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The NUP214-ABL1 fusion protein is a constitutively active protein tyrosine kinase that is found in 6% of patients with T-cell acute lymphoblastic leukemia and that promotes proliferation and survival of T-lymphoblasts. Although NUP214-ABL1 is sensitive to ABL1 kinase inhibitors, development of resistance to these compounds is a major clinical problem, underlining the need for additional drug targets in the sparsely studied NUP214-ABL1 signaling network. In this work, we identify and validate the SRC family kinase LCK as a protein whose activity is absolutely required for the proliferation and survival of T-cell acute lymphoblastic leukemia cells that depend on NUP214-ABL1 activity. These findings underscore the potential of SRC kinase inhibitors and of the dual ABL1/SRC kinase inhibitors dasatinib and bosutinib for the treatment of NUP214-ABL1-positive T-cell acute lymphoblastic leukemia. In addition, we used mass spectrometry to identify protein interaction partners of NUP214-ABL1. Our results strongly support that the signaling network of NUP214-ABL1 is distinct from that previously reported for BCR-ABL1. Moreover, we found that three NUP214-ABL1-interacting proteins, MAD2L1, NUP155, and SMC4, are strictly required for the proliferation and survival of NUP214-ABL1-positive T-cell acute lymphoblastic leukemia cells. In conclusion, this work identifies LCK, MAD2L1, NUP155 and SMC4 as four new potential drug targets in NUP214-ABL1-positive T-cell acute lymphoblastic leukemia.

Background Activating NOTCH1 mutations are common in T-cell acute lymphoblastic leukemia. Inhibition of NOTCH1 signaling with γ-secretase inhibitors causes cell cycle block, but only after treatment for several days. We further documented the effects of γ-secretase inhibitor treatment on T-cell acute lymphoblastic leukemia cell lines and tested whether combining γ-secretase inhibitors with other anti-cancer drugs offers a therapeutic advantage.Design and Methods The effect of γ-secretase inhibitor treatment and combinations of γ-secretase inhibitors with chemotherapy or glucocorticoids was assessed on T-cell acute lymphoblastic leukemia cell lines. We sequenced NOTCH1 in T-cell acute lymphoblastic leukemia cases with ABL1 fusions and tested combinations of γ-secretase inhibitors and the ABL1 inhibitor imatinib in a T-cell acute lymphoblastic leukemia cell line.Results γ-secretase inhibitor treatment for 5–7 days reversibly inhibited cell proliferation, caused cell cycle block in sensitive T-cell acute lymphoblastic leukemia cell lines, and caused differentiation of some T-cell acute lymphoblastic leukemia cell lines. Treatment for 14 days or longer was required to induce significant apoptosis. The cytotoxic effects of the chemotherapeutic agent vincristine were not significantly enhanced by addition of γ-secretase inhibitors to T-cell acute lymphoblastic leukemia cell lines, but γ-secretase inhibitor treatment sensitized cells to the effect of dexamethasone. NOTCH1 mutations were identified in all T-cell acute lymphoblastic leukemia patients with ABL1 fusions and in a T-cell acute lymphoblastic leukemia cell line expressing NUP214-ABL1. In this cell line, the anti-proliferative effect of imatinib was increased by pre-treatment with γ-secretase inhibitors.Conclusions Short-term treatment of T-cell acute lymphoblastic leukemia cell lines with γ-secretase inhibitors had limited effects on cell proliferation and survival. Combinations of γ-secretase inhibitors with other drugs may be required to obtain efficient therapeutic effects in T-cell acute lymphoblastic leukemia, and not all combinations may be useful.