gene expression analysis was carried out using Illumina microarrays. mRNA and protein analysis was carried out by RT-PCR and immunoblotting, respectively. Results: The HTS campaign identified a novel glucocorticoid sensitiser, 2-((4,5-dihydro-1H-imidazol-2-yl)thio)-N-isopropyl-N-phenylacetamide (GCS-3). The sensitizing effect was specific to glucocorticoids, with synergy observed when GCS-3 was combined with dexamethasone or prednisolone, but not with other chemotherapeutic drugs. Synergy was observed in a range of dexamethasone-resistant and -sensitive xenografts representative of B-ALL, T-ALL, early T-cell precursor ALL and Philadelphia chromosome positive ALL. GCS-3 required a functional GR to sensitise ALL xenografts to dexamethasone. Gene expression analysis showed that GCS-3 in the presence of dexamethasone upregulated a previously published gene expression signature induced by glucocorticoid treatment of B-ALL patients (Rhein et al. Leukemia, 2007, 21:897-905). Moreover, GCS-3 in combination with dexamethasone significantly downregulated C-MYC and upregulated BIM expression in a glucocorticoid-resistant ALL xenograft. Treatment with the GR antagonist, RU486, significantly abrogated the cytotoxic effects of the GCS-3/dexamethasone combination. Although RU486 increased C-MYC expression in the dexamethasone treated cells, there was no increase in C-MYC expression in the GCS-3/dexamethasone combination. RU486 suppressed the marked induction of BIM following treatment with the GCS-3/dexamethasone combination, indicating that BIM upregulation is critical for the cytotoxicity of the GCS-3/dexamethasone combination. The GCS-3/dexamethasone combination significantly increased binding of the GR to an intronic binding site at the BIM locus which is associated with BIM transcription and glucocorticoid sensitivity in pediatric ALL. Conclusion: This study describes the potential of the novel glucocorticoid sensitizing agent, GCS-3, as a biological tool to understand glucocorticoid action and resistance. Citation Format: Richard B Lock, Cara E Toscan, Duohui Jing, Chelsea Mayoh. Reversal of glucocorticoid resistance in pediatric acute lymphoblastic leukemia is dependent on restoring BIM expression [abstract]. In: Proceedings of the AACR-NCI-EORTC International Conference on Molecular Targets and Cancer Therapeutics; 2019 Oct 26-30; Boston, MA. Philadelphia (PA): AACR; Mol Cancer Ther 2019;18(12 Suppl):Abstract nr A123. doi:10.1158/1535-7163.TARG-19-A123