Mathilde Broekhuis

Inhibition of glycolysis modulates prednisolone resistance in acute lymphoblastic leukemia cells.

Treatment failure in pediatric acute lymphoblastic leukemia (ALL) is related to cellular resistance to glucocorticoids (eg, prednisolone). Recently, we demonstrated that genes associated with glucose metabolism are differentially expressed between prednisolone-sensitive and prednisolone-resistant precursor B-lineage leukemic patients. Here, we show that prednisolone resistance is associated with increased glucose consumption and that inhibition of glycolysis sensitizes prednisolone-resistant ALL cell lines to glucocorticoids. Treatment of prednisolone-resistant Jurkat and Molt4 cells with 2-deoxy-D-glucose (2-DG), lonidamine (LND), or 3-bromopyruvate (3-BrPA) increased the in vitro sensitivity to glucocorticoids, while treatment of the prednisolone-sensitive cell lines Tom-1 and RS4; 11 did not influence drug cytotoxicity. This sensitizing effect of the glycolysis inhibitors in glucocorticoid-resistant ALL cells was not found for other classes of antileukemic drugs (ie, vincristine and daunorubicin). Moreover, down-regulation of the expression of GAPDH by RNA interference also sensitized to prednisolone, comparable with treatment with glycolytic inhibitors. Importantly, the ability of 2-DG to reverse glucocorticoid resistance was not limited to cell lines, but was also observed in isolated primary ALL cells from patients. Together, these findings indicate the importance of the glycolytic pathway in glucocorticoid resistance in ALL and suggest that targeting glycolysis is a viable strategy for modulating prednisolone resistance in ALL.

Polycomb Cbx family members mediate the balance between haematopoietic stem cell self-renewal and differentiation

The balance between self-renewal and differentiation of adult stem cells is essential for tissue homeostasis. Here we show that in the haematopoietic system this process is governed by polycomb chromobox (Cbx) proteins. Cbx7 is specifically expressed in haematopoietic stem cells (HSCs), and its overexpression enhances self-renewal and induces leukaemia. This effect is dependent on integration into polycomb repressive complex-1 (PRC1) and requires H3K27me3 binding. In contrast, overexpression of Cbx2, Cbx4 or Cbx8 results in differentiation and exhaustion of HSCs. ChIP-sequencing analysis shows that Cbx7 and Cbx8 share most of their targets; we identified approximately 200 differential targets. Whereas genes targeted by Cbx8 are highly expressed in HSCs and become repressed in progenitors, Cbx7 targets show the opposite expression pattern. Thus, Cbx7 preserves HSC self-renewal by repressing progenitor-specific genes. Taken together, the presence of distinct Cbx proteins confers target selectivity to PRC1 and provides a molecular balance between self-renewal and differentiation of HSCs.

Counting stem cells: methodological constraints

The number of stem cells contributing to hematopoiesis has been a matter of debate. Many studies use retroviral tagging of stem cells to measure clonal contribution. Here we argue that methodological factors can impact such clonal analyses. Whereas early studies had low resolution, leading to underestimation, recent methods may result in an overestimation of stem-cell counts. We discuss how restriction enzyme choice, PCR bias, high-throughput sequencing depth and tagging method could affect the conclusions of clonal studies.

Asymmetry in skeletal distribution of mouse hematopoietic stem cell clones and their equilibration by mobilizing cytokines

Upon transplant, functional HSC clones preferentially expand in certain skeletal locations, exhibiting only limited migration toward other niches.

Pyruvate kinase M2 and prednisolone resistance in acute lymphoblastic leukemia.

Treatment of childhood acute lymphoblastic leukemia (ALL) combines different classes of chemotherapeutic agents, such as Vinca alkaloids, anthracyclines and glucocorticoids (GCs). Although such therapy nowadays cures the majority of the patients, combination chemotherapy still fails in approximately

Tracing dynamics and clonal heterogeneity of Cbx7-induced leukemic stem cells by cellular barcoding.

Summary Accurate monitoring of tumor dynamics and leukemic stem cell (LSC) heterogeneity is important for the development of personalized cancer therapies. In this study, we experimentally induced distinct types of leukemia in mice by enforced expression of Cbx7. Simultaneous cellular barcoding allowed for thorough analysis of leukemias at the clonal level and revealed high and unpredictable tumor complexity. Multiple LSC clones with distinct leukemic properties coexisted. Some of these clones remained dormant but bore leukemic potential, as they progressed to full-blown leukemia after challenge. LSC clones could retain multilineage differentiation capacities, where one clone induced phenotypically distinct leukemias. Beyond a detailed insight into CBX7-driven leukemic biology, our model is of general relevance for the understanding of tumor dynamics and clonal evolution.