Treatment with tyrosine kinase inhibitors results in a survival benefit in patients with chronic myeloid leukemia (CML). However, relapse due to persistent leukemic stem cells (LSCs) requires additional selective targets for efficient eradication of the disease. Metabolomic analyses on patient-derived CML LSCs reveal that these have an increased dependency on oxidative metabolism that renders them sensitive to treatment with tigecycline, an FDA-approved inhibitor of mitochondrial translation. These findings uncover a new metabolic vulnerability in CML and provide a rational approach for further clinical evaluation. Treatment of chronic myeloid leukemia (CML) with imatinib mesylate and other second- and/or third-generation c-Abl-specific tyrosine kinase inhibitors (TKIs) has substantially extended patient survival1. However, TKIs primarily target differentiated cells and do not eliminate leukemic stem cells (LSCs)2,3,4. Therefore, targeting minimal residual disease to prevent acquired resistance and/or disease relapse requires identification of new LSC-selective target(s) that can be exploited therapeutically5,6. Considering that malignant transformation involves cellular metabolic changes, which may in turn render the transformed cells susceptible to specific assaults in a selective manner7, we searched for such vulnerabilities in CML LSCs. We performed metabolic analyses on both stem cell–enriched (CD34+ and CD34+CD38−) and differentiated (CD34−) cells derived from individuals with CML, and we compared the signature of these cells with that of their normal counterparts. Through combination of stable isotope–assisted metabolomics with functional assays, we demonstrate that primitive CML cells rely on upregulated oxidative metabolism for their survival. We also show that combination treatment with imatinib and tigecycline, an antibiotic that inhibits mitochondrial protein translation, selectively eradicates CML LSCs both in vitro and in a xenotransplantation model of human CML. Our findings provide a strong rationale for investigation of the use of TKIs in combination with tigecycline to treat patients with CML with minimal residual disease.

The first checkpoint during T cell development, known as β selection, requires the successful rearrangement of the TCR-β gene locus. Notch signaling has been implicated in various stages during T lymphopoiesis. However, it is unclear whether Notch receptor-ligand interactions are necessary during β selection. Here, we show that pre-TCR signaling concurrent with Notch receptor and Delta-like-1 ligand interactions are required for the survival, proliferation, and differentiation of mouse CD4⁻CD8⁻ thymocytes to the CD4⁺CD8⁺ stage. Furthermore, we address the minimal signaling requirements underlying β selection and show a hierarchical positioning of key proximal signaling molecules. Collectively, our results demonstrate an essential role for Notch receptor-ligand interactions in enabling the autonomous signaling capacity of the pre-TCR complex.

Bipotent progenitors for T and natural killer (NK) lymphocytes are thought to exist among early precursor thymocytes. The identification and functional properties of such a progenitor population remain undefined. We report the identification of a novel developmental stage during fetal thymic ontogeny that delineates a population of T/NK-committed progenitors (NK1.1⁺/CD117⁺/CD44⁺/CD25⁻). Thymocytes at this stage in development are phenotypically and functionally distinguishable from the pool of multipotent lymphoid-restricted (B, T, and NK) precursor thymocytes. Exposure of multipotent precursor thymocytes or fetal liver– derived hematopoietic progenitors to thymic stroma induces differentiation to the bipotent developmental stage. Continued exposure to a thymic microenvironment results in predominant commitment to the T cell lineage, whereas coculture with a bone marrow–derived stromal cell line results in the generation of mature NK cells. Thus, the restriction point to T and NK lymphocyte destinies from a multipotent progenitor stage is marked by a thymus-induced differentiation step.

In chronic myeloid leukemia (CML), tyrosine kinase inhibitor (TKI) treatment induces autophagy that promotes survival and TKI-resistance in leukemic stem cells (LSCs). In clinical studies hydroxychloroquine (HCQ), the only clinically approved autophagy inhibitor, does not consistently inhibit autophagy in cancer patients, so more potent autophagy inhibitors are needed. We generated a murine model of CML in which autophagic flux can be measured in bone marrow-located LSCs. In parallel, we use cell division tracing, phenotyping of primary CML cells, and a robust xenotransplantation model of human CML, to investigate the effect of Lys05, a highly potent lysosomotropic agent, and PIK-III, a selective inhibitor of VPS34, on the survival and function of LSCs. We demonstrate that long-term haematopoietic stem cells (LT-HSCs: Lin⁻Sca-1⁺c-kit⁺CD48⁻CD150⁺) isolated from leukemic mice have higher basal autophagy levels compared with non-leukemic LT-HSCs and more mature leukemic cells. Additionally, we present that while HCQ is ineffective, Lys05-mediated autophagy inhibition reduces LSCs quiescence and drives myeloid cell expansion. Furthermore, Lys05 and PIK-III reduced the number of primary CML LSCs and target xenografted LSCs when used in combination with TKI treatment, providing a strong rationale for clinical use of second generation autophagy inhibitors as a novel treatment for CML patients with LSC persistence.