Abstract

Acute lymphoblastic leukemia (ALL) results from transformation of immature B or T cells, and is the most common pediatric cancer. Though the current cure rate of ALL is 80-90%, it is important to understand the underlying biology of ALL in order to develop refined therapies for patients who fail to respond to conventional chemotherapy as well as to reduce its toxicity. The IL-7 receptor (IL-7R) signaling pathway is necessary for the proliferation and survival of T cells. Together with collaborators, we have shown that 9% of patients with T cell acute lymphoblastic leukemia (T-ALL) have gain of function mutations in IL-7R alpha. These mutations promote homodimerization of IL-7R alpha subunits, resulting in constitutive activation of this pathway via Janus Kinase 1 (JAK1). As the JAK-STAT pathway is downstream of the IL-7 receptor, we hypothesized that JAK inhibitors could be used to treat T-ALL patients with IL-7R mutations. To demonstrate this, we first established a cell line model of T-ALL driven by constitutive IL-7R signaling. The model cells were established by transforming the D1 thymocyte cell line with a mutated IL-7R alpha derived from a patient sequence. These cells termed "D1_hIL7R_P1" are also GFP+, which allows us to monitor the proliferation of the cells in vivo.I have showed that D1_hIL7R_P1 cells delivered intravenously result in an aggressive leukemia with morbidity within 18-21 days. Ruxolitinib, a JAK1 inhibitor, inhibits survival, proliferation, and STAT5 activation of D1_hIL7R_P1 cells in vitro. To treat this leukemia in vivo, I administered ruxolitinib for 5 days at a dose of 150 mg/kg to mice starting 8 days after D1_hIL7R_P1 engraftment. Tissues were then harvested for analysis of GFP+ cells as a measure of leukemic burden via flow cytometry. Ruxolitinib reduced leukemic cells from 15% in the blood (as demonstrated in untreated mice) to 5%, from 30% to 10% in the spleen, and 40% to 20% in the lungs. With these promising results, I have acquired T-ALL patient samples that can be xenografted into NSG mice. TALL#5 cell line successfully engrafts in 30 days and can be detected in the bone marrow, spleen, and peripheral blood by flow cytometry. Additionally, TALL#5 expresses human IL-7R alpha and is sensitive to ruxolitinib treatment in vitro. I have also identified two T-ALL cell lines, DND41 and KOPTK1, which express high levels of IL-7R alpha. These cell lines are also potential candidates for studying the effects of ruxolitinib on T-ALL in NSG mice. TALL#5, DND41, and KOPTK1 will be engrafted into mice, and after leukemia is established, mice will be treated with ruxolitinib. If successful, this will give us insight into the effectiveness of JAK inhibitors in treating ALL, and which patients can be recruited for future clinical trials.

Disclosures

Off Label Use: Ruxolitinib is a JAK1/2 inhibitor that is FDA approved for myelofibrosis. My experiments involve pre-clinical studies looking at the efficacy of ruxolitinib in leukemic mice..

Author notes

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Asterisk with author names denotes non-ASH members.