Vascular endothelial cells are a critical component of the hematopoietic microenvironment that regulates blood cell production. Recent studies suggest the existence of functional cross-talk between ...hematologic malignancies and vascular endothelium. Here we show that human acute myeloid leukemia (AML) localizes to the vasculature in both patients and in a xenograft model. A significant number of vascular tissue-associated AML cells (V-AML) integrate into vasculature in vivo and can fuse with endothelial cells. V-AML cells acquire several endothelial cell-like characteristics, including the upregulation of CD105, a receptor associated with activated endothelium. Remarkably, endothelial-integrated V-AML shows an almost fourfold reduction in proliferative activity compared with non-vascular-associated AML. Primary AML cells can be induced to downregulate the expression of their hematopoietic markers in vitro and differentiate into phenotypically and functionally defined endothelial-like cells. After transplantation, these leukemia-derived endothelial cells are capable of giving rise to AML. These novel functional interactions between AML cells and normal endothelium along with the reversible endothelial cell potential of AML suggest that vascular endothelium may serve as a previously unrecognized reservoir for AML.
High-fidelity replication of DNA, and its accurate segregation to daughter cells, is critical for maintaining genome stability and suppressing cancer. DNA replication forks are stalled by many DNA ...lesions, activating checkpoint proteins that stabilize stalled forks. Stalled forks may eventually collapse, producing a broken DNA end. Fork restart is typically mediated by proteins initially identified by their roles in homologous recombination repair of DNA double-strand breaks (DSBs). In recent years, several proteins involved in DSB repair by non-homologous end joining (NHEJ) have been implicated in the replication stress response, including DNA-PKcs, Ku, DNA Ligase IV-XRCC4, Artemis, XLF and Metnase. It is currently unclear whether NHEJ proteins are involved in the replication stress response through indirect (signaling) roles, and/or direct roles involving DNA end joining. Additional complexity in the replication stress response centers around RPA, which undergoes significant post-translational modification after stress, and RAD52, a conserved HR protein whose role in DSB repair may have shifted to another protein in higher eukaryotes, such as BRCA2, but retained its role in fork restart. Most cancer therapeutic strategies create DNA replication stress. Thus, it is imperative to gain a better understanding of replication stress response proteins and pathways to improve cancer therapy.
Chk1 both arrests replication forks and enhances repair of DNA damage by phosphorylating downstream effectors. Although there has been a concerted effort to identify effectors of Chk1 activity, ...underlying mechanisms of effector action are still being identified. Metnase (also called SETMAR) is a SET and transposase domain protein that promotes both DNA double-strand break (DSB) repair and restart of stalled replication forks. In this study, we show that Metnase is phosphorylated only on Ser495 (S495) in vivo in response to DNA damage by ionizing radiation. Chk1 is the major mediator of this phosphorylation event. We had previously shown that wild-type (wt) Metnase associates with chromatin near DSBs and methylates histone H3 Lys36. Here we show that a Ser495Ala (S495A) Metnase mutant, which is not phosphorylated by Chk1, is defective in DSB-induced chromatin association. The S495A mutant also fails to enhance repair of an induced DSB when compared with wt Metnase. Interestingly, the S495A mutant demonstrated increased restart of stalled replication forks compared with wt Metnase. Thus, phosphorylation of Metnase S495 differentiates between these two functions, enhancing DSB repair and repressing replication fork restart. In summary, these data lend insight into the mechanism by which Chk1 enhances repair of DNA damage while at the same time repressing stalled replication fork restart.
A novel secreted cytokine, termed IL-17F, was cloned using nested RACE PCR. This cytokine bears homology to IL-17. IL-17F was expressed only in activated CD4(+) T cells and activated monocytes. ...Recombinant human IL-17F did not stimulate the proliferation of hematopoietic progenitors or the migration of mature leukocytes. However, it markedly inhibited the angiogenesis of human endothelial cells and induced endothelial cells to produce IL-2, TGF-beta, and monocyte chemoattractant protein-1.
MZF1 is a transcription factor belonging to the Krüppel family of zinc finger proteins, expressed in totipotent hemopoietic cells as well as in myeloid progenitors. Here we have inactivated Mzfi1 by ...gene targeting. Mzf1(-/-) mice develop lethal neoplasias characterized by the infiltration and complete disruption of the liver architecture by a monomorphic population of cells of myeloid origin reminiscent of human chloromas. Mzf1 inactivation results in a striking increase of the autonomous cell proliferation and of the ability of Mzf1(-/-) hemopoietic progenitors to sustain long-term hemopoiesis. These findings demonstrate that Mzf1 can act as a tumor/growth suppressor in the hemopoietic compartment.
Acute promyelocytic leukemia (APL) is characterized by a block in myeloid cell differentiation. As a result of a chromosomal translocation in these patients, the promyelocytic leukemia protein PML is ...disrupted as are the nuclear bodies it forms. Disruption of PML and PML nuclear bodies in APL is linked to a loss of growth control and subsequent leukemogenesis. PML contains a zinc-binding domain known as the RING which is required for formation of these bodies. Using yeast 2-hybrid techniques, we found that PML and a related RING protein, Z, bind the proline rich homeodomain protein (PRH) through their RING domains. Previous reports indicate that PRH functions in hematopoiesis and may act as a transcriptional repressor. Our data indicate that PML and Z both bind the repressor domain of PRH and are the first protein partners reported for PRH. We observe that PRH has a punctate pattern in both the nucleus and cytoplasm of chronic myelogenous leukemia K562 cells and in the APL cell line, NB4. Immunoprecipitation and co-localization studies indicate that PML and PRH interact in both cell lines. The effect on cell growth by PML and the hematopoietic actions of PRH raises the possibility that the interaction between PML and PRH represents a link between growth control and hematopoiesis.
A total of 50 consecutive patients (median age, 57.5 years) with AML (n=30) or myelodysplasia (MDS, n=20) underwent HLA matched related donor (MRD, n=27) or unrelated donor (MUD, n=23) peripheral ...blood hematopoietic cell transplantation after nonmyeloablative CY/fludarabine (Flu) conditioning. GVHD prophylaxis included CsA (n=19)+/-mycophenolate mofetil (n=31). At a median follow-up of 59 months, 21 patients (42%) were alive without evidence of disease. By Kaplan-Meier analysis, year 1-4 disease-free survival (DFS) and OS estimates were 0.50/0.58, 0.40/0.46, 0.37/0.43 and 0.37/0.41. MUD recipients were engrafted quickly (13.5 days) compared to MRD recipients (16 days) and relapsed/progressed less frequently (P=0.005). Overall grade 3/4 acute GVHD (aGVHD) occurred in 26% in the absence of antecedent mucositis and was associated with chronic GVHD (cGVHD) and poor OS. Extensive cGVHD developed in 51.2% of 100 day survivors. Rates of aGVHD, cGVHD and survival were similar between MRD and MUD recipients. Of 14 survivors with cGVHD, 5 (35.7%) experienced resolution off immunosuppression, suggesting that tolerance with HLA matched grafts is possible at an advanced age by this method. This study provides further evidence for prolonged DFS after CY/Flu MRD allotransplantation for AML/MDS, and extends the findings to older patients and those with unrelated donors.
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