Targeting CDK6 and BCL2 Exploits the MYB Addiction of Ph+ Acute Lymphoblastic Leukemia

Philadelphia chromosome–positive acute lymphoblastic leukemia (Phþ ALL) is currently treated with BCR-ABL1 tyrosine kinase inhibitors (TKI) in combination with chemotherapy. However, most patients develop resistance to TKI through BCR-ABL1–dependent and –independent mechanisms. Newly developed TKI can target Phþ ALL cells with BCR-ABL1–dependent resistance; however, overcoming BCR-ABL1–independent mechanisms of resistance remains challenging because transcription factors, which are difficult to inhibit, are often involved. We show here that (i) the growth of Phþ ALL cell lines and primary cells is highly dependent on MYB-mediated transcriptional upregulation of CDK6, cyclin D3, and BCL2, and (ii) restoring their expression in MYB-silenced Phþ ALL cells rescues their impaired proliferation and survival. Levels of MYB and CDK6 were highly correlated in adult Phþ ALL (P ¼ 0.00008). Moreover, Phþ ALL cells exhibited a specific requirement for CDK6 but not CDK4 expression, most likely because, in these cells, CDK6 was predominantly localized in the nucleus, whereas CDK4 was almost exclusively cytoplasmic. Consistent with their essential role in Phþ ALL, pharmacologic inhibition of CDK6 and BCL2 markedly suppressed proliferation, colony formation, and survival of Phþ ALL cells ex vivo and in mice. In summary, these findings provide a proof-of-principle, rational strategy to target the MYB addiction of Phþ ALL. © 2017 American Association for Cancer Research

Targeting the cdk6 dependence of ph+ acute lymphoblastic leukemia

Ph+ ALL is a poor-prognosis leukemia subtype driven by the BCR-ABL1 oncogene, either the p190-or the p210-BCR/ABL isoform in a 70:30 ratio. Tyrosine Kinase inhibitors (TKIs) are the drugs of choice in the therapy of Ph+ ALL. In combination with standard chemotherapy, TKIs have markedly improved the outcome of Ph+ ALL, in particular if this treatment is followed by bone marrow transplantation. However, resistance to TKIs develops with high frequency, causing leukemia relapse that results i

Apoptosis: a Janus bifrons in T-cell immunotherapy

Immunotherapy has revolutionized the treatment of cancer. In particular, immune checkpoint blockade, bispecific antibodies, and adoptive T-cell transfer have yielded unprecedented clinical results in hematological malignancies and solid cancers. While T cell-based immunotherapies have multiple mechanisms of action, their ultimate goal is achieving apoptosis of cancer cells. Unsurprisingly, apoptosis evasion is a key feature of cancer biology. Therefore, enhancing cancer cells’ sensitivity to apoptosis represents a key strategy to improve clinical outcomes in cancer immunotherapy. Indeed, cancer cells are characterized by several intrinsic mechanisms to resist apoptosis, in addition to features to promote apoptosis in T cells and evade therapy. However, apoptosis is double-faced: when it occurs in T cells, it represents a critical mechanism of failure for immunotherapies. This review will summarize the recent efforts to enhance T cell-based immunotherapies by increasing apoptosis susceptibility in cancer cells and discuss the role of apoptosis in modulating the survival of cytotoxic T lymphocytes in the tumor microenvironment and potential strategies to overcome this issue

Structure of Nascent Chromatin Is Essential for Hematopoietic Lineage Specification

Summary: The role of chromatin structure in lineage commitment of multipotent hematopoietic progenitors (HPCs) is presently unclear. We show here that CD34+ HPCs possess a post-replicative chromatin globally devoid of the repressive histone mark H3K27me3. This H3K27-unmodified chromatin is required for recruitment of lineage-determining transcription factors (TFs) C/EBPα, PU.1, and GATA-1 to DNA just after DNA replication upon cytokine-induced myeloid or erythroid commitment. Blocking DNA replication or increasing H3K27me3 levels prevents recruitment of these TFs to DNA and suppresses cytokine-induced erythroid or myeloid differentiation. However, H3K27me3 is rapidly associated with nascent DNA in more primitive human and murine HPCs. Treatment of these cells with instructive cytokines leads to a significant delay in accumulation of H3K27me3 in nascent chromatin due to activity of the H3K27me3 demethylase UTX. Thus, HPCs utilize special mechanisms of chromatin modification for recruitment of specific TFs to DNA during early stages of lineage specification. : Petruk et al. find that hematopoietic progenitor cells possess a state of post-replicative chromatin globally devoid of the repressive histone mark H3K27me3. This de-condensed chromatin is required for recruitment of lineage-determining transcription factors to DNA just after replication in early stages of cytokine-induced myeloid or erythroid lineage specification. Keywords: DNA replication, nascent DNA, nascent chromatin, H3K27me3, KDMs, HMTs, hematopoietic progenitors, myeloid and erythroid differentiation, transcription factor

Generation and application of signaling pathway reporter lines in zebrafish

In the last years, we have seen the emergence of different tools that have changed the face of biology from a simple modeling level to a more systematic science. The transparent zebrafish embryo is one of the living models in which, after germline transformation with reporter protein-coding genes, specific fluorescent cell populations can be followed at single-cell resolution. The genetically modified embryos, larvae and adults, resulting from the transformation, are individuals in which time lapse analysis, digital imaging quantification, FACS sorting and next-generation sequencing can be performed in specific times and tissues. These multifaceted genetic and cellular approaches have permitted to dissect molecular interactions at the subcellular, intercellular, tissue and whole-animal level, thus allowing integration of cellular and developmental genetics with molecular imaging in the resulting frame of modern biology. In this review, we describe a new step in the zebrafish road to system biology, based on the use of transgenic biosensor animals expressing fluorescent proteins under the control of signaling pathway-responsive cis-elements. In particular, we provide here the rationale and details of this powerful tool, trying to focus on its huge potentialities in basic and applied research, while also discussing limits and potential technological evolutions of this approach

The zebrafish, a teleost model recapitulating the mammalian molecular events during endocrine development and function

In the last decades, the tropical teleost zebrafish (Danio rerio) has proved to be an excellent vertebrate system to model mammalian molecular events occurring during embryonic development, organ formation and adult physiology, either under normal or pathological conditions. Low costs and small dimensions, external fertilization and high fecundity, tissue transparency, rapid development, availability of mutant and transgenic lines, easy manipulability for gene perturbation and pharmacological screening, are some examples of the advantages characterizing this model organism. In the past 15 years, the use of zebrafish in the endocrinology field has been mainly focused on the analysis of endocrine organ development. Our team and other research groups have elucidated the main steps leading to the formation of endocrine glands such as pancreatic islets and thyroid, hypothalamus and pituitary, interrenal gland and gonads. Comparison of the zebrafish endocrine system to that of mammals has demonstrated that the systems are sufficiently similar for zebrafish to be employed as a model for endocrine research. In more recent years, new zebrafish-based tools have been generated to elucidate in vivo the molecular cross-talks occurring among cells, tissues and organs. Signalling pathway reporter lines represent an interesting implementation of classical transgenesis to visualize in vivo, in an intact organism, the anatomical regions that, in a given time interval, are responding to a specific signalling pathway. To generate these transgenic fish lines, signal specific responsive sequences, identified at the genomic level, are multimerized and placed upstream of reporter genes, typically encoding for fluorescent proteins such as GFP or mCherry. At present, a series of pathway reporter lines are already available, among which Bmp, Shh, FGF, Notch, TGF\u3b2, Wnt, hypoxia and glucocorticoid signalling. Our group is currently generating and validating additional lines, among which the cAMP/CREB pathway, while others, such as thyroid hormone and Foxo signalling, are in the planning phase. Preliminary results and envisaged applications will be presented and discussed

Inability to switch from ARID1A-BAF to ARID1B-BAF impairs exit from pluripotency and commitment towards neural crest formation in ARID1B-related neurodevelopmental disorders

Subunit switches in the BAF chromatin remodeler are essential during development. ARID1B and its paralog ARID1A encode for mutually exclusive BAF subunits. De novo ARID1B haploinsufficient mutations cause neurodevelopmental disorders, including Coffin-Siris syndrome, which is characterized by neurological and craniofacial features. Here, we leveraged ARID1B+/- Coffin-Siris patient-derived iPSCs and modeled cranial neural crest cell (CNCC) formation. We discovered that ARID1B is active only during the first stage of this process, coinciding with neuroectoderm specification, where it is part of a lineage-specific BAF configuration (ARID1B-BAF). ARID1B-BAF regulates exit from pluripotency and lineage commitment by attenuating thousands of enhancers and genes of the NANOG and SOX2 networks. In iPSCs, these enhancers are maintained active by ARID1A-containing BAF. At the onset of differentiation, cells transition from ARID1A- to ARID1B-BAF, eliciting attenuation of the NANOG/SOX2 networks and triggering pluripotency exit. Coffin-Siris patient cells fail to perform the ARID1A/ARID1B switch, and maintain ARID1A-BAF at the pluripotency enhancers throughout all stages of CNCC formation. This leads to persistent NANOG/SOX2 activity which impairs CNCC formation. Despite showing the typical neural crest signature (TFAP2A/SOX9-positive), ARID1B-haploinsufficient CNCCs are also aberrantly NANOG-positive. These findings suggest a connection between ARID1B mutations, neuroectoderm specification and a pathogenic mechanism for Coffin-Siris syndrome