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γ-secretase inhibitors reverse glucocorticoid resistance in T cell acute lymphoblastic leukemia

Abstract

Gamma-secretase inhibitors (GSIs) block the activation of the oncogenic protein Notch homolog-1 (NOTCH1) in T cell acute lymphoblastic leukemia (T-ALL). However, limited antileukemic cytotoxicity and severe gastrointestinal toxicity have restricted the clinical application of these targeted drugs. Here we show that combination therapy with GSIs plus glucocorticoids can improve the antileukemic effects of GSIs and reduce their gut toxicity in vivo. Inhibition of NOTCH1 signaling in glucocorticoid-resistant T-ALL restored glucocorticoid receptor autoupregulation and induced apoptotic cell death through induction of the gene encoding BCL-2–like apoptosis initiator-11 (BCL2L11). GSI treatment resulted in cell cycle arrest and accumulation of goblet cells in the gut mediated by upregulation of the gene encoding the transcription factor Krüppel-like factor-4 (Klf4), a negative regulator of the cell cycle required for goblet cell differentiation. In contrast, glucocorticoid treatment induced transcriptional upregulation of cyclin D2 (Ccnd2) and protected mice from developing the intestinal goblet cell metaplasia typically induced by inhibition of NOTCH signaling with GSIs. These results support a role for glucocorticoids plus GSIs in the treatment of glucocorticoid-resistant T-ALL.

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Figure 1: GSIs reverse glucocorticoid resistance in T-ALL cells.
Figure 2: Inhibition of NOTCH1-HES1 signaling restores glucocorticoid receptor autoregulation.
Figure 3: BCL2L11 upregulation reverses glucocorticoid resistance in T-ALL cells treated with dexamethasone plus CompE.
Figure 4: Interaction of NOTCH inhibition and dexamethasone treatment in tumor response and gut toxicity in vivo.
Figure 5: NOTCH-HES1 signaling regulates KLF4 expression in the gut.
Figure 6: Glucocorticoid-induced Cnnd2 upregulation mediates the enteroprotective effect of dexamethasone against GSI-induced gut toxicity.

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Acknowledgements

We thank U. Klein for help with flow cytometry analysis; T. Matos for the processing of histological, histochemical and imunohistochemical assays; P. Sicinski (Dana-Farber Cancer Institute) for the Ccnd2-knockout mouse; T. Honjo (Kyoto University) and B. Reitzis (Columbia University) for the Rbpjflox/flox conditional knockout mouse; T. Ludwig (Columbia University) for the Cre-Tam mouse line; A. Kung (Dana-Farber Cancer Institute) for the FUW-Luc neo vector; J. Aster (Brigham and Women's Hospital) for the MigR1 and MigR1-ICN1 retroviral constructs and for the NOTCH1-TAD antibody; W. Hahn (Dana-Farber Cancer Institute) for the pLKO shRNA vector; R. Dalla Favera (Columbia University) for the pCM8-cMYB vector; M.J. Garabedian (New York University) for the pCMV-HA-hGR vector; and N. Wu and C. Wa for pharmacokinetic and drug metabolism analysis. This work was supported by the US National Institutes of Health (R01CA120196 and R01CA129382 to A.F. and R56AI070310, 1R01CA105129 and 1R01CA133379 to I.A.); the Wipe Out Leukemia Forever Foundation (A.F.), the Leukemia and Lymphoma Society (grants 1287-08 and 6237-08 to A.F.), the Charlotte Geyer Foundation (A.F.), the Cancer Research Institute (A.F.), the Swim Across America Foundation (A.F.), the Golfers Against Cancer Foundation (A.F.), the American Cancer Society (RSG0806801 to I.A.), the Città della Speranza Foundation, a Cassa di Risparmio di Padova e Rovigo research grant, a Programmi di Ricerca Scientifica di Rilevante Interesse Nazionale, and the Fondo de Investigacion Sanitaria (grant CD07/00033 to P.J.R.). A.F. and I.A. are Leukemia & Lymphoma Society Scholars. T.P. is a recipient of a Young Investigator Award from the Alex's Lemonade Stand Foundation. I.H. is supported by the Dutch Cancer Society.

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Contributions

A.F. supervised the project. P.J.R., V.T., T.P., M.L.S, K.B., C.S. and I.H. designed and performed experiments. M.C. performed histological and immunohistochemical studies. E.H. and E.d.S. assisted in bioimaging analysis of tumor xenografts. G.B. provided clinical samples. J.M. provided clinical samples and supervised drug sensitivity assays in these cells. A.F. and I.A. supervised the analysis of Ccnd2−/− mice. C.C.-C. supervised histological and immunohistochemical studies. W.A. designed and performed experiments on the regulation of Klf4 by NOTCH1 and HES1. A.F. wrote the manuscript.

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Correspondence to Adolfo Ferrando.

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Real, P., Tosello, V., Palomero, T. et al. γ-secretase inhibitors reverse glucocorticoid resistance in T cell acute lymphoblastic leukemia. Nat Med 15, 50–58 (2009). https://doi.org/10.1038/nm.1900

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