STAT5A and STAT5B—Twins with Different Personalities in Hematopoiesis and Leukemia

The transcription factors STAT5A and STAT5B have essential roles in survival and proliferation of hematopoietic cells—which have been considered largely redundant. Mutations of upstream kinases, copy number gains, or activating mutations in STAT5A, or more frequently in STAT5B, cause altered hematopoiesis and cancer. Interfering with their activity by pharmacological intervention is an up-and-coming therapeutic avenue. Precision medicine requests detailed knowledge of STAT5A’s and STAT5B’s individual functions. Recent evidence highlights the privileged role for STAT5B over STAT5A in normal and malignant hematopoiesis. Here, we provide an overview on their individual functions within the hematopoietic system

Twins with different personalities : STAT5B-but not STAT5A-has a key role in BCR/ABL-induced leukemia

Deregulation of the Janus kinase/signal transducers and activators of transcription (JAK/STAT) signaling pathway is found in cancer with STAT5A/B controlling leukemic cell survival and disease progression. As mutations in STAT5B, but not STAT5A, have been frequently described in hematopoietic tumors, we used BCR/ABL as model systems to investigate the contribution of STAT5A or STAT5B for leukemogenesis. The absence of STAT5A decreased cell survival and colony formation. Even more drastic effects were observed in the absence of STAT5B. STAT5B-deficient cells formed BCR/ABL(+) colonies or stable cell lines at low frequency. The rarely evolving Stat5b(-/-) cell lines expressed enhanced levels of BCR/ABL oncoprotein compared to wild-type cells. In line, Stat5b(-/-) leukemic cells induced leukemia with a significantly prolonged disease onset, whereas Stat5a(-/-) cells rapidly caused a fatal disease superimposable to wild-type cells. RNA-sequencing (RNA-seq) profiling revealed a marked enhancement of interferon (IFN)-alpha and IFN-gamma signatures in Stat5b(-/-) cells. Inhibition of IFN responses rescued BCR/ABL(+) colony formation of Stat5b(-/-)-deficient cells. A downregulated IFN response was also observed in patients suffering from leukemia carrying STAT5B mutations. Our data define STAT5B as major STAT5 isoform driving BCR/ABL(+) leukemia. STAT5B enables transformation by suppressing IFN-alpha/gamma, thereby facilitating leukemogenesis. Our findings might help explain the high frequency of STAT5B mutations in hematopoietic tumors.Peer reviewe

Cancers / Loss of STAT3 in lymphoma relaxes NK cell-mediated tumor surveillance

The transcription factors and proto-oncogenes STAT3 and STAT5 are highly activated in hematological malignancies and represent promising therapeutic targets. Whereas the importance of STAT5 as tumor promoter is beyond doubt, the role of STAT3 in hematological cancers is less well understood. Both, enforced as well as attenuated expression of STAT3 were reported in hematopoietic malignancies. Recent evidence implicates STAT3 as key player for tumor immune surveillance as it both mediates the production of and response to inflammatory cytokines. Here we investigated the effects of STAT3 deletion in a BCR/ABL-induced lymphoma model, which is tightly controlled by natural killer (NK) cells in vivo. Upon STAT3 deletion tumor growth is significantly enhanced when compared to STAT3-expressing controls. The increased tumor size upon loss of STAT3 was accompanied by reduced NK cell infiltration and decreased levels of the cytokine IFN- and the chemokine RANTES. Upon transplantation into NK cell-deficient mice differences in lymphoma size were abolished indicating that STAT3 expression in the tumor cells controls NK cell-dependent tumor surveillance. Our findings indicate that STAT3 inhibition in lymphoma patients will impair NK cell-mediated tumor surveillance, which needs to be taken into account when testing STAT3 inhibitors in preclinical or clinical trials.(VLID)217340

A robust approach for the generation of functional hematopoietic progenitor cell lines to model leukemic transformation

Studies of molecular mechanisms of hematopoiesis and leukemogenesis are hampered by the unavailability of progenitor cell lines that accurately mimic the situation in vivo. We now report a robust method to generate and maintain LSK (Lin-, Sca-1+, c-Kit+) cells, which closely resemble MPP1 cells. HPCLSKs reconstitute hematopoiesis in lethally irradiated recipient mice over >8 months. Upon transformation with different oncogenes including BCR/ABL, FLT3-ITD, or MLL-AF9, their leukemic counterparts maintain stem cell properties in vitro and recapitulate leukemia formation in vivo. The method to generate HPCLSKs can be applied to transgenic mice, and we illustrate it for CDK6-deficient animals. Upon BCR/ABLp210 transformation, HPCLSKs Cdk6-/- induce disease with a significantly enhanced latency and reduced incidence, showing the importance of CDK6 in leukemia formation. Studies of the CDK6 transcriptome in murine HPCLSK and human BCR/ABL+ cells have verified that certain pathways depend on CDK6 and have uncovered a novel CDK6-dependent signature, suggesting a role for CDK6 in leukemic progenitor cell homing. Loss of CDK6 may thus lead to a defect in homing. The HPCLSK system represents a unique tool for combined in vitro and in vivo studies and enables the production of large quantities of genetically modifiable hematopoietic or leukemic stem/progenitor cells

The different functions of Stat5 and chromatin alteration through Stat5 proteins

International audienceStat5 proteins modulate gene transcription upon cytokine- and growth factor action. Stat5a and Stat5b proteins alone are weak activators of transcription. They can modify chromatin organization through oligomerization and they act predominantly in co-operation and interaction with other proteins. The conservative view of exclusively nuclear functions of Stat5 was challenged by the observation of additional Stat5 effects in the cytoplasm, resulting in activation of the PI3K-Akt pathway. We summarize biological consequences of mutations in conserved domains of Stat5 or of deletions in the N- or C-terminal domains with impact on target gene transcription. Formation of higher-order oligomers is dramatically changed upon amino- or carboxyterminal deletions in Stat5 proteins. Mutations in or deletion of the Stat5 N-terminus leads to diminished leukemogenic potential of oncogenic Stat5, probably due to the inability to form Stat5 tetramers. The Stat5 N-terminal domain prevents persistent activation and can act as a DNA-docking platform for the glucocorticoid receptor (GR). The corresponding protocols should facilitate follow-up studies on Stat5 proteins and their contribution to normal physiological versus pathological processes through differential chromatin binding

CDK6 as a key regulator of hematopoietic and leukemic stem cell activation

The cyclin-dependent kinase 6 (CDK6) and CDK4 have redundant functions in reg- ulating cell-cycle progression. We describe a novel role for CDK6 in hematopoietic and leukemic stem cells (hematopoietic stem cells [HSCs] and leukemic stem cells [LSCs]) that exceeds its function as a cell-cycle regulator. Although hematopoiesis appears normal under steady-state conditions, Cdk62/2 HSCs do not efficiently repopulate upon competitive transplantation, and Cdk6-deficient mice are significantly more sus- ceptible to 5-fluorouracil treatment. We find that activation of HSCs requires CDK6, which interferes with the transcription of key regulators, including Egr1. Transcrip- tional profiling of HSCs is consistent with the central role of Egr1. The impaired repopulation capacity extends to BCR-ABLp2101 LSCs. Transplantation with BCR- ABLp2101–infected bone marrow from Cdk62/2 mice fails to induce disease, although recipient mice do harbor LSCs. Egr1 knock-down in Cdk62/2 BCR-ABLp2101 LSKs significantly enhances the potential to form colonies, underlining the importance of the CDK6-Egr1 axis. Our findings define CDK6 as an important regulator of stem cell activation and an essential component of a transcriptional complex that suppresses Egr1 in HSCs and LSCs.Depto. de Bioquímica y Biología MolecularFac. de Ciencias BiológicasTRUEpu

Isolation, Maintenance and Expansion of Adult Hematopoietic Stem/Progenitor Cells and Leukemic Stem Cells

Hematopoietic stem cells (HSCs) are rare, self-renewing cells that perch on top of the hematopoietic tree. The HSCs ensure the constant supply of mature blood cells in a tightly regulated process producing peripheral blood cells. Intense efforts are ongoing to optimize HSC engraftment as therapeutic strategy to treat patients suffering from hematopoietic diseases. Preclinical research paves the way by developing methods to maintain, manipulate and expand HSCs ex vivo to understand their regulation and molecular make-up. The generation of a sufficient number of transplantable HSCs is the Holy Grail for clinical therapy. Leukemia stem cells (LSCs) are characterized by their acquired stem cell characteristics and are responsible for disease initiation, progression, and relapse. We summarize efforts, that have been undertaken to increase the number of long-term (LT)-HSCs and to prevent differentiation towards committed progenitors in ex vivo culture. We provide an overview and compare methods currently available to isolate, maintain and enrich HSC subsets, progenitors and LSCs and discuss their individual advantages and drawbacks