Management of Patients with Acute Coronary Syndrome and Cancer.

Cancer patients with acute coronary syndrome (ACS) have significantly greater mortality compared with non-cancer patients. This risk is partly directly attributable to the malignancy; however these patients are frequently undertreated with respect to guideline recommended treatments for ACS due to higher bleeding risks from anemia and thrombocytopenia. Due to exclusion from large clinical trials, there is a paucity of data regarding how to best treat these complex and high-risk patients. PURPOSE OF REVIEW: To review the literature and identify risk factors among cancer patients associated with poor outcomes, pathophysiology of chemotherapy and radiation therapy contributing to accelerated coronary artery disease and ACS, and data regarding outcomes with medical therapy and invasive management. RECENT FINDINGS: Despite an elevated bleeding risk, many cancer patients may benefit from ACC/AHA guideline-directed management for ACS including aspirin, P2Y12 inhibitor, statin, and beta-blocker therapies. Cancer patients with ACS are a uniquely vulnerable population who are often undertreated, and with improved cancer treatments, this population is expected to increase. These patients should be included in future randomized trials to better understand how to balance the complexities of increased bleeding and thrombosis risks during ACS.

Aryl-hydrocarbon receptor activity modulates prolactin expression in the pituitary.

Pituitary tumors account for 15% of intracranial neoplasms, however the extent to which environmental toxicants contribute to the proliferation and hormone expression of pituitary cells is unknown. Aryl-hydrocarbon receptor (AhR) interacting protein (AIP) loss of function mutations cause somatotrope and lactotrope adenomas in humans. AIP sequesters AhR and inhibits its transcriptional function. Because of the link between AIP and pituitary tumors, we hypothesize that exposure to dioxins, potent exogenous ligands for AhR that are persistent in the environment, may predispose to pituitary dysfunction through activation of AhR. In the present study, we examined the effect of AhR activation on proliferation and endogenous pituitary hormone expression in the GH3 rat somatolactotrope tumor cell line and the effect of loss of AhR action in knockout mice. GH3 cells respond to nM doses of the reversible AhR agonist ß-naphthoflavone with a robust induction of Cyp1a1. Although mRNA levels of the anti-proliferative signaling cytokine TGFbeta1 are suppressed upon ß-naphthoflavone treatment, we did not observe an alteration in cell proliferation. AhR activation with ß-naphthoflavone suppresses Ahr expression and impairs expression of prolactin (PRL), but not growth hormone (GH) mRNA in GH3 cells. In mice, loss of Ahr similarly leads to a reduction in Prl mRNA at P3, while Gh is unaffected. Additionally, there is a significant reduction in pituitary hormones Lhb and Fshb in the absence of Ahr. Overall, these results demonstrate that AhR is important for pituitary hormone expression and suggest that environmental dioxins can exert endocrine disrupting effects at the pituitary.

A Src family kinase-Shp2 axis controls RUNX1 activity in megakaryocyte and T-lymphocyte differentiation.

Hematopoietic development occurs in complex microenvironments and is influenced by key signaling events. Yet how these pathways communicate with master hematopoietic transcription factors to coordinate differentiation remains incompletely understood. The transcription factor RUNX1 plays essential roles in definitive hematopoietic stem cell (HSC) ontogeny, HSC maintenance, megakaryocyte (Mk) maturation, and lymphocyte differentiation. It is also the most frequent target of genetic alterations in human leukemia. Here, we report that RUNX1 is phosphorylated by Src family kinases (SFKs) and that this occurs on multiple tyrosine residues located within its negative regulatory DNA-binding and autoinhibitory domains. Retroviral transduction, chemical inhibitor, and genetic studies demonstrate a negative regulatory role of tyrosine phosphorylation on RUNX1 activity in Mk and CD8 T-cell differentiation. We also demonstrate that the nonreceptor tyrosine phosphatase Shp2 binds directly to RUNX1 and contributes to its dephosphorylation. Last, we show that RUNX1 tyrosine phosphorylation correlates with reduced GATA1 and enhanced SWI/SNF interactions. These findings link SFK and Shp2 signaling pathways to the regulation of RUNX1 activity in hematopoiesis via control of RUNX1 multiprotein complex assembly.

Direct recruitment of polycomb repressive complex 1 to chromatin by core binding transcription factors.

Polycomb repressive complexes (PRCs) play key roles in developmental epigenetic regulation. Yet the mechanisms that target PRCs to specific loci in mammalian cells remain incompletely understood. In this study we show that Bmi1, a core component of Polycomb Repressive Complex 1 (PRC1), binds directly to the Runx1/CBFß transcription factor complex. Genome-wide studies in megakaryocytic cells demonstrate significant chromatin occupancy overlap between the PRC1 core component Ring1b and Runx1/CBFß and functional regulation of a considerable fraction of commonly bound genes. Bmi1/Ring1b and Runx1/CBFß deficiencies generate partial phenocopies of one another in vivo. We also show that Ring1b occupies key Runx1 binding sites in primary murine thymocytes and that this occurs via PRC2-independent mechanisms. Genetic depletion of Runx1 results in reduced Ring1b binding at these sites in vivo. These findings provide evidence for site-specific PRC1 chromatin recruitment by core binding transcription factors in mammalian cells.

Numb deletion in POMC-expressing cells impairs pituitary intermediate lobe cell adhesion, progenitor cell localization, and neuro-intermediate lobe boundary formation.

The pituitary gland contains six distinct hormone-secreting cell types that are essential for basic physiological processes including fertility and responding to stress. Formation of hormone-secreting cells during development relies on Notch signaling to prevent progenitors from prematurely differentiating. The nature of the signal curtailing Notch signaling in the pituitary is unknown, but a good candidate is the endocytic adaptor protein NUMB. NUMB targets Notch for proteolytic degradation, but it also has a broad range of actions, including stabilizing adherens junctions through interactions with cadherins and influencing cell proliferation by stabilizing expression of the tumor suppressor protein p53. Here, we show that NUMB and its closely related homolog, NUMBLIKE, are expressed in undifferentiated cells during development and later in gonadotropes in the anterior lobe and melanotropes of the intermediate lobe. All four isoforms of NUMB, are detectable in the pituitary, with the shorter forms becoming more prominent after adolescence. Conditionally deleting Numb and Numblike in the intermediate lobe melanotropes with Pomc Cre mice dramatically alters the morphology of cells in the intermediate lobe, coincident with impaired localization of adherens junctions proteins including E-CADHERIN, N-CADHERIN, ß-CATENIN, and α-CATENIN. Strikingly, the border between posterior and intermediate lobes is also disrupted. These mice also have disorganized progenitor cells, marked by SOX2, but proliferation is unaffected. Unexpectedly, Notch activity appears normal in conditional knockout mice. Thus, Numb is critical for maintaining cell-cell interactions in the pituitary intermediate lobe that are essential for proper cell placement.

Insights into GATA-1-mediated gene activation versus repression via genome-wide chromatin occupancy analysis.

The transcription factor GATA-1 is required for terminal erythroid maturation and functions as an activator or repressor depending on gene context. Yet its in vivo site selectivity and ability to distinguish between activated versus repressed genes remain incompletely understood. In this study, we performed GATA-1 ChIP-seq in erythroid cells and compared it to GATA-1-induced gene expression changes. Bound and differentially expressed genes contain a greater number of GATA-binding motifs, a higher frequency of palindromic GATA sites, and closer occupancy to the transcriptional start site versus nondifferentially expressed genes. Moreover, we show that the transcription factor Zbtb7a occupies GATA-1-bound regions of some direct GATA-1 target genes, that the presence of SCL/TAL1 helps distinguish transcriptional activation versus repression, and that polycomb repressive complex 2 (PRC2) is involved in epigenetic silencing of a subset of GATA-1-repressed genes. These data provide insights into GATA-1-mediated gene regulation in vivo.

The role and regulation of friend of GATA-1 (FOG-1) during blood development in the zebrafish.

The nuclear protein FOG-1 binds transcription factor GATA-1 to facilitate erythroid and megakaryocytic maturation. However, little is known about the function of FOG-1 during myeloid and lymphoid development or how FOG-1 expression is regulated in any tissue. We used in situ hybridization, gain- and loss-of-function studies in zebrafish to address these problems. Zebrafish FOG-1 is expressed in early hematopoietic cells, as well as heart, viscera, and paraspinal neurons, suggesting that it has multifaceted functions in organogenesis. We found that FOG-1 is dispensable for endoderm specification but is required for endoderm patterning affecting the expression of late-stage T-cell markers, independent of GATA-1. The suppression of FOG-1, in the presence of normal GATA-1 levels, induces severe anemia and thrombocytopenia and expands myeloid-progenitor cells, indicating that FOG-1 is required during erythroid/myeloid commitment. To functionally interrogate whether GATA-1 regulates FOG-1 in vivo, we used bioinformatics combined with transgenic assays. Thus, we identified 2 cis-regulatory elements that control the tissue-specific gene expression of FOG-1. One of these enhancers contains functional GATA-binding sites, indicating the potential for a regulatory loop in which GATA factors control the expression of their partner protein FOG-1.

Differentiation-dependent interactions between RUNX-1 and FLI-1 during megakaryocyte development.

The transcription factor RUNX-1 plays a key role in megakaryocyte differentiation and is mutated in cases of myelodysplastic syndrome and leukemia. In this study, we purified RUNX-1-containing multiprotein complexes from phorbol ester-induced L8057 murine megakaryoblastic cells and identified the ets transcription factor FLI-1 as a novel in vivo-associated factor. The interaction occurs via direct protein-protein interactions and results in synergistic transcriptional activation of the c-mpl promoter. Interestingly, the interaction fails to occur in uninduced cells. Gel filtration chromatography confirms the differentiation-dependent binding and shows that it correlates with the assembly of a complex also containing the key megakaryocyte transcription factors GATA-1 and Friend of GATA-1 (FOG-1). Phosphorylation analysis of FLI-1 with uninduced versus induced L8057 cells suggests the loss of phosphorylation at serine 10 in the induced state. Substitution of Ser10 with the phosphorylation mimic aspartic acid selectively impairs RUNX-1 binding, abrogates transcriptional synergy with RUNX-1, and dominantly inhibits primary fetal liver megakaryocyte differentiation in vitro. Conversely, substitution with alanine, which blocks phosphorylation, augments differentiation of primary megakaryocytes. We propose that dephosphorylation of FLI-1 is a key event in the transcriptional regulation of megakaryocyte maturation. These findings have implications for other cell types where interactions between runx and ets family proteins occur.

Identification of ZBP-89 as a novel GATA-1-associated transcription factor involved in megakaryocytic and erythroid development.

A complete understanding of the transcriptional regulation of developmental lineages requires that all relevant factors be identified. Here, we have taken a proteomic approach to identify novel proteins associated with GATA-1, a lineage-restricted zinc finger transcription factor required for terminal erythroid and megakaryocytic maturation. We identify the Krüppel-type zinc finger transcription factor ZBP-89 as being a component of multiprotein complexes involving GATA-1 and its essential cofactor Friend of GATA-1 (FOG-1). Using chromatin immunoprecipitation assays, we show that GATA-1 and ZBP-89 cooccupy cis-regulatory elements of certain erythroid and megakaryocyte-specific genes, including an enhancer of the GATA-1 gene itself. Loss-of-function studies in zebrafish and mice demonstrate an in vivo requirement for ZBP-89 in megakaryopoiesis and definitive erythropoiesis but not primitive erythropoiesis, phenocopying aspects of FOG-1- and GATA-1-deficient animals. These findings identify ZBP-89 as being a novel transcription factor involved in erythroid and megakaryocytic development and suggest that it serves a cooperative function with GATA-1 and/or FOG-1 in a developmental stage-specific manner.

Antagonism of FOG-1 and GATA factors in fate choice for the mast cell lineage.

The zinc finger transcription factor GATA-1 requires direct physical interaction with the cofactor friend of GATA-1 (FOG-1) for its essential role in erythroid and megakaryocytic development. We show that in the mast cell lineage, GATA-1 functions completely independent of FOG proteins. Moreover, we demonstrate that FOG-1 antagonizes the fate choice of multipotential progenitor cells for the mast cell lineage, and that its down-regulation is a prerequisite for mast cell development. Remarkably, ectopic expression of FOG-1 in committed mast cell progenitors redirects them into the erythroid, megakaryocytic, and granulocytic lineages. These lineage switches correlate with transcriptional down-regulation of GATA-2, an essential mast cell GATA factor, via switching of GATA-1 for GATA-2 at a key enhancer element upstream of the GATA-2 gene. These findings illustrate combinatorial control of cell fate identity by a transcription factor and its cofactor, and highlight the role of transcriptional networks in lineage determination. They also provide evidence for lineage instability during early stages of hematopoietic lineage commitment.

Coregulator-dependent facilitation of chromatin occupancy by GATA-1.

Coregulator recruitment by DNA-bound factors results in chromatin modification and protein-protein interactions, which regulate transcription. However, the mechanism by which the Friend of GATA (FOG) coregulator mediates GATA factor-dependent transcription is unknown. We showed previously that GATA-1 replaces GATA-2 at an upstream region of the GATA-2 locus, and that this GATA switch represses GATA-2. Genetic complementation analysis in FOG-1-null hematopoietic precursors revealed that FOG-1 is not required for establishment or maintenance of the active GATA-2 domain, but is critical for the GATA switch. Analysis of GATA factor binding to additional loci also revealed FOG-1-dependent GATA switches. Thus, FOG-1 facilitates chromatin occupancy by GATA-1 at sites bound by GATA-2. We propose that FOG-1 is a prototype of a new class of coregulators termed chromatin occupancy facilitators, which confer coregulation in certain contexts via enhancing trans-acting factor binding to chromatin in vivo.