DNMT3A Mutations Enhance T-ALL Survival, Which Is Reversed By JAK/STAT Inhibition

T-cell acute lymphoblastic leukemia (T-ALL) is an aggressive neoplasm of T-cell progenitors, which accounts for approximately 15% and 25% of pediatric and adult acute lymphoblastic leukemia (ALL) cases respectively. In T-ALL, accumulation of genomic abnormalities often leads to the aberrant expression of select groups of transcription factors that result in increased proliferation, cell survival and impaired differentiation of T-cell progenitors, which give rise to this disease

The Notch signaling pathway is essential for T-lymphopoiesis and T-cell differentiation from thymic precursors. Moreover, gain-of-function mutations in this pathway act as the major genetic lesions that drive T-ALL development (over 60% of adult cases). The de novo DNA methyltransferase enzyme (DNMT3A) is one of the three genes in mammals encoding for enzymes with DNA methyltransferase activity and mutations in this gene are frequently associated with NOTCH1 mutations in adult T-ALL. Interestingly, DNMT3A mutations are rare in pediatric and adolescent T-ALL. Prognosis of pediatric and adolescent T-ALL cases has seen major improvements during recent years with the introduction of high-dose, multi-agent chemotherapy regimens which has not carried over to adult T-ALL patients. These factors suggest the involvement of genetic differences between adult and pediatric T-ALL patients for the differential clinical outcomes

Previous work in our lab has shown that mouse T-ALL cells genetically deficient for Dnmt3a are less apoptotic and have increased JAK/STAT signaling. Here, we demonstrate that DNMT3A-mutant human T-ALL patient derived xenografts (PDXs) are resistant to the chemotherapeutic drugs dexamethasone (DEX) and doxorubicin compared to T-ALL patient specimens with wild-type DNMT3A. However, when primary DNMT3A-mutant human T-ALL cells were treated with dexamethasone or doxorubicin in combination with the JAK/STAT inhibitor ruxolitinib (RUX), we observed the survival benefit of DNMT3A-mutant T-ALL cells were mitigated, suggesting enhanced JAK/STAT signaling promotes survival in T-ALL patient cells with DNMT3A mutations.

To investigate the genes and pathways that endows DNMT3A-mutant T-ALL cells chemoresistance, gene expression profiling was performed by RNA-seq on DNMT3A-mutant, and wild-type human T-ALL cells treated with DEX+RUX. Gene set enrichment analysis (GSEA) showed that MTORC1, IL2-STAT5, E2F and G2M pathways were significantly downregulated in DEX+RUX treated DNMT3A-mutant PDX samples. E2F and G2M pathways are associated with cell cycle progression and further analysis of these two pathways revealed BIRC5 as a common gene that is downregulated in both these pathways following DEX+RUX treatment. Furthermore, our RNA-seq analysis demonstrated that the expression of BIRC5 in DNMT3A-mutant PDXs is downregulated upon DEX+RUX treatment compared to the DNMT3A-wildtype group. Moreover, baseline BIRC5 expression is higher in DNMT3A-mutant T-ALL PDXs compared to the DNMT3A-wildtype group. Following treatment with the BIRC5 inhibitor YM155, the viability of DNMT3A-mutant T-ALL cells decrease significantly compared to DNMT3A-wildtype PDXs, which were relatively unaffected. In addition, we observed a similar reduction in cell viability in DNMT3A-mutant PDXs following BIRC5 siRNA nucleofection compared to the DNMT3A-wildtype group. Cumulatively, these studies reveal that enhanced JAK/STAT signaling promotes survival and chemoresistance in T-ALL patient cells with DNMT3A mutations and identify the JAK/STAT target gene BIRC5 as a specific genetic dependency of DNMT3A-mutant T-ALL cells. These data provide a critical first step towards a novel target for precision medicine approaches for this patient group.

Challen:Incyte: Consultancy, Other: Sponsored Research agreements.