T cell acute lymphoblastic leukaemia (T-ALL) is an aggressive haematological malignancy affecting 10-15% of pediatric ALL patients. Current cure rates of pediatric patients is 80% but increasing understanding of the molecular mechanisms of T-ALL provides possibilities for more effective and targeted therapies. During the last decade targeted therapies with tyrosine kinase inhibitors (TKI) have proven to be effective in BCR-ABL1 fusion positive leukemias. Several studies have suggested TKI dasatinib to be effective also in the treatment of NUP214-ABL1 positive T-ALLs, which comprise approximately 4-10% of the T-ALL cases.
Materials and Methods
In silico drug screening was performed by comparing gene expression profiles of 4769 leukemic samples to a library of 13384 compounds and their known targets from the Drug signature database (DsigDB). These compounds included FDA approved therapeutic molecules and molecules under studies. Findings were validated in an ex vivo drug screen, consisting of 20 T-ALL bone marrow samples and 9 healthy bone marrow controls. Samples were treated for 72 hours with five different concentrations of dasatinib in 10-fold dilutions (0.1-1000nM), cell viability was measured and the data normalized to negative (DMSO) and positive (benzethonium chloride) controls. The effect of dasatinib was further explored in vitro by treating one NUP214-ABL1 fusion positive and six fusion negative T-ALL cell lines with dasatinib (1-1000nM). Gene expression levels of the known dasatinib targets in these cell lines were measured by Global Run On sequencing (GRO-seq) assay and qRT-PCR.
In order to find novel targeted therapies for T-ALL, we performed an in silico drug target screen. A dasatinib-targetable gene LCK was strongly expressed in a number of T-ALL cases whereas normal T-lymphoid cells had lower expression. Chemical screen data of the target specificity of dasatinib showed high inhibition of LCK with percent of control (POC) value of 1, meaning that 0.1µM concentration of dasatinib decreases the kinase activity of LCK to 1% in comparison to control. In vitro dasatinib decreased cell viability in fusion negative Jurkat and MOLT-16 cells, and also in fusion positive cell line Peer. GRO-seq and qRT-PCR confirmed the expression of LCK and several other known dasatinib targets, including other SRC family kinases, in Jurkat and MOLT-16 cell lines. However, some LCK-expressing T-ALL cell lines were less sensitive to dasatinib. In further validation, ex vivo drug testing of patient samples revealed a marked response in 6/20 patient samples with IC50 values ranging between 1.3 - 8.2nM, while in healthy bone marrow controls IC50 values were >1000nM.
Our in silico drug screen identified dasatinib as a potential targeted therapy for a subgroup of T-ALL cases, and this finding was further supported by both ex vivo and in vitro studies. The exact mechanism remains to be elucidated but a number of SRC family kinases, which could potentially be targeted by dasatinib, showed expression in T-ALL samples.
Heckman:Celgene: Research Funding; Pfizer: Research Funding.
Asterisk with author names denotes non-ASH members.