IL-7 Receptor Mutations and Steroid Resistance in Pediatric T-Cell Acute Lymphoblastic Leukemia: a Genome Sequencing Study. IL7R signaling mutations confer steroid resistance in T-ALL

BACKGROUND: Pediatric acute lymphoblastic leukemia (ALL) is the most common childhood cancer and the leading cause of cancer-related mortality in children. T-cell acute lymphoblastic leukemia (T-ALL) represents about 15% of pediatric ALL cases and is considered a high-risk disease entity. T-ALL is often associated with resistance to treatment modalities, including steroids, which are currently the cornerstone for treating ALL; moreover, the initial steroid response strongly predicts survival and cure. However, the cellular mechanisms underlying steroid resistance in T-ALL patients are poorly understood. In this study, we combined various genomic datasets in order to identify candidate genetic mechanisms that underlie steroid resistance in children undergoing T-ALL treatment. METHODS AND FINDINGS: We performed whole-genome sequencing on paired pre-treatment (diagnostic) and post-treatment (remission) samples from 13 patients. In additions, we performed targeted exome sequencing of pre-treatment samples from 69 additional T-ALL patients. We then integrated mutation data with copy-number data for 151 mutated genes, and this integrated dataset was then correlated with clinical outcome and in vitro drug response. Our analysis revealed that mutations in JAK1 and KRAS, two genes encoding components of the interleukin 7 receptor (IL7R) signaling pathway, are associated with steroid resistance and poor outcome. We then sequenced other genes in this pathway, including IL7Ra, JAK1, JAK3, NF1, NRAS, KRAS, and AKT, in these 69 T-ALL patients and an additional 77 T-ALL patients. We identified mutations in 32% of patients, the majority of whom had a specific T-ALL subtype (ETP-ALL or TLX). Based on the outcome of these patients and their prednisolone responsiveness measured in vitro, we then confirmed that these mutations are associated with both steroid resistance and poor outcome. To functionally explore how these mutations cause steroid resistance and subsequent poor outcome, we expressed wild-type and mutant IL7R signaling molecules in two steroid-sensitive T-ALL cell lines (SUPT1 and P12 Ichikawa cells) using inducible lentiviral expression constructs. We found that expressing mutant IL7Ra, JAK1, or NRAS, wild-type NRAS, or wild-type AKT specifically induced steroid resistance in both lines without affecting sensitivity to vincristine or asparaginase. In contrast, wild-type IL7R, JAK1, and JAK3, as well as mutant JAK3 and mutant AKT, had no effect. We then performed a functional study to examine the mechanisms underlying steroid resistance and found that resistance was not due to a change in the steroid receptor’s ability to activate downstream targets. Rather, steroid resistance was associated with strong activation of MEK-ERK and AKT, downstream components of IL7R signaling pathway, thereby inducing a robust anti-apoptotic response by upregulating MCL1 and BCL-XL expression. Both the MEK-ERK and AKT pathways also inactivate BIM, an essential molecule for steroid-induced death, and inhibit GSK-3B, an important regulator of pro-apoptotic BIM. Importantly, treating our cell line models with IL7R signaling inhibitors restored steroid sensitivity. To show clinical relevance, we treated primary T-ALL cells obtained from 11 patients with steroids either alone or in combination with IL7R signaling inhibitors; we found that including MEK, AKT, mTOR, or a dual PI3K/mTOR inhibitor strongly increased steroid-induced cell death. Therefore, combining these inhibitors with steroid treatment may enhance steroid sensitivity in patients with ALL. CONCLUSIONS: Using an unbiased sequencing approach, we found that specific mutations in IL7R signaling molecules underlie steroid resistance in T-ALL. Therefore, future prospective clinical studies should include MEK, AKT, mTOR or a dual PI3K/mTOR inhibitor in order to restore—or even enhance—steroid sensitivity and improve clinical outcome.